Fermentations

Appl Microbiol Biotechnol, 2003 Jun, 61(5-6), 385 - 92 Epub 2003 Apr 03.
Industrial production of beta-lactam antibiotics; Elander RP; The industrial production of beta-lactam antibiotics by fermentation over the past 50 years is one of the outstanding examples of biotechnology . Today, the beta-lactam antibiotics, particularly penicillins and cephalosporins, represent the world's major biotechnology products with worldwide dosage form sales of approximately 15 billion US dollars or approximately 65% of the total world market for antibiotics . Over the past five decades, major improvements in the productivity of the producer organisms, Penicillium chrysogenum and Acremonium chrysogenum (syn . Cephalosporium acremonium) and improved fermentation technology have culminated in enhanced productivity and substantial cost reduction . Major fermentation producers are now estimated to record harvest titers of 40-50 g/l for penicillin and 20-25 g/l for cephalosporin C . Recovery yields for penicillin G or penicillin V are now >90% . Chemical and enzymatic hydrolysis process technology for 6-aminopenicillanic acid or 7-aminocephalosporanic acid is also highly efficient (approximately 80-90%) with new enzyme technology leading to major cost reductions over the past decade . Europe remains the dominant manufacturing area for both penicillins and cephalosporins . However, due to ever increasing labor, energy and raw material costs, more bulk manufacturing is moving to the Far East, with China, Korea and India becoming major production countries with dosage form filling becoming more dominant in Puerto Rico and in Ireland.

Curr Med Chem Anti-Canc Agents, 2002 Mar, 2(2), 255 - 66
Discovery of antitumor indolocarbazoles: rebeccamycin, NSC 655649, and fluoroindolocarbazoles; Long BH et al.; A fermentation directed product search for potential anticancer drugs conducted by Bristol-Myers in the 1970s and early 1980s resulted in the identification of a novel indolocarbazole (IC) rebeccamycin (RBM) as a potential drug development candidate . Subsequently, an analog program designed to impart distal site in vivo antitumor activity resulted in the discovery of diethylaminoethyl analog of RBM (DEAE-RBM), which is presently undergoing clinical evaluation as NSC 655649 and BMY-27557 . Strong DNA intercalation is the primary mechanism of action of DEAE-RBM resulting in the potent catalytic inhibition of both topoisomerases I and II . Precursor feeding fermentation experiments with fluorine-substituted tryptophans yielded novel fluoroindolocarbazoles (FICs) . These FICs were identified as targeting topoisomerase (topo) I in a mechanism-based screen and their action on topo I was confirmed by production of topo I-mediated single-strand breaks in DNA at sites essentially identical to those induced by camptothecin . Topo I dependent cytotoxicity was demonstrated for specific FICs using a P388 and camptothecin-resistant P388/CPT45 pair of cell lines, the latter expresses little or no functional topo I . For example, topo I selectivity was greatest with 3,9-difluoro substituted FIC and was least significant and least cytotoxic with 4,8-difluoro substituted FIC . The review focuses on the discovery of the rebeccamycin class of compounds and their structure-activity relationships leading to the development of the clinical candidate BMY-27557 (NSC 655649), as well as the lead identification of the fluoroindolocarbazole class of compounds.

Arch Microbiol, 2003 Apr, 179(4), 227 - 33 Epub 2003 Feb 28.
Use of the mannitol pathway in fructose fermentation of Oenococcus oeni due to limiting redox regeneration capacity of the ethanol pathway; Richter H et al.; The heterolactic bacterium Oenococcus oeni ferments fructose by a mixed heterolactic/mannitol fermentation . For heterolactic fermentation of fructose, the phosphoketolase pathway is used . The excess NAD(P)H from the phosphoketolase pathway is reoxidized by fructose (yielding mannitol) . It is shown here that, under conditions of C-limitation or decreased growth rates, fructose can be fermented by heterolactic fermentation yielding nearly stoichiometric amounts of lactate, ethanol and CO(2) . Quantitative evaluation of NAD(P)H-producing (phosphoketolase pathway) and -reoxidizing (ethanol, mannitol and erythritol pathways) reactions demonstrated that at high growth rates or in batch cultures the ethanol pathway does not have sufficient capacity for NAD(P)H reoxidation, requiring additional use of the mannitol pathway to maintain the growth rate . In addition, insufficient capacities to reoxidize NAD(P)H causes inhibition of growth, whereas increased NAD(P)H reoxidation by electron acceptors such as pyruvate increases the growth rate.

J Biol Chem, 2003 Jul 11, 278(28), 26146 - 58 Epub 2003 Apr 03.
Multiple pathways are co-regulated by the protein kinase Snf1 and the transcription factors Adr1 and Cat8; Young ET et al.; ADR1 and CAT8 encode carbon source-responsive transcriptional regulators that cooperatively control expression of genes involved in ethanol utilization . These transcription factors are active only after the diauxic transition, when glucose is depleted and energy-generating metabolism has shifted to the aerobic oxidation of non-fermentable carbon sources . The Snf1 protein kinase complex is required for activation of their downstream target genes described previously . Using DNA microarrays, we determined the extent to which these three factors collaborate in regulating the expression of the yeast genome after glucose depletion . The expression of 108 genes is significantly decreased in the absence of ADR1 . The importance of ADR1 during the diauxic transition is illustrated by the observation that expression of almost one-half of the 40 most highly glucose-repressed genes is ADR1-dependent . ADR1-dependent genes fall into a variety of functional classes with carbon metabolism containing the largest number of members . Most of the genes in this class are involved in the oxidation of different non-fermentable carbon sources . These microarray data show that ADR1 coordinates the biochemical pathways that generate acetyl-CoA and NADH from non-fermentable substrates . Only a small number of ADR1-dependent genes are also CAT8-dependent . However, nearly one-half of the ADR1-dependent genes are also dependent on the Snf1 protein kinase for derepression . Many more genes are SNF1-dependent than are either ADR1- or CAT8-dependent suggesting that SNF1 plays a broader role in gene expression than either ADR1 or CAT8 . The largest class of SNF1-dependent genes encodes regulatory proteins that could extend SNF1 dependence to additional pathways.

Appl Environ Microbiol, 2003 Apr, 69(4), 2161 - 5
Karyotype rearrangements in a wine yeast strain by rad52-dependent and rad52-independent mechanisms; Carro D et al.; Yeast strains isolated from the wild may undergo karyotype changes during vegetative growth, a characteristic that compromises their utility in genetic improvement projects for industrial purposes . Karyotype instability is a dominant trait, segregating among meiotic derivatives as if it depended upon only a few genetic elements . We show that disrupting the RAD52 gene in a hypervariable strain partially stabilizes its karyotype . Specifically, RAD52 disruption eliminated recombination at telomeric and subtelomeric sequences, had no influence on ribosomal DNA rearrangement rates, and reduced to 30% the rate of changes in chromosomal size . Thus, there are at least three mechanisms related to karyotype instability in wild yeast strains, two of them not requiring RAD52-mediated homologous recombination . When utilized for a standard sparkling-wine second fermentation, Deltarad52 strains retained the enological properties of the parental strain, specifically its vigorous fermentation capability . These data increase our understanding of the mechanisms of karyotype instability in yeast strains isolated from the wild and illustrate the feasibility and limitations of genetic remediation to increase the suitability of natural strains for industrial processes.

Physiol Plant, 2003 Apr, 117(4), 508 - 520
Tolerance of crop plants to oxygen deficiency stress: fermentative activity and photosynthetic capacity of entire seedlings under hypoxia and anoxia; Mustroph A et al.; The study investigates the reactions of rice, wheat and maize to anoxia (plants without access to oxygen) and hypoxia (roots with very limited access to oxygen) . We studied the adaptations of these intact crop plants because they are known to differ widely in their tolerance to oxygen deficiency . In hypoxia, there was an accumulation of sugars, especially in wheat and maize, although both flood-sensitive species significantly increased the activities of fermentative and glycolytic enzymes, clearly more than in rice . In rice, avoiding an oxygen limitation due to the effective aeration system (30% of root cross-sectional area) may have accounted for only a minor metabolic reaction to hypoxia . In anoxia, maize and wheat quickly lost viability and nearly all photosynthetic capacity, while most rice leaves stayed turgid and green, losing only 50% of the photosynthetic capacity . A strong metabolic arrest under anoxia was obvious for the sucrolytic, glycolytic and fermentative enzymes in all tested species, but was most pronounced in rice . Of the 14 enzymes studied, rice showed the lowest activity increase in hypoxia for 11 enzymes, and the strongest activity decrease in anoxia for 8 enzymes . However, rice was able even under anoxia to keep a 1/4 of the ATP level of the aerated control, while it was at the detection limit in maize and wheat . It appears that in anoxic rice, the switch to metabolic dormancy and maintenance of basic shoot meristems diminishes the needs for energy and substrate . Additionally, rice already has lower sugar demand under hypoxia, and sugar supply appears to be sustained under anoxia by a functioning anaerobic amylase and by the photosynthetically active shoot.

Biotechnol Prog, 2003 Mar-Apr, 19(2), 662 - 4
Mixed culture bioconversion of 16-dehydropregnenolone acetate to androsta-1,4-diene-3,17-dione: optimization of parameters; Banerjee T et al.; Bioconversion of 16-dehydropregnenolone acetate (16-DPA) to androsta-1,4-diene-3,17-dione (ADD), an intermediate for the production of female sex hormones, by mixed culture of Pseudomonas diminuta MTCC 3361 and Comamonas acidovorans MTCC 3362 is reported . Various physicochemical parameters for the bioconversion of 16-DPA to ADD have been optimized in shake flask cultures . Nutrient broth inoculated with actively growing co-culture proved ideal for bacterial growth and bioconversion . A temperature range of 35-40 degrees C was most suitable; higher or lower temperatures adversely affected the bioconversion . Dimethylformamide below 2% concentration was the most suitable carrier solvent . Maximum conversion was recorded at 0.5 mg mL(-1) 16-DPA . A pH of 5.0 yielded a peak conversion of 62 mol % in 120 h incubation period . Addition of 9alpha-hydroxylase inhibitors failed to prevent further breakdown of ADD to nonsteroidal products . 16-DPA conversion in a 5 L fermenter followed a similar trend.

Biotechnol Prog, 2003 Mar-Apr, 19(2), 599 - 611
Evaluation of kinetic models for industrial acetic fermentation: proposal of a new model optimized by genetic algorithms; Gonzalez-Saiz JM et al.; The most important kinetic models developed for acetic fermentation were evaluated to study their ability to explain the behavior of the industrial process of acetification . Each model was introduced into a simulation environment capable of replicating the conditions of the industrial plant . In this paper, it is proven that these models are not suitable to predict the evolution of the industrial fermentation by the comparison of the simulation results with an average sequence calculated from the industrial data . Therefore, a new kinetic model for the industrial acetic fermentation was developed . The kinetic parameters of the model were optimized by a specifically designed genetic algorithm . Only the representative sequence of industrial concentrations of acetic acid was required . The main novelty of the algorithm is the four-composed desirability function that works properly as the response to maximize . The new model developed is capable of explaining the behavior of the industrial process . The predictive ability of the model has been compared with that of the other models studied.

Biotechnol Prog, 2003 Mar-Apr, 19(2), 469 - 76
Co-extraction during reactive extraction of phenylalanine using Aliquat 336: interfacial mass transfer; Pursell MR et al.; Reactive liquid-liquid extraction can be used to separate hydrophilic fermentation products that would not otherwise partition into nonpolar solvents . However, during extraction of the target solute other compounds present in the extraction medium will also react with the ion exchange reagent and are thus co-extracted . In this study the effect of co-extraction on the interfacial flux of the target solute phenylalanine has been investigated for reactive extraction using Aliquat 336 . The effect of co-extracting compounds has been included in a new interfacial flux balance, and experimental results reveal that the interfacial concentrations are equal to the final equilibrium conditions of the system . Using this information a simple mass transfer model has been developed from which film mass transfer coefficients may be determined . Co-extraction of other compounds present in the feed was found to reduce the interfacial flux of the target solute by reducing the driving force . Co-extraction did not affect the value of the film mass transfer coefficient, and therefore, co-extraction does not effect the transport properties of the solute to the interface . Extraction from a multicomponent fermentation broth resulted in a reduced flux, which arises from a reduction in the driving force caused by high levels of co-extraction . Furthermore, the flux was also reduced as the result of a mass transfer resistance caused by soluble surface-active compounds present in the fermentation broth adsorbing to the interface . The biomass associated with the fermentation broth was also found to reduce the solute flux, and it is believed that this is due to blockage of the interfacial area.

Biotechnol Prog, 2003 Mar-Apr, 19(2), 428 - 35
Production and characterization of exopolysaccharides from an enthomopathogenic fungus Cordyceps militaris NG3; Kim SW et al.; Optimization of the submerged culture conditions for the mycelial growth and production of exopolysaccharide (EPS) from a newly isolated Cordyceps species (C . militaris NG3) was studied in flask cultures . The optimal temperature and initial pH for EPS production were 30 degrees C and 8.0, respectively . Sucrose (30 g/L) and corn steep powder (10 g/L) were the most suitable carbon and nitrogen source for both mycelial growth and EPS production . There was a distinguishable morphological changes in mycelium grown between organic and inorganic nitrogen sources . A smooth pellet growth with heavy hyphal thickness was observed in the medium containing organic nitrogen sources, whereas irregular pellets with less hairy region were formed in the medium containing inorganic nitrogen sources . More highly branched cells appearing in the medium of organic nitrogen sources seemed a favorable morphological form for both EPS production and mycelial growth . Under optimal culture conditions, the maximum concentrations of mycelial growth and EPS were 17.6 and 3.4 g/L in a 5-L stirred-tank fermenter . Four groups of EPSs (designated as Fr-I, Fr-II, Fr-III, and Fr-IV) were obtained from the culture filtrates by size exclusion chromatography (SEC), and their molecular characteristics were examined by a multiangle laser-light scattering (MALLS) and refractive index (RI) detector system . The weight-average molar masses of the Fr-I, Fr-II, Fr-III, and Fr-IV of EPS were determined to be 2.262 x 10(6), 3.348 x 10(5), 1.049 x 10(5), and 5.059 x 10(4) g/mol, respectively . All four EPSs showed very low polydispersity indices ranging from 1.00 to 1.18 . The SEC/MALLS analysis revealed that the molecular shape of the Fr-I was a rigid sphere suspected to be an aggregate of complex polysaccharides, the Fr-II and Fr-III were nearly globular in shape, and the Fr-IV was an almost rodlike structure.

Biotechnol Prog, 2003 Mar-Apr, 19(2), 405 - 9
Evolution of culture broth rheological properties during propagation of the entomopathogenic nematode Steinernema carpocapsae, in submerged monoxenic culture; Chavarria-Hernandez N et al.; This article presents the evolution of culture broth rheological properties during monoxenic cultures of Steinernema carpocapsae in cylindrical bottles agitated orbitally . Rheological properties were evaluated in simple-shear flow conditions and were well-modeled by the Ostwald-de Waele model . Rheological properties varied from slightly dilatant, n = 1.2 (-), to moderately pseudoplastic flow behavior, n = 0.6 (-) . Nematode concentrations increased from 750 +/- 190 to 130 900 +/- 6900 nematodes/mL, and the apparent viscosity (eta(a)) evolved from 4.5 +/- 0.7 to 46.6 +/- 3.2 mPa.s during the fermentations . Rheological behavior did not appear to be strongly influenced by nematode number and/or its stage of development; however, the release of substances from the decomposition of nematode cadavers appeared to be of great importance . Among the different developmental stages of the nematodes, only juveniles of the first stage (J1) were highly susceptible to the shearing conditions tested (shear stress, tau(r)()(theta), from 0.9 to 3.5 Pa during periods of 80-100 min), resulting in the viability loss of 85% of J1 nematodes.

Biotechnol Prog, 2003 Mar-Apr, 19(2), 320 - 4
Effect of flow rate pattern on glucose-6-phosphate dehydrogenase synthesis in fed-batch culture of recombinant Saccharomyces cerevisiae; Miguel AS et al.; A strain of genetically modified Saccharomyces cerevisiae (S . cerevisiae) W303 181 was used to improve glucose-6-phosphate dehydrogenase (G6PDH) production in aerobic culture . Fed-batch cultures were carried out in a 5 L fermentor at variable values of the parameter K, namely, 0.2, 0.3, 0.5, 0.7, and 0.8 h(-)(1) . The highest G6PDH production (1164 U/L) and specific activity (517 U/g(cell)) were obtained using the following conditions: glucose, 5.0 g/L; adenine, 8 microg/mL; histidine, 8 microg/mL; tryptophan, 8 microg/mL; temperature, 30 degrees C; inoculum, 1.28 g/L; pH, 5.7; agitation, 400 rpm; aeration, 2.2 vvm; and K, 0.2 h(-)(1) . The exponential feeding pattern increased cell density (2.14 g/L), enzyme productivity (149.27), and biomass yield (0.18 g(glu)/g(cell)( )(mass)) . The level of G6PDH in the genetically modified S . cerevisiae was approximately 4.1-fold higher than that found in a commercial strain.

Biotechnol Prog, 2003 Mar-Apr, 19(2), 312 - 9
Production of phytase by Mucor racemosus in solid-state fermentation; Bogar B et al.; Phytase production was studied by three Mucor and eight Rhizopus strains by solid-state fermentation (SSF) on three commonly used natural feed ingredients (canola meal, coconut oil cake, wheat bran) . Mucor racemosus NRRL 1994 (ATCC 46129) gave the highest yield (14.5 IU/g dry matter phytase activity) on coconut oil cake . Optimizing the supplementation of coconut oil cake with glucose, casein and (NH(4))(2)SO(4), phytase production in solid-state fermentation was increased to 26 IU/g dry matter (DM) . Optimization was carried out by Plackett-Burman and central composite experimental designs . Using the optimized medium phytase, alpha-amylase and lipase production of Mucor racemosus NRRL 1994 was compared in solid-state fermentation and in shake flask (SF) fermentation . SSF yielded higher phytase activity than did SF based on mass of initial substrate . Because this particular isolate is a food-grade fungus that has been used for sufu fermentation in China, the whole SSF material (crude enzyme, in situ enzyme) may be used directly in animal feed rations with enhanced cost efficiency.

Biotechnol Prog, 2003 Mar-Apr, 19(2), 285 - 92
Oil and air dispersion in a simulated fermentation broth as a function of mycelial morphology; Lucatero S et al.; The culture conditions of a multiphase fermentation involving morphologically complex mycelia were simulated in order to investigate the influence of mycelial morphology (Trichoderma harzianum) on castor oil and air dispersion . Measurements of oil drops and air bubbles were obtained using an image analysis system coupled to a mixing tank . Complex interactions of the phases involved could be clearly observed . The Sauter diameter and the size distributions of drops and bubbles were affected by the morphological type of biomass (pellets or dispersed mycelia) added to the system . Larger oil drop sizes were obtained with dispersed mycelia than with pellets, as a result of the high apparent viscosity of the broth, which caused a drop in the power drawn, reducing oil drop break-up . Unexpectedly, bubble sizes observed with dispersed mycelia were smaller than with pellets, a phenomenon which can be explained by the segregation occurring at high biomass concentrations with the dispersed mycelia . Very complex oil drops were produced, containing air bubbles and a high number of structures likely consisting of small water droplets . Bubble location was influenced by biomass morphology . The percentage (in volume) of oil-trapped bubbles increased (from 32 to 80%) as dispersed mycelia concentration increased . A practically constant (32%) percentage of oil-trapped bubbles was observed with pelleted morphology at all biomass concentrations . The results evidenced the high complexity of phases interactions and the importance of mycelial morphology in such processes.

Sheng Wu Gong Cheng Xue Bao, 2002 Nov, 18(6), 761 - 6
{Progress in purification of human serum albumin}; Lu XL et al.; Human serum albumin(HSA) has been used clinically to treat a number of diseases with high dosage . Extremely pure puoduct is required in large-scale production . Plasma-derived HSA(pHSA) has long been produced by precipitation methods . Among them cold ethanol precipitation is dominant . However, chromatographic purification of HSA has been increasingly studied in the last few years . Application of chromatography, especially ionexchange, affinity, and size-exclusion, has opened a new area in the production of pHSA . A new challenge is the purification of recombinant HSA(rHSA) . A successful approach involves STREAMLINE expanded bed adsorption to direct capture the target product from the fermentation broth . This novel process eliminates the need to separate the cells by centrifugation or membrane filtration . Ion exchange chromatography and hydrophobic chromatography play a central role in the purification scheme . Integration with other chromatographic techniques such as size-exclusion, metal chelate, and affinity gives improved purification results . Though innovative, the purification of rHSA still needs further improvement and optimization to increase product purity and process recovery.

Sheng Wu Gong Cheng Xue Bao, 2002 Nov, 18(6), 729 - 34
{Study on fermentation condition of alkaline protease gene engineering strain and the purification and characterization of recombinant enzyme}; Tang XM et al.; In a 5L fermentor the production conditions of alkaline protease gene engineering strain BA071 were investigated . The maximum activity of alkaline protease reached 24,480 u/mL in 40 hours of fermentation by combination of enhancing aeration and changing the agitation rate . The fast purification method of recombinant protease was conducted with FPLC (Fast Protein Liquid Chromatography) . The crude enzyme, treated with ammonium sulfate fractionation and decolored with DEAE-A-50 and polyethylene glycol concentration, was purified with CM-Sephadex-C-50 and Sephadex-G-75 . The purified enzyme appears homologous on SDS-PAGE . The purity of enzyme was increased 76.2 times . SDS-PAGE analysis showed that the molecular weights of expressed recombinant products were about 28 kD . The optimal reaction pH and temperature of recombinant enzyme were at pH11 and 60 degrees C, respectively . The recombinant enzyme exhibited high temperature tolerance and was stable at a wide range of pH . Ca2+, MG2+ can enhance the stability of the recombinant enzyme . While the protease activity of the enzyme was strongly inhibited by Hg2+, Ag+, PMFS {symbol: see text} DFP, and was not affected by SDS and Urea.

Sheng Wu Gong Cheng Xue Bao, 2002 Nov, 18(6), 651 - 5
{Progress in biotechnological production of pyruvic acid}; Liu LM et al.; Pyruvate, an important organic acid, is widely used in the industries of pharmaceuticals, chemicals, agrochemicals, food additives and so on . Compared with the chemical method, biotechnological production of pyruvic acid is an alternative approach because of the low cost and high product quality . In this article, biosynthesis of pyruvate, including direct fermentative production and resting cell method as well as enzymatic method, was discussed . Furthermore, a comparison of these different methods was proposed . Since, a multi-vitamin auxotrophic strain of Torulopsis glabrata is the most competitive strain for industrial production of pyruvate, emphasis was therefore placed on the development of strains screening and fermentation optimization . Finally, some suggestions were put forward to improve the research in this field in the near future.

Biotechnol Bioeng, 2003 Jun 20, 82(6), 702 - 9
Methodology for real-time, multianalyte monitoring of fermentations using an in-situ mid-infrared sensor; Kornmann H et al.; An in-situ, mid-infrared sensor was used to monitor the major analyte concentrations involved in the cultivation of Gluconacetobacter xylinus and the production of gluconacetan, a food-grade exopolysaccharide . To predict the analyte concentrations, three different sets of standard spectra were used to develop calibration models, applying partial least-squares regression . It was possible to build a valid calibration model to predict the 700 spectra collected during the complete time course of the cultivation, using only 12 spectra collected every 10 h as standards . This model was used to reprocess the concentration profiles from 0 to 15 g/L of nine different analytes with a mean standard error of validation of 0.23 g/L . However, this calibration model was not suitable for real-time monitoring as it was probably based on non-specific spectral features, which were correlated only with the measured analyte concentrations . Valid calibration models capable of real-time monitoring could be established by supplementing the set of 12 fermentation spectra with 42 standards of measured analytes . A pulse of 5 g/L ethanol showed the robustness of the model to sudden disturbances . The prediction of the models drifted, however, toward the end of the fermentation . The most robust calibration model was finally obtained by the addition of 34 standard spectra of non-measured analytes . Although the spectra did not contain analyte-specific information, it was believed that this addition would increase the variability space of the calibration model . Therefore, an expanded calibration model containing 88 spectra was used to monitor, in real time, the concentration profiles of fructose, acetic acid, ethanol and gluconacetan and allowed standard errors of prediction of 1.11, 0.37, 0.22, and 0.79 g/L, respectively .

Biotechnol Bioeng, 2003 Jun 20, 82(6), 640 - 52
Toward consistent and productive complex media for industrial fermentations: studies on yeast extract for a recombinant yeast fermentation process; Zhang J et al.; Yeast extract (YE) is commonly used as a key component in the complex media for industrial fermentations . However, the lot-to-lot variation of this raw material frequently requires extensive "use testing" of many lots to identify only the few that support desired fermentation performance . Through extensive fermentation studies and chemical analyses, we have identified adenine and two metabolizable carbon sources, trehalose and lactate, as the principle components in YE that affect the production of a recombinant protein antigen by a yeast strain . Adenine is required for culture growth and the relationship between biomass and measured adenine can be expressed by a Michaelis-Menten model, while the slowly metabolized trehalose serves to maintain the energy supply to the continued antigen synthesis . The rapidly utilized lactate exerts an indirect positive effect by sparing some of the accumulated ethanol from being consumed for growth to being utilized in the product formation . The effects of these YE components are mutually dependent . Based on the database generated from 40 lots at laboratory scale, a relatively high level of carbon sources in YE (trehalose plus lactate, >9.5% w/w) and an intermediate level of adenine (0.14-0.24% w/w) appear to be the minimal requirement of a good lot for this recombinant yeast fermentation . Many poor lots were improved in lab fermenters by rational supplementation of trehalose, lactate, or adenine to compensate for their insufficiencies . At the large production scale, predictions based on adenine and trehalose/lactate contents in various YE lots used correlated reasonably well with culture growth and antigen yield, illustrating the feasibility of such a simple chemical/biochemical analysis as a rapid and reliable initial screening tool . Without incurring any compositional change to an established manufacturing medium, this study demonstrates an effective approach to achieve consistency in fermentations employing complex nutrients and to improve fermentation productivities supported by suboptimal lots of raw material .

J Nutr, 2003 Apr, 133(4), 1120 - 6
Ingestion of water-soluble soybean fiber prevents gastrectomy-induced iron malabsorption, anemia and impairment of voluntary running exercise performance in rats; Shiga K et al.; We examined the effects of feeding water-soluble soybean fiber (WSSF), a highly fermentable dietary fiber, on gastrectomy-induced iron malabsorption, anemia and impairment of exercise performance in rats in two separate experiments . The study was designed as a 2 x 2 factorial arrangement with operation (total gastrectomy) and diet (WSSF) under sedentary (Experiment 1) and exercised (Experiment 2) conditions . In Experiment 1, gastrectomy reduced net iron absorption, hemoglobin concentration, hematocrit and hemoglobin regeneration efficiency (P < 0.01) . However, in rats fed a WSSF diet (50 g/kg diet), iron absorption and the hematological variables of the gastrectomized rats were comparable to those in the sham-operated rats, demonstrating that ingestion of WSSF promotes iron absorption and prevents anemia after gastrectomy . Feeding WSSF increased pools of organic acids and soluble iron in cecal contents and decreased the pH of the cecal contents (P < 0.001) . Of the many cecal variables measured, net iron absorption in gastrectomized rats was most closely correlated (r = 0.614, P < 0.01) with the short-chain fatty acid pool in the cecum . Cecal fermentation of WSSF may contribute to improvements in gastrectomy-induced nutritional defects . In Experiment 2, we examined voluntary running exercise performance in totally gastrectomized rats fed diets with or without WSSF . Total gastrectomy severely impaired running performance (P < 0.001), and WSSF feeding largely restored the lowered performance . We conclude that feeding WSSF improves anemia and impaired voluntary running performance in totally gastrectomized rats.

J Nutr, 2003 Apr, 133(4), 1105 - 12
Extent of hypophagia caused by propionate infusion is related to plasma glucose concentration in lactating dairy cows; Oba M et al.; Two experiments were conducted to evaluate how dose-response effects of intraruminal infusion of propionate on feeding behavior and plasma metabolites are altered by diets differing in fermentability . Twelve ruminally cannulated Holstein cows were used in each experiment . Cows were fed diets containing either steam flaked corn or dry cracked corn (30% of dietary dry matter) in expt . 1, and diets differing in forage-to-concentrate ratio (66:34 vs . 36:64) in expt . 2 . For both experiments, the experimental design was a crossover for dietary treatment, and a 6 x 6 Latin square for infusion treatment within a diet for each period . Infusion treatments were mixtures of sodium propionate and sodium acetate, containing propionate at 0, 0.2, 0.4, 0.6, 0.8 and 1.0 as a fraction of total volatile fatty acids infused . Treatment solutions were infused into the rumen continuously for 18 h starting 6 h before feeding at a rate of 23.1 mmol/min . Although propionate production from ruminal fermentation was expected to be different, dietary treatments did not affect dry matter intake (DMI) responses to propionate infusion for either experiment . However, propionate infusion decreased DMI linearly in expt . 1, but did not decrease DMI at lower rates of propionate infusion, which were much more effective at increasing plasma glucose concentration in expt . 2 . Propionate had a smaller hypophagic effect at low concentrations of plasma glucose and had a greater hypophagic effect at elevated concentrations of plasma glucose, which could be explained by changes in the metabolism of propionate in the liver.

Int J Food Microbiol, 2003 May 25, 83(1), 1 - 14
Phenotypic and genotypic characterization of Oenococcus oeni strains isolated from Italian wines; Guerrini S et al.; A phenotypic and genotypic characterization of 84 Oenococcus oeni isolates from Italian wines of different oenological areas was carried out . Numerical analysis of fatty acid profiles grouped the isolates into two clusters at low level of similarity (63%), the minor cluster containing seven isolates besides the type and the reference strains . Forthy-eight O . oeni isolates, representative of the two clusters, showed no differences in their metabolic properties (heterolactic fermentation pattern, citrate degradation capability and formation of some secondary metabolites) . Moreover, the analysis of species-specific randomly amplified polymorphic DNA and 16S-23S rDNA intergenic spacer region polymorphism as well as the sequence-specific separation of V3 region from 16S rDNA by denaturing gradient gel electrophoresis demonstrated a substantial homogeneity among the isolates . On the basis of ApaI Pulsed Field Gel Electrophoresis (PFGE) restriction patterns, the 84 isolates were grouped into five different clusters at 70% similarity, but no correlation with the phenotypic groups could be demonstrated . However, by combining phenotypic and genotypic data, the 84 O . oeni isolates grouped into eight phenotypic-genotypic combined profiles and a relationship between the origin of the isolates and their combined profile became evident, so that a sort of strain specificity can be envisaged for each wine-producing area.

Vet J, 2003 May, 165(3), 221 - 7
Effect of forage type on claw horn lesion development in dairy heifers; Offer JE et al.; In a continuous design study the claw health of 54 Holstein-Friesian heifer calves was recorded from three months of age until six months after first calving (30 months of age) . Pre-calving heifers were either fed a wet, fermented grass silage-based diet (WF) or a dry, unfermented straw and concentrate based diet (DU), apart from grazing during their first summer . Approximately one month before calving both groups were fed a silage-based diet and afterwards all received a silage and concentrate lactation ration . Claws were examined four times during rearing, once pre-calving, and four times during lactation . Both white line and sole lesions were significantly worse for WF than DU both during rearing and throughout first lactation although the effect was not as consistent over time for white line lesions . It is concluded that for optimal claw health youngstock diets should not be heavily based on wet grass silage (less than 25% DM).

Gastroenterology, 2003 Apr, 124(4), 894 - 902
Colonic fermentation influences lower esophageal sphincter function in gastroesophageal reflux disease; Piche T et al.; BACKGROUND & AIMS: Colonic fermentation of carbohydrates is known to influence gastric and esophageal motility in healthy subjects . This study investigated the effects of colonic fermentation induced by oral administration of fructooligosaccharides (FOS) in patients with gastroesophageal reflux disease (GERD) . METHODS: In the cross-over design used in the study, 9 patients with symptomatic GERD were administered a low-residue diet (i.e., 10 g fiber/day) during 2, 7-day periods, receiving either 6.6 g of FOS or placebo 3 times daily after meals . Each period was separated by a wash out of at least 3 weeks . On day 7, esophageal motility and pH were recorded in fasting conditions and after a test meal containing 6.6 g of FOS or placebo . Breath hydrogen concentrations (reflecting colonic fermentation) and plasma concentrations of glucagon-like peptide 1 (GLP-1), peptide YY, and cholecystokinin were monitored . RESULTS: Compared with placebo, FOS led to a significant increase in the number of transient lower esophageal sphincter relaxations (TLESRs) and reflux episodes, esophageal acid exposure, and the symptom score for GERD . The integrated plasma response of GLP-1 was significantly higher after FOS than placebo . CONCLUSIONS: Colonic fermentation of indigestible carbohydrates increases the rate of TLESRs, the number of acid reflux episodes, and the symptoms of GERD . Although different mechanisms are likely to be involved, excess release of GLP-1 may account, at least in part, for these effects.

Exp Biol Med (Maywood), 2003 Apr, 228(4), 365 - 9
Influence of pH treatments on survival of Escherichia coli O157:H7 in continuous cultures of rumen contents; Thran BH et al.; The pH (i.e., 5.5, 5.75, 6.0, 6.25, 6.5, 6.75, 7.0, and 7.25) effect on Escherichia coli O157:H7 in an artificial rumen model was investigated . Eight fermenters were inoculated with bovine rumen fluid and were supplied with a diet (75 g of dry matter daily in 12 equal portions {every 2 hr}) containing similar forage-to-concentrate ratio . After an adaptation period (i.e., 3 days for adjusting the rumen fluid {pH 6.2} microbial population to the test pH and 4 days for adjustment to the diet at the test pH), each fermenter was inoculated with 10(9) cells of E . coli O157:H7 . Samples were collected hourly for 12 hr and every 2 hr for an additional 12 hr and were analyzed by flow cytometer . E . coli O157:H7 could not be quantified after 24 hr, and detection was only possible after enrichment . Because the pathogen could not be detected 5 days postinoculation (i.e., Day 13), the fermenters were reinoculated with E . coli O157:H7 on Days 17 and 22 . E . coli O157:H7 numbers decreased from 10(6) to 10(4)/ml of fermenter contents in a quadratic (P < 0.05) fashion over the 24-hr sampling period, and the rate of reduction was slower (P < 0.05) for pH 7.0 than for other pH treatments . Results suggested that E . coli O157:H7 population were decreased by competitive exclusion and were not affected by culture pH.

J Agric Food Chem, 2003 Apr 9, 51(8), 2402 - 8
Evolution of volatile byproducts during wine fermentations using immobilized cells on grape skins; Mallouchos A et al.; A biocatalyst was prepared by immobilization of Saccharomyces cerevisiae cells on grape skins . Repeated batch fermentations were conducted using this biocatalyst as well as free cells, at 25, 20, 15, and 10 degrees C . Solid phase microextraction (SPME) was used in monitoring the evolution of volatile byproducts . The effect of immobilization and temperature on evolution patterns of volatiles was obvious . The major part of esters was formed after consumption of 40-50% of the sugars . Similar processes were observed for amyl alcohols and 2-phenylethanol, whereas 1-propanol and 2-methyl-1-propanol were formed during the whole alcoholic fermentation period at an almost constant formation rate . Acetaldehyde and acetoin were synthesized in the early stages of fermentation . Afterward, their amount decreased . In most cases, immobilized cells exhibited higher formation rates of volatiles than free cells . The final concentration of esters was higher in wines produced by immobilized biocatalyst . Their amount increased with temperature decrease . The opposite was observed for higher alcohols.

J Agric Food Chem, 2003 Apr 9, 51(8), 2300 - 3
Glyoxal/glycolaldehyde: a redox system involved in malolactic fermentation of wine; Flamini R et al.; To verify the presence of glycolaldehyde in wine resulting from reduction of glyoxal after malolactic fermentation, sterilized solutions of synthetic cultures were inoculated with a lactic bacterium of the type Oenococcus oeni . Fermentation was also carried out on solutions with glyoxal added . The resulting glycolaldehyde concentrations turned out to be associated with the amounts of glyoxal present, and glyoxal was seen to decrease as glycolaldehyde increased . In addition, it was observed that glyoxal principally forms from breakdown of sugars and that reduction to glycolaldehyde is mainly promoted by bacterial activity . Finally, the ability of glycolaldehyde to induce browning of (+)-catechin in a model wine system was verified and turned out to be about 10 times higher than that of ascorbic acid.

J Agric Food Chem, 2003 Apr 9, 51(8), 2242 - 5
Lignin degradation by Agaricus bisporus accounts for a 30% increase in bioavailable holocellulose during cultivation on compost; ten Have R et al.; The common mushroom Agaricus bisporus is a non-white rot saphrophytic fungus that can degrade lignin to free and utilize holocellulose embedded in fermented straw as present in compost . A new method is described to estimate the actual amount of bioavailable holocellulose in 3.8 kg compost cultures spawned with A . bisporus Horst U1 prior to and during a cultivation with two cycles of mushroom harvesting . The method shows that the initial amount of bioavailable holocellulose per culture, accounting for 130 +/- 22 g, is lower than the total holocellulose consumption by A . bisporus accounting for 182 +/- 15 g . This difference is explained by a 30% increase in bioavailable holocellulose . The increase is caused by the degradation of 95 +/- 3 g of holocellulose-shielding lignin . The results are discussed within the scope of the A . bisporus mushroom yield and lignin degradation by white rot fungi during growth on lignocellulose-containing materials.

J Antibiot (Tokyo), 2003 Jan, 56(1), 9 - 15
Isolation and characterization of two new antifungal antibiotics from a basidiomycete; Chu M et al.; Two novel antibiotics, Sch 484129 (1) and Sch 484130 (2), were isolated from the fermentation broth of a fungal culture, which was identified as a Basidiomycete . The new antibiotics were obtained by ethyl acetate extraction followed by reversed phase HPLC purification . Structure elucidation of 1 and 2 was accomplished by spectroscopic data analyses . Derivatizations of the major component 1 were performed in order to provide definitive structural information . Both components were identified as glycolipids and displayed antifungal activity against Saccharomyces and Aspergillus strains.

J Antibiot (Tokyo), 2003 Jan, 56(1), 1 - 5
Byssochlamysol, a new antitumor steroid against IGF-1-dependent cells from Byssochlamys nivea . I . Taxonomy, fermentation, isolation and biological activity; Mori T et al.; A new antitumor steroid, byssochlamysol, was isolated from the mycelium of Byssochlamys nivea M#5187 . Byssochlamysol inhibited IGF-1-dependent growth of MCF-7 human breast cancer cells with an IC50 of 20 ng/ml, whereas serum-dependent cell growth was not inhibited by less than 10 microg/ml of byssochlamysol . This substance induced apoptosis in IGF-1-dependent Colo320DM human colon cancer cells.

Pharm Res, 2003 Mar, 20(3), 429 - 34
Effect of colonic lactulose availability on the timing of drug release onset in vivo from a unique colon-specific drug delivery system (CODES); Yang L et al.; PURPOSE: To test the hypothesis that the onset of drug release in vivo from a unique colon-specific drug delivery system (CODES) would depend on the colonic availability rate of lactulose . The site specificity of drug release in canine GI tract was also estimated . METHODS: CODES tablets were prepared by tableting the granulation or acetaminophen and lactulose, followed with film coating . The pharmacokinetic performance of different CODES formulations was evaluated in six beagle dogs under fasted conditions . The release of acetaminophen and lactulose was also characterized in vitro . RESULTS: The onset of acetaminophen release in beagle dogs was found to be dependent on the coating level of Eudragit E and lactulose loading in the core tablet . At Eudragit E coating levels of 4%, 8% and 12% (coating weight gain), the onset of in vivo drug release occurred 5.5 (+/- 1.9) h . 4.8 (+/-1.0) h . and 7.5 (+/-1.0) h, respectively, after dosing . A similar trend was observed when the loading of lactulose in the core tablet decreased from 78% to 58% and 38% . However, the rate and extent of acetaminophen absorption did not vary significantly in each situation based on the values of AUC and Cmax . CONCLUSION: The onset of drug release in vivo from CODES tablets is predominantly dependent on colonic availability rate of lactulose because drug release from this system is triggered by localized drop of colonic pH from the fermentation of lactulose.

Food Nutr Bull, 2003 Mar, 24(1), 104 - 25
Household-level technologies to improve the availability and preparation of adequate and safe complementary foods; Mensah P et al.; Plant-based complementary foods are the main source of nutrients for many young children in developing countries . They may, however, present problems in providing nutritionally adequate and safe diets for older infants and young children . The high starch content leads to low-nutrient diets that are bulky and dense, with high levels of antinutritive factors such as phytates, tannins, lectins, and enzyme inhibitors . Phytates impair mineral bioavailability, lectins interfere with intestinal structure, and enzyme inhibitors inhibit digestive enzymes . In addition, there is often microbial contamination, which leads to diarrhea, growth-faltering, and impaired development, and the presence of chemical contaminants may lead to neurological disease and goiter . The fact that some fruits containing carotenoids are only available seasonally contributes to the vulnerability of children receiving predominantly plant-based diets . Traditional household food technologies have been used for centuries to improve the quality and safety of complementary foods . These include dehulling, peeling, soaking, germination, fermentation, and drying . While modern communities tend to reject these technologies in favor of more convenient fast-food preparations, there is now a resurgence of interest in older technologies as a possible means of improving the quality and safety of complementary foods when the basic diet cannot be changed for economic reasons . This paper describes the biology, safety, practicability, and acceptability of these traditional processes at the household or community level, as well as the gaps in research, so that more effective policies and programs can be implemented to improve the quality and safety of complementary foods.

Appl Microbiol Biotechnol, 2003 Feb, 60(6), 700 - 7 Epub 2002 Dec 18.
Purification, molecular characterization and reactivity with aromatic compounds of a laccase from basidiomycete Trametes sp . strain AH28-2; Xiao YZ et al.; A recently isolated basidiomycete, Trametes sp . strain AH28-2, can be induced to produce a high level of laccases when grown on a cellobiose-asparagine liquid medium . After induction by kraft lignin, two major isozymes were detected in the fermentation supernatant of the fungus . The principal component laccase A, which accounts for about 85% of the total activity, can be purified to electrophoretic homogeneity by three chromatographic steps: DEAE-Sepharose FF, Superdex-200 and Mono-Q . The solution containing purified laccase is blue in color, and the ratio of absorbance at 280 nm to that at 600 nm is 22 . The molecular mass of laccase A is estimated to be 62 kDa by SDS-PAGE, 57 kDa by FPLC, and measured as 58522 Da by MALDI mass spectrum . Laccase A is a monomeric glycoprotein with a carbohydrate content of 11-12% and an isoelectric point of 4.2 . The optimum pH and temperature for oxidizing guaiacol are 4.5 and 50 degrees C, respectively . The half-life of the enzyme at 75 degrees C is 27 min . The enzyme shows a good stability from pH 4.2 to pH 8.0 . The K(m) values of the enzyme toward substrates 2,2'-azino-bis (3-ethylbenzothazoline-6-sulfonate) (ABTS), guaiacol and 2,6-dimethoxyphenol are 25, 420 and 25.5 microM, respectively, and the corresponding V(max) values are 670, 66.8, and 79 microM min(-1) x mg(-1), respectively . Laccase A activity is strongly inhibited by 0.1 mM NaN(3) or 0.1 mM cyanide . Two units of laccase A alone is able to completely oxidize 100 micromol 2,6-chlorophenol in 6 h . In the presence of 1 mM ABTS and 1-hydroxybenzotriazole, 15.0 U laccase A is able to oxidize 45% and 70% of 50 micromol fluorene in 12 and 18 h, respectively . The laccase A gene was cloned by a PCR method, and preliminary analysis of its sequence indicates 87.0% similarity to the corresponding segment in the phenoloxidase gene from Coriolus hirsutus.

Appl Microbiol Biotechnol, 2003 Feb, 60(6), 643 - 53 Epub 2002 Dec 18.
New developments in oxidative fermentation; Adachi O et al.; Oxidative fermentations have been well established for a long time, especially in vinegar and in L-sorbose production . Recently, information on the enzyme systems involved in these oxidative fermentations has accumulated and new developments are possible based on these findings . We have recently isolated several thermotolerant acetic acid bacteria, which also seem to be useful for new developments in oxidative fermentation . Two different types of membrane-bound enzymes, quinoproteins and flavoproteins, are involved in oxidative fermentation, and sometimes work with the same substrate but produce different oxidation products . Recently, there have been new developments in two different oxidative fermentations, D-gluconate and D-sorbitol oxidations . Flavoproteins, D-gluconate dehydrogenase, and D-sorbitol dehydrogenase were isolated almost 2 decades ago, while the enzyme involved in the same oxidation reaction for D-gluconate and D-sorbitol has been recently isolated and shown to be a quinoprotein . Thus, these flavoproteins and a quinoprotein have been re-assessed for the oxidation reaction . Flavoprotein D-gluconate dehydrogenase and D-sorbitol dehydrogenase were shown to produce 2-keto- D-gluconate and D-fructose, respectively, whereas the quinoprotein was shown to produce 5-keto- D-gluconate and L-sorbose from D-gluconate and D-sorbitol, respectively . In addition to the quinoproteins described above, a new quinoprotein for quinate oxidation has been recently isolated from Gluconobacter strains . The quinate dehydrogenase is also a membrane-bound quinoprotein that produces 3-dehydroquinate . This enzyme can be useful for the production of shikimate, which is a convenient salvage synthesis system for many antibiotics, herbicides, and aromatic amino acids synthesis . In order to reduce energy costs of oxidative fermentation in industry, several thermotolerant acetic acid bacteria that can grow up to 40 degrees C have been isolated . Of such isolated strains, some thermotolerant Acetobacter species were found to be useful for vinegar fermentation at a high temperature such 38-40 degrees C, where mesophilic strains showed no growth . They oxidized higher concentrations of ethanol up to 9% without any appreciable lag time, while alcohol oxidation with mesophilic strains was delayed or became almost impossible under such conditions . Several useful Gluconobacter species of thermotolerant acetic acid bacteria are also found, especially L-erythrulose-producing strains and cyclic alcohol-oxidizing strains . Gluconobacter frateurii CHM 43 is able to rapidly oxidize meso-erythritol at 37 degrees C leading to the accumulation of L-erythrulose, which may replace dihydroxyacetone in cosmetics . G . frateuriiCHM 9 is able to oxidize cyclic alcohols to their corresponding cyclic ketones or aliphatic ketones, which are known to be useful for preparing many different physiologically active compounds such as oxidized steroids or oxidized bicyclic ketones . The enzymes involved in these meso-erythritol and cyclic alcohol oxidations have been purified and shown to be a similar type of membrane-bound quinoproteins, consisting of a high molecular weight single peptide . This is completely different from another quinoprotein, alcohol dehydrogenase of acetic acid bacteria, which consists of three subunits including hemoproteins.

Braz J Biol, 2002 Nov, 62(4A), 633 - 40 Epub 2003 Feb 11.
Biochemical and hematological responses of the banded knife fish Gymnotus carapo (Linnaeus, 1758) exposed to environmental hypoxia; Moraes G et al.; Oxygen of tropical freshwater environments fluctuates drastically . Eutrophic lakes and ponds of warm waters frequently reach very low oxygen concentrations . This is the most common habitat of the banded knife fish "tuvira" Gymnotus carapo . This electric fish is reported to present bimodal breathing to cope with low environmental oxygen . Biochemical responses can be also observed in fishes facing hypoxia but none were studied in tuvira . In the present study, haematological and metabolic changes were investigated in two groups of fish exposed to hypoxia for 1 and 3 hours . Haematocrit, red blood cells and haemoglobin concentration indicated erythrocyte release from hematopoietic organs and swelling of red blood cells . Glycogen, glucose, lactate, pyruvate, and amino acids were quantified in liver, kidney and white muscle . The metabolic profile of G . carapo to cope with hypoxia suggested liver gluconeogenesis probably supported by proteolysis . The kidney and liver presented the same biochemical trend suggesting similar metabolic role for both organs . Glucogenolysis followed by glucose fermentation and protein mobilisation was observed in the white muscle . The air breathing behaviour of tuvira works in parallel with metabolism to prevent damages from hypoxia . Metabolic adjustments are observed when the air taking is avoided.

Appl Microbiol Biotechnol, 2003 Mar, 61(1), 69 - 76 Epub 2003 Jan 24.
Towards a cost effective strategy for cutinase production by a recombinant Saccharomyces cerevisiae: strain physiological aspects; Ferreira BS et al.; Although the physiology and metabolism of the growth of yeast strains has been extensively studied, many questions remain unanswered where the induced production of a recombinant protein is concerned . This work addresses the production of a Fusarium solani pisi cutinase by a recombinant Saccharomyces cerevisiae strain induced through the use of a galactose promoter . The strain is able to metabolise the inducer, galactose, which is a much more expensive carbon source than glucose . Both the transport of galactose into the cell-required for the induction of cutinase production-and galactose metabolism are highly repressed by glucose . Different fermentation strategies were tested and the culture behaviour was interpreted in view of the strain metabolism and physiology . A fed-batch fermentation with a mixed feed of glucose and galactose was carried out, during which simultaneous consumption of both hexoses was achieved, as long as the glucose concentration in the medium did not exceed 0.20 g/l . The costs, in terms of hexoses, incurred with this fermentation strategy were reduced to 23% of those resulting from a fermentation carried out using a more conventional strategy, namely a fed-batch fermentation with a feed of galactose.

Biotechnol Appl Biochem, 2003 Jun, 37(Pt 3), 245 - 9
Large-scale capture and partial purification of plasmid DNA using anion-exchange membrane capsules; Zhang S et al.; The demand for larger quantities of high-purity plasmids continues to increase . Substantial quantities of plasmid DNA are needed to support viral and plasmid-based gene-therapy programmes, including drug delivery, preclinical and clinical studies and production of DNA vaccines . Reliance on fermentation, which generates large lysate volumes, for producing the needed quantities of plasmid DNA is becoming more widespread . Development of an efficient capture-unit operation for very large plasmid DNA molecules from these large lysate volumes can present a considerable challenge . Use of conventional chromatographic beaded media for plasmid capture is problematic, owing to their restrictive pores, which limits access of the large DNA molecules to only those binding sites on the beads' outer surface . Anion-exchange membranes, on the other hand, have large convective pores that allow the plasmid DNA to readily access all of the membrane's anionic binding sites, even at very high flow rates . The convenience of anion-exchange membranes, configured in ready-to-use capsules, can greatly simplify large-scale plasmid purification strategies . The effectiveness of membrane-based technology for the capture of a pCAT reporter plasmid from large volumes of clarified Escherichia coli lysate was demonstrated . The captured and eluted plasmid pool was found to have been concentrated 10-fold with a reduction in endotoxin of four orders of magnitude.

Int J Syst Evol Microbiol, 2003 Jan, 53(Pt 1), 125 - 31
Pseudoalteromonas agarivorans sp . nov., a novel marine agarolytic bacterium; Romanenko LA et al.; The phenotypic, genomic and phylogenetic characteristics of four aerobic, Gram-negative, non-fermentative, motile, non-pigmented, agarolytic Pseudoalteromonas-like bacteria, isolated from marine environments, have been investigated . These bacteria share DNA-DNA similarities above 86% . Comparative 16S rDNA sequence analysis of strain KMM 255T revealed its membership of the genus Pseudoalteromonas; it shares 99.9% sequence similarity with Pseudoalteromonas distincta, Pseudoalteromonas elyakovii, Pseudoalteromonas atlantica and Pseudoalteromonas espejiana . DNA-DNA reassociation levels obtained for strain KMM 255T and type strains of these four species and other Pseudoalteromonas species were below 45% . The marine isolates differed from known species of the genus by the fact that the cells are motile by means of a single flagellum or two to four polar unsheathed flagella and by an inability to utilize most organic compounds . On the basis of phenotypic, DNA-DNA hybridization and phylogenetic data, it is concluded that the isolates represent a novel species within the genus Pseudoalteromonas, for which the name Pseudoalteromonas agarivorans sp . nov . is proposed . The type strain is strain KMM 255T (= DSM 14585T).

Appl Microbiol Biotechnol, 2003 Apr, 61(2), 163 - 78 Epub 2003 Jan 09.
Global metabolic regulation analysis for Escherichia coli K12 based on protein expression by 2-dimensional electrophoresis and enzyme activity measurement; Peng L et al.; Regulation of the main metabolic pathways of Escherichia coli K12 was investigated based on 2-dimensional electrophoresis (2DE) and the measurement of enzyme activities . The cells were grown aerobically in different carbon sources, such as glucose, acetate, gluconate or glycerol . Microaerobic cultivation was also conducted with glucose as a carbon source . Fifty-two proteins could be identified based on 2DE, and 26 enzyme activities from the main metabolic pathways-including glycolysis, pentose phosphate pathway, TCA cycle, Entner-Doudoroff pathway and fermentative pathway-were assayed . These enzyme activities, together with global and quantitative protein expression, gave us a clear picture of metabolic regulation . The results show that, compared with the control experiment with glucose as a carbon source under aerobic conditions, glycolytic enzymes were slightly up-regulated (<2-fold), TCA cycle enzymes were significantly down-regulated (2- to 10-fold), and fermentative enzymes such as pfl and adhE were highly up-regulated (>10-fold) under microaerobic conditions in glucose medium . When acetate was used as a carbon source, pfkA, pykF, ppc and zwf were down-regulated, while fbp, pckA, ppsA and mez were significantly up-regulated . Glyoxylate enzymes such as aceA and aceB were strongly up-regulated (>10-fold) and TCA-cycle-related enzymes were also up-regulated to some extent . With gluconate as a carbon source, edd, eda, fbp and TCA cycle enzymes were up-regulated . With glycerol as a carbon source, fbp and TCA cycle enzymes were up-regulated, while ackA was significantly down-regulated . Protein abundance obtained by 2DE correlated well with enzyme activity, with a few exceptions (e.g., isocitrate dehydrogenase), during aerobic growth on acetate.

Appl Microbiol Biotechnol, 2003 Apr, 61(2), 110 - 7 Epub 2003 Jan 29.
Continuous gluconic acid production by isolated yeast-like mould strains of Aureobasidium pullulans; Anastassiadis S et al.; By extensive microbial screening, about 50 strains with the ability to secrete gluconic acid were isolated from wild flowers . The strains belong to the yeast-like mould Aureobasidium pullulans (de Bary) Arnaud . In shake flask experiments, gluconic acid concentrations between 23 and 140 g/l were produced within 2 days using a mineral medium . In batch experiments, various important fermentation parameters influencing gluconic acid production by A . pullulans isolate 70 (DSM 7085) were identified . Continuous production of gluconic acid with free-growing cells of the isolated yeast-like microorganisms was studied . About 260 g/l gluconic acid at total glucose conversion could be achieved using continuous stirred tank reactors in defined media with residence times (RT) of about 26 h . The highest space-time-yield of 19.3 g l(-1) x h(-1)) with a gluconic acid concentration of 207.5 g/l was achieved with a RT of 10.8 h . The possibility of gluconic acid production with biomass retention by immobilised cells on porous sinter glass is discussed . The new continuous gluconate fermentation process provides significant advantages over traditional discontinuous operation employing Aspergillus niger . The aim of this work was the development of a continuous fermentation process for the production of gluconic acid . Process control becomes easier, offering constant product quality and quantity.

Br J Nutr, 2003 Apr, 89(4), 445 - 53
Supplementation of barley straw with Sesbania pachycarpa leaves in vitro: effects on fermentation variables and rumen microbial population structure quantified by ribosomal RNA-targeted probes; Muetzel S et al.; Tropical livestock is often maintained on roughage-based diets deficient in N, and therefore requires supplementation with protein-rich substrates to achieve reasonable production levels . The optimum inclusion rate of a potential supplement is usually determined by in vivo feeding trials or by in vitro incubation of the diet components to estimate the feed value of the complete diet . The present work simulates a supplementation experiment in vitro, by incubating a pure roughage (barley straw), a pure supplement (Sesbania pachycarpa leaves) and mixtures of the two, with increasing inclusion levels of the supplement, in a short-term batch incubation system . Fermentation kinetics were followed by the release of fermentation endproducts (gas and short-chain fatty acids) . Microbial biomass was estimated using ribosomal (r) RNA as internal marker for bacteria and eukaryotes separately . Cell-wall-degrading subpopulations were quantified by hybridisation with taxon-specific oligonucleotide probes targeting Chytridiomycetes, Fibrobacter spp., Ruminococcus albus and R . flavefaciens . Carboxymethylcellulase (CMCase) was assayed as an indicator for cell-wall-degrading activity . The addition of S . pachycarpa leaves stimulated fermentation in all cases . Gas production, and especially rRNA concentration, showed clear maxima at 40 % S . pachycarpa inclusion, rates that significantly exceeded the values interpolated from the incubations of the pure substrates . Short-chain fatty acid yield changed only slightly, but in the same way . The analysis of the microbial population structure showed that the positive effects were mainly mediated through enhanced growth of Ruminococcus spp . Increasing proportions of S . pachycarpa leaves in the diet led to a drastic decline in the total eukaryotic population . This points to a defaunation, which may also have added to the positive effects . The eukaryotic subpopulation of the rumen fungi were affected to a lesser degree . Although the cell-wall-degrading organisms showed positive responses to the supplementation, the CMCase activity was not affected significantly by the supplementation . The present work shows that it is possible to predict optimum inclusion levels for a new feed supplement in vitro and thus reduce in vivo experiments . It was also demonstrated that true supplementation effects occur particularly for the microbial biomass production, which is the primary source of amino acids for the ruminant animal . The analysis of microbial population structure in context with conventional metabolic measurements adds valuable information to interpret the observed effects on production-related variables.

Oral Microbiol Immunol, 2003 Apr, 18(2), 127 - 30
Amended biochemical characteristics and phylogenetic position of Treponema medium; Nakazawa F et al.; Umemoto et al . (1997, Int J Syst Bacteriol 47, pp . 67-72) proposed spirochete strain G7201, isolated from the periodontal pocket of an adult patient, as a new species, Treponema medium . They deposited this strain in the American Type Culture Collection (ATCC) as type strain ATCC 700293T . Recently, ATCC suggested that there is a discrepancy between the previous report and the results obtained by ATCC in biochemical tests on T . medium ATCC 700293T . In this study, we re-examined and verified the biochemical characteristics of T . medium . The fermentation pattern of carbohydrates of T . medium resembled that of Treponema vincentii and Treponema denticola, but T . medium was clearly differentiated from T . vincentii in the production of indole, and from T . denticola in the hydrolysis of esculin . Also, sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE) protein profile analysis and phylogenetic comparison of 16S rDNA sequences revealed that T . medium is clearly differentiated from any established treponemal species, which supports the validity of the proposal of Treponema medium as a new species.

Oral Microbiol Immunol, 2003 Apr, 18(2), 109 - 13
Acid-neutralizing activity during amino acid fermentation by Porphyromonas gingivalis, Prevotella intermedia and Fusobacterium nucleatum; Takahashi N; Acid-neutralizing activity during amino acid fermentation by washed cells of Porphyromonas gingivalis, Prevotella intermedia and Fusobacterium nucleatum was studied . When the washed cells of these strains were anaerobically incubated in the presence of aspartylaspartic acid or glutamylglutamic acid for P . gingivalis, aspartic acid for P . intermedia and glutamic acid for F . nucleatum at an initial pH of 5.0 or 5.5, the pH of the incubation mixtures rose toward neutral . F . nucleatum had the highest acid-neutralizing activity, followed by P . intermedia and P . gingivalis . The P . intermedia and F . nucleatum cells were used to measure the amounts of base produced at a fixed pH of 5.0 . These cells generated significant amounts of base at pH 5.0 along with the production of organic acids and ammonia from aspartic or glutamic acid . Acid-base balance theoretically calculated from the amounts of consumed substrate and end products implies that the acid-neutralizing activity was derived from the decrease in acidity during the fermentation of amino acid into organic acids and ammonia.

Free Radic Res, 2003 Jan, 37(1), 91 - 7
The antioxidant cocktail, effective microorganism X (EM-X), protects retinal neurons in rats against N-methyl-D-aspartate excitotoxicity in vivo; Aruoma OI et al.; Injection of the glutamate agonist N-methyl-D-aspartate (NMDA) into the vitreous body of rats resulted in severe degeneration of neurons in the retina, with a loss of 81% of ganglion cells and 43% of non-ganglion cells . The cocktail EM-X is a novel antioxidant drink derived from ferment of unpolished rice, papaya and sea-weeds with effective microorganisms (EM-X) . In animals treated with an intraperitoneal injection of EM-X, the loss of ganglion cells was reduced to 55% and that of non-ganglion cells to 34% when compared to untreated NMDA-injected retinas . Cell degeneration resulting from NMDA excitotoxicity, is thought to be mediated via oxidative stress mechanisms . The neuroprotective effect of the EM-X in this system is therefore likely to be due, at least in part, to its flavonoids, saponins, vitamin E and ascorbic content.

Electrophoresis, 2003 Jan, 24(1-2), 303 - 10
Monitoring of protein profiles for the optimization of recombinant fermentation processes using public domain databases; Durrschmid K et al.; The expression of human superoxide dismutase in fed-batch fermentation of E . coli HMS174(DE3)(pET3ahSOD) was studied as model system . Due to the frequently used strong T7 promoter system a high metabolic load is exerted, which triggers stress response mechanisms and finally leads to the differentiation of the host cell . As a consequence, host cell metabolism is partly shifted from growth to survival accompanied by significant alterations of the protein pattern . In terms of process optimization two-dimensional electrophoresis deserves as a powerful tool to monitor these changes on protein level . For the analysis of samples derived from different states of recombinant protein production wide-range Immobiline Dry Strips pH 3-10 were used . In order to establish an efficient procedure for accelerated process optimization and to avoid costly and time-consuming analysis like mass spectrometry (MS), a database approach for the identification of significant changes of the protein pattern was evaluated . On average, 935 spots per gel were detected, whereby 50 are presumably stress-relevant . Out of these, 24 proteins could be identified by using the SWISS-2DPAGE database . The identified proteins are involved in regulatory networks, energy metabolism, purine and pyrimidine nucleotide synthesis and translation . By this database approach, significant fluctuations of individual proteins in relation to recombinant protein production could be identified . Seven proteins show strong alterations (>100%) directly after induction and can therefore be stated as reliable marker proteins for the assessment of stress response . For distinctive interpretation of this highly specific information, a bioinformatic and statistic tool would be essential in order to perceive the role and contribution of individual proteins in stress response.

Biotechnol Bioeng, 2003 Jun 5, 82(5), 552 - 63
Heat and water transfer in a rotating drum containing solid substrate particles; Schutyser MA et al.; In previous work we reported on the simulation of mixing behavior of a slowly rotating drum for solid-state fermentation (SSF) using a discrete particle model . In this investigation the discrete particle model is extended with heat and moisture transfer . Heat transfer is implemented in the model via interparticle contacts and the interparticle heat transfer coefficient is determined experimentally . The model is shown to accurately predict heat transfer and resulting temperature gradients in a mixed wheat grain bed . In addition to heat transfer, the addition and subsequent distribution of water in the substrate bed is also studied . The water is added to the bed via spray nozzles to overcome desiccation of the bed during evaporative cooling . The development of moisture profiles in the bed during spraying and mixing are studied experimentally with a water-soluble fluorescent tracer . Two processes that affect the water distribution are considered in the model: the intraparticle absorption process, and the interparticle transfer of free water . It is found that optimum distribution can be achieved when the free water present at the surface of the grains is quickly distributed in the bed, for example, by fast mixing . Alternatively, a short spraying period, followed by a period of mixing without water addition, can be applied . The discrete particle model developed is used successfully to examine the influence of process operation on the moisture distribution (e.g., fill level and rotation rate) . It is concluded that the extended discrete particle model can be used as a powerful predictive tool to derive operating strategies and criteria for design and scale-up for mixed SSF and other processes with granular media .

Biotechnol Bioeng, 2003 Jun 5, 82(5), 543 - 51
Oil/water and pre-emulsified oil/water (PIT) dispersions in a stirred vessel: Implications for fermentations; Erler ST et al.; This study examines dispersions of rapeseed oil (RSO) in water by mechanical agitation under conditions mimicking those found in certain antibiotic fermentations; for example, in the presence of air, antifoam, and finely divided CaCO(3) particles . A problem with residual oil has been reported for such fermentations, and it has been suggested that the use of pre-emulsified oil can reduce this problem . Hence, the dispersion of a pre-emulsified oil produced by the "phase inversion temperature (PIT) method" has been evaluated . In both cases, the volume fraction of oil was 2% . For the RSO systems, a relatively high agitation speed was required to disperse the oil, especially in the presence of the particles and, when the agitation was stopped, separation occurred rapidly . The Sauter mean drop diameters depended on the system, being at an average energy dissipation rate of approximately 0.9 W kg(-1), 180 microm for RSO/water, 130 microm for RSO/water(antifoam)/air, 580 microm for RSO/water/CaCO(3), and 850 microm for RSO/water(antifoam)/air/CaCO(3) . For the same four systems, the PIT emulsion, once dispersed, was very stable and the drop size was essentially independent of the operating conditions, with a Sauter mean diameter of approximately 0.3 microm . The implications of these findings for fermentations in which oil is used as a carbon source are assessed .

Biotechnol Bioeng, 2003 Jun 5, 82(5), 533 - 42
Coextraction during reactive extraction of phenylalanine using Aliquat 336: modeling extraction equilibrium; Pursell MR et al.; Reactive liquid-liquid extraction can be used to recover hydrophilic fermentation products that would not otherwise partition into nonpolar solvents through an ion-exchange reaction at the two-phase interface . However, the ion-exchange reagent may not be specific to the solute of interest and other compounds present may also be extracted . In this study, the effect on solute extraction of other compounds present in the extraction medium was investigated for phenylalanine extraction using Aliquat 336 . The extent of extraction at equilibrium was modeled using the equilibrium constants for the reactions present in the process . The interaction of different species within a multicomponent medium was examined using the model and experimental results . It was found that the extent of extraction and coextraction is controlled by the thermodynamics of each extraction reaction and, due to the formation of a common product, the interaction between each of the reactions . The main competition to reactive extraction will come from hydrophobic anionic compounds that will be present in fermentation broth .

J Chromatogr B Analyt Technol Biomed Life Sci, 2003 Mar 25, 786(1-2), 229 - 36
Optimisation of expression and immobilized metal ion affinity chromatographic purification of recombinant (His)6-tagged cytochrome P450 hydroperoxide lyase in Escherichia coli; Delcarte J et al.; Fatty acid hydroperoxide lyase (HPL) is a cytochrome P450 acting on fatty acid's hydroperoxides in many plants . The optimisation of the expression of recombinant (His)(6)-tagged HPL in Escherichia coli is described: the highest HPL production yield were obtained with TB medium supplemented with 2.5 mM delta-aminolevulinic acid and 0.5 mM IPTG . For the first time, the time course expression of a plant P450 in a bench-scale fermentor is detailed and the amount of recombinant HPL production is 16.3 mg/l . The UV-Visible spectrum of the recombinant (His)(6)-tagged HPL have been recorded after a Ni(2+)-based affinity chromatography (IMAC).

J Vet Med A Physiol Pathol Clin Med, 2003 Feb, 50(1), 8 - 13
The influence of a fermentation-resistant glucose diet on the glucose concentration and other metabolites in portal and jugular blood in cows; Braun U et al.; The goal of this study was to determine the effect of fermentation-resistant glucose on the glucose concentration and other metabolites in portal and jugular blood in 15 non-lactating cows . In all cows, an indwelling catheter was placed in the left jugular vein and the portal vein for collection of blood samples . Five control cows were fed hay as a normal diet, five control cows were fed straw to induce an energy deficit and five cows were fed hay and they received additionally 2000 g of a fermentation-resistant D-glucose product . The glucose concentration in jugular and portal blood was not influenced by feeding . The concentration of urea and bile acids were significantly higher in portal blood than jugular blood . There was no difference between portal and jugular blood of glucose and total solids . Diet had a significant effect on the concentrations of ammonia, urea, free fatty acids and triglycerides . The concentrations of ammonia and urea were higher in blood of cows fed straw than in blood of cows fed either hay or a fermentation-resistant glucose product . The concentration of urea remained constant in cows fed hay, but increased in cows fed straw and decreased in cows fed a fermentation-resistant glucose product . The concentration of free fatty acids and triglycerides were significantly higher in cows fed a fermentation-resistant glucose product than in cows fed hay . In the present study, a single administration of 300 g of fermentation-resistant glucose did not affect the concentration of blood glucose . Therefore, despite ongoing promotion of such products, there is no indication at this time that administration of fermentation-resistant glucose to cows at the start of lactation results in an increase in blood glucose concentration.

J Dairy Sci, 2003 Feb, 86(2), 644 - 52
Effects of abomasal infusion of conjugated linoleic acid on milk fat concentration and yield from pasture-fed dairy cows; Mackle TR et al.; The aim of this study was to investigate the effects of conjugated linoleic acid supplementation on the synthesis of milk fat in pasture-fed Friesian cows . In four cows, a commercial mixture containing 62.3% (wt/vol) conjugated linoleic acid was infused intraabomasally to avoid rumen fermentation and biohydrogenation . The design was a 4 x 4 Latin square in which each cow received infusions of 0, 20, 40, and 80 g/d of conjugated linoleic acid mixture for 4 d . Cows were fed freshly cut ryegrass/white clover pasture ad libitum . Milk fat concentration was decreased by 36, 43, and 62% and milk fat yield was decreased by 32, 36, and 60% by the 20, 40, and 80 g of conjugated linoleic acid/d treatments . Dry matter intake, milk protein concentration, and protein yield were unaffected by treatments; however, milk yield was increased by 11% during the 40-g conjugated linoleic acid/d treatment . The effects of conjugated linoleic acid infusion were most pronounced in reducing de novo fatty acid synthesis and desaturation . Results show that the inhibitory effect of this conjugated linoleic acid mixture on milk fat synthesis occurs in pasture-fed cows, and demonstrate the potential to dramatically alter gross milk composition . This technology could offer a management tool to manipulate milk composition and energy demands of pasture-fed cows.

J Dairy Sci, 2003 Feb, 86(2), 630 - 43
Effects of particle size of alfalfa-based dairy cow diets on chewing activity, ruminal fermentation, and milk production; Beauchemin KA et al.; Effects offorage particle size measured as physically effective NDF and ratio of alfalfa silage to alfalfa hay of diets on feed intake, chewing activity, particle size reduction, salivary secretion, ruminal fermentation, and milk production of dairy cows were evaluated using a 4 x 4 Latin square design with a 2 x 2 factorial arrangement of treatments . The diets consisted of 60% barley-based concentrate and 40% forage, comprised either of 50:50 or 25:75 of alfalfa silage:alfalfa hay, and alfalfa hay was either chopped or ground . Various methods were used to determine physically effective NDF content of the diets . Cows surgically fitted with ruminal and duodenal cannulas were offered ad libitum access to these total mixed diets . The physically effective NDF content of the diets was significantly lower when measured using the Penn State Particle Separator than when measured based on particles retained on 1.18-mm screen . Intake of DM was increased by increasing the ratio of silage to hay but was not affected by physically effective NDF content of diets . Eating time (hours per day) was not affected by the physically effective NDF content of diets, although cows spent more time eating per unit of DM or NDF when consuming high versus low alfalfa hay diets . Ruminating time (hours per day) was increased with increased physically effective NDF content of the diets . Rumen pH was affected more by changing dietary particle size than altering the ratio of silage to hay . Feeding chopped hay instead of ground hay improved ruminal pH status: time during which ruminal pH was above 6.2 increased and time during which ruminal pH was below 5.8 decreased . Milk production was increased by feeding higher concentrations of alfalfa silage due to increased DM intake, but was not affected by dietary particle size . Feed particle size, expressed as mean particle length or physically effective NDF was moderately correlated with ruminating time but not with eating time . Although physically effective NDF and chewing time were not correlated to mean rumen pH, they were negatively correlated to the area between the curve and pH 5.8, indicating a positive effect on reducing the risk of acidosis . Milk fat content was correlated to rumen pH but not to physically effective NDF or chewing activity . These results indicate that increasing physically effective NDF content of the diets increased chewing activity and improved rumen pH status but had limited effect on milk production and milk fat content.

J Dairy Sci, 2003 Feb, 86(2), 593 - 609
Effects of tallow in diets based on corn silage or alfalfa silage on digestion and nutrient use by lactating dairy cows; Ruppert LD et al.; Six multiparous Holstein cows (average 31 days in milk; 36.3 kg/d of milk) fitted with ruminal cannulas were used in a 6 x 6 Latin square with 21-d periods to investigate the effects of diets that varied in forage source and amount of supplemental tallow . Isonitrogenous diets in a 2 x 3 factorial arrangement were based on either high corn silage (40:10 corn silage to alfalfa silage, % of dry matter) or high alfalfa silage (10:40 corn silage to alfalfa silage, % of dry matter) and contained 0, 2, or 4% tallow . Intakes of dry matter and total fatty acids were lower when cows were fed the high corn silage diet . Tallow supplementation linearly decreased dry matter intake . Milk yield was unaffected by diet; yields of milk fat and 3.5% fat-corrected milk were higher for the high alfalfa silage diet but were unaffected by tallow . Milk fat percentage was higher for the high alfalfa silage and tended to decrease when tallow was added to the high corn silage diet . Contents of trans-C18:1 isomers in milk fat were increased by high corn silage and tallow, and tended to be increased more when tallow was fed in the high corn silage diet . Ruminal pH and acetate:propionate were lower when high corn silage was fed . Ruminal acetate:propionate decreased linearly as tallow increased; the molar proportion of acetate was decreased more when tallow was added to the high corn silage diet . Ruminal liquid dilution rates were higher for the alfalfa silage diet; ruminal volume and solid passage rates were similar among diets . Total tract apparent digestibilities of dry matter, organic matter, crude protein, starch, energy, and total fatty acids were unaffected by diet . Digestibilities of neutral detergent fiber, acid detergent fiber, hemicellulose, and cellulose were lower when high corn silage was fed . The high alfalfa silage diet increased intakes of metabolizable energy and N, and increased milk energy and productive N . Tallow decreased the amount of N absorbed but had few other effects on utilization of energy or N . Tallow linearly increased concentrations of nonesterified fatty acids and cholesterol in plasma; cholesterol was increased by high alfalfa silage . Overall, forage source had more pronounced effects on production and metabolism than did tallow supplementation . Few interactions between forage source and tallow supplementation were detected except that ruminal fermentation and milk fat content were affected more negatively when tallow was fed in the high corn silage diet.

J Dairy Sci, 2003 Feb, 86(2), 565 - 75
Fibrolytic enzymes and parity effects on feeding behavior, salivation, and ruminal pH of lactating dairy cows; Bowman GR et al.; Four multiparous and four primiparous lactating dairy cows fitted with ruminal cannulas were used in a duplicated 4 x 4 Latin square design to study the effects of parity and inclusion of a fibrolytic enzyme product (Agribrands International, St . Louis, MO) on feeding and chewing behavior, salivation, and ruminal pH . Diets consisting of rolled barley, barley silage, and alfalfa haylage (55% forage, DM basis) differed in enzyme application: 1) control, 2) enzyme applied to concentrate (45% of TMR), 3) enzyme applied to supplement (4% of TMR), and enzyme applied to a premix (0.2% of TMR) . Enzyme supplementation did not alter daily time spent eating or ruminating, but when enzymes were added to the ration daily, saliva production increased, with no difference among enzyme application treatments . Multiparous cows consumed a greater amount of feed, but spent a similar amount of time eating, compared to primiparous cows . Primiparous cows had shorter ruminating episodes, resulting in lower daily ruminating time compared with multiparous cows . Primiparous cows had lower daily saliva output compared with multiparous cows . These results indicate that application of this fibrolytic enzyme product did not alter the physical structure of the feed, as measured by feeding and chewing variables . The increase in total saliva production observed in cows fed enzyme-supplemented diets may be attributed to a physiological response to compensate for the increase in fermentation products during digestion . The increased intake for multiparous cows is attributed to increased eating rate and not to increased time spent eating . The higher DMI of multiparous cows resulted in increased rumination time needed to process the additional feed and increased salivation to buffer the greater production of VFA.

FEMS Microbiol Lett, 2003 Mar 14, 220(1), 75 - 80
A novel computerised image analysis method for the measurement of production of conidia from the aphid pathogenic fungus Erynia neoaphidis; Bonner TJ et al.; A semi-automated method has been developed for the quantification and measurement of conidia discharged by the aphid pathogen Erynia neoaphidis . This was used to compare conidiation by E . neoaphidis-mycosed pea aphid cadavers, mycelial plugs cut from agar plates, mycelial pellets from shake flasks and by mycelial pellets from different phases of liquid batch fermenter culture . Aphid cadavers discharged significantly more and significantly smaller conidia than plugs or pellets . The volume of conidia discharged was stable over the period of discharge (80 h), but more detailed analysis of the size frequency distribution showed that more very small and very large conidia were discharged after 5 h incubation than after 75 h incubation . Biomass harvested at the end of the exponential growth phase in batch fermenter culture produced significantly more conidia than biomass from any other growth phase . The implications of these findings for the development of production and formulation processes for E . neoaphidis as a biological control agent are discussed.

J Agric Food Chem, 2003 Mar 26, 51(7), 2076 - 82
Natural abundance hydrogen isotope affiliation between the reactants and the products in glucose fermentation with yeast; Pionnier S et al.; In glucose fermentation, the hydrogen source of products such as ethanol and glycerol is the medium and the sugar . The site-specific natural isotope ratios of the products, (D/H)(i), and that of the medium and sugar, (D/H)(k), may be related by a matrix, A, of redistribution coefficients, a(ik), that characterizes the specific genealogies of the hydrogen atoms . (D/H)(i) = {A}(D/H)(k), where (D/H)(i) and (D/H)(k) are the column vectors of the isotope ratios of sites i and k that can be measured by (2)H NMR . The complete redistribution matrix was determined in a set of isotope labeling experiments . Thus, we obtained a mathematical model representing the hydrogen isotope affiliation during alcoholic fermentation . It not only provides information about the biochemical reaction mechanism but also can be used to estimate the isotopic data of the products, based on those of the substrate and the medium . The results prove, in a quantitative way, that the metabolites contain isotopic information about the precursor in a biotransformation and can be used to identify its origin . The method established for the study of the hydrogen-transfer mechanism can be applied to other chemical and biochemical reactions.

J Agric Food Chem, 2003 Mar 26, 51(7), 2040 - 8
Clarification of Muscat musts using wheat proteins and the flotation technique; Marchal R et al.; The bovine spongiform encephalopathy (BSE, mad cow) crisis has led some wine-makers to question gelatin as a fining agent and to reject the use of these animal proteins . The search for a substitute for gelatin was begun by comparing vegetable proteins (particularly gluten) with gelatin fining treatments . Clairette de Die (French-controlled appellation) is a sparkling sweet wine . The alcoholic fermentation begins in a tank, continues in a crown-cap bottle, and stops before the complete consumption of sugar . All particles present in the bottle have to be removed before corking, and it is important to have a must as clear as possible . This study concerned the clarifying of a Muscat must, treated with pectinases, using a very efficient flotation technique . The must is fined with bentonite/silica/protein (fish gelatin, wheat gluten, or lupin isolate) to induce flocculation and then pressurized (6 bar) . After depressurization, microbubbles cling to flocculates and climb to the top of the flotation tank (this flotation foam is clarified using a rotary filter) . At laboratory scale, gluten (20 g/hL) and gelatin (10 g/hL) (each combined with bentonite and silica gel) gave turbidities of 50 and 35 NTU, respectively (6.5 and 4.1% of that of the nontreated must) . The Muscat must was also clarified by static settling . The turbidity decreased by 86% for the gluten/bentonite fining and by 60% for the gelatin/bentonite fining . Visually, gluten flocculation takes longer to occur and flocculates sedimentation is longer than with gelati, but the removal of insoluble particles is more complete and leads to lower turbidities . At an industrial scale, gluten (20 g/hL) and gelatin (10 g/hL) (each combined with bentonite and silica gel) gave turbidities of 60 and 48 NTU, respectively . Turbidities measured in the tanks 14 h after the flotation showed a better efficiency for the wheat gluten (24 NTU) compared to gelatin (28 NTU) . This is explained by the static settling that completed the clarifying effect of the flotation . The last experiment showed comparable efficiencies for wheat gluten and lupin proteins . BSE caused a situation of crisis (in Europe particularly) leading the public and wine-makers to lose their confidence in the use of gelatin as fining agent and to reject animal proteins in general . It is proposed here that vegetable proteins could efficiently replace gelatin.

J Agric Food Chem, 2003 Mar 26, 51(7), 1889 - 93
Optimization of making barrel-fermented dry muscatel wines; Aleixandre JL et al.; The optimization of making barrel-fermented muscatel wines requires determining what type of must clarification is most suitable for the quality of the wine, as well as what type of barrel will yield the most acceptable wines . This is achieved by adding pectolytic enzymes to clarify part of the muscatel must statically; the rest is clarified by vacuum filtration . The musts obtained are fermented in French and American oak barrels and, once fermentation has ceased, they are kept with their lees for 2 months, with periodic stirring . Eleven conventional parameters and 31 volatile compounds were quantified, and a sensory analysis of the wines was produced, which led us to conclude that static clarification with pectolytic enzymes from the muscatel musts produces the best-structured wines and the larger content of higher alcohols, esters, and terpenic compounds . The wines fermented in American oak barrels received the highest overall marks, which may be due to the greater aromatic complexity given off by the compounds in the wood.

J Agric Food Chem, 2003 Mar 26, 51(7), 1864 - 73
Factors affecting the levels of tea polyphenols and caffeine in tea leaves; Lin YS et al.; An isocratic HPLC procedure was developed for the simultaneous determination of caffeine and six catechins in tea samples . When 31 commercial teas extracted by boiling water or 75% ethanol were analyzed by HPLC, the levels of (-)-epigallocatechin 3-gallate (EGCG), and total catechins in teas were in the order green tea (old leaves) > green tea (young leaves) and oolong tea > black tea and pu-erh tea . Tea samples extracted by 75% ethanol could yield higher levels of EGCG and total catechins . The contents of caffeine and catechins also have been measured in fresh tea leaves from the Tea Experiment Station in Wen-Shan or Taitung; the old tea leaves contain less caffeine but more EGCG and total catechins than young ones . To compare caffeine and catechins in the same tea but manufactured by different fermentation processes, the level of caffeine in different manufactured teas was in the order black tea > oolong tea > green tea > fresh tea leaf, but the levels of EGCG and total catechins were in the order green tea > oolong tea > fresh tea leaf > black tea . In addition, six commercial tea extracts were used to test the biological functions including hydroxyl radical scavenging, nitric oxide suppressing, and apoptotic effects . The pu-erh tea extracts protected the plasmid DNA from damage by the Fenton reaction as well as the control at a concentration of 100 microg/mL . The nitric oxide suppressing effect of tea extracts was in the order pu-erh tea >/= black tea > green tea > oolong tea . The induction of apoptosis by tea extract has been demonstrated by DNA fragmentation ladder and flow cytometry . It appeared that the ability of tea extracts to induce HL-60 cells apoptosis was in the order green tea > oolong > black tea > pu-erh tea . All tea extracts extracted by 75% ethanol have stronger biological functions than those extracted by boiling water.

J Anim Sci, 2003 Feb, 81(2), 537 - 44
Diet and procedures used to detach particle-associated microbes from ruminal digesta influence chemical composition of microbes and estimation of microbial growth in Rusitec fermenters; Ranilla MJ et al.; Four different detachment methods were evaluated for their ability to remove particle-associated microorganisms (PAM) from ruminal digesta in semicontinuous fermenters fed two diets differing in their forage:concentrate ratio (80:20 {C20} and 20:80 {C80}) . In the methylcellulose method, ruminal digesta was incubated at 38 degrees C for 15 min with saline solution containing 0.1% methylcellulose before being stored at 4 degrees C for 24 h . In the other procedures, samples were incubated with 0.1% methylcellulose before storage for 24 h at 4 degrees C in different solutions (pH = 2): 1) saline solution with 0.1% Tween 80; 2) saline solution with 0.1% Tween 80 and 1% tertiary butanol; and 3) saline solution with 0.1% Tween 80, 1% methanol, and 1% tertiary butanol . Common to all treatments was subsequent homogenization, followed by filtration and resuspension of the residue five times in the treatment solutions . Microbial removal was estimated indirectly by measuring removal of 15N . There were no differences (P > 0.05) among detachment procedures, neither in the detaching efficiency (mean values of 79.7 and 88.1% for C20 and C80 diets, respectively) nor in the total recovery of PAM (54.9 and 34.9% for C20 and C80, respectively) . There were no differences (P > 0.05) among PAM pellets obtained by the different detachment procedures in their N content, purine bases (PB) concentration, or PB:N ratio . For the C80 diet, 15N enrichment was greater (P < 0.05) in PAM pellets obtained with methylcellulose than in those obtained by the other methods . However, there were no differences (P > 0.05) due to the detachment procedure in the values of daily microbial growth estimated using as reference the different PAM pellets . The PAM pellets for diet C20 presented greater (P < 0.01) N content and lower (P < 0.01) PB concentration than those for diet C80 (mean values of 74.3 vs 49.1 mg of N/g of dry matter, and 22.8 vs 26.0 micromol PB/mg of dry matter, respectively) . Daily microbial growth was greater (P < 0.05) for the C80 diet than for the C20 diet (121 vs 114 mg of microbial N, respectively) . Results suggest that the treatment of ruminal digesta with a saline solution with 0.1% methylcellulose at 38 degrees C for 15 min combined with homogenizing and chilling at 4 degrees C for 24 h removed a major proportion of PAM, although further research is needed to decrease microbial losses during the isolation process.

Lett Appl Microbiol, 2003, 36(4), 203 - 7
Production and partial characterization of a beta-amylase by Xanthophyllomyces dendrorhous; Diaz A et al.; AIMS: The characterization of a beta-amylase produced by Xanthophyllomyces dendrorhous . METHODS AND RESULTS: Growth in different culture media showed that X . dendrorhous produces an amylase whose synthesis is repressed by the carbon source and induced by starch and maltose . Enzymatic assays using substrates with different degrees of polymerization together with viscosity experiments revealed that the enzyme was beta-amylase . According to the biochemical characterization, the enzyme has a molecular weight of 240 kDa and a Km of 1.35 mg ml-1 . The optimum pH and temperature were 5.5 and 50 degrees C, respectively . Using different inhibitors of the enzymatic activity it was shown that cysteine, tryptophan and serine are essential amino acids for catalysis . CONCLUSIONS: Xanthophyllomyces dendrorhous CECT1690 synthesizes and secretes beta-amylase that could be a by-product, in addition to carotenoid pigments, in the fermentation downstream . SIGNIFICANCE AND IMPACT OF THE STUDY: The beta-amylase produced by X . dendrorhous may have certain industrial applications.

Nat Biotechnol, 2003 Apr, 21(4), 422 - 7 Epub 2003 Mar 17.
Semisynthetic production of unnatural L-alpha-amino acids by metabolic engineering of the cysteine-biosynthetic pathway; Maier TH; There is an increasing demand for peptide-mimicking molecules to modulate the interactions between proteins of pharmaceutical and agrochemical interest and their target polypeptides . Unnatural L-alpha-amino acids differing from the 20 naturally proteinogenic amino acids only in their side chain are ideal for this purpose, but their chemical synthesis is complex . Here we describe a fermentation-based approach for biosynthesis of unnatural amino acids after re-engineering the cysteine-biosynthetic pathway in Escherichia coli . O-acetylation of serine, the committed step of the pathway, was released from feedback inhibition by mutating the serine acetyltransferase gene . Next, the naturally broad substrate specificity of O-acetylserine sulfhydrylase was exploited for the direct in vivo incorporation of an unnatural side chain in a semisynthetic fermentation process comparable to the production of beta-lactams . O-acetyl-L-serine extruded from the cells by way of the O-acetylserine efflux protein was amenable to further biotransformations.

Public Health Nutr, 2002 Dec, 5(6B), 1259 - 71
Consumption of dairy products in the European Prospective Investigation into Cancer and Nutrition (EPIC) cohort: data from 35 955 24-hour dietary recalls in 10 European countries; Hjartaker A et al.; OBJECTIVES: To describe and compare the consumption of dairy products in cohorts included in the European Prospective Investigation into Cancer and Nutrition (EPIC) . METHODS: Data from single 24-hour dietary recall interviews collected through a highly standardised computer-based program (EPIC-SOFT) in 27 redefined centres in 10 European countries between 1995 and 2000 . From a total random sample of 36 900, 22 924 women and 13 031 men were selected after exclusion of subjects under 35 and over 74 years of age . RESULTS: A high total consumption of dairy products was reported in most of the centres in Spain and in the UK cohort sampled from the general population, as well as in the Dutch, Swedish and Danish centres . A somewhat low consumption was reported in the Greek centre and in some of the Italian centres (Ragusa and Turin) . In all centres and for both sexes, milk constituted the dairy sub-group with the largest proportion (in grams) of total dairy consumption, followed by yoghurt and other fermented milk products, and cheese . Still, there was a wide range in the contributions of the different dairy sub-groups between centres . The Spanish and Nordic centres generally reported a high consumption of milk, the Swedish and Dutch centres reported a high consumption of yoghurt and other fermented milk products, whereas the highest consumption of cheese was reported in the French centres . CONCLUSION: The results demonstrate both quantitative and qualitative disparities in dairy product consumption among the EPIC centres . This offers a sound starting point for analyses of associations between dairy intake and chronic diseases such as cancer.

Indian J Exp Biol, 2002 Mar, 40(3), 325 - 8
Fermentation of starch to ethanol by an amylolytic yeast Saccharomyces diastaticus SM-10; Sharma S et al.; A total of fifteen yeast strains were isolated from natural sources including fruits, soil, molasses, honey and a variety of indigeneous fermented foods . Screening of these strains for growth, ethanol production and glucoamylase activity led to selection of a yeast strain SM-10 identified as S . diastaticus having maximum glucoamylase activity (80 units ml(-1)) and ethanol production from starch (3.5%) . Ethanol production from wheat flour was found to be 1.75% which could be increased to 5.2% after treatment of wheat flour with pepsin, diastase and glucoamylase.

Microbiology, 2003 Mar, 149(Pt 3), 557 - 67
Beginnings of microbiology and biochemistry: the contribution of yeast research; Barnett JA; With improvements in microscopes early in the nineteenth century, yeasts were seen to be living organisms, although some famous scientists ridiculed the idea and their influence held back the development of microbiology . In the 1850s and 1860s, yeasts were established as microbes and responsible for alcoholic fermentation, and this led to the study of the role of bacteria in lactic and other fermentations, as well as bacterial pathogenicity . At this time, there were difficulties in distinguishing between the activities of microbes and of extracellular enzymes . Between 1884 and 1894, Emil Fischer's study of sugar utilization by yeasts generated an understanding of enzymic specificity and the nature of enzyme-substrate complexes.

Med Dosw Mikrobiol, 2002, 54(3), 225 - 31
{Use of new methods for detection of beta-lactamases with extended-spectrum substrates (ESBLs)}; Rokosz A et al.; The aim of performed examinations was to compare results of two methods applied for the detection of extended-spectrum beta-lactamases (ESBLs) . Two hundred and sixty strains of Gram-negative rods were cultured from clinical specimens obtained from hospitalized patients . These strains were identified as ESBL-positive on the basis of the double-disc method (DDST) according to Jarlier et al . (1988) results . Identification of strains was performed in automatic ATB Expression system (bioMerieux, France) . All strains were determined using a novel test for detection of ESBL-type enzymes (DD) according to Appleton (1999) . Two discs are applied in this test: with cefpodoxime (CPD) and with cefpodoxime/clavulanate (CD 01, diagnostic disc) . Consistent results of two methods (DDST and DD) were obtained in the case of 166 from among 260 of examined strains (60.4%) . Consistent results concerned 161 out of 222 examined strains of enteric rods (72.5%) and only 5 from among 38 of other strains (mostly belonging to the group of non-fermenting rods) . On the basis of performed investigations it can be stated that the novel method of extended-spectrum beta-lactamases (ESBLs) detection (DD) according to Appleton (1999) is more objective and easier for interpretation than the double-disc synergy test (DDST) according to Jarlier et al . (DDST), which is widely applied in the routine microbiological diagnostics.

Med Dosw Mikrobiol, 2002, 54(3), 215 - 23
{Biochemical properties of enterotoxic and enterohemorrhagic E-coli strains}; Karakulska J; The purpose of this paper has been to describe the biological characteristics of thirty strains of E . coli . The E . coli strains were isolated from cases of colibacillosis in animals and from human faeces and cow milk samples . Of the thirty analyzed strains, 19 strains (63%) were found to belong to 7 serogroups: O8, O101, O138, O141, O147, O149 and O157 . In 17 strains (57%) fimbriae F4 was discovered and in 1 strain (3%) the presence of fimbriae F5 and F41 was detected . Serological and biochemical researches, based on the analysis of 35 enzymatic reactions, confirmed that all strains, used in this study, belonged to the species E . coli . The strains demonstrated differences in biochemical activity for 12 substrates . It was found that strains of serotype O157: H7 had biochemical homogeneity, except in their rate of sucrose fermentation and their ability to hydrolyze arginine and sorbitol after longer incubation . On the basis of the biochemical activity, O157: H7 strains were affiliated with biotype C . In identifying serotype O157: H7, SMAC medium with sorbitol and liquid and solid media with MUG reagent were very useful.

Biotechnol Bioeng, 2003 May 20, 82(4), 429 - 37
Microfiltration and ultrafiltration of polysaccharides produced by fermentation using a rotating disk dynamic filtration system; Brou A et al.; The recovery of exopolysaccharides (EPS) produced by Sinorhizobium meliloti bacteria by dynamic microfiltration was investigated using a rotating disk device designed in our laboratory, equipped with a 0.2 microm nylon membrane . This system differs from commercially available systems by the presence of vanes on the disk which produce a very important increase in permeate flux while yielding excellent EPS transmission . For polymers produced under standard fermentation conditions (70 h at 30 degrees C), the mass flux rose to 650 g h(-1) m(-2) using a disk equipped with 2 mm vanes rotating at 2000 rpm against 380 g h(-1) m(-2) with a smooth disk at the same speed . The maximum flux observed was 1560 g h(-1) m(-2) with a 6-mm vanes disk rotating at 3000 rpm and a 36 degrees C broth . An interesting finding was that the permeate flux J(f) for various disks can be correlated by the same function of the mean shear stress at the membrane tau(wm) according to J(f) = 4.6 tau(wm) (0.717) for a 30 degrees C broth, showing that the effect of vanes is merely to increase the shear stress by raising the fluid core velocity between the membrane and the disk . With 6-mm vanes the core angular velocity was found to be 84% of disk velocity vs . 45% for a smooth disk . When the fermentation temperature was increased to 36 degrees C to produce a lower molecular weight polymer, the permeate flux rose by about 250%, much more than what could be expected from the reduction in permeate viscosity and followed the same power law with membrane shear stress as for 30 degrees C . The same device was equipped with a PES 50 kDa membrane to concentrate EPS by ultrafiltration . Permeate fluxes were of the order of 160 L h(-1) m(-2) at 2000 rpm and 30 degrees C with nearly complete EPS rejection . Finally, the net electrical power consumed by the disk was measured by subtracting the power consumed without fluid from the power during filtration at the same speed . This power increases with speed and with the presence of vanes, but since the gain provided by the vanes is very high, the specific energy per m(3) of permeate is minimal with the highest vanes tested (6 mm) and maximal for smooth disks .

Arch Microbiol, 2003 Apr, 179(4), 278 - 94 Epub 2003 Mar 11.
Genome-wide transcriptional changes during the lag phase of Saccharomyces cerevisiae; Brejning J et al.; The set of physiological and metabolic changes occurring immediately after inoculation and during the lag phase is thought to be of vital importance for optimal offset of fermentation . The transcriptional changes taking place during the lag phase after inoculation of a late-respiratory-phase yeast culture into fresh, minimal medium were investigated by use of Yeast GeneFilters . In response to the nutritional up-shift, 240 open reading frames were at least five-fold induced and 122 were at least five-fold repressed . These genes were hierarchically clustered according to their lag-phase expression patterns . The majority of the induced genes were most highly induced early in the lag phase, whereas strong repression generally occurred later . Clustering of the genes showed that many genes with similar roles had similar expression patterns . Repressed genes, however, did not cluster as tightly according to function as induced genes . Genes involved in RNA and protein synthesis and processing showed a peak in expression early in the lag phase, except most ribosomal protein genes, which were induced early and whose expression was sustained . Genes involved in chromatin/chromosome structure showed late induction . The correlation between function and expression pattern for these genes indicates regulation by similar mechanisms . Much of the transcriptional response observed appeared to be due to the presence of glucose in the new medium.

J Appl Microbiol, 2003, 94(4), 708 - 19
Production and molecular characteristics of four groups of exopolysaccharides from submerged culture of Phellinus gilvus; Hwang HJ et al.; AIMS: The objective of the present study was to determine the optimal culture conditions for the production of four groups of exopolysaccharides (EPSs) in Phellinus gilvus by submerged culture and to investigate their molecular properties by multi-angle laser-light scattering (MALLS) analysis . METHODS AND RESULTS: The optimal temperature and initial pH for the production of both mycelial biomass and EPSs in P . gilvus by submerged flask cultures were found to be 30 degrees C and pH 9.0, respectively . Glucose and corn steep powder were the most suitable carbon and nitrogen source for both mycelial biomass and EPS production . Optimal medium composition was determined to be glucose 30 g l-1, corn steep powder 5 g l-1, MgSO4 1.23 g l-1, KH2PO4 0.68 g l-1, and K2HPO4 0.87 g l-1 . Four groups of EPSs (Fr-I, II, III, and IV) were obtained from the culture filtrates by gel filtration chromatography on Sepharose CL-4B and characterized by size exclusion chromatography (SEC) coupled with MALLS . The weight average molar mass (Mw) of Fr-I, Fr-II, Fr-III and Fr-IV were determined to be 8.628 x 106 (+/-129 420), 1.045 x 106 (+/-19 855), 61.09 x 104 (+/-1244), and 33.55 x 104 (+/-134) g mol-1, respectively . CONCLUSIONS: Under optimal culture conditions, the maximum EPS production in a 5-l stirred fermenter indicated 5.3 g l-1 after 11 days of fermentation . The SEC/MALLS analysis revealed that Fr-I, which has extremely high molecular weight, was presumably an aggregate of complex polysaccharides forming a compact globular shape; whereas Fr-II was nearly spherical, Fr-III and Fr-IV were rod-like chains in an aqueous solution . SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report on the production of high amounts of EPSs from liquid-culture of the basidiomycete, P . gilvus . The SEC/MALLS approach used in this study could be useful in providing greater insight into the characterization of the mushroom polysaccharides without carrying out elaborate fractionation procedures prior to analysis.

J Appl Microbiol, 2003, 94(4), 555 - 60
The adaptive response of Escherichia coli O157 in an environment with changing pH; de Jonge R et al.; AIMS: To predict and validate survival of non-acid adapted Escherichia coli O157 in an environment mimicking the human stomach . METHODS AND RESULTS: Survival was predicted mathematically from inactivation rates at various, but constant pH values . Predictions were subsequently validated experimentally in a pH-controlled fermentor . Contrary to prediction, acid-sensitive cultures of E . coli O157 survived for a long period of time and died as rapidly as acid-resistant cultures . Experimental results showed that in an environment with changing pH, acid-sensitive cultures became acid-resistant within 17 min . Cyclo fatty acids was reported to be a factor in acid resistance . As synthesis of cyclo fatty acids does not require de novo enzyme synthesis and thus requires little time to develop, we analysed the membrane fatty acid composition of E . coli O157 during adaptation . No changes in membrane fatty acid composition were observed . CONCLUSIONS: Acid adaptation of E . coli O157 can occur during passage of the human gastric acid barrier, which can take up to 4 h . SIGNIFICANCE AND IMPACT OF THE STUDY: The ability of acid-adapted bacteria to survive the human stomach is an important virulence factor . The ability of non-acid adapted E . coli O157 to adapt within a very short period of time under extreme conditions further contributes to the virulence of E . coli O157.

Steroids, 2003 Mar, 68(3), 271 - 5
Effect of a novel steroid (PM-9) on the inhibition of 5alpha-reductase present in Penicillium crustosum broths; Flores E et al.; The conversion of testosterone (T) to 5alpha-dihydrotestosterone (DHT) has been demonstrated in Penicillium crustosum broth obtained from fermented pistachios, lemons and corn tortillas . Furthermore, the presence of 5alpha-reductase enzyme, which is responsible for this conversion, has been established by electrophoretical techniques in these cultures.5alpha-Reductase enzyme is also present in animal and human androgen-dependent tissues as well as in prostate and seminal vesicles . The increase of the conversion of T to DHT in prostate gland, has been related to some illnesses such as benign prostate hyperplasia and prostate cancer . Furthermore, treatment with 5alpha-reductase inhibitors such as finasteride reduces the prostate growth . These data have stimulated research for the synthesis of new molecules with antiandrogenic activity, whose biological effect needs to be demonstrated.The purpose of this study is to determine the inhibition pattern of 5alpha-reductase in P . crustosum by finasteride and the new steroidal compound PM-9 . K(m) and V(max) values for T, were determined in the broths by Lineweaver-Burk plots using different testosterone concentrations . The inhibition pattern of finasteride and PM-9 was also determined by Lineweaver-Burk using different concentrations of T and inhibitors . Results show that finasteride and PM-9 inhibit 5alpha-reductase present in the broth in a competitive manner.

Bioorg Med Chem, 2003 Apr 3, 11(7), 1577 - 82
Integracides: tetracyclic triterpenoid inhibitors of HIV-1 integrase produced by Fusarium sp; Singh SB et al.; HIV-1 integrase is a critical enzyme in the replication of HIV-1 . It is absent in the host cells and therefore is a good target for treatment of HIV-1 infections . Integracides are members of the tetracyclic triterpenoids family that were isolated from the fermentation broth of a Fusarium sp . Integracide A, a sulfated ester, exhibited significant inhibitory activity against strand transfer reaction of HIV-1 integrase . The discovery, structure elucidation including single crystal X-ray structure and HIV-1 inhibitory activity of these compounds are described.

Yeast, 2003 Mar, 20(4), 295 - 314
Proteome analysis of recombinant xylose-fermenting Saccharomyces cerevisiae; Salusjarvi L et al.; Introduction of an active xylose utilization pathway into Saccharomyces cerevisiae, which does not naturally ferment pentose sugars, is likely to have a major impact on the overall cellular metabolism as the carbon introduced to the cells will now flow through the pentose phosphate pathway . The metabolic responses in the recombinant xylose-fermenting S . cerevisiae were studied at the proteome level by comparative two-dimensional gel electrophoresis of cellular proteins within a pH range of 3-10 . Glucose-limited chemostat cultivations and corresponding chemostat cultivations performed in media containing xylose as the major carbon source were compared . The cultivations were studied in aerobic and anaerobic metabolic steady states and in addition at time points 5, 30 and 60 min after the switch-off of oxygen supply . We identified 22 proteins having a significant abundance difference on xylose compared to glucose, and 12 proteins that responded to change from aerobic to anaerobic conditions on both carbon sources . On xylose in all conditions studied, major changes were seen in the abundance of alcohol dehydrogenase 2 (Adh2p), acetaldehyde dehydrogenases 4 and 6 (Ald4p and Ald6p), and DL-glycerol 3-phosphatase (Gpp1p) . Our results give indications of altered metabolic fluxes especially in the acetate and glycerol pathways in cells growing on xylose compared to glucose .

Eur J Clin Nutr, 2003 Mar, 57(3), 490 - 5
Blood-pressure-lowering effect of a novel fermented milk containing gamma-aminobutyric acid (GABA) in mild hypertensives; Inoue K et al.; OBJECTIVE: To study the effect of a new fermented milk product containing GABA (FMG) on the blood pressure (BP) of patients with mild hypertension . DESIGN: A randomized, placebo-controlled, single-blind trial . SETTING: The study was carried out at the outpatient clinic of the Cardiovascular Disease Center, Tokyo Metropolitan Police Hospital, Japan . SUBJECTS: The study population comprised 39 mildly hypertensive patients (16 women and 23 men) aged 28-81 y (mean, 54.2 y) . INTERVENTIONS: The study consisted of a 12-week period of daily intake of FMG or placebo (weeks 1-12) followed by 2 weeks of no intake (weeks 13 and 14) . We measured the peripheral BP and heart rate of seated patients at weeks 0, 2, 4, 8, 12 and 14 . Routine blood study and urinalysis were performed before and after the intake . RESULTS: There was a significant decrease of BP within 2 or 4 weeks, and it remained decreased throughout the 12-week intake period . For the FMG recipients, the mean decrease after 12 weeks was 17.4+/-4.3 mmHg in the systolic BP (SBP) and 7.2+/-5.7 mmHg in the diastolic BP (DBP) . Both of these values differed statistically from baseline levels (P<0.01), and the SBP of the FMG group differed from the placebo group (P<0.05) . Heart rate, body weight, hematological and blood chemistry variables, and urinalysis results (glucosuria and proteinuria) did not vary both groups throughout the study . CONCLUSION: FMG may contribute to lowering BP in mildly hypertensive people.

Microbiol Mol Biol Rev, 2003 Mar, 67(1), 38 - 51, table of contents
Separation, identification, and characterization of microorganisms by capillary electrophoresis; Desai MJ et al.; The use of capillary electrophoresis (CE) for the analysis, identification, and characterization of microorganisms has been gaining in popularity . The advantages of CE, such as small sample requirements, minimal sample preparation, rapid and simultaneous analysis, ease of quantitation and identification, and viability assessment, make it an attractive technique for the analysis of microbial analytes . As this instrumental method has evolved, higher peak efficiencies have been achieved by optimizing CE conditions, such as pH, ionic strength, and polymer additive concentration . Experimental improvements have allowed better quantitation and more accurate results . Many practical applications of this technique have been investigated . Viability and identification of microbes can be accomplished in a single analysis . This is useful for evaluation of microbial analytes in consumer products . Diagnosis of microbe-based diseases is now possible, in some cases, without the need for culture methods . Microbe-molecule, virus-antibody, or bacteria-antibiotic interactions can be monitored using CE, allowing for the screening of possible drug candidates . Fermentation can be monitored using this system . This instrumental approach can be adapted to many different applications, including assessing the viability of sperm cells . Progress has been made in the development of microelectrophoresis instrumentation . These advances will eventually allow the development of small, dedicated devices for the rapid, repetitive analyses of specific microbial samples . Although these methods may never fully replace traditional approaches, they are proving to be a valuable addition to the collection of techniques used to analyze, quantitate, and characterize microbes . This review outlines the recent developments in this rapidly growing field.

Reprod Nutr Dev, 2002 Nov-Dec, 42(6), 545 - 57
Nitrogen utilisation by dairy cows fed diets differing in crude protein level with a deficit in ruminal fermentable nitrogen; Monteils V et al.; We studied the efficiency of nitrogen utilisation by dairy cows, using three diets differing in the crude protein (CP) level but with similar deficits (10 g x kg(-1) dry matter, DM) in ruminal fermentable nitrogen . There was no difference in milk yield from the cows offered the three diets (130, 145 and 160 g CP x kg(-1) DM) . The milk protein content differed between the two most extreme diets (28.9 vs . 29.9 g x kg(-1), P < 0.05), resulting in higher protein yields for the highest CP treatment (P < 0.01) . The efficiency of nitrogen utilisation, calculated as the proportion of ingested nitrogen recovered in the milk, was significantly higher for the 130 g CP x kg(-1) DM diet than for the other two diets (0.37 vs . 0.33 and 0.32 respectively . P < 0.01) . The different diets also resulted in different levels of nitrogen excretion into the environment (237, 270 and 330 g N x d(-1), P < 0.01) . Hepatic deamination of the amino acids may have generated additional energy to enable the animal to make use of the additional nitrogen in the diet, resulting in an increase in plasma urea concentration.

Reprod Nutr Dev, 2002 Nov-Dec, 42(6), 537 - 44
Rumen escape of methionine and lysine administered intraruminally to growing double-muscled Belgian Blue bulls; Froidmont E et al.; In many dietary conditions, methionine (Met) and lysine (Lys) are the most limiting amino acids (AA) for ruminants . The AA protected from ruminal fermentation are not commercially available, with the exception of Met which is not always economical, especially for meat production . This study measured ruminal escape of free Met and Lys supplemented intraruminally to fast growing bulls . Six double-muscled Belgian Blue bulls, fed a high concentrate diet and fitted with a rumen cannula, received free Met (40 g x d(-1)) and free Lys (60 g x d(-1)), individually or simultaneously, in a duplicated Latin square design . The mean ruminal escape of Met and Lys reached 37 and 45% respectively, and did not differ if administered separately or together . Plasma Lys and Met concentrations were increased by 504 and 126%, respectively . Substantial proportions of free AA escaped ruminal fermentation and were available for absorption from the small intestine when they were administered at physiologically high levels.

Onderstepoort J Vet Res, 2002 Dec, 69(4), 315 - 20
Water absorption in relation to fermentation in the colon of the ostrich (Struthio camelus); Musara C et al.; The colon is a major site for fermentation and water absorption in the ostrich . Water absorption along the colon was evaluated and its relationship to osmolality, Na+ concentration, short chain fatty acid (SCFA) concentration and carbohydrate content of digesta analysed . Mean water content decreased from 5.30 +/- 0.99 to 2.51 +/- 0.13 mf/g dry mass in the first 5 m of the colon . Correspondingly, mean carbohydrate content fell from 529.85 +/- 46.61 to 434.99 +/- 29.89 mg/g dry mass . A significant correlation was shown between the decreases in mean carbohydrate and water content along the colon (r2 = 0.997, P < 0.05) . Changes in mean osmolality (+/- 10 mOsm/kg) and SCFA concentration (+/- 7 mmol/l) were minimal in comparison to the change in Na+ concentration (-54 mmol/l) . These findings reflect a close coupling between SCFA production and absorption on the one hand and water absorption on the other.

Prikl Biokhim Mikrobiol, 2003 Jan-Feb, 39(1), 74 - 84
{Fungal decomposition of oat straw during liquid and solid state fermentation}; Stepanova EV et al.; White rot fungi (Coriolus hirsutus, Coriolus zonatus, and Cerrena maxima from the collection of the Komarov Botanical Institute of the Russian Academy of Sciences) and filamentous fungi (Mycelia sterilia INBI 2-26 and Trichoderma reesei 6/16) were grown on oat straw-based liquid and solid media, as well as in a bench-scale reactor, either individually or as co-cultures . All fungi grew well on solid agar medium supplemented with powdered oat straw as the sole carbon source . Under these conditions, the mould Trichoderma reesei fully suppressed the growth of all basidiomycetes studied; conversely, Mycelia sterilia neither affected the development of any of the cultures, nor did it show any substantial susceptibility to suppression by their presence . Pure solid cultures of basidiomycetes, as well as the co-culture of Coriolus hirsutus and Cerrena maxima caused a notable bleaching of the oat straw during its consumption . When grown on the surface of oat straw-based liquid medium, the basidiomycetes consumed up to 40% polysaccharides without measurable lignin degradation (a concomitant process) . Under these conditions, Mycelia sterilia decomposed no more than 25% lignin in 60 days, but this was observed only after polysaccharide exhaustion and biomass accumulation . In contrast, during solid state straw fermentation, white rot fungi consumed up to 75% cellulose and 55% lignin in 83 days (C . zonarus), whereas the corresponding consumption levels for co-cultures of Mycelia sterilia and Trichoderma reesei equaled 70 and 45%, respectively (total loss of dry weight ranged from 55 to 60%) . Carbon dioxide-monitored solid-state fermentation of oat straw by the co-culture of filamentous fungi was successfully performed in an aerated bench-scale reactor.

Prikl Biokhim Mikrobiol, 2003 Jan-Feb, 39(1), 69 - 73
{Composition and biological activity of submerged mycelium of the xylotrophic basidiomycete Lentinus edodes}; Lobanok AG et al.; The composition of submerged mycelium of Lentinus edodes, grown in laboratory fermenters, has been studied . The mycelium contained 23-24% proteins, 8-9% lipids, up to 1800 mg% phenolic substances, and a significant amount of inorganic substances, including calcium and iron . The fungus produced up to 5.0% intracellular and 3.5-4.0 g/l extracellular polysaccharides . The submerged mycelium stimulated the development of humoral immune response elicited by sheep red blood cells.

Amino Acids, 2003, 24(1-2), 195 - 203
Production of cysteine for bacterial and plant biotechnology: application of cysteine feedback-insensitive isoforms of serine acetyltransferase; Wirtz M et al.; The first step of cysteine biosynthesis in bacteria and plants consists in the formation of O-acetylserine catalyzed by serine acetyltransferase (SAT) . SAT is highly sensitive to feedback inhibition by cysteine as part of the regulatory circuit of cysteine biosynthesis und thus hampers over-expression and fermentation of cysteine in biotechnological production processes . Since plants contain multiple SAT isoforms with different cysteine feedback sensitivity, this resource was exploited to demonstrate the suitability of plant SATs for the production of cysteine in both bacteria and plants . Three new cDNAs encoding SATs were isolated from Nicotiana tabacum . The catalytic activity of SAT4 was insensitive up to 0.6 mM cysteine . Expression of SAT4 in a newly constructed Escherichia coli host strain without endogenous SAT activity yielded a significant accumulation of cysteine in the culture medium compared to expression of cysteine sensitive SATs in the same strain . The application of a similarly insensitive SAT isoform from A . thaliana demonstrated the suitability of this approach to increase cysteine levels in transgenic tobacco plants.

Amino Acids, 2003, 24(1-2), 81 - 7
Branched chain amino acids as source of specific branched chain volatile fatty acids during the fermentation process of fish sauce; Sanceda NG et al.; The source of the formation of branched chain volatile fatty acids (VFA) in fish sauce was investigated . Certain branched VFA were derived from the degradation of specific amino acids as iso-butyric acid from valine and iso-valeric acid from leucine . Short and long straight chain VFA were significantly higher in the linoleic acid added sample than in the control but did not significantly bring changes to the branched chain VFA . It is suggested that straight chain VFA developed from fish fats . Alanine and isoleucine did not have a clear influence on the production of volatile fatty acids.

Appl Environ Microbiol, 2003 Mar, 69(3), 1858 - 60
An improved enrichment broth for isolation of Escherichia coli O157, with specific reference to starved cells, from radish sprouts; Sata S et al.; An enrichment broth was developed for the efficient isolation of Escherichia coli O157 from radish sprouts . The broth was buffered peptone water containing 0.5% sodium thioglycolate (STG-BPW), which was designed to allow growth of E . coli O157 in starved and unstarved states . However, this medium suppressed the growth of non-carbohydrate-fermenting obligate aerobes whose colonial appearance on sorbitol MacConkey agar containing cefixime and tellurite (CT-SMAC) resembled that of E . coli O157 . Both starved and unstarved cells of E . coli O157 experimentally inoculated into radish sprouts were successfully recovered with STG-BPW enrichment in all cases, most of which showed marked disappearance of E . coli O157-like colonies on CT-SMAC.

Nutrition, 2003 Mar, 19(3), 261 - 4
Dietary supplementation with fermented soybeans suppresses intimal thickening; Suzuki Y et al.; Although soy foods have been consumed for more than 1000 y, it is only in the past 20 y that they have made inroads into Western diets . We investigated the effect of dietary supplementation with natto extracts produced from fermented soybeans on intimal thickening of arteries after vessel endothelial denudation . Natto extracts include nattokinase, a potent fibrinolytic enzyme having four times greater fibrinolytic activity than plasmin . Intimal thickening was induced in the femoral arteries by intravenous infusion of rose bengal followed by focal irradiation with a transluminal green light . Dietary natto extract supplementation was started 3 wk before endothelial injury and continued for another 3 wk after . In ex vivo studies, euglobulin clot lysis times were measured 3 wk after the initial supplementation . Neointima formation and thickening were also initiated successfully . The intima media ratio 3 wk after endothelial injury was 0.15 +/- 0.03 in the control group . Dietary natto extract supplementation suppressed intimal thickening (0.06 +/- 0.01; P < 0.05) compared with the control group . Natto extracts shortened euglobulin clot lysis time, suggesting that their thrombolytic activities were enhanced . These findings suggest that natto extracts, because of their thrombolytic activity, suppress intimal thickening after vascular injury as a result of the inhibition of mural thrombi formation.

Mol Cell Biochem, 2003 Jan, 243(1-2), 107 - 12
Antiangiogenic effects of butyric acid involve inhibition of VEGF/KDR gene expression and endothelial cell proliferation; Gururaj AE et al.; The formation of new blood vessels from pre-existing ones is required for the growth of solid tumors and for metastasis . Interaction of tumor-secreted vascular endothelial growth factor (VEGF) with its receptor(s) on endothelial cells triggers endothelial cell proliferation and migration, which facilitate tumor angiogenesis . Butyric acid (BuA), a fermentation product of dietary fibers in the colon, is shown to alter gene expression and is postulated to be anticarcinogenic . The results presented in this paper indicate that BuA can be antiangiogenic in vivo by inhibiting angiogenesis in chorioallantoic membrane assay . BuA was not cytotoxic to endothelial cells but was a potent antiproliferative agent besides being proapoptotic to endothelial cells as verified by FACS analysis . Conditioned media from BuA-treated Ehrlich ascites tumor cells showed a 30% decrease in VEGF concentration when compared with untreated cells . The decrease in VEGF mRNA and its receptor, KDR mRNA levels in EAT and endothelial cells respectively, suggests that the VEGF-KDR system of angiogenesis is the molecular target for the antiangiogenic action of BuA.

Br J Cancer, 2003 Mar 10, 88(5), 748 - 53
Opposing effects of butyrate and bile acids on apoptosis of human colon adenoma cells: differential activation of PKC and MAP kinases; McMillan L et al.; Butyrate, produced in the colon by fermentation of dietary fibre, induces apoptosis in colon adenoma and cancer cell lines, which may contribute to protection against colorectal cancer . However, butyrate is present in the colon along with other dietary factors, including unconjugated bile acids, which are tumour promoters . We have shown previously that the proapoptotic effects of butyrate on AA/C1 human adenoma cells were reduced in the presence of bile acids . To determine the cellular basis of this interaction, we examined the effects of butyrate and the secondary bile acid ursodeoxycholic acid (UDCA) on signalling pathways known to regulate apoptosis using AA/C1 cells . Butyrate activated PKC-delta and p38 MAP (mitogen-activated protein) kinase, whereas UDCA activated PKC-alpha and p42/44 MAP kinase . Butyrate treatment also resulted in the caspase-3-mediated proteolysis of PKC-delta . Butyrate-induced apoptosis was reduced by inhibitors of PKC-delta (Rottlerin), p38 MAP kinase (SB202190) and caspase 3 (DEVD-fmk), whereas the proliferative/survival effects of UDCA were blocked by inhibitors of PKC-alpha (Go6976) and MEK 1 (PD98059) . The effects of butyrate and bile acids are therefore mediated by the differential activation of signalling pathways that are known to regulate apoptosis.

Nat Mater, 2002 Dec, 1(4), 236 - 40
Biosynthesis of novel thermoplastic polythioesters by engineered Escherichia coli; Lutke-Eversloh T et al.; The development of non-petrochemical sources for the plastics industry continues to progress as large multinationals focus on renewable resources to replace fossil carbon . Many bacteria are known to accumulate polyoxoesters as water-insoluble granules in the cytoplasm . The thermoplastic and/or elastomeric behaviour of these biodegradable polymers holds promise for the development of various technological applications . Here, we report the synthesis and characterization of microbial polythioesters (PTEs), a novel class of biopolymers of general technological relevance . Biosynthesis of PTE homopolymers was achieved using a recombinant strain of Escherichia coli that expressed a non-natural pathway consisting of a butyrate kinase, a phosphotransbutyrylase, and a PHA synthase . Different homopolymers were produced, consisting of either 3-mercaptopropionate, 3-mercaptobutyrate, or 3-mercaptovalerate repeating units, if the respective mercaptoalkanoic acids were provided as precursor substrates to the fermentative process . The PTEs contributed up to 30% (w/w) of the cellular dry weight and were identified as hydrophobic inclusions in the cytoplasm . The chemical and stereochemical homogeneity of the purified PTEs were identified by different methods, and the estimated physical properties were compared to the oxypolyester equivalents, revealing low crystalline order and, for the poly(3-mercaptopropionate) improved thermal stability . The ability to produce PTEs through a biosynthetic route opens up new avenues in the field of biomaterials.

Bioresour Technol, 2003 Jul, 88(3), 265 - 8
Improved arachidonic acids production from the fungus Mortierella alpina by glutamate supplementation; Yu LJ et al.; The effect of various concentrations of glutamate on arachidonic acid (AA) production from Mortierella alpina in shaker flask culture was studied . Glutamate supplementation promoted Mortierella growth, accelerated substrate metabolism, and increased AA production, and a concentration of 0.8 g/l glutamate resulted in the greatest AA yield (1.41 g/l) . In 10 l airlift stirred fermenter culture, AA yield in the cultures exposed to 0.8 g/l glutamate was also greater than that in the control (0.56 g/l).

J Agric Food Chem, 2003 Mar 12, 51(6), 1571 - 7
Protective effects of fermented filtrate from Antrodia camphorata in submerged culture against CCl4-induced hepatic toxicity in rats; Song TY et al.; The protective effects and the possible mechanisms of dry matter of fermented filtrate (DMF) from Antrodia camphorata in submerged culture (ACSC) on H(2)O(2)-induced cytotoxicity in HepG2 and carbon tetrachloride (CCl(4))-induced hepatotoxicity in Sprague-Dawley rats were investigated . The results showed that the inhibitory effect of DMF and its crude triterpenoids on lipid peroxidation occurred in a dose-response manner in an AAPH/linoleic acid system . When HepG2 cells were pretreated with DMF at the concentration of 0.10 mg/mL for 4 h and then induced by 1 h of treatment with H(2)O(2) (100 microM), lipid peroxidation was significantly (p < 0.05) decreased, as measured by the formation of malondialdehyde . The oral pretreatment with DMF {0.25 and 0.50 mg/kg of body weight (bw)} for 5 consecutive days prior to the administration of a single dose of 40% CCl(4) (0.10 mL/100 g of bw, ip) significantly prevented the increase in serum levels of hepatic enzyme markers (alanine and aspartate aminotransferase) and liver lipid peroxidation (p < 0.05) . Histopathological evaluation of the rat liver revealed that DMF reduced the incidence of liver lesions, including neutrophil infiltration, hydropic swelling, and necrosis induced by CCl(4) in rats . Moreover, reduced glutathione (GSH)-dependent enzymes (glutathione peroxidase, glutathione reductase, and glutathione S-transferase) and the GSH/GSSG ratio were significantly improved in the oral pretreatment DMF of rats (p < 0.01) . The results suggest that DMF may play a role in preventing oxidative damage in living systems by up-regulating hepatic GSH-dependent enzymes to preserve the normal GSH/GSSH ratio and scavenging free radicals formed during CCl(4) metabolism.

J Antibiot (Tokyo), 2002 Dec, 55(12), 1048 - 52
Selective production of fungal beauveriolide I or III by fermentation in amino acid-supplemented media; Namatame I et al.; Beauveriolides I and III, cyclic depsipeptides composed of L-Phe, L-Ala, D-Leu and (3S,4S)-3-hydroxy-4-methyloctanoic acid, and L-Phe, L-Ala, L-allo-Ile and (3S,4S)-3-hydroxy-4-methyloctanoic acid, respectively, were previously isolated from the culture broth of fungal Beauveria sp . FO-6979 as inhibitors of macrophage foam cell formation . To improve the production of these compounds by fermentation, the culture conditions were studied . The production of both beauveriolides was increased five to ten folds by fermentation in the culture media containing tryptone . Further study revealed that addition of L-Leu/L-Ile, but not D-Leu/D-allo-Ile, to the culture medium yielded a high and selective production of beauveriolide I or III . As a result, regardless of their separation difficulty due to the similar physico-chemical properties, a large amount of beauveriolide I or III was prepared from the culture broth obtained from L-Leu- or L-Ile-supplemented fermentation, respectively, by one step purification using silica gel column chromatography.

Metab Eng, 2002 Jul, 4(3), 230 - 7
The effect of increasing NADH availability on the redistribution of metabolic fluxes in Escherichia coli chemostat cultures; Berrios-Rivera SJ et al.; It is generally known that cofactors play a major role in the production of different fermentation products . This paper is part of a systematic study that investigates the potential of cofactor manipulations as a new tool for metabolic engineering . The NADH/NAD+ cofactor pair plays a major role in microbial catabolism, in which a carbon source, such as glucose, is oxidized using NAD+ and producing reducing equivalents in the form of NADH . It is crucially important for continued cell growth that NADH be oxidized to NAD+ and a redox balance be achieved . Under aerobic growth, oxygen is used as the final electron acceptor . While under anaerobic growth, and in the absence of an alternate oxidizing agent, the regeneration of NAD+ is achieved through fermentation by using NADH to reduce metabolic intermediates . Therefore, an increase in the availability of NADH is expected to have an effect on the metabolic distribution . We have previously investigated a genetic means of increasing the availability of intracellular NADH in vivo by regenerating NADH through the heterologous expression of an NAD(+)-dependent formate dehydrogenase and have demonstrated that this manipulation provoked a significant change in the final metabolite concentration pattern both anaerobically and aerobically (Berrios-Rivera et al., 2002, Metabolic engineering of Escherichia coli: increase of NADH availability by overexpressing an NAD(+)-dependent formate dehydrogenase, Metabolic Eng . 4, 217-229) . The current work explores further the effect of substituting the native cofactor-independent formate dehydrogenase (FDH) by an NAD(+)-dependent FDH from Candida boidinii on the NAD(H/+) levels, NADH/NAD+ ratio, metabolic fluxes and carbon-mole yields in Escherichia coli under anaerobic chemostat conditions . Overexpression of the NAD(+)-dependent FDH provoked a significant redistribution of both metabolic fluxes and carbon-mole yields . Under anaerobic chemostat conditions, NADH availability increased from 2 to 3 mol NADH/mol glucose consumed and the production of more reduced metabolites was favored, as evidenced by a dramatic increase in the ethanol to acetate ratio and a decrease in the flux to lactate . It was also found that the NADH/NAD+ ratio should not be used as a sole indicator of the oxidation state of the cell . Instead, the metabolic distribution, like the Et/Ac ratio, should also be considered because the turnover of NADH can be fast in an effort to achieve a redox balance.

Metab Eng, 2002 Jul, 4(3), 217 - 29
Metabolic engineering of Escherichia coli: increase of NADH availability by overexpressing an NAD(+)-dependent formate dehydrogenase; Berrios-Rivera SJ et al.; Metabolic engineering studies have generally focused on manipulating enzyme levels through either the amplification, addition, or deletion of a particular pathway . However, with cofactor-dependent production systems, once the enzyme levels are no longer limiting, cofactor availability and the ratio of the reduced to oxidized form of the cofactor can become limiting . Under these situations, cofactor manipulation may become crucial in order to further increase system productivity . 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 . However, cofactor manipulations can potentially become a powerful tool for metabolic engineering . Nicotinamide adenine dinucleotide (NAD) functions as a cofactor in over 300 oxidation-reduction reactions and regulates various enzymes and genetic processes . The NADH/NAD+ cofactor pair plays a major role in microbial catabolism, in which a carbon source, such as glucose, is oxidized using NAD+ producing reducing equivalents in the form of NADH . It is crucially important for continued cell growth that NADH be oxidized to NAD+ and a redox balance be achieved . Under aerobic growth, oxygen is used as the final electron acceptor . While under anaerobic growth, and in the absence of an alternate oxidizing agent, the regeneration of NAD+ is achieved through fermentation by using NADH to reduce metabolic intermediates . Therefore, an increase in the availability of NADH is expected to have an effect on the metabolic distribution . This paper investigates a genetic means of manipulating the availability of intracellular NADH in vivo by regenerating NADH through the heterologous expression of an NAD(+)-dependent formate dehydrogenase . More specifically, it explores the effect on the metabolic patterns in Escherichia coli under anaerobic and aerobic conditions of substituting the native cofactor-independent formate dehydrogenase (FDH) by and NAD(+)-dependent FDH from Candida boidinii . The over-expression of the NAD(+)-dependent FDH doubled the maximum yield of NADH from 2 to 4 mol NADH/mol glucose consumed, increased the final cell density, and provoked a significant change in the final metabolite concentration pattern both anaerobically and aerobically . Under anaerobic conditions, the production of more reduced metabolites was favored, as evidenced by a dramatic increase in the ethanol-to-acetate ratio . Even more interesting is the observation that during aerobic growth, the increased availability of NADH induced a shift to fermentation even in the presence of oxygen by stimulating pathways that are normally inactive under these conditions.

J Pharm Biomed Anal, 2003 Mar 10, 31(3), 571 - 7
Application of attenuated total reflectance Fourier transform infrared spectrometry to the determination of cephalosporin C in complex fermentation broths; Kargosha K et al.; An analytical procedure has been developed for quantitative determination of cephalosporin C in complex fermentation broths . The method is based on the partial least-square treatment of data obtained by attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectrometric method in the wavenumber range of 1227-1257 cm(-1) . Absorbance spectra were employed for measurement using a set of eleven binary aqueous standard solutions of cep halosporin C and deacetoxycephalosporin C . The method is simple, rapid and accurate (to within +/-1.49%) . The developed method has been used to measure cephalosporin C in aqueous solution of biosynthetic samples only after freeze drying the sample at -30 degrees C and dissolving it in distilled water.

Food Chem Toxicol, 2003 Apr, 41(4), 477 - 87
Evaluation of the toxicity of concentrated barley beta-glucan in a 28-day feeding study in Wistar rats; Delaney B et al.; Beta-glucans are water-soluble cell-wall polysaccharides consisting of (1-->3,1-->4)-linked beta-D-glucopyranosyl monomers that comprise a considerable proportion of soluble fiber from certain grains including oats and barley . Consumption of foods containing beta-glucan or beta-glucan-enriched fractions prepared from these grains lower serum cholesterol concentrations in humans and in animal models of hypercholesterolemia . The present study was conducted to evaluate the toxicity of beta-glucan-enriched soluble fiber from barley in Wistar rats on dietary administration at concentrations of 0.7, 3.5 and 7% beta-glucan for 28 days . There were no adverse effects on general condition and behavior, growth, feed and water consumption, feed conversion efficiency, red blood cell and clotting potential parameters, clinical chemistry values, and organ weights . Necropsy and histopathology findings revealed no treatment-related changes in any organ evaluated . A dose-dependent increase in full and empty cecum weight was observed . This is a common physiological response of rodents to high amounts of poorly digestible, fermentable carbohydrates, and was of no toxicological concern . The only finding of possible biological relevance was an increase in the number of circulating lymphocytes observed in males . However, the increase was not dose-dependent and was not observed in females . Results of this study demonstrated that consumption of concentrated barley beta-glucan was not associated with any obvious signs of toxicity in Wistar rats even following consumption of large quantities .

Bioorg Med Chem, 2003 Mar 20, 11(6), 1041 - 6
Purification and characterization of an alkaline lipase from a newly isolated Pseudomonas mendocina PK-12CS and chemoselective hydrolysis of fatty acid ester; Jinwal UK et al.; Lipase isolated from a soil isolate, Pseudomonas mendocina (PK-12CS) chemoselectively hydrolyzed the fatty ester group in presence of arbamate of compound 5-amino-2,4-dihydro-3H-1,2,4-triazole-3 ones, a class of compounds which are attractive starting materials for the synthesis of triazole annealed heterocycles . The enzymatic method provides an easy access to the synthesis of N-substituted glycine . Under optimized fermentation conditions the culture produced 3510 Lipolytic Units/mL of cell free fermentation broth in 20 h of fermentation . The purified lipase exhibited molecular mass of 80 kDa on SDS polyacrylamide gel electrophoresis . The enzyme was stable at room temperature for more than a month and expressed maximum activity at 37 degrees C and pH 8.

J Dairy Sci, 2003 Jan, 86(1), 324 - 30
Monensin by fat interactions on trans fatty acids in cultures of mixed ruminal microorganisms grown in continuous fermentors fed corn or barley; Jenkins TC et al.; In previous studies, monensin (M) and unsaturated plant oils independently increased trans fatty acid concentrations in cultures of mixed ruminal microorganisms . This study was conducted to determine if combining M with plant oil yielded interactions on trans fatty acid concentrations in cultures of mixed ruminal microorganisms or their effects were additive . Four continuous fermentors were fed 14 g of dry feed per day (divided equally between two feedings), consisting of alfalfa hay pellets (30% of DM) and either a high corn (HC) or a high barley (BB) concentrate (70% of DM) in each of two fermentors . Within each grain type, one fermentor was supplemented with M (25 ppm), and the other fermentor was supplemented with 5% soybean oil (SBO) during d 5 to 8 . Monensin and SBO were added together in all fermentors during d 9 to 12 . Samples were taken at 2 h after the morning feeding on the last day of each period and analyzed for fatty acids by gas-liquid chromatography . A second run of the fermentors followed the same treatment sequence to give additional replication . Average pH across all treatments was 6.15, which was reduced by M but not affected by SBO . Monensin reduced the ratio of acetate to propionate (A:P), which averaged 2.03 across all treatments; fat decreased A:P in cultures not receiving M but increased it in the presence of M . Monensin and SBO altered the concentration of several trans fatty acids, but the only interaction was a grain x M x SBO interaction for trans-10 C18:1 . The increase in trans-10 C18:1 by the M and SBO combination exceeded the sum of increases in trans-10 C18:1 for each individual feed additive, but only for KB . For the HC diet, M increased trans-10 C18:1 more than fat alone and more than the M and SBO combination . The results of this study show that M and SBO effects are additive for all trans FA except for trans-10 C18:1 . In the case of trans-10 C18:1, M and SBO interacted to give higher trans-10 C18:1 concentrations in ruminal contents than would be expected simply by adding their individual effects, but only for HB . Because some trans fatty acid isomers have been associated with milk fat depression in dairy cows, these results suggest more severe depressions in milk fat content when cows are fed M along with unsaturated plant oils.

J Dairy Sci, 2003 Jan, 86(1), 287 - 308
Corn silage management: effects of hybrid, maturity, inoculation, and mechanical processing on fermentation characteristics; Johnson LM et al.; Two experiments were conducted to evaluate the effects of hybrid, maturity, mechanical processing, and inoculation of corn silage on fermentation characteristics . In experiment 1, Pioneer hybrid 3845 corn silage was harvested at three maturities (hard dough, one-third milkline, two-thirds milkline) . In experiment 2, Pioneer hybrids 3845 and Quanta were harvested at three maturities (one-third milkline, two-thirds milkline, and blackline) . In both experiments, corn silage was harvested at each maturity with and without mechanical processing and with and without inoculation . In experiments 1 and 2, corn silage was harvested at a theoretical length-of-cut of 6.4 and 12.7 mm, respectively . Maturity at harvest tended to have a greater impact on silage fermentation characteristics of corn silage than mechanical processing and inoculation . In experiments 1 and 2, corn silage harvested at the earliest maturity tended to have decreased dry matter content and increased water-soluble carbohydrate concentrations during the ensiling process than corn silage harvested at advanced maturities . In experiment 2, pH levels were lower for corn silage harvested at the early maturity (one-third milkline) compared with advanced maturities (two-thirds milkline and blackline) by 57 d after ensiling . The difference in pH can be explained by the greater concentration of water-soluble carbohydrates at the early maturity (one-third ML) soon after ensiling (2, 3, 6 and 10 d after ensiling) compared with advanced maturities (two-thirds ML and BL) . The increased water-soluble carbohydrate concentrations in the less mature corn silage provided nutrients for bacteria to grow and produce primarily lactic acid (6, 10, and 57 d after ensiling) and some acetic acid (2, 3, 6, and 10 d after ensiling) which reduced the pH of corn silage more than at the advanced maturities . There was a slight change in silage fermentation characteristics when corn silage was inoculated with Pioneer 1132 inoculant in experiment 1 . The inoculated corn silage had increased temperature, lactate and acetate concentrations, and lower water-soluble carbohydrate and pH levels compared with uninoculated corn silage . Dry matter recovery tended to be greater for processed corn silage in experiment 1, and greater for unprocessed corn silage in experiment 2 . It appears that when fermentation was greater (increased temperature and lactate concentration 57 d after ensiling) the dry matter recovery was lower.

J Dairy Sci, 2003 Jan, 86(1), 195 - 207
Effects of diet fermentability on efficiency of microbial nitrogen production in lactating dairy cows; Oba M et al.; Effect of diet fermentability on efficiency of microbial N production was evaluated . Eight ruminally and duodenally cannulated Holstein cows (55 +/- 15.9 days in milk; mean +/- SD) were used in a duplicated 4 x 4 Latin square design with a 2 x 2 factorial arrangement of treatments . Experimental diets contained either ground high moisture corn (HM) or dry ground corn (DG) at two dietary starch concentrations (32 vs . 21%) . All diets were formulated for 18% CP, and the sources of dietary protein were alfalfa silage (50% of forage at DM basis), soybean meal, distillers grain, and blood meal . The amount of OM truly fermented in the rumen varied from 7.7 (DG at 21% dietary starch) to 11.3 kg/d (HM at 32% dietary starch) among treatments, and was greater for high starch diets and HM treatments compared with low starch diets and DG treatments, respectively . Microbial N flow was greater for high starch diets compared with low starch diets, but was not affected by corn grain treatment . Microbial efficiency was lower for HM compared with DG treatment (39.7 vs . 48.4 g of microbial N/kg of true ruminally degraded OM), but was not affected by dietary starch concentration . Microbial efficiency was positively correlated with rate of passage for OM and starch (r = 0.77 and 0.75, respectively) . Rapid passage rate may have decreased microbial turnover in the rumen, enhancing microbial efficiency . Microbial efficiency was negatively correlated with rate of starch digestion (r = -0.55), consistent with the energy spilling theory . However, energy spilling did not appear to be from lack of ammonia or low ruminal pH . Microbial efficiency was not related to ruminal ammonia concentration, daily mean ruminal pH, or minimum ruminal pH . Rate of starch availability and rates of passage for starch and OM from the rumen are important determinants of efficiency of microbial protein synthesis in vivo.

J Dairy Sci, 2003 Jan, 86(1), 184 - 94
Effects of corn grain conservation method on ruminal digestion kinetics for lactating dairy cows at two dietary starch concentrations; Oba M et al.; Effects of conservation method of corn grain and dietary starch concentration on ruminal digestion kinetics were evaluated . Eight ruminally and duodenally cannulated Holstein cows (55 +/- 15.9 days in milk; mean +/- SD) were used in a duplicated 4 x 4 Latin square design with a 2 x 2 factorial arrangement of treatments . Experimental diets contained either ground high moisture corn (HM) or dry ground corn (DG) at two dietary starch concentrations (32 vs . 21%) . Mean particle size and dry-matter concentration of corn grain were 1,863 microm and 63.2%, and 885 microm and 89.7%, for HM and DG, respectively . Starch digestibility in the rumen was greater for HM treatments compared with DG treatments, but starch digestibility in the total tract was not affected by conservation method of corn grain because of compensatory digestion in the intestines . The difference in ruminal starch digestibility between HM and DG treatment was greater for high-starch diets (71.1 vs . 46.9%) compared with low-starch diets (58.5 vs . 45.9%) . This interaction is attributed to a greater difference in first-order digestion rate of starch between HM and DG treatment in high-starch diets (28.2 vs . 14.6%/h) compared with low-starch diets (16.8 vs . 12.2%/h) . This suggests that ruminal starch digestion is a second-order reaction limited by enzyme activities as well as substrate availability; ruminal contents of cows fed low-starch diets may have insufficient amylolytic activity for maximal starch digestion when readily fermentable starch is available . Rate of neutral detergent fiber digestion in the rumen was slower for high-starch diets and HM treatments compared with low-starch diets and DG treatments, respectively . Effects of corn grain conservation method on ruminal digestion kinetics are greatly altered by starch concentration of diets.

J Dairy Sci, 2003 Jan, 86(1), 174 - 83
Effects of corn grain conservation method on feeding behavior and productivity of lactating dairy cows at two dietary starch concentrations; Oba M et al.; Effects of conservation method of corn grain and dietary starch concentration on dry matter intake (DMI) and productivity of lactating dairy cows were evaluated . Eight ruminally and duodenally cannulated Holstein cows (55 +/- 15.9 d in milk; mean +/- SD) were used in a duplicated 4 x 4 Latin square design with a 2 x 2 factorial arrangement of treatments . Experimental diets contained either ground high-moisture corn (HM) or dry ground corn (DG) at two dietary starch concentrations (32 vs 21%) . Mean particle size and dry matter (DM) concentration of corn grain were 1863 pm and 63.2%, and 885 microm and 89.7%, for HM and DG, respectively . DMI was lower for HM compared to DG treatment in high-starch diets (20.8 vs 22.5 kg/d), but similar for the HM and DG treatments in low-starch diets (19.7 vs 19.6 kg/d) . This reduction in DMI is attributed to smaller meal size for HM compared to DG in high-starch diets (1.9 vs 2.3 kg of DM for high-starch diets; 2.1 vs 2.0 kg of DM for low-starch diets) . Faster starch fermentation for HM in high-starch diets might result in satiety with smaller meal size . Milk yield was greater when cows were fed high-starch diets compared to low-starch diets (38.6 vs 33.9 kg/d) regardless of corn grain treatment . High-starch diets increased solids-corrected milk yield by 3.3 kg (35.2 vs 31.9 kg/d) compared to low-starch diets for cows fed DG, but did not increase for cows fed HM . This was because of a lower milk fat concentration for cows fed HM in high-starch diets . Reducing ruminal starch fermentation by substituting DG for HM can increase the productivity of lactating cows fed high-starch diets.

J Pharm Biomed Anal, 2003 Feb 26, 31(2), 291 - 8
LC determination of salinomycin in fermentation broths and premixes; Blazsek M et al.; A simple and rapid high performance liquid chromatography method for the determination of salinomycin in fermentation media of Streptomyces albus strains and in premixes has been developed . This method involves reverse-phase separation of the component analysed with UV detection at 210 nm using methanol and 0.2 M acetate buffer pH 5.8 (100:10, v/v) as the mobile phase . The reliability of the method was confirmed by validation . A linear relationship was obtained within range 0.2-2.0 mg ml(-1) (r=0.9999) . The relative standard deviation of methods within-laboratory reproducibility was 1.6% . The estimated quantitation limit of assay was about 32.5 microg ml(-1) . The method has been successfully used in the determination of salinomycin content in testing production processes and premixes of different commercial brands.

J Environ Sci (China), 2003 Jan, 15(1), 1 - 4
Production of laccase by Coriolus versicolor and its application in decolorization of dyestuffs: (I) . Production of laccase by batch and repeated-batch processes; Lin JP et al.; The production of laccase by Coriolus versicolor was studied . The effect of cultivation conditions on laccase production by Coriolus versicolor was examined to obtain optimal medium and cultivation conditions . Both batch and repeated-batch processes were performed for laccase production . In repeated-batch fermentation with self-immobilized mycelia, total of 14 cycles were performed with laccase activity in the range between 3.4 and 14.8 U/ml.

Vopr Virusol, 2003 Jan-Feb, 48(1), 9 - 14
{Analysis of hepatitis C virus proteins in hepatic cells of patients with chronic hepatitis C}; Masalova OV et al.; A correlation between the detection of proteins and an activity of the pathological process was analyzed in a study of the content of the C virus hepatitis (CVH) proteins in hepatic cells of patients with chronic C hepatitis (CCH) . The expression of CVH proteins in frozen sections of biopsy samples of 69 CCH patients was evaluated by using the immune-histological method involving original monoclonal antibodies (MCA) to 5 CVH proteins . The results of the detection of proteins in patients were compared with an activity and stage of CCH (by using histological tests and a level of alanine aminotransferase--AAT) . A set of the CVH proteins were found in the liver of 74% of patients, i.e . core proteins, NS3, NS4A, NS4B and NS5A--in 28, 43, 43, 55 and 58%, respectively . All studied proteins were detected in the cytoplasm of hepatocytes . Proteins were found in the liver more often as compared with the detection rate of CVH RNA in the blood serum (61%) . This demonstrates a high sensitivity of the discussed test at detecting the CVH infection . The accumulation of the core protein was shown to correlate with the presence of the replicative form of CVH RNA in the liver and with a higher level of AAT . The quantity of NS5 A-expressing cells correlated directly with a CCH stage . The quantity of NSB- and NS3-positive hepatocytes correlated negatively with an activity of the inflammatory-and-necrotic processes in the liver . Hyper-fermentation was found more often among the antigen-positive patients . The CCH histological activity was proven to be reliably higher at a simultaneous detection of CCH proteins in the liver and of CVH RNA--in the serum.

J Biol Chem, 2003 May 9, 278(19), 17203 - 9 Epub 2003 Feb 26.
Functional characterization and localization of acetyl-CoA hydrolase, Ach1p, in Saccharomyces cerevisiae; Buu LM et al.; Acetyl-CoA hydrolase (Ach1p), catalyzing the hydrolysis of acetyl-CoA, is presumably involved in regulating intracellular acetyl-CoA or CoASH pools; however, its intracellular functions and distribution remain to be established . Using site-directed mutagenesis analysis, we demonstrated that the enzymatic activity of Ach1p is dependent upon its putative acetyl-CoA binding sites . The ach1 mutant causes a growth defect in acetate but not in other non-fermentable carbon sources, suggesting that Ach1p is not involved in mitochondrial biogenesis . Overexpression of Ach1p, but not constructs containing acetyl-CoA binding site mutations, in ach1-1 complemented the defect of acetate utilization . By subcellular fractionation, most of the Ach1p in yeast was distributed with mitochondria and little Ach1p in the cytoplasm . By immunofluorescence microscopy, we show that Ach1p and acetyl-CoA binding site-mutated constructs, but not its N-terminal deleted construct, are localized in mitochondria . Moreover, the onset of pseudohyphal development in homozygote ach1-1 diploids was abolished . We infer that Ach1p may be involved in a novel acetyl-CoA biogenesis and/or acetate utilization in mitochondria and thereby indirectly affect pseudohyphal development in yeast.

Am J Physiol Gastrointest Liver Physiol, 2003 Jul, 285(1), G45 - 53 Epub 2003 Feb 26.
Anion-dependent Mg2+ influx and a role for a vacuolar H+-ATPase in sheep ruminal epithelial cells; Schweigel M et al.; The K+-insensitive component of Mg2+ influx in primary culture of ruminal epithelial cells (REC) was examined by means of fluorescence techniques . The effects of extracellular anions, ruminal fermentation products, and transport inhibitors on the intracellular free Mg2+ concentration ({Mg2+}i), Mg2+ uptake, and intracellular pH were determined . Under control conditions (HEPES-buffered high-NaCl medium), the {Mg2+}i of REC increased from 0.56 +/- 0.14 to 0.76 +/- 0.06 mM, corresponding to a Mg2+ uptake rate of 15 microM/min . Exposure to butyrate did not affect Mg2+ uptake, but it was stimulated (by 84 +/- 19%) in the presence of CO2/HCO(-)3 . In contrast, Mg2+ uptake was strongly diminished if REC were suspended in HCO(-)3-buffered high-KCl medium (22.3 +/- 4 microM/min) rather than in HEPES-buffered KCl medium (37.5 +/- 6 microM/min) . After switching from high- to low-Cl- solution, {Mg2+}i was reduced from 0.64 +/- 0.09 to 0.32 +/- 0.16 mM and the CO2/HCO(-)3-stimulated Mg2+ uptake was completely inhibited . Bumetanide and furosemide blocked the rate of Mg2+ uptake by 64 and 40%, respectively . Specific blockers of vacuolar H+-ATPase reduced the {Mg2+}i (36%) and Mg2+ influx (38%) into REC . We interpret this data to mean that the K+-insensitive Mg2+ influx into REC is mediated by a cotransport of Mg2+ and Cl- and is energized by an H+-ATPase . The stimulation of Mg2+ transport by ruminal fermentation products may result from a modulation of the H+-ATPase activity.

J Biol Inorg Chem, 2003 Apr, 8(4), 469 - 74 Epub 2003 Feb 19.
Characterization of a {2Fe-2S} protein encoded in the iron-hydrogenase operon of Thermotoga maritima; Pan G et al.; Thermotoga maritima grows optimally at 80 degrees C by fermenting carbohydrates to organic acids, CO(2), and H(2) . The production of H(2) is catalyzed by a cytoplasmic, heterotrimeric (alphabetagamma) Fe-hydrogenase . This is encoded by three genes, hydC (gamma), hydB (beta) and hydA (alpha), organized within a single operon that contains five additional open reading frames (ORFs) . The recombinant form of the first ORF of the operon, TM1420, was produced in Escherichia coli . It has a molecular mass of 8537+/-3 Da as determined by mass spectrometry, in agreement with the predicted amino acid sequence . Purified TM1420 is red in color, has a basic p I (8.8), and contains 1.9 Fe atoms/mol that are present as a single {2Fe-2S} cluster, as determined by UV-visible absorption and EPR spectroscopy . The protein contains five cysteine residues, but their arrangement is characteristic of a subunit or domain rather than of a ferredoxin-type protein . The reduction potential of the {2Fe-2S} cluster (-233 mV at pH 6.5 and 25 degrees C) is pH independent but decreases linearly with temperature to -296 mV (-1.15 mV/ degrees C) at 80 degrees C . TM1420 is not reduced, in vitro, by the Fe-hydrogenase nor by a pyruvate ferredoxin oxidoreductase . The protein was unstable at 70 degrees C under anaerobic conditions with a half-life of approximately 30 min . The basic nature of TM1420, its instability at the growth temperature of T . maritima, and the unusual spacing of its cysteine residues suggest that this protein does not function as a ferredoxin-type electron carrier for the Fe-hydrogenase . Instead, TM1420 is more likely part of a thermostable multi-protein complex that is involved in metal cluster assembly of the hydrogenase holoenzyme.

J Pediatr Gastroenterol Nutr, 2003 Mar, 36(3), 329 - 37
Nondigestible carbohydrates in the diets of infants and young children: a commentary by the ESPGHAN Committee on Nutrition; Aggett PJ et al.; The consumption of nondigestible carbohydrates is perceived as beneficial by health professionals and the general public, but the translation of this information into dietary practice, public health recommendations, and regulatory policy has proved difficult . Nondigestible carbohydrates are a heterogeneous entity, and their definition is problematic . Without a means to characterize the dietary components associated with particular health benefits, specific attributions of these cannot be made . Food labeling for "fiber" constituents can be given only in a general context, and the development of health policy, dietary advice, and education, and informed public understanding of nondigestible carbohydrates are limited . There have, however, been several important developments in our thinking about nondigestible carbohydrates during the past few years . The concept of fiber has expanded to include a range of nondigestible carbohydrates . Their fermentation, fate, and effects in the colon have become a defining characteristic; human milk, hitherto regarded as devoid of nondigestible carbohydrates, is now recognized as a source for infants, and the inclusion of nondigestible carbohydrates in the diet has been promoted for their "prebiotic" effects . Therefore, a review of the importance of nondigestible carbohydrates in the diets of infants and young children is timely . The aims of this commentary are to clarify the current definitions of nondigestible carbohydrates, to review published evidence for their biochemical, physiologic, nutritional, and clinical effects, and to discuss issues involved in defining dietary guidelines for infants and young children.

Intern Med J, 2003 Mar, 33(3), 103 - 9
Rapamycin in cardiovascular medicine; Ruygrok PN et al.; The cellular action of rapamycin (sirolimus), a natural fermentation product produced by Streptomyces hygroscopicus, is mediated by binding to the FK506 binding protein . By inhibiting a kinase known as the target of rapamycin, it restricts the proliferation of smooth-muscle cells by blocking cell-cycle progression at the G1/S transition . The finding that rapamycin possesses both anti-proliferative and antimigratory activity suggests that it could contribute to the control of arterial re-narrowing after percutaneous intervention and control the vascular manifestations of chronic rejection in transplanted hearts . The first clinical trials of implantation of rapamycin- coated stents in obstructive coronary artery lesions have been reported and, in selected patient groups, it appears that the restenosis process has been abolished . Studies are underway to establish the benefits of rapamycin-coated stents in day-to-day interventional practice, including small vessels, long lesions and patients with multivessel disease . With the addition of novel antiplatelet agents and delivery systems, it is possible that the two major limitations of percutaneous coronary intervention - restenosis and stent thrombosis - will be overcome . Cardiac graft loss due to intimal hyperplasia and accelerated atherosclerosis remains the major limitation to long-term survival following cardiac transplantation . Animal studies of rapamycin have suggested that this process can be reduced or abolished . Human studies of the efficacy of rapamycin in preventing both acute rejection and allograft arterial disease are in progress . Concerns regarding toxicity, carcinogenicity, delayed healing and endothelialization remain . As with any new agent or technology, we must remain vigilant to late adverse side-effects.

Appl Biochem Biotechnol, 2003 Feb, 104(2), 141 - 8
Corn-milling pretreatment with anhydrous ammonia; Taylor F et al.; Exposure to anhydrous ammonia has been suggested as a pretreatment for corn milling . Batches of corn were exposed to ammonia under controlled conditions . The amounts of ammonia absorbed and reacted with the corn were measured . The amounts were not more than are needed as nutritional supplement for yeast fermentation to ethanol . Loosening of the hull was observed qualitatively, and subsequent shearing in a disk mill followed by steeping for 2, 4, 6, or 8 h showed that germ could be recovered at higher yield and after a shorter steeping time compared to untreated control batches . Quality of oil was not affected by treatment with ammonia.

J Environ Sci (China), 2003 Jan, 15(1), 5 - 8
Production of laccase by Coriolus versicolor and its application in decolorization of dyestuffs: (II) . Decolorization of dyes by laccase containing fermentation broth with or without self-immobilized mycelia; Lin JP et al.; The capability of decolorization for commercial dyes by Coriolus versicolor fermentation broth containing laccase with or without immobilized mycelium was evaluated . With cell-free fermentation broth containing laccase, high decolorization ratio was achieved foracid orange 7, but not for the other dyes concerned . The immobilized mycelium was proved to be more efficient than the cell-free system . All the four dyestuffs studied were found being decolourized with certain extent by immobilized mycelium . The repeated-batch decolorization was carried out with satisfactory results . The experimental data showed that the continuous decolorization of wastewater from a printing and dyeing industry was possible by using the self-immobilized C . versicolor.

Org Lett, 2002 Oct 31, 4(22), 3823 - 5
Toward a stable apoptolidin derivative: identification of isoapoptolidin and selective deglycosylation of apoptolidin; Pennington JD et al.; {formula: see text} Isoapoptolidin was isolated from crude fermentation extracts of the apoptolidin-producing microorganism Nocardiopsis sp . Apoptolidin isomerizes to isoapoptolidin upon treatment with methanolic triethylamine to establish a 1.4:1 equilibrium mixture of isoapoptolidin and apoptolidin . Semisynthesis of a peracetylated and deglycosylated derivative of apoptolidin is also described.

Biotechnol Bioeng, 2003 May 5, 82(3), 359 - 69
Engineering the aveC gene to enhance the ratio of doramectin to its CHC-B2 analogue produced in Streptomyces avermitilis; Stutzman-Engwall K et al.; Avermectin and its analogues are produced by the actinomycete Streptomyces avermitilis and are major commercial products for parasite control in the fields of animal health, agriculture, and human infections . Historically, the avermectin analogue doramectin (CHC-B1), which is sold commercially as Dectomax is co-produced during fermentation with the undesired analogue CHC-B2 at a CHC-B2:CHC-B1 ratio of 1.6:1 . Although the identification of the avermectin gene cluster has allowed for characterization of most of the biosynthetic pathway, the mechanism for determining the avermectin B2:B1 ratio remains unclear . The aveC gene, which has an essential role in avermectin biosynthesis, was inactivated by insertional inactivation and mutated by site-specific mutagenesis and error-prone PCR . Several unrelated mutations were identified that resulted in improved ratios of the desirable avermectin analogue CHC-B1, produced relative to the undesired CHC-B2 fermentation component . High-throughput (HTP) screening of cultures grown on solid-phase fermentation plates and analysis using electrospray mass spectrometry was implemented to significantly increase screening capability . An aveC gene with mutations that result in a 4-fold improvement in the ratio of doramectin to CHC-B2 was identified . Subsequent integration of the enhanced aveC gene into the chromosome of the S . avermitilis production strain demonstrates the successful engineering of a specific biosynthetic pathway gene to significantly improve fermentation productivity of a commercially important product .

Water Res, 2003 Mar, 37(6), 1433 - 41
Scale-up of anaerobic digestion of the biowaste fraction from domestic wastes; Gallert C et al.; In the City of Karlsruhe/Germany anaerobic digestion of 7200 ta(-1) of separately collected biowaste has proven its feasibility at an organic loading rate (OLR) of up to 8.5 kg CODm(-3)d(-1) . An extension of biowaste collection over the whole city area would increase the amount of biowaste to 12,000 ta(-1), leading to an OLR of the existing anaerobic reactor of up to 15 kg CODm(-3)d(-1) . To test, whether the increased amount of biowaste could be stabilized in the existing plant, biowaste suspensions were digested in a laboratory reactor at a maximum OLR, that exceeded the future OLR of the full-scale plant.The laboratory reactor was started with effluent of the full-scale biowaste digester . Like in full-scale, biowaste suspension from the hydropulper was added in a fed-batch mode . The elimination of organic material (measured as COD, chemical oxygen demand) and the volumetric gas production were linearly increasing with the OLR from 4.3 to 19 kg CODm(-3)d(-1) . Thus, safe operation of the full-scale plant at an OLR of 15 kg CODm(-3)d(-1) should be possible, leaving still some reserve capacity.To determine the metabolic reserves for fatty acid degradation during digestion at an OLR of 10 kg CODm(-3)d(-1), digester effluent was supplemented with either 40 mmoll(-1) acetate, propionate, i-butyrate or n-butyrate . Results of these batch assays indicated a rapid degradation of all fatty acids and fatty acid conversion rates, that would allow a stable anaerobic fermentation at 15 kg CODm(-3)d(-1)OLR.On the basis of the laboratory results the OLR of the full-scale methane reactor was increased to 15 kg CODm(-3)d(-1) . After 7 months, results of full-scale digestion were still consistent with the previously obtained laboratory results.

J Environ Biol, 2002 Jul, 23(3), 271 - 4
Bacteriological analysis of a pond water from Gobardanga, West Bengal; Tarafdar B et al.; The present communication deals with the bacteriological examination of a pond water, which is being used as one of the sources of drinking water, by multiple tube fermentation test to detect the coliform bacteria . The test was performed sequentially in three stages: presumptive, confirmed and completed coliform test . The results show that as the most probable number (MPN) index per 100 ml is 23, that pond water should be treated as nonpotable.

J Anim Sci, 2003 Jan, 81(1), 329 - 36
Effects of urea infusion and ruminal degradable protein concentration on microbial growth, digestibility, and fermentation in continuous culture; Griswold KE et al.; The effects of urea and rumen-degradable protein (RDP) on microbial growth, digestibility, and fermentation were examined using dual-flow continuous culture . The experimental design was a 4 x 4 Latin square with a 2 x 2 factorial arrangement of treatments . Factors were urea infusion (0.4 g/L of artificial saliva) and RDP concentration, and the treatments were as follows: 1) low RDP (8% of dietary dry matter) without urea (LDNU), 2) high RDP (11% of dietary dry matter) without urea (HDNU), 3) low RDP (8% of dietary dry matter) with urea (LDU), and 4) high RDP (11% of dietary dry matter) with urea (HDU) . The LDNU (i.e., negative control) and HDNU treatments were formulated to be nitrogen limiting . Results indicated that infusion of urea increased all digestibility measurements (P < 0.05), which in turn increased (P < 0.05) volatile fatty acid, NH3 nitrogen, trichloroacetic acid-soluble nitrogen, and soluble protein concentrations . Increasing dietary RDP improved dry matter and organic matter digestibility (P < 0.05) but did not alter acid detergent fiber or nonfiber carbohydrate digestibilities (P > 0.05) . Isobutyrate concentration decreased (P = 0.05) with increased RDP . Increased dietary RDP increased crude protein degradation and soluble protein concentration (P < 0.05), but NH3 nitrogen, trichloroacetic acid-soluble nitrogen, and peptide nitrogen were unaffected by changing RDP levels . Microbial growth efficiency was 19.9, 24.9, 28.0, and 32.2 g N/g organic matter truly digested for LDNU, HDNU, LDU, and HDU, respectively, and was significantly improved both by urea infusion (P = 0.002) and increased RDP concentration (P = 0.021) . The interactions of urea and RDP (P < 0.05) were explained by the high digestibility of neutral detergent fiber, nonstructural carbohydrate, and especially hemicellulose, with the HDNU treatment . The results of this study indicated that hemicellulose-degrading bacteria were able to effectively compete with nonstructural carbohydrate-degrading bacteria for available peptide and amino acid nitrogen . Further, the extent of protein degradation was dependent on the availability of NH3 nitrogen in the system.

J Anim Sci, 2003 Jan, 81(1), 323 - 8
Effects of 9,10 anthraquinone on ruminal fermentation, total-tract digestion, and blood metabolite concentrations in sheep; Kung L Jr et al.; The objective of this study was to evaluate the effects of adding 9,10 anthraquinone, a known inhibitor of methanogenesis and sulfate reduction, on blood metabolites, digestibility, and distribution of gas in sheep . In all experiments, we fed a complete pelleted diet that contained 17.5% crude protein and 24.5% acid detergent fiber . In an 8-wk study, feeding up to 66 ppm (dry matter basis) of 9,10 anthraquinone had no adverse effects on blood metabolites including indicators of normal enzyme function, mineral concentrations, and hematological measurements . Feeding 9,10 anthraquinone had no effect on average daily gain, although sheep fed a diet containing 66 ppm of 9,10 anthraquinone numerically gained the least weight . The ruminal molar proportions of acetic acid were decreased (P < 0.05) and the molar proportions of propionic acid were increased (P < 0.05) in sheep fed 1.5 and 66 ppm 9,10 anthraquinone when compared to those fed an unsupplemented diet . In a digestion trial, 9,10 anthraquinone (33 and 66 ppm) had no effect on the apparent digestion of nutrients in the total gastrointestinal tract . In a metabolism study, ruminal gasses were collected by rumenocentesis and analyzed for methane and hydrogen concentrations . Feeding 500 ppm of 9,10 anthraquinone to sheep resulted in a decrease (P < 0.07) in the concentration of methane, but an increase (P < 0.05) in hydrogen concentration of ruminal gas throughout the 19 d of feeding . There was no indication of ruminal adaptation throughout this time . These results are the first to show that 9,10 anthraquinone can partially inhibit in vivo rumen methanogenesis, which supports previous in vitro findings . In addition, at the concentrations used in this study, 9,10 anthraquinone was not toxic to ruminants.

J Anim Sci, 2003 Jan, 81(1), 182 - 90
Effects of dietary fermentable carbohydrates on behavior and heat production in group-housed sows; Rijnen MM et al.; The effects of dietary nonstarch polysaccharides (NSP) on behavior and heat production in group-housed sows were studied . Twelve groups of six nonpregnant sows were fed one of four experimental diets that were similar in composition except for starch and NSP contents . Exchanging sugar beet pulp silage (SBPS) for tapioca created the difference in dietary starch and NSP ratio . On a dry matter (DM) basis, diets contained 0, 10, 20, or 30% SBPS . Sows were group-housed . Intake of fermentable NSP (fNSP) for diets containing 0, 10, 20, or 30% SBPS averaged 7.06, 9.18, 11.61, and 13.73 g x kg(-0.75) d(-1), respectively . Sows were fed, once a day at 0800 . Dry matter intake for diets containing 0, 10, 20, or 30% SBPS, averaged 38.05, 38.38, 38.53, and 38.35 g x kg(-075) x d(-1), respectively, and ME intake averaged 523, 518, 514, and 493 kJ x kg(-0.75) x d(-1), respectively . On average, sows spent 177 min/d on physical activity, of which 8.8% was spent on eating . Time spent in physical activity was affected by diet (P = 0.005) . Sows fed 0 or 10% SBPS spent more time on physical activity than sows fed 20 or 30% SBPS (P = 0.002) . Energy cost of physical activity averaged 464 kJ x kg(-0.75) x d(-1) (standard estimated mean of 31) and was similar for diets (P = 0.679) . Total heat production (HP) and activity-related heat production (AHP) were affected by diet (P < 0.05) . Sows tended to be quieter when fNSP intake increased (P = 0.063) . The effect of fNSP intake on HP and AHP was not constant during the day . During the night period, fNSP intake did not affect HP and AHP (P > 0.10) . During the day period, increased fNSP intake decreased HP (P = 0.006) and tended to decrease AHP (P = 0.062) . During eating, increased fNSP intake increased HP (P = 0.012) and tended to increase AHP (P = 0.074) . Despite similar DMI, sows fed 0 or 10% SBPS spent less time eating than sows fed 20 or 30% SBPS (P = 0.009) . Feed consumption rate was higher (P = 0.003) in groups fed 0 or 10% SBPS than in groups fed 20 or 30% SBPS . Feed consumption rate decreased by 0.19 g DM x kg(-0.75) . min(-1) (P = 0.003) for each gram of fNSP intake . The energy saving effect of physical activity on the NE value of fNSP from SBPS ranged between 2.3 and 3.7 kJ/g of fNSP intake . In conclusion, intake of fNSP from SBPS affected energy expenditure for physical activity (P = 0.063); however, this effect was not constant during the day.

J Expo Anal Environ Epidemiol, 2003 Jan, 13(1), 66 - 73
Fluoride concentrations in three types of commercially packed tea drinks in Taiwan; Lung SC et al.; Tea is a popular drink around the world . It is also one of the major sources of fluoride intake . The objectives of this study were to assess fluoride concentrations in popular non-, semi-, and full-fermented tea drinks sold on the Taiwan market . Concentration differences among three types of commercially available tea drinks (tea leaf, tea bag, and packaged tea beverage) were explored . Several influential factors in intake concentrations were evaluated . The acute threshold intake (ATI) and allowable daily intake (ADI) of those tea drinks were also estimated . For each commercial type, samples from the most popular tea in one particular fermentation degree (non, semi, and full) were randomly purchased and analyzed for fluoride concentrations . Fluoride levels in different rounds of tea, in different containers, and with different ratios of water and tea leaf were also assessed . In total, 132 tea samples were analyzed . The mean fluoride concentrations in leaf tea without the first round, leaf tea with the first round, bagged tea, and packaged tea were 7.04, 7.79, 5.37, and 25.7 mg/l, respectively . Most of the intake concentrations in those samples exceeded 4 mg/l F, the lower bound of fluoride levels reported in the literatures to be associated with a lower IQ in children and a higher risk of bone fracture . Fluoride concentrations in packaged tea were the highest among the three types of commercially available tea . For studied leaf and bagged tea, almost a constant amount of fluoride was infused from the same amount of tea leaf regardless of the water volume . Besides this, making tea with glass or pottery tea makers would not affect fluoride intake concentrations . Acute intoxication is unlikely to occur . However, tea lovers in high fluoride content areas shall consider limit their consumption of tea drinks to avoid potential chronic effects.

Philos Trans R Soc Lond B Biol Sci, 2003 Jan 29, 358(1429), 205 - 13; discussion 213-5
Biochemical and evolutionary aspects of anaerobically functioning mitochondria; van Hellemond JJ et al.; Mitochondria are usually considered to be the powerhouses of the cell and to be responsible for the aerobic production of ATP . However, many eukaryotic organisms are known to possess anaerobically functioning mitochondria, which differ significantly from classical aerobically functioning mitochondria . Recently, functional and phylogenetic studies on some enzymes involved clearly indicated an unexpected evolutionary relationship between these anaerobically functioning mitochondria and the classical aerobic type . Mitochondria evolved by an endosymbiotic event between an anaerobically functioning archaebacterial host and an aerobic alpha-proteobacterium . However, true anaerobically functioning mitochondria, such as found in parasitic helminths and some lower marine organisms, most likely did not originate directly from the pluripotent ancestral mitochondrion, but arose later in evolution from the aerobic type of mitochondria after these were already adapted to an aerobic way of life by losing their anaerobic capacities . This review will focus on some biochemical and evolutionary aspects of these fermentative mitochondria, with special attention to fumarate reductase, the synthesis of the rhodoquinone involved, and the enzymes involved in acetate production (acetate : succinate CoA-transferase and succinyl CoA-synthetase).

Berl Munch Tierarztl Wochenschr, 2003 Jan-Feb, 116(1-2), 50 - 4
Effect of dolomite, magnesium oxide (MgO) and chalk (CaCO3) on in vitro fermentation of amorphous and crystalline cellulose and meadow hay using inoculum from sheep; Varadyova Z et al.; Some minerals can influence some biochemical parameters of rumen fermentation . The objective of this experiment was to determine the effect of different amounts (0.1, 0.25 and 0.5 g) of dolomite and to compare the effect of dolomite, magnesium oxide (MgO) and chalk (CaCO3) upon the end products of rumen fermentation in vitro . Amorphous and crystalline cellulose as well as meadow hay were used as substrates and incubated with buffered rumen fluid in sealed fermentation bottles . In dependence on the amount of dolomite and the kind of substrate an inhibitory effect of dolomite on methane production was evident . Significant differences of methane production were found between the controls, crystalline cellulose and meadow hay with 0.5 g of dolomite . An increase of total gas production was observed for cellulose with both 0.25 and 0.5 g of dolomite and also for meadow hay with 0.5 g of dolomite . It can be concluded that there was a remarkable effect of dolomite on methane production and also a slight effect of magnesium oxide and chalk as compared to the effect of dolomite on the fermentation parameters of incubated substrates.

J Dent Hyg, 2002 Fall, 76(4), 276 - 85
Xylitol for caries prevention; Peldyak J et al.; Xyitol is a naturally occurring sweetener which is essentially not fermentable by the caries-inducive oral microflora . When tested as a sucrose replacer, or even as a small dietary addition, systematic xylitol use leads to impressive reductions in caries incidence . Xylitol is compatible and complementary with all current oral hygiene recommendations . The appealing sensory and functional properties of xylitol facilitate a wide array of applications that promote oral health.

J Agric Food Chem, 2003 Feb 26, 51(5), 1336 - 40
Biotransformation of an organochlorine insecticide, endosulfan, by Anabaena species; Lee SE et al.; This study assesses the role of the blue-green algal species present in the soil in the dissipation of endosulfan and its metabolites in the soil environment . Two Anabaena species, Anabaena sp . PCC 7120 and Anabaena flos-aquae, were used in this study . Anabaena sp . PCC 7120 produced three principal biotransformation compounds, chiefly endosulfan diol (endodiol), and minor amounts of endosulfan hydroxyether and endosulfan lactone . Trace amounts of endosulfan sulfate were detected . In comparison, the biotransformation of endosulfan by Anabaena flos-aquae yielded mainly endodiol with minor amounts of endosulfan sulfate . An unknown compound was produced up to 70% from endosulfan spiked in the medium inoculated by A . flos-aquae after 8 days of incubation . Therefore, the endosulfan fate was dependent on the species . Within 1 day of incubation, two Anabaena species produced low amounts of beta-endosulfan after application of alpha-endosulfan . These results suggest the presence of isomerase in the Anabaena species . Further studies using a fermentor to control the medium pH at 7.2 to minimize chemical hydrolysis of endosulfan revealed a major production of endodiol with minor amounts of endosulfan sulfate and the unknown compound . These results showed that the production of the unknown compound might be dependent on the alkaline pH in the medium and that the production of endodiol by A . flos-aquae might be biologically controlled . This study showed that two algal species could contribute in the detoxification pathways of endosulfan in the soil environment.

J Agric Food Chem, 2003 Feb 26, 51(5), 1302 - 6
Color characteristics of monascus pigments derived by fermentation with various amino acids; Jung H et al.; Various pigment colors were produced by Monascus fermentations with separate addition of 20 amino acids . The color characteristics and structures of the pigment derivatives were investigated . When each amino acid was added to the fermentation broth as a precursor, pigment extracts with different hue and chroma values were obtained depending on the content ratios of yellow, orange, and red colors in the fermentation broth . The yellow and orange pigments were identical regardless of amino acid addition . The red compounds varied on the basis of the type of amino acid added . LC-MS and (1)H and (13)C NMR structural analyses confirmed that the derivative pigments contained the moieties of the added amino acids . L, a, and b values of the CIELAB color system for the derivative pigments were measured . Values of hue and chroma were then calculated . The colors of the derivative pigments were in the range of orangish red to violet red . The hydrophilicities/hydrophobicities of the derivative pigments could be predicted from their log P values, which were estimated using computer programs.

Res Vet Sci, 2003 Apr, 74(2), 113 - 8
Identification and quantification of amines in the equine caecum; Bailey SR et al.; Acute laminitis has been associated with the release of compounds, as yet unidentified, produced by hindgut fermentation which affect blood flow to the digit . The objectives of this study were to identify amine compounds in equine caecal and colonic contents, some of which are known to have vasoactive properties . In addition, the concentrations of amines in caecal contents of horses fed either grass or hay diets were compared . Fifteen amines were identified in equine hindgut contents in concentrations greater than 1 microM . The caecal concentrations of phenylethylamine, isoamylamine, cadaverine, diaminoheptane and spermidine were significantly higher in horses on spring/summer grass compared with those on winter grass or hay . These data show that many amines are present in the equine hindgut, some of which may have the potential to cause peripheral vasoconstriction if released into the circulation from the gastrointestinal tract.

Biol Bull, 2003 Feb, 204(1), 1 - 9
Iron-dependent hydrogenases of Entamoeba histolytica and Giardia lamblia: activity of the recombinant entamoebic enzyme and evidence for lateral gene transfer; Nixon JE et al.; Entamoeba histolytica and Spironucleus barkhanus have genes that encode short iron-dependent hydrogenases (Fe-hydrogenases), even though these protists lack hydrogenosomes . To understand better the biochemistry of the protist Fe-hydrogenases, we prepared a recombinant E . histolytica short Fe-hydrogenase and measured its activity in vitro . A Giardia lamblia gene encoding a short Fe-hydrogenase was identified from shotgun genomic sequences, and RT-PCR showed that cultured entamoebas and giardias transcribe short Fe-hydrogenase mRNAs . A second E . histolytica gene, which encoded a long Fe-hydrogenase, was identified from shotgun genomic sequences . Phylogenetic analyses suggested that the short Fe-hydrogenase genes of entamoeba and diplomonads share a common ancestor, while the long Fe-hydrogenase gene of entamoeba appears to have been laterally transferred from a bacterium . These results are discussed in the context of competing ideas for the origins of genes encoding fermentation enzymes of these protists.

Nutr Cancer, 2002, 43(2), 214 - 26
Growth inhibition and induction of apoptosis in MCF-7 breast cancer cells by fermented soy milk; Chang WH et al.; The effect of a fermented soy milk product (FSP) on various human breast carcinoma cell lines was investigated, and it was shown to have a growth-inhibitory effect, especially on MCF-7 cells . Thus the MCF-7 cell line was used to study the mechanism of action . In female severe combined immune deficiency mice implanted with MCF-7 cells, pretreatment with FSP significantly inhibited tumor growth . The inhibitory effect of FSP on MCF-7 cells seemed to be caused by the additive effects of a wide variety of constituents . The active components of FSP are mainly in the water phase, and the lipid-soluble fraction, which includes the soy isoflavones such as genistein and daidzein, is relatively ineffective . A variety of methods were used to demonstrate that FSP caused apoptotic cell death in MCF-7 cells . FSP induced generation of reactive oxygen species (ROS) . Growth inhibition and ROS generation induced by FSP could be inhibited by catalase and deferoxamine, indicating that the ROS production probably was the cause of this apoptotic cell death . This study suggests that FSP retards tumor growth in vivo and can trigger apoptosis in vitro . It may, therefore, be a potential nutritional supplement in chemotherapy.

J Appl Microbiol, 2003, 94(3), 432 - 48
Transcription profile of brewery yeast under fermentation conditions; James TC et al.; AIMS: Yeast strains, used in the brewing industry, experience distinctive physiological conditions . During a brewing fermentation, yeast are exposed to anaerobic conditions, high pressure, high specific gravity and low temperatures . The purpose of this study was to examine the global gene expression profile of yeast subjected to brewing stress . METHODS AND RESULTS: We have carried out a microarray analysis of a typical brewer's yeast during the course of an 8-day fermentation in 15 degrees P wort . We used the probes derived from Saccharomyces cerevisiae genomic DNA on the chip and RNA isolated from three stages of brewing . This analysis shows a high level of expression of genes involved in fatty acid and ergosterol biosynthesis early in fermentation . Furthermore, genes involved in respiration and mitochondrial protein synthesis also show higher levels of expression . CONCLUSIONS: Surprisingly, we observed a complete repression of many stress response genes and genes involved in protein synthesis throughout the 8-day period compared with that at the start of fermentation . SIGNIFICANCE AND IMPACT OF THE STUDY: This microarray data set provides an analysis of gene expression under brewing fermentation conditions . The data provide an insight into the various metabolic processes altered or activated by brewing conditions of growth . This study leads to future experiments whereby selective alterations in brewing conditions could be introduced to take advantage of the changing transcript profile to improve the quality of the brew.

J Appl Microbiol, 2003, 94(3), 349 - 59
Fermentative activity and production of volatile compounds by Saccharomyces grown in synthetic grape juice media deficient in assimilable nitrogen and/or pantothenic acid; Wang XD et al.; AIMS: To understand the impact of assimilable nitrogen and pantothenic acid on fermentation rate and synthesis of volatile compounds by Saccharomyces under fermentative conditions . METHODS AND RESULTS: A 2 x 3 factorial experimental design was employed with the concentrations of yeast assimilable nitrogen (YAN) (60 and 250 mg l(-1)) and pantothenic acid (10, 50 and 250 microg l(-1)) as variables . In media containing 250 microg l(-1) pantothenic acid, H2S production by two different species of Saccharomyces decreased when YAN was increased from 60 to 250 mg l(-1) . Conversely, H2S production was significantly higher when the concentration of assimilable nitrogen was increased if pantothenic acid was deficient (10 or 50 microg l(-1)) . Yeast synthesis of other volatile compounds were impacted by both assimilable nitrogen and pantothenic acid . CONCLUSIONS: While growth and fermentative rate of Saccharomyces was more influenced by nitrogen than by pantothenic acid, complicated interactions exist between these nutrients that affect the synthesis of volatile compounds including H2S . SIGNIFICANCE AND IMPACT OF THE STUDY: This study has important implications for the winemaking industry where a better understanding of the nutritional requirements of Saccharomyces is necessary to reduce fermentation problems and to improve final product quality.

Acta Microbiol Pol, 2002, 51(3), 225 - 35
Numerical modeling of ferrous-ion oxidation rate in Acidithiobacillus ferrooxidans ATCC 23270: optimization of culture conditions through statistically designed experiments; Abdel-Fattah YR et al.; Statistically designed experimental strategy has been performed in order to evaluate and optimize nutritional and environmental parameters that affect ferrous ion oxidation rate in Acidithiobacillus ferrooxidans ATCC 23270 . Plackett-Burman design was carried out to evaluate efficiently the biological significance of 10 culture conditions influencing ferrous-ion oxidation rate of A . ferrooxidans grown for 5 days in shake-flask batch mode on the newly modified 9-K media . Among ten fermentation factors examined, the most significant variables influencing ferrous-ion oxidation rate were statistically elucidated to be pH and calcium nitrate as positive contributors, whereas trace metals solution and potassium chloride were the most significant negative contributors . The optimal levels of the most significant three nutritional factors were further predicted from a polynomial model created from the data obtained from three level factorial design, a Box-Behnken design . Predicted optimal ferrous-ion oxidation rate Q(Fe2+) was recorded to be 0.148 (g Fe2+/l/hr) . On verifying the predicted value, an experiment was performed under optimal predicted conditions and showed an actual experimental Q(Fe2+) of 0.152 g/l/hr, which was 2.7% over the predicted value . Our optimized medium formula gave overall five folds increase in ferrous-ion oxidation rates over the previously published data of standard 9-K medium on batch culture of A . ferrooxidans ATCC 23270 with higher mu(max) (hr(-1)) of 0.177 which was achieved within 75 h incubation in shake-flask culture.

Indian J Exp Biol, 2002 Sep, 40(9), 1038 - 42
Infra-red spectroscopic analyses of banana waste degraded by oyster mushroom; Reddy GV et al.; Carbon, hydrogen and nitrogen analyses of banana leaf and pseudostem biomass revealed their potentiality as substrates for microorganisms . Infra-red (IR) spectra of both biomass show presence of cellulose, xylan and lignin . IR spectra of leaf and pseudostem biomass degraded in solid state fermentation (SSF) by two Pleurotus species (P . sajor-caju and P . ostreatus) for 40 days showed the utilization of cellulose, xylan and lignin by these microbes . Dynamics of various lignocellulolytic enzymes of Pleurotus species and analyses of carbon, hydrogen and nitrogen contents of degraded biomass supported the same . Both the Pleurotus species exhibited lignin consumption ability on both the substrates.

Indian J Exp Biol, 2002 Sep, 40(9), 1032 - 7
Beta-fructofuranosidase production by 2-deoxyglucose resistant mutants of aspergillus niger in submerged and solid-state fermentation; Ashokkumar B et al.; Aspergillus niger produces extracellular beta-fructofuranosidase under submerged (SmF) and solid state fermentation (SSF) conditions . After UV mutagenesis of conidiospores of A . niger, 2-deoxyglucose (10 g/l) resistant mutants were isolated on Czapek's minimal medium containing glycerol as a carbon source and the mutants were examined for improved production of beta-fructofuranosidase in SmF and SSF conditions . One of such mutant DGRA-1 overproduced beta-fructofuranosidase in both SmF and SSF conditions . In SmF, the mutant DGRA-1 showed higher beta-fructofuranosidase productivity (110.8 U/l/hr) than the wild type (48.3 U/l/hr) . While in SSF the same strain produced 322 U/l/hr of beta-fructofuranosidase, 2 times higher than that of wild type (154.2 U/l/hr) . In SmF, both wild type and mutants produced relatively low level of beta-fructofuranosidase in medium containing sucrose with glucose than from the sucrose medium . However in SSF, the DGRA-1 mutant grown in sucrose and sucrose+ glucose did not show any difference with respect to beta-fructofuranosidase production . These results indicate that the catabolite repression of beta-fructofuranosidase synthesis is observed in SmF whereas in SSF such regulation was not prominent.

Genetics, 2003 Jan, 163(1), 79 - 89
Genetic diversity in yeast assessed with whole-genome oligonucleotide arrays; Winzeler EA et al.; The availability of a complete genome sequence allows the detailed study of intraspecies variability . Here we use high-density oligonucleotide arrays to discover 11,115 single-feature polymorphisms (SFPs) existing in one or more of 14 different yeast strains . We use these SFPs to define regions of genetic identity between common laboratory strains of yeast . We assess the genome-wide distribution of genetic variation on the basis of this yeast population . We find that genome variability is biased toward the ends of chromosomes and is more likely to be found in genes with roles in fermentation or in transport . This subtelomeric bias may arise through recombination between nonhomologous sequences because full-gene deletions are more common in these regions than in more central regions of the chromosome.

FEMS Microbiol Lett, 2003 Jan 28, 218(2), 385 - 93
Regulation of the flavorubredoxin nitric oxide reductase gene in Escherichia coli: nitrate repression, nitrite induction, and possible post-transcription control; da Costa PN et al.; Escherichia coli flavorubredoxin is a new type of cytoplasmic nitric oxide (NO) reductase, which shows NO reductase activity within the range of the canonical membrane-bound heme b(3)-iron NO reductases . Using reverse-transcription polymerase chain reaction we show that although the flavorubredoxin gene (flrd) is transcribed in both aerobic and anaerobic conditions, anaerobiosis induced transcription up to 12-fold, under fermentative conditions; a 28-fold stimulation was observed in an E . coli fnr mutant strain, showing that the flavorubredoxin gene is negatively regulated by FNR . The level of anaerobic transcription was repressed three-fold by nitrate, but induced 47-fold by nitrite . The transcription factors NarL and NarP are not essential for flrd expression . Furthermore, the addition of NO within the physiological range of concentrations does not induce anaerobic transcription of flrd . Since two other E . coli proteins are known to exhibit NO reductase activity, flavohemoglobin and the pentaheme cytochrome c nitrite reductase, we have also compared the concentrations of their mRNAs with those of flavorubredoxin, under the same growth conditions . Transcription of the putative transcriptional activator of flavorubredoxin, ygaA, is also regulated by the absence of oxygen and the presence of nitrite . Levels of FlRd protein did not correlate with mRNA levels . The results reveal that a complex regulation of flavorubredoxin expression is operative, possibly by both transcriptional and post-transcriptional mechanisms.

Biochim Biophys Acta, 2003 Feb 17, 1610(1), 133 - 40
Expression and purification of truncated, non-glycosylated turkey beta-adrenergic receptors for crystallization; Warne T et al.; In order to purify milligram quantities of turkey beta-adrenergic receptor (betaAR) for structural analysis, we have expressed mutant betaARs using the baculovirus system . The initial betaAR construct was truncated at both N- and C-termini thus removing an N-glycosylation site . Cys 116 was mutated to leucine and a histidine tag was added at the C-terminus resulting in the betaAR construct 20-424/His6 . Expression of this construct in Sf9 cells produced 0.5 mg of unpurified receptor per liter of culture which necessitated the use of a fermenter for large-scale production . The yield was improved more than 2-fold to 1.2 mg/l culture by using Tni cells which facilitated the production of receptor on a 4 litre scale in shake cultures . The receptor was purified to homogeneity with 35% recovery giving a yield of 2 mg receptor . A further deletion at the N-terminus (betaAR 34-424/His6) eliminated proteolysis which had been observed with the original construct and also increased expression more than 5-fold to 360 pmol/mg solubilized membrane protein . This expression level is one of the highest reported for a G protein-coupled receptor (GPCR) and has enabled us to purify 10 mg betaAR for large-scale crystallization experiments .

Biotechnol Bioeng, 2003 Apr 20, 82(2), 152 - 69
Metabolic analysis of the synthesis of high levels of intracellular human SOD in Saccharomyces cerevisiae rhSOD 2060 411 SGA122; Gonzalez R et al.; The synthesis of human superoxide dismutase (SOD) in batch cultures of a Saccharomyces cerevisiae strain using a glucose-limited minimal medium was studied through metabolic flux analysis . A stoichiometric model was built, which included 78 reactions, according to metabolic pathways operative in these strains during respirofermentative and oxidative metabolism . It allowed calculation of the distribution of metabolic fluxes during diauxic growth on glucose and ethanol . Fermentation profiles and metabolic fluxes were analyzed at different phases of diauxic growth for the recombinant strain (P+) and for its wild type (P-) . The synthesis of SOD by the strain P+ resulted in a decrease in specific growth rate of 34 and 54% (growth on glucose and ethanol respectively) in comparison to the wild type . Both strains exhibited similar flux of glucose consumption and ethanol synthesis but important differences in carbon distribution with biomass/substrate yields and ATP production 50% higher in P- . A higher contribution of fermentative metabolism, with 64% of the energy produced at the phosphorylation level, was observed during SOD production . The flux of precursors to amino acids and nucleotides was higher in the recombinant strain, in agreement with the higher total RNA and protein levels . Lower specific growth rates in strain P+ appear to be related to the decrease in the rate of synthesis of nonrecombinant protein, as well as a decrease in the activities of the pentose phosphate (PP) pathway and TCA cycle . A very different way of entry into the stationary phase was observed for each strain: in the wild-type strain most metabolic fluxes decreased and fluxes related to energy reserve synthesis increased, while in the P+ strain the flux of 22 reactions (including PP pathway and amino acids biosynthesis) related to SOD production increased their fluxes . Changes in SOD production rates at different physiological states appear to be related to the differences in building blocks availability between respirofermentative and oxidative metabolism . Using the present expression system, ideal conditions for SOD synthesis are represented by either active growth during respirofermentative metabolism or transition from a growing to a nongrowing state . An increase in SOD flux could be achieved using an expression system nonassociated to growth and potentially eliminating part of the metabolic burden .

FEMS Microbiol Lett, 2003 Jan 21, 218(1), 59 - 64
Production of D-(--)-3-hydroxyalkanoic acid by recombinant Escherichia coli; Zhao K et al.; Pathways for extracellular production of chiral D-(-)-3-hydroxybutyric acid (3HB) and D-(-)-3-hydroxyalkanoic acid (mcl-3HA) were constructed by co-expression of genes of beta-ketothiolase (phbA), acetoacetyl-CoA reductase (phbB) and 3-hydroxyacyl-ACP CoA transacylase (phaG), respectively, in Escherichia coli strain DH5alpha . The effect of acrylic acid and glucose on production of both 3HB and mcl-3HA was investigated . It was found that the addition of acrylic acid significantly increased production of 3HB and mcl-3HA consisting of 3-hydroxyoctanoic acid and 3-hydroxydecanoic acid in a ratio of 1:3 from 199 mg x l(-1) to 661 mg x l(-1) and from 27 mg x l(-1) to 135 mg x l(-1), respectively, in shake flask studies when glucose was present in the medium at the very beginning of fermentation . The timing of glucose addition had no effect on 3HB production . In contrast, mcl-3HA production was affected by glucose addition, an mcl-3HA concentration of 193 mg x l(-1) was obtained when glucose was added to the culture at 12 h . A more than seven-fold increase was obtained when compared with that in medium containing glucose at the beginning of fermentation . However, a decrease in production of 3HB and mcl-3HA was found when glucose was added at 12 h to the culture containing acrylic acid . The repressive effect of acrylic acid on acetic acid production was also evaluated and discussed.

Yao Xue Xue Bao, 2001 Oct, 36(10), 787 - 9
{Study on the structural property of polysacchrides 139A--a kind of IL-1R antagonists from microorganism}; Chen J et al.; AIM: To study the structural property of the bioactive component from the fermentation broth of Streptomyces 139 which can inhibit IL-1R . METHODS: Polysaccharide 139A is extracted and purified by means of macroreticular resin Diaion HP-20, cation exchange and DEAE Sephadex A-25 chromatography . The sugar compositions were analyzed with GC, the linkage positions of 139A were determined by methylation and GC-MS . RESULTS: The bioactive component which can inhibit IL-1R is a kind of polysaccharides . CONCLUSION: The components of 139A repeat units are conjectured.

J Plant Res, 2002 Oct, 115(5), 315 - 20 Epub 2002 Jul 18.
Oxygen stress and adaptation of a semi-aquatic plant: rice ( Oryza sativa); Das A et al.; One of the major abiotic stresses that affects plant growth and development is anoxia or hypoxia . Rice is a semi-aquatic plant bestowed with the capability of overcoming oxygen limitation for a considerable period of time . For instance, it can withstand submergence stress either by inherent metabolic adaptations (resistant type), or by keeping its leaves above the water surface by continuously elongating the stem (avoiding type) . In the former case, an interplay of several metabolic pathways engaged in anaerobic fermentation keeps the submerged plant alive for a certain period of time . In the latter type, also known as deepwater rice, continuous stem elongation brought about by a series of reactions in planta enables the shoot to remain above the water surface and thus maintain respiration and photosynthesis . However, the earliest event, i.e., sensing the oxygen level that brings about all the changes, has not been clearly understood . This paper intends to evaluate the metabolic adaptations of rice plants to oxygen constraints.

Biotechnol Appl Biochem, 2003 Feb, 37(Pt 1), 91 - 102
Elicitation of plants and microbial cell systems; Radman R et al.; Plants show physiological and morphological responses to a range of physical and chemical factors known as 'elicitors' . These responses have been considered as defence reactions 'elicited' by the plants' biochemical factory to ensure their survival, persistence and competitiveness . Recently examples have been cited of elicitation in some fungal and bacterial cultures . Through a chronological survey, this Review considers examples of elicitors and elicitation and describes suggested mechanisms of elicitation in plants and microbial cell cultures . The majority of research in this field has been carried out on the plant systems using complex (undefined) biotic elicitors . Carbohydrates are the main class of compounds used as defined elicitors . This Review focuses on carbohydrates as compounds initiating a defence response in cell cultures . Physiological changes brought about on the plant and microbial cultures include expression of novel metabolites and overproduction of already known products . Recent reports confirming elicitation in microbial cultures are of potential importance, as the relative ease of fermentation and scale-up could open an opportunity for the introduction of useful novel metabolites as well as enhancement of commercially useful bioproducts . In this context, a sound knowledge of the elicitor molecules' structure-function relationships and mechanisms of elicitation is essential.

Bioresour Technol, 2003 Jun, 88(2), 153 - 6
Using tea waste as a new casing material in mushroom (Agaricus bisporus (L.) Sing.) cultivation; Gulser C et al.; In this study, the possibility of using tea production waste as a new casing material in mushroom (Agaricus bisporus) cultivation was investigated . Some physical and chemical characteristics of tea waste, fermented tea waste and a mixture of tea waste with peat were compared with that of peat casing, as were their effects on yield . The highest yield was obtained from peat casing . Using tea production waste alone as a casing was not acceptable for assured yield when it was compared with peat . But, a mixture of tea production waste with peat in 1:1 (v:v) ratio increased the yield . There was no significant difference between the mushroom yields of tea production waste+peat and peat casing materials at the end of 30 and 40 days . High salt content, organic and inorganic compounds in casing materials caused reduction of yields . However, a high iron content in casing material gave a significant positive correlation with total yield at 40 days.

Br J Nutr, 2003 Feb, 89(2), 181 - 8
Effect of the addition of malate on in vitro rumen fermentation of cereal grains; Carro MD et al.; Batch cultures of mixed rumen micro-organisms were used to study the effects of different concentrations of malate (Rumalato(R); Norel & Nature S.A., Barcelona, Spain; composed of disodium malate-calcium malate (0.16:0.84, w/w)) on the fermentation of four cereal grains (maize, barley, wheat and sorghum) . Rumen contents were collected from four Merino sheep fed lucerne hay ad libitum and supplemented with 300 g concentrate/d . Rumalato(R) was added to the incubation bottles to achieve final concentrations of 0, 4, 7 and 10 mm-malate . Gas production was measured at regular intervals up to 120 h . Malate increased (P<0.01) the average fermentation rate of all substrates, and the lag time decreased (P<0.05) linearly with increasing concentrations of malate for all substrates, with the exception of sorghum . In 17 h incubations, the final pH and total volatile fatty acid production increased (P<0.001) linearly for all substrates as malate concentration increased from 0 to 10 mm . Propionate and butyrate production increased (P<0.05), while the value of the acetate : propionate ratio and l-lactate concentrations decreased (P<0.05) linearly with increasing doses of malate . Malate treatment increased (P<0.05) the CO2 production and decreased the production of CH4, although this effect was not significant (P>0.05) for maize . Malate at 4 and 7 mm increased (P<0.05) optical density of the cultures measured at 600 nm for maize, with no differences for the other substrates . The results indicate that malate may be used as a feed additive for ruminant animals fed high proportions of cereal grains, because it increased pH and propionate production and decreased CH4 production and l-lactate concentrations; however, in general, no beneficial effects of 10 compared with 7 mm-malate were observed.

Bioresour Technol, 2003 May, 88(1), 17 - 25
Availability of corn stover as a sustainable feedstock for bioethanol production; Kadam KL et al.; The amount of corn stover that can be sustainably collected is estimated to be 80-100 million dry tonnes/yr (t/yr), a majority of which would be available to ethanol plants in the near term as only a small portion is currently used for other applications . Potential long-term demand for corn stover by non-fermentative applications in the United States is estimated to be about 20 million dry t/yr, assuming that corn stover-based products replace 50% of both hardwood pulp and wood-based particleboard, and that 50% of all furfural production is from corncobs . Hence, 60-80 million dry t/yr of corn stover should be available to fermentative routes . To achieve an ethanol production potential of 11 billion L (3 billion gal) per year (a target level for a non-niche feedstock), about 40% of the harvestable corn stover is needed . This amount should be available as long as the diversion of corn stover to non-ethanol fermentative products remains limited.

Biotechnol Prog, 2003 Jan-Feb, 19(1), 238 - 42
Effect of time of harvest of budded virus on the selection of baculovirus FP mutants in cell culture; Lua LH et al.; Rapid formation and selection of FP (few polyhedra) mutants occurs during serial passaging of Helicoverpa armigera nucleopolyhedrovirus (HaSNPV) in insect cell culture . The production of HaSNPV for use as biopesticides requires the passaging of the virus over a number of passages to produce enough virus inoculum for large-scale fermentation . During serial passaging in cell culture, FP mutants were rapidly selected, resulting in declined productivity and reduced potency of virus . Budded virus (BV) is usually harvested between 72 and 96 h postinfection (hpi) in order to obtain a high titer virus stock . In this study, the effect of time of harvest (TOH) for BV on the selection rate of HaSNPV FP mutants during serial passaging was investigated . BV were harvested at different times postinfection, and each series was serially passaged for six passages . The productivity and percentage of FP mutants at each passage were determined . It was found that the selection of FP mutants can be reduced by employing an earlier TOH for BV . Serial passaging with BV harvested at 48 hpi showed a slower accumulation of FP mutants compared to that of BV harvested after 48 hpi . Higher cell specific yields were also maintained when BV were harvested at 48 hpi . When BV that were formed between 48 and 96 hpi were harvested and serially passaged, FP mutants quickly dominated the virus population . This suggests that the BV formed and released between 48 and 96 hpi are most likely from FP mutant infected cells.

Biotechnol Prog, 2003 Jan-Feb, 19(1), 175 - 8
Improved fermentation processes for NS0 cell lines expressing human antibodies and glutamine synthetase; Dempsey J et al.; To meet the increasing requirement for therapeutic antibodies to conduct clinical trials, an enhanced culture medium and fed-batch process was developed for GS-NS0 cell lines . This process was shown to produce high concentrations of monoclonal antibodies for several cell lines expressing different antibodies . Cells were adapted to growth in a glutamine- and serum-free medium containing bovine serum albumin (BSA), cholesterol, and transferrin . A number of amino acids were found to be depleted during cell culture . The concentrations of these amino acids were increased, and further cell culture analyses were performed . This process of cell growth and analysis was repeated over multiple cycles until no depletion was detected . This resulted in an amino acid supplement that was shown to be generic and enhanced antibody productivity up to 5-fold for the three cell lines tested . Transferrin was replaced using tropolone, a lipophilic iron chelator and ferric ammonium citrate . Cell growth was equivalent to that in transferrin-containing medium over the wide ranges tested . A concentrated feed solution, based on the amino acid supplement and the components of the serum- and protein-free supplements, was formulated . Addition of this feed in response to metabolic requirements resulted in a harvest titer a further 2-fold higher than the enhanced culture medium . Harvest antibody titers of up to 600 mg/L were achieved for three cell lines expressing different antibodies, representing an increase of 10-fold over the starting concentrations.

J Gastroenterol, 2002 Nov, 37 Suppl 14, 38 - 42
Role of the innate immune system in the development of chronic colitis; Kanai T et al.; Based on Pasteur's work on the microbial nature of fermentation, it was widely believed that the presence of bacteria in the intestine was essential for the life of the host . It has also been known for decades that gut commensal microbes effect the activation and development of the systemic immune system through gut-associated lymphoid tissues (GALT) . Recent extensive studies have shown that recognition of microbes is mediated by a set of germline-encoded receptors, Toll-like receptors (TLRs), in mammals . This article reviews the role of the innate immunity system in the development of GALT and the pathogenesis of inflammatory bowel diseases (IBD).

Biometals, 2003 Jun, 16(2), 251 - 61
Cadmium stress studies: media development, enrichment, consortia analysis, and environmental relevance; Knotek-Smith HM et al.; The effects of Cadmium (Cd) toxicity on bacterial consortia originating from an-aerobic sewage sludge and cultivated under differing enrichment conditions were studied . Cultures were enriched in minimal media developed specifically for Cd stress studies . At inoculation all Cd was soluble in free ion or chelated form . Electron donors and acceptors were varied to obtain each physiological enrichment type . Adaptation leading to higher levels of Cd resistance of the consortia over time was observed under all physiological conditions . Initial and increased Cd tolerances were consistently greatest in multiphysiological enrichments (MPH) . Sulfate reducing (SRB), methanogenic (MET), and fermentative (FRM) enrichments had less tolerance however, the level of tolerance to the Cd varied from one inoculation to the next . The Cd remained soluble as free Cd in MPH and FRM conditions and was precipitated significantly in SRB and moderately in MET conditions . Denaturing gradient gel electrophoresis (DGGE) of polymerase chain reaction (PCR) amplified 16S rRNA of the SRB, MPH, and FRM enrichments were followed over time . The consortia underwent succession under all physiological conditions when compared with the profile of the inoculum . Microbial population diversity decreased as the consortia were subcultured . The effects of chelators in the MPH medium were also evaluated . The addition of chelators transiently decreased toxicity . Effects of MPH medium on the Cd sorption capacity of soil were evaluated . Microbial growth decreased the amount of Cd left in solution.

Zhong Yao Cai, 1997 Sep, 20(9), 459 - 61
{Pharmacological study of the rhizome powder of Pinellia pedatisecta processed by different procedures}; He Y et al.; The potentiation of hypnotic activity of a barbituate and acute tocicity by the rhizome powder of Pinellia pedatisecta Schott processed by pigbile with two different traditional medicinal procedures (fermentation mixing and heating alternatively) were studied in mice . The results showed that aquatic extracts of two processed drugs could potentiate the action of hypnotics; and ethanol extracts of the drug processed with mixing and heating alterntively could significantly enhance this action, compared with that of the drug pricessed with fermentation; while no acute toxicity was measured by the two processed drugs.

Zhong Yao Cai, 1997 Aug, 20(8), 394 - 6
{Improvement of product technology on prepared soypean}; Lin S et al.; This paper reports the new prepared method of Prepared Soypean by purebred culture of Aspergillus niger . The technology condition is that temperture is 28 +/- 2 degrees C and comparative humidity is 95%, time of fermented culture is 15-20 days . After improvement, polluttion of other bacteria is easily avoided.

Appl Environ Microbiol, 2003 Feb, 69(2), 740 - 6
Molecular analysis of a Saccharomyces cerevisiae mutant with improved ability to utilize xylose shows enhanced expression of proteins involved in transport, initial xylose metabolism, and the pentose phosphate pathway; Wahlbom CF et al.; Differences between the recombinant xylose-utilizing Saccharomyces cerevisiae strain TMB 3399 and the mutant strain TMB 3400, derived from TMB 3399 and displaying improved ability to utilize xylose, were investigated by using genome-wide expression analysis, physiological characterization, and biochemical assays . Samples for analysis were withdrawn from chemostat cultures . The characteristics of S . cerevisiae TMB 3399 and TMB 3400 grown on glucose and on a mixture of glucose and xylose, as well as of S . cerevisiae TMB 3400 grown on only xylose, were investigated . The strains were cultivated under chemostat conditions at a dilution rate of 0.1 h(-1), with feeds consisting of a defined mineral medium supplemented with 10 g of glucose liter(-1), 10 g of glucose plus 10 g of xylose liter(-1) or, for S . cerevisiae TMB 3400, 20 g of xylose liter(-1) . S . cerevisiae TMB 3400 consumed 31% more xylose of a feed containing both glucose and xylose than S . cerevisiae TMB 3399 . The biomass yields for S . cerevisiae TMB 3400 were 0.46 g of biomass g of consumed carbohydrate(-1) on glucose and 0.43 g of biomass g of consumed carbohydrate(-1) on xylose . A K(s) value of 33 mM for xylose was obtained for S . cerevisiae TMB 3400 . In general, the percentage error was <20% between duplicate microarray experiments originating from independent fermentation experiments . Microarray analysis showed higher expression in S . cerevisiae TMB 3400 than in S . cerevisiae TMB 3399 for (i) HXT5, encoding a hexose transporter; (ii) XKS1, encoding xylulokinase, an enzyme involved in one of the initial steps of xylose utilization; and (iii) SOL3, GND1, TAL1, and TKL1, encoding enzymes in the pentose phosphate pathway . In addition, the transcriptional regulators encoded by YCR020C, YBR083W, and YPR199C were expressed differently in the two strains . Xylose utilization was, however, not affected in strains in which YCR020C was overexpressed or deleted . The higher expression of XKS1 in S . cerevisiae TMB 3400 than in TMB 3399 correlated with higher specific xylulokinase activity in the cell extracts . The specific activity of xylose reductase and xylitol dehydrogenase was also higher for S . cerevisiae TMB 3400 than for TMB 3399, both on glucose and on the mixture of glucose and xylose.

Curr Pharm Des, 2003, 9(4), 347 - 58
Role of dietary fiber and short-chain fatty acids in the colon; Andoh A et al.; Luminal nutrition is important for maintenance of gastrointestinal mucosal structure and function . In particular, short chain fatty acids (SCFAs), metabolic products of anaerobic bacterial fermentation of dietary fiber and resistant starch, are particularly important as the preferred respiratory fuel of the colonocytes . A variety of biological effects of SCFAs have been reported, and there is now increasing number of experimental works showing new aspects of these molecules . For example, as the mechanisms mediating anti-inflammatory effects of SCFAs, several investigators identified the inhibitory effect of butyrate on proinflammatory cytokine-induced NF-kappaB activation . Various inflammatory responses are now discussed with the central role of NF-kappaB activation, and thus the inhibition of NF-kappaB activation represents the efficacy of dietary fiber and SCFAs in the treatment with inflammatory bowel disease . Furthermore, recent advance in molecular technology has identified mechanisms mediating anti-tumor effects of SCFAs . SCFAs modulate expression of cell cycle-regulating proteins and induce apoptosis in colon cancer cells . SCFAs increase the susceptibility of colon cancer cells to complement-mediated cell injury . In this review, new aspects of functions of SCFAs are focused and summarized.

Zhong Yao Cai, 1998 Aug, 21(8), 379 - 81
{The study on the fermentation medium of Ganoderma lucidum}; Li G et al.; This study has explored the fermentation medium of Ganoderma lucidum including inorganic salts, carbon source, nitrogen source and vitamins . Then a suitable formula of fermented medium was obtained by comparing different production slats of Gandoerma powder during fermentation.

Biotechnol Bioeng, 2003 Apr 5, 82(1), 111 - 7
Pulsed addition of limiting-carbon during Aspergillus oryzae fermentation leads to improved productivity of a recombinant enzyme; Bhargava S et al.; Fungal morphology in many filamentous fungal fermentations leads to high broth viscosity which limits oxygen mass transfer, and often results in reduced productivity . The objective in this study was to determine if a simple, fed-batch, process strategy-pulsed addition of limiting-carbon source-could be used to reduce fungal broth viscosity, and increase productivity of an industrially relevant recombinant enzyme (glucoamylase) . As a control, three Aspergillus oryzae fed-batch fermentations were carried out with continuous addition of limiting-carbon . To determine the effect of pulse-feeding, three additional fermentations were carried out with limiting-carbon added in 90-second pulses, during repeated five-minute cycles . In both cases, overall carbon feed-rate was used to control dissolved oxygen concentration, such that increased oxygen availability led to increased addition of limiting-carbon . Pulse-fed fermentations were found to have smaller fungal mycelia, lower broth viscosity, and improved oxygen mass transfer . As a result, more carbon was added to pulse-fed fermentations that led to increased enzyme productivity by as much as 75% . This finding has significant implications for the bioprocessing industry, as a simple process modification which is likely to cost very little to implement in most production facilities, has the potential to substantially increase productivity .

Am J Physiol Endocrinol Metab, 2003 Jun, 284(6), E1177 - 80 Epub 2003 Feb 04.
Lysine synthesized by the gastrointestinal microflora of pigs is absorbed, mostly in the small intestine; Torrallardona D et al.; This study used a digesta transfer protocol to determine the site of absorption of lysine synthesized by the gastrointestinal microflora of pigs . Eight pigs were used, four with reentrant cannulas in the terminal ileum, two with simple T cannulas in the terminal ileum, and two intact . All pigs were given, for 5 days, the same low-protein diet that included fermentable carbohydrates . The diet of two pigs with reentrant cannulas (donor) and of the two intact (control) pigs was supplemented with (15)NH(4)Cl . The two other pigs with reentrant cannulas (acceptor pigs) and those with simple cannulas (used to supply unlabeled digesta) were given the same diet but unlabeled NH(4)Cl . Ileal digesta were collected continuously from all of the reentrant cannulas and kept on ice . All digesta from each donor pig were reheated and returned to the distal cannula of its companion acceptor, whose ileal digesta were discarded . Unlabeled ileal digesta from the pigs with simple cannulas were instilled into the distal cannulas of the donor pigs . At the end of the experiment, the average (15)N enrichment in the plasma free lysine of control pigs was 0.0407 atom % excess (APE); that of donor pigs was 0.0322 APE (79% of controls), whereas that of acceptor pigs was only 0.0096 APE (24% of controls) . Due to nitrogen recycling, acceptor pigs had labeled lysine in the digesta of the stomach and small intestine, and donor pigs had labeled lysine in the digesta of the large intestine . If account is taken of the higher (15)N enrichment of microbial lysine in the large compared with the small intestine, it can be estimated that >90% of the absorption of microbial lysine took place in the small intestine.

J Agric Food Chem, 2003 Feb 12, 51(4), 1085 - 9
Ingestion of water-soluble soybean fiber prevents osteopenia and hypercholesterolemia induced by ovariectomy in rats; Mitamura R et al.; This study examined the effects of feeding water-soluble soybean fiber (WSSF), a highly fermentable dietary fiber with low viscosity, on intestinal calcium (Ca) absorption, bone mineral content, and serum total cholesterol levels using female Sprague-Dawley rats (5 weeks old) with or without ovariectomy (OVX) . The rats in each group were fed a WSSF (50 g/kg of diet) or WSSF-free diet for 4 weeks . Ca absorption was higher in OVX rats fed the WSSF diet than in OVX rats fed the WSSF-free diet . Femoral Ca content in both sham and OVX rats fed the WSSF diet was higher than that in rats fed the WSSF-free diet . The serum total cholesterol levels were elevated after OVX compared with that in the sham rats, and this elevation was prevented by the WSSF diet . In conclusion, ingestion of WSSF prevented osteopenia and hypercholesterolemia induced by ovariectomy in rats.

J Agric Food Chem, 2003 Feb 12, 51(4), 916 - 22
Effect of organic acids and nitrogen source on alcoholic fermentation: study of their buffering capacity; Torija MJ et al.; The effect of tartaric acid and other organic acids on alcoholic fermentation was studied . Organic acids in media with high sugar concentrations and ammonium as the sole nitrogen source had an enormous impact on Saccharomyces cerevisiae metabolism during alcoholic fermentation . The main effect on yeast metabolism was the quick acidification of the media in the absence of organic acids . All of the organic acids used in this study (tartaric, malic, citric, and succinic acids) showed a buffering capacity, but not all of the acids had the same one . However, the results suggested that buffering should not be considered the only effect of organic acids on yeast metabolism . Nitrogen source also had a great influence on media pH . Ammonium consumption by yeasts produced a greater acidification of the media than when amino acids were used.

Bioorg Med Chem Lett, 2003 Feb 10, 13(3), 573 - 5
Condensed aromatic peptide family of microbial metabolites, inhibitors of CD28-CD80 interactions; Hegde VR et al.; Three condensed aromatic peptides SCH79235 (1), SCH79236 (2), and SCH204698 (3) were isolated from the fermentation broth of a Streptomycete microorganism . The structure of SCH204698 (3) was established by extensive NMR spectral data . All these compounds exhibited good activity against CD28-CD80 binding with an IC(50) of 0.42, 0.38 and 0.22 microM, respectively.

Mem Inst Oswaldo Cruz, 2002 Dec, 97(8), 1085 - 9 Epub 2003 Jan 20.
Phenotypic and genotypic characteristics of shiga toxin-producing Escherichia coli strains isolated from children in São Paulo, Brazil; Guth BE et al.; The biochemical and serological characteristics, virulence properties, and genetic relatedness of Shiga toxin-producing Escherichia coli (STEC) strains isolated in S o Paulo, from April 1989 through March 1990, were determined . This is also the first report on clinic findings of human STEC infections in Brazil . The only three STEC strains identified in that period were lysine decarboxylase negative, belonged to serotype O111ac: non-motile, were Stx1 producers, carried the eae and astA genes, and 2 of them also presented the EHEC-hly sequence . The children carrying STEC were all boys, with less than two years old, and had no previous history of hospitalization . None of them presented blood in stools . Vomiting, cough and coryza were the most common clinical manifestations observed . Although the STEC strains were isolated during summer months, and presented similar phenotypic and genotypic characteristics, carbohydrate fermentation patterns and PFGE analysis suggested that these diarrheal episodes were not caused by a single clone.

J Biol Chem, 2003 Apr 11, 278(15), 12820 - 5 Epub 2003 Jan 31.
Localization and function of the yeast multidrug transporter Tpo1p; Albertsen M et al.; In Saccharomyces cerevisiae four transporters, Tpo1p-Tpo4p, all members of the major facilitator superfamily, have been shown to confer resistance to polyamines . It was suggested that they act by pumping their respective substrate into the lumen of the vacuole depending on the proton gradient generated by the V-ATPase . Using sucrose gradient ultracentrifugation we found that an hemagglutinin (HA)-tagged Tpo1p as well as its HA-tagged Tpo2p-4p homologues co-localize with plasma membrane markers . Because the HA-tagged Tpo1p carrier protein proved to be functional in conferring resistance to polyamines in TPO1 knockouts, a function of Tpo1p in transport of polyamines across the plasma membrane seemed to be likely . The polyamine transport activity of wild type cells was compared with the respective activity of a TPO1 knockout strain . The results obtained strongly suggest that Tpo1p is a plasma membrane-bound exporter, involved in the detoxification of excess spermidine in yeast . When studying polyamine transport of wild type cells, we furthermore found that S . cerevisiae is excreting putrescine during the fermentative growth phase.

Sheng Wu Gong Cheng Xue Bao, 2002 Sep, 18(5), 622 - 5
{Analysis on metabolic flux shift during guanosine fermentation}; Cai XP et al.; Taking the typical metabolic control product-guanosine as an example, the method of metabolic flux shift investigation based on process multi-levels parameter correlation analysis was established . The metabolic pathway, multi-parameter correlation, accumulation of amino acid and organic acid during guanosine fermentation process were integratively analyzed . The metabolic flux shift from HMP to EMP was ascertained, which was assumed to be caused by the accumulation of ammonium ion . The subsequent optimization based on controlling flux distribution between EMP and HMP did improve the yield by 35% when the metabolic flux shift was prevented.

Sheng Wu Gong Cheng Xue Bao, 2002 Sep, 18(5), 583 - 7
{Characterization of inductive synthesis of levoglucosan kinase by a combined strategy of enzymological and fast atom bombardment mass spectrometric analysis}; Zhuang XL et al.; Levoglucosan is the main product derived from pyrolysis of cellulose . A mutant Aspergillus niger CBX-209 could grow on levoglucosan well fermenting it into citric acid with a yield comparable to that on glucose . Levoglucosan hydrolase was absent by measuring glucose formation with the glucose oxidase and peroxidase coupling system . Cell extracts were partly purified by ammonium sulfate fractionation and ion-exchange chromatograph . Direct formation of glucose 6-phosphate from levoglucosan in the presence of ATP and MgCl2 was observed when it was reacted with partly purified enzyme by a combined strategy of enzymological and fast atom bombardment mass spectrometric analysis . These data showed that the mutant used a novel enzyme, levoglucosan kinase, to convert levoglucosan into glucose 6-phosphate . Levoglucosan kinase was an inductive enzyme.

Sheng Wu Gong Cheng Xue Bao, 2002 Sep, 18(5), 541 - 5
{Fusion expression of human thymosin alpha 1 in Escherichia coli}; Xiu ZY et al.; Engineering E . coli strain, BL21 (DE3)/pGEX-4T-human Thymosin alpha 1, was constructed by oligonucleotide annealing and PCR amplifying the target gene, then ligating it with pGEX-4T-3 vector and transferring into BL21 host . The yield of fusion protein of GST-Thymosin alpha 1 expressed from BL21 (DE3)/pGEX-4T-thymosin alpha 1 is about 35%-40% of total protein after fermentation . Following the simple cut of thrombin or CNBr, about 0.2 g/L thymosin alpha 1 can be harvested . The product is checked by MS and activity test, which indicates that the recombinant product has full biological activity of native thymosin alpha 1.

Res Microbiol, 2002 Dec, 153(10), 667 - 77
Dependency of sugar transport and phosphorylation by the phosphoenolpyruvate-dependent phosphotransferase system on membranous phosphatidyl glycerol in Escherichia coli: studies with a pgsA mutant lacking phosphatidyl glycerophosphate synthase; Aboulwafa M et al.; It has been reported that phosphatidyl glycerol (PG) is specifically required for the in vitro activities of the hexose-phosphorylating Enzymes II of the Escherichia coli phosphoenolpyruvate-dependent sugar transporting phosphotransferase system (PTS) . We have examined this possibility by measuring the properties of a null pgsA mutant that lacks detectable PG . The mutant showed lower in vitro phosphorylation activities towards several sugars when both PEP-dependent and sugar-phosphate-dependent {14C}sugar phosphorylation reactions were measured . The order of dependency on PG for the different enzymes II was: IIMannose > IIGlucose > IIFructose > IIMannitol . Nonsedimentable (40000 rpm for 2 h) Enzymes II exhibited a greater dependency on PG than pelletable Enzymes II . Western blot analyses showed that the glucose Enzyme II is present in normal amounts . Transport and fermentation measurements revealed diminished activities for all Enzymes II . Thermal stability of all of these enzymes except the mannitol-specific Enzyme II was significantly decreased by the pgsA mutation, and sensitivity to detergent treatments was enhanced . Sugar transport proved to be the most sensitive indicator of proper Enzyme II-phospholipid association . Our results show that PG stimulates but is not required for Enzyme II function in E . coli.

Wei Sheng Wu Xue Bao, 2002 Aug, 42(4), 418 - 24
{Study on effect of Vitreoscilla hemoglobin gene expression on growth and metabolism of Streptomyces aureofaciens}; Meng C et al.; Vitreoscilla hemoglobin gene was expressed in S . aureofaciens through promoter of tetracycline resistance gene . Characteristics of S . aureofaciens growth and metabolism were studied in 1 m3 fermenter . In high dissolved oxygen concentrations, expression of hemoglobin gene had little effect on growth and metabolism of S . aureofaciens, and there were no obvious differences between engineering strain and control . The chlortetracycline of engineering strain was 22905 u/mL and of control was 22896 u/mL respectively . Under conditions of low dissolved oxygen, expression of hemoglobin enhanced growth, maintenance of energetic mycelium configuration and chlortetracycline yield of S . aureofaciens: the mycelium concentrations of engineering strain were more about 5%-10% and yield was more 11.4% than control.

Wei Sheng Wu Xue Bao, 2002 Feb, 42(1), 81 - 7
{Production of D-pantolactone hydrolase by Fusarium moniliforme SW-902}; Tang Y et al.; Fusarium moniliforme SW-902, a high-yielding strain for producing D-pantolactone hydrolase was screened . The conditions for enzyme formation of the strain were studied . The suitable carbon source of the medium was glycerol and the suitable nitrogen source was peptone . The optimum temperature and initial pH for enzyme formation were 26 degrees C and pH8.0 respectively . When the organism was cultured in the 60 L-fermentor under the optimum conditions for 45 h, about 7.18 g dry cells/L and 0.92 IU/g dry cells weight were obtained.

Biotechnol Bioeng, 2003 Mar 30, 81(7), 818 - 28
Modeling of yeast metabolism and process dynamics in batch fermentation; Sainz J et al.; Much is known about yeast metabolism and the kinetics of industrial batch fermentation processes . In this study, however, we provide the first tool to evaluate the dynamic interaction that exists between them . A stoichiometric model, using wine fermentation as a case study, was constructed to simulate batch cultures of Saccharomyces cerevisiae . Five differential equations describe the evolution of the main metabolites and biomass in the fermentation tank, while a set of underdetermined linear algebraic equations models the pseudo-steady-state microbial metabolism . Specific links between process variables and the reaction rates of metabolic pathways represent microorganism adaptation to environmental changes in the culture . Adaptation requirements to changes in the environment, optimal growth, and homeostasis were set as the physiological objectives . A linear programming routine was used to define optimal metabolic mass flux distribution at each instant throughout the process . The kinetics of the process arise from the dynamic interaction between the environment and metabolic flux distribution . The model assessed the effect of nitrogen starvation and ethanol toxicity in wine fermentation and it was able to simulate fermentation profiles qualitatively, while experimental fermentation yields were reproduced successfully as well .

Biotechnol Bioeng, 2003 Mar 30, 81(7), 753 - 67
Combined transcriptome and proteome analysis of Escherichia coli during high cell density culture; Yoon SH et al.; Combined transcriptome and proteome analysis was carried out to understand metabolic and physiological changes of Escherichia coli during the high cell density cultivation (HCDC) . The expression of genes of TCA cycle enzymes, NADH dehydrogenase and ATPase, was up-regulated during the exponential fed-batch period and was down-regulated afterward . However, expression of most of the genes involved in glycolysis and pentose phosphate pathway was up-regulated at the stationary phase . The expression of most of amino acid biosynthesis genes was down-regulated as cell density increased, which seems to be the major reason for the reduced specific productivity of recombinant proteins during HCDC . The expression of chaperone genes increased with cell density, suggesting that the high cell density condition itself can be stressful to the cells . Severe competition for oxygen at high cell density seemed to make cells use cytochrome bd, which is less efficient but has a high oxygen affinity than cytochrome bo(3) . Population cell density itself strongly affected the expression of porin protein genes, especially ompF, and hence the permeability of the outer membrane . Expression of phosphate starvation genes was most strongly up-regulated toward the end of cultivation . It was also found that sigma(E) (rpoE) plays a more important role than sigma(S) (rpoS) at the stationary phase of HCDC . These findings should be invaluable in designing metabolic engineering and fermentation strategies for the production of recombinant proteins and metabolites by HCDC of E . coli .

Yeast, 2003 Feb, 20(3), 207 - 19
Transcription patterns of PMA1 and PMA2 genes and activity of plasma membrane H+-ATPase in Saccharomyces cerevisiae during diauxic growth and stationary phase; Fernandes AR et al.; PMA1 and PMA2 genes encode Saccharomyces cerevisiae plasma membrane H(+)-ATPase (PM-H(+)-ATPase), an enzyme with critical physiological roles both in the absence or presence of environmental stress . The two PM-H(+)-ATPase isoforms differ in their biochemical characteristics but, under all the growth conditions that were examined so far, PMA2 expression is negligible and Pma1p accounts for practically the totality of cell PM-H(+)-ATPase activity . In the present work, we have compared gene expression levels and activity of this proton pump in yeast cells cultivated under fermentative or respiratory growth and under carbon starvation . The expression levels of both PMA1 and PMA2 genes were consistently higher (2.5-4.5-fold) in cells cultivated under respiratory metabolism (in ethanol-based medium or after the diauxic shift), than in cells cultivated under fermentative metabolism (during the full period of growth in a medium where glucose is not the limiting nutrient or during the first period of diauxic growth in low-glucose-based medium) . The moderate upregulation of PMA1 and PMA2 transcription in cells grown on ethanol compared with those grown on glucose was reflected in the increased content and activity of PM-H(+)-ATPase . In diauxic growth, during transition to stationary phase after ethanol depletion, a further strong activation (eight-fold) of PMA2 gene transcription was observed . Although PMA2 transcription still remains quite below (20-fold) PMA1 transcription, this is the first environmental condition, identified so far, that leads to a significant PMA2 expression, suggesting that this PM-H(+)-ATPase isoform may play some role during carbon starvation .

Eur J Clin Nutr, 2002 Dec, 56 Suppl 4, S50 - 5
Fermentation, fermented foods and lactose intolerance; Solomons NW; Lactose (milk sugar) is a fermentable substrate . It can be fermented outside of the body to produce cheeses, yoghurts and acidified milks . It can be fermented within the large intestine in those people who have insufficient expression of lactase enzyme on the intestinal mucosa to ferment this disaccharide to its absorbable, simple hexose sugars: glucose and galactose . In this way, the issues of lactose intolerance and of fermented foods are joined . It is only at the extremes of life, in infancy and old age, in which severe and life-threatening consequences from lactose maldigestion may occur . Fermentation as part of food processing can be used for preservation, for liberation of pre-digested nutrients, or to create ethanolic beverages . Almost all cultures and ethnic groups have developed some typical forms of fermented foods . Lessons from fermentation of non-dairy items may be applicable to fermentation of milk, and vice versa.

Eur J Clin Nutr, 2002 Dec, 56 Suppl 4, S27 - 33
The effect of fermented milk on interferon production in malnourished children and in anorexia nervosa patients undergoing nutritional care; Solis B et al.; For several years cytokine production has been associated with infections but it was not suspected that some types of food could also induce cytokines, even in a state of non-infection . Lactic bacteria can induce interferon (IFN) production in human healthy subjects, thus, a better protection against infections would be expected . Therefore, we planned to evaluate the effect of two diets including yoghurt or milk on IFN-gamma production during nutritional recovery in two different situations of malnutrition: (1) children with diarrhoea; and (2) patients with anorexia nervosa (AN) . Both the diet including yoghurt of that including milk seemed to increase IFN-gamma production at the end of nutritional recovery in the malnourished children with diarrhoea . The significance of interferon production and the lymphocyte subset increase should be explored to know if a better resistance against pathogens is related to them . Regulation of intestinal absorption and moderate stimulation of interferon production make the yoghurt-based diet a good choice in the nutritional care of children . In the same way, an increase in the IFN-gamma production was observed in AN patients consuming yoghurt . This increase of IFN-gamma production could be considered a biological marker to detect the effect of probiotics on the immune response, especially in the improvement of a deficient nutritional status.

Proc Natl Acad Sci U S A, 2003 Feb 4, 100(3), 825 - 32 Epub 2003 Jan 29.
Engineering the metabolism of Escherichia coli W3110 for the conversion of sugar to redox-neutral and oxidized products: homoacetate production; Causey TB et al.; Microbial processes for commodity chemicals have focused on reduced products and anaerobic conditions where substrate loss to cell mass and CO(2) are minimal and product yields are high . To facilitate expansion into more oxidized chemicals, Escherichia coli W3110 was genetically engineered for acetate production by using an approach that combines attributes of fermentative and oxidative metabolism (rapid growth, external electron acceptor) into a single biocatalyst . The resulting strain (TC36) converted 333 mM glucose into 572 mM acetate, a product of equivalent oxidation state, in 18 h . With excess glucose, a maximum of 878 mM acetate was produced . Strain TC36 was constructed by sequentially assembling deletions that inactivated oxidative phosphorylation (deltaatpFH), disrupted the cyclic function of the tricarboxylic acid pathway (deltasucA), and eliminated native fermentation pathways (deltafocA-pflB deltafrdBC deltaldhA deltaadhE) . These mutations minimized the loss of substrate carbon and the oxygen requirement for redox balance . Although TC36 produces only four ATPs per glucose, this strain grows well in mineral salts medium and has no auxotrophic requirement . Glycolytic flux in TC36 (0.3 micromol.min(-1).mg(-1) protein) was twice that of the parent . Higher flux was attributed to a deletion of membrane-coupling subunits in (F(1)F(0))H(+)-ATP synthase that inactivated ATP synthesis while retaining cytoplasmic F(1)-ATPase activity . The effectiveness of this deletion in stimulating flux provides further evidence for the importance of ATP supply and demand in the regulation of central metabolism . Derivatives of TC36 may prove useful for the commercial production of a variety of commodity chemicals.

Can J Microbiol, 2002 Nov, 48(11), 965 - 70
Increasing fermentation efficiency at high sugar concentrations by supplementing an additional source of nitrogen during the exponential phase of the tequila fermentation process; Arrizon J et al.; In the tequila industry, fermentation is traditionally achieved at sugar concentrations ranging from 50 to 100 g x L(-1) . In this work, the behaviour of the Saccharomyces cerevisiae yeast (isolated from the juices of the Agave tequilana Weber blue variety) during the agave juice fermentation is compared at different sugar concentrations to determine if it is feasible for the industry to run fermentation at higher sugar concentrations . Fermentation efficiency is shown to be higher (above 90%) at a high concentration of initial sugar (170 g x L(-1)) when an additional source of nitrogen (a mixture of amino acids and ammonium sulphate, different than a grape must nitrogen composition) is added during the exponential growth phase.

Wei Sheng Wu Xue Bao, 1999 Aug, 39(4), 355 - 61
{Fermentation conditions for production of glutathione by recombinant Escherichia coli}; Li Y et al.; The fermentation conditions for production of glutathione by recombinant E . coli were investigated, the addition of certain materials, such as yeast extract, precusor amino acids and ATP were mainly focused on . The results showed that the addition of precusor amino acids and ATP could promote intracellular GSH accumulation . DCW and intracellular GSH content would be 24% and 1.4 times higher than that of fermentation without additions when 2.0 g/L ATP and 9 mmol/L precusor amino acids were added at the begining and 12 h of fermentation, respectively . By using the optimized combination of additives obtained from orthogonal experiments, the maximal DCW and total GSH in broth could be improved 10% and 26% higher than the best results in orthogonal experiments, respectively . Based on the analysis of glucose utilizing ability of this strain, an exponential fed-batch culture process was conducted . DCW and total GSH in broth culd be 8.3 and 4.6 times higher than that of shaking flask culture and finally reached 80 g/L and 880 mg/L, respectively.

Wei Sheng Wu Xue Bao, 1999 Aug, 39(4), 350 - 4
{Screening of acidic xylanase producing strain and studies on its enzyme production conditions}; Chen H et al.; From 150 fungal strains, the authors found 8 strains contained mainly of xylanase activity over 100 U/mL in which the No . 149 strain was the highest xylanase producer . Which tentatively identified as Aspergillas niger . The appropriate medium composition was as follows: wheat bran hemicellulose 4%; NaNO3 1%; wheat bran 1% prepared in Mandels nutritional solution without (NH4)2SO4 and urea . After cultivated in shake-flask at 28 degrees C-32 degrees C for 60 h, the activity reached the highest value of 357.2 U/mL . The optimum pH of xylanase was 4.6 and it was stable at pH3-11 . The fermented broth of strain 149 contained in addition to xylanase (relative activity 100) also included amylase(1.8), mannanase(0.98), beta-xylosidase(0.94) and cellulase(0.17).

Wei Sheng Wu Xue Bao, 1999 Dec, 39(6), 495 - 502
{Cloning of mitomycin C resistance gene from Streptoverticillium caespitosum ATCC27422 and studying its function}; Huang J et al.; Streptoverticillium caespitosum is the major producer of anti-cancer drug mitomycin . It demonstrated that there exists a strong restriction system for the transformation of foreign DNA in this strain . Using the blocked mutant S-6 which is sensitive to mitomycin C (MC) and does not produce mitomycin as a recepient and plasmid pIJ699 as vector, the gene library of the strain ATCC27422 was constructed . A clone of mitomycin C resistance gene(mcr) was obtained by screening from this library . The mitomycin C resistance gene cloned on the 6.2 kb fragment of plasmid pLX5 could be expressed in the Strptonmyces lividans TK54 . It was the first time to introduce successfully the plasmid pLX5 into the wild type strain ATCC27422 by electroporation technique . And the transformant's resistance to mitomycin C has risen greatly: the MIC has gone up from 200 micrograms/ml to more than 1000 micrograms/ml . The result of fermentation showed that the yield of mitomycin of ATCC27422 harboring plasmid pLX5 was more than the wild strain ATCC27422's . It suggested that there is some relationship between the level of the resistance to mitomycin C and its biosynthesis.

Wei Sheng Wu Xue Bao, 1999 Oct, 39(5), 478 - 82
{Progress in research of Vitreoscilla hemoglobin and Vitreoscilla hemoglobin gene}; Yu H et al.; Vitreoscilla is a Gram-negative obligate aerobic bacterium, whose most important property is the ability to produce the Vitreoscilla hemoglobin (VHb) . VHb is encoded by a single gene called Vitreoscilla hemoglobin gene (vgb) containing the natural promoter, and the expression of vgb gene is regulated by dissolved oxygen at the level of transcription . The oxygen binding VHb participate in the metobolism process of cells and make them grow well in the hypoxic habitats . It also facilitates the bacteria growing and the potein producing obviously . The potential application prospect for the oxygen regulated promoter of vgb and the physiological function of VHb in the fermentation industry is reviewed.

Wei Sheng Wu Xue Bao, 1999 Oct, 39(5), 454 - 60
{Breeding of yeast for beer manufacturing by inactivated protoplast fusion}; Zhou D et al.; QSB-XI6, one of fusants, was obtained by PEG-induced fusion between UV-inactivated protoplasts of Saccharomyces cerevisiae LQ16 (Ile-, Datr) and heat-inactivated protoplasts of S . cerevisiae QSB7(Ala-, H2S-) . The cell volume, bicmass, genotype and DNA content of the fusant were measured in comparison with of its parents and the results showed that QSB-XI6 was a fusant . The fermentation ability and the flocculent capacity of this fusant were higher than those of its parents . The diacetyl content of the beer produced by the fusant was measured and some other compositions of the beer was also analysed by gas chromatography . The sensory evaluation of the beer was much better and the flavour was distinctive . The results of many production trials in succession showed that all good characteristics of QSB-XI6 were stable.

Wei Sheng Wu Xue Bao, 1999 Apr, 39(2), 174 - 7
{The effects on ethanol fermentation of Saccharomyces cerevisiae by adding Ca2+ and inositol}; Zhao B et al.; In this paper we report the effects of Ca2+ and inositol upon growth, ethanol fermentation and ethanol tolerance of Saccharomyces cerevisiae . The result shows that by adding inositol has notable effects on growth and ethanol tolerance of Saccharomyces cerevisiae (p < 0.05), but has little effects on ethanol fermentation (p = 0.3933) . By adding Ca2+ has remarkable effects on ethanol fermentation and ethanol tolerance (p < 0.05), but has little effect on yeast growth (p = 0.1544) . By means of adding Ca2+ and inositol has more effects on the growth, ethanol fermentation and ethanol tolerance than the condition of only adding Ca2+ or inositol separately.

Wei Sheng Wu Xue Bao, 1999 Apr, 39(2), 148 - 53
{Solid-state fermentation with Penicillium sp . PT95 for carotenoid production}; Han J et al.; A preliminary study on solid-state fermentation (SSF) with Penicillium sp PT95 for carotenoid production was performed . The results showed that the production of carotenoid in sclerotia of PT95 was more efficient in corn meal medium than in either wheat bran medium or cottonseed hull medium . Addition of nitrogen and carbon sources as well as vegetable oil to media was required for increasing the dry weight of sclerotia and carotenoid yield . Among several tested compounds for nitrogen and carbon sources, sodium nitrate and maltose were the best . Through orthogonal experiments, the optimum culture medium was obtained by supplement of NaNO3 3g, maltose 10 g, soybean oil 2.5 g to per liter of salt solution . Under the optimum culture conditions, the sclerotia dry weight increased from 5.36 g to 9.70 g per 100 g dry substrate, the carotenoid yield from 2149 micrograms to 5260 micrograms per 100 g dry substrate, the proportion of beta-carotene in carotenoids from 61.4% to 71.3%.

Wei Sheng Wu Xue Bao, 1999 Feb, 39(1), 68 - 74
{A new osmotolerant and glycerol-highly-producing species--Candida glycerolgenesis Zhuge sp . nov.}; Wang Z et al.; The strain WL2002-5 isolated from natural samples and overproduced glycerol from glucose was identified systematically . The WL2002-5 fermented glucose and weakly fermented sucrose; assimilated glucose, sucrose, ethanol, weakly assimilated glycerol and citrate, did not assimilated nitrate, erythritol, arabinitol, mannitol . It grew well on the slopes containing 500 g/L of glucose or 10 mL/L acetate or at the temperature up to 40 degrees C . The minimum water activity for its growth was 0.890 . It had a negative reaction with DBB and did vegetative reproduction by budding and easily formed Candida-type pseudohyphae but had no sexual reproduction . The molecular weight of its mitochondrial DNA was 20 kb . We conclude that WL2002-5 is a new species of Candida and nominated it Candida glycerolgenesis Zhuge sp . nov..

J Exp Bot, 2003 Feb, 54(383), 739 - 47
Antisense SNF1-related (SnRK1) protein kinase gene represses transient activity of an alpha-amylase (alpha-Amy2) gene promoter in cultured wheat embryos; Laurie S et al.; A DNA fragment corresponding to part of an SNF1 (sucrose non-fermenting-1)-related protein kinase (SnRK1) transcript was amplified by a polymerase chain reaction (PCR) from a wheat (Triticum aestivum) endosperm cDNA library . It was used to construct a chimaeric gene, pUasSnRKN, comprising a ubiquitin promoter, the SnRK1 PCR product in the antisense orientation and the nopaline synthase (Nos) gene terminator . This construct was used in transient gene expression experiments in cultured wheat embryos together with a series of reporter gene constructs . These included the wheat alpha amylase gene alpha-Amy2 promoter with UidA (Gus) coding region (palpha2GT), rice actin promoter with Gus (pActIDGus), ubiquitin promoter with Gus (pAHC25) and actin promoter with green fluorescent protein (GFP) gene (pAct1Is-GFP1) . All of the reporter genes were found to be active when bombarded into scutellae isolated from immature grains at 25 d post-anthesis and incubated on MS medium for 24 h prior to bombardment . However, co-bombardment of palpha2GT with equimolar amounts of pUasSnRKN resulted in no detectable Gus activity, indicating that the antisense SnRK1 construct repressed the alpha-Amy2 promoter . Co-bombardment with pUasSnRKN had no effect on the activity of the other promoters used in the study . A triple bombardment with palpha2GT, pAct1Is-GFP-1 and pUasSnRKN resulted in clear green fluorescence, indicating that the bombarded cells were still viable, but no Gus activity . RT-PCR analysis showed clearly that the antisense SnRK1 gene was expressing . Northern and RT-PCR analyses confirmed that SnRK1 and both alpha-amylase genes, alpha-Amy1 and alpha-Amy2, are expressed in cultured wheat embryos harvested from grain 25 d post-anthesis . Expression of alpha-Amy1 and alpha-Amy2 was up-regulated by sugar starvation.

Water Res, 2003 Mar, 37(5), 1118 - 24
Enhanced settleability and dewaterability of fungal treated domestic wastewater sludge by liquid state bioconversion process; Alam MZ et al.; A study was conducted to evaluate the settleability and dewaterability of fungal treated and untreated sludge using liquid state bioconversion process . The fungal mixed culture of Aspergillus niger and Penicillium corylophilum was used for fungal pretreatment of wastewater sludge . The fungal strains immobilized/entrapped on sludge particles with the formation of pellets and enhanced the separation process . The results presented in this study showed that the sludge particles (pellets) size of 2-5mm of diameter were formed with the microbial treatment of sludge after 2 days of fermentation that contained maximum 33.7% of total particles with 3-3.5mm of diameter . The settling rate (measured as total suspended solids (TSS) concentration, 130 mg/l) was faster in treated sludge than untreated sludge (TSS concentration, 440 mg/l) after 1 min of settling time . In 1 min of settling operation, 86.45% of TSS was settled in treated sludge while 4.35% of TSS settled in raw sludge . Lower turbidity was observed in treated sludge as compared to untreated sludge . The results to specific resistance to filtration (SRF) revealed that the fungal inoculum had significant potentiality to reduce SRF by 99.8% and 98.7% for 1% w/w and 4% w/w of TSS sludge, respectively . The optimum fermentation period recorded was 3 days for 1% w/w sludge and 6 days for 4% w/w sludge, respectively, for dewaterability test.

Water Res, 2003 Mar, 37(5), 1091 - 9
Recovery of ammonium sulfate from fermentation waste by electrodialysis; Lee HJ et al.; Electrodialysis experiments of the lysine fermentation waste were performed to generate demineralized feed and ammonium sulfate, which can be utilized as a fertilizer and an animal feed, respectively . The electrodialysis performances were compared for different ion exchange membranes in terms of ammonium sulfate removal rate, resistance and conductivity change . Analysis of fouling phenomena revealed that organics fouled ion exchange membranes reversibly in electrodialysis of the fermentation waste . In this study, mitigation of membrane fouling with the pulsed electric field was examined for the electrodialysis of the fermentation waste containing strong foulants . The half-wave power reduced membrane fouling significantly . For a quantitative measure of the membrane fouling tendency, a membrane fouling index for electrodialysis was used . This study showed the potential use of pulsed power as an effective fouling mitigation method for the electrodialysis of fermentation waste.

Anticancer Res, 2002 Nov-Dec, 22(6C), 4215 - 9
Stimulation of the maturation of dendritic cells in vitro by a fermented mistletoe extract; Stein GM et al.; BACKGROUND: Dendritic cells (DC) play a key role during the initiation of specific immune responses . In cancer patients, however, an alteration of their function was observed . In our investigation we analysed the influence of a fermented mistletoe extract often used for adjuvant treatment of cancer patients on the generation and maturation of DC . MATERIALS AND METHODS: Monocytes from healthy individuals were incubated with a fermented mistletoe extract in the presence or absence of GM-CSF/IL-4 . Surface marker expression was measured by flow cytometry . RESULTS: While there was no relevant effect on the generation of DC in the absence or presence of GM-CSF/IL-4 in 5-day cultures, the mistletoe extract significantly stimulated the maturation of pre-generated immature DC, as evidenced by a heightened expression of CD83 . Like the positive control TNF-alpha, the mistletoe extract significantly activated CD80 and CD86 as well as HLA class I and II molecules on these cells . CONCLUSION: Our data clearly demonstrate an influence of the mistletoe extract on the maturation of DC, but it remains to be elucidated whether the function of DC is also activated and, especially, whether this effect can be observed in tumour patients as well.

Wei Sheng Wu Xue Bao, 2001 Dec, 41(6), 750 - 2
{Selection of beta-glucanase-producing Trichoderma köningii T199 and its fermentation conditions}; Shi J et al.; A mutant strain T199 producing about 8 times as much beta-glucanase as its parent strain trichoderma koningii T3 was obtained by treatment with ultraviolet light and N-methyl-N-nitro-N-nitrosoguanidine . Fermentation was conducted in 250 mL flask, each containing 30 mL of medium consisted of 5% corn cob powder, 3% wheat bran and 1.4% nitrogen source No . 10 ((NH4)2SO4 10%, Peptone 20%, Yeast extract 15%) . The optima culture conditions were as below: initial pH 5.0, 30 degrees C, shaking speed 280 r/min, and cultivation time 5 d . Enzyme activity toward CMC-Na, lichenin, laminarin and barley beta-glucan at pH 5.0 and 60 degrees C for 10 min were 300, 1100, 12 and 1600 IU/mL, respectively . The optima pH and temperature for enzyme action toward barley beta-glucan were pH 5.0 and 70 degrees C, respectively . The enzyme was stable under below 50 degrees C and at pH 4.5-6.5.

Wei Sheng Wu Xue Bao, 2001 Dec, 41(6), 709 - 15
{Production of vitamin C precursor--2-keto-L-gulonic acid from D-sorbitol by mixed culture of microorganisms}; Yin G et al.; Gluconobacter oxydans SCB329 only produce a little amount of 2-Keto-L-gulonic acid(2-KLG) from D-Sorbitol when growing alone; while Gluconobacter sp . SCB110 can transform D-Sorbitol to L-Sorbose and can not produce 2-KLG . 2-Keto-L-gulonic acid, the precursor of L-Ascorbic acid (Vitamin C) synthesis, was prepared directly with a high efficiency from D-Sorbitol by mixed culture of microorganism, which comprised Gluconobacter sp . SCB110 and Gluconobacter oxydans SCB329 . The fermentation product from the mixed culture broth in the D-Sorbotol-containing medium was identified as 2-Keto-L-gulonic acid by HPLC, elementary analysis and infra-red adsorption spectrum.

Wei Sheng Wu Xue Bao, 2001 Dec, 41(6), 704 - 8
{Study on the organic acids metabolism in Candida glycerolgenesis under different fermentation conditions}; Jin H et al.; The organic acids in Candida glycerolgenesis fermentation medium affected both the glycerol yield and quality . When other fermentation condition was constant, the total acidity in medium increased when the volumetric ratio of medium to flask and corn steep liquid concentration were increased . When the volumetric ratio of medium to flask was 0.20 and the corn steep liquid concentration was 8 g/L, there have high yield of pyruvate and lactate produced at growth phase, 4.1 g/L and 1.0 g/L, respectively, which are two times than control sample at usual fermentation condition, hence, the glycerol yield was lower . However, the volumetric ratio of medium to flask was 0.08 and the corn steep liquid concentration was 4 g/L, the yield of pyruvate and lactate were lower and the glycerol yield was higher, but the acetate could be accumulated to 2.6 g/L . The organic acids also could be metabolized in final fermentation process, such as the initial glucose concentration was 100 g/L, after the glucose was completely utilized, both the glycerol and organic acids concentration decreased, and pyruvate or lactate could be converted to acetate in following fermentation process . In addition, the additive also can change the production of organic acids, with the addition of 1% oleic acid or VB1 the acetate concentration could be decreased and increased the production of pyruvate, hence, improved the yield of glycerol . While the fluoro-pyruvate or sulphite were added in medium, the acetate was increased largely and ketoglutarate was decreased accordingly, also increase the glycerol yields about 20%.

Wei Sheng Wu Xue Bao, 2001 Oct, 41(5), 635 - 41
{Isolation and fermentation conditions of strains producing 1-phenyl-2-amino-ethanol alcohol dehydrogenase}; Wang J et al.; A Arachnia sp . P163 producing alcohol dehydrogenase which is able to reduce aminoacetophenone to R-1-phenyl-2-aminoethanol was obtained from soil and cultures . The maximum activity of enzyme was produced by the LB medium containing 1% sodium citrate and peptone, 0.1% phenylaminoethanol as inducer at 30 degrees C for 48 hs.

Wei Sheng Wu Xue Bao, 2001 Oct, 41(5), 548 - 52
{Introduction of Vitreoscilla hemoglobin gene in a recombinant E . coli for PHB production}; Yu H et al.; Vitreoscilla hemoglobin gene (vgb) was introduced into the recombinant Escherichia coli VG1(pTU14) for production of poly-beta-hydroxybutyrate (PHB), in order to enhance the oxygen uptake of strain from molecular level and to solve the problem of oxygen limitation in process of fermentation . The carbon monoxide difference spectra analysis of Vitreoscilla hemoglobin (VHb) showed that vgb could be successfully expressed in VG1 (pTU14), and this expression was under the regulation of dissolved oxygen (DO) . The higher amounts of VHb could be got correspond to the lower level of DO . Because of the introduction of vgb, cell growth and PHB accumulation of VG1 (pTU14) were all strongly promoted during the process of batch culture.

Wei Sheng Wu Xue Bao, 2001 Oct, 41(5), 530 - 5
{Studies on the genome size and structure of Gluconobacter oxydans SCB329}; Lin W et al.; In the "Two-step fermentation" of Vitamin C synthesis, Gluconobacter oxydans SCB329 is responsible for the production of 2-keto-L-gulonic acid (2-KLG), which is an important precuror of vitamin C synthesis . The intact chromosome was prepared from logarithmic phase cells by agaraseembedded method and was analysized by restriction endonucleases and contour-clamped homogeneous electric field pulsed-field gel electrophoresis (PFGE) . Spe I (5-ACTAGT) produced 24 fragments, ranging in size from 10 to 320 kilobases (kb) . Xba I (5-TCTAGA) yielded 40 fragments (4 to 200 kb) . A total genome size of approximately 2,700 kb was determined by summing the fragment length . Analysis of the entire genome of SCB329 by PFGE revealed that the genome of SCB329 consist of a chromosome which is 2,500 Kb in length and a large plasmid which is 245 kb . After linearization of the DNA by DNase I and S1 nuclease, in contrast with the band which can not be viewed, the band of chromosome and plasmid were appeared, this suggest that structure of the chromosome and the plasmid were circular.

Bioseparation, 2001, 10(6), 379 - 87
Molecular imprinting and solid phase extraction of flavonoid compounds; Weiss R et al.; Molecularly imprinted polymers (MIPs) for quercetin have been successfully prepared by a thermal polymerization method using 4-vinylpyridine (4-VP) and ethylene glycol dimethacrylate (EDMA) as functional monomer and cross-linker, respectively . The obtained molecularly imprinted polymers were evaluated by HPLC using organic eluents, with respect to their selective recognition properties for quercetin and related compounds of the flavonoid class . Two equivalent control polymers, a blank polymer and a polymer imprinted with a structural analogous template, were synthesized, in order to confirm the obtained results . Furthermore, preliminary experiments confirm the applicability of the prepared MIPs for solid phase extraction (SPE), as rapid and facile clean-up of wine samples for HPLC analysis is an envisaged field of application . The successful preparation of molecularly imprinted polymers for flavones provides an innovative opportunity for the development of advanced separation materials, with applications in the field of wine and fermentation analysis.

J Environ Qual, 2003 Jan-Feb, 32(1), 269 - 77
Methane emissions of beef cattle on forages: efficiency of grazing management systems; DeRamus HA et al.; Fermentation in the rumen of cattle produces methane (CH4) . Methane may play a role in global warming scenarios . The linking of grazing management strategies to more efficient beef production while reducing the CH4 emitted by beef cattle is important . The sulfur hexafluoride (SF6) tracer technique was used to determine the effects of best management practices (BMP) grazing compared with continuous grazing on CH4 production in several Louisiana forages during 1996-1998 . Cows and heifers (Bos taurus) grazed common bermudagrass {Cynodon dactylon (L.) Pers.}, bahiagrass (Paspalum notatum Flugge), and ryegrass (Lolium multiflorum Lam.) pastures and were wintered on bahiagrass hay with supplements of protein molasses blocks (PMB), cottonseed meal and corn (CSMC), urea and corn (URC), or limited ryegrass grazing (LRG) . Daily CH4 emissions were between 89 and 180 g d(-1) for young growing heifers and 165 to 294 g d(-1) for mature Simbrah cows . Heifers on "ad lib" ryegrass in March and April produced only one-tenth the CH4 per kg of gain as heifers on LRG of 1 h . Using BMP significantly reduced the emission of CH4 per unit of animal weight gain . Management-intensive grazing (MIG) is a BMP that offers the potential for more efficient utilization of grazed forage crops via controlled rotational grazing and more efficient conversion of forage into meat and milk . Projected CH4 annual emissions in cows reflect a 22% reduction from BMP when compared with continuous grazing in this study . With the BMP application of MIG, less methane was produced per kilogram of beef gain.

Wei Sheng Wu Xue Bao, 1998 Aug, 38(4), 313 - 7
{Isolation of a lipase-producing Pseudomonas strain and optimization of its fermentation conditions}; Gao X et al.; A lipase-producing bacterium strain was isolated from soil and was identified as Pseudomonas sp. . Its lipase yield was improved 2.25-fold by combined treatment of UV irradiation and NTG . The lipase fermentation condition for the mutant strain was optimized with Plackett-Burman design and Response Surface Analysis (RSA), and the formula of the optimum medium suitable for industrial scale fermentation was thereby established . A maximum yield of 87.5 U/ml was obtained.

Wei Sheng Wu Xue Bao, 1998 Oct, 38(5), 371 - 5
{Growth of Methylomonas Z201 cells and production of epoxypropane in monophasic and biphasic fermentation systems}; Xia S et al.; The growth of Methylomonas Z201 cells and production of epoxypropane in mono- and biphasic fermentation systems were studied . In monophasic fermentation systems, the inhibitions of propene and epoxypropane on the growth of Methylomonas Z201 cells were observed and the concentration of epoxypropane reached 1.3 mmol/L . In biphasic fermentation systems, hexadecane acted as the "reservoir" of growth substrate (methane) and reactants (propene, molecular oxygen), the decrease of propene and epoxypropane in aqueous phase reduced the inhibition effect of propene and epoxypropane on the growth of cells, the concentrations of epoxypropane in both water phase and hexadecane phase reached 1.7 mmol/L and 2.6 mmol/L . In both monophasic and biphasic fermentation systems, the operational stability of cells was enhanced compared to that of resting cells.

Wei Sheng Wu Xue Bao, 1998 Feb, 38(1), 44 - 51
{Degradation of phytic acid in rapeseed meal by two strains of molds}; Xiang W et al.; By using selective media, two strains of molds which are able to degrade phytic acid were isolated from soil samples . These molds are able to use inositol as the sole carbon and energy sources for growth . In broth cultures, the degradation rates of phytic acid by these molds were 74.4% and 95.0%, respectively . However, in solid-state fermentation which used rapeseed meal as a raw material, the degradation rate was about 40% . Several metal ions tested were able to enhance the degradation rate of phytic acid . Factors such as temperature, pH and moisture content which affect the growth of molds were also explored . These two strains of molds were initially identified as Paecilomyces sp . and Penicillium sp. . They do not produce any detectable amount of aflatoxins.

Wei Sheng Wu Xue Bao, 1998 Jun, 38(3), 237 - 9
{Studies on the fermentation technology of microbiological preparation No.633}; Xia Z et al.; This paper reports the fermentation techmology of bacterial feed No.633 . A new fermentational technic, which was composed of fermentation medium, optimum pH, temperature, airate flow, pot pressure and fermentation time, was established.

Wei Sheng Wu Xue Bao, 2001 Jun, 41(3), 315 - 9
{Cloning and expression of VHB gene in D-arabitol producing yeast}; He P et al.; Recombinant plasmid pVgb-EX2 containing Vitreoscilla hemoglobin gene vgb and formaldehyde resistant gene SFA1 was constructed and transformed into D-arabitol producing yeast strain Saccharomyces sp . X-62 . The fact that the amount of VHb in transformant cells was considerably higher than that in control cells indicated that gene vgb was expressed in transformant cells . D-arabitol productivity and yield of fermentation by transformants were improved . The most improvement of D-arabitol productivity in repeat experiments reached 27.3% . It appeared that the fermentation productivity of D-arabitol was relative to the amount of VHb in cells under experimental conditions.

Wei Sheng Wu Xue Bao, 2000 Dec, 40(6), 610 - 3
{Construction of cDNA library of Mortierella and screening of delta 9 fatty acid desaturase cDNA sequence}; Zhang Y et al.; Mortierella species have potential for fermentative production of polyunsaturated fatty acids including gamma-linolenic acid, Arachidonic acid and EPA, etc . In order to clone genes encoding enzymes in the unsaturated fatty acid biosynthetic pathway, cDNA library of Mortierella was constructed using lambda gt 10 vector . Using cDNA encoding conserved region of delta 9 fatty acid desaturase gene as probe, Mortierella cDNA library was screened . After two rounds of screening one positive clone was identified which has insert length of larger than 1.6 kb.

Wei Sheng Wu Xue Bao, 2001 Apr, 41(2), 229 - 33
{A study on the breeding of new Ganoderma varieties by UV induced mutagenesis}; Li G et al.; UV induced mutagenesis of Ganoderma's protoplast was carried out . By crude screen and careful screen, we obtained two mutants, 43020# and 43026$, whose dry weight and polysaccharides contents are higher than original strain . Results of continued cultivation for 10 generation on PDA slants and successive tests of shaking bottles, and successive 3 time pilot scale tests at 3 tons, showed that these two mutants are excellent strains with steady properties, high production and high content of polysaccharides . This study has provided a rapid and effective method for breeding of Ganoderma varieties which is suitable to industrial fermentation.

Wei Sheng Wu Xue Bao, 2000 Aug, 40(4), 440 - 3
{Screening and fermentation conditions of a bacterial strain over-producing xylanase}; Xu Z et al.; A xylanase producer WXULI-11 was isolated and identified as Pseudomonas sp . By combination treatment of WXULI-11 with UV and NTG, a mutant WLUN024 was obtained and its enzyme activity reached 354 IU/mL in 36 h by shaking flask . The mechanism of its xylanase production was primarily studied . The results showed that this enzyme was induced by xylan, xylose and some other xylosic material, while xylose had "two-way effect" on xylanase synthesis . Effects of nitrogen and other factors on enzyme production were also investigated.

Wei Sheng Wu Xue Bao, 2000 Apr, 40(2), 217 - 20
{Isolation, purification and bioactivities of exopoly saccharides from fermented broth of Ganoderma lucidum}; Li P et al.; The exopolysaccharides of Ganoderma lucidum(GLEP) extracted from the fermentation broth after removing protein by Sevage and protease digestion procedures, were applied to a column of DEAE-cellulose(OH- form), and eluted stepwise with distilled water, sodium hydrogen carbonate (0.1 mol/L, 0.3 mol/L, 0.5 mol/L successively) and 0.1 mol/L sodium hydroxide . Five fractions were obtained, and the main fraction was known as GLEP-I, furthermore subjected to chromatography on a column of SepharoseC1-6B, eluted at a flow rate of 30 mL/(cm2.h), the relative viscosity of sample solution of 1.5 . Two fractions, GLEP-IFr1 and GLEP-IFr2 with a ratio of 3.8:1, were obtained . Molecular weight of GLEP-IFr1 and GLEP-IFr2 was estimated to be 38,000 and 22,000 Dalton respectively by Membrane Osmometer . The animal test showed that GLEP-IFr1 could inhibited the growth of Sarcoma 180 tumor in mice . The average inhibition ratio was 57.4% (i.p . 10 mg/kg for 10 days) . The result of immunological activity showed that GLEP-IFr1 could significantly improve macrophage cytophagy.

Wei Sheng Wu Xue Bao, 2000 Apr, 40(2), 180 - 7
{The key enzymes of metabolisms of glycerol in Candida glycerolgenesis}; Wang Z et al.; The specific enzymes for glycerol formation and dissimilation in Candida glycerolgenesis were studied . C . glycerolgenesis hardly assimilated glycerol without glucose and a little amount of glucose remarkably stimulated this dissimilation pathway and the activity and expression of mitochondrial glycerol-3-phosphate dehydrogenase was induced strongly by glycerol-3-phosphate and repressed by glucose metabolism . During glycerol production fermented by C . glycerolgenesis, the activity of cytoplasmic glycerol 3-phosphate dehydrogenase maintained at a higher level and existed two peaks of activity, which appeared at 36 h and 60 h, respectively . The activity level of the first activity peak determines glycerol productivity and becomes a key enzyme of glycerol formation . The activity of glycerol 3-phosphatase maintained at a higher level just between 18 h and 48 h and appeared on peak activity at 36 h . In that period, the activity of glycerol 3-phosphatase was up to more than two times of cytoplasmic glycerol 3-phosphate dehydrogenase and that was inosculated with the phase of rapid accumulation of glycerol during glycerol production by C . glycerolgenesis . In the phase of slowly accumulation of glycerol between 48 h and 72 h, the activity of glycerol 3-phosphatase decreased below that of cytoplasmic glycerol 3-phosphate dehydrogenase . So, cytoplasmic glycerol 3-phosphate dehydrogenase is a key enzyme of glycerol formation in the phase of rapid accumulation of glycerol and glycerol 3-phosphatase becomes a more effective factor after glycerol fermentation reached into slowly accumulation phase . In a conclusion, it is a fundamental for C . glycerolgenesis over-producing high level of glycerol that its cytoplasmic glycerol 3-phosphate dehydrogenase maintains at high activity and the activity of glycerol 3-phosphatase is much higher than that of cytoplasmic glycerol 3-phosphate dehydrogenase.

Wei Sheng Wu Xue Bao, 1998 Dec, 38(6), 484 - 6
{Selection of ningnanmycin overproduction strains by resistance of self-secondary metabolites}; Chen J et al.; The S . noursei var . xichangensis FR-5 protoplasts was treated by UV, NTG in sequence, and the mutated strains were selected by use of self-secondary metabolites, the microorganisms were isolated by fermentation after agarblock rational method . The seven strains of microorganisms which were one fold production higher than that of original strain were obtained from 122 mutants . The method should be a simple, high speed selection model of breed Ningnanmycin overproduction strains of microorganisms.

Wei Sheng Wu Xue Bao, 1998 Dec, 38(6), 449 - 53
{Nuclease P1 production by immobilized Penicillium citrinum cells}; Xia L; Mycelia of Penicillium citrinum were adsorbed and immobilized efficiently within porous polyurethane . Nuclease P1 produced by the immobilized cells were studied in shaking flasks . The suitable glucose and peptone contents in medium were 10 g/L and 1 g/L, respectively . And the shaking speed was 180-200 r/min . After 48 h fermentation, the nuclease P1 activity reached 513.3 U/ml, the productivity was 3.6 times higher than that of the free cells . The production cost is obviously reduced . In repeated batch fermentation, the immobilized cells kept the capacity after 28 batches for 56 days with an average nuclease P1 activity of 507.4 U/ml.

Wei Sheng Wu Xue Bao, 2000 Feb, 40(1), 100 - 4
{The selection of fluoroacetate-resistant mutant from E . coli MMR204 and its influence on the expression of heterologous GL-7ACA acylase}; Zhu T et al.; In the cultivation of gene engineered strain of Escherichia coli on glucose medium, excretion and accumulation of acetic acid inhibit not only cell growth but also the the expression of heterologous protein . It is obvious that the desirable host strain maintaining acetate at a low level is one of the approaches to increase the production of recombinant protein . The present article deals with the selection of mutants of E . coli DP19, DP8, which grow on the medium containing pyruvate as the sole carbon source in the presence of 50 mmol/L fluoroacetic acid . It is shown that mutant DP19 is defective in its phosphotransacetylase(PTA) activity and accumulates less acetate in the medium, while DP8 is defective in acetate kinase (ACK) and accumulates similar level of acetate comparing with its parent . Using pta- mutant E . coli DP19 as host, the expression of GL-7ACA acylase gene on the recombinant plasmid pMR24 is improved, and the yield of enzyme activity in flask fermentation is about twice as much as its parent.

Wei Sheng Wu Xue Bao, 2000 Feb, 40(1), 21 - 5
{A Carnobacterium-like organism isolated form a patient with multiple bacterial synergistic gangrene}; Xu J et al.; An atypical lactic-acid producing gram positive rod Y6 strain was studied in this report, which was isolated form clinic sample of a patient with multiple bacterial synergistic gangrene, and could not be identified by routine method . A 1.5 kb of 16S rDNA of Y6 strain was synthesized and sequenced . Comparative 16S rDNA sequence analyses revealed that stain Y6 is most closely related to the genus of Carnobacterium . The overall similarity value between Y6 strain and Carnobacterium species are 93% to 97% . The signature nucleotides in 16S rDNA primary sequence of strain Y6 and that of genus of Carnobacterium were identical . The biological features of Y6 strain are very similar to that of Carnobacterium, such as lactic acid as main end product of in PYG broth, no gas produced form fermentation of glucose, catalase negative, no motility . Data suggested that Y6 strain is very similar to the genus of Carnobacterium, of which no strain has been isolated form clinical sample so far . Based on the results obtained, we names Y6 strain as Carnobacterium-like organism.

Wei Sheng Wu Xue Bao, 2000 Oct, 40(5), 528 - 34
{The important role of vitamins in the over-production of pyruvic acid}; Li Y et al.; The effect of nicotinic acid, thiamine, pyridoxine, biotin and riboflavin on the production of pyruvic acid by Torulopsis glabrata WSH-IP303 with glucose as carbon source and NH4Cl as sole nitrogen source was investigated . By using orthogonal experiment method, thiamine was confirmed to be the most important factor affecting the production of pyruvic acid . Based on a certain concentration range of thiamine (0.01-0.015 mg/L), glucose consumption rate can be enhanced by increasing the concentration of nicotinic acid . When the concentration of nicotinic acid, thiamine, pyridoxine, biotin and riboflavin were 8, 0.015, 0.4, 0.04 and 0.1 mg/L, respectively, the concentration and yield to glucose of pyruvic acid reached 52.4 g/L and 0.525 g/g at 48 h in flask culture, respectively . Batch culture was conducted in a 2.5 L fermentor with initial glucose concentration of 120 g/L . By adopting the optimal concentration combination of vitamins, the concentration and yield to glucose of pyruvic acid reached 69.4 g/L and 0.593 g/g at 57.5 h, which were increased by 32.4% and 13% than the best results in flask culture, respectively.

Wei Sheng Wu Xue Bao, 2000 Oct, 40(5), 500 - 6
{Studies on the correlation between production of L-malic acid and some cytosolic enzymes in the L-malic acid producing strain Aspergillus sp . N1-14}; Zhou X et al.; The cytosol enzymatic study in the case of high L-malic acid(LMA) production of Aspergillus sp . N1-14' was reported . The activities of 4 kind enzymes that catalyse the CO2 fixation reactions have been detected, which are pyruvate carboxylase(PC), phosphoenolpyruvate carboxlase (PEPC), phosphoenolpyurvate carboxykinase(PCK) and malic enzyme(ME) . With the exception of ME, the linear correlation was found between activities of three carboxlases and the production rate of LMA . The activity of malate dehydrogenase(MDH) was at the level of 2-3 exponential higher than that of the other analysed enzymes, while the activity of succinate dehydrogenase(SDH) was much lower, and as a discrepancy, SDH was in a positive correlation to the content of LMA in fermenting slurry(r = 0.9252) . It is shown that the accumulated LMA acted as an activator of SDH . Through dynamic study, it is found that, in contrast with the slow and even increase of biomass, the content of cytosol protein(Cp) sharply fluctuated mainly due to the changes of aeration conditions . The data of the linear correlation coefficients(r) of activities of cytosol enzymes to Cp(PC r = 0.9563, PEPC r = 0.7688, PCK r = 0.7300, MDH r = 0.3920, SDH r = -0.2086) exhibited an inner law of protein synthesis . Experiment of increasing the amount of spore inoculum resulted in increase of LMA and decrease of SA . After fermenting 120 h in a 5 L stirred fermentor, with 3-fold of original spore inoculum 105.88 g/L of LMA was achieved, the overall productivity was 0.883 g/(L.h), the converting rate of glucose to LMA was 78.43% . This result supports the exist of a inner law of protein synthesis in the early period of LMA fermentation by Aspergillus sp . N1-14'.

Eur J Clin Nutr, 2003 Jan, 57(1), 1 - 17
Dairy products and colorectal cancer . A review of possible mechanisms and epidemiological evidence; Norat T et al.; OBJECTIVES: This review provides an overview of the principal hypotheses and epidemiological evidence of the possible links between colorectal cancer and intake of milk and/or dairy products . METHODS: The first section outlines the main hypotheses about the possible effect of calcium, vitamin D, fats and other milk components . The possible role of acid lactic bacteria in fermented products is also discussed . The second section is a summary of the published epidemiological evidence . The results on milk, cheese and yoghurt are summarized using a meta-analytical approach . The results of studies on calcium and vitamin D are briefly described . RESULTS: Case-control studies are heterogeneous and, on average, do not provide evidence of association between total intake of total dairy products, milk, cheese or yoghurt and colorectal cancer risk . The average result from cohort studies support the hypothesis of a protective effect of total dairy products (odds ratio (OR): 0.62; 95% confidence interval (CI): 0.52-0.74; P heterogeneity test: 0.93) and for milk (OR: 0.80; 95% CI: 0.68-0.95; P heterogeneity: 0.77) . No association was found between cheese (OR: 1.10; 95% CI: 0.88-1.36; P heterogeneity: 0.55) or yoghurt (OR: 1.03; 95% CI: 0.83-1.28; P heterogeneity: 0.69) in cohort studies . CONCLUSIONS: Cohort studies consistently found a protective effect of total dairy products and milk intake, but the evidence is not supported by case-control studies . No relationship was found with cheese or yoghurt intake . As the number of cohort studies is still limited, their results need to be confirmed by other prospective studies.

Biotechnol Appl Biochem, 2003 Jun, 37(Pt 3), 311 - 5
Biosynthesis of rifamycin SV by Amycolatopsis mediterranei MTCC17 in solid cultures; Krishna PS et al.; Studies were performed on the production of rifamycin SV, an ansamycin compound, extensively used for curing tuberculosis, leprosy and several other mycobacterial infections, using a strain of Amycolatopsis mediterranei MTCC17 in solid cultures . Wheat bran was employed as a solid substrate . The culture produced 4 g of rifamycin SV/kg of substrate . Pre-treatment of the substrate with dilute HCl was found to increase the yield of rifamycin SV by 300% (from 4 to 12 g x kg of substrate(-1)) . Various process parameters were tested to establish the best conditions for the maximum production of the compound and a initial moisture level of 80%, inoculum size of 40%, initial substrate pH of 7.0, incubation temperature of 26 degrees C and a 7 day fermentation period were found to be optimal . Different solvents were used for the extraction of rifamycin SV from the fermented matter and methanol was found to be most suitable . Under optimized conditions, the yield of rifamycin SV further increased from 12 to 32 g x kg of substrate(-1), showing an 8-fold increase from the initial value.

Biochem Soc Trans, 2003 Feb, 31(Pt 1), 220 - 3
AMP-activated protein kinase and hepatic genes involved in glucose metabolism; Ferre P et al.; Mammalian AMP-activated protein kinase presents strong structural and functional similarities with the yeast sucrose non-fermenting 1 (Snf1) kinase involved in the derepression of glucose-repressed genes . It is now clearly established that AMP-activated protein kinase in the liver decreases glycolytic/lipogenic gene expression as well as genes involved in hepatic glucose production . This is achieved through a decreased transcriptional efficiency of transcription factors such as sterol-regulatory-element-binding protein-1c, carbohydrate-response-element-binding protein, hepatocyte nuclear factor 4 alpha or forkhead-related protein . Clearly, the long-term consequences of AMP-activated protein kinase activation have to be taken into account if activators of this enzyme are to be designed as anti-diabetic drugs.

Biochem Soc Trans, 2003 Feb, 31(Pt 1), 178 - 81
Snf1 protein kinase: a key player in the response to cellular stress in yeast; Sanz P; Sucrose non-fermenting 1 (Snf1) protein kinase, a yeast homologue of mammalian AMP-activated protein kinase, plays a main role in transcriptional activation and repression of gene expression . In addition, Snf1 kinase has a broad role in the cellular response to several forms of stress, such as nutrient limitation, salt stress and heat shock.

Biochem Soc Trans, 2003 Feb, 31(Pt 1), 175 - 7
Role of the yeast Snf1 protein kinase in invasive growth; Kuchin S et al.; The sucrose non-fermenting 1 (Snf1) protein kinase of Saccharomyces cerevisiae is important for transcriptional, metabolic and developmental responses to glucose limitation . Here we discuss the role of the Snf1 kinase in regulating filamentous invasive growth . Haploid invasive growth occurs in response to glucose limitation and requires FLO11, a gene encoding a cell-surface adhesin . Snf1 regulates transcription of FLO11 by antagonizing the function of two repressors, Nrg1 and Nrg2 . Snf1 and the Nrg repressors also affect diploid pseudohyphal differentiation, which is a response to nitrogen limitation, suggesting an unexpected signalling role for the Snf1 kinase.

ISA Trans, 2003 Jan, 42(1), 135 - 47
A method for detection and diagnosis on batch fermentations; Dondo RG; In this work we present some basic ideas about detection and diagnosis of faults and abrupt dynamic changes in batch fermentations . Our work focuses on the simultaneous use of two detection methods (residual based and balances based) within the estimation procedure . The idea behind the use of both methods is that the weakness of one of them can be compensated by the use of the other one . Thus the simultaneous use of both methods allows detecting and possibly isolating a wide range of faults . Observations such as the effect of nonlinearities on the detection tests and robustness to model uncertainty are discussed . Numerical results on a particular case, the xanthan gum batch fermentation, are presented . Simulated faults and abnormal behaviors were promptly detected but diagnostics showed mixed results.

J Antibiot (Tokyo), 2002 Nov, 55(11), 969 - 74
PF1163A, a novel antifungal agent, inhibit ergosterol biosynthesis at C-4 sterol methyl oxidase; Nose H et al.; PF1163A and B are a pair of antifungal agents isolated from a fermentation broth of Penicillium sp . PF1163A inhibited ergosterol synthesis in Saccharomyces cerevisiae, resulting in an accumulation of 4,4-dimethylzymosterol and a decrease of ergosterol . The ERG25 strain overexpressing the ERG25 gene was resistant to PF1163A . ERG25p is a C-4 sterol methyl oxidase known to be essential for the viability of yeast and fungi because of the known role of ERG25 gene disruption in S . cerevisiae-led lethality . ERG25p is the enzyme responsible for the first step in the removal of the two methyl groups at the C-4 position of sterol . From the results obtained here, we conclude that PF1163A is a novel natural antifungal that inhibits C-4 sterol methyl oxidase.

J Antibiot (Tokyo), 2002 Nov, 55(11), 941 - 51
Thielavins as glucose-6-phosphatase (G6Pase) inhibitors: producing strain, fermentation, isolation, structural elucidation and biological activities; Sakemi S et al.; High-throughput screening of microbial extracts using rat hepatic microsomal glucose-6-phosphatase (G6Pase) led us to find thielavin B as a G6Pase inhibitor with inhibition of glucose output from glucagon-stimulated hepatocytes . Further searching for more potent analogs identified 11 new thielavins F-P in addition to the known thielavins A and B from a fungus Chaetomium carinthiacum ATCC 46463 . Thielavin G showed the strongest activity as a G6Pase inhibitor (IC50=0.33 microM), while the IC50 of thielavin B was 5.5 microM . According to the structure-activity relationship, including authentic thielavins C, D and 3 partial hydrolysates from thielavins A and B, 3 benzoic acid-units and carboxylic acid functions are essential for G6Pase inhibition.

Isotopes Environ Health Stud, 2002 Sep, 38(3), 139 - 47
Production of 13C labelled pea flour for use in human digestion and fermentation studies; Edwards CA et al.; Stable isotope breath tests offer a new approach to the study of digestion and fermentation of carbohydrates in man . In this study, 13C labelled peas were grown by pulsing 250 ml 13CO2 into a sealed growth chamber . A second pulse was added to a portion of the peas to increase the 13C enrichment . This generated pea flour with an enrichment of 2.36 at.% excess (range 2.09-2.71 n = 3) and 8.64 atom % excess (range 7.37-9.78 n = 3) respectively . This represented incorporation of an absolute yield of 3.8% of the 13CO2 into peas in the 'once-labelled' treatment and 7.5% in the 'twice-labelled' treatment . Ingestion of a mixture of the labelled pea flour (300 mg) by two volunteers generated measurable 13CO2 excretion for breath test analysis . The profile of breath 13CO2 enrichment increased to a maximum within three hours after consuming the pea flour followed by a decrease almost back to baseline by 13 hours . Breath 13CO2 appeared to rise again after this apparent nadir at 13 hours until the end of the sampling period . Mathematical analysis of the data suggested that two peaks best described the profile of breath 13CO2 up to 13 hours . A third peak was necessary to describe the late rise in breath 13CO2 enrichment . This use of 13C enriched pea flour may provide a useful non invasive method for measurement of digestion and fermentation in vivo.

Se Pu, 2001 Nov, 19(6), 549 - 51
{Separation of xylose and xylitol in fermentation liquid by capillary zone electrophoresis}; Chen GH et al.; Xylitol may be produced with microbial fermentation technology when xylose is used as the raw material . It is important in scientific research and production fields concerned with xylose and xylitol to develop the method of separation and detection . The anionic complexes with strong UV absorption at 195 nm will form by dissolving xylose and xylitol in borax solution . They may be separated with borax buffer by use of capillary zone electrophoresis . It was shown that the resolution between xylose and xylitol gradually increased with the increase of borax concentration, but its maximum concentration was 130 mmol/L at room temperature . The resolution depended on the pH of running buffer with a maximum at pH 9.55 . The resolution was independent of cetyltrimethylammonium bromide when its concentration was between 4 x 10(-6) mmol/L-8 x 10(-4) mmol/L . So the optimum conditions were as follows: borax concentration of 130 mmol/L, pH 9.55 and cetyltrimethylammonium bromide concentration of 5 x 10(-5) mmol/L in running buffer; separating voltage of -12 kV; column temperature of 25 degrees C . Xylose and xylitol could be separated on base line in 6 min under these conditions . Samples from fermentation process and the recoveries of spiked samples were determined . The relative standard deviations of the results were between 1.42%-3.11% for xylose, and 0.62%-1.32% for xylitol . The recoveries were between 96.0%-108.0% for xylose and 94.0%-109.0% for xylitol.

J Ind Microbiol Biotechnol, 2003 Jan, 30(1), 41 - 6 Epub 2003 Jan 03.
Production of the secondary metabolites gamma-aminobutyric acid and monacolin K by Monascus; Su YC et al.; gamma-Aminobutyric acid (GABA), a hypotensive agent, and monacolin K, a cholesterol-lowering drug, can be produced by Monascus spp . Under optimal culture conditions, the products of fermentation using Monascus spp . may serve as a multi-functional dietary supplement and can prevent heart disease . In this study, Monascus purpureus CCRC 31615, the strain with the highest amount of monacolin K, was identified from 16 strains using solid fermentation . Its GABA productivity was particularly high . Addition of sodium nitrate during solid-state fermentation of M . purpureus CCRC 31615 improved the productivity of monacolin K and GABA to 378 mg/kg and 1,267.6 mg/kg, respectively . GABA productivity increased further to 1,493.6 mg/kg when dipotassium hydrophosphate was added to the medium.

J Ind Microbiol Biotechnol, 2003 Jan, 30(1), 22 - 7 Epub 2003 Jan 03.
Lactic acid production by Saccharomyces cerevisiae expressing a Rhizopus oryzae lactate dehydrogenase gene; Skory CD; This work demonstrates the first example of a fungal lactate dehydrogenase (LDH) expressed in yeast . A L(+)-LDH gene, ldhA, from the filamentous fungus Rhizopus oryzae was modified to be expressed under control of the Saccharomyces cerevisiae adh1 promoter and terminator and then placed in a 2 micro -containing yeast-replicating plasmid . The resulting construct, pLdhA68X, was transformed and tested by fermentation analyses in haploid and diploid yeast containing similar genetic backgrounds . Both recombinant strains utilized 92 g glucose/l in approximately 30 h . The diploid isolate accumulated approximately 40% more lactic acid with a final concentration of 38 g lactic acid/l and a yield of 0.44 g lactic acid/g glucose . The optimal pH for lactic acid production by the diploid strain was pH 5 . LDH activity in this strain remained relatively constant at 1.5 units/mg protein throughout the fermentation . The majority of carbon was still diverted to the ethanol fermentation pathway, as indicated by ethanol yields between 0.25-0.33 g/g glucose . S . cerevisiae mutants impaired in ethanol production were transformed with pLdhA68X in an attempt to increase the lactic acid yield by minimizing the conversion of pyruvate to ethanol . Mutants with diminished pyruvate decarboxylase activity and mutants with disrupted alcohol dehydrogenase activity did result in transformants with diminished ethanol production . However, the efficiency of lactic acid production also decreased.

J Nat Prod, 2003 Jan, 66(1), 136 - 9
New dimeric naphthopyrones from Aspergillus niger; Akiyama K et al.; Three new dimeric naphthopyrones, asperpyrones A (1), B (2), and C (3), together with two known compounds, fonsecinone A (4) and aurasperone A (5), have been isolated from okara that was fermented with Aspergillus niger JV-33-48 . Compounds 1, 4, and 5 showed inhibitory activity on Taq DNA polymerase.

J Nat Prod, 2003 Jan, 66(1), 121 - 4
Nodulisporic acids C, C1, and C2: a series of D-ring-opened nodulisporic acids from the fungus Nodulisporium sp; Ondeyka JG et al.; Nodulisporic acids C (4a), C1 (5a), and C2 (6a), a series of D-ring-opened nodulisporic acids, were isolated from fermentations of a mutant culture of Nodulisporium sp . MF5954 . Nodulisporic acid C, the most potent of the three, showed good activity against fleas with an LD90 of 10 micro g/mL . These compounds tend to be unstable in the free acid form and were isolated as stable sodium salts.

J Anim Sci, 2002 Dec, 80(12), 3336 - 46
Effects of oscillating dietary protein on ruminal fermentation and site and extent of nutrient digestion in sheep; Ludden PA et al.; Eight cannulated wethers (BW = 52.5 +/- 5.7 kg) were used in a replicated 4 x 4 Latin square designed experiment to evaluate the effects of oscillating dietary protein concentrations on ruminal fermentation, site and extent of digestion, and serum metabolite concentrations . Four treatments consisted of a 13, 15, or 17% CP diet fed daily or a regimen in which dietary CP was oscillated between 13 and 17% on a 48-h basis (ACP) . All diets consisted of 65% bromegrass hay (10.5% CP, 61.9% NDF, 37.2% ADF) plus 35% corn-based supplement and were formulated to contain the same amount of degradable intake protein (9.6% of DM) plus additional undegradable intake protein (SoyPLUS, West Central Cooperative, Ralston, IA) to accomplish CP levels above 13% . Each of four experimental periods were 16 d in duration with 12 d for diet adaptation followed by 4 d for sample collection . All wethers were fed at 3.0% of initial BW (DM basis) throughout the experiment, resulting in an average organic matter intake of 1.39 kg/d across treatments . When compared to the 15% CP daily treatment, feeding ACP had no effect (P > or = 0.10) on ruminal or lower tract N, NDF, ADF, or OM digestion . True ruminal OM digestion responded quadratically (P = 0.07) to increasing dietary CP, reaching a maximum of 52.0% of OM intake with the 15% CP treatment . Sheep fed ACP tended to have lower (P = 0.08) ruminal NH3 N concentrations and an overall higher (P = 0.0001) molar proportion of acetate compared to those fed 15% CP daily . Total VFA concentrations were not affected (P > or = 0.45) by increasing dietary CP . Microbial efficiency did not differ (P > or = 0.55); thus, bacterial N flow at the duodenum responded quadratically (P = 0.04) to increasing dietary CP . Nonbacterial N (P = 0.001) and total N (P = 0.01) flows at the duodenum and total tract N digestibility (P < or = 0.04) increased linearly as dietary CP increased . Wethers fed ACP maintained a lower (P = 0.002) serum glucose and lower (P = 0.0006) serum urea N compared to those fed 15% CP daily . Because the CP content of the diet was increased at the expense of corn, the response to increased CP observed in this experiment is most likely due to negative associative effects of supplemental starch on ruminal fermentation and microbial growth . Oscillating the CP content of the diet on a 48-h basis has little effect on digestion or N utilization in sheep compared with feeding the same quantity of protein on a daily basis.

Se Pu, 2001 Jan, 19(1), 16 - 20
{Study of the adsorption properties in chromatographic separation of the citric acid}; Peng QJ et al.; A new method studying the change of thermodynamic properties at different temperatures to screen suitable stationary phases for chromatographic separation of the citric acid (CA) on an analytical column has been developed . The applied method is mainly based on using CA with a series of solute concentrations (0 g/L-250 g/L) as mobile phase to seriously overpass the unloaded stationary phase to find the breakthrough curves of CA . The adsorption isotherms were calculated from the retention times of breakthrough curve . The bed porosity of column was calculated from the elution time of a peak of the blue dextran as non-adsorbed tracer . The stationary phase, which has been selected through this method, for the separation of CA from fermentation broth, showed a very high adsorption capacity for CA, even under very low concentrations, so it can even be used in the separation and purification process of minute amounts of CA . The separation effect of this stationary phase for citric acid can be greatly enhanced through the technique of change of temperature . This new separation process can shorten the old process path, improve the yield of finished products more than 10%, and reduce the cost more than 10% . At the same time, the pollution of the production of citric acid can be reduced . It turned out that this method will play an important role in the synthesis of stationary phase for the separation of citric acid and the study of its structure.

Se Pu, 2001 Jan, 19(1), 13 - 5
{Determination of enantiomeric purity for lactic acid in fermentation broth by Rhizopus oryzae with high performance liquid chromatography}; Bai DM et al.; A procedure for the resolution of DL-lactic acid and the determination of D-isomer ratio in L-lactic acid fermentation broth by Rhizopus oryzae is described . The effects of pH of mobile phase and concentration of chiral mobile phase additives on resolution of DL-lactic acid were investigated . The optical isomers of lactic acid were resolved by RP-HPLC with 2,3,6-tri-O-beta-cyclodextrin(TM-beta-CD) as a chiral mobile phase additive, and C18 column as stationary phase, and detected at wavelength 210 nm . The results showed that a correction factor should be introduced into the equation for calculation of the percentage of D-lactic acid, because the UV absorption of D-lactic acid and L-lactic acid might not be the same when TM-beta-CD was present . Quantitation was achieved with external standard method, the average recovery was 100.4%, and the relative standard deviation was 0.82% . This method can be used for the determination of the percentage of D-isomer in L-lactic acid fermentation broth by Rhizopus oryzae, and it is simple, rapid and accurate . The results showed that the mass fraction of D-isomer in the fermented broth increased during the period of storage.

Se Pu, 2000 Nov, 18(6), 527 - 8
{Determination of lactic acid in fermentation broth of Rhizopus oryzae by reversed-phase high performance liquid chromatography (RP-HPLC)}; Bai DM et al.; A method for determining lactic acid in fermentation broth of Rhizopus oryzae by RP-HPLC is described . The operating conditions were Wakosil-II 5 C18 RS column(4.6 mm i.d . x 150 mm, 5 microns) at room temperature, 0.01 mol/L phosphoric acid solution (pH 2.5) as mobile phase with a flow rate of 1.0 mL/min and UV detection at 210 nm . The retention time of lactic acid was 3.820 min . This method is simple, rapid and accurate . The results will not be affected by other components in the broth . The relative standard deviation was 0.22% (n = 5), and the recovery was over 99%.

Se Pu, 2002 Jan, 20(1), 46 - 8
{Determination of organic acids in fermentation broth of spiramycin by high performance liquid chromatography}; Li YY et al.; A method for determining organic acids in spiramycin fermentation broth by high performance liquid chromatography is described . The operating conditions were Zorbax 300-SB C18 column (5 microns, 4.6 mm i.d . x 15 cm) at 35 degrees C, 0.01 mol/L phosphoric acid buffer solution (pH 2.32) and methanol as mobile phase at a flow rate of 0.6 mL/min and UV detection at 210 nm . The relative standard deviations were 0.33%-0.10% and the recoveries were 99.95%-100.08% . It's a simple, rapid and accurate method.

Se Pu, 2000 Mar, 18(2), 170 - 2
{Simultaneous analysis of low concentrations of glucose, ethanol and glycerol by high performance liquid chromatographic method}; Li WJ et al.; A method based on high performance liquid chromatography (HPLC) for the analysis of glucose, ethanol and glycerol simultaneously is presented . The baseline separation was achieved at room temperature (25-30 degrees C) on a Beckman mu-Spherogel carbohydrate column (6.5 mm i.d . x 30 cm) with H2SO4-H2O(0.5:1,000, V/V) as mobile phase at a flow rate of 1.000 mL/min . All the substances were detected with the Beckman 186 Refractive Index Detector . Under these conditions, distinct peaks of glucose, ethanol and glycerol were resolved within 15 min . The retention time in the particular condition was (6.04 +/- 0.04) min for glucose, (13.36 +/- 0.08) min for ethanol and (8.72 +/- 0.05) min for glycerol . The detection limit for glucose, ethanol and glycerol were found to be 10(-5) g, 10(-4) g and 10(-5) g respectively . A series of experiments have been performed to investigate the glucose consumption and the ethanol, glycerol production in the random growth of Saccharomyces cerevisiae with the method above . Only 0.5 mL sample was needed each time . In 30 hours after inoculation, the cell density increased from 10(6) mL-1 to 10(7) mL-1 . It was found that the glucose concentration decreased approximately linearly after 8 hours, and ethanol and glycerol was detected after the delay period as expected . The results show that the method proposed is rapid and convenient with enough accuracy . The method may be used to investigate the metabolic behavior of yeast in the fermentation process at the early stage or detect the slight changes of concentrations of some objective substances in particular culture experiments.

Se Pu, 2000 Jan, 18(1), 88 - 9
{Derivative gas chromatographic analysis of fructooligosaccharide in fermented sucrose}; Cai WM et al.; As a new sweetener, fructooligosaccharide is paid more and more attention for its health improvement property . It includes trisaccharide, tetrasaccharide and pentasaccharide, and can be produced from sucrose by the fermentation of microorganism . In order to analyze the content of fructooligosaccharide in fermented sugar by gas chromatography, fructooligosaccharide was transformed into trimethylsilyl derivatives . Based on the modified gas chromatograph SP2308, and under the chosen chromatographic conditions with 0.53 mm capillary column of OV-101, the contents of fructose, glucose, sucrose and fructooligosaccharide were determined by programmed temperature chromatography . The recovery of fructooligosaccharide was satisfactory.

Am J Physiol Gastrointest Liver Physiol, 2003 Jun, 284(6), G989 - 95 Epub 2003 Jan 22.
Upregulation of activin A gene by butyrate in human colon cancer cell lines; Sonoyama K et al.; Activin A has been reported to play a role in the progression of colorectal cancer . Because dietary fiber protects against colorectal cancer, we hypothesized that butyrate, a fermentation product of dietary fiber, may affect the expression of activin A in colon cancer cells . Semiquantitative RT-PCR demonstrated that the activin A gene was upregulated by sodium butyrate in the human colon cancer cell lines HT-29 and Caco-2 in a concentration- and time-dependent manner . However, the activin A gene did not respond to sodium butyrate in the human normal colonic cell line FHC, rat normal intestinal epithelial cell (IEC) line IEC-6, and the explant of rat colon . Flow cytometry and agarose gel electrophoresis of genomic DNA revealed that cell cycle arrest and apoptosis were induced by sodium butyrate but not exogenous activin A in HT-29 cells, indicating that activin A could not act as an autocrine factor in colon cancer cells . By assuming that activin A promotes colorectal cancer spread as a paracrine factor, our findings suggest that butyrate could act as a tumor promoter in some circumstances.

J Agric Food Chem, 2003 Jan 29, 51(3), 784 - 92
Characteristics of salt-fermented sauces from shrimp processing byproducts; Kim JS et al.; A salt-fermented sauce from shrimp processing byproducts (heads, shells, and tails) was prepared and characterized . Three types of sauces were prepared; sauce C, with 30 g of salt/100 g of byproduct (high salt); sauce E, with 30 g of salt and 0.2 g of sodium erythorbate (high salt); and sauce L, with 20 g of salt, 0.2 g of sodium erythorbate, 6 g of sorbitol, 0.5 mL of lactic acid, and 5 mL of ethanol (low salt) . Sauces C and E showed higher exopeptidase activities than sauce L, whereas sauce L showed the highest endopeptidase activity . After 3 months of fermentation, the amino N content of sauce increased from 150-200 to 500-600 mg/100 g and the nonprotein nitrogen content increased from 300 to 950-1050 mg/100 g . Volatile basic nitrogen content increased significantly from 18 to 60 mg/100 g . The total carotenoids retained in sauces C, E, and L were 26.3, 76.2, and 73%, respectively, thus indicating that the addition of sodium erythorbate to sauces E and L retarded oxidation . Water activities of sauces C, E, and L were 0.753, 0.751, and 0.773, respectively . According to the omission test, the taste of sauces was influenced by the content of free amino acids, mainly glutamic acid and aspartic acid . All three sauces examined showed a 35% higher total amino acid content than commercial salt-fermented shrimp sauces . Therefore, shrimp processing byproducts may lend themselves to the preparation of high-quality salt-fermented sauces.

J Agric Food Chem, 2003 Jan 29, 51(3), 654 - 8
Storage of immobilized yeast cells for use in wine-making at ambient temperature; Kourkoutas Y et al.; A comparative study of the storage and reuse of immobilized yeast cells on apple pieces, kissiris, and gamma-alumina was carried out . The immobilized biocatalysts were allowed to remain in the fermented alcoholic liquid after the end of each fermentation batch for extended periods at 30 degrees C before reactivation in batch fermentation for wine-making . The results showed that the biocatalysts were able to reactivate and ferment after successively increased periods of storage compared to free cell systems both on glucose medium and on grape must . In glucose medium, apple-, kissiris-, and gamma-alumina-supported biocatalysts reactivated after 120, 80, and 83 days, respectively . Possible storage periods for grape must were lower but remained high . Immobilized yeast biocatalyst on apple pieces produced wines with an improved volatiles composition compared to kissiris- and gamma-alumina-supported biocatalysts . There were no significant negative effects on the fermentation activity and volatile byproduct composition.

Appl Microbiol Biotechnol, 2003 Jan, 60(5), 560 - 3 Epub 2002 Nov 20.
Increasing ethanol productivity during xylose fermentation by cell recycling of recombinant Saccharomyces cerevisiae; Roca C et al.; The influence of cell recycling of xylose-fermenting Saccharomyces cerevisiae TMB3001 was investigated during continuous cultivation on a xylose-glucose mixture . By using cell recycling at the dilution rate ( D) of 0.05 h(-1), the cell-mass concentration could be increased from 2.2 g l(-1) to 22 g l(-1) . Consequently, the volumetric ethanol productivity increased ten-fold, from 0.5 g l(-1) h(-1) to 5.35 g l(-1) h(-1) . By increasing the biomass concentration, the xylose consumption rate increased from 0.75 g xylose l(-1) h(-1) without recycling to 1.9 g l(-1) h(-1) with recycling . The specific ethanol productivity was in the range of 0.23-0.26 g g(-1) h(-1) with or without cell recycling, showing that an increased cell-mass concentration did not influence the efficiency of the yeast.

Nat Biotechnol, 2003 Feb, 21(2), 150 - 6 Epub 2003 Jan 21.
Integrating transcriptional and metabolite profiles to direct the engineering of lovastatin-producing fungal strains; Askenazi M et al.; We describe a method to decipher the complex inter-relationships between metabolite production trends and gene expression events, and show how information gleaned from such studies can be applied to yield improved production strains . Genomic fragment microarrays were constructed for the Aspergillus terreus genome, and transcriptional profiles were generated from strains engineered to produce varying amounts of the medically significant natural product lovastatin . Metabolite detection methods were employed to quantify the polyketide-derived secondary metabolites lovastatin and (+)-geodin in broths from fermentations of the same strains . Association analysis of the resulting transcriptional and metabolic data sets provides mechanistic insight into the genetic and physiological control of lovastatin and (+)-geodin biosynthesis, and identifies novel components involved in the production of (+)-geodin, as well as other secondary metabolites . Furthermore, this analysis identifies specific tools, including promoters for reporter-based selection systems, that we employed to improve lovastatin production by A . terreus.

J Appl Microbiol, 2003, 94(2), 312 - 20
Raw potato starch and short-chain fructo-oligosaccharides affect the composition and metabolic activity of rat intestinal microbiota differently depending on the caecocolonic segment involved; Le Blay GM et al.; AIMS: In vitro studies have suggested that fructo-oligosaccharides (FOS) and resistant starch (two fermentable non-digestible carbohydrates) display different fermentation kinetics . This study investigated whether these substrates affect the metabolic activity and bacterial composition of the intestinal microflora differently depending on the caecocolonic segment involved . METHODS AND RESULTS: Eighteen rats were fed a low-fibre diet (Basal) or the same diet containing raw potato starch (RPS) (9%) or short-chain FOS (9%) for 14 days . Changes in wet-content weights, bacterial populations and metabolites were investigated in the caecum, proximal and distal colon and faeces . Both substrates exerted a prebiotic effect compared with the Basal diet . However, FOS increased lactic acid-producing bacteria (LAPB) throughout the caecocolon and in faeces, whereas the effect of RPS was limited to the caecum and proximal colon . As compared with RPS, FOS doubled the pool of caecal fermentation products, while the situation was just the opposite distally . This difference was mainly because of the anatomical distribution of lactate, which accumulated in the caecum with FOS and in the distal colon with RPS . Faeces reflected these impacts only partly, showing the prebiotic effect of FOS and the metabolite increase induced by RPS . CONCLUSIONS: This study demonstrates that FOS and RPS exert complementary caecocolonic effects . SIGNIFICANCE AND IMPACT OF THE STUDY: The RPS and FOS combined ingestion could be beneficial by providing health-promoting effects throughout the caecocolon.

Radiats Biol Radioecol, 2002 Nov-Dec, 42(6), 628 - 31
{Coding in molecular biology and radiation}; Eingorin MIa; In work, on the basis of known base biological laws, the new natural three-dimensional designs are under construction: codonogram and aminogram . Codonogram and aminogram have revealed the new laws of coding and constructions in molecular biology . The secondary coding codonogram, at the expense of redundancy, has enabled for DNA, RNA, and mRNA to construct the hidden coding layers (HLC) . HLC are constructed on a base of pirimidine and purine . In interaction with ferments HLC, should guarantee: addressation, synchronization, correction and other functional transformations of a gene . Radiation can damage a layer mRNA of synthesis protein, hidden layers of coding and structure of the ferments of the cell . In work are given: kodonogram, aminogram UGC, basis of construction HLC, the references to the literature with an example of decoding.

Biotechnol Bioeng, 2003 Mar 20, 81(6), 738 - 47
Potential inhibitors from wet oxidation of wheat straw and their effect on ethanol production of Saccharomyces cerevisiae: wet oxidation and fermentation by yeast; Klinke HB et al.; Alkaline wet oxidation (WO) (using water, 6.5 g/L sodium carbonate and 12 bar oxygen at 195 degrees C) was used as pretreatment method for wheat straw (60 g/L), resulting in a hydrolysate and a cellulosic solid fraction . The hydrolysate consisted of soluble hemicellulose (8 g/L), low-molecular-weight carboxylic acids (3.9 g/L), phenols (0.27 g/L = 1.7 mM) and 2-furoic acid (0.007 g/L) . The wet oxidized wheat straw hydrolysate caused no inhibition of ethanol production by Saccharomyces cerevisiae ATCC 96581 . Nine phenols and 2-furoic acid, identified to be present in the hydrolysate, were each tested in concentrations of 50-100 times the concentration found in the hydrolysate for their effect on fermentation by yeast . At these high concentrations (10 mM), 4-hydroxybenzaldehyde, vanillin, 4-hydroxyacetophenone and acetovanillone caused a 53-67% decrease in the volumetric ethanol productivity in S . cerevisiae compared to controls with an ethanol productivity of 3.8 g/L . The phenol acids (4-hydroxy, vanillic and syringic acid), 2-furoic acid, syringaldehyde and acetosyringone were less inhibitory, causing a 5-16% decrease in ethanol productivity . By adding the same aromatic compounds to hydrolysate (10 mM), it was shown that syringaldehyde and acetovanillone interacted negatively with hydrolysate components on the ethanol productivity . Fermentation in WO hydrolysate, that had been concentrated 6 times by freeze-drying, lasted 4 hours longer than in regular hydrolysate; however, the ethanol yield was the same . The longer fermentation time could not be explained by an inhibitory action of phenols alone, but was more likely caused by inhibitory interactions of phenols with carboxylic acids, such as acetic and formic acid .

Biotechnol Bioeng, 2003 Mar 20, 81(6), 726 - 31
Enzymatic hydrolysis of water-soluble wheat arabinoxylan . 1 . Synergy between alpha-L-arabinofuranosidases, endo-1,4-beta-xylanases, and beta-xylosidase activities; Sorensen HR et al.; Hydrolysis of arabinoxylan is an important prerequisite for improved utilization of wheat hemicellulose in the ethanol fermentation industry . This study investigates the individual and combined efficiencies of three commercial, cellulytic and hemicellulytic enzyme preparations, Celluclast 1.5 L, Ultraflo L, and Viscozyme L, in catalyzing the liberation of arabinose and xylose from water-soluble wheat arabinoxylan . Ultraflo L was the best enzyme preparation for releasing arabinose, liberating 53 wt% of the theoretical maximum after 48 h of reaction (10 wt% enzyme/substrate ratio, 40 degrees C, pH 6) . Celluclast 1.5 L was superior to the other enzyme preparations in releasing xylose, liberating 26 wt% of the theoretical maximum after 48 h of reaction (10 wt% enzyme/substrate ratio, 50 degrees C, pH 5) . The 50:50 mixtures of the enzyme preparations showed no synergistic cooperation in arabinose release, but a synergistic interaction in xylose release was found between Ultraflo L and Celluclast 1.5 L . On the basis of high-performance anion exchange chromatography (HPAEC) analysis of the hydrolysates after enzymatic reaction, we propose that the observed synergism between Celluclast 1.5 L and Ultraflo L is the result of positive interaction between alpha-L-arabinofuranosidase and endo-1,4-beta-xylanase activities present in Ultraflo L that released arabinose, xylobiose and xylotriose, and beta-xylosidase activities in Celluclast 1.5 L, capable of catalyzing the hydrolysis of xylobiose and xylotriose to xylose .

Biotechnol Bioeng, 2003 Mar 20, 81(6), 666 - 72
High-cell-density fed-batch cultivation of the docosahexaenoic acid producing marine alga Crypthecodinium cohnii; De Swaaf ME et al.; The heterotrophic marine alga Crypthecodinium cohnii is known to produce docosahexaenoic acid (DHA), a polyunsaturated fatty acid with food and pharmaceutical applications, during batch cultivation on complex media containing sea salt, yeast extract, and glucose . In the present study, fed-batch cultivation was studied as an alternative fermentation strategy for DHA production . Glucose and acetic acid were compared as carbon sources . For both substrates, the feed rate was adapted to the maximum specific consumption rate of C . cohnii . In glucose-grown cultures, this was done by maintaining a significant glucose concentration (between 5 and 20 g/L) throughout fermentation . In acetic acid-grown cultures, the medium feed was automatically controlled via the culture pH . A feed consisting of acetic acid (50% w/w) resulted in a higher overall volumetric productivity of DHA (r(DHA)) than a feed consisting of 50% (w/v) glucose (38 and 14 mg/L/h, respectively) . The r(DHA) was further increased to 48 mg/L/h using a feed consisting of pure acetic acid . The latter fermentation strategy resulted in final concentrations of 109 g/L dry biomass, 61 g/L lipid, and 19 g/L DHA . These are the highest biomass, lipid, and DHA concentrations reported to date for a heterotrophic alga . Vigorous mixing was required to sustain aerobic conditions during high-cell-density cultivation . This was complicated by culture viscosity, which resulted from the production of viscous extracellular polysaccharides . These may present a problem for large-scale industrial production of DHA . Addition of a commercial polysaccharide-hydrolase preparation could decrease the viscosity of the culture and the required stirring .

Zhongguo Zhong Yao Za Zhi, 2001 Jun, 26(6), 381 - 4
{Studies on polysaccharides in different development stages of Armillaria mellea (Vahl.: Fr.) Quel.}; Chen XM et al.; OBJECTIVE: To study the polysaccharides in different development stages of Armillaria mellea . METHOD: Polysaccharides in rhizomorph, fruit-body, mycelia and its fermenting liquor of A . mellea were extracted, isolated and purified . The properties contents, molar ratio and molecular weight of the polysaccharides were determined by IR spectra, HPLC, GPC and gel chromatography . RESULT: The polysaccharides in mycelia and its fermenting liquor contained only glucose, the sugar contents were 9.00% in mycelia and 0.87 g.(100 ml)-1 in fermenting liquor respectively . The polysaccharides in both rhizomorph and fruit-body consisted of glucose and xylose, and the molar ratio was 1:14 in rhizomorph and 1:10 in fruit-body respectively; the polysaccharide contents were 1.12% in rhizomorph and 2.27% in fruit-body . The molecular weight of these polysaccharides was about 10,000-70,000 . CONCLUSION: The experimental results supply important scientific data for developing A . mellea as a medicine.

FEBS Lett, 2003 Jan 16, 534(1-3), 87 - 92
Identification of an Arabidopsis mitochondrial succinate-fumarate translocator; Catoni E et al.; Complementation of a yeast acr1 mutant carrying a deletion of the succinate/fumarate carrier gene enabled functional identification of a mitochondrial succinate translocator from Arabidopsis thaliana (AtmSFC1) . Thus complementation of yeast mutants is applicable also for identification and characterization of organellar transporters . Reverse transcription polymerase chain reaction and promoter-GUS fusion showed expression of AtmSFC1 in 2 day old dark grown seedlings, which declined in cotyledons during further development, consistent with a role in export of fumarate for gluconeogenesis during lipid mobilization at early germination of Arabidopsis seeds . In mature plants, expression was found in developing and germinating pollen, suggesting a role in ethanolic fermentation.

Pharmacol Res, 2003 Jan, 47(1), 49 - 52
Study of anti-inflammatory activity of Tibetan mushroom, a symbiotic culture of bacteria and fungi encapsulated into a polysaccharide matrix; Diniz RO et al.; Tibetan mushroom (TM) is a fermented beverage composed by a dozen of bacteria and yeasts living together into polysaccharide grains secreted by them . TM is similar to kefir, a probiotic beverage originated in the Caucasian mountains exhibiting some anti-bacterial, anti-mycotic, anti-neoplastic and immunomodulatory effects . Aiming to evaluate a plausible anti-inflammatory property of TM we conducted cotton-induced granuloma and paw edema assays in rats, the latter using carrageenin, dextran and histamine as stimuli . TM samples were thawed and continuously cultured during 15 days into molasses solutions (50 g/l) . The experiments used TM suspensions after 24h fermentation and TM grains mechanically disintegrated . The results showed a significant inhibition on the formation of granuloma tissue for the test group as compared to the negative control group . TM suspensions presented an inhibition of 43% for the inflammatory process . Rat paw edema also showed significant decreases with the mediators . The edema induced by carrageenin was inhibited 62% at the 3rd hour . The edema dextran-induced was completely inhibited at 1h and antagonized the histamine edema 52% at 1h.

Wei Sheng Yan Jiu, 2001 May, 30(3), 190 - 2
{Health effects of indigestible carbohydrates on colon}; Hong J et al.; Indigestible carbohydrates are closely related to the growth of microbes in colon and the health status of colon . People are now becoming more and more interested in indigestible carbohydrates and many studies have been carried out . In this review, the classification and the digestion of indigestible carbohydrates, and the health effects of their fermented products on colon were introduced.

J Antibiot (Tokyo), 2002 Oct, 55(10), 899 - 903
Strain diversity within a microbial collection; DiTullio D et al.; Random subsets of a modest-sized microbial collection have been examined for culture redundancy, initially by morphology, both to the naked eye and microscopically, of cultures grown on a variety of agar-based solid media . Subsequent analysis, by simple TLC, of the extractable metabolites produced by morphologically similar cultures grown in submerged shaken fermentation was carried out . Apparent duplicate cultures were further examined on Biolog SF-P MicroPlates for differentiation . The results were subjected to a statistical analysis and the contribution of each stage in the process to resolving culture uniqueness was noted . A statistical extrapolation of the results, from the subsets of each culture type, to the total for that type within the entire collection, with 95% confidence limits, is presented . Morphological comparison, on four different agar media, gives a significant underestimation of the metabolic diversity of the collection . The weighted mean from the three types of cultures indicate that the expected content of the collection is approximately 93% unique strains.

J Antibiot (Tokyo), 2002 Oct, 55(10), 847 - 54
CJ-15,208, a novel kappa opioid receptor antagonist from a fungus, Ctenomyces serratus ATCC15502; Saito T et al.; A novel kappa opioid receptor binding inhibitor CJ-15,208 (I) was isolated from the fermentation broth of a fungus, Ctenomyces serratus ATCC15502 . The structure of I was determined to be a cyclic tetrapeptide consisting of one tryptophan, one D-proline, and two L-phenylalanine . Compound I was a selective binding inhibitor for the kappa opioid receptor: 47 nM (IC50) for kappa, 260 nM for mu, and 2,600 nM for delta . In the electrically-stimulated twitch response assay of rabbit vas deferens I recovered the suppression by a kappa agonist asimadoline with an ED50 of 1.3 microM, indicating that it is a kappa antagonist.

Water Sci Technol, 2002, 46(11-12), 153 - 7
Anaerobic acidification of a synthetic wastewater in batch reactors at 55 degrees C; Yu HQ et al.; Experiments were conducted to study the acidogenesis of a dairy wastewater in batch reactors at pH 5.5 and 55 degrees C . There was a biased fermentation sequence for carbohydrate and protein, and the protein fermentation was delayed by carbohydrate . The production of hydrogen was exclusively from the fermentation of carbohydrate . Acetate and butyrate concentrations both increased rapidly at the beginning and peaked at some points, then declined in the reactors fed with 8 g-COD (chemical oxygen demand)/l, or higher concentrations . Butanol and propanol fractions increased with the substrate concentration . The metabolism shifted from the volatile fatty acid-producing pathways to the alcohol-producing pathways when the substrate concentration increased beyond 8 g-COD/l . The acidogenic biomass yield was in the range 0.19-0.25 mg-VSS/mg-COD.

Adv Biochem Eng Biotechnol, 2003, 79, 1 - 35
Amino acid production processes; Ikeda M; With the exploitation of new uses and the growing markets of amino acids, amino acid production technology has made large progress during the latter half of the 20th century . Fermentation technology has played crucial roles in this progress, and currently the fermented amino acids represent chief products of biotechnology in both volume and value . This area is highly competitive in the world market and process economics are of primary importance . For cost-effective production, many technologies have been developed to establish high-productive fermentation and recovery processes . The producer organisms used in large-scale, well-established processes have been developed to a high level of production efficiency . The tools of genetic engineering of amino acid-producing organisms have been well developed and are now being applied for enlargement of biosynthetic and transport capacity, which is beginning to have a great impact on the amino acid industry . Furthermore, the rapid strides in genome analysis are bound to revolutionize the strain improvement methodology.

Environ Sci Technol, 2002 Dec 15, 36(24), 5511 - 6
Green technology for conversion of food scraps to biodegradable thermoplastic polyhydroxyalkanoates; Du G et al.; A new technology is developed and demonstrated that couples anaerobic digestion of food scraps with production of biodegradable thermoplastics, polyhydroxyalkanoates (PHAs) . The food wastes were digested in an anaerobic reactor producing four major organic acids . The concentrations of acetic, propionic, butyric, and lactic acids reached 5.5, 1.8, 27.4, and 32.7 g/L, respectively . The fermentative acids were transferred through membranes via molecule diffusion into an air-bubbling reactor where the acids were utilized to produce PHAs in an enriched culture of Ralstonia eutropha . With a silicone rubber membrane, butyric acid and small amounts of acetic and propionic acids were transferred and used, producing a homopolymer PHA, poly(3-hydroxybutyrate) . The dry cell weight and PHA content reached 11.3 g/L and 60.2% (w/w), respectively . With a dialysis membrane, the mass transfer rates of fermentative acids were enhanced, and the PHA production was significantly improved . The dry cell weight and its PHA content reached 22.7 g/L and 72.6% (w/w), respectively . The formed PHA was a copolymer of 3-hydroxybutyrate and 3-hydroxyvalerate (HV) with 2.8 mol % HV monomer unit . The polymer content (72.6% of dry cell mass) reported in this study is the highest one obtained from organic wastes and is comparable with the PHA content from pure glucose fermentation.

Biol Pharm Bull, 2003 Jan, 26(1), 84 - 7
Hypocholesterolemic effect of hot-water extract from mycelia of Cordyceps sinensis; Koh JH et al.; This study was conducted to investigate the hypocholesterolemic effect of the hot-water fraction (HW) from cultured mycelia of Cordyceps sinensis in a 5 l fermenter . The composition of HW was mainly carbohydrate (83.9%) and protein (11.8%) on a dry basis, and the carbohydrate of HW consisted of glucose, mannose, galactose, and arabinose in the molecular ratio of 1.0 : 0.8 : 0.5 : 0.1, respectively . In mice fed a cholesterol-free diet and those fed a cholesterol-enriched diet, body and liver weights were not significantly different from those of the controls . The serum total cholesterol (TC) of all mice groups administered HW (150 and 300 mg/kg/d, respectively) with the cholesterol-enriched diet decreased more than in the control group . Among the mice fed the cholesterol-enriched diet, HW also increased the high-density lipoprotein (HDL) cholesterol level, but decreased the very low-density lipoprotein plus low-density lipoprotein (VLDL+LDL) cholesterol level . The changes in HDL- and VLDL+LDL-cholesterol levels consequently decreased the atherogenic value . The results indicate that HW in rats administered a cholesterol-enriched diet decreased the plasma cholesterol level . The 300 mg/kg dose had a significant effect on the serum TC level.

Mol Microbiol, 2003 Jan, 47(2), 495 - 506
The product of the UTH1 gene, required for Bax-induced cell death in yeast, is involved in the response to rapamycin; Camougrand N et al.; A yeast mutant was isolated that was resistant to Bax-induced cell death . It supports a mutation leading to decreased amounts of the protein Uth1p . A strain in which the UTH1 gene is disrupted also exhibits resistance to Bax expression . The absence of Uth1p does not change the mitochondrial localization of Bax, its insertion in the mitochondrial outer membrane or its cytochrome c-releasing activity . On the other hand, the absence of Uth1p does prevent the appearance of other hallmarks related to Bax expression in yeast, such as oxidation of mitochondrial lipid, production of reactive oxygen species and maintenance of plasma membrane properties after ethanol stress . The absence of Uth1p was also found to induce resistance to rapamycin, a specific inducer of autophagy . This resistance only appears when cells are grown under respiratory conditions, but not under fermentative conditions, suggesting that Uth1p acts in an autophagic pathway involving mitochondria, in accordance with its main localization in the outer mitochondrial membrane . Taken together, these data show that Bax is able to activate a death pathway related to autophagy in yeast, which also exhibits typical hallmarks of apoptosis, revealing a possible dual function of Bax in both types of death . This hypothesis is discussed in the light of observations suggesting a co-regulation of apoptosis and autophagy in mammalian cells.

Yeast, 2003 Jan 30, 20(2), 157 - 69
Identification of multicopy suppressors of cell cycle arrest at the G1-S transition in Saccharomyces cerevisiae; Munoz I et al.; Inactivation of HAL3 in the absence of SIT4 function leads to cell cycle arrest at the G(1)-S transition . To identify genes potentially involved in the control of this phase of the cell cycle, a screening for multicopy suppressors of a conditional sit4 hal3 mutant (strain JC002) has been developed . The screening yielded several genes known to perform key roles in cell cycle events, such as CLN3, BCK2 or SWI4, thus proving its usefulness as a tool for this type of studies . In addition, this approach allowed the identification of additional genes, most of them not previously related to the regulation of G(1)-S transition or even without known function (named here as VHS1-3, for viable in a hal3 sit4 background) . Several of these gene products are involved in phospho-dephosphorylation processes, including members of the protein phosphatase 2A and protein phosphatases 2C families, as well as components of the Hal5 protein kinase family . The ability of different genes to suppress sit4 phenotypes (such as temperature sensitivity and growth on non-fermentable carbon sources) or to mimic the functions of Hal3 was evaluated . The possible relationship between the known functions of these suppressor genes and the progress through the G(1)-S transition is discussed .

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