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| Appl Biochem Biotechnol, 2002 Spring, 98-100, 909 - 20 Enhancement of oxygen transfer by pressure pulsation in aqueous glycerol fermentation; Huang WC et al.; Aeration plays an important role in the production of glycerol by fermentation with yeast . Effective aeration depends on a number of factors, such as amount of air, fineness of air dispersion, rate of agitation, and time of gas-liquid contact . This investigation dealt with the effect of periodic variation in gas pressure on oxygen transfer measured by sulfite oxidation and glycerol fermentation in stirred tanks . The oxygen transfer rate measured with the sulfite oxidation method was improved by 20-30% under the condition of pressure pulsation (PP) at 30 degrees C . The yield and productivity of glycerol were increased by about 26 and 6.8%, respectively, in 48 h by employing a glucose concentration of 250 g/L with PP at 30 degrees C. Appl Biochem Biotechnol, 2002 Spring, 98-100, 875 - 83 Xylose reductase activity of Candida guilliermondii during xylitol production by fed-batch fermentation: selection of process variables; Rodrigues DC et al.; Xylose reductase activity of Candida guilliermondii FTI 20037 was evaluated during xylitol production by fed-batch fermentation of sugarcane bagasse hydrolysate . A 2(4-1) fractional factorial design was used to select process variables . The xylose concentrations in the feeding solution (S(F)) and in the fermentor (S0), the pH, and the aeration rate were selected for optimization of this process, which will be undertaken in the near future . The best experimental result was achieved at S(F) = 45 g/L, S0 = 40 g/L, pH controlled at 6.0, and aeration rate of 1.2 vvm . Under these conditions, the xylose reductase activity was 0.81 U/mg of protein and xylitol production was 26.3 g/L, corresponding to a volumetric productivity of 0.55 g/(L x h) and a xylose xylitol yield factor of 0.68 g/g. Appl Biochem Biotechnol, 2002 Spring, 98-100, 849 - 61 Recirculation of process streams in fuel ethanol production from softwood based on simultaneous saccharification and fermentation; Alkasrawi M et al.; The effect of process stream recirculation on ethanol production from steam- pretreated softwood based on simultaneous saccharification and fermentation (SSF) was investigated for two process configurations . In the first configuration, a part of the stillage stream after distillation was recycled and, in the second configuration, the liquid after SSF was recycled . The aim was to minimize the energy consumption in the distillation of the fermentation broth and in the evaporation of the stillage, as well as the use of fresh water . However, recirculation leads to an increased concentration of nonvolatiles in the first configuration, and of both volatiles and nonvolatiles in the second configuration . These substances might be inhibitory to the enzymes and the yeast in SSF . When 60% of the fresh water was replaced by stillage, the ethanol yield and the productivity were the same as for the configuration without recirculation . The ethanol production cost was reduced by 17% . In the second configuration, up to 40% of the fresh water could be replaced without affecting the final ethanol yield, although the initial ethanol productivity decreased . The ethanol production cost was reduced by 12% . At higher degrees of recirculation, fermentation was clearly inhibited, resulting in a decrease in ethanol yield while hydrolysis seemed unaffected. Appl Biochem Biotechnol, 2002 Spring, 98-100, 815 - 32 Enhancing the enzymatic hydrolysis of cellulosic materials using simultaneous ball milling; Mais U et al.; One of the limiting factors restricting the effective and efficient bioconversion of softwood-derived lignocellulosic residues is the recalcitrance of the substrate following pretreatment . Consequently, the ensuing enzymatic process requires relatively high enzyme loadings to produce monomeric carbohydrates that are readily fermentable by ethanologenic microorganisms . In an attempt to circumvent the need for larger enzyme loadings, a simultaneous physical and enzymatic hydrolysis treatment was evaluated . A ball-mill reactor was used as the digestion vessel, and the extent and rate of hydrolysis were monitored . Concurrently, enzyme adsorption profiles and the rate of conversion during the course of hydrolysis were monitored . alpha-Cellulose, employed as a model substrate, and SO2-impregnated steam-exploded Douglas-fir wood chips were assessed as the cellulosic substrates . The softwood-derived substrate was further posttreated with water and hot alkaline hydrogen peroxide to remove >90% of the original lignin . Experiments at different reaction conditions were evaluated, including substrate concentration, enzyme loading, reaction volumes, and number of ball beads employed during mechanical milling . It was apparent that the best conditions for the enzymatic hydrolysis of alpha-cellulose were attained using a higher number of beads, while the presence of air-liquid interface did not seem to affect the rate of saccharification . Similarly, when employing the lignocellulosic substrate, up to 100% hydrolysis could be achieved with a minimum enzyme loading (10 filter paper units/g of cellulose), at lower substrate concentrations and with a greater number of reaction beads during milling . It was apparent that the combined strategy of simultaneous ball milling and enzymatic hydrolysis could improve the rate of saccharification and/or reduce the enzyme loading required to attain total hydrolysis of the carbohydrate moieties. Appl Biochem Biotechnol, 2002 Spring, 98-100, 747 - 51 Production of coconut aroma by fungi cultivation in solid-state fermentation; de Alberto AA et al.; The production of 6-pentyl-alpha-pyrone (6-PP), an unsaturated D-lactone with a strong coconut-like aroma was studied and compared with liquid and solid substrates . A fungi strain that produces coconut aroma compound was selected . The liquid medium of the submerged culture was used to impregnate a solid support of sugarcane bagasse in SSF (Solid State Fermentation) . This substrate was adequate for growth and aroma production; the concentration obtained using SSF was higher than using liquid fermentation process . In the present work, it is demonstrated that, by solid-state-fermentation process, it is possible to produce 6-PP . The amount of 6-PP produced using a solid state substrate, following a 5 d culture, was 3 mg/g dry matter . Therefore, the amount of 6-PP produced during solid-state-fermentation process is higher than that reported in literature for submerged process. Appl Biochem Biotechnol, 2002 Spring, 98-100, 733 - 46 Modeling simultaneous saccharification and fermentation of softwood; Pettersson PO et al.; Simultaneous saccharification and fermentation (SSF) of wood has been modeled for the past 15-20 years, but the substrates used for model evaluation have so far not included pretreated softwood . In the present study, data from lab-scale batch SSF of SO2-impregnated, steam-pretreated spruce chips were used to evaluate a model found in the literature . The model, which was somewhat modified, consists of a number of nonlinear, coupled ordinary differential equations, which were solved numerically . Some parameter values were fitted to data by use of least-squares minimization . A difficulty in parameter estimation was the lack of cellobiose measurements, something that was relieved by adding assumptions about parameter relations . The simulated concentration profiles agreed well with the measured concentrations of glucose and ethanol . It is therefore concluded that the basic model features apply to softwood SSF . The model predicts rate saturation with respect to enzyme concentration at concentrations above 60 FPU/g cellulose, although this was not observed in the experimental data, which only comprised enzyme concentrations up to 32 FPU/g cellulose. Appl Biochem Biotechnol, 2002 Spring, 98-100, 717 - 32 Ethanol production from olive oil extraction residue pretreated with hot water; Ballesteros I et al.; The olive pulp fraction contained in the residue generated in olive oil extraction by a two-step centrifugation process can be upgraded by using the cellulose fraction to produce ethanol and recovering high value phenols (tyrosol and hydroxytyrosol) . Olive pulp was pretreated in a laboratory scale stirred autoclave at different temperatures (150-250 degrees C) . Pretreatment was evaluated regarding cellulose recovery, enzymatic hydrolysis effectiveness, ethanol production by a simultaneous saccharification and fermentation process (SSF), and phenols recovery in the filtrate . The pretreatment of olive pulp using water at temperatures between 200 degrees C and 250 degrees C enhanced enzymatic hydrolysis . Maximum ethanol production (11.9 g/L) was obtained after pretreating pulp at 210 degrees C in a SSF fed-batch procedure . Maximum hydroxytyrosol recovery was obtained in the liquid fraction when pretreated at 230 degrees C. Appl Biochem Biotechnol, 2002 Spring, 98-100, 699 - 716 Comparison of the fermentability of enzymatic hydrolyzates of sugarcane bagasse pretreated by steam explosion using different impregnating agents; Martin C et al.; Sugarcane bagasse is a potential lignocellulosic feedstock for ethanol production, since it is cheap, readily available, and has a high carbohydrate content . In this work, bagasse was subjected to steam explosion pretreatment with different impregnation conditions . Three parallel pretreatments were carried out, one without any impregnation, a second with sulfur dioxide, and a third with sulfuric acid as the impregnating agent . The pretreatments were performed at 205 degrees C for 10 min . The pretreated material was then hydrolyzed using cellulolytic enzymes . The chemical composition of the hydrolyzates was analyzed . The highest yields of xylose (16.2 g/100 g dry bagasse), arabinose (1.5 g/100 g), and total sugar (52.9 g/100 g) were obtained in the hydrolysis of the SO2 -impregnated bagasse . The H2SO4 -impregnated bagasse gave the highest glucose yield (35.9 g/100 g) but the lowest total sugar yield (42.3 g/100 g) among the three methods . The low total sugar yield from the H2SO4-impregnated bagasse was largely due to by-product formation, as the dehydration of xylose to furfural . Sulfuric acid impregnation led to a three-fold increase in the concentration of the fermentation inhibitors furfural and 5-hydroxymethylfurfural (HMF) and a two-fold increase in the concentration of inhibitory aliphatic acids (formic, acetic, and levulinic acids) compared to the other two pretreatment methods . The total content of phenolic compounds was not strongly affected by the different pretreatment methods, but the quantities of separate phenolic compounds were widely different in the hydrolyzate from the H2SO4-impregnated bagasse compared with the other two hydrolyzates . No major differences in the content of inhibitors were observed in the hydrolyzates obtained from SO2-impregnated and non-impregnated bagasse . The fermentability of all three hydrolyzates was tested with a xylose-utilizing Saccharomyces cerevisiae strain with and without nutrient supplementation . The hydrolyzates of SO2-impregnated and nonimpregnated bagasse showed similar fermentability, whereas the hydrolyzate of H2SO4-impregnated bagasse fermented considerably poorer. Appl Biochem Biotechnol, 2002 Spring, 98-100, 663 - 77 Growth of lignocellulosic-fermenting fungi on different substrates under low oxygenation conditions; Pavarina EC et al.; Four soil fungi able to grow under low oxygenation conditions were selected and used in studies to determine the production of enzymes that promote the degradation of lignocellulosic materials . The capacity of these fungi to ferment such materials was also investigated . The fungi were grown in sugarcane bagasse and sawdust at final concentrations of 4 and 10%, as the carbon sources . The strains were cultivated under microaerophilic and combined conditions of oxygenation (aerobic followed by microaerophilic conditions) . The results obtained with the basidiomycete specie, Trichocladium canadense, Geotrichum sp., and Fusarium sp . suggest that they prefer lower oxygen concentration for growth and enzyme production . Lignocellulolytic activities were detected in all strains but varied with the carbon source used . The highest levels of these activities were produced by the Basidiomycete specie and Fusarium sp . Ethanol and other nongaseous fermentation products were detected following high-performance liquid chromatography analysis using a supelcogel C-610H column, demonstrating the fermentative capability of these strains . In view of their ability to produce enzymes necessary for the breakdown of lignocellulosic materials and to utilize most of the degradation products for growth, these strains have a great potential for biotechnological application. Appl Biochem Biotechnol, 2002 Spring, 98-100, 599 - 610 Pressure pulsation in solid-phase fermentation; Yang X et al.; Inadequate dissipation of heat generated by biologic activities has prevented the use of solid-phase fermentation in large-scale applications . This study deals with the cooling effects of pressure pulsation on solid, porous beds . Pressure pulsation also enhances evaporation of medium moisture, which is also described . Computer software has been developed for on-line control of heat accumulation and moisture loss involving automatic variation in pressure pulsation frequency and airflow direction as well as replenishment of water . Aspergillus niger NRRL3 was cultivated on a moist, solid medium made of wheat bran and ground corncobs to produce cellobiase . During 100 h of fermentation, the maximum temperature inside the solid bed was kept under 40 degrees C, and the medium water content was successfully maintained between 61 and 65%, which was optimal for cell growth . Cells grew heavily on the solid-phase substrate and distributed uniformly . With good on-line control of temperature and moisture, the 12-L fermentor provided a better environment for enzyme production than 250 mL flasks did. Appl Biochem Biotechnol, 2002 Spring, 98-100, 59 - 72 SO2-catalyzed steam explosion of corn fiber for ethanol production; Bura R et al.; Corn fiber, a by-product of the corn wet-milling industry, represents a renewable resource that is readily available in significant quantities and could potentially serve as a low-cost feedstock for the production of fuel-grade alcohol . In this study, we used a batch reactor to steam explode corn fiber at various degrees of severity to evaluate the potential of using this feedstock in the bioconversion process . The results indicated that maximum sugar yields (soluble and following enzymatic hydrolysis) were recovered from corn fiber that was pretreated at 190 degrees C for 5 min with 6% SO2 . Sequential SO2-catalyzed steam explosion and enzymatic hydrolysis resulted in very high conversion (81%) of all polysaccharides in the corn fiber to monomeric sugars . Subsequently, Saccharomyces cerevisiae was able to convert the resultant corn fiber hydrolysates to ethanol very efficiently, yielding 90-96% of theoretical conversion during the fermentation process. Appl Biochem Biotechnol, 2002 Spring, 98-100, 563 - 75 Treatment with lignin residue: a novel method for detoxification of lignocellulose hydrolysates; Bjorklund L et al.; Acid hydrolysis of lignocellulose to hydrolysates intended for production of fuel ethanol results in the formation of byproducts in addition to fermentable sugars . Some of the byproducts, such as phenolic compounds and furan aldehydes, are inhibitory to the fermenting microorganism . Detoxification of the hydrolysates may be necessary for production of ethanol at a satisfactory rate and yield . The lignin residue obtained after hydrolysis is a material with hydrophobic properties that is produced in large amounts as a byproduct within an ethanol production process based on lignocellulosic raw materials . We have explored the possibility of using this lignin residue for detoxification of spruce dilute-acid hydrolysates prior to fermentation with Saccharomyces cerevisiae . Three dilute-acid hydrolysates of spruce were treated with lignin residue, which in all cases resulted in improved fermentability in terms of productivity and yield of ethanol . The effect was improved by washing the lignin before treatment, by using larger amounts of lignin in the treatment, and by performing the treatment at low temperature . Treatment with the lignin residue removed up to 53% of the phenolic compounds and up to 68% of the furan aldehydes in a spruce dilute-acid hydrolysate . A larger fraction of furfural was removed compared to the less hydrophobic 5-hydroxymethylfurfural. Appl Biochem Biotechnol, 2002 Spring, 98-100, 509 - 23 Use of measurement uncertainty analysis to assess accuracy of carbon mass balance closure for a cellulase production process; Schell DJ et al.; Closing carbon mass balances is a critical and necessary step for verifying the performance of any conversion process . We developed a methodology for calculating carbon mass balance closures for a cellulase production process and then applied measurement uncertainty analysis to calculate 95% confidence limits to assess the accuracy of the results . Cellulase production experiments were conducted in 7-L fermentors using Trichoderma reesei grown on pure cellulose (Solka-floc), glucose, or lactose . All input and output carbon-containing streams were measured and carbon dioxide in the exhaust gas was quantified using a mass spectrometer . On Solka-floc, carbon mass balances ranged from 90 to 100% closure for the first 48 h but increased to 101 to 135% closure from 72 h to the end of the cultivation at 168 h . Carbon mass balance closures for soluble sugar substrates ranged from 92 to 127% over the entire course of the cultivations . The 95% confidence intervals (CIs) for carbon mass balance closure were typically +/-11 to 12 percentage points after 48 h of cultivation . Many of the carbon mass balance results did not bracket 100% closure within the 95% CIs . These results suggest that measurement problems with the experimental or analytical methods may exist . This work shows that uncertainty analysis can be a useful diagnostic tool for identifying measurement problems in complex biochemical systems. Appl Biochem Biotechnol, 2002 Spring, 98-100, 5 - 21 Two-step steam pretreatment of softwood with SO2 impregnation for ethanol production; Soderstrom J et al.; Two-step steam pretreatment of softwood was investigated with the aim of improving the enzymatic digestibility for ethanol production . In the first step, softwood was impregnated with SO2 and steam pretreated at different severities . The first step was performed at low severity to hydrolyze the hemicellulose and release the sugars into the solution . The combination of time and temperature that yielded the highest amount of hemicellulosic sugars in the solution was determined . In the second step, the washed solid material from the optimized first step was impregnated once more with SO2 and steam pretreated under more severe conditions to enhance the enzymatic digestibility . The investigated temperature range was between 180 and 220 degrees C, and the residence times were 2, 5 and 10 min . The effectiveness of pretreatment was assessed by both enzymatic hydrolysis of the solids and simultaneous saccharification and fermentation (SSF) of the whole slurry after the second pretreatment step, in the presence of antibiotics . For each pretreatment combination, the liquid fraction was fermented to determine any inhibiting effects . At low severity in the second pretreatment step, a high conversion of cellulose was obtained in the enzymatic hydrolysis step, and at a high severity a high conversion of cellulose was obtained in the second pretreatment step . This resulted in an overall yield of sugars that was nearly constant over a wide range of severity . Compared with the one-step steam pretreatment, the two-step steam pretreatment resulted in a higher yield of sugar and in a slightly higher yield of ethanol . The overall sugar yield, when assessed by enzymatic hydrolysis, reached 80% . In the SSF configuration, an overall ethanol yield of 69% was attained. Appl Biochem Biotechnol, 2002 Spring, 98-100, 49 - 58 Dilute-acid hydrolysis of sugarcane bagasse at varying conditions; Neureiter M et al.; Sugarcane bagasse, a byproduct of the cane sugar industry, is an abundant source of hemicellulose that could be hydrolyzed to yield a fermentation feedstock for the production of fuel ethanol and chemicals . The effects of sulfuric acid concentration, temperature, time, and dry matter concentration on hemicellulose hydrolysis were studied with a 20-L batch hydrolysis reactor using a statistical experimental design . Even at less severe conditions considerable amounts (>29%) of the hemicellulose fraction could be extracted . The percentage of soluble oligosaccharides becomes very low in experiments with high yields in monosaccharides, which indicates that the cellulose fraction is only slightly affected . For the sugar yields, acid concentration appears to be the most important parameter, while for the formation of sugar degradation products, temperature shows the highest impact . It could be demonstrated that the dry matter concentration in the reaction slurry has a negative effect on the xylose yield that can be compensated by higher concentrations of sulfuric acid owing to a positive interaction between acid concentration and dry matter contents. Appl Biochem Biotechnol, 2002 Spring, 98-100, 489 - 96 Use of immobilized Candida yeast cells for xylitol production from sugarcane bagasse hydrolysate: cell immobilization conditions; Carvalho W et al.; Candida guilliermondii cells were immobilized in Ca-alginate beads and used for xylitol production from concentrated sugarcane bagasse hydrolysate . A full factorial design was employed to determine whether variations in the immobilization conditions would have any effects on the beads, chemical stability and on the xylitol production rates . Duplicate fermentation runs were carried out in 125-mL Erlenmeyer flasks maintained in a rotatory shaker at 30 degrees C and 200 rpm for 72 h . Samples were periodically analyzed to monitor xylose and acetic acid consumption, xylitol production, free cell growth, and bead solubilization . Concentrations of sodium alginate at 20.0 g/L and calcium chloride at 11.0 g/L and bead curing time of 24 h represented the most appropriate immobilization conditions within the range of conditions tested. Appl Biochem Biotechnol, 2002 Spring, 98-100, 449 - 58 Influence of oxygen availability on cell growth and xylitol production by Candida guilliermondii; Faria LF et al.; Oxygen availability is the most important environmental parameter in the production of xylitol by yeasts, directly affecting yields and volumetric productivity . This work evaluated the cell behavior in fermentations carried out with different dissolved oxygen concentrations (0.5-30.0% of saturation), as well as a limited oxygen restriction (0% of saturation), at several oxygen volumetric transfer coefficients (12 < or = kLa < or = 70 h(-1)) . These experiments allowed us to establish the specific oxygen uptake rate limits to ensure high yields and volumetric productivity . When oxygen availability was limited, the specific oxygen uptake rate values were between 12 and 26 mg of O2/of g cell x h, resulting in a yield of 0.71 g of xylitol/xylose consumed, and 0.85 g/{L x h} for the volumetric productivity . According to the results, the effective control of the specific oxygen uptake rate makes it possible to establish complete control over this fermentative process, for both cell growth and xylitol production. Appl Biochem Biotechnol, 2002 Spring, 98-100, 403 - 13 Xylose reductase and xylitol dehydrogenase activities of Candida guilliermondii as a function of different treatments of sugarcane bagasse hemicellulosic hydrolysate employing experimental design; Alves LA et al.; The sugarcane bagasse hydrolysate, which is rich in xylose, can be used as culture medium for Candida guilliermondii in xylitol production . However, the hydrolysate obtained from bagasse by acid hydrolysis at 120 degrees C for 20 min has by-products (acetic acid and furfural, among others), which are toxic to the yeast over certain concentrations . So, the hydrolysate must be pretreated before using in fermentation . The pretreatment variables considered were: adsorption time (15,37.5, and 60 min), type of acid used (H2So4 and H3Po4), hydrolysate concentration (original, twofold, and fourfold concentrated), and active charcoal (0.5, 1.75 and 3.0%) . The suitability of the pretreatment was followed by measuring the xylose reductase (XR) and xylitol dehydrogenase (XD) activity of yeast grown in each treated hydrolysate . The response surface methodology (2(4) full factorial design with a centered face) indicated that the hydrolysate might be concentrated fourfold and the pH adjusted to 7.0 with CaO, followed by reduction to 5.5 with H3PO4 . After that it was treated with active charcoal (3.0%) by 60 min . This pretreated hydrolysate attained the high XR/XD ratio of 4.5. Appl Biochem Biotechnol, 2002 Spring, 98-100, 229 - 41 Enhanced formation of extracellular laccase activity by the white-rot fungus Trametes multicolor; Hess J et al.; The white-rot fungus Trametes multicolor MB 49 has been identified as an excellent producer of the industrially important enzyme laccase . The formation of extracellular laccase could be considerably stimulated by the addition of Cu(II) to a simple, glycerol-based culture medium . In this study, optimal concentrations of copper were found to be 0.5-1 mM, which were added during the growth phase of the fungus . Other medium components important for laccase production are the carbon and nitrogen sources employed . When using an optimized medium containing glycerol (40 g/L), peptone from meat (15 g/L), and MgSO4 x 7H2O and stimulating enzyme formation by the addition of 1.0 mM Cu, maximal laccase activities obtained in shake-flask cultures were approx 85 U/mL . These results, however, could not be scaled up to a laboratory fermentor cultivation . Laccase production by T . multicolor decreased considerably when the fungus was grown in a stirred-tank reactor, presumably because of damage of the mycelia caused by shear stress and/or changes in the morphology of the fungus. Appl Biochem Biotechnol, 2002 Spring, 98-100, 205 - 13 Effect of kLa on the production of glucose 6-phosphate dehydrogenase from Saccharomyces cerevisiae grown by fermentation process; Silva DP et al.; In a 5-L fermentor (NBS-MF 105), Saccharomyces cerevisiae (0.7 g/L) was inoculated into a liquid medium (pH 4.0) containing 17 g/L of glucose, 2.55 g/L of yeast extract, 4.25 g/L of peptone, 2.04 g/L of Na2HPO4 x 12H2O, 4.34 g/L of (NH4)2SO4 and 0.064 g/L of MgSO4 x 7H2O and aerobically cultivated at 35 degrees C for 22 h . Agitation and aeration were adjusted to attain initial kLa values of 15, 60, 135, and 230 h(-1) . The glucose 6-phosphate dehydrogenase (G6PDH) productivity (PrG6PDH) obtained for kLa values of 15, 60, 135, and 230 h(-1) was 10.6, 31.8, 30.3, and 23.3 U/({Lx h}), respectively, whereas the cell productivity (Pr(x)) for the same kLa values were 0.24, 0.69, 0.69, and 0.49 g/{L x h}, respectively . Thus, both events are coupled and depend on the dissolved oxygen in the medium. Appl Biochem Biotechnol, 2002 Spring, 98-100, 1161 - 72 Combined sedimentation and filtration process for cellulase recovery during hydrolysis of lignocellulosic biomass; Knutsen JS et al.; A combined sedimentation and ultrafiltration process was investigated for recovering cellulase enzymes during the hydrolysis of lignocellulosic biomass . Lignocellulosic particles larger than approx 50 microm in length were first removed via sedimentation using an inclined settler . Ultrafiltration was then used to retain the remaining lignocellulosic particles and the cellulose enzymes, while transmitting fermentable sugars and other small molecules . The permeate flux from the ultrafiltration step for a feed consisting of 0.22 w/v% cellulase is 64+/-5 L/m2-h, while that for a feed consisting of the settler overflow from a mixture 0.22 w/v% cellulase and 10 wt% lignocellulose fed to the settler is 130+/-20 L/m2-h . The higher permeate flux in the latter case is presumably due to binding of a portion of the cellulase enzymes to the lignocellulosic particles during hydrolysis and filtration, preventing the enzymes from fouling the membrane . A filter paper activity assay shows little loss in enzymatic activity throughout the combined sedimentation/ultrafiltration separation process. Appl Biochem Biotechnol, 2002 Spring, 98-100, 1105 - 14 Mixed submerged fermentation with two filamentous fungi for cellulolytic and xylanolytic enzyme production; Garcia-Kirchner O et al.; The efficient saccharification of lignocellulosic materials requires the cooperative actions of different cellulase enzyme activities: exoglucanase, endoglucanase, beta-glucosidase, and xylanase . Previous studies with the fungi strains Aureobasidium sp . CHTE-18, Penicillium sp . CH-TE-001, and Aspergillus terreus CH-TE-013, selected mainly because of their different cellulolytic and xylanolytic activities, have demonstrated the capacity of culture filtrates of cross-synergistic action in the saccharification of native sugarcane bagasse pith . In an attempt to improve the enzymatic hydrolysis of different cellulosic materials, we investigated a coculture fermentation with two of these strains to enhance the production of cellulases and xylanases . The 48-h batch experimental results showed that the mixed culture of Penicillium sp . CH-TE-001 and A . terreus CH-TE-013 produced culture filtrates with high protein content, cellulase (mainly beta-glucosidase), and xylanase activities compared with the individual culture of each strain . The same culture conditions were used in a simple medium with mineral salts, corn syrup liquor, and sugarcane bagasse pith as the sole carbon source with moderate shaking at 29 degrees C . Finally, we compared the effect of the cell-free culture filtrates obtained from the mixed and single fermentations on the saccharification of different kinds of cellulosic materials. Appl Biochem Biotechnol, 2002 Spring, 98-100, 1049 - 59 Oxygen uptake rate in production of xylitol by Candida guilliermondii with different aeration rates and initial xylose concentrations; Gimenes MA et al.; The global oxygen uptake rate (OUR) and specific oxygen uptake rates (SOUR) were determined for different values of the volumetric oxygen mass transfer coefficient (15, 43, and 108 h(-1)), and for varying initial xylose concentrations (50, 100, 150, and 200 g/L) in shaking flasks . The initial cell concentration was 4.0 g/L, and there was only significant growth in the fermentation with the highest oxygen availability . In this condition, OUR increased proportionally to cell growth, reaching maximum values from 2.1 to 2.5 g of O2/(L x h) in the stationary phase when the initial substrate concentration was raised from 50 to 200 g/L, respectively . SOUR showed different behavior, growing to a maximum value coinciding with the beginning of the exponential growth phase, after which point it decreased . The maximum SOUR values varied from 265 to 370 mg of O2/(g of cell x h), indicating the interdependence of this parameter and the substrate concentration . Although the volumetric productivity dropped slightly from 1.55 to 1.18 g of xylitol/(L x h), the strain producing capacity (Y(P/X)) rose from 9 to 20.6 g/g when the initial substrate concentration was increased from 50 to 200 g/L . As for the xylitol yield over xylose consumed (Y(P/S)), there was no significant variation, resulting in a mean value of 0.76 g/g . The results are of interest in establishing a strategy for controlling the dynamic oxygen supply to maximize volumetric productivity. Appl Biochem Biotechnol, 2002 Spring, 98-100, 1009 - 23 Hybrid neural modeling of bioprocesses using functional link networks; Harada LH et al.; The objective of this work was to develop a model for an extractive ethanol fermentation in a simple and rapid way . This model must be sufficiently reliable to be used for posterior optimization and control studies . A hybrid neural model was developed, combining mass and energy balances with neural networks, which describe the process kinetics . To determine the best model, two structures of neural networks were compared: the functional link networks and the feedforward neural networks . The two structures are shown to describe well the process kinetics, and the advantages of using the functional link networks are discussed. Nahrung, 2002 Apr, 46(2), 92 - 5 Effect of soaking, germination, cooking and fermentation on antinutritional factors in cowpeas; Ibrahim SS et al.; The present work deals with the study of efficacy of some treatments, namely soaking (in water and bicarbonate solution), ordinary and pressure cooking, germination and fermentation in reducing or removal of antinutritional factors usually present in cowpeas (protease inhibitors, tannins, phytic acid and flatus-producing oligosaccharides (raffinose and stachyose) . The results showed that long-time soaking (16 h) in bicarbonate solution caused remarkable reduction in the antinutritional factors . Pressure cooking was more effective than ordinary . Cooking pregerminated cowpeas was most effective . Fermentation completely removed trypsin inhibitor, oligosaccharides and reduced remarkably phytic acid . However, tannins noticeably increased. Nahrung, 2002 Apr, 46(2), 64 - 7 Effect of bottle colour and storage conditions on browning of orange wine; Selli S et al.; We investigated the effect of bottle colour, storage temperature and storage time on the browning of orange wine . Kozan orange variety was used for the production of wine . The fermentation was performed spontaneously at 20 degrees C . The wine was bottled in three differently coloured bottles: clear white, green and brown . Then each was divided into two parts: one part was stored at 13-14 degrees C in the cellar and the other at 23-26 degrees C in the laboratory for 150 days . During the storage, browning index and ascorbic acid content of the wines were measured at day 0, 75 and 150 . The results showed that the use of brown bottles and the short-time storage reduced the browning in orange wines, however, storage at two different temperatures did not significantly affect the browning index. Yao Xue Xue Bao, 1998 Jul, 33(7), 481 - 5 {Fibrinolytic properties and thrombolytic effect of a novel fibrinolytic enzyme from Streptomyces sp . Y405}; Wang M et al.; A novel protease (designated as SW-1) purified from the fermentation broth of Streptomyces sp . strain Y405 showed fibrinolytic activity . In vitro, on plasminogen-free fibrin plate which had been heated at 85 degrees C for 30 minutes to eliminate plasminogen, SW-1 showed the same fibrinolytic activity as the mixture of SW-1 and plasminogen . Without adding plasminogen into the fibrin clot tube SW-1 showed the same effect as with plasminogen (P > 0.05) . The results might indicate that SW-1 is a fibrinolytic enzyme which degrades fibrin directly, but not a plasminogen activator which degrades fibrin by activating plasminogen . SW-1 was found also to degrade fibrinogen . In vivo, the thrombolytic effect of SW-1 (4000 u.kg-1) on venous thrombus in rat was most remarkable versus that of control(P < 0.001), but was not divergent versus that of urokinase at the some dose(P > 0.05) . SW-1 was shown to enhance plasminogen and plasmin activity but did not show significant effect on endogenous t-PA or alpha 2-PI . SW-1 also caused decrease of fibrinogen level in plasma, indicating that it degrades the fibrin in thrombus and at the same time degrades the circulating fibrinogen to certain extent . The changes of these biochemical data demonstrated the thrombolytic properties of SW-1 in vivo. Appl Microbiol Biotechnol, 2002 May, 58(6), 761 - 6 Epub 2002 Mar 20. Xylanase production in solid state fermentation by Aspergillus niger mutant using statistical experimental designs; Park YS et al.; The initial moisture content, cultivation time, inoculum size and concentration of basal medium were optimized in solid state fermentation (SSF) for the production of xylanase by an Aspergillus niger mutant using statistical experimental designs . The cultivation time and concentration of basal medium were the most important factors affecting xylanase activity . An inoculum size of 5 x 10(5) spores/g, initial moisture content of 65%, cultivation time of 5 days and 10 times concentration of basal medium containing 50 times concentration of corn steep liquor were optimum for xylanase production in SSF . Under the optimized conditions, the activity and productivity of xylanase obtained after 5 days of fermentation were 5,071 IU/g of rice straw and 14,790 IU l(-1) h(-1), respectively . The xylanase activity predicted by a polynomial model was 5,484 IU/g of rice straw. Appl Microbiol Biotechnol, 2002 May, 58(6), 735 - 42 Epub 2002 Mar 20. A theoretical analysis of the biosynthesis of actinorhodin in a hyper-producing Streptomyces lividansstrain cultivated on various carbon sources; Bruheim P et al.; A stoichiometric equation for the biosynthesis of actinorhodin (ACT) was derived taking into consideration both the requirements of the carbon precursors (acetyl-CoA) and reducing power (NADPH) . The estimate for reducing power was derived from a detailed molecular analysis of each step in the ACT biosynthetic pathway . Even though ACT is slightly more oxidized than most carbon substrates, e.g . glucose, reducing power (NADPH and NADH) is necessary due to reducing steps and to monooxygenase steps . The equation was used to evaluate, in a metabolic network context, the experimental results from batch fermentations with eight different carbon sources using a Streptomyces lividans 1326 derived strain containing the pathway-specific activator gene ( actII-ORF4) on a multicopy plasmid (pIJ68) . The yield of ACT on the various carbon sources ranged from 0.04 to 0.18 Cmol ACT/Cmol carbon source in the stationary phase . Glucose was the best carbon source and supported a yield of 25% of the maximum theoretical yield . There are no obvious constraints in the primary metabolic pathways that can explain why the various carbon sources allowed different levels of ACT production, because their potential for supplying acetyl-CoA and NADPH are far from fully utilized . For the observed ACT yields, there is an excess production of NADPH that has to be reoxidized either by a transhydrogenase or a NADPH oxidase . This study discusses the central metabolic pathways, focusing on providing precursors for ACT synthesis. Comp Biochem Physiol A Mol Integr Physiol, 2002 Jun, 132(2), 333 - 40 Production of short-chain fatty acids and gas from various oligosaccharides by gut microbes of carp (Cyprinus carpio L.) in micro-scale batch culture; Kihara M et al.; We studied the metabolism of various oligosaccharides by carp (Cyprinus carpio) hindgut microbes by measuring gas productivity and organic acid production in gut contents using a 50-microl-scale batch culture system . Carp hindgut contents were incubated with 500 microg each of raffinose, lactosucrose, kestose, lactulose, gentiobiose, 4'-galactosyllactose and 6'-galactosyllactose and soybean-, xylo-, and isomalto-oligosaccharides or none (blank culture) at 25 degrees C for 6 h . The time-course of gas release from the culture (Y microl/culture) was expressed as an exponential function of incubation time (t) {Y=A+Bx(1-e(-kt))}; A, B and k are constants) . Potential production of gas (A+B) from soybean-oligosaccharide and raffinose was larger than for the other saccharides except for kestose, and blank culture . The rate constant of gas (k) for lactosucrose was larger than that for isomalto- and xylo-oligosaccharide, lactulose, kestose or blank culture . Net production of total SCFA (sum of acetic, propionic and n-butyric acid weights) from cultures with soybean- and isomalto-oligosaccharides, raffinose, gentiobiose and lactosucrose was greater than that from blank culture . These results suggested that soybean-oligosaccharide and raffinose were potentially highly fermentable oligosaccharides for carp hindgut microbes . Chemical structures of oligosaccharides seem to play an important role in the fermentability . It is also likely that oligosaccharide utilization differs between mammals and teleosts. J Vet Med A Physiol Pathol Clin Med, 2002 Apr, 49(3), 157 - 60 Studies on the pH value of abomasal contents in dairy cows during the first 3 weeks after calving; Van Winden SC et al.; The pH value of the abomasal contents in adult cattle is normally constant and has a value of 2 . Abomasal contents with pH values of 5.5 and higher could give rise to bacterial fermentation with subsequent gas production . The accumulation of gas is thought to form a key event in the pathogenesis of abomasal displacement . The aim of the present study was to determine the pH values of abomasal contents of dairy cows in the first 3 weeks after calving . The pH of the abomasal contents was, over several days . higher than the pH value of the abomasal juice in mid-lactation cows . The highest pH values were measured on day 14 after calving, thereafter the pH declined . Possible explanations for the rise in the pH value of the abomasal contents are discussed. J Vet Med A Physiol Pathol Clin Med, 2002 Apr, 49(3), 151 - 6 Bile acid extraction rate in the liver of cows fed high-fat diet and lipid profiles in the portal and hepatic veins; Mohamed T et al.; The purposes of this study are to assess the responses of increased supplemental dietary fat in the cow, without upsetting rumen fermentation, on the bile acid (BA) extraction rate in the liver and to determine whether this diet would affect the postprandial lipid profiles in the portal and hepatic venous blood . Six Holstein cows were equipped with catheters fitted in the portal and hepatic veins . Two cows each were assigned randomly to a sequence of three dietary treatments of 21-day period . The methodology of this study was based on the supplementation of the basal concentrate diet with 0 (control), 5, or 10% calcium salts of fatty acids (CSFA) . The total bile acids were significantly increased in the portal and hepatic veins with the 5% CSFA diet, whereas no increase occurred with the 10% CSFA diet . Data obtained in this study showed that 10% CSFA diet failed to stimulate BA secretion to exceed the values obtained with 5% CSFA-diet . Moreover, there was no change in the hepatic extraction rate of BA in animals fed either the 0 or 5% CSFA diets which ranged from 2.4 to 6.5-fold and 3.1 to 7.3-fold, respectively . However, the extraction rate increased sharply with the 10% CSFA diet (27-fold) . The median portal and hepatic concentrations of total lipids, triglycerides, total cholesterol, phospholipids and non-esterified fatty acids did not show any significant increase during feeding of the control diet . Moreover, feeding either the 5 or 10% CSFA diet did not significantly increase these values in either vein. Indian J Exp Biol, 2001 Jul, 39(7), 691 - 6 Gluconic acid production by Aspergillus niger mutant ORS-4.410 in submerged and solid state surface fermentation; Singh OV et al.; Aspergillus niger ORS-4.410, a mutant of Aspergillus niger ORS-4 was produced by repeated irradiation with UV rays . Treatments with chemical mutagnes also resulted into mutant strains . The mutants differed from the parent strain morphologically and in gluconic acid production . The relationship between UV treatment dosage, conidial survival and frequency of mutation showed the maximum frequency of positive mutants (25%) was obtained along with a conidial survival of 59% after second stage of UV irradiation . Comparison of gluconic acid production of the parent and mutant ORS-4.410 strain showed a significant increase in gluconic acid production that was 87% higher than the wild type strain . ORS-4.410 strain when transferred every 15 days and monitored for gluconic acid levels for a total period of ten months appeared stable . Mutant ORS-4.410 at 12% substrate concentration resulted into significantly higher i.e . 85-87 and 94-97% yields of gluconic acid under submerged and solid state surface conditions respectively . Further increase in substrate concentration appeared inhibitory . Maximum yield of gluconic acid was obtained after 6 days under submerged condition and decreased on further cultivation . Solid state surface culture condition on the other hand resulted into higher yield after 12 days of cultivation and similar levels of yields continued thereafter. J Anim Sci, 2002 May, 80(5), 1223 - 32 Isolated canine and murine intestinal cells exhibit a different pattern of fuel utilization for oxidative metabolism; Beaulieu AD et al.; The amount and type of dietary fiber influences the end-products of fermentation and thus fuel availability to intestinal tissue . Metabolic fuel usage was studied in intestinal cells isolated from dogs consuming a commercial diet or from rats consuming either a commercial rat diet or dog diet to examine preferential fuel usage, the effect of diet, and species differences . Production of 14CO2 was measured by incubating cells in media containing either D-{U-14C}glucose, {1-14C}n-butyrate, L-{U-14C}glutamine, or {1-14C}propionate with or without competing substrates . The presence of a mixture of 5 mM each of glucose, butyrate, propionate, and acetate and 1 mM glutamine in the media decreased CO2 production from glucose, glutamine, and propionate by canine enterocytes (P < 0.05) and from glutamine and propionate by canine colonocytes (P < 0.05) . The presence of glutamine in the media decreased glucose oxidation by murine enterocytes, regardless of the diet . Similarly, glutamine decreased glucose oxidation by murine colonocytes (P < 0.05), but only when the rats had consumed the rat diet . Regardless of diet, murine colonocytes oxidized more butyrate (P < 0.01) than did enterocytes, and murine enterocytes tended (P < 0.07) to oxidize more glucose than did colonocytes . The proportion of propionate in colonic contents was higher in dogs than in rats (P < 0.02), and the proportion of butyrate tended to be higher in contents from rats than in those from dogs (P < 0.08) . Colonic and cecal wet weights were decreased (P < 0.05) when rats were fed the commercial dog diet . Preferred utilization of substrates by isolated canine enterocytes and colonocytes differed from that of murine intestinal cells . These differences were only partially overcome by feeding the same diet to each species. J Anim Sci, 2002 May, 80(5), 1179 - 86 Fermented wheat in liquid diets: effects on gastrointestinal characteristics in weanling piglets; Scholten RH et al.; Effects of adding fermented wheat to liquid diets on gastrointestinal characteristics in weanling piglets were studied . Gastrointestinal characteristics of 40 28-d-old weanling piglets were measured at the day of weaning (d 0) and at d 4 and 8 after weaning . Piglets were group-housed and fed twice daily . Feeding level was based on the average metabolic BW of piglets per group . Groups were fed a liquid diet with either 45% unfermented wheat (FERM_0) or 45% fermented wheat (FERM_45) . The other 55% of the diet was identical . To ferment wheat, water at 30 degrees C was added to milled wheat in a 1:2.2 ratio . The pH and contents of lactic acid, acetic acid, propionic acid, and butyric acid were measured in the digesta of the stomach, three parts of the small intestine, cecum, and large intestine . In addition, changes in microbial populations in the digesta were studied during the period after weaning . Moreover, villus height, crypt depth, and villus shape were studied in the small intestine . Piglets fed FERM_45 showed lower gastric pH (P < 0.05) at d 4 and higher gastric lactic acid content (P < 0.001) at both d 4 and 8 . Piglets fed FERM_45 showed in the first part of the small intestine higher villus height (P < 0.01) at d 8 and higher villus:crypt ratio (P < 0.001) at both d 4 and 8 . Villus shape tended (P < 0.10) to be favorable for piglets fed FERM_45 . The present study indicates that feeding a partly fermented liquid diet to weanling piglets may be a concept to prevent undesirable changes in mucosal architecture after weaning. J Antibiot (Tokyo), 2002 Mar, 55(3), 249 - 55 Watasemycins A and B, new antibiotics produced by Streptomyces sp . TP-A0597; Sasaki O et al.; Two novel antibiotics, watasemycins A and B, were isolated from the fermentation broth of an actinomycete strain . The producing strain TP-A0597 was isolated from the seawater sample collected in Toyama Bay, Japan and identified as Streptomyces sp . based on the taxonomic study . The new antibiotics were obtained by solvent extraction and chromatographic purification and spectroscopic analyses identified that they were new analogs of thiazostatins . Watasemycin possesses a methyl group at 5'-position of thiazostatin instead of a hydrogen atom . Watasemycins showed antibiotic activity against Gram-positive and negative bacteria and yeast. J Antibiot (Tokyo), 2002 Mar, 55(3), 239 - 48 Novel stachyflin derivatives from Stachybotrys sp . RF-7260 . Fermentation, isolation, structure elucidation and biological activities; Minagawa K et al.; Stachybotrys sp . RF-7260 was found to produce stachyflins, novel anti-influenza virus agents, under solid-state fermentation conditions . Feeding DL-lysine to a culture of Stachybotrys sp . RF-7260 induced the formation of the novel compounds, SQ-02-S-L2 and -L1, and feeding DL-valine the formation of SQ-02-S-VI and -V2 . The structures of these metabolites were determined by detailed 2D NMR analyses in comparison with acetylstachyflin . SQ-02-S-L2 and -L1 have the lysine moiety and SQ-02-S-V1 has the valine moiety . SQ-02-S-V2 has an amidine moiety instead of the lactam moiety in acetylstachyflin . SQ-02-S-L2, -L1 and -V1, substituted on the lactam amide hydrogen, displayed only a low level of the antiviral activity . However, deacetyl SQ-02-S-V2 showed potent antiviral activity similar to stachyflin. Extremophiles, 2002 Apr, 6(2), 161 - 6 Effect of glucose, maltose, soluble starch, and CO2 on the growth of the hyperthermophilic archaeon Pyrococcus furiosus; Biller KF et al.; The hyperthermophilic archaeon Pyrococcus furiosus was cultivated in batch and continuous fermentations on different carbon substrates . The cultivation of P furiosus on soluble starch as the only carbon source resulted in cell densities three times higher than in cultivations on maltose, 1.06 x 10(10) cells/ml compared to 3.4 x 10(9) cells/ml . The yield coefficient, Y(x/y) = 0.12 g/g, and the growth rate, mu = 0.33 h(-1), were almost equal on soluble starch and on maltose, but on glucose no growth could be detected . An inhibitory effect of glucose, when added to other carbon substrates, also could not be found . Isobutyric and isovaleric acid were detected as novel metabolites produced by P . furiosus . Inhibitory effects of these acids, as well as of the well-known products acetic acid, propionic acid, and alanine, could be precluded . Concentrations of 10% CO2 in the gas supply respective to the exhaust gas enhanced the growth of P furiosus significantly . The maximum cell number was two orders of magnitude higher than was observed with pure nitrogen . Further increase of the CO2 concentration up to 100% had no significant effect on the growth of P . furiosus. J Chromatogr B Analyt Technol Biomed Life Sci, 2002 Apr 25, 770(1-2), 255 - 9 A simple, optimized method for the determination of sulphide in whole blood by GC-mS as a marker of bowel fermentation processes; Hyspler R et al.; Hydrogen sulphide is produced in human large intestine by anaerobic fermentation and may play a pathogenic role . An analytical method for determination of sulphide in whole blood using an extractive alkylation technique was optimised and validated for this purpose . The sample was mixed with organic phase containing pentafluorobenzyl bromide as an alkylating agent . The benzalkonium chloride was used as a phase-transfer catalyst . The quantitative determination was performed using GC-MS technique in selected ion monitoring mode . The blood levels of sulphide of healthy controls were measured (35-80 microM/l) . The method is versatile, reproducible (RSD=2.7%) and suitable for research of anaerobic fermentation in vivo. Infect Immun, 2002 Jun, 70(6), 3101 - 10 Purification, characterization, and immunogenicity of the refolded ectodomain of the Plasmodium falciparum apical membrane antigen 1 expressed in Escherichia coli; Dutta S et al.; The apical membrane antigen 1 (AMA1) has emerged as a promising vaccine candidate against malaria . Advanced evaluation of its protective efficacy in humans requires the production of highly purified and correctly folded protein . We describe here a process for the expression, fermentation, refolding, and purification of the recombinant ectodomain of AMA1 (amino acids 83(Gly) to 531(Glu)) of Plasmodium falciparum (3D7) produced in Escherichia coli . A synthetic gene containing an E . coli codon bias was cloned into a modified pET32 plasmid, and the recombinant protein was produced by using a redox-modified E . coli strain, Origami (DE3) . A purification process was developed that included Sarkosyl extraction followed by affinity purification on a Ni-nitrilotriacetic acid column . The recombinant AMA1 was refolded in the presence of reduced and oxidized glutathione and further purified by using two ion-exchange chromatographic steps . The final product, designated AMA1/E, was homogeneous, monomeric, and >99% pure and had low endotoxin content and low host cell contamination . Analysis of AMA1/E showed that it had the predicted primary sequence, and tertiary structure analysis confirmed its compact disulfide-bonded nature . Rabbit antibodies made to the protein recognized the native parasite AMA1 and inhibited the growth of the P . falciparum homologous 3D7 clone in an in vitro assay . Reduction-sensitive epitopes on AMA1/E were shown to be necessary for the production of inhibitory anti-AMA1 antibodies . AMA1/E was recognized by a conformation-dependent, growth-inhibitory monoclonal antibody, 4G2dc1 . The process described here was successfully scaled up to produce AMA1/E protein under GMP conditions, and the product was found to induce highly inhibitory antibodies in rabbits. Br J Nutr, 2002 May, 87(5), 421 - 6 A modified dot-blot method of protein determination applied in the tannin-protein precipitation assay to facilitate the evaluation of tannin activity in animal feeds; Hoffmann EM et al.; Tannins have received considerable attention from animal nutritionists as potential agents for modifying ruminal fermentation patterns, or for exploring new feed resources . This group of secondary plant compounds is defined by their ability to form complexes with proteins . A widely accepted method for assaying the biological activity of extracted tannins is the precipitation of bovine serum albumin . The protein carries a radioactive label (125I) to allow direct quantification from the precipitate . Tannin-protein complexes dissolve in sodium dodecylsulfate solution . A dot-blot assay for protein determination, which is based on the reversible binding of a fluorochrome, benzoxanthene yellow, to the protein spots and is not disturbed by the presence of detergents, can replace the radioactive method by a fluorimetric measurement . A novel alternative to the last part of the dot-blot assay is to scan the stained protein spots in situ using a video camera and computer image analysis . Several filter sets were tested and, within a concentration range of 0.1-2.0 mg protein/ml, each of them yielded results identical to the original method while the time required was only 30 % of the working time consumed by the original procedure . The modified dot-blot assay should be applicable to the evaluation of tannin activity in all shrub and tree foliages considered as animal feed. Physiol Plant, 2002 May, 115(1), 119 - 124 Ethanol sensitivity of rice and oat coleoptiles; Kato-Noguchi H; The ability to avoid the ethanol-induced injury was evaluated in rice (Oryza sativa L.) and oat (Avena sativa L.) coleoptiles . The growth of the rice and oat coleoptiles was inhibited by ethanol exogenously applied at concentrations greater than 200 and 30 mM, respectively . At 300 mM ethanol, oat coleoptiles were brown and flaccid but rice coleoptiles did not show any visible symptoms of toxicity . The acetaldehyde level in rice and oat coleoptiles was increased by exogenously applied ethanol and the increases were greater in oat than in rice coleoptiles under aerobic and anaerobic conditions . At 300 mM ethanol, the acetaldehyde concentrations in the rice and oat coleoptiles were 46 and 87 nmol g-1 FW under aerobic conditions, respectively, and 52 and 124 nmol g-1 FW under anaerobic conditions, respectively . The activity of alcohol dehydrogenase (ADH; EC 1.1.1.1) in the direction of ethanol to acetaldehyde was greater in oat than in rice coleoptiles and ADH protein in oat coleoptiles was more induced by exogenously applied ethanol than that in rice coleoptiles . These results suggest that in vivo conversion rate of ethanol to acetaldehyde by ADH is lower in rice than oat coleoptiles, which may be one of the reasons that ethanol sensitivity of rice is much lower than that of oat coleoptiles . The great ability of rice to avoid the ethanol-induced injuries may contribute its anoxia tolerance when glycolysis and ethanolic fermentation replace the Krebs cycle as the main source of energy under anaerobic conditions. J Agric Food Chem, 2002 May 22, 50(11), 3277 - 82 Impact odorants contributing to the fungus type aroma from grape berries contaminated by powdery mildew (Uncinula necator); incidence of enzymatic activities of the yeast Saccharomyces cerevisiae; Darriet P et al.; Powdery mildew due to the fungus Uncinula necator is an important disease for the vineyard . The development of the fungus at the surface of the berries leads to the occurrence of a very characteristic and sometimes intense mushroom-type odor cited as an important default for grapes quality . Gas chromatography/olfactometry, gas chromatography, and multidimensional gas chromatogaphy/mass spectrometry techniques were used to investigate the most important odorants of grapes diseased by powdery mildew . Among 22 odorants detected, strongly odorant compounds were identified or tentatively identified in purified extracts obtained from grapes diseased by powdery mildew . Aroma extraction dilution analysis (AEDA) analysis revealed that 1-octen-3-one (mushroom odor), (Z)-1,5-octadien-3-one (geranium-leaf odor), and an unidentified odorous zone (fishy-mushroom like odor) were the most potent volatiles of the diseased grapes . In the presence of nonproliferating Saccharomyces cerevisiae yeast cells, and consequently during alcoholic fermentation, the enzymatic reduction of 1-octen-3-one and (Z)-1,5-octadien-3-one to much less odorant compounds, namely 3-octanone and (Z)-5-octen-3-one, was shown . Those results explain to some extent the disappearance of the fungal aroma specific to powdery mildew grapes during alcoholic fermentation. J Agric Food Chem, 2002 May 22, 50(11), 3186 - 92 Ellagic acid and flavonoid antioxidant content of muscadine wine and juice; Talcott ST et al.; Antioxidant properties of flavonoids and ellagic acid were characterized in eight wines and juices produced by various processing methodologies from red and white muscadine grape cultivars (Vitis rotundifolia) . Juices and wines were produced by hot- and cold-pressed techniques, and additional wine was produced following on-hull fermentation for 3, 5, and 7 days . Chromatographic conditions were developed to simultaneously separate anthocyanins, ellagic acid, and flavonols and correlated to a measurement of overall antioxidant capacity (AOX), and their changes were monitored after storage for 60 days at 20 and 37 degrees C . Regression coefficients between concentrations of individual polyphenolics and AOX ranged from 0.55 for ellagic acid to 0.90 for kaempferol . Both red and white wines had higher AOX values after storage than juices made from an identical grape press, despite lower concentrations of individual polyphenolic compounds . Red wines fermented on-hull had higher initial concentrations of antioxidant polyphenolics as compared to a corresponding hot-pressed juice, but changes in AOX during storage were more affected by time than by storage temperature despite lower concentrations of flavonoids and ellagic acid present at 37 degrees C as compared to 20 degrees C . Oxidative or polymerization reactions significantly decreased levels of monomeric anthocyanins during storage with the greatest losses observed for delphinidin and petunidin 3,5-diglucosides . Processing methods for muscadine wine and juice production were important factors influencing concentrations of antioxidant flavonoids and ellagic acid, while the role of fermentation and time had the greatest influence on retention of AOX properties during storage. Metab Eng, 2002 Apr, 4(2), 182 - 92 Metabolic engineering through cofactor manipulation and its effects on metabolic flux redistribution in Escherichia coli; San KY et al.; Applications of genetic engineering or metabolic engineering have increased in both academic and industrial institutions . Most current metabolic engineering studies have focused on enzyme levels and on the effect of the amplification, addition, or deletion of a particular pathway . Although it is generally known that cofactors play a major role in the production of different fermentation products, their role has not been thoroughly and systematically studied . It is conceivable that in cofactor-dependent production systems, cofactor availability and the proportion of cofactor in the active form may play an important role in dictating the overall process yield . Hence, the manipulation of these cofactor levels may be crucial in order to further increase production . We have demonstrated that manipulation of cofactors can be achieved by external and genetic means and these manipulations have the potential to be used as an additional tool to achieve desired metabolic goals . We have shown experimentally that the NADH/NAD(+) ratio can be altered by using carbon sources with different oxidation states . We have shown further that the metabolite distribution can be influenced by a change in the NADH/NAD(+) ratio as mediated by the oxidation state of the carbon source used . We have also demonstrated that the total NAD(H/(+)) levels can be increased by the overexpression of the pncB gene . The increase in the total NAD(H/(+)) levels can be achieved even in a complex medium, which is commonly used by most industrial processes . Finally, we have shown that manipulation of the CoA pool/flux can be used to increase the productivity of a model product, isoamyl acetate. Bioelectrochemistry, 2002 May 15, 56(1-2), 127 - 9 Monitoring of ethanol during fermentation using a microbial biosensor with enhanced selectivity; Tkac J et al.; The present study is concerning the construction of ferricyanide-mediated Gluconobacter oxydans cell ethanol biosensor . The size exclusion effect of a cellulose acetate membrane was used for elimination of glucose interferences during ethanol assays in real samples . A typical response time of the biosensor was 13 s with a high sensitivity of 3.5 microA mM(-1) . The microbial biosensor exhibits a very low detection limit of 0.85 microM and a wide linear range from 2 to 270 microM . The operational stability was excellent . During 8.5 h of repetitive ethanol assays, no decrease in the sensor sensitivity was observed . The biosensor was successfully used in the off-line monitoring of ethanol fermentation with a good agreement with HPLC measurements (R(2)=0.998). Int J Biochem Cell Biol, 2002 Aug, 34(8), 901 - 6 A lipoprotein family from Mycoplasma fermentans confers host immune activation through Toll-like receptor 2; Seya T et al.; Mycoplasma have been reported to be associated with human diseases . Three forms of a mycoplasma lipopeptide/protein with the ability to modulate the host immune system were independently identified and named macrophage-activating lipopeptide 2 (MALP-2), P48 and M161Ag (identical to MALP-404) . Although these molecules had polypeptides of different sizes, they exerted similar immunomodulatory effects on macrophages/dendritic cells, such as cytokine induction, NO production and maturation of antigen-presenting cells (APCs) . M161Ag exhibited complement-activating ability and bound macrophages via complement C3b/C3bi and their receptors . The diacylated N-terminal palmitates were involved in these activities . Toll-like receptor 2 (TLR2) was found to be responsible for these functional features of these mycoplasma products, except for complement activation . Here, we summarize the functional properties of this family of proteins, namely pathogen-associated molecular pattern (PAMP) and discuss its relationship to the reported pathogenesis of latent mycoplasma infection. Trends Biotechnol, 2002 Jun, 20(6), 257 - 60 Impact of targeted vector design on Co/E1 plasmid replication; Grabherr R et al.; The demands for recombinant proteins, in addition to plasmid DNA, for therapeutic use are steadily increasing . Bacterial fermentation processes have long been and still are the major tool for production of these molecules . The key objective of process optimization is to attain a high yield of the required quality, which is determined, to a large extent, by plasmid replication rates, metabolic capacity and the properties of the specific gene construct . When high copy number plasmids are used, the metabolic capacity of the host cell is often overstrained and efficient protein production is impaired . The plasmid copy number is the key parameter in the exploitation of the host cell, and can be maximized by optimal control of the flux ratios between biosynthesis of host cell proteins and recombinant proteins. Bioresour Technol, 2002 Apr, 82(2), 171 - 6 Effect of temperature, anaerobiosis, stirring and salt addition on natural fermentation silage of sardine and sardine wastes in sugarcane molasses; Zahar M et al.; Conditions for a natural fermentation during ensilage of sardines or their waste in sugarcane molasses (60:40 w/w) were evaluated regarding the effect of temperature (15, 25 and 35 degrees C), anaerobiosis (closed vs . open jars), daily stirring of the mixture, and salt addition to the initial mix at 5% (w/w) level . Successful natural fermentation took place in sardine silages incubated at 25 or 35 degrees C in open jars to reach a pH of 4.4 in about 2 and 1 weeks, respectively . For samples kept at 15 degrees C, the pH decline was very slow and pH did not decrease below 5.5 after one month of incubation . At 25 degrees C, the most favorable conditions for silage of sardine waste in cane molasses, as evidenced by the fastest decline in pH to a stable value of about 4.4, were achieved in closed jars and with daily stirring of the mix . The pH 4.4 was reached in one week with an advance of at least 3 days compared to the other conditions (open jars and closed jars without daily stirring) . Addition of salt at 5% (w/w) in the mix before incubation inhibited the fermentation process. Bioresour Technol, 2002 Apr, 82(2), 131 - 7 Bio-degradation of lignin in olive pomace by freshly-isolated species of Basidiomycete; Haddadin MS et al.; The solid waste (pomace) from olive oil processing cannot be used directly as an animal feed, but it was thought that an appropriate series of fermentations might improve its nutritional value . As a first step, typical samples of pomace were subjected, after an alkaline pre-treatment, to delignification by Phanerochaete chrysosporium (ATCC 19343), Oxysporus sp., Schizophyllum commune, Hyphoderma sp . or Ganoderma sp.; the last four species being freshly isolated from decaying wood collected in a woodland in Jordan . The relative activity of the species was judged by the levels of ligninase or laccase secreted and the extent of lignin degradation under a range of experimental conditions . Oxysporus sp . (ca . 69%) and S . commune (ca . 53%) gave significantly higher levels of breakdown of the lignified material than the other recent isolates . P . chrysosporium (ca . 60%) was not as active as in previously reported studies, and it may be that culturing the fungus on a standard laboratory medium had reduced its ability to generate ligninase . Further work is needed to establish whether the delignified pomace could be further processed into a feed for poultry. J Antibiot (Tokyo), 2002 Feb, 55(2), 155 - 64 Stachyflin and acetylstachyflin, novel anti-influenza A virus substances, produced by Stachybotrys sp . RF-7260 . I . Isolation, structure elucidation and biological activities; Minagawa K et al.; Two novel compounds, stachyflin and acetylstachyflin, have been isolated by solid-state fermentation of Stachybotrys sp . RF-7260 . The structures of both metabolites, determined by detailed NMR analyses and X-ray crystallographic analysis, are novel with a pentacyclic moiety including cis-fused decalin . The absolute stereochemistry of stachyflins was determined by circular dichroism analysis . Stachyflin showed antiviral activity against influenza A virus (H1N1) in vitro with an IC50 value of 0.003 microM . Acetylstachyflin was about 77-fold less active than stachyflin. J Antibiot (Tokyo), 2002 Feb, 55(2), 141 - 6 Migrastatin and a new compound, isomigrastatin, from Streptomyces platensis; Woo EJ et al.; Streptomyces platensis (strain NRRL 18993), a producer of dorrigocins, was shown to produce migrastatin, a cyclic congener of dorrigocin A previously reported from a different organism . Additionally a new compound isomeric to migrastatin, isomigrastatin, was also isolated and its structure was determined to be a cyclic form of dorrigocin B . Both compounds were fully characterized from MS and NMR data . Product titers of both were improved by the addition of XAD-16 resin to the fermentation medium. J Antibiot (Tokyo), 2002 Feb, 55(2), 121 - 7 CJ-21,164, a new D-glucose-6-phosphate phosphohydrolase inhibitor produced by a fungus Chloridium sp; Kim YJ et al.; A new D-glucose-6-phosphate phosphohydrolase (G6Pase) inhibitor, CJ-21,164 (1) was isolated from the fermentation broth of the fungus Chloridium sp . CL48903 . The structure was elucidated to be a novel tetramer of the salicylic acid derivatives by spectroscopic analyses . Compound I inhibited G6Pase in rat liver microsomes with an IC50 of 1.6 microM . Glucose output from hepatocytes isolated from rat liver was inhibited when I was present in the incubation medium, consistent with the role of I as a G6Pase inhibitor. Breast Cancer Res Treat, 2002 Feb, 71(3), 203 - 17 Chemopreventive and adjuvant therapeutic potential of pomegranate (Punica granatum) for human breast cancer; Kim ND et al.; Fresh organically grown pomegranates (Punica granatum L.) of the Wonderful cultivar were processed into three components: fermented juice, aqueous pericarp extract and cold-pressed or supercritical CO2-extracted seed oil . Exposure to additional solvents yielded polyphenol-rich fractions ('polyphenols') from each of the three components . Their actions, and of the crude whole oil and crude fermented and unfermented juice concentrate, were assessed in vitro for possible chemopreventive or adjuvant therapeutic potential in human breast cancer . The ability to effect a blockade of endogenous active estrogen biosynthesis was shown by polyphenols from fermented juice, pericarp, and oil, which inhibited aromatase activity by 60-80% . Fermented juice and pericarp polyphenols, and whole seed oil, inhibited 17-beta-hydroxysteroid dehydrogenase Type 1 from 34 to 79%, at concentrations ranging from 100 to 1,000 microg/ml according to seed oil >> fermented juice polyphenols > pericarp polyphenols . In a yeast estrogen screen (YES) lyophilized fresh pomegranate juice effected a 55% inhibition of the estrogenic activity of 17-beta-estradiol; whereas the lyophilized juice by itself displayed only minimal estrogenic action . Inhibition of cell lines by fermented juice and pericarp polyphenols was according to estrogen-dependent (MCF-7) >> estrogen-independent (MB-MDA-231) > normal human breast epithelial cells (MCF-10A) . In both MCF-7 and MB-MDA-231 cells, fermented pomegranate juice polyphenols consistently showed about twice the anti-proliferative effect as fresh pomegranate juice polyphenols . Pomegranate seed oil effected 90% inhibition of proliferation of MCF-7 at 100 microg/ml medium, 75% inhibition of invasion of MCF-7 across a Matrigel membrane at 10 microg/ml, and 54% apoptosis in MDA-MB-435 estrogen receptor negative metastatic human breast cancer cells at 50 microg/ml . In a murine mammary gland organ culture, fermented juice polyphenols effected 47% inhibition of cancerous lesion formation induced by the carcinogen 7,12-dimethylbenz{a}anthracene (DMBA) . The findings suggest that clinical trials to further assess chemopreventive and adjuvant therapeutic applications of pomegranate in human breast cancer may be warranted. J Anim Sci, 2002 Apr, 80(4), 1135 - 43 Effect of extruded full-fat soybeans on conjugated linoleic acid content of intramuscular, intermuscular, and subcutaneous fat in beef steers; Madron MS et al.; Crossbred Angus steers (n = 30) were used to determine whether the conjugated linoleic acid (CLA) content of beef fat could be increased by feeding varying levels of extruded full-fat soybeans as a source of polyunsaturated fatty acids for rumen biohydrogenation . Diets were as follows: 1) control, 2) 12.7% extruded full-fat soybeans (LESB), and 3) 25.6% extruded full-fat soybeans (HE SB) . Steers were individually housed and fed the diets for 111 d during the finishing period . Over the experimental period, treatment groups were similar in ADG (1.7 +/- 0.1 kg/d) and had a similar slaughter weight (603 +/- 11.6 kg) . Dressing percentage averaged 61.6% and carcass composition averaged 14.3% protein, 30.9% lipid, and 54.8% water . At slaughter, the intramuscular, intermuscular, and subcutaneous fat depots were sampled from the rib longissimus, eye of round, and chuck tender muscles . Across all fat depots, the CLA content differed (P < 0.05), averaging 6.6, 6.7, and 7.7 mg/g of fatty acids for the control, LESB, and HESB diets, respectively . There were significant differences in CLA content between fat depots within a cut, but differences were relatively small and the hierarchy in fat depots was not consistent among cuts . The cis-9, trans-11 isomer was the predominant CLA isomer and its content in fat was related to trans-11 C18:1 content (r = 0.53; P < 0.001) . There was substantial individual variation in CLA content and this varied from 2.6 to 17.0 mg/g fatty acids across all treatments and fat depots . Overall, results demonstrated that including extruded full-fat soybeans in the diet of finishing steers increased the CLA content of beef fat . Differences were relatively small and the relationship of this to rumen fermentation and endogenous synthesis of CLA is considered. J Anim Sci, 2002 Apr, 80(4), 1117 - 23 Carbohydrate fermentation and nitrogen metabolism of a finishing beef diet by ruminal microbes in continuous cultures as affected by ethoxyquin and(or) supplementation of monensin and tylosin; Han H et al.; Long-term feedlot studies have shown positive effects (i.e., improved ADG and reduced morbidity and mortality) of dietary supplementation with ethoxyquin (AGRADO) . This may be due to improving the antioxidant capacity at the ruminal, postruminal, or postabsorption levels . This study was designed to investigate the role of ethoxyquin at the rumen level . A finishing diet (12.5% CP; DM basis) was formulated to contain (on a DM basis) 77.5% flaked corn, 10% corn cobs, 10% protein/vitamin/mineral supplement, and 2.5% tallow . In a randomized complete block design experiment, the treatments were arranged as a 2 x 2 factorial . The main factors were two ethoxyquin treatments (without or with 150 ppm) and two monensin/tylosin treatments (without or with monensin and tylosin at 0.0028 and 0.0014% of dietary DM, respectively) . Eight dual-flow, continuous culture fermenters were used in two experimental periods (blocks; 8 d each with 5 d for adjustment and 3 d for sample collection) to allow for four replications for each treatment . No interactions (P > 0.05) were detected for any of the measurements evaluated . Therefore, results of the main factors were summarized . Ethoxyquin supplementation improved (P < 0.05) true digestibility of OM (from 38.8 to 45.0%) but it did not alter (P > 0.05) concentrations of total VFA (averaging 131 mM) or acetate (averaging 58.8 mM) . Ethoxyquin decreased (P < 0.05) propionate concentration from 51.1 to 42.4 mM and increased (P < 0.05) butyrate concentration from 18.4 to 22.9 mM . Digestion of total nonstructural carbohydrates was not altered (P > 0.05) by the treatments and averaged 86% . With the exception of increased (P < 0.05) concentration of propionate (from 42.0 to 51.5 mM) and decreased (P < 0.05) concentration of butyrate (from 25.9 to 16.3 mM), no effects (P > 0.05) were detected for monensin/tylosin . Ruminal N metabolism, including efficiency of bacterial protein synthesis (averaging 21.2 g N/kg OM truly digested), was not affected (P > 0.05) by the treatments . Results suggest positive effects of ethoxyquin on ruminal digestion of OM and unique changes in VFA production. J Anim Sci, 2002 Apr, 80(4), 1105 - 11 Utilization of distillers grains from the fermentation of sorghum or corn in diets for finishing beef and lactating dairy cattle; Al-Suwaiegh S et al.; Beef finishing and dairy lactation experiments were conducted to evaluate the nutritional value of distillers grains (DG) from sorghum or corn fermentation, in both wet (35.4% DM) and dry (92.2% DM) form (dairy trial only) . In the finishing experiment, 60 yearling steers were used in a completely randomized design with three diets that were fed for 127 d: 1) control diet with 86% (DM basis) dry-rolled corn and no DG; 2) 30% of ration DM as wet corn DG in place of dry-rolled corn; and 3) 30% of ration DM as wet sorghum DG in place of dry-rolled corn . All diets contained a minimum of 6.8% degradable intake protein and 13.0% CP . Steers fed DG had 10% greater ADG (< 0.01) and 8% greater efficiency of gain (P < 0.01) than steers fed the control diet . Wet corn and sorghum DG resulted in similar ADG and efficiency of gain . Hot carcass weights, fat thickness, and yield grades were greater for steers fed DG than for controls (P < 0.07) . Improvements in ADG and feed efficiency observed when DG replaced dry-rolled corn indicated that the NEg content of wet DG is approximately 29% greater than that of dry-rolled corn . In the dairy lactation experiment, 16 lactating Holstein cows (eight multiparous, including four fistulated) were used in a replicated 4 x 4 Latin square design with 4-wk periods . Corn and sorghum DG were fed at 15% of the ration DM in either wet or dry form . Diets were fed as total mixed rations that contained 50% of a 1:1 mixture of alfalfa and corn silages, 24.3% ground corn, and 9.1% soybean meal (DM basis) . There was no effect of source or form of DG on DMI, ruminal pH and VFA, or in situ digestion kinetics of NDF from DG . Efficiency of milk production was unaffected by diet . Corn and sorghum DG resulted in relatively similar performance when fed to beef or dairy cattle in this study. Plant J, 2002 May, 30(3), 329 - 36 The ethanolic fermentation pathway supports respiration and lipid biosynthesis in tobacco pollen; Mellema S et al.; Rapid pollen tube growth requires a high rate of sugar metabolism to meet energetic and biosynthetic demands . Previous work on pollen sugar metabolism showed that tobacco pollen carry out efficient ethanolic fermentation concomitantly with a high rate of respiration (Bucher et al., 1995) . Here we show that the products of fermentation, acetaldehyde and ethanol, are further metabolised in a pathway that bypasses mitochondrial PDH . The enzymes involved in this pathway are pyruvate decarboxylase, aldehyde dehydrogenase and acetyl-CoA synthetase . Radiolabelling experiments show that during tobacco pollen tube growth label of 14C-ethanol is incorporated into CO2 as well as into lipids and other higher molecular weight compounds . A role for the glyoxylate cycle appears unlikely since activity of malate synthase, a key enzyme of the glyoxylate cycle, could not be detected. Biosci Biotechnol Biochem, 2002 Feb, 66(2), 378 - 84 Transepithelial transport of the bioactive tripeptide, Val-Pro-Pro, in human intestinal Caco-2 cell monolayers; Satake M et al.; Some of the food-derived tripeptides with angiotensin converting enzyme (ACE)-inhibitory activity have been reported to be hypotensive after being orally administered . The mechanism for the intestinal transport of these tripeptides was studied by using monolayer-cultured human intestinal Caco-2 cells which express many enterocyte-like functions including the peptide transporter (PepT1)-mediated transport system . Val-Pro-Pro, an ACE-inhibitory peptide from fermented milk, was used as a model tripeptide . A significant amount of intact Val-Pro-Pro was transported across the Caco-2 cell monolayer . This transport was hardly inhibited by a competitive substrate for PepT1 . Since no intact Val-Pro-Pro was detected in the cells, Val-Pro-Pro apically taken by Caco-2 cells via PepT1 was likely to have been quickly hydrolyzed by intracellular peptidases, producing free Val and Pro . These findings suggest that PepT1-mediated transport was not involved in the transepithelial transport of intact Val-Pro-Pro . Paracellular diffusion is suggested to have been the main mechanism for the transport of intact Val-Pro-Pro across the Caco-2 cell monolayer. Biosci Biotechnol Biochem, 2002 Feb, 66(2), 307 - 18 L-erythrulose production by oxidative fermentation is catalyzed by PQQ-containing membrane-bound dehydrogenase; Moonmangmee D et al.; Thermotolerant Gluconobacter frateurii CHM 43 was selected for L-erythrulose production from mesoerythritol at higher temperatures . Growing cells and the membrane fraction of the strain rapidly oxidized mesoerythritol to L-erythrulose irreversibly with almost 100% of recovery at 37 degrees C . L-Erythrulose was also produced efficiently by the resting cells at 37 degrees C with 85% recovery . The enzyme responsible for mesoerythritol oxidation was found to be located in the cytoplasmic membrane of the organism . The EDTA-resolved enzyme required PQQ and Ca2+ for L-erythrulose formation, suggesting that the enzyme catalyzing meso-erythritol oxidation was a quinoprotein . Quinoprotein membrane-bound mesoerythritol dehydrogenase (QMEDH) was solubilized and purified to homogeneity . The purified enzyme showed a single band in SDS-PAGE of which the molecular mass corresponded to 80 kDa . The optimum pH of QMEDH was found at pH 5.0 . The Michaelis constant of the enzyme was found to be 25 mM for meso-erythritol as the substrate . QMEDH showed a broad substrate specificity toward C3-C6 sugar alcohols in which the erythro form of two hydroxy groups existed adjacent to a primary alcohol group . On the other hand, the cytosolic NAD-denpendent meso-erythritol dehydrogenase (CMEDH) of the same organism was purified to a crystalline state . CMEDH showed a molecular mass of 60 kDa composed of two identical subunits, and an apparent sedimentation constant was 3.6 s . CMEDH catalyzed oxidoreduction between mesoerythritol and L-erythrulose . The oxidation reaction was observed to be reversible in the presence of NAD at alkaline pHs such as 9.0-10.5 . L-Erythrulose reduction was found at pH 6.0 with NADH as coenzyme . Judging from the catalytic properties, the NAD-dependent enzyme in the cytosolic fraction was regarded as a typical pentitol dehydrogenase of NAD-dependent and the enzyme was independent of the oxidative fermentation of L-erythrulose production. Int J Food Microbiol, 2002 May 5, 75(1-2), 27 - 38 Protein expression during lag phase and growth initiation in Saccharomyces cerevisiae; Brejning J et al.; In order to obtain a better understanding of the biochemical events taking place in Saccharomyces cerevisiae during the lag phase, the proteins expressed during the first hours after inoculation were investigated by two-dimensional (2-D) gel electrophoresis and compared to those expressed in late respiratory growth phase . The studies were performed on a haploid strain (S288C) grown in defined minimal medium . Some of the abundant proteins, whose expression relative to total protein expression was induced during the lag phase, were identified by MALDI MS, and the expression of the corresponding genes was assessed by Northern blotting . The rate of protein synthesis was found to increase strongly during the lag phase and the number of spots detected on 2-D gels increased from 502 spots just after inoculation to 1533 spots at the end of the lag phase . During the first 20 min, the number of detectable spots was considerably reduced compared to the number of spots detected from the yeast in respiratory growth just prior to harvest and inoculation (747 spots), indicating an immediate pausing or shutdown in synthesis of many proteins just after inoculation . In this period, the cells got rid of most of their buds . The MALDI MS-identified, lag phase-induced proteins were adenosine kinase (Ado1p), whose cellular role is presently uncertain, cytosolic acetaldehyde dehydrogenase (Ald6p) and (DL)-glycerol-3-phosphatase 1, both involved in carbohydrate metabolism, a ribosomal protein (Asc1p), a fragment of the 70-kDa heat shock protein Ssb1, and translationally controlled tumour protein homologue (Yk1056cp), all involved in translation, and S-adenosylmethionine synthetase I involved in biosynthesis reactions . The level of mRNA of the corresponding genes was found to increase strongly after inoculation . By pattern matching using previously published 2-D maps of yeast proteins, several other lag phase-induced proteins were identified . These were also proteins involved in carbohydrate metabolism, translation, and biosynthesis reactions . The identified proteins together with other, yet unidentified, lag phase-induced proteins are expected to be important for yeast growth initiation and could be valuable biological markers for yeast performance . Such markers would be highly beneficial in the control and optimisation of industrial fermentations. Steroids, 2002 Jun, 67(7), 549 - 54 Two new C-nor/D-homo corticosteroids formed by dehydration rearrangement of a 12beta-hydroxy corticoid; Fu X et al.; Two new C-nor/D-homo corticosteroids were isolated from the filtrate of the triene carbonate intermediate during the production of betamethasone . Their structures were elucidated based on NMR spectroscopic studies and mass spectral analyses . The mechanism of formation of each of these two steroids was postulated as a Wagner-Meerwein rearrangement of the corresponding 12beta-hydroxy steroid . The new 12beta-hydroxy corticosteroid was also isolated from the fermentation process of betamethasone synthesis . Its structure was identified via NMR and mass spectroscopic studies . Treatment of the new 12beta-hydroxy corticoid with PCl(5) at low temperature provided two C-nor/D-homo compounds in a ratio of 1:1 . Conversion of 12beta-hydroxy corticoid into a corresponding 12beta-mesylate followed by heating in HOAc to 110 degrees C in the presence of NaOAc produced a mixture of the rearranged products in a 1:4 ratio. Appl Biochem Biotechnol, 2002 Feb, 97(2), 105 - 21 Low-temperature brewing by freeze-dried immobilized cells; Bekatorou A et al.; We propose a novel biocatalyst in brewing . A cryotolerant strain of Saccharomyces cerevisiae was immobilized on delignified cellulosic material followed by freeze-drying of the immobilized cells without the use of any cryoprotectant . The freeze-dried immobilized biocatalyst was used in repeated-batch fermentation of wort and showed reduced fermentation time and increased productivities as compared with free freeze-dried cells (FFDCs) . It also demonstrated suitability for low-temperature brewing (5 and 0 degrees C) . The fermentation time in repeated-batch fermentations at 15 degrees C was 1.5-2 d for a period of 13 mo, showing a high operational stability of the system . At 0 degrees C the freeze-dried immobilized biocatalyst showed a 2- to 3.5-fold decrease in fermentation time in comparison with FFDCs . Polyphenol contents, bitterness, and diacetyl concentration were lower in beers produced by freeze-dried immobilized cells as compared with FFDCs . At 0 degrees C polyphenols were 40% lower than at 15 degrees C . Higher alcohols were reduced and ethyl acetate increased in comparison with FFDCs . Amyl alcohols at 0 degrees C were lower than half of their content at 15 degrees C, while ethyl acetate was 31 mg/L at 0 degrees C and 18 mg/L at 15 degrees C . These data justify the improved aroma and taste of beers produced by freeze-dried immobilized biocatalyst mainly at low temperatures. Biotechnol Bioeng, 2002 Jun 20, 78(6), 658 - 69 An assessment of seed quality and its influence on productivity estimation in an industrial antibiotic fermentation; Cunha CC et al.; This study investigates the benefits of including seed quality information into data-based models for final productivity estimation in an industrial antibiotic fermentation process . Multiway principal component analysis is applied to assess the seed quality using routinely gathered plant data . Multiway partial least-squares regression is then used to estimate the final productivity using data from the main fermentation only . The issue of selecting appropriate process variables as inputs is investigated . Subsequently, seed characteristics are included into the estimation models to assess the benefits of including information from this stage for productivity estimation . It is shown that it is possible to extract seed fermentation features related to the final productivity both at pilot and production scales . It is postulated that significant influential variations are mirrored in monitored variables during the main fermentation, and therefore seed quality is implicitly accounted for . Int J Cancer, 2002 May 20, 99(3), 424 - 6 Novel epidemiologic evidence for the association between fermented fish sauce and esophageal cancer in South China; Ke L et al.; Previous studies have suggested that fermented fish sauce is related to an increased risk for nasopharyngeal, thyroid and gastric cancers and has suspicious carcinogenic and promoting effects in the laboratory, but these reports have not investigated the association between this agent and esophageal cancer in population . Therefore, we investigated the relationship between esophageal cancer and consumption of fermented fish sauce, alcohol and tobacco after adjusting other risk factors using data from a large hospital-based case-control study in Chaoshan area of China . The subjects for analysis included 1,248 cases (median age 58.5 {range 29-82} years, 936 males, 312 females, some 50% with the habit of fermented fish sauce eating) with squamous cell carcinoma of the esophagus and the same amount of controls matched by sex and age . A significant increase in risk (OR 3.21; 95% CI 2.45-4.19) for eating fermented fish sauce compared to not eating and a significant dose-response relationship was found with the consumption of fermented fish sauce (p for trend < 0.001) . In comparison based on the binary variables, the OR for the subjects with fermented fish sauce eating and current smoking was 15.5 with a 95% CI of 8.14-29.3, relative to those exposed to neither habit . The joint effect of fermented fish sauce (F) and smoking (S) was more than additive (F * S > F + S: 15.5 > 1 + (5.6 - 1) + (6.6 - 1)), but the effect for fermented fish sauce and alcohol was not found . It is concluded that risks of esophageal cancer in the population were substantially associated with fermented fish sauce . Further epidemiologic and experimental study are required to find a biologic causal relationship between them . Bioresour Technol, 2002 May, 82(3), 277 - 84 Oyster mushroom cultivation with rice and wheat straw; Zhang R et al.; Cultivation of the oyster mushroom, Pleurotus sajor-caju, on rice and wheat straw without nutrient supplementation was investigated . The effects of straw size reduction method and particle size, spawn inoculation level, and type of substrate (rice straw versus wheat straw) on mushroom yield, biological efficiency, bioconversion efficiency, and substrate degradation were determined . Two size reduction methods, grinding and chopping, were compared . The ground straw yielded higher mushroom growth rate and yield than the chopped straw . The growth cycles of mushrooms with the ground substrate were five days shorter than with the chopped straw for a similar particle size . However, it was found that when the straw was ground into particles that were too small, the mushroom yield decreased . With the three spawn levels tested (12%, 16% and 18%), the 12% level resulted in significantly lower mushroom yield than the other two levels . Comparing rice straw with wheat straw, rice straw yielded about 10% more mushrooms than wheat straw under the same cultivation conditions . The dry matter loss of the substrate after mushroom growth varied from 30.1% to 44.3% . The straw fiber remaining after fungal utilization was not as degradable as the original straw fiber, indicating that the fungal fermentation did not improve the feed value of the straw. Proteomics, 2001 Sep, 1(9), 1133 - 48 Similarity of the Escherichia coli proteome upon completion of different biopharmaceutical fermentation processes; Champion KM et al.; A comprehensive view of the physiological state of Escherichia coli cells at the completion of fermentation processes for biopharmaceutical production was attained via two-dimensional gel electrophoretic analysis of cellular proteins . For high cell density fermentations in which phosphate is depleted to induce recombinant protein expression from the alkaline phosphatase promoter, proteome analysis confirms that phosphate limitation occurs . Known phosphate starvation inducible proteins are observed at high levels; these include the periplasmic phosphate binding protein and the periplasmic phosphonate binding protein . The phn (EcoK) locus of these E . coli K-12 strains remains cryptic, as demonstrated by failure to grow with phosphonate as the sole phosphorus source . Proteome analysis also provided evidence that cells utilize alternative carbon and energy sources during these fermentation processes . To address regulatory issues in the biopharmaceutical industry, comparative electrophoretic analyses were conducted on a qualitative basis for four different fermentation processes . Using this approach, the protein profiles for these processes were found to be highly similar, with the vast majority (85-90%) of proteins detected in all profiles . The observed similarity in proteomes suggests that multiproduct host cell protein immunoassays are a feasible means of quantifying host-derived polypeptides from a variety of biopharmaceutical fermentation processes. Nat Prod Lett, 2002 Apr, 16(2), 87 - 93 3,4-Dihydroxy-7,8-dihydro-beta-ionone 3-O-beta-D-glucopyranoside and other glycosidic constituents from apple leaves; Stingl C et al.; 3,4-Dihydroxy-7,8-dihydro-beta-ionone 3-O-beta-D-glucopyranoside 1 has been isolated from a methanolic extract of apple (Malus domestica) leaves by XAD-2 adsorption chromatography and subsequent purification by high speed countercurrent chromatography (HSCCC) and HPLC . Under acidic conditions this glycoside gives rise to a number of volatile compounds including 1,1,6-trimethyl-1,2-dihydronaphthalene (TDN) 2, which is known as an off-flavor compound in wine . In the course of the study, six additional glycoconjugates were isolated from apple leaves, i.e . roseoside 3, 3-oxo-alpha-ionol beta-D-glucopyranoside 4, benzoic acid beta-D-glucose ester 5, kaempferol 3-O-rhamnoside 6, (+)-syringaresinol beta-D-glucopyranoside 7 and 4-hydroxy-5-(3''-methyl-2''-butenyl)-benzoic acid methylester 3-beta-D-glucopyranoside 8 . The latter compound has been isolated for the first time in nature . In order to avoid the 'kerosene-off-flavor' caused by TDN, the methanolic extract has been subjected to yeast fermentation . This treatment reduced the amount of TDN formed and resulted in an improved flavor impression of the methanolic extract. Microbiology, 2002 May, 148(Pt 5), 1397 - 406 Identification of novel sulfur-containing bacterial polyesters: biosynthesis of poly(3-hydroxy-S-propyl-omega-thioalkanoates) containing thioether linkages in the side chains; Ewering C et al.; This study describes the biosynthesis of novel sulfur-containing polyhydroxyalkanoates (PHAs), which consist exclusively of hydroxypropylthioalkanoic acid containing thioether groups in the side chains . In addition, the utilization of alkylthioalkanoic acids (=thia fatty acids) by various bacteria was investigated . Based on feedings with propylthiooctanoic acid (PTO) or propylthiohexanoic acid, the metabolically engineered PHA-negative mutant PHB(-)4 of Ralstonia eutropha, which harbours plasmid pBBR1::phaC1 expressing the PHA synthase of Pseudomonas mendocina, synthesized two novel poly(3-hydroxy-S-propyl-omega-thioalkanoic) acids {poly(3HPTA)s} . A terpolyester consisting of 3-hydroxypropylthiobutyric acid (3HPTB), 3-hydroxypropylthiohexanoic acid (3HPTHx) and 3-hydroxypropyl- thiooctanoic acid (3HPTO) was synthesized from PTO, whereas a co-polyester of 3HPTB and 3HPTHx was synthesized from propylthiohexanoic acid . Fed-batch fermentation of R . eutropha PHB(-)4(pBBR1::phaC1) on PTO was done on a 26-litre scale, providing a cell density of 7.3 g l(-1), from which 45 g of the novel poly(3HPTB-co-3HPTHx-co-3HPTO) were isolated . The chemical structures of the poly(3HPTA)s were identified by gas chromatography/mass spectrometry, elemental sulfur analysis, partial pyrolysis and detailed mass spectrometric analysis, exhibiting 3HPTB, 3HPTHx and 3HPTO as constituents . These novel, hitherto undescribed, constituents of PHAs were randomly distributed in the co-polyesters. Med Lav, 2002 Jan-Feb, 93(1), 26 - 33 {Acute poisoning with carbon dioxide: report of 2 fatal cases}; Romeo L et al.; BACKGROUND: Carbon dioxide (CO2), a very high density gas, tends to stratify at the lowest levels of the atmosphere . It can be produced by neutral geothermal emissions, fermentative processes or by human and industrial activity . When carbon dioxide concentrations rise to a very high level in a confined and poorly ventilated space, the anoxic hazard is a very important cause of severe accidents that can involve workers and rescuers . At CO2 levels higher than 20% there is a very high risk of a fatal accident, also considering the odourless feature of this gas . OBJECT: Two fatal accidents in workers are described which occurred during inspection of a concrete well, built as a part of sewerage network in a rural area . In the weeks after the accident, composition and concentration of gases inside the well were analysed . We also considered the influence of an organic fertilizer called "pollina" which was found on the ground around the concrete well, in order to ascertain whether fermentation could alter the gas composition inside the well . METHODS: Samples of air and water were collected in the well and samples of the organic fertilizer (pollina) on the ground surrounding the concrete well were also taken . Different quantities of organic fertilizer (pollina) with or without water were incubated in airtight glass bottles and placed in a dark room at 20 degrees C temperature; analysis of air inside the glass bottles was performed after 7 and 18 days of incubation . All the samples of air and water were analysed by gas-chromatographic-mass-spectrometry . RESULTS: Analysis of the air samples collected inside the well after 2, 16 and 18 weeks after the accident showed a low concentration of O2 (range 4.2-9%), a high concentration of CO2 (range 5.9-12.3%), a normal level of N2 (range 78-85%) and a concentration of N2O between 0.03 and 0.19% . In water collected 2 weeks after the accident at the bottom of the concrete well, CO2 and N2O concentrations of respectively 222 mg/L and 2 mg/L were measured . In the bottles with "pollina" we found, at different times of incubation, high concentrations of CO2 (highest value 25.2%), low levels of O2 (lowest value 0.5%) and negligible concentrations of N2O (< 0.015%) . CONCLUSIONS: All these findings suggest that the atmosphere inside the concrete well was altered by the fermentative processes of pollina . The death of the two workers, caused by a poorly oxygenated atmosphere with a high concentration of carbon dioxide, can be classified under the confined space hypoxic syndrome (CSHS). J Ind Microbiol Biotechnol, 2002 May, 28(5), 297 - 301 Enhanced production of heterologous macrolide aglycones by fed-batch cultivation of Streptomyces coelicolor; Desai RP et al.; A media development program for the enhanced production of macrolide aglycones by Streptomyces coelicolor is described . Shake flask studies utilizing a yeast extract and a bakers' yeast increased production by 200% and 80%, respectively . However, ammonia generation and high pH were identified as potential problems in these enriched media . Studies in pH-controlled fermentors revealed that production stage pH significantly affects macrolide titers, with low pH (5.5) being more productive than high pH (6.5) . Implementation of glucose feeding in shake flask cultures reduced ammonia generation and controlled production stage pH, resulting in significantly enhanced productivities . The combined effects of media supplementation and glucose feeding resulted in a three to five-fold overall improvement in total macrolide aglycone titers, and is the first reported high-level (>1 g/l) production of recombinant polyketides in a heterologous host. J Ind Microbiol Biotechnol, 2002 May, 28(5), 280 - 3 Production of testosterone from phytosterol using a single-step microbial transformation by a mutant of Mycobacterium sp; Lo CK et al.; A testosterone (TS)-producing mutant, ST2, was derived from a phytosterol-assimilating and androst-4-ene-3,17-dione (AD)-producing bacterium, Mycobacterium sp . B-3805S, using nitrosoguanidine (NTG) mutagenesis . Production of TS from phytosterol using a single-step microbial transformation process by ST2 was investigated in a 5-l surface-aeration microprocessor-controlled fermentor loaded with a synthetic medium supplemented with 0.1% phytosterol, 2% glucose and 1% peptone at 30 degrees C . An increase in dissolved oxygen at the initial stage of fermentation favored the side-chain degradation of phytosterol to AD . Later in the fermentation, a decrease in the dissolved oxygen to zero resulted in a decrease in pH to 6.0 as well as the reduction of AD to TS . Under optimal fermentation conditions, the maximum conversion ratio of phytosterol to TS was 31% after 120 h cultivation . It was concluded that the control of dissolved oxygen in the fermentation culture is the most important parameter for production of TS from phytosterol via AD . TS was isolated from the fermentation culture by addition of Amberlite XAD-7 resin and was further purified by flash chromatography on a silica gel column . After crystallization, TS was obtained as needle crystals with the correct melting point. Mol Microbiol, 2002 Jan, 43(2), 449 - 58 Differential roles of two SARP-encoding regulatory genes during tylosin biosynthesis; Bate N et al.; The tylosin biosynthetic gene cluster of Streptomyces fradiae is remarkable in harbouring at least five regulatory genes, two of which (tylS and tylT) encode proteins of the Streptomyces antibiotic regulatory protein (SARP) family . The aim of the present work was to assess the respective contributions of TylS and TylT to tylosin production . A combination of targeted gene disruption, fermentation studies and gene expression analysis via reverse transcriptase-polymerase chain reaction (RT-PCR) suggests that tylS is essential for tylosin production and controls the expression of tylR (previously shown to be a global activator of the biosynthetic pathway) plus at least one other gene involved in polyketide metabolism or regulation thereof . This is the first demonstration of a SARP acting to control another regulatory gene during antibiotic biosynthesis . In contrast, tylT is not essential for tylosin production. Eur J Biochem, 2002 Apr, 269(8), 2052 - 9 ORF6 from the clavulanic acid gene cluster of Streptomyces clavuligerus has ornithine acetyltransferase activity; Kershaw NJ et al.; The clinically used beta-lactamase inhibitor clavulanic acid is produced by fermentation of Streptomyces clavuligerus . The orf6 gene of the clavulanic acid biosynthetic gene cluster in S . clavuligerus encodes a protein that shows sequence homology to ornithine acetyltransferase (OAT), the fifth enzyme of the arginine biosynthetic pathway . Orf6 was overexpressed in Escherichia coli (at approximately 15% of total soluble protein by SDS/PAGE analysis) indicating it was not toxic to the host cells . The recombinant protein was purified (to > 95% purity) by a one-step technique . Like other OATs it was synthesized as a precursor protein which underwent autocatalytic internal cleavage in E . coli to generate alpha and beta subunits . Cleavage was shown to occur between the alanine and threonine residues in a KGXGMXXPX--(M/L)AT (M/L)L motif conserved within all identified OAT sequences . Gel filtration and native electrophoresis analyses implied that the ORF6 protein was an alpha2beta2 heterotetramer and direct evidence for this came from mass spectrometric analyses . Although anomalous migration of the beta subunit was observed by standard SDS/PAGE analysis, which indicated the presence of two bands (as previously observed for other OATs), mass spectrometric analyses did not reveal any evidence for post-translational modification of the beta subunit . Extended denaturation with SDS before PAGE resulted in observation of a single major beta subunit band . Purified ORF6 was able to catalyse the reversible transfer of an acetyl group from N-acetylornithine to glutamate, but not the formation of N-acetylglutamate from glutamate and acetyl-coenzyme A, nor (detectably) the hydrolysis of N-acetylornithine . Mass spectrometry also revealed the reaction proceeds via acetylation of the beta subunit. Microb Ecol, 2002 Jan, 43(1), 92 - 106 Epub 2001 Dec 07. Functional groups and activities of bacteria in a highly acidic volcanic mountain stream and lake in Patagonia, Argentina; Wendt-Potthoff K et al.; Acidic volcanic waters are naturally occurring extreme habitats that are subject of worldwide geochemical research but have been little investigated with respect to their biology . To fill this gap, the microbial ecology of a volcanic acidic river (pH approximately equal to 0-1.6), Rio Agrio, and the recipient lake Caviahue in Patagonia, Argentina, was studied . Water and sediment samples were investigated for Fe(II), Fe(III), methane, bacterial abundances, biomass, and activities (oxygen consumption, iron oxidation and reduction) . The extremely acidic river showed a strong gradient of microbial life with increasing values downstream and few signs of life near the source . Only sulfide-oxidizing and fermentative bacteria could be cultured from the upper part of Rio Agrio . However, in the lower part of the system, microbial biomass and oxygen penetration and consumption in the sediment were comparable to non-extreme aquatic habitats . To characterize similarities and differences of chemically similar natural and man-made acidic waters, our findings were compared to those from acidic mining lakes in Germany . In the lower part of the river and the lake, numbers of iron and sulfur bacteria and total biomass in sediments were comparable to those known from acidic mining lakes . Bacterial abundance in water samples was also very similar for both types of acidic water (around 10(5) mL(-1)) . In contrast, Fe(II) oxidation and Fe(III) reduction potentials appeared to be lower despite higher biogenic oxygen consumption and higher photosynthetic activity at the sediment-water interface . Surprisingly, methanogenesis was detected in the presence of high sulfate concentrations in the profundal sediment of Lake Caviahue . In addition to supplementing microbiological knowledge on acidic volcanic waters, our study provides a new view of these extreme sites in the general context of aquatic habitats. Eur J Gastroenterol Hepatol, 2002 May, 14(5), 535 - 42 Predominance of caecal injury in a new dextran sulphate sodium treatment in rats: histopathological and fermentative characteristics; Moreau NM et al.; OBJECTIVES : Cyclic administrations of dextran sulphate sodium (DSS) alternating with distilled water usually induce chronic colitis after a few weeks . In order to obtain stable chronic colitis (without recovery or relapse) in a few days, a new continuous DSS treatment was tested and characterized . Short-chain fatty acids (SCFAs), which remain poorly documented in experimental colitis, were also investigated . METHODS : Thirty-six Sprague-Dawley rats were treated with 5% DSS for 7 days (DI) followed by 3% DSS for 7 days (DM) or 14 days (DF) . Control rats received only water . Inflammatory injuries in the caecum and the colon were assessed by macroscopic (colon length, caecum weight, damages score) and histological parameters . SCFAs (acetate, propionate, butyrate) were quantified individually in caecal, proximal and distal contents . RESULTS : Macroscopic and histological observations revealed that this continuous DSS treatment induced acute inflammation (DI) followed rapidly by chronic active colitis . The latter was uncommonly predominant in the caecum and the distal colon, and was also associated with some fermentative disturbances . Caecal SCFA concentrations decreased with DSS at DI and DM . The molar ratio of caecal butyrate increased with DSS . Acetate decreased in the colon while propionate increased . CONCLUSION : This new DSS treatment is able to induce in a few days stable chronic inflammation with caecal and distal predominant injuries, and mild fermentative caeco-colonic alterations . This model could contribute to the study of potential anti-inflammatory effects of prebiotics. J Nutr, 2002 May, 132(5), 1012 - 7 The effects of short-chain fatty acids on human colon cancer cell phenotype are associated with histone hyperacetylation; Hinnebusch BF et al.; The short-chain fatty acid (SCFA) butyrate is produced via anaerobic bacterial fermentation within the colon and is thought to be protective in regard to colon carcinogenesis . Although butyrate (C4) is considered the most potent of the SCFA, a variety of other SCFA also exist in the colonic lumen . Butyrate is thought to exert its cellular effects through the induction of histone hyperacetylation . We sought to determine the effects of a variety of the SCFA on colon carcinoma cell growth, differentiation and apoptosis . HT-29 or HCT-116 (wild-type and p21-deleted) cells were treated with physiologically relevant concentrations of various SCFA, and histone acetylation state was assayed by acid-urea-triton-X gel electrophoresis and immunoblotting . Growth and apoptotic effects were studied by flow cytometry, and differentiation effects were assessed using transient transfections and Northern blotting . Propionate (C3) and valerate (C5) caused growth arrest and differentiation in human colon carcinoma cells . The magnitude of their effects was associated with a lesser degree of histone hyperacetylation compared with butyrate . Acetate (C2) and caproate (C6), in contrast, did not cause histone hyperacetylation and also had no appreciable effects on cell growth or differentiation . SCFA-induced transactivation of the differentiation marker gene, intestinal alkaline phosphatase (IAP), was blocked by histone deacetylase (HDAC), further supporting the critical link between SCFA and histones . Butyrate also significantly increased apoptosis, whereas the other SCFA studied did not . The growth arrest induced by the SCFA was characterized by an increase in the expression of the p21 cell-cycle inhibitor and down-regulation of cyclin B1 (CB1) . In p21-deleted HCT-116 colon cancer cells, the SCFA did not alter the rate of proliferation . These data suggest that the antiproliferative, apoptotic and differentiating properties of the various SCFA are linked to the degree of induced histone hyperacetylation . Furthermore, SCFA-mediated growth arrest in colon carcinoma cells requires the p21 gene. J Nutr, 2002 May, 132(5), 967 - 73 Dietary fructans, but not cellulose, decrease triglyceride accumulation in the liver of obese Zucker fa/fa rats; Daubioul C et al.; This study was designed to compare the effects of dietary supplementation with nondigestible carbohydrates, differing in fermentability by colonic bacteria, on hepatic steatosis in growing obese Zucker rats . Male Zucker fa/fa rats were divided into three groups: a control group that received the basal diet, a fructan group that received 10 g highly fermented Synergy 1/100 g diet and a cellulose group that received 10 g poorly fermented Vivapur Microcrystalline cellulose/100 g diet . Rats consuming fructan had a lower energy intake, a lower body weight and less triacylglycerol accumulation in the liver as assessed in vivo by nuclear magnetic resonance (NMR) spectroscopy, and ex vivo by biochemical and histochemical analysis compared with the control and/or cellulose groups . The high fermentation of fructans compared with cellulose was reflected by greater cecal contents and by a twofold greater propionate concentration in the portal vein of rats fed fructan compared with those fed cellulose . By measuring the capacity of hepatocytes isolated from liver of Zucker rats to synthesize triglycerides or total lipids from different precursors, we showed that propionate, at the concentrations measured in the portal vein of rats treated with fructan, selectively decreased the incorporation of acetate into total lipids, a phenomenon that could contribute, along with the lower energy intake, to less triglyceride accumulation in the liver of obese Zucker rats fed dietary fructans. J Immunol Methods, 2002 Apr 1, 262(1-2), 217 - 27 Large-scale bacterial fermentation and isolation of scFv multimers using a heat-inducible bacterial expression vector; Bayly AM et al.; This protocol describes optimised large-scale bacterial fermentation conditions for recombinant single-chain Fv molecule (scFv) monomers and multimers (diabodies and triabodies) . The heat-inducible bacterial secretion vector, pPOW3, utilising the temperature-regulated tandem lambda promoters is particularly suited to the large-scale fermentation of single-chain antibodies, providing low-cost recombinant protein synthesis . The protein expressed by this vector is secreted into the periplasm where it is found as both the soluble and insoluble protein that is associated with the cell membranes . A protein fractionation method for the rapid extraction and affinity purification of the soluble protein fraction and the urea solubilization and refolding of the insoluble protein fraction expressed from single-chain antibody (Ab) fragment gene constructs is described . This method is simple to perform and utilises inexpensive reagents to provide cost-effective protein synthesis. J Gastroenterol Hepatol, 2002 Mar, 17(3), 249 - 52 Physiological small bowel malabsorption of carbohydrates protects against large bowel diseases in Africans; Segal I; In African black people there is a paucity of 'developed society' large bowel diseases such as diverticular disease, colorectal adenomas and carcinomas, ulcerative colitis and Crohn's disease . Appendicitis has an incidence of about 5-10% of the number likely to be observed in a white population . The conundrum is that the disparity exists despite many Africans having adopted an urbanized lifestyle with major changes in their dietary pattern . Dietary fiber intake, which was previously 30-35 g, has decreased to 12-14 g daily . Studies on small bowel function in black people have shown that physiological malabsorption of lactose, fructose, sucrose and maize (the staple food) occurs . It is hypothesized that the increased concentration of substrate available for fermentation in the colon compensates for the low dietary fiber intake, is protective to the large bowel and is a factor in the prevention of 'developed society' large bowel diseases in the African population . J Agric Food Chem, 2002 May 8, 50(10), 3030 - 6 Evaluation of the nutritional characteristics of a finger millet based complementary food; Mbithi-Mwikya S et al.; Finger millet (Eleusine coracana), kidney beans (Phaseolus vulgaris), peanuts (Arachis hypogoea), and mango (Mangifera indica) were processed separately and then combined, on the basis of their amino acid scores and energy content, into a complementary food for children of weaning age . The finger millet and kidney beans were processed by germination, autoclaving, and lactic acid fermentation . A mixture containing, on a dry matter basis, 65.2, 19.1, 8.0, and 7.7% of the processed finger millet, kidney beans, peanuts, and mango, respectively, gave a composite protein with an in vitro protein digestibility of 90.2% and an amino acid chemical score of 0.84 . This mixture had an energy density of 16.3 kJ.g(-1) of dry matter and a decreased antinutrient content and showed a measurable improvement in the in vitro extractability for calcium, iron, and zinc . A 33% (w/v) pap made from a mix of the processed ingredients had an energy density of 5.4 kJ.g(-1) of pap, which is sufficient to meet the energy requirements of well-nourished children of 6-24 months of age at three servings a day and at the FAO average breast-feeding frequency. J Agric Food Chem, 2002 May 8, 50(10), 2891 - 9 Relationship between varietal amino acid profile of grapes and wine aromatic composition . Experiments with model solutions and chemometric study; Hernandez-Orte P et al.; Synthetic solutions containing amino acids, sugar, water, and yeast nutrients have been fermented by Saccharomyces cerevisiae, and the volatile composition of the fermented media has been analyzed by GC . Eleven amino acid compositions imitating the characteristic amino acid profile of 11 different grape varieties were tested . Significant differences in the levels of some important volatile compounds (ethanol, ethyl acetate, acetic acid, higher alcohols and some of their acetates, methionol, isobutyric acid, ethyl butyrate, and hexanoic and octanoic acids) were found . The levels of some of the volatile compounds are well correlated with the aromatic composition of wines made with grapes of the same varieties . A multiple linear regression study produced good models for most of the odorants for which the level was related to the must amino acid composition . Partial least-squares regression models confirm that amino acid composition explains a high proportion of the variance in the volatile composition and show that the relationship between both sets of variables is highly multivariate . According to the different models, the levels of some byproducts of fatty acid synthesis are related to threonine and serine, the level of beta-phenyletanol is closely related to the level of phenylalanine, and methionol is strongly correlated to the must methionine contents . The addition of selected amino acids to different musts confirms the previous observations. J Basic Microbiol, 2002, 42(2), 83 - 90 Production of laccase as the sole phenoloxidase by a Brazilian strain of Pleurotus pulmonarius in solid state fermentation; Marques De Souza CG et al.; The production of laccase by a Brazilian strain of Pleurotus pulmonarius was studied in solid state fermentation using wheat bran as substrate . Among oxidative and hydrolytic enzymes tested (laccase, aryl alcohol oxidase, lignin peroxidase, Mn peroxidase, xylanase and cellulase), laccase was the main enzyme produced by P . pulmonarius . The most suitable condition for maximum production of laccase (8,600 U/g substrate) was initial moisture content of 75% and 5 days of cultivation at 30 degrees C . The optimum pH and temperature for laccase activity were found to be 6.5 and 50 degrees C, respectively . P . pulmonarius laccase was stable at 50 degrees C for more than 6 hours, and it retained about 73% and 18% of its activity when heated for 1 h at 55 and 60 degrees C, respectively . The enzyme was greatly stable at alkaline pH, but not at acidic pH . The laccase activity appear to be correlated with the ability of crude extract to decolourize several industrial dyes. Biochemistry, 2002 May 7, 41(18), 5873 - 82 Adenosine triphosphate-induced electron transfer in 2-hydroxyglutaryl-CoA dehydratase from Acidaminococcus fermentans; Hans M et al.; 2-hydroxyglutaryl-CoA dehydratase from Acidaminococcus fermentans catalyzes the chemical difficult elimination of water from (R)-2-hydroxyglutaryl-CoA to glutaconyl-CoA . The enzyme consists of two oxygen-sensitive protein components, the homodimeric activator (A) with one {4Fe-4S}1+/2+ cluster and the heterodimeric dehydratase (D) with one nonreducible {4Fe-4S}2+ cluster and reduced riboflavin 5'-monophosphate (FMNH2) . For activation, ATP, Mg2+, and a reduced flavodoxin (16 kDa) purified from A . fermentans are required . The {4Fe-4S}(1+/2+) cluster of component A is exposed to the solvent since it is accessible to iron chelators . Upon exchange of the bound ADP by ATP, the chelation rate is 8-fold enhanced, indicating a large conformational change . Oxidized component A exhibits ATPase activity of 6 s(-1), which is completely abolished upon reduction by one electron . UV-visible spectroscopy revealed a spontaneous one-electron transfer from flavodoxin hydroquinone (E(0)' = -430 mV) to oxidized component A, whereby the {4Fe-4S}2+ cluster of component A became reduced . Combined kinetic, EPR, and Mossbauer spectrocopic investigations exhibited an ATP-dependent oxidation of component A by component D . Whereas the {4Fe-4S}2+ cluster of component D remained in the oxidized state, a new EPR signal became visible attributed to a d1-metal species, probably Mo(V) . Metal analysis with neutron activation and atomic absorption spectroscopy gave 0.07-0.2 Mo per component D . In summary, the data suggest that in the presence of ATP one electron is transferred from flavodoxin hydroquinone via the {4Fe-4S}1+/2+ cluster of component A to Mo(VI) of component D, which is thereby reduced to Mo(V) . The latter may supply the electron necessary for transient charge reversal in the unusual dehydration. Sheng Wu Gong Cheng Xue Bao, 2002 Jan, 18(1), 74 - 8 {Studies on the optimal expression condition, purification and its characterization of ScFv-2F3}; Luo YM et al.; The expression vectors of the gene encoding ScFv-2F3 were transformed into E . coli BL21(DE3) . Clones of higher expression were first selected, then were grown in the presence of IPTG at 37 degrees C to induce its expression . The culture conditions were carefully optimized . It was found that optimal conditions were as follows: the induction was started as OD590 reached to 1.0-1.8; the concentration of IPTG was 0.3-0.5 mmol/L and induction time is 7 h . The yield of ScFv-2F3 expressed in the selected clones is about 20% of the total proteins . The optimal culture conditions were successfully applied to fermenter of 50 L . The conditions of washing the inclusion bodies were also optimized . A two-step method was used to renature the inclusion body . The expression product of interest and its biological activities were characterized with Western blotting and ELISA . A novel selenium-containing single-chain abzyme with GPX activity was prepared. Arch Microbiol, 2002 May, 177(5), 381 - 91 Epub 2002 Mar 05. Desulfotignum phosphitoxidans sp . nov., a new marine sulfate reducer that oxidizes phosphite to phosphate; Schink B et al.; A new sulfate-reducing bacterium was isolated from marine sediment with phosphite as sole electron donor and CO(2) as the only carbon source . Strain FiPS-3 grew slowly, with doubling times of 3-4 days, and oxidized phosphite, hydrogen, formate, acetate, fumarate, pyruvate, glycine, glutamate, and other substrates nearly completely, with concomitant reduction of sulfate to sulfide . Acetate was formed as a side product to a small extent . Glucose, arabinose, and proline were partly oxidized and partly fermented to acetate plus propionate . Growth with phosphite, hydrogen, or formate was autotrophic . Also, in the presence of sulfate, CO dehydrogenase was present, and added acetate did not increase growth rates or growth yields . In the absence of sulfate, phosphite oxidation was coupled to homoacetogenic acetate formation, with growth yields similar to those in the presence of sulfate . Cells were small rods, 0.6 - 0.8 x 2-4 microm in size, and gram-negative, with a G+C content of 53.9 mol% . They contained desulforubidin, but no desulfoviridin . Based on sequence analysis of the 16S rRNA gene and the sulfite reductase genes dsrAB, strain FiPS-3 was found to be closely related to Desulfotignum balticum . However, physiological properties differed in many points from those of D . balticum . These findings justify the establishment of a new species, Desulfotignum phosphitoxidans. Appl Environ Microbiol, 2002 May, 68(5), 2294 - 9 Mechanisms for accessing insoluble Fe(III) oxide during dissimilatory Fe(III) reduction by Geothrix fermentans; Nevin KP et al.; Mechanisms for Fe(III) oxide reduction were investigated in Geothrix fermentans, a dissimilatory Fe(III)-reducing microorganism found within the Fe(III) reduction zone of subsurface environments . Culture filtrates of G . fermentans stimulated the reduction of poorly crystalline Fe(III) oxide by washed cell suspensions, suggesting that G . fermentans released one or more extracellular compounds that promoted Fe(III) oxide reduction . In order to determine if G . fermentans released electron-shuttling compounds, poorly crystalline Fe(III) oxide was incorporated into microporous alginate beads, which prevented contact between G . fermentans and the Fe(III) oxide . G . fermentans reduced the Fe(III) within the beads, suggesting that one of the compounds that G . fermentans releases is an electron-shuttling compound that can transfer electrons from the cell to Fe(III) oxide that is not in contact with the organism . Analysis of culture filtrates by thin-layer chromatography suggested that the electron shuttle has characteristics similar to those of a water-soluble quinone . Analysis of filtrates by ion chromatography demonstrated that there was as much as 250 microM dissolved Fe(III) in cultures of G . fermentans growing with Fe(III) oxide as the electron acceptor, suggesting that G . fermentans released one or more compounds capable of chelating and solubilizing Fe(III) . Solubilizing Fe(III) is another strategy for alleviating the need for contact between cells and Fe(III) oxide for Fe(III) reduction . This is the first demonstration of a microorganism that, in defined medium without added electron shuttles or chelators, can reduce Fe(III) derived from Fe(III) oxide without directly contacting the Fe(III) oxide . These results are in marked contrast to those with Geobacter metallireducens, which does not produce electron shuttles or Fe(III) chelators . These results demonstrate that phylogenetically distinct Fe(III)-reducing microorganisms may use significantly different strategies for Fe(III) reduction . Thus, it is important to know which Fe(III)-reducing microorganisms predominate in a given environment in order to understand the mechanisms for Fe(III) reduction in the environment of interest. Physiol Plant, 2002 Apr, 114(4), 524 - 532 Imazethapyr, an inhibitor of the branched-chain amino acid biosynthesis, induces aerobic fermentation in pea plants; Gaston S et al.; Acetolactate synthase (ALS; EC 4.1.3.18) inhibition is the primary mechanism of action of imazethapyr (IM) . However, the precise mechanisms that links ALS inhibition with plant death have not been elucidated . Supply of IM to pea (Pisum sativum L) plants produced an immediate cessation of growth, caused a 50% inhibition of the in vivo ALS activity within 1 day of treatment, and a remarkable accumulation (2.7-times) of free amino acids after 3 days . Carbohydrates (soluble and starch) were accumulated in both leaves and roots . Accumulation of soluble sugars in roots preceded that of starch in leaves, suggesting that the accumulation of carbohydrates in leaves is not the reason for the arrested root growth . A transient pyruvate accumulation was observed in roots, 1 day after the onset of IM supply . This was coincident with an increase in pyruvate decarboxylase (EC 4.1.1.1), and later increases in alcohol dehydrogenase (EC 1.1.1.1), lactate dehydrogenase (EC 1.1.1.27), and alanine amino transferase (EC 2.6.1.2) activities . This enhancement of fermentative activities was coincident with a slight decrease in aerobic respiration . The overall data suggest that the impairment of ALS activity may lead to a fermentative metabolism that may be involved in growth inhibition and plant death. Org Lett, 2002 May 2, 4(9), 1431 - 4 Integramides A and B, two novel non-ribosomal linear peptides containing nine C(alpha)-methyl amino acids produced by fungal fermentations that are inhibitors of HIV-1 integrase; Singh SB et al.; {structure: see text} . Integramides A and B are two novel 16-mer linear peptides rich in C(alpha)-methyl amino acids that were isolated from fungal extracts of Dendrodochium sp . by employing a bioassay-guided isolation procedure using recombinant HIV-1 integrase . The structure and stereochemistry were elucidated by a combination of 2D NMR and ESI- and FAB-MS including MS/MS studies and by Marfey's method . Integramides A and B inhibited the coupled reaction of HIV-1 integrase with IC50 values of 17 and 10 microM, respectively. J Nat Prod, 2002 Apr, 65(4), 570 - 2 Large-scale isolation and crystallization of epothilone D from Myxococcus xanthus cultures; Arslanian RL et al.; The introduction of the epothilone polyketide synthase (PKS) into Myxococcus xanthus has enabled the heterologous production of epothilone D (1) on a large scale . To isolate this valuable product from the fermentation medium, an economical, scalable, and high-yielding purification process was developed . With the crystallization of 1 from a binary solvent system that consisted of ethanol and water, the product was recovered as white crystals with a final purity of > or =97% (w/w) . This is the first reported crystallization of 1. Food Nutr Bull, 2002 Mar, 23(1), 94 - 105 Iron-deficiency anemia in young working women can be reduced by increasing the consumption of cereal-based fermented foods or gooseberry juice at the workplace; Gopaldas T; This efficacy for both employers and employees (young working women 18 to 23 years of age) was undertaken to determine whether culturally acceptable dietary changes in lunches in the workplace and at home could bring about a behavioral change and improvement in their iron-deficiency anemia status . Maximum weight was given to increasing consumption of iddli, a popular cereal-based-fermented food, or of gooseberry juice . Four small factories were selected in periurban Bangalore, with a sample of 302 women . The 180-day interventions were supervised at the workplace . In unit 1 (72 women), the intervention consisted of iddli four times a week plus information, education, and communication (IEC) related to iron-deficiency anemia . Unit 2 (80 women) received 20 ml of gooseberry juice (containing 40 mg of vitamin C) three times a week plus IEC once a month . Women in unit 3 (70 women), the positive control, received 400 mg albendazole once plus ferrous sulfate tablets (60 mg elemental iron) two times a week . No IEC was given . Unit 4 (70 women) served as the negative control and received no intervention . The pre-post impact measures were dietary and nutrient intake, knowledge and practice, and hemoglobin status . In units 1, 2, and 3, the hemoglobin status of the women improved significantly from 11.10 to 12.30 g/dl, 11.20 to 12.70 g/dl, and 11.50 to 13.00 g/dl, respectively . In unit 4 there was no change: the values were 10.90 g/dl before and after intervention . The results show that the type of workplace lunch was of greater significance than IEC . Knowledge gains were impressive, but behavioral change was not sustained . It was concluded that the hemoglobin levels of the workers can easily be improved by cost-effective workplace lunches that also lead to better employer-employee relations. J Appl Microbiol, 2002, 92(5), 801 - 11 Influence of ciliate protozoa on biochemical changes and hydrolytic enzyme profile in the rumen ecosystem; Santra A et al.; AIMS: To assess the effect of presence or absence of rumen protozoa on fermentation characteristics and enzyme profile in growing lambs . METHODS AND RESULTS: Weaner lambs (G1, G2, G3, G4, G5 and G6 groups) were defaunated by oral administration of sodium laurel sulphate (at 8 g 100 kg(-1) body weight) . The lambs of G4, G5 and G6 groups were refaunated . The roughage and concentrate ratio in the diet of G1 and G4, G2 and G5, and G3 and G6 were 50:50 (R1), 65:35 (R2) and 80:20 (R3), respectively . Daily dry matter intake was similar in defaunated and faunated lambs . However, digestibility of organic matter (OM), cellulose and gross energy were lower in defaunated lambs while crude protein (CP) digestibility was similar in both defaunated and faunated lambs . The rumen pH and NH3-N were lower (P < 0.01) while TVFA, total-N and TCA-ppt-N were higher (P < 0.01), in defaunated lambs . Ruminal activity of carboxymethyl cellulase was lower (P < 0.01) in defaunated lambs and amylase, xylanase, protease and urease were similar in faunated and defaunated lambs . Nutrient utilization, rumen metabolites and ciliate protozoal count were higher, whereas digestibility of fibre fractions was lower in high rather than low concentrate fed lambs . The rumen protozoa present before defaunation were B-type and the protozoa which re-established on refaunation were also B-type . CONCLUSIONS: Absence of ciliate protozoa decreased nutrient digestibility and increased ruminal TVFA and total-N with lower NH3-N concentration, indicating better energy and protein utilization in defaunated lambs . SIGNIFICANCE AND IMPACT OF THE STUDY: Defaunation improved energy and protein utilization in lambs. J Anim Physiol Anim Nutr (Berl), 2002 Apr, 86(3-4), 57 - 73 Effects of different fibre sources and fat addition on cholesterol and cholesterol-related lipids in blood serum, bile and body tissues of growing pigs; Kreuzer M et al.; Knowledge is limited on the efficacy of hindgut-fermentable dietary fibre to reduce blood, bile and body tissue cholesterol levels . In three experiments with growing pigs the effects of different kinds and levels of bacterially fermentable fibre (BFS) on cholesterol metabolism were examined . Various diets calculated to have similar contents of metabolizable energy were supplied for complete fattening periods . In the first experiment, a stepwise increase from 12 to 20% BFS was performed by supplementing diets with fermentable fibre from sugar beet pulp (modelling hemicelluloses and pectin) . Beet pulp, rye bran (modelling cellulose) and citrus pulp (pectin) were offered either independently or in a mixture in the second experiment . These diets were opposed to rations characterized in carbohydrate type by starch either mostly non-resistant (cassava) or partly resistant (maize) to small intestinal digestion . The third experiment was planned to explore the interactions of BFS from citrus pulp with fat either through additional coconut oil/palm kernel oil blend or full-fat soybeans . In all experiments the increase of the BFS content was associated with a constant (cellulose) or decreasing (hemicelluloses, pectin) dietary proportion of non-digestible fibre . In experiment 1 an inverse dose-response relationship between BFS content and cholesterol in blood serum and adipose tissue as well as bile acid concentration in bile was noted while muscle cholesterol did not respond . In experiment 2 the ingredients characterized by cellulose and hemicelluloses/pectin reduced cholesterol-related traits relative to the low-BFS-high-starch controls whereas, except in adipose tissue cholesterol content, the pectinous ingredient had the opposite effect . However, the changes in serum cholesterol mainly affected HDL and not LDL cholesterol . Adipose tissue cholesterol also was slightly lower with partly resistant starch compared to non-resistant starch in the diet . Experiment 3 showed that the use of citrus pulp increased serum cholesterol concentrations when levels were low in the corresponding low-BFS diets (low-fat and soy bean diets), but caused no further increase in the coconut-oil/palm kernel oil blend diet . From the present results it seems that fermentable hemicelluloses have a more favourable effect of decreasing metabolic cholesterol and related traits than hardly digestible fibre, fermentable cellulose or, particularly, pectin . Furthermore, some types of fibre expressed a certain potential to reduce cholesterol content of fat pork and pork products by up to 10% (experiment 1) and 25% (experiment 2). J Exp Bot, 2002 May, 53(371), 1099 - 107 Legume embryos develop in a hypoxic environment; Rolletschek H et al.; Specific morphological and biochemical characteristics of seeds can cause oxygen deficiency within maternal and embryonic tissues . In this study, optical sensors were used to measure O(2) profiles across developing seeds of Vicia faba and Pisum sativum and developmental and environmental modulations of internal O(2) levels were studied . In addition, the metabolic state of developing embryos was analysed by monitoring adenylate energy charge, adenylate nucleotides and the levels of nucleotide sugars . Within the seed coat O(2) concentration decreased sharply to approximately 3% towards the inner border . Lowest O(2) levels were detected within the endospermal cavity between the seed coat and embryo . It is probable that low seed coat permeability provides an hypoxic environment for legume embryo development . The O(2) concentration in embryonic tissue changed during development with the lowest levels in the early stages . Measured in darkness, the levels were below 3%, but increased upon illumination indicating that photosynthesis significantly contributes to internal O(2) levels . Only in very young embryos were ATP levels and energy charge low . Otherwise they were maintained at a constant higher value . ADP-glucose and UDP-glucose did not show large fluctuations . Throughout embryo development fermentative activity did not play a major role . Obviously, specific mechanisms prevent seed tissues from becoming anoxic during development . The possible role of low oxygen on seed metabolism and on the control of seed development in legumes is discussed. Biochem J, 2002 Aug 1, 365(Pt 3), 629 - 38 Expression and regulation of the AMP-activated protein kinase-SNF1 (sucrose non-fermenting 1) kinase complexes in yeast and mammalian cells: studies using chimaeric catalytic subunits; Daniel T et al.; Mammalian AMP-activated protein kinase (AMPK) and yeast SNF1 (sucrose non-fermenting 1) kinase are members of a highly conserved protein kinase family that plays an important role in energy homoeostasis . AMPK and SNF1 kinase are heterotrimeric complexes consisting of a catalytic subunit and two regulatory subunits . We swapped the C-terminal regulatory domains of the catalytic subunits of AMPK (alpha) and SNF1 kinase (Snf1) and compared the expression and regulation of these chimaeric proteins with the native catalytic subunits in both mammalian and yeast cells . In mammalian cells, alpha1-Snf1 yielded a functional kinase complex following co-expression with the yeast regulatory subunits Sip2 and Snf4 . Unlike native AMPK, the alpha 1-Snf1 complex was not activated by the stresses that deplete intracellular AMP . Significantly, hyperosmotic stress led to the marked activation of both the alpha 1-Snf1 complex and AMPK, without a detectable change in adenine nucleotide levels, indicating that an alternative, non-AMP-dependent, pathway was responsible for activation . alpha1-Snf1 was able to restore growth of snf1 mutant yeast on raffinose and phosphorylated the transcriptional repressor protein Mig1 . Co-expression of the AMPK trimeric complex in yeast yielded an activity, increased by low glucose, that was similar to native SNF1 kinase . Importantly, expression of AMPK restored growth of a snf1 mutant on raffinose . Our results provide clues to the regulation of AMPK and SNF1 kinase and demonstrate that, in mammalian cells, there are at least two pathways that can activate AMPK, namely one that involves an increase in the AMP/ATP ratio and one that is independent of this ratio . In yeast, the glucose signalling pathway is able to activate AMPK, suggesting that the mammalian and yeast kinase pathways are conserved. Life Sci Space Res, 1980, 18, 89 - 100 The influence of life on the evolution of the atmosphere; Walker JC; The early history of life on earth may have been characterized by coevolution of microbial metabolism and atmospheric composition . Metabolic developments affected the composition of the atmosphere, and the resulting changes in the atmosphere stimulated the evolution of new metabolic capabilities . The first organisms eked out an existence by deriving energy from the fermentation of organic compounds abiotically synthesized . The abiotic source was meager, however, and when autotrophy arose, life was freed from its dependence on abiotic synthesis . The expanded level of biological activity made possible by autotrophy resulted in an increased rate of burial of reduced organic matter in sea floor sediments . The resultant drain on the concentration of electron donors in the biosphere caused a decline in the hydrogen content of the atmosphere . Biological productivity was limited by the supply of reduced compounds . This paper explores the biogeochemical circulation of electron donors in the primitive anaerobic ocean, concluding that their shortage was so critical as to provide strong selective pressure for the evolution of algal photosynthesis. Mol Microbiol, 2002 Apr, 44(1), 143 - 55 The roles of the polytopic membrane proteins NarK, NarU and NirC in Escherichia coli K-12: two nitrate and three nitrite transporters; Clegg S et al.; Two polytopic membrane proteins, NarK and NarU, are assumed to transport nitrite out of the Escherichia coli cytoplasm, but how nitrate enters enteric bacteria is unknown . We report the construction and use of four isogenic strains that lack nitrate reductase Z and the periplasmic nitrate reductase, but express all combinations of narK and narU . The active site of the only functional nitrate reductase, nitrate reductase A, is located in the cytoplasm, so nitrate reduction by these four strains is totally dependent upon a mechanism for importing nitrate . These strains were exploited to determine the roles of NarK and NarU in both nitrate and nitrite transport . Single mutants that lack either NarK or NarU were competent for nitrate-dependent anaerobic growth on a non-fermentable carbon source, glycerol . They transported and reduced nitrate almost as rapidly as the parental strain . In contrast, the narK-narU double mutant was defective in nitrate-dependent growth unless nitrate transport was facilitated by the nitrate ionophore, reduced benzyl viologen (BV) . It was also unable to catalyse nitrate reduction in the presence of physiological electron donors . Synthesis of active nitrate reductase A and the cytoplasmic, NADH-dependent nitrite reductase were unaffected by the narK and narU mutations . The rate of nitrite reduction catalysed by the cytoplasmic, NADH-dependent nitrite reductase by the double mutant was almost as rapid as that of the NarK+-NarU+ strain, indicating that there is a mechanism for nitrite uptake by E . coli that is in-dependent of either NarK or NarU . The nir operon encodes a soluble, cytoplasmic nitrite reductase that catalyses NADH-dependent reduction of nitrite to ammonia . One additional component that contributes to nitrite uptake was shown to be NirC, the hydrophobic product of the third gene of the nir operon, which is predicted to be a polytopic membrane protein with six membrane-spanning helices . Deletion of both NarK and NirC decreased nitrite uptake and reduction to a basal rate that was fully restored by a single chromosomal copy of either narK or nirC . A multicopy plasmid encoding NarU complemented a narK mutation for nitrite excretion, but not for nitrite uptake . We conclude that, in contrast to NirC, which transports only nitrite, NarK and NarU provide alternative mechanisms for both nitrate and nitrite transport . However, NarU might selectively promote nitrite ex-cretion, not nitrite uptake. Lett Appl Microbiol, 2002, 34(5), 323 - 8 Typing of Saccharomyces cerevisiae and Kloeckera apiculata strains from Aglianico wine; Caruso M et al.; AIMS: Kloeckera apiculata and Saccharomyces cerevisiae yeast species are dominant, respectively, at the early and at the following stages of wine fermentation . In the present study, PCR fingerprinting and NTS region amplification and restriction were applied as techniques for monitoring yeast population performing Aglianico of Vulture grape must fermentation . METHODS AND RESULTS: Thirty S . cerevisiae and 30 K . apiculata strains were typed by PCR fingerprinting with (GAC)5 and (GTG)5 primers and by complete NTS region amplification followed by restriction with HaeIII and MspI enzymes . S . cerevisiae strains generated two patterns with (GAC)5 primer, while (GTG)5 primer yielded a higher genetic polymorphism . Conversely, in K . apiculata Aglianico wine strains (GAC)5 and (GTG)5 primers generated the same profile for all strains . Restriction analysis of the amplified NTS region gave the same profile for all strains within the same species, except for one strain of S . cerevisiae . CONCLUSIONS: The PCR fingerprinting technique was useful in discriminating at strain level S . cerevisiae, particularly with the primer (GTG)5 . RFLP patterns generated from the NTS region of the two species can be more easily compared than the patterns resulting from PCR fingerprinting, thus RFLP is more suitable for the rapid monitoring of the species involved in different stages of fermentation . SIGNIFICANCE AND IMPACT OF THE STUDY: The molecular techniques used allow discrimination of S . cerevisiae at strain level and monitoring of the ratio of S . cerevisiae/K . apiculata during the fermentation process . Thus, their application can assure technological adjustments in a suitable time. J Appl Microbiol, 2002, 92(4), 602 - 10 Selenium incorporation into Saccharomyces cerevisiae cells: a study of different incorporation methods; Ponce de Leon CA et al.; AIMS: To study the effects of the selenium enrichment protocols in yeast at various points in the cell cycle, total selenium accumulation and the forms of selenium incorporated . METHODS AND RESULTS: The use of selenized yeast as enriched selenium supplements in human nutrition has become a topic of increasing interest over the last decade . Four enrichment procedures have been evaluated using sodium selenite as the selenium source: enrichment during the growth phase; enrichment at the non-growth phase, both of these at different selenium levels; enrichment by seeding in a fermentable carbon source (glucose); Se-enrichment with a non-fermentable carbon source (glycerol) . A nitric acid digestion of the yeast samples prepared under different conditions has been performed in order to evaluate the total selenium incorporated into the yeast cells . Also, an enzymatic digestion of the yeast samples with pepsin has been carried out as an initial step to begin the process of determining which of the different possible selenium species are formed . The cell count evaluations of the selenium-enriched yeast showed that the growth phase, seeding and the use of YEPG media is influenced by the addition of Se, while the non-growth phase is not . Total selenium incorporation studies showed that seeding the yeast permits more accumulation of selenium . Speciation studies of the enriched yeast showed that the growth phase increases the formation of L-Se-methionine . CONCLUSIONS: When the aim of enriching yeast with selenium is the formation of L-Se-methionine, the best enrichment procedure is using the growth phase with small concentrations of sodium selenite . SIGNIFICANCE AND IMPACT OF THE STUDY: The use of selenium supplements is widespread and most of the supplements use selenium-enriched yeast in their formulation . Studies made on supplements do not have the appropriate Se-species for optimal absorption in the human body . This study presents and compares methods for the best selenium yeast enrichment that could ultimately be used in selenium supplement formulations. Addiction, 2002 Apr, 97(4), 381 - 8 Fermenting fruit and the historical ecology of ethanol ingestion: is alcoholism in modern humans an evolutionary hangover? Dudley R. In the field of addiction research, the possibility of ancestral exposure to psychoactive compounds has generally been excluded . A paleobiological approach to the human diet, however, illustrates the potential utility of historical data in interpreting modern-day addictive behaviors . Low-level dietary exposure to ethanol via ingestion of fermenting fruit has probably characterized the predominantly frugivorous anthropoid lineage for about 40 million years . Potentially adaptive primate behaviors associated with the natural occurrence of ethanol include the olfactory use of ethanol plumes to localize fruit crops, the use of ethanol as an appetitive stimulant to facilitate rapid consumption of transient nutritional resources, and the physiological exploitation of the caloric benefits of ethanol . Such behavioral and energetic advantages probably pertain to all animal taxa that consume fermenting fruit, and may have been retained in modern humans in spite of considerable dietary diversification over the last several million years . In contemporary human environments, excessive consumption of ethanol would then represent maladaptive cooption of ancestrally advantageous behaviors given essentially ad libitum access to a compound otherwise found only within scarce nutritional substrates . Epidemiologically demonstrated health benefits of low-level alcohol consumption are consistent with an ancient and potentially adaptive exposure of primate frugivores to this most common of the psychoactive substances. Appl Biochem Biotechnol, 2001 Spring, 91-93, 777 - 85 Solid-state fermentation of phytase from cassava dregs; Hong K et al.; Phytases produced by numerous microorganisms and plants degrade phytic acid that has chelated with metal ions in food and feed . It is important to study phytase for the role of metal ions in nutrition of animals and humans as well as in the reduction of organic phosphate content of aqueous environment . This article reports on solid-state fermentation of phytase from a new substrate of cassava dregs . Large quantities of cassava dregs are produced in tropical areas as a byproduct of cassava starch processing . Protein and inorganic salts were found to be low in cassava dregs . Cassava dregs could be employed for phytase synthesis after the addition of a nitrogen source and mineral salts . Ammonium nitrate was the best nitrogen source among the nitrogen sources investigated, including beef extract, yeast extract, urea, ammonium nitrate, sodium nitrate, and ammonium sulfate . Sodium dodecyl sulfate promoted phytase production from cassava dregs . A maximum phytase yield of 6.73 U/g of dry mass was obtained . The obtained phytase was stable at feed-processing temperature, since 70% of initial enzyme activity was maintained after 30 min of treatment at 75 degrees C. Appl Biochem Biotechnol, 2001 Spring, 91-93, 729 - 37 Activity of xylose reductase from Candida mogii grown in media containing different concentrations of rice straw hydrolysate; Mayerhoff ZD et al.; Xylose reductase (XR) activity was evaluated in extracts of Candida mogii grown in media containing different concentrations of rice straw hydrolysate . Results of XR activity were compared to xylitol production and a similar behavior was observed for these parameters . Highest values of specific production and productivity were found for xylose reductase (35 U/g of cell and 0.97 U/{g of cell x h}, respectively) and for xylitol (5.63 g/g of cell and 0.13 g/{g of cell x h}) in fermentation conducted in medium containing 49.2 g of xylose/L . The maximum value of XR:XD ratio (1.82) was also calculated under this initial xylose concentration with 60 h of fermentation. Appl Biochem Biotechnol, 2001 Spring, 91-93, 681 - 9 Xylanase production by Aspergillus awamori in solid-state fermentation and influence of different nitrogen sources; Lemos JL et al.; The use of purified xylan as a substrate for bioconversion into xylanases increases the cost of enzyme production . Consequently, there have been attempts to develop a bioprocess to produce such enzymes using different lignocellulosic residues . Filamentous fungi have been widely used to produce hydrolytic enzymes for industrial applications, including xylanases, whose levels in fungi are generally much higher than those in yeast and bacteria . Considering the industrial importance of xylanases, the present study evaluated the use of milled sugarcane bagasse, without any pretreatment, as a carbon source . Also, the effect of different nitrogen sources and the C:N ratio on xylanase production by Aspergillus awamori were investigated, in experiments carried out in solid-state fermentation . High extracellular xylanolytic activity was observed on cultivation of A . awamori on milled sugarcane bagasse and organic nitrogen sources (45 IU/mL for endoxylanase and 3.5 IU/mL for beta-xylosidase) . Endoxylanase and beta-xylosidase activities were higher when sodium nitrate was used as the nitrogen source, when compared with peptone, urea, and ammonium sulfate at the optimized C:N ratio of 10:1 . The use of yeast extract as a supplement to the these nitrogen sources resulted in considerable improvement in the production of xylanases, showing the importance of this organic nitrogen source on A . awamori metabolism. Appl Biochem Biotechnol, 2001 Spring, 91-93, 655 - 69 Physiological aspects involved in production of xylanolytic enzymes by deep-sea hyperthermophilic archaeon Pyrodictium abyssi; Andrade CM et al.; Xylanases (EC 3.2.1.8) catalyze the hydrolysis of xylan, the major constituent of hemicellulose . The use of these enzymes could greatly improve the overall economics of processing lignocellulosic materials for the generation of liquid fuels and chemicals . The hyperthermophilic archaeon Pyrodictium abyssi, which was originally isolated from marine hot abyssal sites, grows optimally at 97 degrees C and is a prospective source of highly thermostable xylanase . Its endoxylanase was shown to be highly thermostable (over 100 min at 105 degrees C) and active even at 110 degrees C . The growth of the deep-sea archaeon P . abyssi was investigated using different culture techniques . Among the carbohydrates used, beech wood xylan, birch wood glucuronoxylan and the arabinoxylan from oats pelt appeared to be good inducers for endoxylanase and beta-xylosidase production . The highest production of arabinofuranosidase, however, was detected in the cell extracts after growth on xylose and pyruvate, indicating that the intermediate of the tricarboxylic acid cycle acted as a nonrepressing carbon source for the production of this enzyme . Electron microscopic studies did not show a significant difference in the cell surface (e.g., xylanosomes) when P . abyssi cells were grown on different carbohydrates . The main kinetic parameters of the organism have been determined . The cell yield was shown to be very low owing to incomplete substrate utilization, but a very high maximal specific growth rate was determined (micromax = 0.0195) at 90 degrees C and pH 6.0 . We also give information on the problems that arise during the fermentation of this hyperthermophilic archaeon at elevated temperatures. Appl Biochem Biotechnol, 2001 Spring, 91-93, 627 - 42 Influence of operating conditions and vessel size on oxygen transfer during cellulase production; Schell DJ et al.; The production of low-cost cellulase enzyme is a key step in the development of an enzymatic-based process for conversion of lignocellulosic biomass to ethanol . Although abundant information is available on cellulase production, little of this work has examined oxygen transfer . We investigated oxygen transfer during the growth of Trichoderma reesei, a cellulase-producing microorganism, on soluble and insoluble substrates in vessel sizes from 7 to 9000 L . Oxygen uptake rates and volumetric mass transfer coefficients (kLa) were determined using mass spectroscopy to measure off gas composition . Experimentally measured kLa values were found to compare favorably with a kLa correlation available in the literature for a non-Newtonian fermentation broth during the period of heavy cell growth. Appl Biochem Biotechnol, 2001 Spring, 91-93, 605 - 13 Effect of agitation and aeration on production of hexokinase by Saccharomyces cerevisiae; Silva DP et al.; A batch culture of Saccharomyces cerevisiae for the production of hexokinase was carried out in a 5-L fermentor containing 3 L of culture medium, which was inoculated with cell suspension (about 0.7 g/L), and left fermenting at 35 degrees C and pH 4.0 . The aeration and agitation were adjusted to attain k(L)a values of 15, 60, 135, and 230 h(-1) . The highest hexokinase productivity (754.6 U/{L x h}) and substrate-cell conversion yield (0.21 g/g) occurred for a k(L)a of 60 h(-1) . Moreover, the formation of hexokinase and cell growth are coupled events, which is in accordance with the constitutive character of this enzyme . Hexokinase formation for kLa > 60 h(-1) was not enhanced probably owing to saturation of the respiratory pathway by oxygen. Appl Biochem Biotechnol, 2001 Spring, 91-93, 353 - 65 Kinetics of ethanol fermentation with high biomass concentration considering the effect of temperature; Atala DI et al.; A model of ethanol fermentation considering the effect of temperature was developed and validated . Experiments were performed in a temperature range from 28 to 40 degrees C in continuous mode with total cell recycling using a tangential microfiltration system . The developed model considered substrate, product and biomass inhibition, as well as an active cell phase (viable) and an inactive (dead) phase . The kinetic parameters were described as functions of temperature. Appl Biochem Biotechnol, 2001 Spring, 91-93, 35 - 49 Detoxification of lignocellulose hydrolysates with ion-exchange resins; Nilvebrant NO et al.; Lignocellulose hydrolysates contain fermentation inhibitors causing decreased ethanol production . The inhibitors include phenolic compounds, furan aldehydes, and aliphatic acids . One of the most efficient methods for removing inhibiting compounds prior to fermentation is treatment of the hydrolysate with ion-exchange resins . The performance and detoxification mechanism of three different resins were examined: an anion exchanger, a cation exchanger, and a resin without charged groups (XAD-8) . A dilute acid hydrolysate of spruce was treated with the resins at pH 5.5 and 10.0 prior to ethanolic fermentation with Saccharomyces cerevisiae . In addition to the experiments with hydrolysate, the effect of the resins on selected model compounds, three phenolics (vanillin, guaiacol, and coniferyl aldehyde) and two furan aldehydes (furfural and hydroxymethyl furfural), was determined . The cation exchanger increased ethanol production, but to a lesser extent than XAD-8, which in turn was less effective than the anion exchanger . Treatment at pH 10.0 was more effective than at pH 5.5 . At pH 10.0, the anion exchanger efficiently removed both anionic and uncharged inhibitors, the latter by hydrophobic interactions . The importance of hydrophobic interactions was further indicated by a substantial decrease in the concentration of model compounds, such as guaiacol and furfural, after treatment with XAD-8. Appl Biochem Biotechnol, 2001 Spring, 91-93, 269 - 82 Enzymatic hydrolysis of ammonia-treated sugar beet pulp; Foster BL et al.; Sugar beet pulp is a carbohydrate-rich coproduct generated by the table sugar industry . Beet pulp has shown promise as a feedstock for ethanol production using enzymes to hydrolyze polymeric carbohydrates and engineered bacteria to ferment sugars to ethanol . In this study, sugar beet pulp underwent an ammonia pressurization depressurization (APD) pretreatment in which the pulp was exploded by the sudden evaporation of ammonia in a reactor vessel . APD was found to substantially increase hydrolysis efficiency of the cellulose component, but when hemicellulose- and pectin-degrading enzymes were added, treated pulp hydrolysis was no better than the untreated control. Appl Biochem Biotechnol, 2001 Spring, 91-93, 237 - 52 Ethanol production from lignocellulosic byproducts of olive oil extraction; Ballesteros I et al.; The recent implementation of a new two-step centrifugation process for extracting olive oil in Spain has substantially reduced water consumption, thereby eliminating oil mill wastewater . However, a new high sugar content residue is still generated . In this work the two fractions present in the residue (olive pulp and fragmented stones) were assayed as substrate for ethanol production by the simultaneous saccharification and fermentation (SSF) process . Pretreatment of fragmented olive stones by sulfuric acid-catalyzed steam explosion was the most effective treatment for increasing enzymatic digestibility; however, a pretreatment step was not necessary to bioconvert the olive pulp into ethanol . The olive pulp and fragmented olive stones were tested by the SSF process using a fed-batch procedure . By adding the pulp three times at 24-h intervals, 76% of the theoretical SSF yield was obtained . Experiments with fed-batch pretreated olive stones provided SSF yields significantly lower than those obtained at standard SSF procedure . The preferred SSF conditions to obtain ethanol from olives stones (61% of theoretical yield) were 10% substrate and addition of cellulases at 15 filter paper units/g of substrate. Food Addit Contam, 2002 Apr, 19(4), 387 - 99 Fate of patulin in the presence of the yeast Saccharomyces cerevisiae; Moss MO et al.; Patulin is known to become analytically non-detectable during the production of cider from contaminated apple juice . The fate of {14C}-labelled patulin during the alcoholic fermentation of apple juice was studied . Three commercial cider strains of Saccharomyces cerevisiae degraded patulin during active fermentative growth, but not when growing aerobically . The products of patulin degradation were more polar than patulin itself and remained in the clarified fermented cider . Patulin did not appear to bind to yeast cells or apple juice sediment in these model experiments . HPLC analysis of patulin-spiked fermentations showed the appearance of two major metabolites, one of which corresponded by both TLC and HPLC to E-ascladiol prepared by the chemical reduction of patulin using sodium borohydride . Using a diode array detector, both metabolites had a lambda(max) = 271 nm, identical to that of ascladiol . The nmr spectrum of a crude preparation of these metabolites showed signals corresponding to those of the E-ascladiol prepared chemically and a weaker set of signals corresponding to those reported in the literature for Z-ascladiol. Toxicol Sci, 2002 May, 67(1), 144 - 52 Evaluation of spinosad in a two-generation dietary reproduction study using Sprague-Dawley rats; Hanley TR Jr et al.; Spinosad, an insecticide derived from a naturally occurring bacterium via fermentation, represents a new class of insecticides acting by a novel mode of action . A dietary study was conducted in Sprague-Dawley rats in which groups of 30 rats/sex/dosage level were given diets that provided 0, 3, 10, or 100 mg spinosad/kg body weight/day, 7 days/week, for 2 successive generations . Following 10 weeks of dietary exposure, the P1 generation was mated twice to produce F1a and F1b litters . After weaning, groups of 30 rats/sex/dosage level were selected from the F1a litters, given diets containing spinosad for 12 weeks, and mated to produce the F2 generation . Dietary administration of spinosad to rats at a dosage of 100 mg/kg/day over 2 generations produced parental toxicity and effects on the offspring . Among adult males, body weights and weight gains were decreased 2-9% relative to controls, with P1 males more affected than P2 . Absolute and relative liver, kidney, heart, spleen, and thyroid weights were increased by from 12% to as much as 240% of control values . Histologic changes consistent with cationic amphiphilic compounds were noted in the kidneys, lungs, mesenteric lymph nodes, spleen, and thyroid of P1 and P2 males and females . In females given 100 mg/kg/day, though premating body weights were not affected, weight gains during the F1a and F1b gestation periods were depressed 15-16% . Increased incidences of dystocia, and vaginal bleeding and mortality occurred during parturition and lactation at 100 mg/kg/day . Effects on the offspring (decreased litter size and survival through day 4 of lactation) were limited to the high-dosage group . Signs indicative of poor maternal care noted in the pups (stomachs void of milk, cold, thin, etc.) were observed at 100 mg/kg/day . Early postnatal effects on the offspring were considered likely secondary to the effects in maternal animals around the time of parturition . At 100 mg/kg/day, weight gain in pups was depressed throughout lactation, with statistically significantly decreased weights noted toward the latter half of the lactation period . There were no treatment-related effects on adults or their offspring at 3 or 10 mg/kg/day in either generation . Based on these results, spinosad is not considered a selective reproductive toxicant, (i.e., no effects on reproductive parameters were noted below a level that produced toxicity in the adults) and the no observed effect level (NOEL) for both parental and reproductive/perinatal toxicity was 10 mg/kg/day. Antimicrob Agents Chemother, 2002 May, 46(5), 1218 - 25 Susceptibilities of Mycoplasma fermentans and Mycoplasma hyorhinis to membrane-active peptides and enrofloxacin in human tissue cell cultures; Nir-Paz R et al.; Mycoplasmas, which are bacteria that are devoid of a cell wall and which belong to the class Mollicutes, are pathogenic for humans and animals and are frequent contaminants of tissue cell cultures . Although contamination of cultures with mycoplasma can easily be monitored with fluorescent dyes that stain DNA and/or with molecular probes, protection and decontamination of cultures remain serious challenges . In the present work, we investigated the susceptibilities of Mycoplasma fermentans and Mycoplasma hyorhinis to the membrane-active peptides alamethicin, dermaseptin B2, gramicidin S, and surfactin by growth inhibition and lethality assays . In the absence of serum, the four peptides killed mycoplasmas at minimal bactericidal concentrations that ranged from 12.5 to 100 microM, but in all cases the activities were decreased by the presence of serum . As a result, under standard culture conditions (10% serum) only alamethicin and gramicidin S were able to inhibit mycoplasma growth (MICs, 50 microM), while dermaseptin B2 and surfactin were ineffective . Furthermore, 8 days of treatment of HeLa cell cultures experimentally contaminated with either mycoplasma species with 70 microM enrofloxacin cured the cultures of infection, whereas treatment with alamethicin and gramicidin S alone was not reliable because the concentrations and treatment times required were toxic to the cells . However, combination of alamethicin or gramicidin S with 70 microM enrofloxacin allowed mycoplasma eradication after 30 min or 24 h of treatment, depending on the mycoplasma and peptide considered . HeLa cell cultures experimentally infected with mycoplasmas should prove to be a useful model for study of the antimycoplasma activities of antibiotics and membrane-active peptides under conditions close to those found in vivo. FEBS Lett, 2002 Apr 10, 516(1-3), 172 - 8 The roles of hydrogenases 3 and 4, and the F0F1-ATPase, in H2 production by Escherichia coli at alkaline and acidic pH; Bagramyan K et al.; The hyc operon of Escherichia coli encodes the H2-evolving hydrogenase 3 (Hyd-3) complex that, in conjunction with formate dehydrogenase H (Fdh-H), constitutes a membrane-associated formate hydrogenlyase (FHL) catalyzing the disproportionation of formate to CO2 and H2 during fermentative growth at low pH . Recently, an operon (hyf) encoding a potential second H2-evolving hydrogenase (Hyd-4) was identified in E . coli . In this study the roles of the hyc- and hyf-encoded systems in formate-dependent H2 production and Fdh-H activity have been investigated . In cells grown on glucose under fermentative conditions at slightly acidic pH the production of H2 was mostly Hyd-3- and Fdh-H-dependent, and Fdh-H activity was also mainly Hyd-3-dependent . However, at slightly alkaline pH, H2 production was found to be largely Hyd-4, Fdh-H and F0F1-ATPase-dependent, and Fdh-H activity was partially dependent on Hyd-4 and F0F1-ATPase . These results suggest that, at slightly alkaline pH, H2 production and Fdh-H activity are dependent on both the F0F1-ATPase and a novel FHL, designated FHL-2, which is composed of Hyd-4 and Fdh-H, and is driven by a proton gradient established by the F0F1-ATPase. Crit Rev Biotechnol, 2002, 22(1), 65 - 84 Food-derived carbohydrates--structural complexity and functional diversity; Tharanathan RN; Carbohydrates are biomolecules abundantly available in nature . They are found in bewildering types ranging from simple sugars through oligo- and polysaccharides to glycoconjugates and saccharide complexes, each exhibiting characteristic bio-physiological and/or nutritional functions both in in vivo and in vitro systems . For example, their presence or inclusion in food dictates the texture (body) and gives desirable customer appeal (satisfaction), or their inclusion in the diet offers beneficial effects of great therapeutic value . Thus, carbohydrates are integrally involved in a multitude of biological functions such as regulation of the immune system, cellular signaling (communication), cell malignancy, antiinfection responses, host-pathogen interactions, etc . If starch is considered the major energy storage carbohydrate, the gums/mucilages and other non-starch carbohydrates are of structural significance . The most investigated properties of starch are its gelatinization and melting behavior, especially during food processing . This has led to the development of the food polymer science approach, which has enabled a new interpretive and experimental frame work for the study of the plasticizing influence of simple molecules such as water, sugars, etc . on food systems that are kinetically constrained . Starch, although considered fully digestible, has been challenged, and starch is found to be partly indigestible in the GI tract of humans . This fraction of starch-resisting digestion in vivo is known as resistant starch (RS) . The latter, due to its excellent fermentative capacity in the gut, especially yielding butyric acid is considered a new tool for the creation of fiber-rich foods, which are of nutraceutical importance . By a careful control of the processing conditions the content of RS, a man-made fiber, can be increased to as high as 30% . Arabinoxylans are the major endospermic cell wall polysaccharides of cereals . In wheat they are found complexed with ferulic acid esters, which after oxidative coupling in vivo mediated by H2O2 and peroxidases or even by photochemical means give cross linked diferuloyl derivatives . The latter confer strength and extensibility to the cell wall and offer resistance for digestibility by ruminants . They also help blocking of the ingress of pathogens . The ester bound ferulic acid after oxidation in vivo generates reactive oxygen species that contribute to the fragmentation of non-starch polysaccharides (hemicelluloses), and thereby reduces the product viscosity, a property seen during long-term storage of rice . In plant tissues, the arabinogalactans are implicated in such diverse functions as cell-cell adhesion, nutrition of growing pollen tubes, response to microbial infections, and also as markers of identity expressed in the terminal sequences of saccharide chains. Oncol Rep, 2002 May-Jun, 9(3), 613 - 6 Inhibition by long-term fermented miso of induction of gastric tumors by N-methyl-N'-nitro-N-nitrosoguanidine in CD (SD) rats; Ohara M et al.; The present study was designed to investigate the effects of fermented miso in the diet on the induction of gastric tumors by N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) in male CD (SD) rats . A total of 120 animals, 6 weeks of age, were divided into 6 groups and given MNNG (100 ppm) in the drinking water for 16 weeks . Starting 1 week before the carcinogen treatment the rats were fed a normal control MF solid diet, or the same diet containing 10% long-term fermented, medium- or short-term fermented miso, or 1% NaCl until the end of the MNNG exposure period . They were then maintained on the MF control diet and normal tap water until the autopsy time point at 52 weeks . The long-term fermented miso significantly reduced the size of the gastric tumors as compared with the other groups . The present results thus indicate that dietary supplementation with long-term fermented miso could act as a chemopreventive agent for gastric carcinogenesis. Int J Pharm, 2002 Apr 26, 237(1-2), 23 - 33 Susceptibility of the H2-receptor antagonists cimetidine, famotidine and nizatidine, to metabolism by the gastrointestinal microflora; Basit AW et al.; The H(2)-receptor antagonist ranitidine has previously been shown to be a substrate for colonic bacterial metabolism . The objective of the present study was to assess the in vitro stability of the other H(2)-receptor antagonists, cimetidine, famotidine and nizatidine, to colonic bacteria . One hundred milligrams of each drug were introduced into individual batch culture fermenters (100 ml) consisting of buffer medium inoculated with freshly voided human faeces (10% w/v) . Control experiments, equivalent drug quantities in buffer medium without the presence of faeces, were also run in parallel . Samples were removed at set time intervals over a 24 h period and were subsequently analysed by HPLC . A selection of the samples removed from the fermenters was also subjected to analysis by UV spectroscopy and mass spectrometry . Following an initial dissolution phase in the fermentation system, a marked decline in nizatidine concentration was noted over time with virtually no drug remaining after 12 h, thereby suggesting degradation and metabolism of the drug by colonic bacteria . No such decline in concentration was noted for cimetidine or famotidine or for any of the drugs in the control buffer systems . The metabolic reaction pathway for nizatidine was complex, although UV and mass spectrometry analysis indicated that metabolism was initiated via cleavage of an N-oxide bond within the molecule . These results in combination with those obtained from a previous study indicate that of the four commercially available H(2)-receptor antagonists, nizatidine and ranitidine are susceptible to metabolism by colonic bacteria, which in turn has ramifications for drug delivery and absorption. Appl Microbiol Biotechnol, 2002 Mar, 58(4), 539 - 42 A lipoxygenase inhibitor from Aspergillus niger; Sekhar RK et al.; A lipoxygenase-1 (LOX-1) inhibitor was isolated from the fermented broth of Aspergillus niger CFTRI 1105 . It was purified, using column and preparative thin layer chromatography . 1H NMR and GC-MS examination revealed the structure of the inhibitor to be 2-(2'-methyl, 4'-hydroxyphenyl), 2-(4"hydroxyphenyl)-propane with a molecular weight of 242 and the molecular formula C,6H18O2 . This bisphenol-derivative inhibitor shows 50% inhibition of soybean LOX-I at 0.98 mM concentration . The activity of this inhibitor was compared with commercial bisphenol A and its structural analogues, butylhydroxyanisole and butylhydroxytoluene in an attempt to understand the role of functional groups affecting lipoxygenase activity. Appl Microbiol Biotechnol, 2002 Mar, 58(4), 421 - 7 Epub 2002 Feb 08. Myco-protein from Fusarium venenatum: a well-established product for human consumption; Wiebe MG; Fusarium venenatum A3/5 was first chosen for development as a myco-protein in the late 1960s . It was intended as a protein source for humans and after 12 years of intensive testing, F . venenatum A3/5 was approved for sale as food by the Ministry of Agriculture, Fisheries and Food in the United Kingdom in 1984 . Today, myco-protein is produced in two 150,000 l pressure-cycle fermenters in a continuous process which outputs around 300 kg biomass/h . The continuous process is typically operated for around 1,000 h . One factor which has limited the length of production runs was the appearance of highly branched mutants in the population . Several factors affect the time of appearance of such mutants and a number of strategies for delaying their appearance have been investigated . After reduction of the RNA content, the fungal biomass is mixed with egg albumin and made into a variety of products . Consumption of these can lead to reduced blood cholesterol and to lower energy intake in a subsequent meal . E venenatum myco-protein is now used in products available in six European countries and there are plans for it to be sold in France, the United States and Germany. J Dairy Sci, 2002 Mar, 85(3), 632 - 41 Effect of supplemental tallow on performance of dairy cows fed diets with different corn silage:alfalfa silage ratios; Onetti SG et al.; A study was conducted to investigate the response to supplemental tallow of lactating cows fed basal diets with different alfalfa silage:corn silage ratios . We postulated that supplemental tallow will have decreasing negative effects on rumen fermentation, dry matter intake (DMI), and milk fat percentage as the dietary ratio of alfalfa silage:corn silage is increased . Eighteen Holstein cows averaging 134 +/- 14 d in milk were used in a replicated 6 x 6 Latin square design with 21-d periods . Treatments were arranged as a 2 x 3 factorial with 0 or 2% tallow (DM basis) and three forage treatments: 1) 50% of diet DM as corn silage, 2) 37.5% corn silage and 12.5% alfalfa silage, and 3) 25% corn silage and 25% alfalfa silage . Cows were allowed ad libitum consumption of a total mixed ration . Diets were formulated to contain 18% crude protein and 32% neutral detergent fiber . No fat x forage treatment interactions were observed . Fat supplemented cows had lower DMI and produced more milk with less milk fat content relative to non-supplemented cows . Concentration of trans-octadecenoic acids was higher in milk fat of tallow-supplemented cows . Tallow supplementation had no effect on ruminal pH and acetate:propionate ratio, but tended to decrease total volatile fatty acid (VFA) concentration in the rumen . Increasing the proportion of alfalfa silage increased DMI, milk fat percentage, and milk fat yield regardless of the fat content of the diet . Total VFA concentration and acetate:propionate ratio in the rumen were increased in response to higher levels of alfalfa in the diets . These results suggest that replacing corn silage with alfalfa silage did not alleviate the negative response of dairy cows to tallow supplementation at 2% of diet DM. J Dairy Sci, 2002 Mar, 85(3), 574 - 9 Effects of pH and pH fluctuations on microbial fermentation and nutrient flow from a dual-flow continuous culture system; Calsamiglia S et al.; Eight dual-flow continuous culture fermenters (1400 ml) were used in two consecutive periods to study the effects of pH and pH fluctuations on microbial fermentation and nutrient flow . Fermenters were maintained at 39 degrees C, with solid and liquid dilution rates of 5 and 10%/h, respectively, and fed continuously a 60% alfalfa hay and 40% concentrate diet (18.9% crude protein, 36.6% neutral detergent fiber, 17.6% acid detergent fiber) . Treatments were high pH (constant at 6.4); low pH (constant at 5.7); cycles of 4 h at pH 6.4 and 4 h at pH 5.7; and pH constant at 6.4, except for two 30-min drops per day to pH 5.7, followed by a 3-h slow recovery to pH 6.4 . The low pH (constant at 5.7) produced lower apparent dry matter, neutral detergent fiber and acid detergent fiber digestion, lower total and branch-chained volatile fatty acid concentrations, and lower acetate and higher propionate proportions than high pH (constant at 6.4) . There were no differences in these estimates between constant high pH and the two treatments that alternated high pH and low pH . The constant low pH reduced protein degradation and increased nonammonia N and dietary N flow compared with constant high pH . The pH treatments had no effect on bacterial N flow or efficiency of microbial protein synthesis . Flow of essential amino acids was highest for constant low pH and lowest for constant high pH . Results indicate that constant low pH reduced fiber and protein digestion and increased the flow of total and some individual amino acids . However, the effects of transitory decreases of pH were either small or insignificant with the conditions tested in this study. Biotechnol Bioeng, 2002 May 20, 78(4), 395 - 402 Equilibrium modeling of extractive enzymatic hydrolysis of penicillin G with concomitant 6-aminopenicillanic acid crystallization; Diender MB et al.; In the present downstream processing of penicillin G, penicillin G is extracted from the fermentation broth with an organic solvent and purified as a potassium salt via a number of back-extraction and crystallization steps . After purification, penicillin G is hydrolyzed to 6-aminopenicillanic acid, a precursor for many semisynthetic beta-lactam antibiotics . We are studying a reduction in the number of pH shifts involved and hence a large reduction in the waste salt production . To this end, the organic penicillin G extract is directly to be added to an aqueous immobilized enzyme suspension reactor and hydrolyzed by extractive catalysis . We found that this conversion can exceed 90% because crystallization of 6-aminopenicillanic acid shifts the equilibrium to the product side . A model was developed for predicting the equilibrium conversion in batch systems containing both a water and a butyl acetate phase, with either potassium or D-p-hydroxyphenylglycine methyl ester as counter-ion of penicillin G . The model incorporates the partitioning equilibrium of the reactants, the enzymatic reaction equilibrium, and the crystallization equilibrium of 6-aminopenicillanic acid . The model predicted the equilibrium conversion of Pen G quite reasonably for different values of pH, initial penicillin G concentration and phase volume ratio . The model can be used as a tool for optimizing the enzymatic hydrolysis . Waste Manag, 2002, 22(1), 1 - 5 Energy recovery from grass using two-phase anaerobic digestion; Yu HW et al.; Municipal solid wastes are major sources of air, water and soil contamination . There is a need for alternative waste management techniques to better utilize the waste and minimize its adverse environmental impact . A two-phase pilot-scale bio-fermentation system was used to evaluate the feasibility of producing methane from grass waste, a major constituent of solid wastes . The bi-phasic system consists of a solid phase and a methane phase . Leachate is re-circulated through the solid phase until a desired level of volatile fatty acid (VFA) is accumulated in the leachate . The leachate is then transferred to the methane reactor where the VFA is converted to methane . The results showed that 67% of the volatile solids in the waste can be converted into soluble chemical oxygen demand in a period of six months . The system produced an average of 0.15 m3 of methane per kg of grass . The average methane concentration in the produced gas was 71% . A mathematical model was developed to estimate the methane and carbon dioxide concentrations in the gas phase as a function of reactor properties. Rev Argent Microbiol, 2002 Jan-Mar, 34(1), 15 - 21 {Production of pectinases by Penicillium simplicissimum A3263 in an amaranth-seed flour medium}; Pastor MD et al.; The present work studies the production of pectinases using a strain of Penicillium simplicissimum A3263 and considering the influence of adding Amaranthus cruentus seed meal in a selected medium . We also considered the influence of aeration on enzyme production . Research was oriented towards the production of pectin lyase, the enzyme having the highest commercial value . This work was carried out in Erlenmeyer flasks in rotary shaker to select the medium and in a mechanically stirred fermentor to study aeration . The microorganism was developed as pellets of 1 mm diameter . Enzyme levels were of the order of 8216.21 pectin lyase units and 167.57 polygalacturonase units per gram of fungal biomass, respectively, using a medium containing 40 g/l of amaranth seed meal . As for the influence of aeration, it was determined that the higher values were obtained at 750 rpm corresponding to an oxygen absorption rate of 2691 ml O2/lh for an air flow of 1 l/l.min . The results obtained are considered very important in view of the fact that they exceeded in 550% those obtained by other authors. Wei Sheng Yan Jiu, 1998 Sep 30, 27(5), 351 - 3 {Preparation of morchella liquid by submersed fermentation and and analysis of its nutrient contents}; Yu Q; Natural morchella esculenta fruits were collected and its spores were isolated to get pure mycelium . Morchella liquid was prepared by submersed fermentation technology . The quantitative analysis showed that the liquid is relatively high in nutrient contents and several kinds of other essential components . The nutrient contents of the liquid are as follows (g/L): carbohydrates 63.1, reduced saccharide 25.6, protein 8.3, dry material 104.7, minerals 1.5 . There are also several high active micromaterials as follows: morchella lentinan 1.45 g/L, 10 kinds of essential and semiessential amino acid, 12 kinds of mineral (Fe, Zn Se, Cr, etc.) . The amount of soluble proteins are high in the liquid(one wider zone and four narrow zones on electrophoresis with molecular weights of 31.0 kd, 97.4 kd, 66.2 kd and 52.9 kd respectively) . The morchella liquid can be used as a new kind of base material for health foods and a flavoring agent. Yao Xue Xue Bao, 1998, 33(2), 92 - 7 {Evaluation of penicilloyl proteins of allergic impurity in gene engineering drugs}; Hu C et al.; It is well known that penicillins are necessary for prevention of contamination during cell culture or fermentation in the production of gene engineering drugs . As penicillins are easily combined with proteins forming allergic impurities, the benzyl penicilloyl proteins in aqueous solution, an evaluation system should be established for control of the gene engineering drugs . An enzyme linked immunosorbent assay (ELISA) with high specificity and high sensitivity in vitro, by which the conjugated penicillins of 0.3 ppm in a sample could be detected and a passive cutaneous anaphylaxis (PCA) test, a classical method for evaluation of allergic reaction in vivo are integral parts of this system . Some domestic gene engineering drugs such as erythropoietin (EPO), G-CSF, GM-CSF and 125SerIL-2 are evaluated with this system . No matter whether the expression products was produced in secretion manner or in inclusion body, it was possible that some residues of allergic impurities may remain in the finished products if an unsuitable process of isolation and purification was used . The maximum absolute content of the conjugated penicillins in a sample was up to about one per thousand. Se Pu, 1998 Nov, 16(6), 539 - 40 {Simultaneous determination of aspartame and amino acids in fermented milk beverages by HPLC}; Zhang W et al.; An RP-HPLC method for the simultaneous determination of aspartame and amino acids in fermented milk beverages were established . Samples were prepared by mixing with methanol at a ratio of 1:1 and centrifuged at 4,000 r/min for 15 min . Ten microL supernatant was moved into a sample tube, dried and derivatized according to PICO-TAG procedure developed by Waters . A Novapak C18 column (3.9 mm x 150 mm) was used instead of PICO TAG column and the gradient elution program was modified correspondingly . Column temperature was maintained at 38 degrees C and the components were detected at 254 nm . Linearity for aspartame at the range in 1-100 mg/L is A = 333 C + 20 with r = 0.9996 where A is the integrated area of chromatographic peak of aspartame and C is the corresponding mass concentration . Repeatability for 8 injections was tested and the RSD of 3.2% was obtained . The recovery of aspartame for 5 samples with the method ranged from 94.2% to 98.7%. J Physiol Anthropol Appl Human Sci, 2002 Jan, 21(1), 45 - 50 Effect of postprandial posture on digestion and absorption of dietary carbohydrate; Hirota N et al.; The effect of postprandial body posture on digestion and absorption of dietary carbohydrate were examined through breath hydrogen test on 6 female subjects . During the experiment, the participants either sat on a chair or lay on their backs for the first 4 hr (from 08:00 to 12:00) after eating the test breakfast meal . They then remained sedentary on a sofa for 6 hr (12:00 to 18:00) . Participants' end alveolar breath samples were collected for 10 hr (every 15 min from 08:00 to 12:30, and then every 30 min until 18:00) . The experiment was conducted on two consecutive days using a randomized, crossover study design . The results demonstrated that in the supine position orocecal transit time of the test meal was significantly slower than in the sitting position (260 +/- 21 min and 238 +/- 20 min, respectively, p < 0.01) . In addition, afternoon breath hydrogen excretion due to a partial malabsorption of dietary carbohydrate and its fermentation in the colon was significantly larger in the sitting position (144.0 +/- 24.1 ppm.hr) than in the supine position (110.0 +/- 26.1 ppm.hr, p < 0.05) . These results support the hypothesis that there was a marked effect of postprandial body posture on the function of the digestive system . The present findings suggest that the postprandial supine position is preferable to the sitting position for the digestion and absorption of dietary carbohydrate. J Ind Microbiol Biotechnol, 2002 Jan, 28(1), 17 - 20 Modulation of epothilone analog production through media design; Frykman S et al.; Recently, the epothilone polyketide synthase (PKS) was successfully introduced into a heterologous production host for the large-scale production of epothilone D . We have found that at least three other epothilones can also be produced as the major fermentation product of this recombinant strain by supplementation of specific substrates to the production media . Addition of acetate or propionate to the media results in modulation of the epothilone D:C ratio, whereas addition of L-serine with either acetate or propionate yields epothilone H(1) or H(2) as the major product . This strategy permits production of at least four novel epothilones by culturing a single host with a genetically modified epothilone PKS in various media. J Ind Microbiol Biotechnol, 2002 Jan, 28(1), 12 - 6 Production of a novel FK520 analog in Streptomyces hygroscopicus: improving titer while minimizing impurities; Regentin R et al.; FK520, also called ascomycin, is an immunosuppressive agent produced by Streptomyces hygroscopicus . Engineering the polyketide synthase genes of the parent strain generated novel FK520 analogs with the potential for improved in vivo stability . By replacing the acyl transferase (AT) domain in the polyketide synthase module 8 with an AT specific for methylmalonyl CoA (the rapamycin AT 3), the strain produced 13-desmethoxy-13-methyl-FK520 (13dmmFK520) . Process development and scale-up studies of this recombinant S . hygroscopicus strain producing 13dmmFK520 are described here . Production kinetics and compound stability in fermentation broth were significantly different compared to the native FK520 . Fermentation of the new strain resulted in the synthesis of a contaminating substance that co-purified with the 13dmmFK520 . To optimize 13dmmFK520 production and to facilitate purification, growth parameters and media development were examined . Although a medium was identified that increased product titers by ca . 300%, the ratio of impurity to product was doubled . Lower dissolved oxygen (20% compared to 50% and 80%) increased titers by 20% with no appreciable effect on the concentration of impurity . Increasing the fermentation pH from 6.0 to 6.5 did not change the 13dmmFK520 titer, but reduced the impurity-to-product ratio by approximately 450%. Anal Bioanal Chem, 2002 Jan, 372(2), 276 - 83 Epub 2001 Dec 15. Enzyme sensor array for the determination of biogenic amines in food samples; Lange J et al.; An enzyme sensor array for the simultaneous determination of the three biogenic amines (histamine, tyramine and putrescine) by pattern recognition using an artificial neural network and its application to different food samples is described . A combination of a monoamine oxidase, a tyramine oxidase and a diamine oxidase (with specific activities sufficient for rapid detection) are immobilised each on a separate screen-printed thick-film electrode via transglutaminase and glutaraldehyde to compare these cross-linking reagents with regard to their suitability . To calculate the amount of a specific biogenic amine, the raw data from multichannel software were transferred to a neural network . The sensor array takes 20 min to complete (excluding statistical data analysis) with only one extraction and subsequent neutralisation step required prior to sensor measurement . The lower detection limits with the enzyme sensor were 10 mg/kg for histamine and tyramine, and 5 mg/kg for putrescine with a linear range up to 200 mg/kg for histamine and tyramine and 100 mg/kg for putrescine . The application area of the enzyme sensor array was tested from fish to meat products, sauerkraut, beer, dairy products, wine and further fermented foods and compared with the data of conventional LC analyses (mean correlation coefficient: 0.854). Appl Microbiol Biotechnol, 2002 Mar, 58(3), 291 - 6 Epub 2002 Jan 12. High-level ethanol production from starch by a flocculent Saccharomyces cerevisiae strain displaying cell-surface glucoamylase; Kondo A et al.; A Strain of host yeast YF207, which is a tryptophan auxotroph and shows strong flocculation ability, was obtained from SaccharomYces diastaticus ATCC60712 and S . cerevisiae W303-1B by tetrad analysis . The plasmid pGA11, which is a multicopy plasmid for cell-surface expression of the Rhyzopus oryzae glucoamylase/alpha-agglutinin fusion protein, was then introduced into this flocculent yeast strain (YF207/pGA11) . Yeast YF207/pGA11 grew rapidly under aerobic condition (dissolved oxygen 2.0 ppm), using soluble starch . The harvested cells were used for batch fermentation of soluble starch to ethanol under anaerobic condition and showed high ethanol production rates (0.71 g h(-1) l(-1)) without a time lag, because glucoamylase was immobilized on the yeast cell surface . During repeated utilization of cells for fermentation, YF207/pGA11 maintained high ethanol production rates over 300 h . Moreover, in fed-batch fermentation with YF207/pGA11 for approximately 120 h, the ethanol concentration reached up to 50 g l(-1) . In conclusion, flocculent yeast cells displaying cell-surface glucoamylase are considered to be very effective for the direct fermentation of soluble starch to ethanol. Biotechnol Prog, 2002 Mar-Apr, 18(2), 269 - 75 Automated production support for the bioprocess industry; Lennox B et al.; This paper describes the application of Artificial Intelligence and Multivariate Statistical Techniques to two industrial fermentation systems . In the first example, an Expert System is shown to provide tighter control of an important process parameter . This is shown to lead to improved consistency of operation . In the second application, Principal Component Analysis is applied to a final stage fermentation production facility . The results presented indicate that the algorithm can provide concise indicators of process faults that can be presented to the operators to assist them in taking suitable corrective actions. Biotechnol Prog, 2002 Mar-Apr, 18(2), 234 - 9 Production of fructose and ethanol from sugar beet molasses using Saccharomyces cerevisiae ATCC 36858; Atiyeh H et al.; The production of enriched fructose syrups and ethanol from beet molasses using Saccharomyces cerevisiae ATCC 36858 was studied . In batch experiments with a total sugar concentration between 94.9 and 312.4 g/L, the fructose yield was above 93% of the theoretical value . The ethanol yield and volumetric productivity in the beet molasses media with sugar concentration below 276.2 g/L were in the range of 59-76% of theoretical value and between 0.48 and 2.97 g of ethanol/(L x h), respectively . The fructose fraction in the carbohydrates content of the produced syrups was more than 95% when the total initial sugar concentration in the medium was below 242.0 g/L . Some oligosaccharides and glycerol were also produced in all tested media . Raffinose and the produced oligosaccharides were completely consumed by the end of the fermentation process when the total initial sugar concentration was below 190.1 g/L . The glycerol concentration was below 16.1 g/L . The results could be useful for a potential industrial production of ethanol and high-fructose syrup from sugar beet molasses. Biol Chem, 2002 Feb, 383(2), 283 - 90 Cultivation in glucose-deprived medium stimulates mitochondrial biogenesis and oxidative metabolism in HepG2 hepatoma cells; Weber K et al.; In order to test the hypothesis that an imbalance between energy requirement and energy supply regulates mitochondrial genes and ultimately mitochondrial biogenesis, energy supply was challenged in HepG2 cells by withdrawal of glucose from the culture medium, making the cells exclusively dependent on mitochondrial ATP production . Such cells showed a 2-fold increase of cytochrome c oxidase activity, elevated levels of mitochondrial DNA, mitochondrial DNA encoded mRNAs and proteins, as well as the nuclear encoded mitochondrial transcription factor A . Lactate production was significantly reduced and glutamine was consumed as an alternative substrate for oxidative metabolism . Long-term adapted cells formed exclusively monolayers, while they normally grow in multilayers forming tumor spheroids . Also, long-term adapted cells proliferated significantly faster . No differences for the ATP/ADP ratio were observed, indicating that this is not the primary signal initiating the adaptative processes . These results show that mitochondrial biogenesis and oxidative metabolism are stimulated in HepG2 cells grown in the absence of fermentable glucose, probably in order to compensate for the diminished supply of glycolytic ATP. Int J Food Microbiol, 2002 Mar, 73(2-3), 363 - 6 The use of predictive microbiology by the Australian meat industry; Sumner J et al.; In Australia, the key regulatory body, the Meat Standards Committee (MSC) of the Agricultural and Resource Management Council of Australia and New Zealand (ARMCANZ) and the Export Meat Industry Advisory Council (EMIAC) have accepted in principle the usefulness of predictive microbiology for science-based regulation . The predictive microbiology approach is being used in a range of areas including hot-boning, distribution of meat, retailing of meat, fermentation, plant breakdowns and extended chilling regimes. Int J Syst Evol Microbiol, 2002 Mar, 52(Pt 2), 507 - 11 Weissella kimchii sp . nov., a novel lactic acid bacterium from kimchi; Choi HJ et al.; A gram-positive, catalase-negative, non-sporulating, facultatively anaerobic, short rod-shaped bacterium, with cells measuring 0.3-0.5 x 1-2 microm and designated strain CHJ3T, was isolated from partially fermented kimchi, a traditional Korean fermented vegetable food . The strain produced CO2 gas, D-lactate from glucose and dextran from sucrose and hydrolysed aesculin and arginine . It also fermented N-acetylglucosamine, amygdalin, arbutin, cellobiose, D-fructose, galactose, beta-gentiobiose, gluconate, D-glucose, maltose, D-mannose, salicin, sucrose and D-xylose . The G+C content of the DNA was 48.2 mol% . Phylogenetic analysis of 16S rRNA showed that strain CHJ3T is a member of the genus Weissella . The nearest phylogenetic relative of strain CHJ3T was Weissella confusa, with 16S rRNA similarity of 98.3% . However, strain CHJ3T could be differentiated from W . confusa on the basis of some phenotypic characteristics, analysis of whole-cell protein patterns and DNA-DNA hybridization data . These data suggest that strain CHJ3T be classified in the genus Weissella as a novel species, Weissella kimchii sp . nov . The type strain is CHJ3T (= KCCM 41287T = DSM 14295T = KCTC 3746T). Int J Syst Evol Microbiol, 2002 Mar, 52(Pt 2), 377 - 81 Comamonas koreensis sp . nov., a non-motile species from wetland in Woopo, Korea; Chang YH et al.; A bacterial strain, designated YH12T, was isolated from a wetland sample collected from Woopo, Republic of Korea, and characterized using a polyphasic approach . Analysis of 16S rDNA indicated that the isolate formed a monophyletic clade with the members of the genus Comamonas . The closest phylogenetic relative among the valid species was Comamonas testosteroni, with 96.6% 16S rDNA similarity . The chemotaxonomic properties of the wetland isolate supported its membership of the genus Comamonas, as it contained ubiquinone Q-8 as a major respiratory quinone and hexadecanoic, methylene-hexadecanoic and octadecenoic acids as major cellular fatty acids . The G+C content of the DNA was 66 mol% . The isolate is a gram-negative, non-pigmented, rod-shaped, oxidase- and catalase-positive, non-motile, non-endospore-forming and non-fermentative bacterium . The phenotypic properties of the isolate were compared with those of the type strains of Comamonas terrigena, C . testosteroni and Delftia acidovorans . A number of tests, including motility, can differentiate our isolate from related taxa . On the basis of the 16S rDNA phylogenetic, chemotaxonomic and phenotypic evidence given in this study, it is proposed that strain YH12T (= KCTC 12005T = IMSNU 11158T) be assigned as the type strain of a novel species of the genus Comamonas, Comamonas koreensis sp . nov. Int J Food Microbiol, 2002 Mar 25, 74(1-2), 57 - 64 The enumeration and identification of acetic acid bacteria from South African red wine fermentations; Du Toit WJ et al.; Acetic acid bacteria are microorganisms that can profoundly influence the quality of wine . Surprisingly, little research has been done on these microorganisms in the winemaking field . The object of this study was to investigate the occurrence of acetic acid bacteria in South African red wine fermentations and to identify the dominant species occurring . Acetic acid bacteria were isolated and enumerated from small-scale and commercial red must fermentations in 1998 and 1999, respectively . The initial occurrence of acetic acid bacteria in the must was shown to vary with cell numbers ranging from 10(6)-10(7) to 10(4)-10(5) cfu/ml for the 1998 and 1999 musts, respectively . The acetic acid bacteria decreased to 10(2)-10(3) cfu/ml in musts having a low pH (< or = 3.6), whereas in some musts having a high pH (> or = 3.7), the cell numbers increased during fermentation . During the process of cold soaking, the cell numbers of acetic acid bacteria also increased until inoculation with commercial wine yeast . Gluconobacter oxydans dominated in the fresh must and Acetobacter pasteurianus and A . liquefaciens during fermentation . This study showed that A . liquefaciens and A . hansenii were present in significant numbers, which has not been reported before. Z Naturforsch {C}, 2002 Jan-Feb, 57(1-2), 109 - 12 Improvement in xylitol production from sugarcane bagasse hydrolysate achieved by the use of a repeated-batch immobilized cell system; Carvalho W et al.; Candida guilliermondii cells were immobilized in Ca-alginate beads and used for xylitol production from concentrated sugarcane bagasse hydrolysate during five successive fermentation batches, each lasting 48 hours . The bioconversion efficiency of 53.2%, the productivity of 0.50 g/l x h and the final xylitol concentration of 23.8 g/l obtained in the first batch increased to 61.5%, 0.59 g/l x h and 28.4 g/l, respectively, in the other four batches (mean values), with variation coefficients of up to 2.3%. J Agric Food Chem, 2002 Apr 10, 50(8), 2267 - 73 Fermentation of cottonseed and other feedstuffs in cattle rumen fluid; Schneider IC et al.; Bovine rumen fluid was fermented anaerobically over 48 h with cottonseed, corn, alfalfa, or a mixture of these substrates in anaerobic mineral buffer . Samples taken at different incubation times were derivatized with n-butanol and subjected to gas chromatography and mass spectroscopy . No unusual fermentation end-products from the cottonseed substrate were detected . Cottonseed supported rumen fermentation at levels comparable to those of the other substrates . Major components were usually found in the decreasing order of acetate, propionate, butyrate, and valerate, although acetate and propionate concentrations decreased late in the alfalfa and mixed-feed fermentations, eventually allowing butyrate concentrations to exceed those of propionate . As expected, lactate was produced in high concentrations when corn was fermented . The minor components 2-methylpropionate, 2- and 3-methylbutyrate, phenylacetate, phenylpropionate, and caproate also accumulated, with their relative concentrations varying with the substrate . Succinate was produced in substantial amounts only when corn and alfalfa were fermented; it did not accumulate when cottonseed was the substrate . Samples containing cottonseed were derivatized and subjected to reversed-phase high-performance liquid chromatography, revealing that gossypol concentrations did not change during fermentation. Int J Food Microbiol, 2002 Mar 25, 74(1-2), 73 - 8 Thermal inactivation of poliovirus type 1 in water, milk and yoghurt; Strazynski M et al.; Loss of infectivity of poliovirus type 1, strain Sabin, during heating, freezing, and storage in water, milk and yoghurt was determined by plaque-titration in Vero cell cultures . The heating experiments simulated the conditions arising during the processing of milk and yoghurt, for example high-temperature heating (95 degrees C, 15 and 30 s), short-time pasteurization (72 degrees C, 15 and 30 s), long-time pasteurization (62 degrees C, 30 min), and yoghurt-fermentation (42 degrees C, 30 min and 180 min) . Only high-temperature heating, long-time pasteurization and short-time pasteurization for 30 s proved to be reliable methods of inactivating polioviruses present in water, milk and yoghurt completely . Short-time pasteurization for 15 s and the conditions of yoghurt-fermentation failed to cause complete inactivation of polioviruses . Additionally, polioviruses mixed in milk or yoghurt withstood these procedures with significantly lower reductions of infectivity than in water . Heating at 55 degrees C for 30 min resulted in complete inactivation of polioviruses, regardless of the suspending medium . The infectivity of polioviruses is scarcely affected by freezing (-20 degrees C, 30 min) and storage (24 days) at low temperatures (4 degrees C) and high humidity (a(w) = 0.99). In Vitro Cell Dev Biol Anim, 2002 Feb, 38(2), 79 - 85 Comparative PCR analysis for detection of mycoplasma infections in continuous cell lines; Uphoff CC et al.; Mycoplasma contamination of cell lines is one of the major problems in cell culturing . About 15-35% of all cell lines are infected with a limited number of mycoplasma species of predominantly human, swine, or bovine origin . We examined the mycoplasma contamination status in 495 cell cultures by polymerase chain reaction (PCR) assay, microbiological culture method, and deoxyribonucleic acid-ribonucleic acid (DNA-RNA) hybridization, and in 103 cell cultures by PCR and DNA-RNA hybridization, in order to determine the sensitivity and specificity of the PCR assay in routine cell culture . For those two cohorts, results for the three or two assays were concordant in 92 and 91% of the cases, respectively . The sensitivity (detection of true positives) of this PCR detection assay was 86%, and the specificity (detection of true negatives) was 93%, with positive and negative predictive values (probability of correct results) of 73 and 97%, respectively . PCR defined the mycoplasma status with 92% accuracy (detection of true positives and true negatives) . The mycoplasma contaminants were speciated by analyzing the PCR amplification fragment using several restriction enzymes . Most of the cultures (47%) were infected with Mycoplasma fermentans, followed by M . hyorhinis (19%), M . orale (10%), M . arginini (9%), Acholeplasma laidlawii (6%), and M . hominis (3%) . To sum up, PCR represents a sensitive, specific, accurate, inexpensive, and quick mycoplasma detection assay that is suitable for the routine screening of cell cultures. J Dairy Res, 2001 Nov, 68(4), 653 - 61 Polysaccharide production by kefir grains during whey fermentation; Rimada PS et al.; Fermentation of deproteinised whey with kefir grains CIDCA AGK1 was studied focusing on polysaccharide production from lactose . Kefir grains were able to acidify whey at different rates depending on the grain/whey ratio . During fermentation, kefir grains increased their weight and a water-soluble polysaccharide was released to the media . Exopolysaccharide concentration increased with fermentation time, reaching values of 57.2 and 103.4 mg/l after 5 days of fermentation in cultures with 10 and 100 g kefir grains/l, respectively . The polysaccharide fraction quantified after fermentation corresponded to the soluble fraction, because part of the polysaccharide became a component of the grain . Weight of kefir grains varied depending on the time of fermentation . Polysaccharide production was affected by temperature . Although the highest concentration of polysaccharide in the media was observed at 43 degrees C at both grain/whey ratios, the weight of the grains decreased in these conditions . In conclusion, kefir grains were able to acidify deproteinised whey, reducing lactose concentration, increasing their weight and producing a soluble polysaccharide. Nutr Hosp, 2002 Feb, 17 Suppl 1, 17 - 22 {Fiber in the treatment of obesity and its comorbidities}; Trallero Casanas R; A considerable part of the health-care significance of obesity lies in its co-morbidity . Obesity is associated with high blood pressure, type 2 diabetes mellitus and dyslipidaemia, representing both individually and when taken together risk factors for cardiovascular disease, the main cause of death in industrialized countries . The influence of fibre consumption on body weight, the risk of type 2 diabetes and cardiovascular disease has been shown in important epidemiological surveys, but the mechanisms whereby fibre exercises this protective effect have not been definitively clarified . For the most part, the ability of fibre to slow down the process of absorption of nutrients by modifying the metabolic response to intake, its ability to increase the excretion of biliary acid and the metabolic effects of the short-chain fatty acids produced in the bacterial fermentation of fibre have all focussed the attention of the working hypotheses analyzed in the studies published in the last few years . This review is intended to identify the importance of fibre for the prevention and treatment of obesity and its comorbidities on the basis of the available evidence at present. Curr Microbiol, 2002 May, 44(5), 374 - 8 Biogenic amine production by Oenococcus oeni; Guerrini S et al.; The biogenic amine-producing capability of several Oenococcus oeni strains, originally isolated from different Italian wines, was determined . The amine-producing capability was quali-quantitatively variable among the strains: out of the 44 strains investigated under optimal growth conditions, more than 60% were able to produce histamine, at concentrations ranging from 1.0 to 33 mg/L, and about 16% showed the additional capability to form both putrescine and cadaverine, to different extents and variable relative proportions . The amine-producing behavior of the strains was confirmed under stress culture conditions, while performing malolactic fermentation . In wine, one randomly chosen strain was very effective in forming putrescine from ornithine . The formation of putrescine from arginine by some strains has been also demonstrated . Consequently, O . oeni can really and significantly contribute to the overall biogenic amine content of wines . Practical consequences of these findings are discussed. Curr Microbiol, 2002 May, 44(5), 341 - 9 Isolation from a shea cake digester of a tannin-tolerant Escherichia coli strain decarboxylating p-hydroxybenzoic and vanillic acids; Chamkha M et al.; A facultatively anaerobic, mesophilic, Gram-negative, non-motile, non-sporulated bacterium, designated strain C2, was isolated from an anaerobic digester fed with shea cake rich in tannins and aromatic compounds and previously inoculated with anaerobic sludge from the pit of a slaughterhouse, after enrichment on tannic acid . The straight rods occurred singly or in pairs . Strain C2 fermented numerous carbohydrates (fructose, galactose, glucose, lactose, mannose, maltose, melibiose, raffinose, rhamnose, ribose, saccharose, sorbitol, trehalose, and xylose) and peptides (Biotrypcase, Casamino acids, and yeast extract), producing acid and gas, and had a G + C content of 51.6 +/- 0.1 mol % . Strain C2 was very closely related to Escherichia coli (= DSM 30083(T)) phylogenetically (similarity of 99%), genotypically (DNA homology of 79%), and phenotypically . The isolate tolerated tannic acid (hydrolyzable tannin) and decarboxylated by non-oxidative decarboxylation only p-hydroxybenzoic and vanillic acids to their corresponding phenol and guaicol, under anaerobic and aerobic conditions without further degradation . Adding glucose increased growth and the rate of conversion . High concentrations of p-hydroxybenzoic acid or vanillic acid inhibited growth, and decarboxylation could not occur completely, suggesting phenol toxicity . In contrast, the type strain of E . coli cannot metabolize p-hydroxybenzoic and vanillic acids, anaerobically or aerobically, with or without glucose added. Drug Dev Ind Pharm, 2002, 28(2), 157 - 64 Polysaccharides as excipients for colon-specific coatings . Permeability and swelling properties of casted films; Cavalcanti OA et al.; Oligosaccharides such as inulin (In) and polysaccharides such as galactomannans, combined with polymethacrylates on isolated films for film coatings, were obtained from aqueous-based solvents and investigated as potential vehicles for colonic drug delivery . These compositions, which are susceptible to fermentation by colonic microflora, constitute promising excipients for the development of new colon-specific therapeutic systems . The characteristics of several compositions have been demonstrated in permeability and swelling studies on isolated films composed of a polymethacrylate associated with In or galactomannans of mesquite seed gum (MSG) . Results reported prove a dependency of the properties of mixed films on the polymethacrylate-polysaccharide concentration ratio and on the composition of the dissolution media . An increase in permeability through the mixed films was observed in a simulated colonic environment for the following compositions: Eudragit RS30D-MSG 70:30 w/w; Eudragit RS30D-In 90:10 w/w; Eudragit RS30D-In 76:24 w/w. J Org Chem, 2002 Apr 5, 67(7), 2257 - 62 Biosynthetic precursors of the lipase inhibitor lipstatin; Schuhr CA et al.; Three putative intermediates in the biosynthesis of the lipase inhibitor lipstatin were synthesized in stable isotope-labeled form and were added to fermentation cultures of Streptomyces toxytricini . Biosynthetic lipstatin was isolated and analyzed by NMR spectroscopy . {3,10,11,12-(2)H}-(3S,5Z,8Z)-3-hydroxytetradeca-5,8-dienoic acid (9) was shown to serve as a direct biosynthetic precursor of lipstatin . {7,8-(2)H(2)}Hexylmalonate (11) was also incorporated into lipstatin, albeit at a relatively low rate . The leucine moiety of {(13)C-formyl,(15)N}-N-formylleucine (10) was diverted to lipstatin under loss of the (13)C-labeled formyl residue. Org Lett, 2002 Apr 4, 4(7), 1123 - 6 Structure, stereochemistry, and biological activity of integramycin, a novel hexacyclic natural product produced by Actinoplanes sp . that inhibits HIV-1 integrase; Singh SB et al.; {structure: see text} HIV-1 integrase is a critical enzyme for viral replication, and its inhibition is an emerging target for potential antiviral chemotherapy . We have discovered a novel inhibitor, integramycin, from screening of fermentation extracts using an in vitro assay . Integramycin possesses a hexacyclic ring system and exhibited an IC50 value of 4 microM against HIV-1 integrase (strand transfer) . The isolation, structure elucidation, stereochemistry, conformation, and biological activity has been described. Protein Expr Purif, 2002 Apr, 24(3), 453 - 9 High-level expression and secretion of recombinant mouse endostatin by Escherichia coli; Xu R et al.; The expression of murine endostatin was achieved by placing its gene downstream of an alkaline phosphatase gene (phoA) promoter . To ensure proper folding and secretion of the recombinant protein, the mouse endostatin was fused with alkaline phosphatase signal peptide . SDS/polyacrylamide gel electrophoresis analysis of the culture medium of recombinant Escherichia coli cells revealed that endostatin was efficiently secreted . The signal peptide was efficiently cleaved during secretion as demonstrated by N-terminal amino acid sequencing . The maximum yield of secreted endostatin during fermentation was 40 mg/liter . Up to 28 mg of endostatin was purified from 1 liter of cell culture broth . The biological activity of recombinant protein was tested in a bovine aortic endothelial (BAE) cell proliferation assay . The recombinant endostatin inhibited the growth of BAE cells stimulated by basic fibroblast growth factor, and its ED50 was comparable to that from a previous report . Flow cytometric measurements of BAE cells cultivated in medium with endostatin demonstrated a cell cycle arrest mainly in the G0/G1 phase and a decrease in the S phase . Adv Exp Med Biol, 2002, 504, 217 - 26 Mycotoxins and fermentation--beer production; Wolf-Hall CE et al.; Along with food safety issues due to mycotoxins, the effects of Fusarium infections on malt and beer quality can be disastrous . While some of the Fusarium head blight mycotoxins, such as DON, present in infected barley may be lost during steeping, the Fusarium mold is still capable of growth and mycotoxin production during steeping, germination and kilning . Therefore, detoxification of grain before malting may not be practical unless further growth of the mold is also prevented . Methods for reducing the amount of mold growth during malting are needed . Physical, chemical and biological methods exist for inhibiting mold growth in grain . Irradiation is a promising means for preventing Fusarium growth during malting, but its effects on malt quality and mycotoxin production in surviving mold need to be evaluated in more detail . Chemical treatments such as ozonation, which do not leave chemical residues in beer, also appear to be promising . Although biological control methods may be desirable, the effects of these inhibitors on malt and beer quality require further investigation . In addition, storage studies are needed to determine the effect of biological control on Fusarium viability and malt quality . It may also be possible to incorporate detoxifying genes into fermentation yeasts, which would result in detoxification of mycotoxins present in wort . Development of these types of technological interventions should help improve the safety of products, such as beer, made from Fusarium infected grain. Adv Exp Med Biol, 2002, 504, 205 - 16 Effects of processing on zearalenone; Ryu D et al.; Zearalenone (ZEN), a common contaminant of all major cereal grains worldwide, is produced by some plant pathogenic molds including Fusarium graminearum and F . culmorum . The biological activity of this mycotoxin is mainly attributed to its estrogenic activity that modulates/disrupts endocrine function in animals and possibly humans . Efforts have been made to reduce the level of ZEN by various chemical, physical, and biological processing methods . Some chemical treatments were shown to be effective in reducing zearalenone content in artificially or naturally contaminated foods . During physical processing, the fate of ZEN depended on its distribution in the food matrix and its chemical properties such as heat stability and solubility . For example, wet milling of contaminated corn resulted in starch that was essentially toxin-free . In contrast, animal feed fractions such as bran and germ, by-products of the wet milling process, tended to concentrate ZEN . Extrusion cooking, a complex process where food is subjected to heat, high pressures and shear stress, reduced ZEN levels in food as well as its estrogenic activity . Fermentation of foods with bacteria and yeast resulted in reduction in ZEN levels . However, fermentation can result in the conversion of ZEN to more potent derivatives such as cc-zearalenol . Further efforts are needed to identify effective methods for removing/detoxifying ZEN in foods. J Clin Microbiol, 2002 Apr, 40(4), 1225 - 9 Escherichia coli ehl1 gene-positive serotype O18ac:H31 associated with an outbreak of diarrhea in a neonatal nursery in Neuquén City, Argentina; Chinen I et al.; Between 9 October and 12 November 1996, an outbreak of bloody diarrhea occurred in the neonatal nursery ward of the Policlinico Neuquen, in Neuquen, a city in the southwestern region of Argentina . Seven patients of the intermediate care unit were affected . Isolates of Escherichia coli O18ac:H31 that were non-lactose and -sorbitol fermenting were recovered from outbreak cases . Although the strains were negative for a number of virulence factors typically found in diarrheagenic groups of E . coli, all isolates from the present neonatal outbreak possessed the enterohemolysin gene, ehl1 . All isolates showed resistance to the antibiotics ampicillin and chloramphenicol . These isolates showed a low adherence property in HeLa cells without any recognizable pattern . In order to evaluate the outbreak dissemination in the neonatology ward, a prevalence study was conducted on 13 November . Stool specimens were obtained from 16 neonates hospitalized in the sector and from 33 medical staff members . E . coli isolates with identical biochemical characteristics of the outbreak strain were recovered from 11 of 16 inpatients and from 4 of 33 staff members during the prevalence study . A total of 15 E . coli strains recovered both from the outbreak and the prevalence study were processed by random amplified polymorphic DNA (RAPD)-PCR and pulsed-field gel electrophoresis (PFGE) . By RAPD-PCR 14 of 15 strains showed patterns with 85 to 100% similarity, and by PFGE these strains were identical, each showing a unique pattern with 15 bands between 40 and 400 kb . One strain isolated from a nurse during the prevalence study presented a pattern not related to the others, and this was characterized as E . coli O81:HNM resistant to ampicillin only and negative for all the virulence factors studied . This outbreak occurred despite strict regulations in place to prevent cross-infection in the hospital . Postoutbreak prevalence studies were performed weekly thereafter without detecting new cases. Comp Biochem Physiol B Biochem Mol Biol, 2002 Apr, 131(4), 613 - 20 Xylose as a nectar sugar: from biochemistry to ecology; Jackson S et al.; Studies of nectar sugar composition in the Proteaceae, an ancient southern hemisphere plant family, have demonstrated that xylose comprises up to 39% of nectar sugar in two genera, Protea and Faurea, and may therefore represent a substantial fraction of the energy available to pollinators of these plants . Although insect and bird pollinators of Protea species are averse to xylose, mice (Aethomys namaquensis) will drink pure xylose, which is metabolized either by gut bacteria or by the mouse tissues . In the form of xylan polymers, the pentose sugar D-xylose is a structural component of plant cell walls, and there is considerable biotechnological interest in xylose fermentation . Bacteria and yeasts convert D-xylose to D-xylulose and thence via the pentose phosphate pathway to fructose-6-phosphate, which is either oxidized or fermented to ethanol . Gut symbionts of rodent pollinators may be analogous to ruminal xylose-metabolizing bacteria . The presence of xylose in Protea and Faurea nectar remains puzzling in view of pollinator aversions: even for rodent pollinators, it is the least preferred nectar sugar . In the generalized pollination systems of the Proteaceae, a coevolutionary explanation for nectar xylose as an attractant for mammalian pollinators is probably less likely than one involving plant physiology, with xylose in phloem sap being secreted passively into the nectar. Arch Tierernahr, 2001, 54(4), 341 - 7 Research note: a method for studying local differences in ruminal fermentation in dairy cattle; Tafaj M et al.; A method was developed for studying local differences in ruminal fermentation . The developed sampler consisted of an acrylic glass container (460 cm3) with an aperture for digesta sampling, which could be opened and closed by the scaled "T" rod . The scale was a reference for defined rumen layers: top, middle, 5 to 10 cm and 25 to 35 cm beneath the top of particles mat, respectively, and bottom 5 to 10 cm above the rumen floor . The repeatability of the method was proved in two rumen cannulated cows . Particle/fluid ratio, pH and sample amount were measured 2 to 2 1/2 h after morning feeding in four replicates each day (over 5 days), rumen layer and animal . No significant differences between replicates were observed . The coefficients of variation (CV) of the particle/fluid ratio varied between 8.7% and 13.6% . Top layer had higher CV than middle and bottom layer . CV of pH ranged between 0.59% and 1.27% . The developed method of sampling showed satisfactory repeatability for investigation of digesta properties and fermentation in different rumen layers. Rapid Commun Mass Spectrom, 2002, 16(8), 743 - 8 Analysis of saccharides in beer samples by flow injection with electrospray mass spectrometry; Mauri P et al.; Saccharides in foods play important roles, as they are essential substrates for fermentation processes . In brewing, the concentration of maltooligosaccharides influences the characteristics of beers and therefore their determination is of great practical interest . Electrospray ionization mass spectrometry (ESI-MS) was applied to identify and characterise maltooligosaccharides in beer samples . The effects due to different cation concentrations and dilution of samples were studied . Furthermore, quantitative analyses of maltooligosaccharides by means of flow-injection ESI-MS (FI/ESI-MS) of 1-microL beer samples (diluted 1000-fold) are described . Biotechnol Bioeng, 2002 May 5, 78(3), 270 - 9 Pilot-scale process sensitivity studies for the scaleup of a fungal fermentation for the production of pneumocandins; Pollard DJ et al.; The filamentous fungus Glarea lozoyensis produces a novel, pharmaceutically important pneumocandin (B(0)) that is used to synthesize a lipopeptide which demonstrates cidal activity against clinically relevant pathogens . A range of unwanted pneumocandin analogs are also produced by the organism . To maintain the unwanted impurities to acceptable levels upon scaleup, a good understanding of the impact of chemical and physical environment on the cell physiology is required, which benefits downstream processing . Pilot-scale studies were performed to determine the impact of dissolved oxygen, temperature, pH, and carbon dioxide on the process . Experiments included multiple fermenters (up to seven) at 0.07 and 0.8 m(3) scale using single source medium sterilization and inoculum . Gas blending was used to separate effects of dissolved oxygen from agitation . The process was significantly influenced by dissolved oxygen level . The critical dissolved oxygen tension (C(crit)) for growth was below 2% air saturation . The C(crit) for production of pneumocandin B(0) was 20% air saturation, with a significant reduction of the specific production rate below this value . In contrast, low dissolved oxygen levels produced a substantial increase of pneumocandins B(1), B(5), and E(0), while high dissolved oxygen levels produced a disproportionate increase of D(5) . This sensivity to dissolved oxygen was independent of agitation within a power range of 2-15 kW/m(3) . Broth viscosity was impacted below 10% dissolved oxygen, suggesting an effect on morphology . The process was shown to be sensitive to temperature but relatively insensitive to pH and carbon dioxide (in the exhaust gas) within the ranges studied . This scaledown analysis explained phenomena seen at pilot scale and helped define operating boundary conditions for successful scale up to 19 m(3) . Biotechnol Bioeng, 2002 May 5, 78(3), 237 - 50 Albumin recovery with centrifugal adsorption technology (CAT); Peet DJ et al.; Centrifugal adsorption technology (CAT) is a new compact, countercurrent technology for efficient adsorption from large liquid streams by using adsorbent particles in the micrometer range . CAT seems particularly suited for the recovery of macromolecules at low concentrations, because the small particle dimensions lead to fast mass transfer rates . In this work, the potential of CAT for protein recovery is studied by model and experiment . A predictive model for the separation performance of CAT is presented, incorporating mass transfer resistance and axial dispersion transport in the liquid and the adsorbent phases . The model calculations were compared to experimental data for the adsorption of bovine serum albumin (BSA) on a standard commercial anion-exchange resin with particle diameter d(p) = 50 microm in a pilot-scale CAT apparatus . The model calculations accurately predicted the separation efficiency of CAT . The experimental set-up is shown to be mass transfer limited for the conducted experiments, which agrees with the model predictions . The model was also used to estimate the dimensions and performance of a CAT apparatus for the large-scale recovery of human serum albumin (HSA) from fermentation broth at the scale of 40 tons per year . The resulting equipment dimensions proved to be very small indeed, making CAT a potentially very attractive technology . Appl Environ Microbiol, 2002 Apr, 68(4), 2018 - 25 The Awa1 gene is required for the foam-forming phenotype and cell surface hydrophobicity of sake yeast; Shimoi H et al.; Sake, a traditional alcoholic beverage in Japan, is brewed with sake yeasts, which are classified as Saccharomyces cerevisiae . Almost all sake yeasts form a thick foam layer on sake mash during the fermentation process because of their cell surface hydrophobicity, which increases the cells' affinity for bubbles . To reduce the amount of foam, nonfoaming mutants were bred from foaming sake yeasts . Nonfoaming mutants have hydrophilic cell surfaces and no affinity for bubbles . We have cloned a gene from a foam-forming sake yeast that confers foaming ability to a nonfoaming mutant . This gene was named AWA1 and structures of the gene and its product were analyzed . The N- and C-terminal regions of Awa1p have the characteristic sequences of a glycosylphosphatidylinositol anchor protein . The entire protein is rich in serine and threonine residues and has a lot of repetitive sequences . These results suggest that Awa1p is localized in the cell wall . This was confirmed by immunofluorescence microscopy and Western blotting analysis using hemagglutinin-tagged Awa1p . Moreover, an awa1 disruptant of sake yeast was hydrophilic and showed a nonfoaming phenotype in sake mash . We conclude that Awa1p is a cell wall protein and is required for the foam-forming phenotype and the cell surface hydrophobicity of sake yeast. Appl Environ Microbiol, 2002 Apr, 68(4), 1604 - 9 Reduced oxidative pentose phosphate pathway flux in recombinant xylose-utilizing Saccharomyces cerevisiae strains improves the ethanol yield from xylose; Jeppsson M et al.; In recombinant, xylose-fermenting Saccharomyces cerevisiae, about 30% of the consumed xylose is converted to xylitol . Xylitol production results from a cofactor imbalance, since xylose reductase uses both NADPH and NADH, while xylitol dehydrogenase uses only NAD(+) . In this study we increased the ethanol yield and decreased the xylitol yield by lowering the flux through the NADPH-producing pentose phosphate pathway . The pentose phosphate pathway was blocked either by disruption of the GND1 gene, one of the isogenes of 6-phosphogluconate dehydrogenase, or by disruption of the ZWF1 gene, which encodes glucose 6-phosphate dehydrogenase . Decreasing the phosphoglucose isomerase activity by 90% also lowered the pentose phosphate pathway flux . These modifications all resulted in lower xylitol yield and higher ethanol yield than in the control strains . TMB3255, carrying a disruption of ZWF1, gave the highest ethanol yield (0.41 g g(-1)) and the lowest xylitol yield (0.05 g g(-1)) reported for a xylose-fermenting recombinant S . cerevisiae strain, but also an 84% lower xylose consumption rate . The low xylose fermentation rate is probably due to limited NADPH-mediated xylose reduction . Metabolic flux modeling of TMB3255 confirmed that the NADPH-producing pentose phosphate pathway was blocked and that xylose reduction was mediated only by NADH, leading to a lower rate of xylose consumption . These results indicate that xylitol production is strongly connected to the flux through the oxidative part of the pentose phosphate pathway. Biotechnol Appl Biochem, 2002 Apr, 35(Pt 2), 133 - 40 Production of non-alcoholic beer using free and immobilized cells of Saccharomyces cerevisiae deficient in the tricarboxylic acid cycle; Navratil M et al.; Production of non-alcoholic beer using Saccharomyces cerevisiae has been studied . Non-recombinant mutant strains with a defect in the synthesis of tricarboxylic-acid-cycle enzymes were used and applied in both free and pectate-immobilized form, using both batch and packed-bed continuous systems . After fermentation, basic parameters of the beer produced by five mutant strains were compared with a standard strain of brewing yeast . Results showed that the beer prepared by mutant yeast cells was characterized by lower levels of total alcohols, with ethanol concentrations between 0.07 and 0.31% (w/w) . The organic acids produced, especially lactic acid, in concentrations up to 1.38 g x l(-1) had a strong protective effect on the microbial stability of the final product and thus the usual addition of lactic acid could be omitted . Application of the yeast mutants appears to be a good alternative to the classical methods for the production of non-alcoholic beer. Ned Tijdschr Tandheelkd, 1993 Jul, 100(7), 320 - 2 {Dietary habits of Turkish and Moroccan children in the Netherlands}; Meulmeester JF et al.; Migration might have considerable implications for nutrition and consequently for health . Because of the higher demands of growth, children are especially vulnerable to changes in nutrition . In order to get an insight into the need for a special nutrition policy for Turkish ad Moroccan children, the dietary habits and food consumption of 8-year-old Turkish and Moroccan children in The Netherlands were assessed . The results showed that the diet of these children was more compatible with the Dutch recommendations for a prudent diet than the diet of Dutch children from a similar socio-economic background . Quite a few of the basic food items Turkish and Moroccan children are accustomed to eat, like rice, cereals, bread, pulses and vegetables, contain valuable nutrients . So, an increased consumption of these items was recommended . The intake of calcium and riboflavin of Turkish and Moroccan children was low and so the use of dairy products--preferably in fermented form like yoghurt--was recommended . This would also enhance the Vitamin D status. Biochem Pharmacol, 2002 Mar 1, 63(5), 1019 - 26 Dimerumic acid as an antioxidant from the mold, Monascus anka: the inhibition mechanisms against lipid peroxidation and hemeprotein-mediated oxidation; Taira J et al.; This study aimed to investigate the antioxidant mechanism of dimerumic acid isolated as the active component with a radical scavenging action from the mold Monascus anka, traditionally used for the fermentation of foods . Dimerumic acid inhibited NADPH- and iron(II)-dependent lipid peroxidation (LPO) of rat liver microsomes at 20 and 200 microM, respectively . When ferrylmyoglobin was incubated with dimerumic acid, the myoglobin was scavenged and an electron spin resonance (ESR) signal with nine peaks was observed . The spin adduct was identified as a nitroxide radical by analysis of hyperfine structure . Similar ESR signal was also detected by incubation of dimerumic acid with peroxyl radicals . Thus, it was clarified that the antioxidant action of dimerumic acid is due to one electron donation of the hydroxamic acid group in the dimerumic acid molecule toward oxidants resulting in formation of nitroxide radical. Sheng Wu Gong Cheng Xue Bao, 2001 Nov, 17(6), 693 - 7 {Analysis of parameters for optimum of ergosterol fermentation}; Gao H et al.; Parameters in the process of ergosterol fermentation are studied . The relationship between biomass, ergosterol content in Saccharomyces cerevisiae and parameters such as DO, pH, OUR, glucose concentration, are discussed . Because of its good manipulation in yeast fermentation process, Do can thus serve as an effective control parameter . DO at about 12% (+/- 1%) can enhance the total yield of ergosterol considerably. Climacteric, 1999 Mar, 2(1), 6 - 12 Hot flushes and other menopausal symptoms in relation to soy product intake in Japanese women; Nagata C et al.; OBJECTIVE: To examine the relationships between dietary intake of soy products and hot flushes and other menopausal symptoms . METHODS: Subjects were 284 women aged 40-59 years who attended a health check-up program provided by a general hospital in Gifu, Japan . They completed a health questionnaire including the Kupperman test of menopausal distress . Diet was assessed by a semiquantitative food frequency questionnaire . RESULTS: Fermented soy product intake but not total soy product intake was significantly negatively correlated with hot flush severity (r = -0.16, p = 0.01) after controlling for age and menopausal status . Neither total soy product intake nor fermented soy product intake was significantly correlated with menopausal index score . Estimated isoflavone intake from total and fermented soy products was significantly lower by 15% (p = 0.02) and 19% (p = 0.01), respectively, in women with hot flushes, compared to those without hot flushes after controlling for covariates . CONCLUSION: The data support a hypothesis that intake of fermented soy products alleviates the severity of hot flushes. Arch Microbiol, 2002 Mar, 177(3), 244 - 50 Epub 2002 Jan 22. Isolation, characterization, and fermentative pattern of a novel thermotolerant Prototheca zopfii var . hydrocarbonea strain producing ethanol and CO2 from glucose at 40 degrees C; Ueno R et al.; A novel thermotolerant strain of the achlorophyllous micro-alga Prototheca was isolated from a hot spring . The isolate was found to produce an appreciable amount of ethanol and CO2 from glucose under anoxic conditions at both 25 and 40 degrees C; this type of alcohol fermentation has not yet been reported in the genus Prototheca . Moreover, it also evolved gas from sucrose after a time lag at 40 degrees C . Its taxonomic characteristics coincided with those of Prototheca zopfii var . hydrocarbonea, and phylogenetic analysis, based on a small-subunit (SSU) rDNA sequence, also revealed a close relationship between the two strains . D-lactic acid, ethanol, CO2 and a trace of acetic acid were produced from glucose, but L-lactic acid, formic acid, and H2 were not . At 25 degrees C, D-lactic acid and ethanol were produced in approximately equimolar amounts under N2/H2/CO2, whereas ethanol production was predominant under N2 . More ethanol was produced at 40 degrees C than at 25 degrees C irrespective of the gas composition in the atmosphere . This is the first report on gas production from glucose and on the changes in the fermentative patterns as a function of temperature for the genus Prototheca. J Anim Physiol Anim Nutr (Berl), 2001 Dec, 85(11-12), 378 - 84 Effect of plant oils and aspartate on rumen fermentation in vitro; Jalc D et al.; The effect of plant oils and aspartate (ASP) on rumen fermentation in vitro . The objective of this study was to determine the effect of plant oils (rapeseed - RO, sunflower - SO; linseed - LO; 10% wt/wt) and 8 mmol sodium aspartate on rumen fermentation of a diet (250 mg) consisting of hay, barley and sugar beet molasses (60 : 30 : 10) . Rumen fluid was collected from two Slovak Merino sheep fed the same diet twice daily . Mixed ruminal micro-organisms were incubated in fermentation fluid (40 ml) containing rumen fluid and McDougall's buffer (1 : 4) . Incubations were carried out in batch cultures for 72 h at 39 degrees C two times in a 3-week intervals . When compared to the control, all supplemented diets (RO, SO, LO, ASP) significantly increased the pH, the mol% of propionate (LO + 8.7%; SO + 10.12%; RO + 8.65%; ASP + 5.86%) and the acetate : propionate ratio and numerically decreased methane production (SO -32.8%; LO, RO -30.08%; ASP -21.56%) . Lactate production was also significantly decreased . Addition of plant oils to aspartate-treated incubations partly inhibited the decrease of n-butyrate, lactate and the increase of pH and in vitro dry matter digestibility (IVDMD) caused by ASP treatment . The effect of combined additives (RO + ASP, SO + ASP, LO + ASP) on methane production SO + ASP (-19.23%) and mol% propionate SO + ASP (+2.66%), LO + ASP (+4.28%) was less effective . All combined additives caused a significant decrease in digestibility of the given feeds . No effect of plant oils and ASP could be observed on the parameters of rumen fermentation (mainly methane and propionate). Indian J Exp Biol, 2001 Nov, 39(11), 1144 - 8 Optimization of some additives to improve protease production under SSF; Tunga R et al.; In a locally isolated Rhizopus oryzae strain highest-production of protease (388.54/g wheat bran) was observed in presence of Tween-80 and dioctyl sodium sulfosuccinate individually at 40mg/g wheat bran concentration . Under solid state fermentation biotin (0.0025mg/g wheat bran); Ca2+ (0.05mg/g wheat bran) and 1-Naphthyl acetic acid (0.01mg/g wheat bran) also showed some inducing effect on the synthesis of the enzyme protease by solid state fermentation. Indian J Exp Biol, 2001 Nov, 39(11), 1136 - 43 Optimisation of fermentation conditions for gluconic acid production by a mutant of Aspergillus niger; Singh OV et al.; Aspergillus niger ORS-4, isolated from the sugarcane industry waste materials was found to produce notable level of gluconic acid . From this strain, a mutant Aspergillus niger ORS-4.410 having remarkable increase in gluconic acid production was isolated and compared for fermentation properties . Among the various substrates used, glucose resulted into maximum production of gluconic acid (78.04 g/L) . 12% concentration led to maximum production . Effect of spore age and inoculum level on fermentation indicated an inoculum level of 2% of the 4-7 days old spores were best suited for gluconic acid production . Maximum gluconate production could be achieved after 10-12 days of the fermentation at 30 degrees C and at a pH of 5.5 . Kinetic analysis of production indicated that growth of the mutant was favoured during initial stages of the fermentation (4-8 days) and production increased during the subsequent 8-12 days of the fermentation . CaCO3 and varying concentrations of different nutrients affected the production of gluconic acid . Analysis of variance for the factors evaluated the significant difference in the production levels. Proc Natl Acad Sci U S A, 2002 Mar 19, 99(6), 3370 - 5 Fast 100-nm resolution three-dimensional microscope reveals structural plasticity of mitochondria in live yeast; Egner A et al.; By introducing beam-scanning multifocal multiphoton 4Pi-confocal microscopy, we have attained fast fluorescence imaging of live cells with axial super resolution . Rapid scanning of up to 64 pairs of interfering high-angle fields and subsequent confocal detection enabled us to perform three to five times finer optical sectioning than confocal microscopy . In conjunction with nonlinear image restoration, we demonstrate, to our knowledge for the first time, three-dimensional imaging of live eukaryotic cells at an equilateral resolution of approximately 100 nm . This imaging mode allowed us to reveal the morphology and size of the green fluorescent protein-labeled mitochondrial compartment of live Saccharomyces cerevisiae (bakers' yeast) growing on different carbon sources . Our studies show that mitochondria of cells grown on medium containing glycerol as the only carbon source, as opposed to glucose-grown cells, exhibit a strongly branched tubular reticulum . We determine the average tubular diameter and find that it increases from 339 +/- 5 nm to 360 +/- 4 nm when changing from glucose to glycerol, that is, from a fermentable to a nonfermentable carbon source . Moreover, this change is associated with a 2.8-fold increase of the surface of the reticulum, resulting in an average increase in volume of the mitochondrial compartment by a factor of 3.0 +/- 0.2. FEBS Lett, 2002 Feb 27, 513(2-3), 193 - 6 Starvation-induced degradation of yeast hexose transporter Hxt7p is dependent on endocytosis, autophagy and the terminal sequences of the permease; Krampe S et al.; The yeast high-affinity glucose transporters Hxt6p and Hxt7p are rapidly degraded during nitrogen starvation in the presence of high concentrations of fermentable carbon sources . Our results suggest that degradation is mainly due to the stimulation of general protein turnover and not caused by a mechanism specifically triggered by glucose . Analysis of Hxt6p/7p stability and cellular distribution in end4, aut2 and apg1 mutants indicates that Hxt7p is internalized by endocytosis, and autophagy is involved in the final delivery of Hxt7p to the vacuole for proteolytic degradation . Internalization and degradation of Hxt7p were blocked after truncation of its N-terminal hydrophilic domain . Nevertheless, this fully functional and stabilized hexose transporter could not maintain fermentation capacity of the yeast cells under starvation conditions, indicating a regulatory constraint on glucose uptake. J Agric Food Chem, 2002 Mar 27, 50(7), 2062 - 7 Effect of caffeic acid on the color of red wine; Darias-Martin J et al.; The copigmentation effect of prefermentation additions of different doses of caffeic acid was investigated during the 1997 harvest . Microfermentation with the major red grape cultivars Listan negro and Negramoll, grown in the Canary Islands, was carried out with the same protocol . Visible and UV spectra were registered periodically . HPLC chromatograms were carried out . The color enhancement of cv . Negramoll wine varied between 13 and 75% (AU at 520 nm), and that of cv . Listan negro wine between 25 and 45% at the end of fermentation . During aging these values were enhanced to reach even >100% in some cases . An initial complex of the 1:1 type, where one molecule of caffeic acid associates with one molecule of anthocyanin, has been identified using the mathematical procedure of Brouillard et al . (J . Am . Chem . Soc . 1989, 111, 2604-2610) . Caffeic acid seems to contribute to color stability and protection against oxidation . The importance of nonpigment composition in pigment extraction and color retention during and after fermentation is demonstrated. J Agric Food Chem, 2002 Mar 27, 50(7), 1833 - 9 Study of catechin and xanthine tea profiles as geographical tracers; Fernandez PL et al.; The contents of gallic acid, epigallocatechin gallate, epigallocatechin, epicatechin, epicatechin gallate, catechin, caffeine, theophylline, and theobromine were determined in a set of 45 tea samples, including fermented (black and red) and nonfermented (green) teas of different geographical origins (i.e., China, Japan, Kenya, Sri Lanka, and India) . A reversed-phase high performance liquid chromatographic method with gradient elution and photometric detection at 275 nm was used to carry out the analysis . Before the HPLC determination, an extraction step was developed using a mixture of acetonitrile and water (60:40, v/v) . Pattern recognition techniques involving principal component analysis (PCA) and linear discriminant analysis (LDA) were applied to differentiate the tea samples according to their geographical origins . Catechins, gallic acid, and tea alkaloids are adequate chemical descriptors to distinguish between fermented and nonfermented tea samples cultivated in different geographical areas. Faraday Discuss, 2001, (120), 261 - 76; discussion 325-51 Synchronization of glycolytic oscillations in a yeast cell population; Dano S et al.; The mechanism of active phase synchronization in a suspension of oscillatory yeast cells has remained a puzzle for almost half a century . The difficulty of the problem stems from the fact that the synchronization phenomenon involves the entire metabolic network of glycolysis and fermentation, and consequently it cannot be addressed at the level of a single enzyme or a single chemical species . In this paper it is shown how this system in a CSTR (continuous flow stirred tank reactor) can be modelled quantitatively as a population of Stuart-Landau oscillators interacting by exchange of metabolites through the extracellular medium, thus reducing the complexity of the problem without sacrificing the biochemical realism . The parameters of the model can be derived by a systematic expansion from any full-scale model of the yeast cell kinetics with a supercritical Hopf bifurcation . Some parameter values can also be obtained directly from analysis of perturbation experiments . In the mean-field limit, equations for the study of populations having a distribution of frequencies are used to simulate the effect of the inherent variations between cells. Appl Biochem Biotechnol, 2002 Jan, 97(1), 1 - 22 Production and characterization of pullulan from beet molasses using a nonpigmented strain of Aureobasidium pullulans in batch culture; Lazaridou A et al.; The production of pullulan from beet molasses by a pigment-free strain of Aztreobasidium pullulans on shake-flask culture was investigated . Combined pretreatment of molasses with sulfuric acid and activated carbon to remove potential fermentation inhibitors present in molasses resulted in a maximum pullulan concentration of 24 g/L, a biomass dry wt of 14 g/L, a pullulan yield of 52.5%, and a sugar utilization of 92% with optimum fermentation conditions (initial sugar concentration of 50 g/L and initial pH of 7.0) . The addition of other nutrients as carbon and nitrogen supplements (olive oil, ammonium sulfate, yeast extract) did not further improve the production of the exopolysaccharides . Structural characterization of the isolated polysaccharides from the fermentation broths by 13C-nuclear magnetic resonance spectroscopy and pullulanase digestion combined with size-exclusion chromatography confirmed the identity of pullulan and the homogeneity (>93% dry basis) of the elaborated polysaccharides by the microorganism . Using multiangle laser light scattering and refractive index detectors in conjunction with high-performance size-exclusion chromatography molecular size distributions and estimates of the molecular weight (Mw = 2.1-4.1 x 10(5)), root mean square of the radius of gyration (R = 30-38 nm), and polydispersity index (Mw/Mn = 1.4-2.4) were obtained . The fermentation products of molasses pretreated with sulfuric acid and/or activated carbon were more homogeneous and free of contaminating proteins . In the concentration range of 2.8-10.0 (w/v), the solution's rheologic behavior of the isolated pullulans was almost Newtonian (within 1 and 1200 s(-1) at 20 degrees C); a slight shear thinning was observed at 10.0 (w/v) for the high molecular weight samples . Overall, beet molasses pretreated with sulfuric acid and activated carbon appears as an attractive fermentation medium for the production of pullulan by A . pullulans. Curr Opin Drug Discov Devel, 2001 Nov, 4(6), 756 - 9 Development of fermentation process for rDNA products; Cossar D; Fermentation process development for recombinant DNA-derived products is becoming increasingly important in the current commercial and regulatory framework . This article provides an overview of the current approach to process development, and the contribution of developmental data to final process validation is highlighted . Cited literature is restricted to between 1995 and 2001. Folia Microbiol (Praha), 2001, 46(5), 427 - 31 D-amino-acid oxidase--an improved production of the enzyme by the yeast Trigonopsis variabilis in a laboratory fermentor; Kujan P et al.; The cultivation of the yeast Trigonopsis variabilis producing D-amino-acid oxidase (an enzyme participating in the transformation of cephalosporin C into 7-aminocephalosporanic acid for the production of beta-lactam antibiotics) was controlled by changes of dissolved oxygen tension and extended fermentation times . The production technology was optimized on a laboratory scale and scale-up parameters were identified. Antimicrob Agents Chemother, 2002 Apr, 46(4), 1112 - 3 Potent in vitro antimalarial activity of metacycloprodigiosin isolated from Streptomyces spectabilis BCC 4785; Isaka M et al.; Bioassay-guided fractionation of the extract from the fermentation broth of Streptomyces spectabilis BCC 4785 led to the isolation of three principle antimalarial agents, metacycloprodigiosin, bafilomycin A(1), and spectinabilin . Metacycloprodigiosin exhibited potent in vitro activity against Plasmodium falciparum K1, with a 50% inhibitory concentration of 0.0050 +/- 0.0010 microg/ml, while its cytotoxicity was much weaker. Nutr Metab Cardiovasc Dis, 2001 Aug, 11(4 Suppl), 118 - 21 Study of the regulation by nutrients of the expression of genes involved in lipogenesis and obesity in humans and animals; Delzenne N et al.; Dietary digestible carbohydrates are able to modulate lipogenesis, by modifying the expression of genes coding for key lipogenic enzymes, like fatty acid synthase . The overall objective of the Nutrigene project (FAIR-CT97-3011) was to study the efficiency of various carbohydrates to modulate the lipogenic capacity and relevant gene expression in rat and human species (control and obese subjects) and to understand the underlying molecular mechanisms involved in the regulation of lipogenic genes by carbohydrates . Key cellular mediators (namely SREBP-1c and 2, AMP activated protein kinase, cholesterol content) of the regulation of lipogenic gene expression by glucose and/or insulin were identified and constitute new putative targets in the development of plurimetabolic syndrome associated with obesity . In humans, hepatic lipogenesis and triglyceride synthesis, assessed in vivo by the use of stable isotopes, was promoted by a high-carbohydrate diet in non obese subjects, and in non alcoholic steatotic patients, but was not modified in the adipose tissue of obese subjects . Non digestible/fermentable carbohydrates, such as fructans, were shown to decrease hepatic lipogenesis in non obese rats, and to lessen hepatic steatosis and body weight in obese Zucker rats . If confirmed in obese humans, this would allow the development of functional food able to counteract the metabolic disturbances linked to obesity. Euro Surveill, 2001 Oct, 6(10), 147 - 51 Escherichia coli O157 infections and unpasteurised milk; Allerberger F et al.; We report on two children with Escherichia coli O157 infection, one of whom developed haemolytic uraemic syndrome (HUS) . Both had drunk raw cows or goats milk in the week before their illness . Molecular subtyping identified a sorbitol fermenting Escherichia coli O157:H isolate from a dairy cow . This isolate differed from Shiga toxin producing O157:H strains isolated from the 6 year old boy with HUS . This result underlines the need to search for other causes of infection, despite documented consumption of unpasteurised milk . In the second patient, human sorbitol non-fermenting O157:H isolates and animal isolates from goats were indistinguishable . The isolation of indistinguishable sorbitol non-fermenting Escherichia coli O157:H from contact animals supports the association between HUS and consumption of raw goats milk, and re-emphasises the importance of pasteurising milk. J Anim Sci, 2002 Mar, 80(3), 628 - 36 Effects of dietary lipid type on muscle fatty acid composition, carcass leanness, and meat toughness in lambs; Ponnampalam EN et al.; Isonitrogenous amounts of two protein sources differing in rumen degradation rate and in lipid composition were fed to sheep with or without a rapidly fermentable cereal grain . The effects on intake, carcass leanness, and muscle fatty acid (FA) composition were examined . Thirty-eight crossbred wether lambs (9 mo, 35 to 48 kg) were allocated by stratified randomization to six treatment groups: 1) basal diet of alfalfa hay:oat hay (20:80) ad libitum = basal; 2) basal + lupin (358 g DM/d) = lupin; 3) basal + fish meal (168 g DM/d) = fish meal; 4) basal + barley (358 g DM/d) = barley; 5) basal + barley + lupin (179 + 179 g DM/d) = barley/lupin; or 6) basal + barley + fish meal (179 + 84 g DM/d) = barley/ fish meal . Lambs were fed individually . Dietary treatments were imposed for 8 wk, and the supplements were offered at 2-d intervals . Daily feed intake and weekly BW of lambs were recorded . At the end of the feeding period lambs were slaughtered after an overnight fast . Hot carcass weight (HCW) and fat depth (GR; total fat and muscle tissue depth at 12th rib, 110 mm from midline) were recorded . At 24 h postmortem samples of longissimus thoracis (LT) and longissimus lumborum (LL) muscles were taken from chilled (4 degrees C) carcasses for the assessment of FA composition and meat tenderness, respectively . Lambs fed lupin or fish meal with or without barley had heavier slaughter weights (P < 0.004) and HCW (P < 0.001) than lambs fed basal or barley when initial BW was included as a covariate . The lupin diet also resulted in heavier carcasses (P < 0.05) than the fish meal or barley/fish meal diets . With GR as an indicator, fish meal and barley/ fish meal diets produced leaner carcasses (P < 0.01) than lupin and barley/lupin lambs . Long-chain n-3 FA content {20:5n-3 (P < 0.001), 22:5n-3 (P < 0.003), and 22:6n-3 (P < 0.001)} in the LT muscle were substantially higher with the fish meal and barley/fish meal diets, whereas muscle total n-6 FA was increased (P < 0.003) by lupin and barley/lupin compared with all other diets . Thus, increased muscle long-chain n-3 FA content occurred without an increase in fatness measured as GR, whereas increased muscle n-6 FA content was associated with an increase in carcass fatness . Under these circumstances, a reduction in carcass fatness had no effect on meat tenderness measured as Warner-Bratzler shear force. Arch Microbiol, 2002 Apr, 177(4), 313 - 21 Epub 2002 Jan 31. The sensor kinase CitA (DpiB) of Escherichia coli functions as a high-affinity citrate receptor; Kaspar S et al.; For the CitA-CitB (DpiB-DpiA) two-component signal transduction system from Escherichia coli, three diverse functions have been reported: induction of the citrate fermentation genes citCDEFXGT, repression of the regulator gene appY, and destabilization of the inheritance of iteron-containing plasmids such as pSC101 . This poses the question of the principal biological role of this system . Here it is shown that the periplasmic domain of the E . coli sensor kinase CitA functions as a high-affinity citrate receptor . Two CitA derivatives were purified by affinity chromatography and subjected to binding studies using isothermal titration calorimetry (ITC) . One of them, termed CitA215MBP, comprised the N-terminal part of CitA (amino acid residues 1-215), including the two transmembrane helices, and was fused to the amino terminus of the E . coli maltose-binding protein lacking its signal peptide . The second CitA derivative, designated CitAP(Ec), encompassed only the periplasmic domain (amino acid residues 38-177) . CitA215MBP bound citrate at 25 degrees C with a K(d) of 0.3 microM and a binding stoichiometry of up to 0.9 in 50 mM sodium phosphate buffer, pH 7 . Binding was driven by the enthalpy change (Delta H of -95.7 kJ mol(-1)), whereas the entropy change was not favorable for binding ( T Delta S of -58.6 kJ mol(-1)) . ITC experiments with CitAP(Ec) yielded similar K(d) values for citrate (0.15-1.0 microM) . Besides citrate, also isocitrate ( K(d) approximately tricarballylate ( K(d) approximately t not malate were bound by CitAP(Ec) . The results favor the assumption that the primary biological function of the CitA-CitB system is the regulation of the citrate fermentation genes. Gut, 2002 Apr, 50(4), 525 - 9 Stimulation of the intestinal Cdx2 homeobox gene by butyrate in colon cancer cells; Domon-Dell C et al.; BACKGROUND: The transcription factor encoded by the intestinal Cdx2 homeobox gene and treatment with sodium butyrate (NaB), a byproduct of fibre fermentation by colonic bacteria, exert similar effects on colon cancer cell lines as they both inhibit cell growth and stimulate cell differentiation and apoptosis . AIM: To investigate whether NaB regulates expression of the Cdx2 gene in colon cancer cell lines . METHODS: Human adenocarcinoma cell lines Caco2 and HT29 were grown in the presence or absence of NaB . Cells were analysed for Cdx2 mRNA expression by reverse transcription-polymerase chain reaction, for protein expression by western blotting and electromobility shift assays, and for transcriptional activity of the Cdx2 promoter by transfection with luciferase reporter plasmids . RESULTS: In HT29 and Caco2 cells, NaB stimulated Cdx2 mRNA and protein expression as well as transcriptional activity of the Cdx2 promoter . Stimulation of the activity of the Cdx2 promoter by NaB was dose and time dependent . The Cdx2 promoter contains discrete regions that participate in or inversely that blunt the stimulatory effect exerted by NaB . In addition, NaB stimulated the transcriptional activity of the Cdx2 promoter downregulated by oncogenic ras . CONCLUSION: This study is the first report of an intestine specific transcription factor, Cdx2, stimulated by butyrate . Thus it provides a new mechanism whereby butyrate controls proliferation and differentiation of colon cancer cells. J Clin Dent, 2002, 13(3), 116 - 8 Dental considerations in sucralose use; Mandel ID et al.; Sucralose is a new type of non-caloric, high-intensity sweetener recently approved for use by the U.S . FDA . Its availability may expand the number of palatable, low-sugar foods and beverages currently on the market . A series of studies has been conducted to assess whether sucralose has cariogenic potential . These include an examination of oral bacterial metabolism, experimental caries in animal models, and the effect of sucralose-containing solutions on human plaque pH in situ . The sum of these studies demonstrates that sucralose is non-cariogenic . Sucralose-based sweeteners that contain bulking ingredients, which allow them to pour and measure more like sugar, do have cariogenic potential due to the presence of added fermentable carbohydrate; however, the data suggest that both the currently marketed sucralose granular and packet products are less cariogenic than sugar . Thus, when used to replace sugar, both sucralose and the tested sucralose-based sweeteners may be useful in the dietary management of caries. J Environ Sci (China), 2002 Jan, 14(1), 141 - 4 Optimization of C/N ratio preparation of protein-rich and multi-enzymes feed thallus through synergic fermentation of mixed distillers' grains; Li ZY et al.; A new procedure of determining optimal C/N (the rate of carbon source to nitrogen source) of mixed distillers' grains for combined bacteria synergic fermentation is established . At the same time an improved method evaluating bacteria growth, called method of dry cell weighing by filtering is developed . For each combination of C and N, their initial and residual contents before and after fermentation respectively are determined . Then followed the calculation of utilization of C and N sources by the compound bacteria . The optimal C/N is finally located from among the utilization of C and N of several combinations and the weight of produced mass of oven-dried thallus . The conditions of fermentation are: inoculum size 10%, temperature 30.0 degrees C, rotational speed 170 r/min, shake culture time 48 h . The best results obtained from orthogonal experiments are: maximum mass of oven dried thallus is 14.693 g in a liter liquid medium, maximum utilization rate of carbon source is 98.13% and maximum utilization rate of nitrogen is 78.14% . Optimal C/N is 5.1. FEMS Microbiol Lett, 2002 Jan 22, 207(1), 69 - 73 Analysis of the Schwanniomyces occidentalis SWA2 gene promoter in Saccharomyces cerevisiae; Carmona TA et al.; The effect of different carbon sources on the expression in Saccharomyces cerevisiae of the SWA2 alpha-amylase gene from Schwanniomyces occidentalis was studied from constructs containing its 5' region (-223 to +15), which were fused in-frame to the lacZ gene coding sequence . Maximal expression was achieved with the non-fermentable substrates ethanol and/or glycerol, whereas lower levels were found with maltose or galactose . In contrast, glucose repressed it, even in the presence of any of these other carbon sources . Deletion analyses of the -233 to -85 SWA2 promoter region permitted the identification of two fragments involved in both glucose repression and ethanol activation . A possible region required for cAMP regulation was localised . The SWA2 promoter contains a MIG1-binding GC box whose deletion caused a five-fold increase in the glucose-repressed reporter expression . Despite this, expression of the SWA2 promoter was not MIG1-dependent. Res Microbiol, 2002 Jan-Feb, 153(1), 53 - 8 Crosses between Saccharomyces cerevisiae and Saccharomyces bayanus generate fertile hybrids; Sebastiani F et al.; Crossings between strains of Saccharomyces cerevisiae and Saccharomyces bayanus were carried out . Genetic, molecular and electrophoretic karyotyping data indicated that interspecific hybrids were obtained . The hybrid cells segregated "grande" and "petite" colonies, and the latter ranged between 20 and 50%; unlike "grande" colonies, "petite" colonies did not sporulate and did not ferment maltose . In the hybrids, the extent of sporulation varied between 10 and 20%; only very rare asci (around 10(-4)) held viable ascospores . Clones from the viable ascospores sporulated and produced asci with viable ascospores able to give mating with spores from both hybrid derivatives and parental species . Fertile asci could derive from allotetraploid cells generated by endomitotic events in allodiploid cells, a mechanism that enables overcoming the species barrier between S . cerevisiae and S . bayanus. J Clin Microbiol, 2002 Mar, 40(3), 922 - 5 Characterization of Shiga toxin-producing Escherichia coli O26 strains and establishment of selective isolation media for these strains; Hiramatsu R et al.; We characterized the carbohydrate-fermenting ability of 31 strains of Shiga toxin-producing Escherichia coli (STEC) O26 isolated from diarrhea patients in Aichi Prefecture, Japan, in order to establish selective isolation media for these strains . None of the 31 STEC O26 strains (24 O26:H11, 7 O26:H-) fermented rhamnose, whereas all of the other 108 STEC strains (100 O157, 8 O111) and all of the non-STEC strains except one (i.e., 58 of 59) fermented rhamnose . The great majority of the STEC O26 strains (96.8% {30 of 31}) showed very high resistance to potassium tellurite (MIC > or = 50 microg/ml), whereas the majority of the non-STEC strains (72.9% {43 of 59}) showed very high sensitivity (MIC < or = 1.56 microg/ml) to this compound . Accordingly, we developed a rhamnose-MacConkey (RMAC) medium in which the lactose in MacConkey medium was replaced by rhamnose, and cefixime-tellurite-RMAC (CT-RMAC) medium in which potassium tellurite (2.5 mg/liter) and cefixime (0.05 mg/liter) were added to RMAC . All of the STEC O26 strains generated colorless (rhamnose-nonfermented) colonies on both media; the vast majority of selected E . coli strains (95.7% {89 of 93; including 26 STEC O157, 8 STEC O111}), other than STEC O26, generated red colonies on RMAC, and most of the non-STEC strains (84.7% {50 of 59}) did not grow on CT-RMAC . We demonstrate that both the RMAC and the CT-RMAC media can be used for the isolation of STEC O26 and that CT-RMAC has better specificity for the routine isolation for STEC O26 in a laboratory. J Agric Food Chem, 2002 Mar 13, 50(6), 1535 - 42 Selection of yeast starter culture strains for the production of marula fruit wines and distillates; Fundira M et al.; Juice of the Sclerocarya birrea subsp . caffra (marula) fruit was fermented by indigenous microflora and different commercial Saccharomyces cerevisiae yeast strains at different temperatures, namely, 15 and 30 degrees C . Volatile acids, esters, and higher alcohols were quantified in the wine and distillates, and the results were interpreted using a multivariate analysis of variance and an average linkage cluster analysis . Significant differences between 15 and 30 degrees C and also among yeasts with respect to volatile compounds were observed . Yeast strains VIN7 and FC consistently produced wines and final distillates significantly different from the other strains . A panel of tasters and marula and brandy producers was asked to select wines and distillates that had an acceptable and typical marula "nose" . They were also asked to detect the differences among wines and distillates fermented with the same yeast strain at different temperatures. Extremophiles, 2002 Feb, 6(1), 39 - 44 Sulfolobus tokodaii sp . nov . (f . Sulfolobus sp . strain 7), a new member of the genus Sulfolobus isolated from Beppu Hot Springs, Japan; Suzuki T et al.; The taxonomic position of a thermoacidophilic crenarchaeote Sulfolobus sp . strain 7, previously isolated from the Beppu Hot Springs in the geothermal area of Kyushu Island, Japan, was investigated by cloning and sequencing, by phylogenetic analysis of the 16S rRNA gene sequence, by DNA-DNA homology with similar species, and by biochemical characterization of the isolate . This isolate is an obligate aerobe and grows optimally at 80 degrees C and pH2.5-3 under aerobic and chemoheterotrophic growth conditions by aerobic respiration rather than simple fermentation . In conjunction with the phenotypic properties, the present phylogenetic analysis based on the 16S rRNA gene sequence and DNA-DNA hybridization experiments indicate that this isolate is related to the described Sulfolobus taxon and should be considered a novel species of the genus . We propose that this isolate is a novel species of the genus Sulfolobus that we name Sulfolobus tokodaii sp . nov . The type strain is strain 7 (JCM 10545). BMC Bioinformatics . 2002;3(1):7 . Epub 2002 Feb 14. Correlating overrepresented upstream motifs to gene expression: a computational approach to regulatory element discovery in eukaryotes; Caselle M et al.; BACKGROUND: Gene regulation in eukaryotes is mainly effected through transcription factors binding to rather short recognition motifs generally located upstream of the coding region . We present a novel computational method to identify regulatory elements in the upstream region of eukaryotic genes . The genes are grouped in sets sharing an overrepresented short motif in their upstream sequence . For each set, the average expression level from a microarray experiment is determined: If this level is significantly higher or lower than the average taken over the whole genome, then the overerpresented motif shared by the genes in the set is likely to play a role in their regulation . RESULTS: The method was tested by applying it to the genome of Saccharomyces cerevisiae, using the publicly available results of a DNA microarray experiment, in which expression levels for virtually all the genes were measured during the diauxic shift from fermentation to respiration . Several known motifs were correctly identified, and a new candidate regulatory sequence was determined . CONCLUSIONS: We have described and successfully tested a simple computational method to identify upstream motifs relevant to gene regulation in eukaryotes by studying the statistical correlation between overepresented upstream motifs and gene expression levels. Appl Microbiol Biotechnol, 2002 Feb, 58(2), 164 - 9 Polygalacturonase production by Aspergillus awamori on wheat in solid-state fermentation; Blandino A et al.; The production of exo-polygalacturonase (exo-PG) and endo-PG by Aspergillus awamori grown on wheat in solid-state fermentation was studied . Endo- and exo-PG activities were detected after 24 h of inoculation . Glucose released from starch hydrolysis acted as a catabolite repressor for the exo-PG enzyme . In contrast, endo-PG production was not affected by glucose repression . When milled grains were used, the particle-size distribution and the chemical composition of the medium influenced the rate of micro-organism growth and therefore the trend followed by endo- and exo-PG production . However, these two parameters did not affect the maximum production of exo-PG and endo-PG . For one of the milled samples, three different moisture contents were used (50, 55, 60%) . Moisture contents of 60% provide a higher yield of pectinases by A . awamori. Appl Microbiol Biotechnol, 2002 Feb, 58(2), 157 - 63 13C-NMR analysis of glucose metabolism during citric acid production by Aspergillus niger; Peksel A et al.; The effect of glucose concentration on glycolytic metabolism under conditions of citric acid accumulation by Aspergillus niger was studied with 13C-labelled glucose . The results show that during cultivation at high glucose (14%, w/v), most of the label in citric acid is in C-2/C-4, and is thus due to the pyruvate carboxylase reaction . However, a significant portion is also present in C-1/C-5, whose origin is less clear but most likely due to reconsumption of glycerol and erythritol . Formation of trehalose and mannitol is high during the early phase of fermentation and declines thereafter . The early fermentation phase is further characterized by a high rate of anaplerosis from oxaloacetate to pyruvate, which also decreases with time . At low glucose concentrations (2%, w/v), which lead to a significantly reduced citric acid yield and formation rate, labelling of citrate in C-2/C-4 is decreased and C-l/C-5 labelling increased . Growth on 2% glucose is also characterized by an appreciable scrambling of mannitol and considerable backflux from mannitol to trehalose (indicating tight glycolytic control at the fructose-6-phosphate step) and an increased anaplerotic formation of pyruvate from oxaloacetate . These data indicate that cultivation on high sugar concentrations shifts control of glycolysis from fructose-6-phosphate to the glyceraldehyde-3-phosphate dehydrogenase step. Syst Appl Microbiol, 2001 Dec, 24(4), 639 - 44 Performance of industrial strains of Saccharomyces cerevisae during wine fermentation is affected by manipulation strategies based on sporulation; Gimren-Alcaniz JV et al.; Genetic manipulation of industrial wine yeast strains has become an essential tool for both the study of the molecular mechanisms underlaying their physiology and the improvement of their fermentative properties . The construction of null mutants for any gene in these usually diploid strains, by using a procedure based on sporulation of a heterozygote lacking one copy of the gene of interest, has been tested as an alternative to the tedious work of sequential disruption of the complete set of copies . Our results indicate that most of the homozygotes resulting from sporulation of wine yeast strains are defective in glucose consumption under microvinification conditions in synthetic must and produce stuck fermentations . These kinds of defects are observed even for strains derived from sporulation of wild type . Alteration of genomic features of wine strains by sporulation is responsible for these defects. Syst Appl Microbiol, 2001 Dec, 24(4), 634 - 8 Yeasts present during wine fermentation: comparative analysis of conventional plating and PCR-TTGE; Fernandez-Gonzalez M et al.; Yeasts isolated from must before and during fermentation at a wine cellar of La Mancha region in Spain were characterised using Polymerase Chain Reaction / Restriction Fragments Lengths Polymorphism and Polymerase Chain Reaction / Temporal Temperature Gradient Gel Electrophoresis . S . cerevisiae strains were differentiated using mtDNA restriction analysis . Direct PCR-TTGE was also used to study biodiversity during wine fermentation, and revealed the variations in the population . It was observed that isolation by conventional plating may afford a skewed view of the strains taking part in wine fermentation. Mol Biotechnol, 2002 Jan, 20(1), 29 - 40 Large-scale extraction of proteins; Cunha T et al.; The production of foreign proteins using selected host with the necessary posttranslational modifications is one of the key successes in modern biotechnology . This methodology allows the industrial production of proteins that otherwise are produced in small quantities . However, the separation and purification of these proteins from the fermentation media constitutes a major bottleneck for the widespread commercialization of recombinant proteins . The major production costs (50-90%) for typical biological product resides in the purification strategy . There is a need for efficient, effective, and economic large-scale bioseparation techniques, to achieve high purity and high recovery, while maintaining the biological activity of the molecule . Aqueous two-phase systems (ATPS) allow process integration as simultaneously separation and concentration of the target protein is achieved, with posterior removal and recycle of the polymer . The ease of scale-up combined with the high partition coefficients obtained allow its potential application in large-scale downstream processing of proteins produced by fermentation . The equipment and the methodology for aqueous two-phase extraction of proteins on a large scale using mixer-settlerand column contractors are described . The operation of the columns, either stagewise or differential, are summarized . A brief description of the methods used to account for mass transfer coefficients, hydrodynamics parameters of hold-up, drop size, and velocity, back mixing in the phases, and flooding performance, required for column design, is also provided. Appl Environ Microbiol, 2002 Mar, 68(3), 1374 - 80 Hindgut fermentation in three species of marine herbivorous fish; Mountfort DO et al.; Symbioses with gut microorganisms provides a means by which terrestrial herbivores are able to obtain energy . These microorganisms ferment cell wall materials of plants to short-chain fatty acids (SCFA), which are then absorbed and used by the host animal . Many marine herbivorous fishes contain SCFA (predominantly acetate) in their hindgut, indicative of gut microbial activity, but rates of SCFA production have not been measured . Such information is an important prerequisite to understanding the contribution that gut microorganisms make in satisfying the energy needs of the fish . We have estimated the rates of acetate production in the gut of three species of temperate marine herbivorous fish from northeastern New Zealand: Kyphosus sydneyanus (family Kyphosidae), Odax pullus (family Odacidae), and Aplodactylus arctidens (family Aplodactylidae) . Ex vivo preparations of freshly caught fish were maintained with their respiratory and circulatory systems intact, radiolabeled acetate was injected into ligated hindgut sections, and gut fluid was sampled at 20-min intervals for 2 h . Ranges for acetate turnover in the hindguts of the studied species were determined from the slope of plots as the log of the specific radioactivity of acetate versus time and pool size, expressed on a nanomole per milliliter per minute basis . Values were 450 to 570 (K . sydneyanus), 373 to 551 (O . pullus), and 130 to 312 (A . arctidens) . These rates are comparable to those found in the guts of herbivorous reptiles and mammals . To determine the contribution of metabolic pathways to the fate of acetate, rates of sulfate reduction and methanogenesis were measured in the fore-, mid-, and hindgut sections of the three fish species . Both rates increased from the distal to proximal end of the hindgut, where sulfate reduction accounted for only a small proportion (<5%) of acetate methyl group transformed to CO(2), and exceeded methanogenesis from acetate by >50-fold . When gut size was taken into account, acetate uptake from the hindgut of the fish species, determined on a millimole per day per kilogram of body weight basis, was 70 (K . sydneyanus), 18 (O . pullus), and 10 (A . arctidens). J Med Microbiol, 2002 Mar, 51(3), 207 - 20 Characterisation of aerobic gram-negative bacteria from subgingival sites of dogs--potential bite wound pathogens; Forsblom B et al.; Ninety-eight aerobic, gram-negative bacterial isolates from subgingival samples from family-owned dogs with naturally occurring periodontitis were characterised phenotypically by conventional biochemical testing, by cellular fatty acid profiling and by the use of commercial identification systems . The majority (48, 81%) of the fermentative isolates but only 18% of the non-fermenters were identified by conventional biochemical testing alone . With additional cellular fatty acid profiling, another 7 (12%) fermentative and 23 (59%) non-fermentative isolates were identified to genus or group level . Cellular fatty acid analysis was essential for the identification of most non-fermenters, many of which are difficult to identify due to a paucity of positive reactions in routine biochemical tests . Commercial identification systems were less useful and did not contribute to further identification of these problematic isolates . This study underlines the difficulties encountered in the identification of canine oral bacteria--a group of potential bite wound pathogens--and presents schemes for microbiology laboratories to characterise such isolates. Yeast, 2002 Mar 15, 19(4), 295 - 301 Decreasing acetic acid accumulation by a glycerol overproducing strain of Saccharomyces cerevisiae by deleting the ALD6 aldehyde dehydrogenase gene; Eglinton JM et al.; Glycerol is a major fermentation product of Saccharomyces cerevisiae that contributes to the sensory character of wine . Diverting sugar to glycerol overproduction and away from ethanol production by overexpressing the glycerol 3-phosphate dehydrogenase gene,GPD2, caused S . cerevisiae to produce more than twice as much acetic acid as the wild-type strain (S288C background) in anaerobic cell culture . Deletion of the aldehyde dehydrogenase gene, ALD6, in wild-type and GPD2 overexpressing strains (GPD2-OP) decreased acetic acid production by three- and four-fold, respectively . In conjunction with reduced acetic acid production, the GPD2-OP ald6Delta strain produced more glycerol and less ethanol than the wild-type . The growth rate and fermentation rate were similar for the modified and wild-type strains, although the fermentation rate for the GPD2 ald6Delta strain was slightly less than that of the other strains from 24h onwards . Analysis of the metabolome of the mutants revealed that genetic modification affected the production of some secondary metabolites of fermentation, including acids, esters, aldehydes and higher alcohols, many of which are flavour-active in wine . Modification of GPD2 and ALD6 expression represents an effective strategy to increase the glycerol and decrease the ethanol concentration during fermentation, and alters the chemical composition of the medium such that, potentially, novel flavour diversity is possible . The implications for the use of these modifications in commercial wine production require further investigation in wine yeast strains . Electrophoresis, 2002 Feb, 23(3), 414 - 20 Comparison of aqueous and nonaqueous carrier electrolytes for the separation of penicillin V and related substances by capillary electrophoresis with UV and mass spectrometric detection; Hilder EF et al.; A method for the determination of penicillin V together with its impurities and by-products formed during biosynthesis, using capillary electrophoresis (CE) with UV and electrospray-mass spectrometric (ESI-MS) detection is presented . Aqueous and nonaqueous electrolytes containing 20 mM ammonium acetate were investigated to determine their suitability for the separation of these analytes . These carrier electrolytes were optimized with respect to the pH and the solvent/s used (water, methanol, acetonitrile, ethanol and isopropanol) and it was shown that although the nonaqueous electrolytes offered unique separation selectivities, the best results in terms of selectivity and sensitivity were obtained for the aqueous system . Finally, the applicability of this method for the analysis of a mixture representative of a real fermentation broth was demonstrated using an aqueous carrier electrolyte with both UV and ESI-MS detection. Biotechnol Bioeng, 2002 Apr 20, 78(2), 172 - 8 Furfural, 5-hydroxymethyl furfural, and acetoin act as external electron acceptors during anaerobic fermentation of xylose in recombinant Saccharomyces cerevisiae; Wahlbom CF et al.; The electron acceptors acetoin, acetaldehyde, furfural, and 5-hydroxymethylfurfural (HMF) were added to anaerobic batch fermentation of xylose by recombinant, xylose utilising Saccharomyces cerevisiae TMB 3001 . The intracellular fluxes during xylose fermentation before and after acetoin addition were calculated with metabolic flux analysis . Acetoin halted xylitol excretion and decreased the flux through the oxidative pentose phosphate pathway . The yield of ethanol increased from 0.62 mol ethanol/mol xylose to 1.35 mol ethanol/mol xylose, and the cell more than doubled its specific ATP production after acetoin addition compared to fermentation of xylose only . This did, however, not result in biomass growth . The xylitol excretion was also decreased by furfural and acetaldehyde but was unchanged by HMF . Thus, furfural present in lignocellulosic hydrolysate can be beneficial for ethanolic fermentation of xylose . Enzymatic analyses showed that the reduction of acetoin and furfural required NADH, whereas the reduction of HMF required NADPH . The enzymatic activity responsible for furfural reduction was considerably higher than for HMF reduction and also in situ furfural conversion was higher than HMF conversion . Biomacromolecules, 2002 Jan-Feb, 3(1), 159 - 66 Characterization of microbial polythioesters: physical properties of novel copolymers synthesized by Ralstonia eutropha; Lutke-Eversloh T et al.; Various samples of polythioesters with different contents of 3-mercaptopropionic acid (3MP) or 3-mercaptobutyric acid (3MB) as one comonomer and with 3-hydroxybutyric acid (3HB) as the second constituent were produced by cultivating cells of Ralstonia eutropha strain H16 in mineral salts medium containing 3MP or 3MB plus gluconate as carbon sources . Fermentations were done also at the 30-L scale . The various samples were cast as films from chloroform and the following were recorded: melting point, solid-state NMR, X-ray diffraction . The copolyester poly(3HB-co-3MP) displayed mutiple melting peaks corresponding to separate phases rich in 3MP and 3HB . The copolyester poly(3HB-co-3MB) displayed very low crystallinity and melting points higher than that of poly(3HB) when the 3HB content was 40% or less. Scanning, 2002 Jan-Feb, 24(1), 1 - 5 Interaction of maize zein with wheat gluten in composite dough and bread as determined by confocal laser scanning microscopy; Bugusu BA et al.; Protein body-free maize zein, when mixed at 35 degrees C (above its glass transition temperature range), significantly (p < 0.01) improved the rheological and leavening properties of sorghum-wheat composite flour dough, resulting in improved loaf volume . Confocal laser scanning microscopy was used to observe the structure of zein fibrils and the interaction between zein and gluten proteins in the composite dough and bread systems . Autofluorescence and immunolocalization techniques were used to locate gluten and zein, respectively . Optical sections were collected every 0.4 microm through the samples and digitally processed to produce reconstructed three-dimensional images . Results showed that zein fibrils form an outer layer that intermittently coats the gluten networks, thereby strengthening them . This type of microstructure is able to withstand the pressure exerted by gas cell expansion during yeast fermentation to increase loaf volume. Arch Tierernahr, 2001, 54(3), 225 - 42 Comparative investigation of salinomycin and flavophospholipol in sheep fed different composed diets; Febel H et al.; The effects of salinomycin and flavophospholipol, and their relationship with the diet, were studied in nine ruminally and duodenally cannulated wethers . Within the composition of the ration, the levels of rumen degradable protein (RDP) and non-structural carbohydrates (NSC) were changed (diet H: 74% RDP and 38% NSC; diet M: 57% RDP and 32% NSC; diet L: 48% RDP and 23% NSC) . There was no clear treatment effect of flavophospholipol on propionate concentration . Salinomycin supplementation appeared to be more effective than flavophospholipol in the increase of propionate concentration at the expense of acetic acid . Salinomycin significantly reduced the ammonia concentration of the rumen fluid . Microbial N content of the duodenal digesta was significantly lower when salinomycin was used . Salinomycin inhibited proteolysis and reduced the efficiency of microbial protein synthesis . The effect of salinomycin on ruminal N metabolism was independent of the composition of substrate . Unlike salinomycin, flavophospholipol tended to increase proteolysis in the rumen and did not inhibit protein synthesis . The effect of salinomycin on ruminal fermentation and the duodenal flow of nutrients were independent of substrate composition. Comb Chem High Throughput Screen, 2002 Feb, 5(1), 65 - 73 Evaluation of a micro volume pulsed ultrafiltration cell for screening ligands in non-covalent complexes; Beverly MB et al.; A pulsed ultrafiltration cell with a 35 microL binding chamber was evaluated for its ability to screen ligands that formed non-covalent complexes with protein targets . The cell was tested with ligands to the targets of carbonic anhydrase and serum albumin . Non-covalent ligand binding to both of these targets was observed and bound ligands were eluted from the cell in less than five min . The cell was also demonstrated to effectively screen a methanolic fermentation broth extract spiked with a known inhibitor to carbonic anhydrase . In addition to detecting specific binding events, the pulsed ultrafiltration method was investigated for its ability to distinguish non-specific binding events . Using carbonic anhydrase with the zinc-binding site removed, it was found that non-specific complexes observed when using electrospray ionization alone were not detected when using the pulsed ultrafiltration mass spectrometry method. J Dairy Sci, 2002 Jan, 85(1), 217 - 26 Influence of corn processing and frequency of feeding on cow performance; Dhiman TR et al.; Twenty cows, including five fitted with rumen cannulae, were used to study the influence of corn processing and frequency of feeding on milk yield and ruminal fermentation characteristics . Cows were assigned to five treatments in a 5 x 5 Latin square experiment . Each period was 3 wk . Cows were fed 45% forage and 55% grain in a total mixed ration . Diets contained 35% corn either coarsely ground and fed once a day (1x), finely ground (FGC) fed 1x, steam-flaked (SFC) fed lx, FGC fed four times a day (4x), or SFC fed 4x . Processing of corn and frequency of feeding had no influence on dry matter intake . Digestibility of starch was increased 6 and 3 percentage units by feeding SFC corn compared with coarsely and finely ground corn, respectively . Cows fed SFC or FGC produced 4% more milk with lower fat content compared with coarsely ground corn . Increasing the feeding frequency did not improve milk fat content . The fat-corrected milk yield was not different among treatments . Feeding SFC resulted in a low acetate-to-propionate ratio in the rumen fluid than FGC . Cows fed SFC produced 45 and 115 g more milk protein per cow/d than cows fed FGC or coarse, respectively . With the value of increased milk protein observed in this study, it would be more economical to feed SFC or finely ground corn to dairy cows compared with coarse ground . The breakeven price of flaking corn in this study was $32 and $12/metric tonne compared with coarse and FGC, respectively . Based on a survey conducted by the authors, the price of flaking corn in the United States ranged between $7 to $22/metric tonne during year 2000. J Dairy Sci, 2002 Jan, 85(1), 204 - 16 Effects of abomasal infusions of histidine, glucose, and leucine on milk production and plasma metabolites of dairy cows fed grass silage diets; Huhtanen P et al.; Our previous study showed that His was the first-limiting amino acid (AA) for milk protein production in cows fed grass silage and cereal-based supplement . The aim of this study was to identify the second-limiting AA and determine whether glucose was limiting responses to His . Abomasal infusion of His (6.5 g/d), glucose (250 g/d), His (6.5 g/d) + glucose (250 g/d), His 6.5 g/d) + Leu (12 g/d) and His (6.5 g/d) + Leu (12 g/d) + glucose (250 g/d) on milk production and utilization of amino acids by mammary gland was in an incomplete 5 x 6 Latin square design with 14-d periods . The diet was based on restrictively fermented grass silage fed ad libitum and 8 kg/d of concentrate comprised of barley, oats, unmolassed sugar beet pulp, urea, and minerals . The infusions did not affect feed intake, diet digestibility, or rumen fermentation pattern . The molar proportion of propionate in rumen VFA was low (15.5%), suggesting that glucose supply from the basal diet could be limiting . Milk and milk protein yields were increased by His infusion . Infusion of His increased plasma His concentration from 19 to 52 microM but decreased extraction efficiency of His . Infusion of glucose increased plasma glucose concentration, milk lactose concentration, and yield and tended to increase milk protein yield . Responses in milk protein yield to combined infusions of His and glucose were additive, suggesting that the utilization of the first-limiting AA His was limited by glucose supply . Infusion of Leu increased plasma Leu concentration but did not produce any further milk protein yield response compared with the infusions without Leu . It was concluded that the efficiency of utilization of the first-limiting AA His could be improved by increasing the supply of glucose, when the basal diet produces a rumen fermentation pattern low in propionate . Leu was not the second-limiting AA in cows fed grass silage-based diets. Plant Foods Hum Nutr, 2002 Winter, 57(1), 63 - 71 Biochemical and sensory evaluation of wheat bran supplemented sorghum bread; Mallasy LO et al.; Addition of wheat bran to sorghum flour (Dabar cultivar) at two extraction rates 72% and 80% resulted in lowering reducing sugars . The percent decrease was 75.6% compared with the control at the end of fermentation period . There was a highly significant (p < or = 0.05) increase in crude fiber content as a result of addition of wheat bran . The increase was from 0.8 to 5.2 and from 0.5 to 5.3% for the 80% S/WB and 72% S/WB blends, respectively . Sorghum bread containing wheat bran was lower in reducing sugars and showed a significant increase (p < 0.05) in starch content . Sorghum bread containing wheat bran resulted in a lower in vitro protein and starch digestibilities. J Nat Prod, 2002 Feb, 65(2), 108 - 14 Emmyguyacins A and B: unusual glycolipids from a sterile fungus species that inhibit the low-pH conformational change of hemagglutinin A during replication of influenza virus; Boros C et al.; Two novel glycolipids, emmyguyacin A (1a) and emmyguyacin B (1b), were isolated at concentrations of 1.51 g/L from a potato dextrose agar fermentation of a sterile fungus species . The compounds inhibit replication of influenza A virus (A/X31) in MDCK cells by inhibiting the pH-dependent conformational change of hemagglutinin A (IC(50) 9 microM) . The structures were deduced using one- and two-dimensional NMR techniques and mass spectrometric analyses on both the parent compounds and a host of degradation products and derivatives . A novel and unusual oxalic acid ester of a monohydroxylated fatty acid (5, 17-oxalyloxydocosanoic acid) is reported . The first isolation and characterization of the fatty acid 17-hydroxydocosanoic acid (3) itself is also reported as a saponification product of 1. J Antibiot (Tokyo), 2001 Dec, 54(12), 1025 - 30 CJ-13,981 and CJ-13,982, new squalene synthase inhibitors; Watanabe S et al.; Two new squalene synthase (SSase) inhibitors, CJ-13,981 (I) and CJ-13,982 (II), were isolated from the fermentation broth of an unidentified fungus CL 15036 . They inhibited human liver microsomal SSase with IC50s of 2.8 and 1.1 microM, respectively, but showed no inhibitory activity against human brain protein farnesyltransferase (PFTase) at 100 microM . Based on FAB-MS and NMR analyses, the structures of I and II were determined to be 3-hydroxy-3,4-dicarboxy-15-hexadecenoic acid and 3-hydroxy-3,4-dicarboxyhexadecanoic acid, respectively. J Antibiot (Tokyo), 2001 Dec, 54(12), 1019 - 24 Neuroprotectins A and B, bicyclohexapeptides protecting chick telencephalic neurons from excitotoxicity . II . Structure determination; Kobayashi H et al.; In the course of our search for neuroprotective agents of microbial origin against kainate-induced neurotoxicity, we have succeeded in the isolation of two new bicyclohexapeptides, neuroprotectins A and B, together with a known compound, complestatin, from the fermentation broth of Streptomyces sp . Q27107 . They are closely related in structure to complestatin and possess an oxindolylalanine moiety in place of the tryptophan residue present in complestatin. J Antibiot (Tokyo), 2001 Dec, 54(12), 1013 - 8 Neuroprotectins A and B, bicyclohexapeptides protecting chick telencephalic neuronal cells from excitotoxicity . I . Fermentation, isolation, physico-chemical properties and biological activity; Kobayashi H et al.; Glutamate, an excitatory amino acid, is known to induce neurotoxicity in central nervous system under abnormal conditions such as ischemia, hypoglycemia, epilepsy, Huntington's chorea, Parkinson's disease and Alzheimer's disease . In our search for neuroprotective agents of microbial origin against excitatory neurotoxins, we have isolated two new bicyclohexapeptides, neuroprotectins A and B, together with a known compound complestatin, from the fermentation broth of Streptomyces sp . Q27107 . Neuroprotectins protected primary cultured chick telencephalic neurons from glutamate- and kainate-induced excitotoxicities in a dose-dependant fashion. Bioresour Technol, 2002 Mar, 82(1), 87 - 93 Effect of pH on hydrogen production from glucose by a mixed culture; Fang HH et al.; The effect of pH on the conversion of glucose to hydrogen by a mixed culture of fermentative bacteria was evaluated . At 36 degrees C, six hours hydraulic retention, over 90% of glucose was degraded at pH ranging 4.0-7.0, producing biogas and an effluent comprising mostly fatty acids . At the optimal pH of 5.5, the biogas comprised 64 +/- 2% of hydrogen with a yield of 2.1 +/- 0.1 mol-H2/mol-glucose and a specific production rate of 4.6 +/- 0.4 l-H2/(g-VSS day) . The effluent was composed of acetate (15.3-34.1%) and butyrate (31.2-45.6%), plus smaller quantities of other volatile fatty acids and alcohols . The diversity of microbial communities increased with pH, based on 16S rDNA analysis by denaturing gradient gel electrophoresis (DGGE). Biochim Biophys Acta, 2002 Jan 15, 1569(1-3), 139 - 50 A novel two-step extraction method with detergent/polymer systems for primary recovery of the fusion protein endoglucanase I-hydrophobin I; Collen A et al.; Extraction systems for hydrophobically tagged proteins have been developed based on phase separation in aqueous solutions of non-ionic detergents and polymers . The systems have earlier only been applied for separation of membrane proteins . Here, we examine the partitioning and purification of the amphiphilic fusion protein endoglucanase I(core)-hydrophobin I (EGI(core)-HFBI) from culture filtrate originating from a Trichoderma reesei fermentation . The micelle extraction system was formed by mixing the non-ionic detergent Triton X-114 or Triton X-100 with the hydroxypropyl starch polymer, Reppal PES100 . The detergent/polymer aqueous two-phase systems resulted in both better separation characteristics and increased robustness compared to cloud point extraction in a Triton X-114/water system . Separation and robustness were characterized for the parameters: temperature, protein and salt additions . In the Triton X-114/Reppal PES100 detergent/polymer system EGI(core)-HFBI strongly partitioned into the micelle-rich phase with a partition coefficient (K) of 15 and was separated from hydrophilic proteins, which preferably partitioned to the polymer phase . After the primary recovery step, EGI(core)-HFBI was quantitatively back-extracted (K(EGIcore-HFBI)=150, yield=99%) into a water phase . In this second step, ethylene oxide-propylene oxide (EOPO) copolymers were added to the micelle-rich phase and temperature-induced phase separation at 55 degrees C was performed . Total recovery of EGI(core)-HFBI after the two separation steps was 90% with a volume reduction of six times . For thermolabile proteins, the back-extraction temperature could be decreased to room temperature by using a hydrophobically modified EOPO copolymer, with slightly lower yield . The addition of thermoseparating co-polymer is a novel approach to remove detergent and effectively releases the fusion protein EGI(core)-HFBI into a water phase. Berl Munch Tierarztl Wochenschr, 2002 Jan-Feb, 115(1-2), 6 - 10 Effect of nisin and monensin on rumen fermentation in the artificial rumen; Jalc D et al.; The objective of this study was to investigate the effect of nisin and monensin on rumen fermentation of diets containing hay and barley (80:20%) in artificial rumen (Rusitec system) . The Rusitec system consisted of four fermentation vessels (V1, V2, V3, V4): V1 was without additives (control), V2 received daily 2 mg of nisin, V3 involved 5 mg of monensin and V4 combination of 2 mg of nisin with 5 mg of monensin . After an adaptation period (7 days), the fermentation parameters were determined for six consecutive days . Compared to control diet, the addition of nisin resulted in an increase (P < 0.05) of hemicellulose degradation, acetate, propionate (mmol.day-1) production and energetic efficiency of VFA (E), decrease of butyrate production . Nisin had no effect on dry matter (DM), organic matter (OM), cellulose and detergent fiber degradation, production of total gas, methane and efficiency of microbial synthesis . The addition of monensin resulted in an decrease of DM, OM (P < 0.05), cellulose, hemicellulose, detergent fiber degradation (P < 0.001), total gas, methane and ammonia nitrogen (NH3-N) production . Monensin also significantly decreased acetate, butyrate, L-lactate (mmol.day-1) production and it increased propionate production (P > 0.001) and efficiency of microbial synthesis . The combined effect of nisin and monensin in V4 was similar to the effect of monensin in V3 compared to control . Then, the effect of additive monensin was dominant over nisin . In conclusion, our results indicate that nisin was less effective than monensin on some fermentation parameters (important for the improvement of the efficiency of utilization of the diet by ruminants) in artificial rumen. Arch Tierernahr, 2001, 54(1), 47 - 59 Sugar beet mash silage as a component of a total-mixed-ration for dairy cows--effects on parameters of digestion and animal performance; Schmidt S et al.; Sugar beet mash silage (BMS) was offered in amounts up to 35% of DM to dairy cows as a component of a total-mixed-ration (TMR) . Barley and molasses in the control ration were replaced by BMS half in ration BMS 1/2 and in total in ration BMS 1/1 on the basis of the calculated contents of net energy for lactation . Two trials were carried out . In trial I each ration was tested on parameters of rumen fermentation and digestibility of crude nutrients using 4 Holstein cows with rumen fistula . Chewing activity was tested on 2 Holstein cows for each ration . With the BMS rations the ruminal NH3 concentration was lower and the drop in pH was less than in the control ration . The pattern of volatile fatty acids in the rumen fluid from the BMS groups tended towards more propionic and butyric acid . The feeding of BMS showed no negative impact on chewing and rumination . Energy digestibility raised significantly from 59.8% in the control ration to 72.6% in the BMS 1/1 ration . In trial II the same rations were fed in a change-overdesign to a herd of 24 Holstein cows to test feed intake and animal performance . The results showed no significant effects of BMS rations on DM intake and milk production . The results of both trials indicate that even high amounts of cereals can be replaced by BMS without negative effects on rumen fermentation, milk yield and milk composition with slight drop in fat content . For a better handling of BMS, it is of advantage to include it in a TMR. Arch Tierernahr, 2001, 54(1), 33 - 46 Effect of antibiotics, 2-bromoethanesulfonic acid and pyromellitic diimide on methanogenesis in rumen ciliate cultures in vitro; Varadyova Z et al.; The effects of penicillin G, streptomycin, chloramphenicol, 2-bromoethanesulfonic acid and pyromellitic diimide on total gas, methane, volatile fatty acid production and food degradability after 24 h of incubation in vitro were investigated in the cultures of two rumen ciliates . The inocula of both rumen ciliates Entodinium caudatum and Epidinium ecaudatum were used at a volume of 34 ml into the 50 ml glass syringes together with the feed and compounds tested . Despite penicillin G--streptomycin treatment methane production in both cultures was significantly decreased by the inhibitors for Epidinium ecaudatum . Methane production of the bacterial fraction of both protozoan species was significantly lower than in the whole cultures . No epifluorescence of methanogens on (or in) the cells of Entodinium caudatum was observed in contrast to Epidinium with which strong epifluorescence of methanogens on the cell surface was detected . Microscopic observation could indicate that the methane production by Entodinium caudatum was probably caused by their intracellular methanogenic activity, while methane production by Epidinium ecaudatum could be related to both the methanogenic bacterial fraction from their external surface and probably also to intracellular activity . Decreased feed degradability and differences in the fermentation end products accompanied the inhibition of methanogenesis in both in vitro cultures . Entodinium caudatum appeared to be more sensitive than Epidinium ecaudatum to the compounds tested. Biosens Bioelectron, 2002 Apr, 17(4), 305 - 13 A generic strategy for subcloning antibody variable regions from the scFv phage display vector pCANTAB 5 E into pASK85 permits the economical production of F(ab) fragments and leads to improved recombinant immunoglobulin stability; Kramer K et al.; Apart from the decisive sensitivity and specificity of immunosensors, the employed antibodies essentially contribute to additional key factors like fabrication costs for sensor chips and sensor stability . A production scheme for recombinant antibody fragments has been optimised with respect to these particular issues of biosensor development . The phagemid vector pCANTAB 5 E is widely used for the selection of antibody fragments from corresponding libraries . However, large-scale production of the selected single-chain F(v) (scFv) fragments is substantially restricted by the high cost for the inducer IPTG and the anti-E-tag antibody . The latter is needed in significant amounts for the purification of the recombinant protein . A generic strategy was established for subcloning scFv variable regions from pCANTAB 5 E into the plasmid pASK85 for the expression of F(ab) fragments . pASK85 bears coding sequences for murine constant domains including a His(6) tag at the carboxyl-terminal end of the constant heavy chain domain . The anti-s-triazine antibody K47H served as a model system in this study . Biosynthesis of the F(ab) fragment in a high cell density fermenter was induced by addition of anhydrotetracycline . The F(ab) fragment was subsequently purified from the periplasmic extract in a single step by immobilized metal affinity chromatography (IMAC) . A yield of 100 microg/lxOD(550) purified F(ab) fragment was obtained employing a standard fermentation scheme . The sensitivity and cross-reactivity of the F(ab) was comparable to the parent scFv when assayed by enzyme immunoassay . However, the F(ab) fragment exhibited significantly improved long-term stability. Biosens Bioelectron, 2002 Apr, 17(4), 297 - 302 Biosensor arrays for simultaneous measurement of glucose, lactate, glutamate, and glutamine; Moser I et al.; For simultaneous measurement of glucose, lactate, glutamine, and glutamate a biosensor array is implemented in a micro flow-system thus giving a microsystem . The microsystem consists of a glass chip with the integrated biosensor array and a bottom part, which comprises a gold counter electrode, a 300 microm thick seal, and electrical interconnection lines . The flow device has a total internal volume of 2.1 or 6 microl when integrated with a mixer on chip . The biosensors with no crosstalking and high long term stability were produced by modifying the electrochemical transducers and utilizing photopatternable enzyme membranes . The use of appropriate miniaturization technology leads to mass producable devices for in vivo and ex vivo applications in whole blood and fermentation broth . Due to a novel glutaminase with an activity optimum in the neutral pH range direct and simultaneous monitoring of glutamine together with glucose, lactate, and glutamate could be performed. Lett Appl Microbiol, 2002, 34(1), 67 - 71 Production of Aspergillus terreus alpha-L-rhamnosidase by solid state fermentation; Elinbaum S et al.; AIMS: The study of production of Aspergillus terreus CECT 2663 alpha-L-rhamnosidase in solid state fermentation using wheat bran, washed sugar cane bagasse and polyurethane foam as substrates or supports for the enzyme production . METHODS AND RESULTS: Cultures were carried out in Petri dishes under controlled temperature and humidity . Naringin or rhamnose were the enzyme inducers and carbon sources . The enzyme activity to inducer ratio was appreciably greater when using sugar cane bagasse or polyurethane foam than wheat bran . The influence of inoculum size, inducer, airflow, humidity and temperature were determined . Under optimum conditions, about four units of enzyme per ml nutrient solution were obtained after 4-6 d . CONCLUSIONS: The activity to inducer ratio was higher, and the cultivation time was shorter in solid state fermentation than those observed in submerged cultures . SIGNIFICANCE AND IMPACT OF THE STUDY: Solid cultures, using naringin as inducer, can be appropriate alpha-L-rhamnosidase production. Lett Appl Microbiol, 2002, 34(1), 56 - 61 Mycelial growth and exo-biopolymer production by submerged culture of various edible mushrooms under different media; Kim SW et al.; AIMS: The effect of synthetic media on the submerged mycelial growth and exo-biopolymer production in various edible mushrooms was investigated in shake flask culture . METHODS AND RESULTS: Among 19 mushrooms examined, the relatively high yield in mycelial biomass and exo-biopolymer production was achieved in potato malt peptone (PMP) medium . In particular, Ganoderma lucidum NO . 1 and Phellinus linteus KCTC 6190 showed favourable growth in PMP medium with exo-biopolymer concentration of 1170 and 1520 mg l(-1), respectively . CONCLUSIONS: Enhanced exo-biopolymer production was achieved from Ganoderma lucidum NO . 1 and Phellinus linteus KCTC 6190 in a 5L batch fermentor, indicating approximately 5000 and 2410 mg l(-1), respectively . SIGNIFICANCE AND IMPACT OF THE STUDY: The exo-biopolymer production and mycelial growth from various mushrooms were found to be strongly controlled by different complex media. Lett Appl Microbiol, 2002, 34(1), 51 - 5 Exopolysaccharide production from Sclerotium glucanicum NRRL 3006 and Botryosphaeria rhodina DABAC-P82 on raw and hydrolysed starchy materials; Selbmann L et al.; AIMS: Evaluation of fermentative usage of raw starchy materials for exopolysaccharide (EPS) production by Sclerotium glucanicum NRRL 3006 and Botryosphaeria rhodina DABAC-P82 . METHODS AND RESULTS: Non-hydrolysed corn starch, soft wheat flour, potato flour, cassava flour, sweet and industrial potato flours, and corn starch hydrolysed to different dextrose equivalent (DE) were tested in shaken culture for EPS production . Both fungal strains produced EPS on all tested materials but the production was maximum on hydrolysed corn starch (30.5 and 19.8 g l(-1) by B . rhodina and S . glucanicum on corn starch at 100 and 62 DE, respectively) . CONCLUSIONS: Raw starchy materials as such and, in particular, partially or totally hydrolysed corn starch could be used profitably for EPS production by S . glucanicum and B . rhodina . SIGNIFICANCE AND IMPACT OF THE STUDY: The excellent EPS production, productivity and yield of B . rhodina DABAC-P82 when grown on 60 g l(-1) of totally hydrolysed corn starch. J Appl Microbiol, 2002, 92(2), 329 - 37 Effects of temperature, pH, water activity and CO2 concentration on growth of Rhizopus oligosporus NRRL 2710; Sparringa RA et al.; AIMS: To investigate the effects of temperature, pH, water activity (aw) and CO2 concentration on the growth of Rhizopus oligosporus NRRL 2710 . METHODS AND RESULTS: Hyphal extension rates from mycelial and spore inocula were measured on media with different aw (approximately 1.0, 0.98 and 0.96) and pH (3.5, 5.5 and 7.5) incubated at 30, 37 or 42 degrees C in atmospheres containing 0.03, 12.5 or 25% (v/v) CO2 . The effects of environmental conditions on hyphal extension rate were modelled using surface response methodology . The rate of hyphal extension was very sensitive to pH, exhibiting a pronounced optimum at pH 5.5-5.8 . The hyphal extension rate was less sensitive to temperature, aw or CO2, exhibiting maximum rates at 42 degrees C, a(w) approximately 1.0 and 0.03% (v/v) CO2 . CONCLUSIONS: The fastest hyphal extension rate (1.7 mm h(-1)) was predicted to occur at 42 degrees C, pH 5.85, a(w) approximately 1.0 and 0.03% CO2 . SIGNIFICANCE AND IMPACT OF THE STUDY: The present work is the first to model the simultaneous effects of temperature, pH, aw and CO2 concentration on mould growth . The information relates to tempe fermentation and to possible control of the microflora in Tanzanian cassava heap fermentations. Bioresour Technol, 2002 Mar, 82(1), 65 - 71 Effects of temperatures and organic loading rates on biomethanation of acidic petrochemical wastewater using an anaerobic upflow fixed-film reactor; Patel H et al.; The effect of temperature and organic loading rate on the rate of methane production from acidic petrochemical wastewater without neutralization was investigated by continuously feeding an anaerobic upflow fixed-film reactor . The temperatures selected for the studies were 25, 37, 45 and 55 degrees C . Organic loading rate (OLR) for each temperature was varied from 3.6 to 21.7 kg COD m(-3) d(-1) . Best performance with respect to COD and BOD reduction, total gas production and methane yield was obtained with the reactor operating at 37 degrees C . OLR could be increased to a maximum of 21.7 kg COD m(-3) d(-1) with 90-95% COD and BOD reduction and methane yield of 0.450 m3 kg(-1) COD d(-1) added . The reactor operating at 55 degrees C gave the highest methane yield of 0.666 m3 kg(-1) COD d(-1) at an OLR of 6 kg COD m(-3) d(-1) . This decreased to 0.110 m3 kg(-1) COD d(-1) when the OLR was increased to 18.1 kg COD m(-3) d(-1) . The reactor operating at 45 degrees C gave a maximum methane yield of 0.416 m3 kg(-1) COD d(-1) added at an OLR of 6 kg COD m(-3) d(-1) . On further increasing the OLR to 9 kg COD m(-3) d(-1), COD reduction was 89%, however, methane yield decreased to 0.333 m3 kg(-1) COD d(-1) added . The highest methane yield of 0.333 m3 kg(-1) COD d(-1) added at an OLR of 6 kg COD m(-3) d(-1) was obtained with reactors operating at 25 degrees C . These studies indicate potential rates of methane production from acidic petrochemical wastewater under different temperatures . This provides a guideline for various kinetic analyses and economic evaluation of the potential feasibility of fermenting acidic wastewater to methane. Am J Ther, 1995 Mar, 2(3), 196 - 216 Pentostatin (2prime prime or minute-Deoxycoformycin): Clinical Pharmacology, Role In Cancer Chemotherapy, and Future Prospects; Spiers AS; Pentostatin (2prime prime or minute-deoxycoformycin, dCF) is a product of the fermentation of Streptomyces antibioticus . It is a tight-binding inhibitor of adenosine deaminase (ADA), an enzyme essential in cellular metabolism of purines . Children with congenital absence of ADA suffer from atrophy of lymphoid tissues and severe combined immune deficiency (SCID) syndrome . It was speculated that pentostatin would be lymphocytotoxic, and this proved to be the case, promoting its investigation in lymphoid neoplasms . It was anticipated that pentostatin would be most active in neoplasms with high intracellular concentrations of ADA---e.g., acute lymphocytic leukemia (ALL), particularly its T cell variety . Although pentostatin proved to be active in ALL, large doses were required and toxic effects outweighted therapeutic benefits . By contrast, pentostatin proved to be exceptionally active in hairy cell leukemia (HCL), a B cell neoplasm with low intracellular concentrations of ADA . Pentostatin has since been shown to possess activity in chronic lymphocytic leukemia, prolymphocytic leukemia, cutaneous T cell lymphomas, adult T cell lymphoma-leukemia, and low-grade non-Hodgkin's lymphomas . It potentiates the activity of vidarabine against viruses and against the cells of acute myeloid leukemia . Pentostatin is inactive in melanoma and renal carcinoma, but has not been adequately evaluated in other solid tumors . The toxic effects of pentostatin include renal failure, central nervous system (CNS) depression, immunosuppression, keratoconjunctivitis, and opportunistic infections . In the absence of pre-existing bone marrow compromise, pentostatin produces only mild myelosuppression . Aside from its use as an antineoplastic agent, pentostatin has potential applications as an immunosuppresive drug, as an antiviral agent, as an antimalarial compound, and in the protection of cells of the CNS from damage induced by ischemia and anoxia. EMBO J, 2002 Feb 15, 21(4), 572 - 9 Conserved properties of hydrogenosomal and mitochondrial ADP/ATP carriers: a common origin for both organelles; van der Giezen M et al.; Mitochondria are one of the hallmarks of eukaryotic cells, exporting ATP in exchange for cytosolic ADP using ADP/ATP carriers (AAC) located in the inner mitochondrial membrane . In contrast, several evolutionarily important anaerobic eukaryotes lack mitochondria but contain hydrogenosomes, peculiar organelles of controversial ancestry that also supply ATP but, like some fermentative bacteria, make molecular hydrogen in the process . We have now identified genes from two species of the hydrogenosome-containing fungus Neocallimastix that have three-fold sequence repeats and signature motifs that, along with phylogenetic analysis, identify them as AACs . When expressed in a mitochondrial AAC- deficient yeast strain, the hydrogenosomal protein was correctly targeted to the yeast mitochondria inner membrane and yielded mitochondria able to perform ADP/ATP exchange . Characteristic inhibitors of mitochondrial AACs blocked adenine nucleotide exchange by the Neocallimastix protein . Thus, our data demonstrate that fungal hydrogenosomes and yeast mitochondria use the same pathway for ADP/ATP exchange . These experiments provide some of the strongest evidence yet that yeast mitochondria and Neocallimastix hydrogenosomes are but two manifestations of the same fundamental organelle. Scand J Gastroenterol, 2002 Jan, 37(1), 74 - 9 Phenol toxicity and conjugation in human colonic epithelial cells; Pedersen G et al.; BACKGROUND: Colonic epithelial cells are exposed to a range of potentially harmful luminal factors . including phenols, but it is unresolved whether these compounds impair the integrity of the epithelium . The aim of this study was to describe the effect of phenol exposure on human colonic epithelial cells in vitro and the conjugation pathways involved in detoxification . METHODS: Primary human colonic epithelial cell cultures or HT-29 cell cultures were exposed to paracetamol, dinitrophenol or phenol (0.1-5 mM) for 24 h . Cell viability was measured using the methyltetrazoleum test . Phenol conjugation products released from cell cultures were identified by high-pressure liquid chromatography . Phenol glucuronidase (PGD) and sulphotransferase (PST) enzyme activities were measured in isolated cell homogenates . RESULTS: Paracetamol, dinitrophenol and phenol (>1.25 mM) significantly impaired the viability of primary colonic epithelial cell cultures . No differences between cell cultures from ulcerative colitis and control patients were observed . Paracetamol (5 mM) also induced significant cell damage in HT-29 cells . Glucuronidation was the preferred conjugation pathway in both cell models, despite the presence of PGD and PST activity . CONCLUSION: Phenols have a direct toxic effect on human colonic epithelial cells in vitro, which supports the view that dietary fermentation metabolites may be involved in the modulation of chronic bowel inflammation. Eur J Nutr, 2001 Oct, 40(5), 189 - 99 History of nutrition and acid-base physiology; Manz F; In the 17th century the notion of nutrition and diet changed in northern European countries . First chemical experiments fostered the idea that salts resulted from a union of acids and bases . Digestion was no more regarded as a process of cooking but a succession of fermentations controlled by a balanced production of acids and alkali . Life seemed to depend on the equilibrium of acids and alkalis . In the 19th century food was systematically analysed for the content of energy and macronutrients and first scientifically based nutritional standards were formulated . The preferred use of processed food from the new food industry resulted in epidemics of nutritional disorders . Acidosis seemed to be a plausible pathogenic factor . Practitioners (S Ishizuka, H Hay, FX Mayr) formulated holistic doctrines integrating the concept of balance of acids and bases and recommending food with an excess of alkali . New micromethods to determine the concentration of electrolytes and blood acid-base status promoted physiological and clinical research into acid-base metabolism in the 1960s . In the new physiologically based terminology of systemic acid-base status, the relationship between blood acid-base status and net acid intake or excretion was, however, incorrectly simplified . In the 1970s metabolic acidosis was observed in patients on chemically defined diets and parenteral nutrition . Based on the data of comprehensive acid-base balance studies, calculation models were used to estimate renal net acid excretion from nutrient intake and to predict the potential renal acid load of single foods . Extrapolating current trends to the future, one can say that acid-base physiology will probably remain a challenge in nutrition and functional medicine over the next few years . The challenge will include new concepts for the manipulation of nutritional acid load in sports, dietetics and preventive medicine as well as new definitions of the upper intake level of potential renal acid load in functional foods and the monitoring of renal net acid excretion in populations. Protein Eng, 2002 Jan, 15(1), 29 - 33 The effect of proline insertions on the thermostability of a barley alpha-glucosidase; Muslin EH et al.; The thermal stability of alpha-glucosidase is important because the conversion of starch to fermentable sugars during industrial production of ethanol (e.g . brewing, fuel ethanol production) typically takes place at temperatures of 65-73 degrees C . In this study we investigate the thermostability of alpha-glucosidases from four plant species, compare their deduced amino acid sequences, and test the effect of substituting a proline for the residue present in the wild-type enzyme on the thermostability of alpha-glucosidase . The alpha-glucosidase from barley (Hordeum vulgare) was significantly less thermostable than the other three alpha-glucosidases . A comparison of the published deduced amino acid sequences of these four alpha-glucosidases revealed conserved proline residues in the three most thermostable alpha-glucosidases that were not found in the barley enzyme . Site-directed mutagenesis was done on recombinant barley alpha-glucosidase to create proteins with prolines at these conserved positions . The thermostability (T(50)) of one of these mutant enzymes, T340P, was 10 degrees C higher than the non-mutated enzyme. Aviakosm Ekolog Med, 2001, 35(5), 3 - 10 {Immobilized biocatalysts in biotechnological processes}; Nazarov NM; In recent years, biotechnology has been stuck with the problem of choosing the most technologically convenient way of utilization of biocatalysts using bacterial cells or structural formations . Immobilization of bio-objects on solid carriers uses the advantages of heterogenic catalysis over homogenous like simple separation of byocatalyst from end product on completion of biotechnological process, and highly effective transformation or synthesis of organic compounds through biomass concentration in fermenter . In addition, immobilized biocatalysts exhibit better stability in case of exposure to aggressive temperatures, active environmental reactions, and "attack" of foreign microflora . Immobilized biocatalyzers can operate repeatedly and owing to the unique specificity show promise for wide application in aerospace and environmental medicine. J Pediatr Gastroenterol Nutr, 2002 Feb, 34(2), 158 - 64 Modeling 13C breath curves to determine site and extent of starch digestion and fermentation in infants; Christian MT et al.; BACKGROUND: The colon salvages energy from starch, especially when the capacity of the small intestine to digest it is limited . The aim of this study was to determine the site and relative extent of starch digestion and fermentation in infants . METHODS: Thirteen infants (10 male and 3 female infants), median age 11.8 months (range, 7.6-22.7 months), were fed a starchy breakfast containing 13C-labeled wheat flour after an overnight fast . Duplicate breath samples were obtained before breakfast and every 30 minutes for 12 hours . Breath 13CO2 enrichment was measured using isotope ratio mass spectrometry, and results were expressed as percentage dose recovered (PDR) for each 30 minutes . The PDR data were analyzed and mathematically modeled assuming either a constant estimate of CO2 production rate or adjusted for physical activity . RESULTS: Mean +/- SD cumulative 13C PDR (cPDR) at 12 hours was 21.3% +/- 8.4% for unadjusted data and 26.5% +/- 11.6% for adjusted data . A composite model of two curves fit significantly better than a single curve . Modeling allowed estimation of cPDRs of small intestine (17.5% +/- 6.5% and 22.7% +/- 9.3% for unadjusted and adjusted data, respectively) and colon (4.6% +/- 2.9% and 6.3% +/- 5.4%) . CONCLUSIONS: Modeling of 13CO2 enrichment curves after ingestion of 13C-enriched wheat flour is an attractive means to estimate the contribution of the upper and lower gut to starch digestion and fermentation. Arzneimittelforschung, 2002, 52(1), 60 - 6 Lekteplase--a secreted tissue plasminogen activator derivative from Escherichia coli; Manosroi J et al.; Fermentation studies of batch-mode cultivation in 4-L fermenters were carried out to obtain an active recombinant DNA-derived tissue plasminogen activator (t-PA) deletion mutant, lekteplase, secreted and correctly folded from Escherichia coli . The OmpA signal sequence was used to deliver the heterologous product composed of kringle 2 plus serine protease domain (K2S) to the medium . Supplementing the complex medium with 10% glycerol and 20 mmol/l magnesium chloride led to an increase in cell numbers with final cell density reaching an OD600 of 24 . The expression level of lekteplase in the medium detected by sandwich ELISA was 100 mg/L . Enzymatic activity of lysine-sepharose purified product was demonstrated by amidolytic assay, in vitro fibrin clot lysis, and copolymerization PAGE. Int J Syst Evol Microbiol, 2002 Jan, 52(Pt 1), 281 - 4 Kockovaella barringtoniae sp . nov., a new basidiomycetous yeast species isolated from a plant leaf collected in a tropical rain forest in Thailand; Fungsin B et al.; One yeast strain characterized by the proliferation of non-ballistosporous stalked conidia, budding cells and ballistoconidia, the presence of xylose in whole-cell hydrolysates, the presence of Q-10 as the major ubiquinone isoprenologue, the inability to ferment sugars and positive diazonium blue B (DBB) and urease reactions was isolated from a plant sample collected in a tropical rain forest in Thailand . The isolate clustered with Kockovaella species in the 18S rDNA-based tree . On the basis of the morphological, biochemical and molecular phylogenetic characteristics, the isolate was assigned to the genus Kockovaella . DNA complementarity experiments showed that the isolate was genetically distinct from known species of the genus Kockovaella . The isolate is described as Kockovaella barringtoniae sp . nov . The type strain is strain TY-278T (= JCM 10998T = TISTR 5770T). Int J Oncol, 2002 Mar, 20(3), 563 - 70 Fermented wheat germ extract induces apoptosis and downregulation of major histocompatibility complex class I proteins in tumor T and B cell lines; Fajka-Boja R et al.; The fermented wheat germ extract (code name: MSC, trade name: Avemar), with standardized benzoquinone content has been shown to inhibit tumor propagation and metastases formation in vivo . The aim of this study was to understand the molecular and cellular mechanisms of the anti-tumor effect of MSC . Therefore, we have designed in vitro model experiments using T and B tumor lymphocytic cell lines . Tyrosine phosphorylation of intracellular proteins and elevation of the intracellular Ca2+ concentration were examined using immunoblotting with anti-phosphotyrosine antibody and cytofluorimetry by means of Ca2+ sensitive fluorescence dyes, Fluo-3AM and FuraRed-AM, respectively . Apoptosis was measured with cytofluorimetry by staining the DNA with propidium iodide and detecting the cell population . The level of the cell surface MHC class I molecules was analysed with indirect immunofluorescence on cytofluorimeter using a monoclonal antibody to the non-polymorphic region of the human MHC class I . MSC stimulated tyrosine phosphorylation of intracellular proteins and the influx of extracellular Ca2+ resulted in elevation of intracellular Ca2+ concentration . Prominent apoptosis of 20-40% was detected upon 24 h of MSC treatment of the cell lines . As a result of the MSC treatment, the amount of the cell surface MHC class I proteins was downregulated by 70-85% compared to the non-stimulated control . MSC did not induce a similar degree of apoptosis in healthy peripheral blood mononuclear cells . Inhibition of the cellular tyrosine phosphatase activity or Ca2+ influx resulted in the opposite effect increasing or diminishing the Avemar induced apoptosis as well as the MHC class I downregulation, respectively . A benzoquinone component (2,6-dimethoxi-p-benzoquinone) in MSC induced similar apoptosis and downregulation of the MHC class I molecules in the tumor T and B cell lines to that of MSC . These results suggest that MSC acts on lymphoid tumor cells by reducing MHC class I expression and selectively promoting apoptosis of tumor cells on a tyrosine phosphorylation and Ca2+ influx dependent way . One of the components in MSC, 2,6-dimethoxi-p-benzoquinone was shown to be an important factor in MSC mediated cell response. Biotechnol Bioeng, 2002 Mar 30, 77(7), 815 - 26 Dependence of morphology on agitation intensity in fed-batch cultures of Aspergillus oryzae and its implications for recombinant protein production; Amanullah A et al.; We previously reported that, although agitation conditions strongly affected mycelial morphology, such changes did not lead to different levels of recombinant protein production in chemostat cultures of Aspergillus oryzae (Amanullah et al., 1999) . To extend this finding to another set of operating conditions, fed-batch fermentations of A . oryzae were conducted at biomass concentrations up to 34 g dry cell weight/L and three agitation speeds (525, 675, and 825 rpm) to give specific power inputs between 1 and 5 kWm(-3) . Gas blending was used to control the dissolved oxygen level at 50% of air saturation except at the lowest speed where it fell below 40% after 60-65 h . The effects of agitation intensity on growth, mycelial morphology, hyphal tip activity, and recombinant protein (amyloglucosidase) production in fed-batch cultures were investigated . In the batch phase of the fermentations, biomass concentration, and AMG secretion increased with increasing agitation intensity . If in a run, dissolved oxygen fell below approximately 40% because of inadequate oxygen transfer associated with enhanced viscosity, AMG production ceased . As with the chemostat cultures, even though mycelial morphology was significantly affected by changes in agitation intensity, enzyme titers (AGU/L) under conditions of substrate limited growth and controlled dissolved oxygen of >50% did not follow these changes . Although the measurement of active tips within mycelial clumps was not considered, a dependency of the specific AMG productivity (AGU/g biomass/h) on the percentage of extending tips was found, suggesting that protein secretion may be a bottle-neck in this strain during fed-batch fermentations . Biotechnol Bioeng, 2002 Mar 30, 77(7), 796 - 805 A mass balance study to assess the extent of contaminant removal achieved in the operations for the primary recovery of plasmid DNA from Escherichia coli cells; Ciccolini LA et al.; Mass balances were performed on an alkaline lysis operation for the primary recovery of supercoiled plasmid DNA as part of a process for plasmid gene preparation . Escherichia coli DH5alpha/pSVbeta was cultured in defined medium by fed-batch fermentation and harvested at the end of the exponential phase . Alkaline lysis of the recombinant cells was performed at fixed shear rates ranging between 46 and 461 s(-1), with neutralization 100 and 300 s after the initiation of the lysis . Mass balance calculations were used to optimize the operating conditions for carrying out the alkaline lysis operation . The results indicated that a plasmid yield of 75% and purity with respect to total DNA of 60% were achievable during the primary recovery operation . The influences of key contaminants, including the soluble proteins and the suspended solids, as they bear on the subsequent purification operations, were evaluated and discussed . Hiroshima J Med Sci, 2001 Dec, 50(4), 83 - 6 Radioprotective effects of miso (fermented soy bean paste) against radiation in B6C3F1 mice: increased small intestinal crypt survival, crypt lengths and prolongation of average time to death; Ohara M et al.; The radioprotective effect of miso, a fermentation product from soy bean, was investigated with reference to the survival time, crypt survival and jejunum crypt length in male B6C3F1 mice . Miso at three different fermentation stages (early-, medium- and long-term fermented miso) was mixed in MF diet into biscuits at 10% and was administered from 1 week before irradiation . Animal survival in the long-term fermented miso group was significantly prolonged as compared with the short-term fermented miso and MF cases after 8 Gy of 60Co-gamma-ray irradiation at a dose rate of 2Gy min(-1) . Delay in mortality was evident in all three miso groups, with significantly increased survival . At doses of 10 and 12 Gy X-irradiation at a dose rate of 4 Gy min(-1), the treatment with long-term fermented miso significantly increased crypt survival . Also the protective influence against irradiation in terms of crypt lengths in the long-term fermented miso group was significantly greater than in the short-term or medium-term fermented miso and MF diet groups . Thus, prolonged fermentation appears to be very important for protection against radiation effects. Appl Microbiol Biotechnol, 2002 Jan, 58(1), 63 - 6 Production of teicoplanin by valine analogue-resistant mutant strains of Actinoplanes teichomyceticus; Jin ZH et al.; Teicoplanin is a glycopeptide antibiotic produced by Actinoplanes teichomyceticus . A strain improvement to increase the productivity of the major component, teicoplanin A2-2, was carried out . As the fatty moiety of teicoplanin A2-2 is derived from L-valine, L-valine analogue (valine hydroxamate)-resistant mutants were derived . One of the mutants, 98-1-227, overproduced valine and produced a higher titer of total teicoplanin with higher A2-2 content . In a pilot fermentor (7 m3), the total productivity of teicoplanin was 1,800 units/ml and the A2-2 content was 58%. Comp Immunol Microbiol Infect Dis, 2002 Jan, 25(1), 49 - 57 Isolation and characterization of Mycoplasma arginini from camels (Camelus dromedarius) with pneumonia; Elfaki MG et al.; Post-mortem examinations of 100 camels with pneumonic lesions were made at a local abattoir for Mycoplasma species . Sixteen isolates with indistinguishable biochemical and immunological characters were identified . The biochemical profile of these isolates showed that they were sensitive to digitonin, negative for urease production, glucose fermentation, and phosphatase activity but were positive for arginine hydrolysis . The identity of these isolates was further confirmed by disk growth inhibition test using a panel of specific antisera against selected reference Mycoplasma spp . Based on the biochemical profile and growth inhibition results, the camel isolates were identified as M . arginini . The pathological findings associated with M . arginini isolation consisted mostly of chronic interstitial pneumonia . The isolation rate of M . arginini from these specimens was 8.8% . These results suggest that the role of M . arginini in pneumonia in camels should be explored in greater detail. J Anim Sci, 2002 Jan, 80(1), 235 - 41 Kinetics of ruminal degradation of wheat and potato starches in total mixed rations; Monteils V et al.; Wheat and potato are rich in starch but their starches differ in their rate of ruminal degradation . Kinetics of in sacco disappearance and profiles of ruminal fermentation were studied for these two concentrates in total mixed rations based on grass silage or corn silage . Wheat starch was more rapidly (34%/h) degraded by rumen microorganisms than potato starch (5%/h) . The differences in starch degradation in sacco were found again in the VFA concentrations, mainly in grass silage-based diets . Overall ruminal pH, total VFA concentration, and proportions of acetate, propionate, and butyrate are more variable for wheat during the kinetic (amplitude and quickness) than for potato in grass silage-based diets . In these diets, risks of acidosis were more elevated with wheat than with potato but the VFA concentrations were also higher . These differences of fermentation profile were so reduced in corn silage-based diets that, in this case, wheat can be substituted by potato without any effect on digestion and no risk of acidosis. Appl Microbiol Biotechnol, 2002 Jan, 58(1), 84 - 8 A spontaneous runaway vector for production-scale expression of bovine somatotropin from Escherichia coi; Trepod CM et al.; An Escherichia coli expression vector was constructed for the production-scale fermentation of recombinant bovine somatotropin (rBST) . Gene expression is regulated by a spontaneous increase in copy number at a constant low temperature without the need for an external inducer . This vector, designated pURA-4, contains the ampicillin resistance gene, the replication origin from pBR322, the R1 temperature-inducible runaway replicon, and a gene encoding rBST . Optimized rBST expression levels of >35% total cell protein were achieved at a constant 28 degrees C . Shake-flask analysis of pURA-4 shows that the copy number spontaneously increases approximately 6-fold during rBST production . Investigation into the mechanism of pURA-4 spontaneous runaway shows that the increase in copy number is directed by the pBR322 ori and not by the R1 replicon . Although the R1 temperature-inducible replicon does not mediate spontaneous runaway, it does have a positive effect on rBST expression . Copy number analysis also confirmed the stability of pURA-4 spontaneous runaway from the shake-flask scale through the production scale. Folia Microbiol (Praha), 2001, 46(4), 333 - 7 Incidence of Candida species isolated from human skin and nails: a survey; Jautova J et al.; Distribution of Candida species was investigated by examining 245 samples from skin lesions and nails . The isolates were identified using standard laboratory methods including germ tube test, micromorphology of colonies on rice agar, the commercial kit, saccharide assimilation and fermentation tests . Eight species of Candida were identified: C . albicans accounted for 56.4% of the isolates, C . parapsilosis 29.1, C . tropicalis 7.8, C . pulcherrima 2.9, C . guilliermondii 1.5, C . krusei and C . zeylanoides for 0.9% each, and C . robusta for 0.5% . The factors significantly associated with colonization were prolonged antibiotic therapy, parenteral nutrition, low birth body mass of infants, intubation, duration of stay in hospital, indwelling intravenous catheter, malignancies, diabetes, surgery, and obesity. Folia Microbiol (Praha), 2001, 46(4), 303 - 8 Production, characterization and properties of polysaccharide depolymerizing enzymes from a strain of Curvularia inaequalis; Gomes E et al.; Xylanase, beta-glucosidase, beta-xylosidase, endoglucanase and polygalacturonase production from Curvularia inaequalis was carried out by means of solid-state and submerged fermentation using different carbon sources . beta-Glucosidase, beta-xylosidase, polygalacturonase and xylanase produced by the microorganisms were characterized . beta-Glucosidase presented optimum activity at pH 5.5 whereas xylanase, polygalacturonase and beta-xylosidase activities were optimal at pH 5.0 . Maximal activity of beta-glucosidase was determined at 60 degrees C, beta-xylosidase at 70 degrees C, and polygalacturonase and xylanase at 55 degrees C . These enzymes were stable at acidic to neutral pH and at 40-45 degrees C . The crude enzyme solution was studied for the hydrolysis of agricultural residues. J Agric Food Chem, 2002 Feb 13, 50(4), 899 - 905 In vitro efficacies of phosphorolytic enzymes synthesized in mycelial cells of Aspergillus niger AbZ4 grown by a liquid surface fermentation; Zyla K et al.; Activities of phytase, a pH 6.0 optimum nonspecific phosphomonoesterase and phosphodiesterase assayed toward bis(p-nitrophenyl)phosphate (phosphodiesterase I) and against p-nitrophenylphosphorylcholine (phosphodiesterase II), were partially purified from mycelial extracts of Aspergillus niger AbZ4 cultivated on a molasses medium by a liquid surface fermentation method . After elimination of phosphate from the medium, 7.3- and 3.5-fold enhancements in specific activities of phytase and phosphodiesterase II were observed . Efficacies of mycelial protein fractions in dephosphorylating a wheat-based broiler feed were determined in vitro according to a procedure that simulated digestion in the intestinal tract of poultry . The addition of 0.052 mg of protein from fractions, each of which was high in either pH 6.0 optimum phosphomonoesterase, phosphodiesterase I, phosphodiesterase II, or phytase per gram of a feed sample resulted in the enhancement of phosphorus release by 10, 11, 27, and 88%, respectively . In the presence of an excess of commercial phytase, the addition of the mycelial fraction high in phytase increased the dephosphorylation rate by 56% . The fraction high in phosphodiesterase II enhanced feed dephosphorylation by 8% in the presence of an excess of commercial phytase and commercial acid phosphatase. J Agric Food Chem, 2002 Feb 13, 50(4), 806 - 12 Generation of Maillard compounds from inulin during the thermal processing of Agave tequilana Weber Var . azul; Mancilla-Margalli NA et al.; During the cooking process of Agave tequilana Weber var . azul to produce tequila, besides the hydrolysis of inulin to generate fermentable sugars, many volatiles, mainly Maillard compounds, are produced, most of which may have a significant impact on the overall flavor of tequila . Exudates (agave juice) from a tequila company were collected periodically, and color, Brix, fructose concentration, and reducing sugars were determined as inulin breakdown took place . Maillard compounds were obtained by extraction with CH(2)Cl(2), and the extracts were analyzed by GC-MS . Increments in color, Brix, and reducing sugars were observed as a function of time, but a decrease in fructose concentration was found . Many Maillard compounds were identified in the exudates, including furans, pyrans, aldehydes, and nitrogen and sulfur compounds . The most abundant Maillard compounds were methyl-2-furoate, 2,3-dihydroxy-3,5-dihydro-6-methyl-4(H)-pyran-4-one, and 5-(hydroxymethyl)furfural . In addition, a series of short- and long-chain fatty acids was also found . A large number of the volatiles in A . tequilana Weber var . azul were also detected in tequila extracts, and most of these have been reported as a powerful odorants, responsible for the unique tequila flavor. J Agric Food Chem, 2002 Feb 13, 50(4), 790 - 5 Fate of spinosad in litter and soils of a mixed conifer stand in the Acadian forest region of New Brunswick; Thompson DG et al.; Spinosad is a natural insecticide, produced via fermentation culture of the actinomycete Saccharopolyspora spinosa, with potential use against a number of forest pests including spruce budworm (Choristoneura fumiferana {Clem}) . Persistence of spinosad was determined in terrestrial fate experiments conducted within a semimature stand of black spruce (Picea mariana {Mill.}) and balsam fir (Abies balsamea {L}) in the Acadian forest region of New Brunswick, Canada . Results of experiments established under full coniferous canopy and in a canopy opening indicated that spinosad dissipated rapidly following hyperbolic kinetics in both litter and soils and was not susceptible to leaching . Time to 50% dissipation estimates for spinosyn A ranged from 2.0 to 12.4 days depending upon matrix and experimental conditions . Spinosyn D dissipated to levels below quantitation limits (0.02 microg/g of dry mass) within 7 days in all cases . Sporadic low-level detection of the demethylated metabolites suggested that parent compounds were degraded in situ. Analyst, 2002 Jan, 127(1), 66 - 9 Rapid determination of underivatized pyroglutamic acid, glutamic acid, glutamine and other relevant amino acids in fermentation media by LC-MS-MS; Qu J et al.; Determination of amino acids in a complex matrix without derivatization is advantageous, however, difficulties are found in both the detection and the separation of those compounds . In this study, a rapid and reliable LC-MS-MS method for the quantitation of underivatized amino acids in exocellular media was established . Injections were made directly after centrifugation of the samples, without further preparation . The separation of seven underivatized amino acids was achieved on a reversed-phase C18 column with pentadecafluorooctanoic acid as a volatile ion-pair reagent, and the specific detection of most amino acids was achieved by MS-MS of the specific transitions {M + H}+-->{M + H - 46}+ . The calibration curves of all analytes were linear over the range of 1.0-1000 microg ml(-1) and the detection limits ranged from 0.1 to 5 ng ml(-1), with an injection volume of 20 microl . The inter-day and intra-day precisions ranged from 2.6 to 5.7% and 4.8 to 8.2%, respectively; the mean recoveries of the seven analytes were 81-104%, 91-107% and 93-101% respectively at the spiked level of 10, 40 and 200 microg ml(-1) . A large number of fermentation samples were analysed using this method . The technique is simple, rapid, selective and sensitive, and shows potential for the high-throughput quantitation of amino acids from other biological matrices. Water Res, 2002 Feb, 36(3), 774 - 8 Luxury uptake of phosphorus by sediment bacteria; Khoshmanesh A et al.; This note reports the results of experiments aimed at confirming the luxury uptake of phosphorus (P) by sediment bacteria as polyphosphate (Poly-P) . Aerobic suspensions of sediments from two different sites were spiked with 1 mg P/L as orthophosphate and augmented with acetate (a fermentation product) or glucose . The orthophosphate was rapidly taken up over a period of a few hours . When these aerobic uptake experiments were made anaerobic and additional organic carbon added, only the acetate-amended sediment released a significant amount of the added phosphorus . It was hypothesised that during the aerobic stage, and with the addition of acetate, some of the phosphorus was accumulated as Poly-P by sediment microorganisms, which was released during the subsequent anaerobic stage (provided acetate was still present) . Two lines of evidence--transmission electron microscope analysis of sediment bacteria and 31P-NMR analysis of sediment extracts--are presented to support the hypothesis that a portion of the phosphorus taken up during the aerobic experiments was stored as Poly-P. Water Res, 2002 Feb, 36(3), 561 - 76 Interactive effects of the electron acceptor sulphate and o-cresol on the methanogenic degradation of hexanoate; Holmes S et al.; A three-stage continuous culture system was used to segregate the component microbial groups of a methanogenic hexanoate-degrading association enriched from anaerobic refuse . The inhibitory effects of o-cresol concentrations (2-20 mM) on the fermentative, acetogenic, sulphate-reducing and methanogenic bacteria were then assessed in the presence of either 1.4 or 3.5 mM sulphate in the influent medium . The sulphate-reducing bacteria (SRB) in the 1.4 mM sulphate-supplemented systems were the most sensitive to o-cresol, with 29.3 and 56.6% inhibition on supplementation with 4 and 6 mM o-cresol, respectively . With 3.5mM supplementation, inhibition was 4.5 and 19.4%, respectively . Methanogenesis was not inhibited by concentrations < 10 mM o-cresol, and complete inhibition was recorded only at concentrations > or = 10 mM . Both fermentation and acetogenesis were affected by inhibition of the electron sinks . The increase in influent sulphate concentration promoted electron flow to sulphidogenesis, as predicted on thermodynamic criteria, but did not affect the relative sensitivity of the different physiological groups.
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