Fermentations

Am J Vet Res, 2000 Aug, 61(8), 858 - 61
In vitro evaluation of intraluminal factors that may alter intestinal permeability in ponies with carbohydrate-induced laminitis; Weiss DJ et al.; OBJECTIVES: To study the in vitro effects of cecal contents incubated with corn starch on colonic permeability in horses . ANIMALS: 4 healthy adult ponies . PROCEDURE: Mucosal specimens were obtained from the right ventral colon and mounted in Ussing chambers . Changes in short circuit current, conductance, and large-molecule permeability in response to addition of cecal contents and cecal contents incubated with corn starch were evaluated for 120 minutes . RESULTS: Incubation of cecal contents with corn starch for 8 hours resulted in a decrease in cecal content pH and an increase in lactic acid concentration . These changes were similar to those reported in vivo for ponies given corn starch . Exposure of colonic mucosa to cecal contents incubated with corn starch resulted in an increase in tissue conductance and permeability of technetium Tc 99m pentetate, compared with mucosa exposed to cecal contents alone . CONCLUSIONS AND CLINICAL RELEVANCE: In vitro exposure of colonic mucosa to cecal contents incubated with starch resulted in increased paracellular permeability . Fermentation of excessive amounts of carbohydrate in the intestinal lumen of horses may directly induce increased intestinal permeability associated with carbohydrate-induced laminitis.

J Biotechnol, 2000 Jul 14, 80(3), 271 - 6
Repeated-batch production of pigments by immobilised Monascus purpureus; Fenice M et al.; Extracellular pigment production by immobilised Monascus purpureus C322 has been studied in repeated-batch processes using different immobilising carriers such as Ca-alginate, polyurethane sponge, active carbon and pearlite . With Ca-alginate, pigment production was maximum (30.5 UA470 as process mean production, three batches) while the cell leakage was negligible (0.4 g 1(-1) free biomass) and the bead mechanical stability good; with this carrier, an extended repeated-batch fermentation (nine batches, 55 days) was carried out: the process pigment productivity was 3.87 UA470 day(-1).

Am J Bot, 2000 Aug, 87(8), 1081 - 1090
Fate of oxygen losses from Typha domingensis (Typhaceae) and Cladium jamaicense (Cyperaceae) and consequences for root metabolism; Chabbi A et al.; The objective of this work was to determine whether radial oxygen loss (ROL) from roots of TYPHA: domingensis and CLADIUM: jamaicense creates an internal oxygen deficiency or, conversely, indicates adequate internal aeration and leakage of excess oxygen to the rhizosphere . Methylene blue in agar was used to visualize the pattern of ROL from roots, and oxidation of a titanium-citrate solution was used to quantify rates of oxygen leakage . TYPHA:'s roots had a higher porosity than CLADIUM:'s and responded to flooding treatment by increasing cortical air space, particularly near the root tips . A greater oxygen release, which occurred along the subapical root axis, and an increase in rhizosphere redox potential (E(h)) over time were associated with the well-developed aerenchyma system in TYPHA: Typha roots, regardless of oxygen release pattern, showed low or undetectable alcohol dehydrogenase (ADH) activity or ethanol concentrations, indicating that ROL did not cause internal deficiencies . CLADIUM: roots also released oxygen, but this loss primarily occurred at the root tips and was accompanied by increased root ADH activity and ethanol concentrations . These results support the hypothesis that oxygen release by CLADIUM: is accompanied by internal deficiencies of oxygen sufficient to stimulate alcoholic fermentation and helps explain CLADIUM:'s lesser flood tolerance in comparison with TYPHA:

J Anim Sci, 2000 Aug, 78(8), 2215 - 22
Influence of feed intake fluctuation and frequency of feeding on nutrient digestion, digesta kinetics, and ruminal fermentation profiles in limit-fed steers; Soto-Navarro SA et al.; Nine crossbred beef steers (344 +/- 26 kg) fitted with ruminal cannulas were used in a randomized complete block design to evaluate the effects of feeding frequency and feed intake fluctuation on total tract digestion, digesta kinetics, and ruminal fermentation profiles in limit-fed steers . In Period 1, steers were allotted randomly to one of four dietary treatments: 1) feed offered once daily at 0800; 2) feed offered once daily at 0800 with a 10% fluctuation in day-to-day feed intake; 3) feed offered twice daily at 0800 and 1700; and 4) feed offered twice daily at 0800 and 1700 with a 10% fluctuation in a day-to-day feed intake . In Period 2, steers were reallocated across treatments . The 90% concentrate diet was fed at 90% of the ad-libitum consumption by each steer . Chromium-EDTA and Yb-labeled steam-flaked corn were intraruminally infused at 0800 on d 1 and 3 and Co-EDTA and Er-labeled steam-flaked corn were infused on d 2 and 4 of the 4-d collection period . Ruminal samples were collected at 0, 3, 6, 9, 12, 15, 18, and 24 h after the 0800 feeding, and total feces were collected for 4 d . Total tract digestibilities of OM, N, and starch were lowest (fluctuation x frequency, P < .05) when feed was offered twice daily with a 10% fluctuation in intake . Ruminal fluid volume and passage rate were not affected (P > .10) by feeding frequency or intake fluctuation . A frequency x fluctuation x sampling time interaction occurred (P < .01) for ruminal pH . Steers fed a constant amount of feed once daily had higher (P < .05) ruminal pH at 0, 3, 18, and 24 h than steers fed once daily with a 10% fluctuation in feed intake . Total VFA concentration was greater (P < .01) at 9 h after the 0800 feeding when feed was offered once vs twice daily . Feeding twice daily increased (P < .05) the molar proportion of acetate and decreased (P < .05) the molar proportion of propionate . Increasing feeding frequency resulted in a more stable ruminal environment; however, the increased acetate:propionate ratio with twice-daily feeding might result in lower efficiency of energy utilization by limit-fed steers.

J Anim Sci, 2000 Aug, 78(8), 2032 - 8
Effects of planting density and processing method on laboratory characteristics of grain sorghum for ruminants; Defoor PJ et al.; Grain sorghum grown in 38-cm (high-density) or 76-cm rows (normal-density) was steam-flaked, harvested as high-moisture grain followed by rolling and ensiling, or dry-rolled . Chemical composition, enzymatic starch availability, CP insolubility, and IVDMD in a reduced-strength buffer were evaluated . High-density planting increased (P < .10) OM and starch concentration and decreased (P < .0001) CP concentration but did not affect (P > .10) P concentration, enzymatic starch availability, or CP insolubility . High-density planting resulted in lower (P < .10) in vitro ruminal culture pH at 6, 12, and 18 h of incubation when grain sorghum was processed by steam flaking, and lower (P < .10) IVDMD at 6, 12, and 18 h of digestion when grain sorghum was processed by dry rolling . Steam flaking decreased (P < .10) CP concentration and solubility and increased (P < .10) OM concentration . High-moisture ensiling decreased (P < .10) the insolubility of CP but did not otherwise seem to alter the chemical composition of grain sorghum relative to dry rolling . Starch was more available (P < .10), and DM was digested more rapidly and extensively (P < .10) in vitro, in steam-flaked sorghum followed by high-moisture sorghum . Based on these data, it seems that planting density primarily affected chemical composition of grain sorghum, whereas processing primarily affected CP insolubility and rate and extent of starch fermentation.

Biosci Biotechnol Biochem, 2000 Jul, 64(7), 1337 - 44
Purification and characterization of acid-stable protopectinase produced by Aspergillus awamori in solid-state fermentation; Nagai M et al.; Aspergillus awamori IFO 4033 produced an acid-stable protopectinase in solid-state fermentation using wheat bran as the medium . The enzyme was purified to a homogeneous preparation with anion-exchange, hydrophobic, and size-exclusion chromatography . The enzyme was a monomeric protein of 52 kDa, by SDS-PAGE analysis, with an isoelectric point of pH 3.7 . The optimum pH for enzyme activity was 2.0, and it was most active at 50 degrees C (at pH 2.0) and was stable up to 50 degrees C (at pH 2.0) . The enzyme showed pectin-releasing activity toward protopectins from various origins, especially on lemon protopectin . An outstanding characteristic of the enzyme was its extreme stability in acidic conditions: the enzyme activity was not lost after incubating at pH 2.0 and 37 degrees C for 24 h.

J Pharm Sci, 2000 Sep, 89(9), 1097 - 105
Determination of erythromycin and related substances in enteric-coated tablet formulations by reversed-phase liquid chromatography; Wardrop J et al.; An isocratic method for the identification and quantitation of erythromycin and related substances in enteric-coated tablet formulations using high-performance liquid chromatography (HPLC) with ultraviolet (UV) detection at 205 nm is described . A novel method for sample preparation using a molecular weight centrifuge filter to reduce the interferences observed from polymeric tablet coating material is also presented . Erythromycin HPLC assays are best run at high pH; therefore, various polymer columns were evaluated . The resulting HPLC method that was developed has several advantages over current pharmacopeial assay methods for enteric-coated erythromycin tablets . Comparative data from both methods for the same batch of EryTab tablets are presented . The method can also be applied to various other erythromycin formulations, including particle-coated tablets, erythromycin stearate tablets, and erythromycin ethylsuccinate suspensions and fermentation broths . A C18 Polymeric column is used with a mobile phase composition of 0.02 M potassium phosphate dibasic buffer (pH 9): acetonitrile (60:40) and flow rate of 1 mL/min . This method is more sensitive, specific, and rugged than the pharmacopeial method.

J Exp Bot, 2000 Aug, 51(349), 1413 - 22
Anoxia tolerance in the aquatic monocot Potamogeton pectinatus absence of oxygen stimulates elongation in association with an unusually large pasteur effect; Summers JE et al.; Elongation by stems of overwintered tubers of Potamogeton pectinatus (L.) is strongly promoted over several days by oxygen-free conditions . Characteristics of the respiration underpinning this unusual response were examined . Anaerobic plants produced ethanol and CO(2) in approximately equimolar amounts, indicating that glycolysis coupled to alcoholic fermentation was the principal CO(2)-producing respiratory pathway . Rates of CO(2) evolution by aerobic and anaerobic whole plants (shoot and tuber) were similar, suggesting a rate of glycolysis three times that of aerobic plants, i.e . a strong Pasteur effect . In the shoot alone, anaerobic CO(2) production was twice the aerobic rate indicating a 6-fold increase in the rate of glycolysis in this tissue . Anoxic stems contained more sucrose at a stronger concentration than slower-growing aerobic stems or anaerobic leaves, demonstrating that sugar supply to the site of most rapid growth exceeded demand in the absence of oxygen . Concentrations of potentially toxic acetaldehyde in the external medium were small (approximately 0.2 mol m(-3)) during anoxia and on return to aerated conditions . Lactic acid was undetectable under anaerobic conditions and in vivo (31)P-NMR analysis of shoots revealed a cytoplasmic acidification of only </=0.2 pH units . In contrast, shoots of Pisum sativum, an anoxia-intolerant species, showed much stronger cytoplasmic acidification when transferred to oxygen-free conditions.

Vopr Pitan, 2000, 69(1-2), 50 - 2
{Effects of mixture of fermented whey SGOL-1-40 ("Sgidolac") and sterilized milk on the course of experimental tuberculosis}; Lind RM et al.; The effect of enzymolic whey SGOL 1-40 ("Sgidolac") and sterilized milk mixture on experimental tuberculosis process with mice has been investigated . The mice of CBA line infected with tuberculosis (Type H37Rv) and been given this mixture (rate SGOL 1-40 to milk 1:5) per os immediately after the infection at the amount of 0.5 g/Kg body weight per day every day, perished on 42-nd day, meanwhile the mice that had received the mixture 3 weeks before the infection and all the period after it, died on 46-th day he mice in the control group (infected and untreated) died on the 38-th day . The positive treatment and prophylactic effect of the SGOL 1-40 and milk mixture on the tuberculosis process has been stated and morphologically proved.

J Chromatogr B Biomed Sci Appl, 2000 Jun 23, 743(1-2), 5 - 12
Effect of biological suspensions on the position of the binodal curve in aqueous two-phase systems; Rito-Palomares M et al.; This study is concerned with the influence of biological suspension on the position of the binodal curve in aqueous two-phase systems (ATPSs) . Three different biological suspensions (i.e., disrupted yeast, E . coli homogenate and fermentation broth from Trichoderma harzianum) were selected and their impact upon ATPS performance was evaluated on the basis of changing volume ratio (Vr) and the position of the binodal curve of biological ATPSs (added with biomass) . Biological ATPSs with initial Vr greater than 1 and long (>40%, w/w) tie-line length (TLL), exhibited significant changes in Vr when compared with that from non-biological systems . Such behaviour was associated with the top phase biomass accumulation . It was shown that the addition of the biological suspensions used in this study to ATPSs caused the binodal curve to displace towards the origin, which was associated with the critical contribution of the bio-polymer (present in the systems) to the phase formation . The practical implementation of ATPSs for the purification of biological materials exploiting the information reported in this study is discussed.

Res Rep Health Eff Inst, 1999 Oct, (89), i - ii, 1-117; discussion 119-33
Reproductive and offspring developmental effects following maternal inhalation exposure to methanol in nonhuman primates; Burbacher T et al.; INTRODUCTION: In an effort to improve air quality and decrease dependence on petroleum, the federal government, industry, and other groups have encouraged development of alternative fuels such as methanol to substitute for gasoline or diesel fuel . Methanol is also a candidate to provide the hydrogen for fuel cells, which are being developed for a variety of power sources (including motor vehicle engines) . Before people are exposed to increased concentrations of methanol, the potential health effects of such exposures require study . Methanol, a simple alcohol containing one carbon atom, occurs naturally in plants and animals and participates in human metabolism . People regularly consume low doses of methanol in fruits, vegetables, and fermented beverages as well as soft drinks and foods sweetened with aspartame (which breaks down to methanol in the gastrointestinal tract) . Despite its ubiquitous presence, methanol can be highly toxic if sufficient quantities are consumed . Ingestion of methanol (usually in the form of wood alcohol or tainted alcoholic beverages) can result in metabolic acidosis, blindness, and even death . Although the body has the capacity to metabolize the low doses of methanol to which people are regularly exposed, it cannot handle high doses because too much methanol overwhelms the body's ability to remove a toxic metabolite (formate) . When formate accumulates, methanol poisoning occurs . One factor that regulates the rate at which formate is removed is the liver level of a derivative of the vitamin folic acid . People who are deficient in folic acid (including 15% to 30% of pregnant women) may be particularly susceptible to the toxic effects of methanol . If methanol were to be widely adopted as a fuel, environmental exposures would increase through ingestion of contaminated drinking water, inhalation of vapors from evaporative and other emissions, and dermal contact . Current concentrations of methanol in ambient air are very low, 1 to 30 parts per billion (ppb) . If all motor vehicles in the United States were converted to 100% methanol fuel, methanol levels in ambient air are estimated to increase approximately 1,000-fold (to 1 to 10 ppm in cities) and in a worst-case situation could occasionally reach concentrations as high as 200 ppm in enclosed spaces (HEI 1987) . Inhaling these concentrations of methanol for short periods of time is not predicted to affect formate production and thus should not present a health risk . However, little is known about the consequences of long-term inhalation of methanol vapors, especially in susceptible populations of pregnant women and developing fetuses . HEI, therefore, developed a research program to address this information gap . APPROACH: Dr . Thomas Burbacher and colleagues of the University of Washington studied the effects of long-term exposure to methanol vapors on metabolism and reproduction in adult female monkeys (Macaca fascicularis) and developmental effects in their offspring, who were exposed prenatally to methanol . The investigators exposed adult female monkeys (11 to 12 animals/group) to one of four concentrations of methanol vapors (0, 200, 600, and 1,800 ppm) for 2.5 hours a day, seven days a week during the following periods: (1) before breeding, (2) during breeding, and (3) during pregnancy . They collected blood from the adults at regular intervals to monitor methanol levels (which served as a marker of internal dose) and formate concentrations . They also conducted pharmacokinetic studies to determine whether methanol disposition (which includes absorption, distribution, metabolism, and excretion) was altered as a result of repeated methanol exposures and to assess pregnancy-related changes . Because high doses of methanol damage the central nervous system, the infants (8 to 9 animals/group) were examined at regular intervals during the first nine months of life to assess their growth and neurobehavioral development . RESULTS: Exposure to methanol vapors did n

Biotechnol Bioeng, 2000 Oct 5, 70(1), 65 - 71
Feasible boundaries of aqueous two-phase systems with NH(3) and CO(2) as recyclable volatile salts; van Berlo M et al.; Aqueous two-phase systems (ATPSs) have great potential for use in the downstream processing of fermentation products . A major drawback of these systems, limiting application in industrial practice up till now, is the consumption of large amounts of auxiliary materials such as polymers and salts . Making use of alternative auxiliaries can diminish this relatively large discharge . A possible approach is to make use of volatile salts induced by combinations of ammonia and carbon dioxide that can be recycled to the extraction system . As part of an ongoing research effort on ATPSs with volatile salts, this work aims at getting more information on the system boundaries or operating conditions of these systems in terms of phase behavior . The results show that the NH(3)/CO(2) ratio is an important parameter that has a large influence on the system boundaries . Both for systems with PEG 2000 and PEG 4000, this ratio has to be larger than about 1.75 to make a liquid-liquid phase separation possible . The most optimal ratio seems to be 2.0 for reasons of solution composition and absence of solid salt .

J Bacteriol, 2000 Sep, 182(17), 4934 - 40
Effects of limited aeration and of the ArcAB system on intermediary pyruvate catabolism in Escherichia coli; Alexeeva S et al.; The capacity of Escherichia coli to adapt its catabolism to prevailing redox conditions resides mainly in three catabolic branch points involving (i) pyruvate formate-lyase (PFL) and the pyruvate dehydrogenase complex (PDHc), (ii) the exclusively fermentative enzymes and those of the Krebs cycle, and (iii) the alternative terminal cytochrome bd and cytochrome bo oxidases . A quantitative analysis of the relative catabolic fluxes through these pathways is presented for steady-state glucose-limited chemostat cultures with controlled oxygen availability ranging from full aerobiosis to complete anaerobiosis . Remarkably, PFL contributed significantly to the catabolic flux under microaerobic conditions and was found to be active simultaneously with PDHc and cytochrome bd oxidase-dependent respiration . The synthesis of PFL and cytochrome bd oxidase was found to be maximal in the lower microaerobic range but not in a delta ArcA mutant, and we conclude that the Arc system is more active with respect to regulation of these two positively regulated operons during microaerobiosis than during anaerobiosis.

Int J Syst Evol Microbiol, 2000 Jul, 50 Pt 4, 1665 - 72
Use of conventional taxonomy, electrophoretic karyotyping and DNA-DNA hybridization for the classification of fermentative apiculate yeasts; Vaughan-Martini A et al.; A taxonomic study was conducted that considered strains of the genera Hanseniaspora/Kloeckera held in the Industrial Yeasts Collection (DBVPG) of the Dipartimento di Biologia Vegetale of the Universita di Perugia, Italy . Standard phenotypic as well as molecular criteria were considered in a effort to revisit the classification of these strains, some of which have been in the collection for about 50 years . Results of salient physiological tests showed that some of the DBVPG and type strains could not be identified by current taxonomic keys . Electrophoretic karyotypes were identical for some species, with the type strains of the seven accepted species showing only five distinct chromosomal patterns . DNA-DNA hybridization analyses, using a non-radioactive dot-blot technique, allowed for the distinction of taxa . The taxonomic implications of these results are discussed.

J Exp Bot, 2000 Apr, 51(345), 785 - 96
Regulation of alcoholic fermentation in coleoptiles of two rice cultivars differing in tolerance to anoxia; Gibbs J et al.; To investigate regulation of anaerobic carbohydrate catabolism in anoxia-tolerant plant tissue, rate of alcoholic fermentation and maximum catalytic activities of four key enzymes were assessed in coleoptiles of two rice cultivars that differ in tolerance to anoxia . The enzymes were ATP-dependent phosphofructokinase (PFK), pyrophosphate-dependent phosphofructokinase (PFP), pyruvate decarboxylase (PDC), and alcohol dehydrogenase (ADH) . During anoxia, rates of coleoptile elongation and ethanol synthesis were faster in the more tolerant variety Calrose than in IR22 . Calrose coleoptiles, in contrast to IR22, also showed a sustained Pasteur effect, with the estimated rate of glycolysis during anoxia being 1.4-1.7-fold faster than that of aerobic coleoptiles . In Calrose after 5 d anoxia, maximum catalytic activities of crude enzyme extracts were (in mumol substrate g-1 fresh weight min.-1) 170-240 for ADH, 4-6 for PDC and PFP and 0.4-0.7 for PFK . During anoxia, activity per coleoptile of all four enzymes increased 3-5.5-fold, suggesting that PFK, and PFP, like PDC and ADH, are synthesised in anoxic rice coleoptiles . Enzyme activities, on a fresh weight basis, were lower in IR22 than in Calrose . In vivo activities of PDC and PFK in anoxic coleoptiles from both cultivars were calculated using in vitro activities, estimated substrate levels, cytoplasmic pH, and S0.5 (the substrate level at which 0.5Vmax is reached, without inferring Michaelis-Menten kinetics) . Data indicated that potential carbon flux through PFK, rather than through PDC, more closely approximated rates of alcoholic fermentation . That PFK is an important site of regulation was supported further for Calrose coleoptiles by a decrease in the concentration of its substrate pool (F-6-P + G-6-P) following the onset of anoxia . By contrast, in IR22, there was little evidence for control by PFK, consistent with recent evidence that suggests substrate supply limits alcoholic fermentation in this cultivar.

Enzyme Microb Technol, 2000 Sep 1, 27(6), 376 - 389
Biocatalytic preparation of a chiral synthon for a vasopeptidase inhibitor: enzymatic conversion of N(2)-
Patel RN, Banerjee A, Nanduri VB, Goldberg SL, Johnston RM, Hanson RL, McNamee CG, Brzozowski DB, Tully TP, Ko RY, LaPorte TL, Cazzulino DL, Swaminathan S, Chen C, Parker LW, Venit JJ.
{4S-(4I,7I,10aJ)}1-Octahydro-5-oxo-4-{phenylmethoxy)carbonyl}amino}-7H-pyrido-{2,1-b} {1,3}thiazepine-7-carboxylic acid methyl ester (BMS-199541-01) is a key chiral intermediate for the synthesis of Omapatrilat (BMS-186716), a new vasopeptidease inhibitor under development . By using a selective enrichment culture technique we have isolated a strain of Sphingomonas paucimobilis SC 16113, which contains a novel L-lysine epsilon-aminotransferase . This enzyme catalyzed the oxidation of the epsilon-amino group of lysine in the dipeptide dimer N(2)-{N{phenyl-methoxy)-carbonyl} L-homocysteinyl} L-lysine)1,1-disulphide (BMS-201391-01) to produce BMS-199541-01 . The aminotransferase reaction required alpha-ketoglutarate as the amino acceptor . Glutamate formed during this reaction was recycled back to alpha-ketoglutarate by glutamate oxidase from Streptomyces noursei SC 6007 . Fermentation processes were developed for growth of S . paucimobilis SC 16113 and S . noursei SC 6007 for the production of L-lysine epsilon-amino transferase and glutamate oxidase, respectively . L-lysine epsilon-aminotransferase was purified to homogeneity and N-terminal and internal peptides sequences of the purified protein were determined . The mol wt of L-lysine epsilon-aminotransferase is 81 000 Da and subunit size is 40 000 Da . L-lysine epsilon-aminotransferase gene (lat gene) from S . paucimobilis SC 16113 was cloned and overexpressed in Escherichia coli . Glutamate oxidase was purified to homogeneity from S . noursei SC 6003 . The mol wt of glutamate oxidase is 125 000 Da and subunit size is 60 000 Da . The glutamate oxiadase gene from S . noursei SC 6003 was cloned and expressed in Streptomyces lividans . The biotransformation process was developed for the conversion of BMS-201391-01 to BMS-199541-01 by using L-lysine epsilon-aminotransferase expressed in E . coli . In the biotransformation process, for conversion of BMS-201391-01 (CBZ protecting group) to BMS-199541-01, a reaction yield of 65-70 M% was obtained depending upon reaction conditions used in the process . Phenylacetyl or phenoxyacetyl protected analogues of BMS-201391-01 also served as substrates for L-lysine epsilon-aminotransferase giving reaction yields of 70 M% for the corresponding BMS-199541-01 analogs . Two other dipeptides N-{N{(phenylmethoxy)carbonyl}-L-methionyl}-L-lysine (BMS-203528) and N,2-{S-acetyl-N-{(phenylmethoxy)carbonyl}-L-homocysteinyl}-L-lysine (BMS-204556) were also substrates for L-lysine epsilon-aminotransferase . N-alpha-protected (CBZ or BOC)-L-lysine were also oxidized by L-lysine epsilon-aminotransferase.

Enzyme Microb Technol, 2000 Sep 1, 27(6), 362 - 370
Production of alcohol from raw wheat flour by Amyloglucosidase and Saccharomyces cerevisiae; Montesinos T et al.; Ethanol production, by a simultaneous saccharification and fermentation process from raw wheat flour, has been performed by Saccharomyces cerevisiae and a low level of amyloglucosidase enzyme . The fermentation time was about 60 h after a 6 h pre-saccharification, with an amyloglucosidase (AMG) level of 270 AGU . kg(-1) starch, but only 31 h with a simultaneous saccharification fermentation process (SSF) . When an AMG level of 540 AGU . kg(-1) starch was used, the time decreased to 21 h, giving an ethanol concentration of 67 g . l(-1) . Sugar composition of the wort after the liquefaction may be responsible of the difference between these two process . Maltose, a fermentable sugar, was produced in high concentration during the liquefaction, allowing a shorter process period, counteracting the effect of the slow starch hydrolysis at 35 degrees C (SSF temperature).

Metab Eng, 1999 Oct, 1(4), 299 - 308
A maximum production strategy of lysine based on a simplified model derived from a metabolic reaction network; Shimizu H et al.; The aim of this study is to develop a strategy for maximum production of a target product with a simplified model derived from a metabolic reaction network through an example of lysine production . Based on the model, a search for the optimal specific growth rate profile was conducted among the available conditions of batch fermentation based on the derived model, when the total fermentation time was fixed . The optimal specific growth rate was obtained as a boundary control: initially, the specific growth rate was maintained at a maximum value and was subsequently switched to a critical value giving the maximum specific production rate . To make the specific growth rate follow this optimal profile as accurately as possible in batch mode, first, an appropriate initial concentration of leucine was employed in the experiment . Second, the feeding strategy of leucine was further studied . The specific growth rate profile with feeding was closer to the optimal one and the amount of lysine produced at the final stage of fermentation was increased about twofold, compared to that in the batch fermentation . Finally, the strategy was summarized as an algorithm for general use of this method.

J Mol Microbiol Biotechnol, 2000 Jan, 2(1), 27 - 32
The economics of acetone-butanol fermentation: theoretical and market considerations; Gapes JR; Acetone-butanol (AB) fermentation was once run commercially in many countries until these chemicals could be made more cheaply from fossil oil sources . Research into the revitalisation of the process has shown that the process could once again be run economically in niche markets if run in a relatively small industrial scale processing low-grade agricultural products . The following analysis is intended to help identify suitable niche markets.

J Mol Microbiol Biotechnol, 2000 Jan, 2(1), 15 - 20
The acetone-butanol fermentation in pilot plant and pre-industrial scale; Nimcevic D et al.; A summary of literature data concerning pilot or preindustrial scale trials of the acetone-butanol fermentation throughout its history is given . The recent pilot plant trials in Austria are also described for the first time . Some aspects of the current development of the acetone-butanol fermentation in general, especially from a technical point of view are also discussed.

J Mol Microbiol Biotechnol, 2000 Jan, 2(1), 5 - 8
The history of the acetone-butanol project in Austria; Gapes JR; The acetone-butanol fermentation and the closely related 2-propanol-butanol fermentation are of interest to Europe in particular for environmental and socioeconomic reasons, but its economic and technical feasibility must be proven before reestablishment as a commercial proposition . In particular the reestablishment of a new, fledgling fermentation industry selling into markets presently serviced by the mature, firmly established and highly capitalised petrochemical industry will require a significant driving force or else commercial users, who frequently have long-term supply contracts, will remain with known and proven suppliers . Hence the present state-of-the-art of the acetone-butanol fermentation is best described as technically and economically difficult but possible in niche markets . The most likely future is for decentral fermentation facilities processing locally made substrates and selling into niche markets.

J Biotechnol, 2000 Jul 28, 81(1), 27 - 34
Effect of nitrogen source and nitrogen concentration on the production of pyruvate by Torulopsis glabrata; Li Y et al.; The effect of nitrogen sources including yeast extract, peptone, soybean hydrolyzate and some inorganic nitrogen sources, as well as the nitrogen concentration on the fermentative production of pyruvate by Torulopsis glabrata WSH-IP12 was investigated . The addition of yeast extract greatly inhibited pyruvate accumulation, while peptone was shown to be the most favorable nitrogen source . In flask culture, 15 g l(-1) peptone was needed to consume 80 g l(-1) glucose with 23.4 g l(-1)of pyruvate accumulated . Pyruvate production was markedly dependent on the ratio of carbon to nitrogen (C:N), its production was improved by increasing the concentration of glucose and peptone proportionally and reduced by exclusively increasing the glucose concentration . In a glucose fed-batch culture, cell growth and pyruvate production slowed after 28 h . However, cell growth and pyruvate production recovered after further nitrogen, in the form of peptone and ammonium sulfate, was added to the culture . A final concentration of pyruvate of 54.5 g l(-1) was achieved at 64 h (yield to glucose consumed of 0.471 g g(-l)) . By using aqueous ammonia instead of potassium hydroxide for pH control, 57.3 g l(-1) pyruvate with a yield of 0.498 g g(-1) was produced by 55 h . This result further indicates that nitrogen level plays an important role in the production of pyruvate.

Plant Sci, 2000 Jul 28, 156(2), 151 - 158
Fermentative metabolism in grape berries: isolation and characterization of pyruvate decarboxylase cDNA and analysis of its expression throughout berry development; Or E et al.; The involvement of pyruvate decarboxylase (PDC) in the control of alcohol production during ripening of fruit tissues under aerobic conditions has been only partially studied . Enzymological studies showed a significant increase in PDC activity during the ripening of oranges and pears, concurrently with the induction of ethanol production . In tomato, on the other hand, the induction of ethanol production and ADH gene expression after the onset of ripening was not accompanied by induction of PDC activity . The isolation of PDC cDNA from fruits has not yet been reported, nor has its expression pattern during fruit development . We report here the isolation of a cDNA clone encoding for a grape PDC and the characterization of its expression throughout berry development . The pattern of PDC gene expression throughout berry development, combined with earlier findings on constitutive PDC activity in the berry, may suggest that PDC is not the limiting factor for the production of ethanol in the berry, which is induced only after the onset of berry ripening . Alternatively, the induction of ADH gene expression, which occurs only after the onset of ripening in both tomatoes and grape berries, may serve as a regulator of ethanol production in response to a ripening-related cue.

Metab Eng, 2000 Jan, 2(1), 69 - 77
Optimization of ethanol production in Saccharomyces cerevisiae by metabolic engineering of the ammonium assimilation; Nissen TL et al.; Ethanol is still one of the most important products originating from the biotechnological industry with respect to both value and amount . In addition to ethanol, a number of byproducts are formed during an anaerobic fermentation of Saccharomyces cerevisiae . One of the most important of these compounds, glycerol, is produced by yeast to reoxidize NADH, formed in synthesis of biomass and secondary fermentation products, to NAD+ . The purpose of this study was to evaluate whether a reduced formation of surplus NADH and an increased consumption of ATP in biosynthesis would result in a decreased glycerol yield and an increased ethanol yield in anaerobic cultivations of S . cerevisiae . A yeast strain was constructed in which GLN1, encoding glutamine synthetase, and GLT1, encoding glutamate synthase, were overexpressed, and GDH1, encoding the NADPH-dependent glutamate dehydrogenase, was deleted . Hereby the normal NADPH-consuming synthesis of glutamate from ammonium and 2-oxoglutarate was substituted by a new pathway in which ATP and NADH were consumed . The resulting strain TN19 (gdh1-A1 PGK1p-GLT1 PGK1p-GLN1) had a 10% higher ethanol yield and a 38% lower glycerol yield compared to the wild type in anaerobic batch fermentations . The maximum specific growth rate of strain TN19 was slightly lower than the wild-type value, but earlier results suggest that this can be circumvented by increasing the specific activities of Gln1p and Glt1p even more . Thus, the results verify the proposed concept of increasing the ethanol yield in S . cerevisiae by metabolic engineering of pathways involved in biomass synthesis.

Metab Eng, 1999 Apr, 1(2), 141 - 52
Redistribution of metabolic fluxes in Escherichia coli with fermentative lactate dehydrogenase overexpression and deletion; Yang YT et al.; Under anaerobic conditions, competition for pyruvate between the branch point enzymes pyruvate formate lyase (PFL, Km = 2 mM) and fermentative lactate dehydrogenase (LDH, Km = 7.2 mM) determines the partition of carbon flux . Two Escherichia coli mutant strains, one deficient in ackA, pta, and ldhA and the other overexpressing LDH, were constructed to systematically analyze the effects of these perturbations in the existing pathways on the redistribution of carbon fluxes . Deletion of the lactate and acetate synthesis pathways was detrimental to cell growth . Carbon flux is forced through ethanol and formate production pathways, resulting in a concomitant increase in those fluxes . In addition, overexpression of LDH simultaneously increases the common flux as well as the flux to the competing acetyl-CoA branch . Overexpression of lactate dehydrogenase (ldhA) in the parent strain increases the lactate synthesis rate from 0.19 to 0.40 mmol/g-biomass-h when the LDH activities increases from 1.3 to 15.3 units . Even an increase of more than 10 times in the LDH activity fails to divert a large fraction of the carbon flux to lactate; the majority of the flux still channels through the acetyl-CoA branch . Overexpression of LDH in the parent strain simultaneously increases the common flux as well as the flux through the acetyl-CoA branch . Subsequently, the flux amplification factors (or deviation indices which can be related to the flux control coefficients) are positive for all three fluxes occurring at the pyruvate node.

Chin J Biotechnol, 1999, 15(3), 165 - 70
Cultivation of encapsulated yeast cells in NaCS-PDMDAAC polyelectrolyte complexes; Zhang H et al.; The cultivation of encapsulated S . cerevisiate and C . utilis in NaCS-PDMDAAC Polyelectrolyte Complexes was studied . The results showed that these two kinds of encapsulated microorganisms showed the same growing properties as in their free cell cultures, and thus NaCS-PDMDAAC microcapsules were suitable for the encapsulation of biological substances . The encapsulated S . cerevisiate cells were fermented sequentially for 16 batches . The highest cell density in the capsules reached 2.64 x 10(10) cells/mL and ethanol concentration reached 47.0 g/L . The encapsulated C . utilis produced the same amount of GSH as the free cell culture.

Biotechnol Prog, 2000 Jul-Aug, 16(4), 661 - 7
Selective flocculation and precipitation for the improvement of virus-like particle recovery from yeast homogenate; Tsoka S et al.; The purification of an intracellular product from a complex mixture of contaminants after cell disruption is a common problem in processes downstream of fermentation systems . This is particularly challenging for the recovery of particulate (80 nm in diameter) multimeric protein products, named virus-like particles (VLPs), from cell debris and other intracellular components . Selective flocculation for debris removal followed by selective precipitation of the target protein can be used as a preclarification step to aid purification . In this paper, selective borax flocculation of cell debris in yeast homogenate, followed by selective poly(ethylene glycol) precipitation of VLPs are defined with a view to demonstrating their potential in aiding the initial clarification stages of the purification sequence . The translation from laboratory scale to pilot scale operation is addressed, demonstrating the challenge of scale-up of solid-liquid separation stages for biological particle processing.

Biotechnol Prog, 2000 Jul-Aug, 16(4), 571 - 6
A recombinant lipoprotein antigen against Lyme disease expressed in E . coli: fermentor operating strategies for improved yield; Madurawe RD et al.; Decorin-binding lipoprotein, lpp-DBP, a bacterial surface adhesin, shows promise as a vaccine against Lyme disease . It is expressed in recombinant E . coli as an undesirable 20.5 KDa apoprotein that is subsequently lipidated in vivo to the desired 22 KDa lpp-DBP form . This study defines fermentation conditions for maximizing lpp-DBP yield . Super broth medium, a low post-induction temperature (30 degrees C), and a glucose feed based on dissolved oxygen resulted in high lpp-DBP yield and minimized apoprotein formation . Since cells lysed within 2-3 h after induction, the cell yield was maximized by growing cells to high cell density prior to induction . Compared to a glucose feed based on maintaining a constant fermentor glucose concentration (Glucose-Stat), feeding based on maintaining a constant dissolved oxygen level (DO-Stat) improved yields . Also, a dissolved oxygen level of 60% (air saturation) was best, as no product degradation was detected by Western blotting and SDS-PAGE . Acetic acid levels under both modes of glucose feed were sufficiently low, and no adverse growth effects were observed.

Biotechnol Prog, 2000 Jul-Aug, 16(4), 553 - 6
Precursor-directed biosynthesis of 6-deoxyerythronolide B analogs in Streptomyces coelicolor: understanding precursor effects; Leaf T et al.; A fermentation process employing precursor-directed biosynthesis is being developed for the manufacture of 6-deoxyerythronolide B (6-dEB) analogues . Through a plasmid-based system in Streptomyces coelicolor, 6-dEB synthesis is catalyzed by 6-dEB synthase (DEBS) . 6-dEB synthesis is abolished by inactivation of the ketosynthase (KS) 1 domain of DEBS but can be restored by providing synthetic activated diketides . Because of its inherent catalytic flexibility, the KS1-deficient DEBS is capable of utilizing unnatural diketides to form various 13-substituted 6-dEBs . Here we characterize process variables associated with diketide feeding in shake-flask experiments . 13-R-6-dEB production was found to depend strongly on diketide feed concentrations, on the growth phase of cultures at feeding time, and on the R-group present in the diketide moiety . In all cases a major portion of the fed diketides was degraded by the cells.

Biotechnol Prog, 2000 Jul-Aug, 16(4), 541 - 7
Dry-grind process for fuel ethanol by continuous fermentation and stripping; Taylor F et al.; Conversion of a high-solids saccharified corn mash to ethanol by continuous fermentation and stripping was successfully demonstrated in a pilot plant consuming 25 kg of corn per day . A mathematical model based on previous pilot plant results accurately predicts the specific growth rate obtained from these latest results . This model was incorporated into a simulation of a complete dry-grind corn-to-ethanol plant, and the cost of ethanol production was compared with that of a conventional process . The results indicate a savings of $0.03 per gallon of ethanol produced by the stripping process . The savings with stripping result from the capacity to ferment a more concentrated corn mash so there is less water to remove downstream.

Food Addit Contam, 2000 Jun, 17(6), 469 - 75
Determination of ethyl carbamate in some fermented Korean foods and beverages; Kim YK et al.; Ethyl carbamate has been associated with cancer for several decades . It is mainly found in fermented foods and beverages . In view of the importance of fermented foods in the Korean diet and the significant level of ethyl carbamate expected, we determined ethyl carbamate concentrations in some of the staple food items and estimated the daily intake for the Korean population . Ethyl carbamate in commercial samples of kimchi, soy sauce, vinegar, soybean paste, and alcoholic beverages were determined by gas chromatography-mass spectrometry/selective ion monitoring (GC-MS/SIM) . Homemade soy sauce and kimchi were also analysed . The maximum ethyl carbamate concentrations observed were 73 micrograms/kg in soy sauce, 7.9 micrograms/kg in soybean paste, 2.5 micrograms/l in vinegar, 16.2 micrograms/kg in kimchi and 15.4 mu/l in Korean traditional alcoholic beverages . Combining these values with the average daily food intake data, we estimated that the maximum daily exposure of Korean population to ethyl carbamate is 2.8 micrograms/day, which is not a negligible amount considering the 'virtually safe dose' derived by animal experiment ranges between 1.2 and 4.8 micrograms/day . It would be desirable to closely monitor ethyl carbamate levels in Korean foods and to find ways to reduce the daily intake.

Rev Latinoam Microbiol, 1998 Jan-Jun, 40(1-2), 109 - 19
{Biosynthesis of congeners during alcohol fermentation}; Santillan-Valverde MC et al.; The flavor of alcoholic beverages is a consequence of a complex mixture of many compounds, including small concentrations of some volatile metabolites known as congeners, which are produced by the yeast during the fermentation . The more important compounds are those that can be found in all the alcoholic beverages in different concentrations, and they can be grouped on the following chemical species: higher alcohols, esters, and carboniles . In the current paper the biochemical pathways that produce these compounds from the raw materials are reviewed . Research done in this field has led to a more complete knowledge concerning to organoleptic profiles of alcoholic beverages and to a better control for the production of the final product.

Syst Appl Microbiol, 2000 Jun, 23(2), 300 - 3
Expression levels and patterns of glycolytic yeast genes during wine fermentation; Puig S et al.; Promoters from glycolytic genes are widely used for gene overexpression in the yeast Saccharomyces cerevisiae . Wine strains are not an exception, and genes of enological interest have been expressed in this way . However, the transcriptional pattern of glycolytic genes has never been studied during wine fermentation . In this work we describe the levels and expression patterns of glycolytic genes for a wine yeast strain during the alcoholic fermentation of three different musts . Results show similar transcriptional patterns for all genes studied: maximal levels of mRNA at the exponential growth stage, and a gradual decrease during the stationary phase, in accordance with the fermentation rates . The glyceraldehyde 3-phosphate dehydrogenase genes reach the highest transcriptional levels during wine fermentation, similarly as previously described for laboratory strains and conditions.

Syst Appl Microbiol, 2000 Jun, 23(2), 165 - 73
The role of cold-shock proteins in low-temperature adaptation of food-related bacteria; Wouters JA et al.; There is a considerable interest in the cold adaptation of food-related bacteria, including starter cultures for industrial food fermentations, food spoilage bacteria and food-borne pathogens . Mechanisms that permit low-temperature growth involve cellular modifications for maintaining membrane fluidity, the uptake or synthesis of compatible solutes, the maintenance of the structural integrity of macromolecules and macromolecule assemblies, such as ribosomes and other components that affect gene expression . A specific cold response that is shared by nearly all food-related bacteria is the induction of the synthesis so-called cold-shock proteins (CSPs), which are small (7 kDa) proteins that are involved in mRNA folding, protein synthesis and/or freeze protection . In addition, CSPs are able to bind RNA and it is believed that these proteins act as RNA chaperones, thereby reducing the increased secondary folding of RNA at low temperatures . In this review established and novel aspects concerning the structure, function and control of these CSPs are discussed . A model for bacterial cold adaptation, with a central role for ribosomal functioning, and possible mechanisms for low-temperature sensing are discussed.

Plant J, 2000 Jul, 23(1), 115 - 22
Functional identification of an Arabidopsis snf4 ortholog by screening for heterologous multicopy suppressors of snf4 deficiency in yeast; Kleinow T et al.; Yeast Snf4 is a prototype of activating gamma-subunits of conserved Snf1/AMPK-related protein kinases (SnRKs) controlling glucose and stress signaling in eukaryotes . The catalytic subunits of Arabidopsis SnRKs, AKIN10 and AKIN11, interact with Snf4 and suppress the snf1 and snf4 mutations in yeast . By expression of an Arabidopsis cDNA library in yeast, heterologous multicopy snf4 suppressors were isolated . In addition to AKIN10 and AKIN11, the deficiency of yeast snf4 mutant to grown on non-fermentable carbon source was suppressed by Arabidopsis Myb30, CAAT-binding factor Hap3b, casein kinase I, zinc-finger factors AZF2 and ZAT10, as well as orthologs of hexose/UDP-hexose transporters, calmodulin, SMC1-cohesin and Snf4 . Here we describe the characterization of AtSNF4, a functional Arabidopsis Snf4 ortholog, that interacts with yeast Snf1 and specifically binds to the C-terminal regulatory domain of Arabidopsis SnRKs AKIN10 and AKIN11.

Z Naturforsch {C}, 2000 May-Jun, 55(5-6), 341 - 6
Oxidative metabolism of ambrox and sclareolide by Botrytis cinerea; Farooq A et al.; Ambrox (1), a perfumery diterpene, was oxidatively metabolised by a plant pathogenic fungus Botrytis cinerea in a xenobiotic fashion to afford a major product, i.e., 1beta-hydroxy-8-epiambrox (13) (60%) along with three minor metabolites 3beta-hydroxyambrox (2), sclareolide (5) and 3beta-hydroxysclareolide (7) . Sclareolide (5), a cytotoxic diterpenoidal lactone was fermented with the same fungus to yield 3beta-hydroxysclareolide (7) (59%) as a major metabolite together with two minor metabolites characterised as 1-ketosclareolide (15), and 3beta,14-dihydroxysclareolide (16).

Anticancer Res, 2000 May-Jun, 20(3B), 2245 - 8
Population thiamine status and varying cancer rates between western, Asian and African countries; Boros LG; The role of food supplements in the form of vitamins has not been extensively investigated in relation to varying cancer rates between populations of different geographical regions . New data indicate that thiamine (vitamin B1), a common food supplement in Western food products, is directly involved in nucleic acid ribose synthesis of tumor cells in its biologically activated form through the non-oxidative transketolase catalyzed pentose cycle reaction . Whether thiamine plays a role in increased cancer rates in the Western World by enhancing tumor cell proliferation, while increased consumption of thiaminase rich food limiting thiamine availability protects against common malignancies in Asia and Africa has not been evaluated . In the Western World, thiamine is a popular vitamin supplement in the form of tablets and it is also added to basic food items such as milled flour, cereals, peanut butter, refreshment drinks and pastas . On the contrary, thiaminase, the natural thiamine-degrading enzyme, is abundantly present in raw and fermented fish, certain vegetables and roasted insects consumed primarily in Africa and Asia . Excess thiamine supplementation in common food products may contribute to the increased cancer rates of the Western World.

Fertil Steril, 2000 Aug, 74(2), 203 - 12
Use of fibrinolytic agents in the prevention of postoperative adhesion formation; Hellebrekers BW et al.; OBJECTIVE: To review the events leading to the formation of adhesions, to describe the development of fibrinolytic agents, to review more than a century of research on the use of fibrinolytic agents in adhesion prevention, and to look at future aspects of adhesion prevention . RESULTS: A better understanding of the pathogenesis of adhesion formation has resulted in the use of fibrinolytic agents in their prevention . Fibrinolytic agents promote fibrinolytic activity during the early period after peritoneal trauma during which an increased formation of fibrin is seen in combination with a deficiency of endogenous fibrinolytic activity . Initially, chemical attacks on fibrin (fibrolysin and hypertonic glucose), foreign digestive ferments (pepsin, trypsin, and papain), and stimulation of intraperitoneal leukocytosis (amniotic fluid) were used . Development of new thrombolytic agents was soon followed by experiments in animal adhesion models and clinical studies to examine their antiadhesion properties . Plasmin preparations (plasmin, actase, and fibrinolysin) and plasmin activators (streptokinase, urokinase, and tissue-type plasminogen activator) were found to be efficacious in preventing adhesion formation in the greater part of reviewed animal and clinical studies . CONCLUSION(S): From the current literature, it can be concluded that postoperative intraperitoneal administration of thrombolytic agents can significantly decrease adhesion formation . Given the large number of experimental studies in animals, future studies should focus on the clinical use of fibrinolytic agents in the prevention of postsurgical adhesion formation.

Biochem J, 2000 Aug 15, 350 Pt 1, 261 - 8
Expression of escherichia coli otsA in a Saccharomyces cerevisiae tps1 mutant restores trehalose 6-phosphate levels and partly restores growth and fermentation with glucose and control of glucose influx into glycolysis; Bonini BM et al.; The TPS1 gene, encoding trehalose-6-phosphate synthase (TPS), exerts an essential control on the influx of glucose into glycolysis in the yeast Saccharomyces cerevisiae . The deletion of TPS1 causes an inability to grow on glucose because of a hyperaccumulation of sugar phosphates and depletion of ATP and phosphate . We show that expression of the Escherichia coli homologue, otsA, in a yeast tps1 mutant results in high TPS activity . Although the trehalose 6-phosphate (Tre6P) level during exponential growth on glucose was at least as high as in a wild-type yeast strain, growth on glucose was only partly restored and the lag phase was much longer . Measurement of the glycolytic metabolites immediately after the addition of glucose showed that in spite of a normal Tre6P accumulation there was still a partial hyperaccumulation of sugar phosphates . Strong elevation of the Tre6P level by the additional deletion of the TPS2 gene, which encodes Tre6P phosphatase, was not able to cause a strong decrease in the sugar phosphate levels in comparison with the wild-type strain . In addition, in chemostat experiments the short-term response to a glucose pulse was delayed, but normal metabolism was regained over a longer period . These results show that Tre6P synthesis from a heterologous TPS enzyme can to some extent restore the control of glucose influx into glycolysis and growth on glucose in yeast . However, they also indicate that the yeast TPS enzyme, as opposed to the E . coli otsA gene product, is able to increase the efficiency of the Tre6P control on glucose influx into yeast glycolysis.

Membr Cell Biol, 2000, 13(4), 511 - 26
Relationship between formate hydrogen lyase and proton-potassium pump under heterolactic fermentation in Escherichia coli: functional multienzyme associations in the cell membrane; Trchounian AA et al.; Anaerobically grown glucose-fermenting E . coli cells produce molecular hydrogen, acidify the medium and uptake potassium ions . It was shown that the H2 release and the proton-potassium exchange with the fixed (2H+/K+) stoichiometry of the initial DCC-sensitive fluxes were lost in mutants with the deleted fdhF gene or the hycA-H operon responsible for the biosynthesis of formate dehydrogenase H (FDH,H) or hydrogenase 3 (H3), respectively, which are the main components of the formate hydrogen lyase FHL(H) . However, both processes occurred in mutants with the deleted hycE, hycF or hycG genes encoding the major and minor components of H3, respectively . The K+ uptake was sensitive to the osmotic shock resulting from glucose addition to the medium and decreased significantly in the presence of valinomycin . The H2 release and the 2H+/K+ exchange were absent in the mutant with the deleted hycB gene encoding the corresponding minor component of H3 . This mutant acidified the medium and uptook K+ with Km typical for TrkA, but the stoichiometry of the DCC-inhibited fluxes was variable, and the K+ gradient between the cytoplasm and the medium in this mutant was lower than in the mutants lacking other minor components of H3 . The results obtained suggest that the hycB gene product, FdhF and HycE, form probably the FHL(H) complex that directly interacts with the H+-ATPase complex F0F1 and the TrkA(H) system of K+ uptake . Such a multienzyme association is responsible for the H2 production and 2H+/K+ exchange . The major and other minor components of H3 have probably no direct role in the H2 production and 2H+/K+ exchange . H2 production by precursor's or hycE mutant's protoplasts treated with toluene was shown to occur upon addition of the thiol reagent dithiothreitol to the medium containing ATP, potassium ions, NAD+, and NADH . H2 production was inhibited by DCC . The quantity of available thiol groups in membrane vesicles of the precursor or the hycE, hycF or hycG mutants, in which the H2 production and 2H+/K+ exchange were observed, was larger than in other mutants . The number of SH groups decreased in the presence of DCC . These results indicate a significance of the thiol groups for the function of the proposed association.

Appl Environ Microbiol, 2000 Aug, 66(8), 3151 - 9
Engineering of the pyruvate dehydrogenase bypass in Saccharomyces cerevisiae: role of the cytosolic Mg(2+) and mitochondrial K(+) acetaldehyde dehydrogenases Ald6p and Ald4p in acetate formation during alcoholic fermentation; Remize F et al.; Acetic acid plays a crucial role in the organoleptic balance of many fermented products . We have investigated the factors controlling the production of acetate by Saccharomyces cerevisiae during alcoholic fermentation by metabolic engineering of the enzymatic steps involved in its formation and its utilization . The impact of reduced pyruvate decarboxylase (PDC), limited acetaldehyde dehydrogenase (ACDH), or increased acetoacetyl coenzyme A synthetase (ACS) levels in a strain derived from a wine yeast strain was studied during alcoholic fermentation . In the strain with the PDC1 gene deleted exhibiting 25% of the PDC activity of the wild type, no significant differences were observed in the acetate yield or in the amounts of secondary metabolites formed . A strain overexpressing ACS2 and displaying a four- to sevenfold increase in ACS activity did not produce reduced acetate levels . In contrast, strains with one or two disrupted copies of ALD6, encoding the cytosolic Mg(2+)-activated NADP-dependent ACDH and exhibiting 60 and 30% of wild-type ACDH activity, showed a substantial decrease in acetate yield (the acetate production was 75 and 40% of wild-type production, respectively) . This decrease was associated with a rerouting of carbon flux towards the formation of glycerol, succinate, and butanediol . The deletion of ALD4, encoding the mitochondrial K(+)-activated NAD(P)-linked ACDH, had no effect on the amount of acetate formed . In contrast, a strain lacking both Ald6p and Ald4p exhibited a long delay in growth and acetate production, suggesting that Ald4p can partially replace the Ald6p isoform . Moreover, the ald6 ald4 double mutant was still able to ferment large amounts of sugar and to produce acetate, suggesting the contribution of another member(s) of the ALD family.

Appl Microbiol Biotechnol, 2000 Jun, 53(6), 646 - 9
Economic considerations in the production of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) by bacterial fermentation; Choi J et al.; The process for the production of poly(3-hydroxybutyrate-co-3-hydroxyvalerate) {P(3HB/V)} by bacterial fermentation and its recovery was analysed . The effects of various factors such as P(3HB/V) content, P(3HB/V) productivity, P(3HB/V) yield and 3-hydroxyvalerate (3HV) fraction in P(3HB/V) on the production cost of P(3HB/V) were examined . The increase in the 3HV yield on a carbon source did not significantly decrease the production cost when the 3HV fraction was 10 mol%, because the cost of the carbon substrate for 3HV was relatively small in terms of the total cost . However, at a 3HV fraction of 30 mol%, the 3HV yield on a carbon source had a significant effect on the total P(3HB/V) production cost . The production cost of P(3HB/V) increased linearly with the increase in the 3HV fraction in P(3HB/V).

Farmaco, 2000 Mar, 55(3), 165 - 7
Recombinant production of PIGF-1 and its activity in animal models; Maglione D et al.; In this paper we review current knowledge on placenta growth factor (PIGF) and summarise our data on its recombinant production in bacteria and its activity . PIGF and vascular endothelial growth factor (VEGF) are both angiogenic factors belonging to the platelet-derived growth factor (PDGF) family . PIGF is a dimeric glycoprotein which shares a number of biochemical and functional features with VEGF . The aminoacidic similarity between the two factors is high (about 50%) in the PDGF-like domain . By alternative splicing of the PIGF mRNA, three forms of PIGF protein are generated which are named PIGF-1, PIGF-2 and PIGF-3 . We have focused our attention on form 1 of human PIGF (PIGF-1) . A large quantity of active recombinant PIGF-1 has been obtained using a bacterial expression system . By optimising the fermentation and purification it was possible to produce about 140 mg/l of culture of active PIGF-1, which is potentially suitable for a pharmaceutical use . The angiogenic activity of two different batches of bacteria-derived PIGF-1 obtained in our laboratory was demonstrated in chick chorionallantoic membrane assays . Finally, in preliminary studies we have shown that bacteria-derived PIGF-1 has a protective effect against myocardial lesions induced by isoprenaline in rat and rabbit.

J Nutr, 2000 Aug, 130(8), 1991 - 5
Fermentation of resistant rice starch produces propionate reducing serum and hepatic cholesterol in rats; Cheng HH et al.; This study was designed to investigate the effects of different proportions of rice starch and cornstarch on lipid metabolism in rats fed high dietary cholesterol . Male Wistar rats were fed a 10 g/100 g fat diet containing 1 g/100 g cholesterol with 0 (control diet), 15, 30, 45 or 63 g/100 g rice starch with an enzyme resistant starch concentration of 1.26, 1.39, 1.52, 1.65 or 1.80 g/100 g, respectively, for 4 wk . Groups fed diets with < 63 g/100 g rice starch were supplemented with cornstarch to 63 g/100 g . The two kinds of starch had different structures as seen using scanning electron microscopy (SEM) . The rice starch was an aggregation (n = 20-60) of smaller granules (3-8 microm in diameter), whereas the cornstarch was composed of larger (5-15 microm in diameter), single granules . The compound rice starch (0.99 kg/L) was larger in size and denser in structure than cornstarch (0.63 kg/L) . Serum total cholesterol concentrations in rats fed both the 45 and 63 g/100 g rice starch diets were significantly lower than in all other groups (P < 0.05) . The serum propionate concentration in the rats fed 63 g/100 g rice starch diets was significantly higher than that of other groups . Hepatic triglyceride and total cholesterol concentrations in rats fed 63 g/100 g rice starch diets were significantly lower than in the control group . These results suggest that, because the compound rice starch was an aggregation of smaller granules, larger in size and denser in structure than cornstarch, it was digested more slowly and altered lipid metabolism . Resistant rice starch may be fermented to produce propionate, which reduces serum and hepatic cholesterol.

J Nutr, 2000 Aug, 130(8), 1887 - 93
Isoflavone aglycone-rich extract without soy protein attenuates atherosclerosis development in cholesterol-fed rabbits; Yamakoshi J et al.; The antiatherogenic effect of soy protein with intact isoflavones is well established, but the effects of isoflavones without soy protein have not been determined . We investigated the antiatherogenic effect of an isoflavone aglycone-rich extract (containing 429.4 mg/g isoflavone aglycones) without soy protein from fermented soy in cholesterol-fed rabbits . We fed 12-wk-old New Zealand white male rabbits diets containing 1 g/100 g cholesterol with 0, 0.33 or 1 g/100 g isoflavone aglycones for 8 wk . We also fed the rabbits a diet containing 1 g/100 g cholesterol with 1.09 g/100 g soy saponin-rich extract, a component other than isoflavone aglycones in the isoflavone aglycone-rich extract . Controls did not consume cholesterol, isoflavone aglycones or saponins . The isoflavone aglycone- and saponin-rich extracts did not affect the serum lipid profile of cholesterol-fed rabbits . The serum concentration of daidzein in its conjugated form was significantly higher in the high isoflavone group than in the low isoflavone group . The level of cholesteryl ester hydroperoxide (ChE-OOH) induced by CuSO(4) in plasma in the high isoflavone group was significantly less than that in the cholesterol group, and the ChE-OOH levels of LDL in the low and high isoflavone groups were significantly less than those in the cholesterol group . The ChE-OOH levels in plasma and LDL in the saponin group did not differ from the cholesterol group . In the aortic arch, the cholesterol concentration was significantly lower in the high isoflavone group, and malondialdehyde concentration was significantly lower in the low and high isoflavone groups compared with the cholesterol group; however these concentrations in the saponin group did not differ from those in the cholesterol group . The atherosclerotic lesion area of the aortic arch was significantly lower in the isoflavone groups (26.3% lower in the low isoflavone group and 36.9% lower in the high isoflavone group) than in the cholesterol group . The lesion areas were not different in the soy saponin and cholesterol groups . Immunohistochemical analysis revealed fewer oxidized LDL-positive macrophage-derived foam cells in atherosclerotic lesions in the aortic arch of isoflavone groups compared with that of the cholesterol group . These results suggest that the antioxidative action of isoflavones and their antioxidative metabolites inhibit the oxidation of LDL, thereby exerting an antiatherosclerotic effect.

J Biol Chem, 2000 Oct 27, 275(43), 33869 - 75
Evolution of the adhE gene product of Escherichia coli from a functional reductase to a dehydrogenase . Genetic and biochemical studies of the mutant proteins; Membrillo-Hernandez J et al.; The multifunctional AdhE protein of Escherichia coli (encoded by the adhE gene) physiologically catalyzes the sequential reduction of acetyl-CoA to acetaldehyde and then to ethanol under fermentative conditions . The NH(2)-terminal region of the AdhE protein is highly homologous to aldehyde:NAD(+) oxidoreductases, whereas the COOH-terminal region is homologous to a family of Fe(2+)-dependent ethanol:NAD(+) oxidoreductases . This fusion protein also functions as a pyruvate formate lyase deactivase . E . coli cannot grow aerobically on ethanol as the sole carbon and energy source because of inadequate rate of adhE transcription and the vulnerability of the AdhE protein to metal-catalyzed oxidation . In this study, we characterized 16 independent two-step mutants with acquired and improved aerobic growth ability on ethanol . The AdhE proteins in these mutants catalyzed the sequential oxidation of ethanol to acetaldehyde and to acetyl-CoA . All first stage mutants grew on ethanol with a doubling time of about 240 min . Sequence analysis of a randomly chosen mutant revealed an Ala-267 --> Thr substitution in the acetaldehyde:NAD(+) oxidoreductase domain of AdhE . All second stage mutants grew on ethanol with a doubling time of about 90 min, and all of them produced an AdhE(A267T/E568K) . Purified AdhE(A267T) and AdhE(A267T/E568K) showed highly elevated acetaldehyde dehydrogenase activities . It therefore appears that when AdhE catalyzes the two sequential reactions in the counter-physiological direction, acetaldehyde dehydrogenation is the rate-limiting step . Both mutant proteins were more thermosensitive than the wild-type protein, but AdhE(A267T/E568K) was more thermal stable than AdhE(A267T) . Since both mutant enzymes exhibited similar kinetic properties, the second mutation probably conferred an increased growth rate on ethanol by stabilizing AdhE(A267T).

Mikrobiologiia, 2000 May-Jun, 69(3), 410 - 4
{Saccharomyces bayanus var . Uvarum comb.nov., a new variety established by genetic analysis}; Naumov GI; Partial genetic isolation of two Saccharomyces bayanus varieties, S . bayanus var . bayanus and S . bayanus var . uvarum comb . nov., was established by hybridological analysis . The hybrids of these two varieties were semisterile: their ascospores were characterized by low survival . Earlier, the new variety was described as a group of cryophilic wine yeast cultivars capable of fermenting melibiose.

Acta Vet Scand, 2000, 41(1), 25 - 40
Functional anatomy of the omasum in high Arctic Svalbard reindeer (Rangifer tarandus platyrhynchus) and Norwegian reindeer (Rangifer tarandus tarandus); Mathiesen SD et al.; The structure and fill of the omasum was investigated in summer and in winter in adult female reindeer living on the polar desert and tundra of the high Arctic archipelago of Svalbard and in sub-Arctic mountain habitats in northern Norway . The mean total mass of the omasum in non-lactating adult female Svalbard reindeer was 467 g (0.65 g per 100 g live body mass (BM)) in September and 477 g (1.03 g per 100 g BM) in April . By contrast, the mean mass of the omasum in non-lactating adult reindeer in northern Norway was 534 g (0.83 g per 100 g BM) in September but only 205 g (0.35 g per 100 g BM p < 0.05) in late March, owing to a decrease in both tissue mass and the wet mass of the contents of the organ . The mean absorptive surface of the omasum in Svalbard reindeer was 2300 cm2 in September and 2023 cm2 in April . In Norwegian reindeer, by contrast, the absorptive surface area decreased from 2201 cm2 in September to 1181 cm2 (p < 0.05) in late March . The marked seasonal decline of omasal tissue and contents in Norwegian reindeer probably results from intake of highly digestible forage plants, including lichens, in winter . Svalbard reindeer, a non-migratory sub-species, survive eating poor quality fibrous vascular plants in winter . The absence of any marked seasonal change in the mass, total absorptive surface area or filling of the omasum in Svalbard reindeer in winter despite a substantial decline in body mass presumably reflects their need to maintain maximum absorption of nutrients, including volatile fatty acids, when feeding on such poorly fermentable forage.

Rapid Commun Mass Spectrom, 2000, 14(15), 1417 - 24
Pyrolysis/gas chromatography/mass spectrometry monitoring of fungal-biotreated distillery wastewater using Trametes sp . I-62 (CECT 20197); Gonzalez T et al.; Distillery wastewaters generated by ethanol production from fermentation of sugar-cane molasses, named vinasses, lead to important ecological impact due to their high content of soluble organic matter and their intense dark-brown color . Taking advantage of the well-known ability of white-rot fungi to degrade an extensive variety of organic pollutants, the capacity of Trametes sp . I-62 (CECT 20197) to detoxify this type of effluents was evaluated . In this work, pyrolysis/gas chromatography/mass spectrometry was applied to the chemical characterization of several fractions of Cuban distillery wastewater as well as to monitoring the changes which occurred after fungal treatment with this white-rot basidiomycete . Maximum effluent decolorization values and chemical oxygen demand reduction attained after seven days of fungal treatment were 73.3 and 61.7%, respectively, when 20% (v/v) of distillery vinasses was added to the culture medium . Under these conditions a 35-fold increase in laccase production by Trametes sp . I-62 was measured, but no manganese peroxidase activity could be detected . The pyrolysis/gas chromatography/mass spectrometry results showed a decrease in a number of pyrolysis products after seven days of fungal treatment, mainly furan derivatives . The decrease in the relative areas of these compounds could be related to the vinasse color-removal associated with melanoidin degradation . All these results indicated the potential use ofTrametes sp . I-62 in the detoxification of recalcitrant distillery vinasses .

Am J Clin Nutr, 2000 Aug, 72(2), 432 - 8
Digestion of so-called resistant starch sources in the human small intestine; Vonk RJ et al.; BACKGROUND: Resistant starch sources, which are only partially digested in the small intestine, can be used to increase colonic availability of short-chain fatty acids . OBJECTIVE: To study the characteristics of the fermentation of resistant starch, the digestion of resistant starch in the small intestine has to be quantified . We compared the metabolic fates of highly digestible cornstarch (DCS), Hylon VII (type 2 resistant starch), and Novelose 330 (type 3 resistant starch), which are of corn origin and, therefore, naturally enriched in (13)C . DESIGN: After administration of 40 g starch or glucose to 7 healthy volunteers, glucose and exogenous glucose concentrations in serum and (13)CO(2) excretion in breath were analyzed for 6 h . (13)C abundance in carbon dioxide was analyzed by isotope ratio mass spectrometry (IRMS) and (13)C abundance in glucose by gas chromatography-combustion IRMS . RESULTS: By comparing the area under the curve (2 h) of exogenous glucose concentration in serum ((13)C glycemic index) after intake of starch or glucose, (13)C glycemic indexes for DCS, Hylon VII, and Novelose 330 were calculated to be 82 +/- 23%, 44 +/- 16%, and 43 +/- 15%, respectively . Comparison of 6-h cumulative percentage dose recovery in breath showed that 119 +/- 28% of DCS, 55 +/- 23% of Hylon VII, and 50 +/- 26% of Novelose 330 was digested in the small intestine . CONCLUSION: The exogenous glucose response in serum and the (13)CO(2) excretion in breath can be used to estimate small intestinal digestion of resistant starch, which amounts to approximately 50%.

J Food Prot, 2000 Jul, 63(7), 934 - 9
Effects of gamma radiation on sensory qualities, microbiological and chemical properties of salted and fermented squid; Byun MW et al.; The effects of gamma radiation on sensory quality, microbial population, and chemical properties of salted and fermented squid were investigated . Squid (Todarodes pacificus) was sliced, washed, and then salted with 5, 10, and 20% (wt/wt) sodium chloride . Salted squid was irradiated with dosages of 0, 2.5, 5.0, and 10 kGy of gamma radiation and fermented at 15 degrees C for 50 days . Proximate composition, salinity, water activity, sensory evaluation, and total microbiological populations were examined . Chemical analyses providing information on degree of fermentation, such as amino nitrogen (AN), volatile basic nitrogen (VBN), trimethylamine (TMA), and hypoxanthine (Hx) were also conducted . Irradiated squid was not different in proximate composition, salinity, and water activity from nonirradiated squid . Sensory evaluation scores, total bacteria populations, and pH values were variable depending on salt concentration and irradiation dose . During fermentation, AN, VBN, TMA, and Hx contents increased rapidly as the salt concentration and irradiation dose decreased . Specifically, these chemical compounds of salted and fermented squid prepared with 10% salt and 10 kGy of gamma radiation maintained the appropriate level of fermentation . The present results showed that the combination of low salt concentration (10%) and gamma radiation was effective in processing salted and fermented squid and extending its shelf life compared to control (20% of salt) without adding any food additives.

FEMS Microbiol Lett, 2000 Aug 1, 189(1), 81 - 7
Direct profiling of the yeast dynamics in wine fermentations; Cocolin L et al.; We present a method to directly characterize the yeast diversity present in wine fermentations by employing denaturing gradient gel electrophoresis (DGGE) of polymerase chain reaction (PCR)-amplified 26S ribosomal RNA (rRNA) genes . PCR-DGGE of a portion of the 26S rRNA gene was shown to distinguish most yeast genera associated with the production of wine . With this method the microbial dynamics in several model wine fermentations were profiled . PCR-DGGE provided a qualitative assessment of the yeast diversity in these fermentations accurately identifying populations as low as 1000 cells ml(-1) . PCR-DGGE represents an attractive alternative to traditional plating schemes for analysis of the microbial successions inherent in the fermentation of wine.

FEMS Microbiol Lett, 2000 Aug 1, 189(1), 9 - 13
Cellular stress responses oscillate in synchronization with the ultradian oscillation of energy metabolism in the yeast Saccharomyces cerevisiae; Wang J et al.; We investigated whether cellular responses to various stress conditions are regulated in synchronization with the ultradian rhythm of respiratory-fermentative metabolism which is coupled to the cell cycle rhythm in continuous cultures of the yeast Saccharomyces cerevisiae . The cellular resistance to heat oscillated with a peak at the late respiro-fermentative phase, which approximately corresponds to the unbudding period of the cell cycle . Cellular resistance to H(2)O(2) and that to the superoxide-generating agent menadione oscillated in the same phase as that of heat resistance . The resistance to cadmium and that to 1-chloro-2,4-dinitrobenzene, an uncoupler of energy metabolism in mitochondria, both oscillated with a peak advanced by about 80 degrees relative to that of heat resistance, approximately covering the respiro-fermentative phase . Thus, cellular resistance to various stresses in S . cerevisiae oscillated in synchronization with the metabolic oscillation in the continuous culture.

FEBS Lett, 2000 Jul 7, 476(3), 134 - 9
Mitochondrial Isa2p plays a crucial role in the maturation of cellular iron-sulfur proteins; Pelzer W et al.; The assembly of iron-sulfur (Fe/S) clusters in a living cell is mediated by a complex machinery which, in eukaryotes, is localised within mitochondria . Here, we report on a new component of this machinery, the protein Isa2p of the yeast Saccharomyces cerevisiae . The protein shares sequence similarity with yeast Isa1p and the bacterial IscA proteins which recently have been shown to perform a function in Fe/S cluster biosynthesis . Like the Isa1p homologue, Isa2p is localised in the mitochondrial matrix as a soluble protein . Deletion of the ISA2 gene results in the loss of mitochondrial DNA and a strong growth defect . Simultaneous deletion of the ISA1 gene does not further exacerbate this growth phenotype suggesting that the Isa proteins perform a non-essential function . When Isa2p was depleted by regulated gene expression, mtDNA was maintained, but cells grew slowly on non-fermentable carbon sources . The maturation of both mitochondrial and cytosolic Fe/S proteins was strongly impaired in the absence of Isa2p . Thus, Isa2p is a new member of the Fe/S cluster biosynthesis machinery of the mitochondrial matrix and may be involved in the binding of an intermediate of Fe/S cluster assembly.

J Bacteriol, 2000 Aug, 182(16), 4632 - 6
Demonstration of a novel glycolytic pathway in the hyperthermophilic archaeon Thermococcus zilligii by (13)C-labeling experiments and nuclear magnetic resonance analysis; Xavier KB et al.; The operation of a novel glycolytic pathway was demonstrated in nongrowing cells of Thermococcus zilligii by analysis of the isotopic enrichment in the end products derived from fermentation of (13)C-labeled glucose . The new pathway involved the formation of formate, derived from C-1 in glucose, via cleavage of a six-carbon carboxylic acid.

Carcinogenesis, 2000 Aug, 21(8), 1513 - 9
Morphodensitometric analysis of protein kinase C beta(II) expression in rat colon: modulation by diet and relation to in situ cell proliferation and apoptosis; Davidson LA et al.; We have recently demonstrated that overexpression of PKC beta(II) renders transgenic mice more susceptible to carcinogen-induced colonic hyperproliferation and aberrant crypt foci formation . In order to further investigate the ability of PKC beta(II) to modulate colonocyte cytokinetics, we determined the localization of PKC beta(II) with respect to cell proliferation and apoptosis along the entire colonic crypt axis following carcinogen and diet manipulation . Rats were provided diets containing either corn oil {containing n-6 polyunsaturated fatty acids (PUFA)} or fish oil (containing n-3 PUFA), cellulose (non-fermentable fiber) or pectin (fermentable fiber) and injected with azoxymethane (AOM) or saline . After 16 weeks, an intermediate time point when no macroscopic tumors are detected, colonic sections were utilized for immunohistochemical image analysis and immunoblotting . Cell proliferation was measured by incorporation of bromodeoxyuridine into DNA and apoptosis by terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling . In the distal colon, PKC beta(II) staining was localized to the upper portion of the crypt . In comparison, proximal crypts had more (P < 0.05) staining in the lower tertile . AOM enhanced (P < 0.05) PKC beta(II) expression in all regions of the distal colonic crypt (upper, middle and lower tertiles) . There was also an interaction (P < 0.05) between dietary fat and fiber on PKC beta(II) expression (corn/pectin > fish/cellulose, fish/pectin > corn/cellulose) in all regions of the distal colonic crypt . With respect to colonic cell kinetics, proliferation paralleled the increase in PKC beta(II) expression in carcinogen-treated animals . In contrast, apoptosis at the lumenal surface was inversely proportional to PKC beta(II) expression in the upper tertile . These results suggest that an elevation in PKC beta(II) expression along the crypt axis in the distal colon is linked to enhancement of cell proliferation and suppression of apoptosis, predictive intermediate biomarkers of tumor development . Therefore, select dietary factors may confer protection against colon carcinogenesis in part by blocking carcinogen-induced PKC beta(II) expression.

Bioelectrochemistry, 2000 Jun, 51(2), 151 - 6
Redox potential is a determinant in the Escherichia coli anaerobic fermentative growth and survival: effects of impermeable oxidant; Bagramyan K et al.; Decrease of redox potential (Eh) down to -550-600 mV in the Escherichia coli culture is observed during growth in either anaerobic or aerobic conditions . The E . coli growth and survival under anaerobic fermentative conditions were found to be strongly inhibited by potassium ferricyanide in the concentration of 1 mM, when Eh was decreased to -50-100 mV . This oxidant also resulted in approximately 2-fold decrease of total and N,N'-dicyclohexylcarbodiimide (DCCD)-inhibited H+ efflux, 2.5-fold inhibition in K+ influx, 1.5-fold less K+ accumulation, and delayed a decrease in Eh to negative values by bacteria . K3{Fe(CN)6} was shown to block an ATP-dependent increase in the amount of accessible thiol groups of membrane vesicles that was inhibited by DCCD, and this inhibition by the oxidant could be recovered by dithiothreitol . These effects were not observed with cells growing under aerobic conditions . The effects of K3{Fe(CN)6}, an impermeable oxidant, might be explained by the fact that redox potential is a determinant in the E . coli anaerobic fermentative growth and survival that has a regulatory role in maintaining H+ and K fluxes and the number of accessible thiol groups on membrane.

J Biotechnol, 2000 Jun 23, 80(2), 135 - 42
Improved secondary metabolite production in the genus Streptosporangium by optimization of the fermentation conditions; Pfefferle C et al.; The cultivation of strains of the genus Streptosporangium in batch fermentations demonstrated that the optimal conditions for secondary metabolite production are completely different to those of the closely related genus Streptomyces . The dissolved oxygen tension (pO(2)) was identified as an important parameter for optimal production of secondary metabolites in submerged cultures . Extreme variations of this parameter by changes in aeration (gas flow), agitation system and stirrer speed showed a tremendous impact in production yields of all investigated strains . Finally, a 20-fold increase in productivity was observed by conditions of controlled oxygen excess compared to optimal fermentation conditions for Streptomyces strains.

FEBS Lett, 2000 Jul 21, 477(3), 219 - 23
Trichodion, a new inhibitor of inflammatory signal transduction pathways from a Trichosporiella species; Erkel G; In a search for new inhibitors of the IFN-gamma mediated signal transduction in HeLa S3 cells using secreted alkaline phosphatase (SEAP) as reporter gene, the novel pyran-dione trichodion was isolated from fermentations of the imperfect fungus Trichosporiella sp . 20-95 . The compound inhibits the IFN-gamma mediated expression of the reporter gene with IC(50) values of 21-42 microM (5-10 microgram/ml) . The NF-kappaB and AP-1 mediated expression of the reporter gene are inhibited with IC(50) values of 42-84 microM (10-20 microgram/ml) and 21 microM (5 microgram/ml) respectively . Western blotting with COX-2 and NOS II antibodies showed that the expression of both proinflammatory enzymes is almost completely inhibited at 21-42 microM (5-10 microgram/ml) in LPS/IFN-gamma stimulated J774 mouse macrophages . Studies on the mode of action of the compound revealed that the inhibition of the NF-kappaB dependent pathway is due to the stabilization of the IkappaB protein and the inhibition of the IFN-gamma dependent signaling is caused by an inhibition of the phosphorylation of the STAT1alpha transcription factor.

J Antibiot (Tokyo), 2000 May, 53(5), 496 - 501
Identification of inhibitors of inducible nitric oxide synthase from microbial extracts; Alvi KA et al.; A new member of the angucycline family, vineomycin C (3), together with four known metabolites saquayamycin A1 (1), A-7884 (2), rabelomycin (5) and xanthomegnin (6) were isolated from microbial extracts . The structures were determined by 1D and 2D NMR techniques and chemical degradation . Compounds 1-3 and 5 were isolated from a fermentation of Streptomyces sp., while 6 was isolated from a fungal fermentation extract . All five compounds have shown potent inhibitory activity in the inducible nitric oxide synthase (iNOS) assay.

J Antibiot (Tokyo), 2000 May, 53(5), 479 - 83
IB-96212, a novel cytotoxic macrolide produced by a marine Micromonospora . II . Physico-chemical properties and structure determination; Canedo LM et al.; IB-96212, is a new member of spiroketal containing macrolide class of fermentation-derived natural products isolated from mycelial extracts of Micromonospora sp . The structure consists of a new aglycone which possesses a 26-membered macrolide ring system and of one deoxy sugar identified as L-rhodinose, this structure represents the first reported spiroketal macrolide natural product related to other macrolides, such as oligomycins, dunaimycins, citovaricin, rutamycin and ossamycin.

J Antibiot (Tokyo), 2000 May, 53(5), 474 - 8
IB-96212, a novel cytotoxic macrolide produced by a marine Micromonospora . I . Taxonomy, fermentation, isolation and biological activities; Fernandez-Chimeno RI et al.; A novel bioactive macrolide, IB-96212 has been isolated from the fermentation broth of a marine actinomycete, L-25-ES25-008 . The strain belongs to the genus Micromonospora . The macrolide showed a very strong cytotoxic activity against P-388, and lower but significant activity against A-549, HT-29, and MEL-28 cell lines . We describe the isolation, taxonomy and fermentation of the producing strain as well as the isolation of IB-96212.

J Dairy Sci, 2000 Jul, 83(7), 1598 - 624
Effects of diet on short-term regulation of feed intake by lactating dairy cattle; Allen MS; Physical and chemical characteristics of dietary ingredients and their interactions can have a large effect on dry matter intake (DMI) of lactating cows . Physical limitations caused by distension of the reticulo-rumen or other compartments of the gastrointestinal tract often limit DMI of high producing cows or cows fed high forage diets . Fermentation acids also limit DMI from a combination of increased osmolality in the reticulo-rumen and specific effects of propionate, although the mechanisms are not clear . The specific physical and chemical characteristics of diets that can affect DMI include fiber content, ease of hydrolysis of starch and fiber, particle size, particle fragility, silage fermentation products, concentration and characteristics of fat, and the amount and ruminal degradation of protein . Site of starch digestion affects the form of metabolic fuel absorbed, which can affect DMI because absorbed propionate appears to be more hypophagic than lactate or absorbed glucose . Dry matter intake is likely determined by integration of signals in brain satiety centers . Difficulty in measurement and extensive interactions among the variables make it challenging to account for dietary effects when predicting DMI . However, a greater understanding of the mechanisms along with evaluation of animal responses to diet changes allows diet adjustments to be made to optimize DMI as well as to optimize allocation of diet ingredients to animals . This paper discusses some of the characteristics of dietary ingredients that should be considered when formulating diets for lactating dairy cows and when allocating feeds to different groups of animals on the farm.

J Dairy Sci, 2000 Jul, 83(7), 1520 - 9
Milk production and composition, rumen fermentation parameters, and grazing behavior of dairy cows supplemented with different forms and amounts of corn grain; Soriano FD et al.; The objectives were to compare milk production and composition, change in body weight and body condition score, rumen fermentation parameters and grazing patterns by cows when supplemented with different forms and amounts of corn grain . In experiment 1, 36 Holstein cows were supplemented with either 6, 6, 6, or 4 kg/d of dry matter of high moisture corn, coarsely ground corn, finely ground corn, or high moisture corn in two equal daily feedings, respectively . Milk yield (30.3 kg/d), milk protein (2.97%), and milk urea N (14.7 mg/dl) were not different among treatments . Body weight change and body condition score change were similar (23.1 kg and -0.24) for the 10-wk study . During experiment 2, four rumen cannulated cows in midlactation were supplemented with 6 kg/d of dry matter from either coarsely ground corn or high moisture corn in two equal feedings after milking . After the p.m . milking, ruminal pH was measured and rumen fluid samples, were collected from cows to determine ammonia N and volatile fatty acids at 0.5, 1, 2, 3, ...8 h post-corn feeding during grazing . Ruminal pH values were similar for corn supplements, and, with one exception, were 6.0 or below between 5 and 8 h . Ruminal ammonia-N concentrations reached a maximum at 7 h also . In experiment 3, 40 cows were observed for grazing behavior every 30 min for two consecutive days . Cows grazed an average of 6.4 h/d, 4.1 h in the afternoon and 2.3 h in the morning . Milk yield, milk composition, change in body weight, and body condition were similar regardless of the type or amount of corn supplemented.

Nahrung, 2000 Jun, 44(3), 207 - 10
Biosynthesis of nutraceutical iso-oligosaccharides by multiple forms of transferase produced by Aspergillus foetidus; Wang XD et al.; Isomalto-oligosaccharide and isofructo-oligosaccharide production was attempted using enzymes produced by Aspergillus foetidus . Four fractions having transferase enzyme activity were obtained from the fermentation broth of Aspergillus foetidus by ammonium sulfate precipitation and DEAE-cellulose chromatography . The optimum temperature was 60 degrees C and pH stability in the range 4 to 6 for various fractions . The pH optima, heat sensitivity and kinetic parameters for the four fractions were however not the same . All four enzyme fractions could not utilize lactose and cellobiose to synthesize isooligosaccharide and showed different transferase activity for maltose and sucrose for synthesis of isooligosaccharides . The HPLC isooligosaccharide product analysis of these transferase enzymes reveal that the four forms of enzymes are distinct and produce oligosaccharides like panose, kestose and nystose or act as hydrolytic enzymes, depending on reaction conditions.

Anal Biochem, 2000 Aug 1, 283(2), 192 - 9
Determination of carbohydrates, sugar alcohols, and glycols in cell cultures and fermentation broths using high-performance anion-exchange chromatography with pulsed amperometric detection; Hanko VP et al.; Cell cultures and fermentation broths are complex mixtures of organic and inorganic compounds . Many of these compounds are synthesized or metabolized by microorganisms, and their concentrations can impact the yields of desired products . Carbohydrates serve as carbon sources for many microorganisms, while sugar alcohols (alditols), glycols (glycerol), and alcohols (methanol and ethanol) are metabolic products . We used high-performance anion-exchange chromatography with pulsed amperometric detection (HPAE-PAD) to simultaneously analyze for carbohydrates, alditols, and glycerol in growing yeast (Saccharomyces cerevisiae) cultures and their final fermentation broths . Both cultures were grown on complex undefined media, aliquots centrifuged to remove particulates, and the supernatants diluted and directly injected for analysis . Pulsed amperometry allowed a direct detection of the carbohydrates, alditols, and glycols present in the cultures and fermentation broths with very little interference from other matrix components . The broad linear range of three to four orders of magnitude allowed samples to be analyzed without multiple dilutions . Peak area RSDs were 2-7% for 2, 3-butanediol, ethanol, glycerol, erythritol, rhamnose, arabitol, sorbitol, galactitol, mannitol, arabinose, glucose, galactose, lactose, ribose, raffinose, and maltose spiked into a heat-inactivated yeast culture broth supernatant that was analyzed repetitively for 48 h . This method is useful for directly monitoring culture changes during fermentation . The carbohydrates in yeast cultures were monitored over 1 day . A yeast culture with medium consisting primarily of glucose and trace levels of trehalose and arabinose showed a drop in sugar concentration over time and an increase in glycerol . Yeast growing on a modified culture medium consisting of multiple carbohydrates and alditols showed preference for specific carbon sources and showed the ability to regulate pathways leading to catalysis of alternative carbon sources .

J Chromatogr A, 2000 Jun 9, 881(1-2), 517 - 30
Liquid chromatographic determination of biogenic amines in fermented foods after derivatization with 3,5-dinitrobenzoyl chloride; Kirschbaum J et al.; The reagent 3,5-dinitrobenzoyl chloride (DNBZ-Cl) was tested for pre-column derivatization of biogenic amines (BAs) . Samples were derivatized within 3 min in 1 M NaOH at ambient temperature by adding 2-propanole and 50 mM DNBZ-Cl in acetonitrile . The reaction was terminated by addition of 2 M HCl . For high-performance liquid chromatography an encapsulated stationary reversed-phase and gradient elution using a ternary gradient system were used . The DNBZ derivatives were quantified by their UV-absorption at 260 nm . The structures of the derivatives were elucidated using coupling of HPLC with electrospray ionization mass spectrometry . Detection limits of BAs were approximately 124-864 microg l(-1) (injected amounts 203-1410 pg) at a signal-to-noise ratio of 3:1 . The coefficients of determination were 0.989-0.996, with the exceptions of cadaverine (0.976) and serotonin (0.965) . The method was applied to the quantitative determination of agmatine, cadaverine, histamine, octopamine, 2-phenylethylamine, putrescine, serotonin, spermidine, spermine, tryptamine and tyramine, in fermented cabbage juices, soy sauces, Misos (soy pastes), fermented fish sauces, and anchovy paste.

J Chromatogr A, 2000 Jun 9, 881(1-2), 285 - 97
Chromatographic determination of riboflavin and its derivatives in food; Gliszczynska-Swiglo A et al.; Three elution methods on two different reversed-phase C18 columns were developed to determine flavin derivatives in raw egg white, raw egg yolk, egg powder, pasteurised milk, fermented milk products and liver (chicken, calf and pig) . Additionally, 11 thin-layer chromatography solvent systems were used to confirm presence of flavins detected in assessed products . It was found that an Alphabond C18 column was not as effective as a Symmetry C18 column . Method A (mobile phase gradient of methanol-0.05 M ammonium acetate, pH 6.0 applied on an Alphabond C18 column) can be used for determination of flavin adenine dinucleotide, flavin mononucleotide, riboflavin 4',5'-cyclic phosphate, riboflavin, 10-formylmethylflavin and 10-hydroxyethylflavin in products that do not contain 7alpha-hydroxyriboflavin . Method B (mobile phase gradient of methanol-demineralized water, on an Alphabond C18 column) can be useful to separate flavin coenzymes from other flavin compounds or to confirm the presence of 7alpha-hydroxyriboflavin and 10-hydroxyethylflavin in analysed samples . Method C (mobile phase gradient of methanol-0.05 M ammonium acetate, pH 6.0, on a Symmetry C18 column) allows separation of all flavins detected in tested products: flavin adenine dinucleotide, flavin mononucleotide, riboflavin 4',5'-cyclic phosphate, riboflavin, 10-formylmethylflavin, 10-hydroxyethylflavin, 7alpha-hydroxyriboflavin, riboflavin-beta-D-galactoside and riboflavin-alpha-D-glucoside.

J Chromatogr A, 2000 Jun 9, 881(1-2), 81 - 91
Chromatographic determination of amino acid enantiomers in beers and raw materials used for their manufacture; Erbe T et al.; Using gas chromatography (GC) on a chiral stationary phase, accompanied by high-performance liquid chromatography, beers and raw materials used for manufacturing (hops, barley grains, malts) were investigated for the pattern and quantities of amino acid enantiomers . Although L-amino acids were most abundant, certain D-amino acids were detected in all beers and most of the raw materials . Highest amounts of D-amino acids were detected in special beers such as Berliner Weisse that underwent bottle-conditioning with lactic cultures, and Belgian fruit beers produced by spontaneous fermentation . It is demonstrated that GC on chiral stationary phases is highly suitable for the quantitative determination of amino acid enantiomers in beers and raw materials used for their manufacture . Quantities, relative amounts and pattern of amino acid enantiomers can serve in particular as chiral markers for the authenticity of special beers.

J Clin Invest, 2000 Jul, 106(2), 281 - 7
Congenital sucrase-isomaltase deficiency arising from cleavage and secretion of a mutant form of the enzyme; Jacob R et al.; Congenital sucrase-isomaltase deficiency (CSID) is an autosomal recessive human intestinal disorder that is clinically characterized by fermentative diarrhea, abdominal pain, and cramps upon ingestion of sugar . The symptoms are the consequence of absent or drastically reduced enzymatic activities of sucrase and isomaltase, the components of the intestinal integral membrane glycoprotein sucrase-isomaltase (SI) . Several known phenotypes of CSID result from an altered posttranslational processing of SI . We describe here a novel CSID phenotype, in which pro-SI undergoes an unusual intracellular cleavage that eliminates its transmembrane domain . Biosynthesis of pro-SI in intestinal explants and in cells transfected with the SI cDNA of this phenotype demonstrated a cleavage occurring within the endoplasmic reticulum due to a point mutation that converts a leucine to proline at residue 340 of isomaltase . Cleaved pro-SI is transported to and processed in the Golgi apparatus and is ultimately secreted into the exterior milieu as an active enzyme . To our knowledge this is the first report of a disorder whose pathogenesis results not from protein malfolding or mistargeting, but from the conversion of an integral membrane glycoprotein into a secreted species that is lost from the cell surface.

Australas Radiol, 1999 Feb, 43(1), 37 - 40
The role of computed tomography in the diagnosis of arterial gas embolism in fatal diving accidents in Tasmania; Oliver J et al.; Four cases of fatal diving accidents in Tasmania are presented, highlighting the role of CT in the investigation of diving fatalities . The CT technique allows rapid diagnosis when arterial gas embolism (AGE) is suspected . The traditional method of investigation, underwater autopsy, is a difficult procedure that requires specialized training in which the subtle diagnosis of AGE may be completely missed . Facilities for performing underwater autopsies are normally available only in tertiary referral centres, and therefore the diagnosis of AGE may be missed due to lack of facilities . The use of CT in the diagnosis of AGE in divers was first utilized in the early 1980s but has still not become widely adopted in forensic practice . This radiological technique has the advantage of being sensitive, quick, reliable, readily available and provides a permanent record . For hospitals that do not have a resident forensic pathologist, a CT scan can be easily performed and interpreted to eliminate the possibility of AGE . There are a number of pitfalls in the diagnosis of AGE with CT, particularly intravascular gas production following postmortem fermentation and off-gassing . Awareness of these pitfalls will help the radiologist in making a correct diagnosis of AGE.

J Chromatogr Sci, 2000 Jul, 38(7), 307 - 14
Determination of major aroma impact compounds in fermented cucumbers by solid-phase microextraction--gas chromatography--mass spectrometry--olfactometry detection; Marsili RT et al.; Purge-and-trap, solid-phase extraction, and solid-phase microextraction-gas chromatography-mass spectrometry (SPME-GC-MS) sample preparation techniques for the analysis of odor impact chemicals in fermented cucumber brine are compared . SPME-GC-MS is coupled with detection frequency olfactometry experiments to determine key impact odor compounds in the brine . The most potent odorants that define the typical characteristic brine aroma are trans-4-hexenoic acid and cis-4-hexenoic acid . Confirmation of key impact odorants in brine is confirmed by recombination experiments.

Trends Biotechnol, 2000 Aug, 18(8), 356 - 60
Defined media and inert supports: their potential as solid-state fermentation production systems; Ooijkaas LP et al.; Solid-state fermentation (SSF) using inert supports impregnated with chemically defined liquid media has several potential applications in both scientific studies and in the industrial production of high-value products, such as metabolites, biological control agents and enzymes . As a result of its more defined system, SSF on inert supports offers numerous advantages, such as improved process control and monitoring, and enhanced process consistency, compared with cultivation on natural solid substrates.

Biochim Biophys Acta, 2000 Jul 14, 1480(1-2), 171 - 81
A non-specific aminopeptidase from Aspergillus; Blinkovsky AM et al.; A fermentation broth supernatant of the Aspergillus oryzae strain ATCC20386 contains aminopeptidase activity that releases a wide variety of amino acids from natural peptides . The supernatant was fractionated by anion exchange chromatography . Based on the primary amino acid sequence data obtained from proteins in certain fractions, polymerase chain reaction (PCR) primers were made and a PCR product was generated . This PCR product was used to screen an A . oryzae cDNA library from which the full length gene was then obtained . Fusarium venenatum and A . oryzae were used as hosts for gene expression . Transformed strains of both F . venenatum and A . oryzae over-expressed an active aminopeptidase (E.C . 3.4.11), named aminopeptidase II . The recombinant enzyme from both fungal hosts appeared as smears on sodium dodecyl sulfate-polyacrylamide gel electrophoresis . After deglycosylation of the N-linked sugars, both samples were a sharp band at approximately 56 kDa and had identical N-terminal amino acid sequences . Aminopeptidase II is a metalloenzyme with, presumably, Zn in the active site . Using various natural peptides and para-nitroanilides (pNAs) of amino acids as substrates, the aminopeptidase was found to be non-specific . Only X-Pro bonds demonstrated resistance to hydrolysis catalyzed by this aminopeptidase . The optimal enzyme activity was observed at pH 9.5 and 55 degrees C . Among amino acid pNAs, Leu-pNA appears to have the highest value of bimolecular constant of 40 min(-1) mM(-1) (k(cat) = 230 min(-1); K(m) = 5.8 mM) at pH 7.5 and 21 degrees C . Among Xaa-Ala-Pro-Tyr-Lys-amide pentapeptides, the velocity of catalytic hydrolysis at pH 7.5 and 21 degrees C was in a decreasing order: Pro, Ala, Leu, Gly and Glu.

Enzyme Microb Technol, 2000 Aug 1, 27(3-5), 337 - 344
A mathematical model of the formation of fermentable sugars from starch hydrolysis during high-temperature mashing; Muller R; During the mashing process of brewing, activity of the amylolytic enzymes decays due to the high temperatures used to gelatinise the starch . Because the different enzymes produce different sugars, high temperatures can be exploited to modify the fermentability of resulting worts . This is especially useful when producing low alcohol beers . The expression a.exp(b.t)-c.exp(d.t) (where t is the temperature of the mash in degrees C) provides a simple but useful description of the activity of the amylases . Combining the activities of alpha- and beta-amylases results in a prediction of the resulting fermentability . A simple modification to the expression accommodates changes in mash thickness . The error of prediction is approximately 3 degrees of fermentability . The model is not appropriate for predicting the fermentability of worts produced at the lower standard mashing temperatures . It can be used without the necessity of analytical parameters so analyses that the brewer would not normally perform are not required . If increased accuracy is needed, the results of two previous mashes can be used to modify the parameters used.

Enzyme Microb Technol, 2000 Aug 1, 27(3-5), 312 - 318
Heterotrophic production of biomass and lutein by Chlorella protothecoides on various nitrogen sources; Shi X et al.; The effects of nitrate, ammonium, and urea as nitrogen sources on the heterotrophic growth of Chlorella protothecoides were investigated using flask cultures . No appreciable inhibitory effect on the algal growth was observed over a nitrogen concentration range of 0.85-1.7 g l(-)(1) . In contrast, differences in specific growth rate and biomass production were found among the cultures with the various nitrogen compounds . The influence of different nitrogen sources at a concentration equivalent to 1.7 g l(-)(1) nitrogen on the heterotrophic production of biomass and lutein by C . protothecoides was investigated using the culture medium containing 40 g l(-)(1) glucose as the sole carbon and energy source in fermentors . The maximum biomass concentrations in the three cultures with nitrate, ammonium, and urea were 18.4, 18.9, and 19.6 g l(-)(1) dry cells, respectively . The maximum lutein yields in these cultures were between 68.42 and 83.81 mg l(-)(1) . The highest yields of both biomass and lutein were achieved in the culture with urea . It was therefore concluded that urea was the best nitrogen source for the production of biomass and lutein . Based on the experimental results, a group of kinetic models describing cell growth, lutein production, and glucose and nitrogen consumption were proposed and a satisfactory fit was found between the experimental results and predicted values . Dynamic analysis of models demonstrated that enhancing initial nitrogen concentration in fermentor cultures, which correspondingly enhances cell growth and lutein formation, may shorten the fermentation cycle by 25-46%.

Biotechnol Bioeng, 2000 Sep 5, 69(5), 537 - 47
Effect of carbon and nitrogen sources on growth dynamics and exopolysaccharide production for the hyperthermophilic archaeon Thermococcus litoralis and bacterium Thermotoga maritima; Rinker KD et al.; Batch and continuous cultures were used to compare specific physiological features of the hyperthermophilic archaeon, Thermococcus litoralis (T(opt) of 85 degrees to 88 degrees C), to another fermentative hyperthermophile that reduces S degrees facultatively, that is, the bacterium Thermotoga maritima (T(opt) of 80 degrees to 85 degrees C) . Under nutritionally optimal conditions, these two hyperthermophiles had similar growth yields on maltose and similar cell formula weights based on elemental analysis: CH(1.7)O(0 . 7)N(0.2)S(0.006) for T . litoralis and CH(1.6)O(0.6)N(0.2)S(0.005) for T . maritima . However, they differed with respect to nitrogen source, fermentation product patterns, and propensity to form exopolysaccharides (EPS) . T . litoralis could be cultured in the absence or presence of maltose on an amino acid-containing defined medium in which amino acids served as the sole nitrogen source . T . maritima, on the other hand, did not utilize amino acids as carbon, energy, or nitrogen sources, and could be grown in a similar defined medium only when supplemented with maltose and ammonium chloride . Not only was T . litoralis unable to utilize NH(4)Cl as a nitrogen source, its growth was inhibited at certain levels . At 1 g/L ( approximately 20 mM) NH(4)Cl, the maximum growth yield (Y(x/s(max))) for T . litoralis was reduced to 13 g cells dry weight (CDW)/mol glucose from 40 g CDW/mol glucose in media lacking NH(4)Cl . Alanine production increased with increasing NH(4)Cl concentrations and was most pronounced if growth on NH(4)Cl was carried out in an 80% H(2) atmosphere . In T . maritima cultures, which would not grow in an 80% H(2) atmosphere, alanine and EPS were produced at much lower levels, which did not change with NH(4)Cl concentration . EPS production rose sharply at high dilution rates for both organisms, such that maltose utilization plots were biphasic . Wall growth effects were also noted, because cultures failed to wash out at dilution rates significantly above maximum growth rates determined from batch growth experiments . This study illustrates the importance of effective cultivation methods for addressing physiological issues related to the growth of hyperthermophilic heterotrophs .

Biotechnol Bioeng, 2000 Sep 5, 69(5), 526 - 36
Effects of Ca(OH)(2) treatments ("overliming") on the composition and toxicity of bagasse hemicellulose hydrolysates; Martinez A et al.; Hemicellulose syrups from dilute sulfuric acid hydrolysates of hemicellulose contain inhibitors that prevent efficient fermentation by yeast or bacteria . It is well known that the toxicity of these hydrolysate syrups can be ameliorated by optimized "overliming" with Ca(OH)(2) . We have investigated the optimization of overliming treatments for sugar cane bagasse hydrolysates (primarily pentose sugars) using recombinant Escherichia coli LY01 as the biocatalyst . A comparison of composition before and after optimal overliming revealed a substantial reduction in furfural, hydroxymethylfurfural, and three unidentified high-performance liquid chromatography (HPLC) peaks . Organic acids (acetic, formic, levulinic) were not affected . Similar changes have been reported after overliming of spruce hemicellulose hydrolysates (Larsson et al., 1999) . Our studies further demonstrated that the extent of furan reduction correlated with increasing fermentability . However, furan reduction was not the sole cause for reduced toxicity . After optimal overliming, bagasse hydrolysate was rapidly and efficiently fermented (>90% yield) by LY01 . During these studies, titration, and conductivity were found to be in excellent agreement as methods to estimate sulfuric acid content . Titration was also found to provide an estimate of total organic acids in hydrolysate, which agreed well with the sum of acetic, levulinic, and formic acids obtained by HPLC . Titration of acids, measurement of pH before and after treatment, and furan analyses are proposed as relatively simple methods to monitor the reproducibility of hydrolysate preparations and the effectiveness of overliming treatments .

Biotechnol Bioeng, 2000 Sep 5, 69(5), 495 - 503
Physiological state-specific models in estimation of recombinant Escherichia coli fermentation performance; Feng M et al.; Implementation of advanced control strategies in bioprocesses is often hindered by the lack of on-line measurements reflecting the physiological state of the culture . Although a number of techniques have been used to estimate key variables from data monitored on-line, these often do not explicitly take into account changes in physiological state and information on many aspects of physiological state that may not be present in on-line data . Here we demonstrate that data obtained from chemical fingerprinting methods, such as pyrolysis mass spectrometry, can be used to identify changes in the physiological state during cultivation . This information can be utilized for the estimation of the physiological state and can enable physiological state-specific-model development for on-line bioprocess control .

Arch Microbiol, 2000 May-Jun, 173(5-6), 398 - 402
CO2-dependent fermentation of phenol to acetate, butyrate and benzoate by an anaerobic, pasteurised culture; Karlsson A et al.; Fermentative degradation of phenol was studied using a non-methanogenic, pasteurised enrichment culture containing two morphologically different bacteria . Phenol was fermented to benzoate, acetate and butyrate and their relative occurrence depended on the concentration of hydrogen . Proportionately more benzoate was formed with high initial levels of H2 . The influence of PH2 on the fermentation pattern was studied both in dense cell suspensions and in growing cultures by addition of hydrogen . An increase in growth yield (OD578) was observed, compared to controls, as a consequence of phenol degradation; however, the increase was less in H2-amended treatments, in which most of the phenol ended up as benzoate . The degradation of phenol in the dense cell suspension experiments was dependent on CO2 . Benzoate was not degraded when added as a substrate to the growing culture . This is, to our knowledge, the first report concerning the fermentative degradation of phenol to nonaromatic products.

Physiol Biochem Zool, 2000 May-Jun, 73(3), 264 - 70
Metabolic adaptations of overwintering European common lizards (Lacerta vivipara); Voituron Y et al.; The European common lizard Lacerta vivipara, a reptile of cold-temperate climates, provides us an interesting model of low-temperature adaptation . Indeed its unique cold-hardiness strategy, which employs both freeze tolerance and freeze avoidance, may be seen as the primary reason for its large distribution, which extends from Spain to beyond the Arctic circle . To study the metabolism supporting this capacity, we used three techniques: two techniques of calorimetry (oxygen consumption and thermogenesis) and nuclear magnetic resonance spectroscopy . These techniques were used to examine the metabolic balance and the different molecular pathways used between three different periods through the year (September, January, and May) . The results show a significant 20% augmentation of winter anaerobic metabolism compared to other periods of the year . This is mainly because of an activation of the lactic fermentation pathway leading to an increase of lactate concentration (>34% in winter) . Furthermore, glucose, which increases some 245% in winter, is used as antifreeze and metabolic substrate . Furthermore, this study provides evidence that the physiological adaptations of the common lizard differ from those of other ectotherms such as Rana sylvatica . Concentrations of alanine and glycerol, commonly used as antifreeze by many overwintering ectotherms, do not increase during winter.

Bioseparation, 2000, 9(2), 81 - 91
The effect of demulsifiers on lysozyme extraction from hen egg white using reverse micelles; Jarudilokkul S et al.; The liquid-liquid extraction of protein from buffered aqueous phases using reverse micelles (RM) has been extensively researched from a fundamental point of view . However, very little effort has been expended at scaling up this process for the extraction of real fermentation broth . When real broths are used with reverse micellar phases there are major problems with emulsion formation . In this study the effect of a variety of demulsifiers on lysozyme extraction was evaluated in terms of their influence on the separating properties of the emulsion, water content (Wo), and, extraction yield and kinetics from both buffer and hen egg white . In addition, the use of a low shear contactor (a Graesser or 'raining bucket') was assessed in terms of its suitability as a RM contactor . It was found that most of the demulsifiers reduced the settling time of the emulsion, and enhanced the yield and kinetics of lysozyme extraction from hen egg white . It was hypothesised that this was due to the demulsifier displacing the lysozyme from the interface and preventing the protein unfolding and precipitating . This effect was found to depend on both the generic type of demulsifier, and its concentration.

J Chromatogr Sci, 2000 Jun, 38(6), 259 - 61
A simple high-performance liquid chromatography method for the analysis of glucose, glycerol, and methanol in a bioprocess; Parpinello GP et al.; A reliable high-performance liquid chromatography-refractive index detection method for the simultaneous analysis of glucose, methanol, and glycerol in a bioprocess fermentation media using direct injection is presented . The validation studies show a satisfactory selectivity, linearity, accuracy, and recovery of the method . The lowest concentration detectable for glucose and methanol is 3.5 and 6.7 mg/100 mL, respectively . This method could be an attractive choice for the analysis of these compounds not only in fermentation media but also in biomedical and environmental samples.

Nutr Cancer, 2000, 36(2), 131 - 49
Are dietary fiber-induced alterations in colonic epithelial cell proliferation predictive of fiber's effect on colon cancer?
Whiteley LO, Klurfeld DM.
Alterations in cell proliferation of the colon have been observed as a result of changes in amount and type of dietary fiber and in relation to risk of developing colon cancer . Although some human observational and intervention studies contribute to the database, most information results from experiments on rodents . Because of numerous contradictory reports linking dietary fiber, cell proliferation, and colon cancer, we undertook a critical review of existing methods in an attempt to explain the inconsistencies . Although there may be some individual types of dietary fiber that protect against chemically induced colon cancer, dietary fiber as a single entity does not appear to afford any consistent protection . Because of significant differences in experimental protocols among laboratories, it is not yet possible to state with certainty that increases in cell proliferation, induced by fiber consumption, are predictive of increased tumorigenesis . Much of what has been observed and interpreted as elevation of risk may simply be normal homeostatic changes in cell proliferation . Even though fermentation to short-chain fatty acids is a mechanistically attractive hypothesis to explain why fiber modulates cytokinetics, data do not consistently support short-chain fatty acids as biological intermediates in risk of colon cancer . The state of the art in this field has not yet progressed to the point where a clear effect of dietary fiber on cytokinetics and colon carcinogenesis can be assessed with any degree of certainty . Additional markers of apoptosis, differentiation, and cell-cell communication may be required for a more accurate analysis of the relation among fiber, cytokinetics, and colon cancer.

J Agric Food Chem, 2000 Jun, 48(6), 2395 - 400
Influence of the simultaneous addition of the protease flavourzyme and the lipase novozym 677BG on dry fermented sausage compounds extracted by SDE and analyzed by GC-MS; Ansorena D et al.; A dry fermented sausage (chorizo de Pamplona) was elaborated with the simultaneous addition of a lipase (Novozym 677BG) and a protease (Flavourzyme) and ripened during 21 days, in contrast to the control without enzymes and ripened during 35 days . Faster and more intense lipolytic and proteolytic activities were observed in the modified sausage, despite its shorter maturation time . At the end of the ripening, a determination of the profile of compounds extracted by simultaneous distillation-extraction with dichloromethane was carried out . The total amounts of extracted compounds (expressed in milligrams of dodecane per gram of dry matter) were 2.5 in the sausage with enzymes and 1.9 in the control . The chemical groups showing increments due to the use of enzymes were esters (103.5% increment) and acids (87% increment) in both cases due to the greater presence of long-chain fatty acid products . However, development of substances originated from further degradation process of amino acids and free fatty acids did not seem to have taken place.

J Agric Food Chem, 2000 Jun, 48(6), 2384 - 6
Evidence of Strecker aldehyde excretion by yeast in cold contact fermentations; Perpete P et al.; A medium containing labeled leucine-d(10) has been used to show that Saccharomyces cerevisiae was able to produce deuterated 3-methylbutanal in a cold contact fermentation . Whereas residual unreduced Strecker aldehydes bound to polyphenols were considered until now as the main defect of alcohol-free beers, yeast emerges from this work as an additional source of worty flavor . When the same experiment was conducted without leucine, 3-methylbutanal excretion also occurred, indicating that valine could be used by yeast as precursor.

J Agric Food Chem, 2000 Jun, 48(6), 2149 - 54
Enzymatic formation of ether linkage producing shoyuflavones from genistein and (+/-)-trans-epoxysuccinic acid; Kinoshita E et al.; The production mechanism of shoyuflavones, conjugated ethers of isoflavones with tartaric acid and isolated from fermented soy sauce, was studied . In the high molecular weight fraction of the culture extract of Aspergillus oryzae, genistein was transformed into shoyuflavone B in the presence of (+/-)-trans-epoxysuccinic acid but not in the low molecular one . Asp . sojae and Asp . tamarii showed high activity similar to Asp . oryzae but none of Asp . niger, Rhizopus oligosporus, and Mucor praini did . The contents of epoxysuccinic acids in the starting materials of soy sauce and the cultures of various Asp . fungi were determined as dimethyl 2-chloro-3-hydroxysuccinate derivatives by GC-MS . Although epoxysuccinic acids were contained in Asp . oryzae, Asp . sojae, and Asp . tamarii cultures, they were not found in soybeans and wheat . A possible producing mechanism for shoyuflavones by enzymatically conjugating isoflavones to (+/-)-trans-epoxysuccinic acid with ether linkage was suggested.

Yi Chuan Xue Bao, 2000, 27(2), 165 - 9
{Alpha-acetolactate decarboxylase from B . licheniformis AS10106: cloning and expression gene in E . coli and S . cerevisiae}; Qin YJ et al.; A genomic library of B . licheniformis AS10106 that contained the alpha-acetolactate decarboxylase gene(alpha-ALDC) was constructed with vector pUC19 and host E . coli JM109 strain . The inserted fragments of foreign DNA ranged from 4 to 10 kb in the 4800 clones thus obtained . Six positive clones were detected after screening the plated library by the method of clony coloration . Subcloning of the DNA fragment containing the alpha-acetolactate decarboxylase gene showed that the alpha-acetolactate decarboxylase gene was on an 1.6 kb BamH I-EcoR I fragment . Preliminary analysis of the enzyme expressed from one recombinant plasmid pGEA showed that the properties of the recombinant enzyme, such as the optimal temperature and pH of reaction, were identical to those of the native enzyme . Using yeast-E . coli shuttle vector pYES2, an expression recombinant plasmid pYEA containing B . licheniformis AS10106 alpha-acetolactate decarboxylase gene was constructed . S . cerevisiae H158 transformed with pYEA had expressed alpha-acetolactate decarboxylase activity and shown the ability to reduce the formation of diacetyl during beer fermentation.

Rev Cubana Med Trop, 1999 Sep-Dec, 51(3), 152 - 5
{Determination of verotoxins in strains of Escherichia coli O157:H7}; Ramirez M et al.; We performed an study to find out the main virulence factor in verotoxigenic Escherichia coli: the production of verotoxins in 50 non sorbitol-fermenting Escherichia coli strains (possible enterohemorrhage) which were isolated from children with acute diarrheas in the City of Havana and referred to the National Reference Laboratory for acute diarrheal diseases in "Pedro Kouri" Institute . By using the agglutination technique with latex particles of E . coli O157:H7, we determined whether the verotoxins belonged to this serotype, we also researched the production of verotoxins in Vero cell culture . Ninety-six percent of the total number of strains were positive in the qualitative determination of this factor which was more frequently observed after 24 hours.

Lett Appl Microbiol, 2000 Jul, 31(1), 73 - 6
Effect of uracil on rifamycin SV production by Amycolatopsis mediterranei MV35R; Murali Krishna PS et al.; The effect of different organic nitrogen compounds on the production of rifamycin SV by Amycolatopsis mediterranei MV35R and their optimum concentrations have been described . Results obtained indicate that rifamycin SV production increased from 4020 mg l-1 to 4575 mg l-1 when organic nitrogen compound uracil was added at 0.2% (w/v) concentration to the fermentation medium by A . mediterranei MV35R . The rifamycin SV yield was enhanced by 505 mg l-1 using uracil (2 g l-1) when compared with barbital.

Lett Appl Microbiol, 2000 Jul, 31(1), 46 - 51
The impact of media composition and petite mutation on the longevity of a polyploid brewing yeast strain; Powell CD et al.; Ageing in Saccharomyces cerevisiae is a finite phenomenon, determined by replicative, rather than chronological lifespan . Yeast physiological condition is known to influence industrial fermentation performance, however, until recently cellular senescence has not been considered as a brewing yeast stress factor . A polyploid lager yeast (BB11) and a brewery isolate, exhibiting petite mutation were analysed for longevity . It was observed that mitochondrial deficiency induced a reduction in lifespan . In addition, replicative capacity was perceived to be dependent on environmental conditions.

Arq Gastroenterol, 1999 Oct-Dec, 36(4), 227 - 37
{Mechanisms of absorption of amino acids and oligopeptides . Control and implications in human diet therapy}; Frenhani PB et al.; The mechanisms involved in the absorption of amino acids and oligopeptides are reviewed regarding their implications in human feedings . Brush border and basolateral membranes are crossed by amino acids and di-tripeptides by passive (facilitated or simple diffusion) or active (Na+ or H+ co-transporters) pathways . Active Na(+)-dependent system occurs mainly at brush border and simple diffusion at basolateral, both membranes have the passive facilitated transport . Free-amino acids use either passive or active transport systems whereas di-tripeptides do mainly active (H+ co-transporter) . Brush border have distinctive transport system for amino acids and di-tripeptides . The former occurs mainly by active Na+ dependently whereas the later is active H(+)-dependent with little affinity for tetra or higher peptides . Free amino acids are transported at different speed by saturable, competitive carriers with specificity for basic, acidic or neutral amino acids . Di and tripeptides have at least two carriers both electrogenic and H(+)-dependent . The basolateral membrane transport of amino acids is mostly by facilitated diffusion while for di-tripeptides it is an active anion exchange associated process . The main regulation of amino acids and di-tripeptide transport is the presence o substrate at the mucosal membrane with higher the substrate higher the absorption . Di and tripeptides are more efficiently absorbed than free amino acids which in turns are better absorbed than oligopeptides . So di-tripeptides result in better N-retention and is particularly useful in cases of lower intestinal absorption capacity . The non-absorbed peptides are digested and fermented by colonic bacteria resulting short-chain fatty acids, dicarboxylic acids, phenolic compounds and ammonia . Short-chain fatty acid provides energy for colonocytes and bacteria and the ammonia not fixed by bacteria returns to the liver for ureagenesis.

Sheng Wu Gong Cheng Xue Bao, 2000 Jan, 16(1), 10 - 2
{The diploid of genetically engineered yeast for expression of hybrid HBsAg gene SA-28}; Chen XK et al.; By mating a yeast haploid expressing hybrid HBsAg gene SA-28, Y19/YFD158, with another hapolid Y95, the diploid Y95xY19/YFD158 was constructed . The experiments showed that the cell density of the diploid in fermentation was three time higher than that of the haploid, the stability of expression vector in the diploid was much higher than in the haploid, the xpression level of hybrid HBsAg gene of the diploid was more than three time higher than that of the haploid and the average cope number in diploid cell was a little lower than that in the haploid.

Eur J Biochem, 2000 Jul, 267(14), 4566 - 76
Reconstitution of ethanolic fermentation in permeabilized spheroplasts of wild-type and trehalose-6-phosphate synthase mutants of the yeast Saccharomyces cerevisiae; Noubhani A et al.; In the yeast Saccharomyces cerevisiae, TPS1-encoded trehalose-6-phosphate synthase (TPS) exerts an essential control on the influx of glucose into glycolysis, presumably by restricting hexokinase activity . Deletion of TPS1 results in severe hyperaccumulation of sugar phosphates and near absence of ethanol formation . To investigate whether trehalose 6-phosphate (Tre6P) is the sole mediator of hexokinase inhibition, we have reconstituted ethanolic fermentation from glucose in permeabilized spheroplasts of the wild-type, tps1Delta and tps2Delta (Tre6P phosphatase) strains . For the tps1Delta strain, ethanol production was significantly lower and was associated with hyperaccumulation of Glu6P and Fru6P . A tps2Delta strain shows reduced accumulation of Glu6P and Fru6P both in intact cells and in permeabilized spheroplasts . These results are not consistent with Tre6P being the sole mediator of hexokinase inhibition . Reconstitution of ethanolic fermentation in permeabilized spheroplasts with glycolytic intermediates indicates additional target site(s) for the Tps1 control . Addition of Tre6P partially shifts the ethanol production rate and the metabolite pattern in permeabilized tps1Delta spheroplasts to those of the wild-type strain, but only with glucose as substrate . This is observed at a very high ratio of glucose to Tre6P . Inhibition of hexokinase activity by Tre6P is less efficiently counteracted by glucose in permeabilized spheroplasts compared to cell extracts, and this effect is largely abolished by deletion of TPS2 but not TPS1 . In permeabilized spheroplasts, hexokinase activity is significantly lower in a tps2Delta strain compared to a wild-type strain and this difference is strongly reduced by additional deletion of TPS1 . These results indicate that Tps1-mediated protein-protein interactions are important for control of glucose influx into yeast glycolysis, that Tre6P inhibition of hexokinase might not be competitive with respect to glucose in vivo and that also Tps2 appears to play a role in the control of hexokinase activity.

Syst Appl Microbiol, 2000 Apr, 23(1), 71 - 85
Molecular typing demonstrates homogeneity of Saccharomyces uvarum strains and reveals the existence of hybrids between S . uvarum and S . cerevisiae, including the S . bayanus type strain CBS 380; Nguyen HV et al.; PCR/RFLP of the NTS2 sequence of rDNA was shown to be suitable for differentiating Saccharomyces sensu stricto species . We previously showed that, within the presently accepted S . bayanus taxon, strains formerly classified as S . uvarum represented a distinct subgroup (Nguyen and Gaillardin, 1997) . In this study, we reidentified 43 more strains isolated recently from wine, cider and various fermentation habitats, and confirmed by karyotyping, hybridization and mtDNA analysis the homogeneity of strains from the S . uvarum subspecies . Molecular typing of nuclear and mitochondrial genomes of strains preserved in collections, and often originating from beer like S . pastorianusNT, revealed the existence of hybrids between S . uvarum and S . cerevisiae . Surprisingly, S . bayanusT CBS380 appeared itself to be a hybrid between S . uvarum and S . cerevisiae . This strain has a mitochondrial genome identical to that of S . uvarum, and a very similar karyotype with 13 isomorphic chromosomes, six of which at least hybridize strongly with S . uvarum chromosomes or with a S . uvarum specific sequence . However, four of the chromosome bands of S . bayanusT bear Y' sequences indistinguishable from those of S . cerevisiae, a feature that is not observed among presently isolated S . uvarum strains . Because of the hybrid nature of S . bayanus(T) and of the scarcity of similar hybrids among present days isolates, we propose to reinstate S . uvarum as a proper species among the Saccharomyces sensu stricto complex.

Syst Appl Microbiol, 2000 Apr, 23(1), 41 - 6
The effect of vitamins and amino acids on glucose uptake in aerobic chemostat cultures of three Saccharomyces cerevisiae strains; de Kock SH et al.; In the respiro-fermentative region of aerobic chemostat cultures at steady state, Saccharomyces cerevisiae CBS 8066 produced high concentrations of ethanol with concomitant low levels of residual glucose which followed Monod kinetics . By contrast, very high residual glucose concentrations were observed in cultures of S . cerevisiae strains ATCC 4126 and NRRL Y132 at dilution rates above 60% of the washout dilution rate, resulting in much lower ethanol concentrations, even though clearly glucose-limited at lower dilution rates in the respiratory region . The addition of a vitamin mixture resulted in decreased residual glucose concentrations in respiro-fermentative cultures of all three strains, but the effect was much more pronounced with strains ATCC 4126 and NRRL Y132 . Meso-inositol was mainly responsible for this effect, although with strain ATCC 4126 other vitamins as well as an amino acid mixture were also required to minimise the steady-state residual glucose levels . The residual glucose concentration in continuous culture was, therefore, greatly dependent on the growth factor requirements of the particular yeast strain, which apparently increased on increasing the dilution rate into the respiro-fermentative region . The strain differences with respect to growth factor requirements at high dilution rates, which were not evident at low dilution rates, had a profound effect on the kinetics of glucose assimilation in aerobic chemostat culture.

FASEB J, 2000 Jul, 14(10), 1279 - 88
Rapid production of the major birch pollen allergen Bet v 1 in Nicotiana benthamiana plants and its immunological in vitro and in vivo characterization; Krebitz M et al.; Type I allergies are immunological disorders that afflict a quarter of the world's population . Improved diagnosis of allergic diseases and the formulation of new therapeutic approaches are based on the use of recombinant allergens . We describe here for the first time the application of a rapid plant-based expression system for a plant-derived allergen and its immunological characterization . We expressed our model allergen Bet v 1, the major birch pollen allergen, in the tobacco-related species Nicotiana benthamiana using a tobacco mosaic virus vector . Two weeks postinoculation, plants infected with recombinant viral RNA containing the Bet v 1 coding sequence accumulated the allergen to levels of 200 microg/g leaf material . Total nonpurified protein extracts from plants were used for immunological characterizations . IgE immunoblots and ELISA (enzyme-linked immunoassay) inhibition assays showed comparable IgE binding properties for tobacco recombinant (r) Bet v 1 and natural (n) Bet v 1, suggesting that the B cell epitopes were preserved when the allergen was expressed in N . benthamiana plants . Using a murine model of type I allergy, mice immunized with crude leaf extracts containing Bet v 1 with purified rBet v 1 produced in E . coli or with birch pollen extract generated comparable allergen-specific IgE and IgG1 antibody responses and positive type I skin test reactions . These results demonstrate that nonpurified Bet v 1 overexpressed in N . benthamina has the same immunogenicity as purified Bet v 1 produced in E . coli or nBet v 1 . We therefore conclude that this plant expression system offers a viable alternative to fermentation-based production of allergens in bacteria or yeasts . In addition, there may be a broad utility of this system for the development of new and low-cost vaccination strategies against allergy.

Appl Environ Microbiol, 2000 Jul, 66(7), 2921 - 7
Antibiosis between ruminal bacteria and ruminal fungi; Dehority BA et al.; Cellulose digestion, bacterial numbers, and fungal numbers were monitored over time in vitro by using a purified cellulose medium with and without antibiotics (penicillin and streptomycin) . All fermentations were inoculated with a 1:10 dilution of whole rumen contents (WRC) . Without antibiotics, cellulose digestion was higher (P < 0.01) at 24, 30, 48, and 72 h; fungi had almost disappeared by 24 h, while bacterial concentrations increased over 100-fold in 24 h and then decreased gradually up to 72 h . In those fermentations with added antibiotics, fungal concentrations increased 4-fold by 30 h and up to 42-fold at 72 h; bacterial concentrations were markedly reduced by 24 h and remained low through 72 h . Similar results were obtained with ground alfalfa as a substrate . In further studies, the in vitro fermentation of purified cellulose without antibiotics was stopped after 18 to 20 h, and the microbial population was killed by autoclaving . Antibiotics were added to half of the tubes, and all tubes were reinoculated with WRC . After 72 h, extensive cellulose digestion had occurred in those tubes without antibiotics, as compared to very low cellulose digestion with added antibiotics . The extent of this inhibition was found to increase in proportion to the length of the initial fermentation period, suggesting the production of a heat-stable inhibitory factor or factors . The inhibitory activity was present in rumen fluid, could be extracted from lyophilized rumen fluid (LRF) with water, and was stable in response to proteolytic enzymes . In addition, the water-extracted residue of LRF was found to contain growth factor activity for rumen fungi in vitro.

J Dairy Sci, 2000 Jun, 83(6), 1350 - 8
Effects of brown midrib 3 mutation in corn silage on productivity of dairy cows fed two concentrations of dietary neutral detergent fiber: 3 . Digestibility and microbial efficiency; Oba M et al.; The effects of digestibility of corn silage neutral detergent fiber (NDF) and dietary NDF content on ruminal digestion kinetics, site of nutrient digestion, and microbial N production efficiency were evaluated with eight multiparous high producing dairy cows in a duplicated 4 x 4 Latin square design with 21-d periods . Experimental diets contained corn silage from a brown midrib (bm3) hybrid or its isogenic normal control at two concentrations of dietary NDF (29 and 38%) . The NDF digestibility estimated by a 30-h in vitro fermentation was higher for bm3 corn silage by 9.4 units (55.9 vs . 46.5%) . Neither ruminal nor total tract NDF digestibility was affected by corn silage treatment . The bm3 corn silage diet decreased starch digestibility in the rumen and in the total tract, but increased postruminal starch digestibility compared with control diet . The bm3 corn silage diets increased microbial N flow to the duodenum and tended to decrease ruminal ammonia concentration . Microbial efficiency was greater for cows fed bm3 corn silage in spite of lower ruminal pH . Higher efficiency of microbial nitrogen production might be attributed to faster passage rate of NDF for cows fed bm3 corn silage compared with those fed control corn silage . Higher in vitro NDF digestibility might predict enhanced NDF fragility and ease of NDF hydrolysis in vivo . Enhanced in vitro NDF digestibility does not necessarily result in increased NDF digestibility either in the rumen or in the total tract, but possibly increases rate of passage and DMI, improving efficiency of microbial N production.

J Dairy Sci, 2000 Jun, 83(6), 1342 - 9
Effects of brown midrib 3 mutation in corn silage on productivity of dairy cows fed two concentrations of dietary neutral detergent fiber: 2 . Chewing activities; Oba M et al.; The effects of digestibility of corn silage neutral detergent fiber (NDF) and dietary NDF content on chewing activities were evaluated with eight multiparous high producing dairy cows in a duplicated 4 x 4 Latin square design with 21-d periods . Experimental diets contained corn silage from a brown midrib (bm3) hybrid or its isogenic normal control at two concentrations of dietary NDF (29 and 38%) . The NDF digestibility estimated by 30-h in vitro fermentation was higher for bm3 corn silage by 9.4 units (55.9 vs . 46.5%) . Feeding behavior of animals and ruminal pH were monitored continuously for 4 d in each period by a computerized data acquisition system . Ruminal digesta were evacuated twice per period to determine the ruminal NDF pool and its turnover rate . There was no effect of NDF digestibility on total chewing time either per day or per kilogram of NDF intake, or on ruminating time either per day or per kilogram of NDF intake . Although bm3 treatments increased the turnover rate of NDF in the rumen, they increased dry matter intake and had no effect on ruminal NDF pool size . Total chewing time and ruminating time per day were related positively to NDF intake and ruminal NDF pool size, but were not related to ruminal turnover rate of NDF . These results provide no evidence that enhanced NDF digestibility decreases the physical effectiveness of NDF of corn silage.

J Dairy Sci, 2000 Jun, 83(6), 1333 - 41
Effects of brown midrib 3 mutation in corn silage on productivity of dairy cows fed two concentrations of dietary neutral detergent fiber: 1 . Feeding behavior and nutrient utilization; Oba M et al.; The effects of digestibility of corn silage neutral detergent fiber (NDF) and dietary NDF content on feeding behavior, dry matter intake (DMI), and energy utilization were evaluated with eight multiparous high producing dairy cows in a duplicated 4 x 4 Latin square design with 21-d periods . Experimental diets contained corn silage from a brown midrib (bm3) hybrid or its isogenic normal control at two concentrations of dietary NDF (29 and 38%) . Both low NDF diets and bm3 corn silage treatments increased DMI and milk yield . However, an interaction between corn silage treatment and dietary NDF content was observed for meal size and for interval between meals, implying that different mechanisms regulating DMI dominate depending on the fermentation characteristics of diets . The bm3 treatment depressed milk fat concentration when fed in a low NDF diet . The bm3 corn silage increased solids-corrected milk yield, but did not affect daily body weight gain compared with control, whereas low NDF diets increased daily body weight gain, but did not affect solids-corrected milk yield compared to high NDF diets . Both bm3 treatments and low NDF diets reduced ruminal pH, but low NDF treatments increased fluctuation of ruminal pH and bm3 treatments did not . Feeding bm3 corn silage increased the energy utilized for milk production, possibly because of a consistent supply of metabolic fuels from the rumen . The beneficial effects of bm3 corn silage on productivity of lactating cows were greater for the cows fed a high NDF diet.

Biochem, Educ. . 2000 May 1, 28(3), 161 - 163
alpha-Amylase production by solid state fermentation: a new practical approach to biotechnology courses; Mulimani VH et al.; A laboratory practical experiment in Biotechnology involving the investigation of alpha-amylase production by solid state fermentation of Gibberella fujikuroi is described.

J Anim Sci, 2000 Jun, 78(6), 1648 - 58
Accounting for the effects of a ruminal nitrogen deficiency within the structure of the Cornell Net Carbohydrate and Protein System; Tedeschi LO et al.; The Cornell Net Carbohydrate and Protein System (CNCPS) prediction of fiber digestion and microbial mass production from ruminally degraded carbohydrate has been adjusted to accommodate a ruminal N deficiency . The steps for the adjustment are as follows: 1) the ruminal available peptide and ammonia pools are used to determine the N allowable microbial growth; 2) this value is subtracted from the energy allowable microbial growth to obtain the reduction in microbial mass; 3) this mass reduction is allocated between pools of bacteria digesting fiber (FC) and nonfiber (NFC) carbohydrate according to their original proportions in the energy allowable microbial growth; 4) the reduction in fermented FC is computed as the FC bacterial mass reduction divided by its yield (g bacteria/g FC digested); and 5) this reduction is added to the FC fraction escaping the rumen . Five published studies included information that allowed us to evaluate the response of animals to added dietary N . These evaluations compared observed and CNCPS-predicted ADG with and without this adjustment . The adjustment decreased the CNCPS overprediction of ADG from 19.2 to 4.7%, mean bias declined from .16 to .04 kg/d, and the r2 of the regression between observed and metabolizable energy (ME) or metabolizable protein allowable ADG was increased from .83 to .88 with the adjustment . When the observed dry matter intake was regressed against CNCPS-predicted DMI with an adjustment for reduction in cell wall digestibility, the r2 was increased from .77 to .88 . These results indicated the adjustment for ruminal nitrogen deficiency increased the accuracy of the CNCPS model in evaluating diets of growing animals when ruminally degraded N is deficient.

J Am Coll Nutr, 2000 Jun, 19(3 Suppl), 300S - 307S
Mechanisms for the impact of whole grain foods on cancer risk; Slavin JL; Dietary guidance recommends consumption of whole grains for the prevention of cancer . Epidemiologic studies find that whole grains are protective against cancer, especially gastrointestinal cancers such as gastric and colonic, and hormonally-dependent cancers including breast and prostate . Four potential mechanisms for the protectiveness of whole grains against cancer are described . First, whole grains are concentrated sources of dietary fiber, resistant starch, and oligosaccharides, fermentable carbohydrates thought to protect against cancer . Fermentation of carbohydrates in the colon results in production of short chain fatty acids that lower colonic pH and serve as an energy source for the colonocytes . Secondly, whole grains are rich in antioxidants, including trace minerals and phenolic compounds, and antioxidants have been proposed to be important in cancer prevention . Thirdly, whole grains are significant sources of phytoestrogens that have hormonal effects related to cancer protection . Phytoestrogens are thought to be particularly important in the prevention of hormonally-dependent cancers such as breast and prostate . Finally, whole grains mediate glucose response, which has been proposed to protect against colon and breast cancer.

Biotechnol Annu Rev, 2000, 5, 1 - 23
Fuzzy control of bioprocess in Japan; Honda H et al.; Process control of bioprocess has been carried out by the judgment of the experts, who are the skilled operators and have lots of experiences for the control of the process . In almost all cases, those experiences are described linguistic IF-THEN rules . Fussy inference is one of the powerful tools to incorporate the linguistic rules to the computer for process control . Fuzzy control are divided into two types; one is the direct fuzzy control of process variables such as sugar feed rate in fed-batch culture and fermentation temperature in batch operation . The other is the indirect control of bioprocess, in which at first the phase recognition is carried out by fuzzy inference and the control strategies constructed in each phase are used for the control of process variables . In Japan, the fuzzy control has already been applied to practical industrial productions, such as pravastatin precursor, vitamin B2, and Japanese sake mashing process . In this review, these industrial approaches of fuzzy control are introduced.

J Bone Miner Metab, 2000, 18(4), 216 - 22
Intake of fermented soybean (natto) increases circulating vitamin K2 (menaquinone-7) and gamma-carboxylated osteocalcin concentration in normal individuals; Tsukamoto Y et al.; Changes in circulating vitamin K2 (menaquinone-7, MK-7) and gamma-carboxylated osteocalcin concentrations in normal individuals with the intake of fermented soybeans (natto) were investigated . Eight male volunteers were given sequentially fermented soybeans (natto) containing three different contents of MK-7 at an interval of 7 days as follows: regular natto including 775 micrograms/100 g (MK-7 x 1) or reinforced natto containing 1298 micrograms/100 g (MK-7 x 1.5) or 1765 micrograms/100 g (MK-7 x 2) . Subsequently, it was found that serum MK-7 and gamma-carboxylated osteocalcin concentrations were significantly elevated following the start of dietary intake of MK-7 (1298 or 1765 micrograms/100 g) . Serum undercarboxylated osteocalcin concentrations were significantly decreased by dietary MK-7 (1765 micrograms/100 g) supplementation . Moreover, the changes in serum MK-7 level with the frequency of dietary natto intake were examined in 134 healthy adults (85 men and 39 women) without and with occasional (a few times per month), and frequent (a few times per week) dietary intake of regular natto including MK-7 (775 micrograms/100 g) . Serum MK-7 and gamma-carboxylated osteocalcin concentrations in men with the occasional or frequent dietary intake of natto were significantly higher than those without any intake . The present study suggests that intake of fermented soybean (natto) increases serum levels of MK-7 and gamma-carboxylated osteocalcin in normal individuals.

Biochem Biophys Res Commun, 2000 Jun 24, 273(1), 45 - 9
Bile acids reduce the apoptosis-inducing effects of sodium butyrate on human colon adenoma (AA/C1) cells: implications for colon carcinogenesis; McMillan L et al.; Butyrate is produced in the colon by fermentation of dietary fibre and induces apoptosis in colon adenoma and cancer cell lines, which may contribute to the protective effect of a high fibre diet against colorectal cancer (CRC) . However, butyrate is present in the colon together with unconjugated bile acids, which are tumour promoters in the colon . We show here that bile acids deoxycholate (DCA) and chenodeoxycholate (CDCA), at levels present in the colon, gave a modest increase in cell proliferation and decreased spontaneous apoptosis in AA/C1 adenoma cells . Bile acids significantly inhibited the induction of apoptosis by butyrate in AA/C1 cells . However, the survival-inducing effects of bile acids on AA/C1 cells could be overcome by increasing the concentration of sodium butyrate . These results suggest that dysregulation of apoptosis in colonic epithelial cells by dietary factors is a key factor in the pathophysiology of CRC .

Mikrobiol Z, 2000 Mar-Apr, 62(2), 19 - 26
{The effect of glucose on the biosynthesis of extracellular enzymes by Streptomyces recifensis var . lyticus 2435 and its mutants}; Zhernosekova IV et al.; Different glucose concentrations have been studied for their effect on biosynthesis of bacterio- and proteolytic enzymes in Streptomyces recifensis var . lyticus 2435 and strains II-29 and 2P-15 obtained from the latter . It has been established that synthesis of enzymes by the parent strain 2435 was subjected to catabolite repression: synthesis of proteolytic and staphylolytic enzymes was the most sensitive to glucose, synthesis of glucosidases was less sensitive . The role of catabolite repression in regulation of synthesis of bacteriolytic enzymes to the latter in strain II-29 and rifamycin-resistant mutant 2P-15 is essentially decreased . As compared with the initial strains, the high level of activity of lytic enzymes in the strain 2P-15 is achieved at higher glucose concentrations in the fermentation medium, synthesis of staphylolysins and glucosideases are inhibited inconsiderably at 2% and 4% content of glucose, in medium respectively and it does not decrease under further increase of glucose concentration.

Mikrobiol Z, 2000 Mar-Apr, 62(2), 11 - 8
{The effect of glucose on the fatty acid level in Saccharomyces cerevisiae and Schwanniomyces occidentalis cells}; Hudz' SP et al.; The influence of glucose on fatty acid contents in the cells of yeast Saccharomyces cerevisiae and Schwanniomyces occidentalis has been studied . It was shown that fatty acid contents in Schw . occidentalis was enhanced with the increase of glucose concentration in the medium . The increase of glucose concentration in the medium during respiro-fermentative growth phase reduces the level of C18 in unsaturated fatty acids of S . cerevisiae; the increase of fatty acids content in total lipids and nonetherified fatty acids was observed under the same conditions in the respiratory phase . It was established that during the growth of S . cerevisiae in the medium with 0.1% glucose the level of nonetherified fatty acids was higher and with 4% glucose--lower in comparison with Schw . occidentalis . The correlation of nonetherified and etherified fatty acid levels in S . cerevisiae is higher in comparison with Schw . occidentalis grown under the same conditions.

J Nat Prod, 2000 Jun, 63(6), 777 - 81
Ribofuranosyl triazolone: a natural product herbicide with activity on adenylosuccinate synthetase following phosphorylation; Schmitzer PR et al.; 2,4-Dihydro-4-(beta-D-ribofuranosyl)-1,2,4(3H)-triazol-3-one (2) was identified as the principal phytotoxic component of a fermentation broth derived from an Actinomadura . The compound is a new natural product, but known by synthesis . Broad-spectrum herbicidal activity was demonstrated in greenhouse tests . Metabolite reversal studies suggested the target site was adenylosuccinate synthetase, which was confirmed by direct measurement of the activity of the 5'-phosphorylated derivative on the isolated enzyme.

J Nat Prod, 2000 Jun, 63(6), 757 - 61
Phytotoxic compounds from the new coprophilous fungus guanomyces polythrix; Macias M et al.; Bioactivity-directed fractionation of the fermentation broth and mycelium of the coprophilous fungus Guanomyces polythrix led to the isolation of several phytotoxic compounds, including five new naphthopyranone derivatives (1-5) . In addition, rubrofusarin B, emodin, citrinin, and 4-hydroxybenzoic acid methyl ester were obtained . The structures of the new compounds were established by spectral and chiroptical methods . The isolates caused significant inhibition of radicle growth of two weed seedlings (Amaranthus hypochondriacus and Echinochloa crusgalli) and interacted with both spinach and bovine brain calmodulins.

J AOAC Int, 2000 May-Jun, 83(3), 635 - 50
Determination of the levels of isoflavonoids in soybeans and soy-derived foods and estimation of isoflavonoids in the Japanese daily intake; Nakamura Y et al.; The levels of 6 kinds of isoflavonoids found in 11 domestic and imported soybeans, and 12 kinds of soybean-based processed foods in Japan were systematically analyzed, and the Japanese daily intake of isoflavonoids from those foods was estimated . The total isoflavonoids (daidzein, glycitein, and genistein) were analyzed with acid hydrolysis and the intact isoflavonoids (daidzein, glycitein, genistein, daidzin, glycitin, and genistin) were analyzed without hydrolysis . This was followed by cleanup with an ODS cartridge column and determined by liquid chromatography with a diode array detector . The highest content of isoflavonoids was found in kinako (a roasted soybean powder) and the lowest was found in soy sauce . The contents and composition of the isoflavonoids in the 11 soybeans varied by species and country of origin . The level of isoflavonoids found in the processed foods varied by manufacturing method or ingredients . The percentage of aglycone tended to be higher in miso (fermented soybean paste) and soy sauce, which are heated and fermented during the manufacturing process . Japanese daily intake of isoflavonoids from soybeans and soybean-based processed foods was estimated as 27.80 mg per day (daidzein 12.02 mg, glycitein 2.30 mg, and genistein 13.48 mg).

Prikl Biokhim Mikrobiol, 2000 May-Jun, 36(3), 287 - 92
{Change in composition of structural components of leaf cellulose sulfate during fermentation bleaching by xylanases}; Aleksandrova GP et al.; The possibility of the use of xylanase preparations for hydrolysing hemicelluloses in a non-bleached kraft pulp in order to facilitate its bleaching was studied . The effects of enzymatic preparations of the fungal and bacterial origins were examined, and the optimal conditions for xylanase activity were determined . UV spectroscopy demonstrated that the treatment of kraft pulp with the enzymatic preparations containing xylanase facilitated the subsequent removal of lignin and increased the brightness by 5% . The effect of enzymatic treatment was retained in the case of peroxide bleaching . The enzymatic preparations studied are promising for the development of chlorine-free pulp bleaching technologies.

Prikl Biokhim Mikrobiol, 2000 May-Jun, 36(3), 243 - 53
{Fermentation waste--a biomodifier of concrete}; Balobova AV et al.; The composition and properties of molasses-containing yeast fermentation waste used as a plasticizer and structural biological additive to concrete mixtures are reviewed . The basic principles of the effects of organic and inorganic components of yeast fermentation waste on the properties of the bonding system are analyzed.

J Antibiot (Tokyo), 2000 Apr, 53(4), 357 - 63
SB-203207 and SB-203208, two novel isoleucyl tRNA synthetase inhibitors from a Streptomyces sp . I . Fermentation, isolation and properties; Stefanska AL et al.; Two novel inhibitors of isoleucyl tRNA synthetase designated SB-203207 and SB-203208 have been detected in the culture of a new Streptomyces species . The fermentation, isolation and some properties of the inhibitors are described.

J Antibiot (Tokyo), 2000 Apr, 53(4), 337 - 44
New Cdc25B tyrosine phosphatase inhibitors, nocardiones A and B, produced by Nocardia sp . TP-A0248: taxonomy, fermentation, isolation, structural elucidation and biological properties; Otani T et al.; Strain TP-A0248 which produces two new Cdc25B tyrosine phosphatase inhibitors and also possessing antifungal activity, designated nocardiones A (1) and B (2), was considered to belong to the genus Nocardia on the basis of literature comparison of chemotaxonomic properties . The nocardiones were isolated by solvent extraction of fermentation broth of Nocardia sp . TP-A0248 and purified by the conventional column chromatography . Spectroscopic studies led to determination that 1 and 2 belong to a class compound of naphtho{1,2-b}furan-4,5-diones . Compound 1 inhibited the activity of Cdc25B, PTP1B and FAP-1 protein tyrosine phosphatases at a concentration of 10 microM . It also showed moderate in vitro antifungal and cytotoxic activity.

J Biotechnol, 2000 Jun 9, 80(1), 55 - 62
Effect of RQ and pre-seed conditions on biomass and galactosyl transferase production during fed-batch culture of S . cerevisiae BT150; Zigov inverted question marka J; A feedback RQ controlled fed-batch process for the recombinant production of a soluble human N-deglycosylated recombinant beta-1, 4-galactosyltransferase (NdrGal-T) with Saccharomyces cerevisiae BT150 was investigated . Several RQ values were tested for optimal production of NdrGal-T . Four times higher volumetric activity was reached at RQ=1.0 (32 U l(-1)) than at all higher RQ values (about 8 U l(-1)) . RQ, 1.0 was the best choice for both, biomass and enzyme production . Optimal concentration of glucose in preculture was 25 g l(-1) . At higher values slightly more ethanol was produced than at lower values of preculture glucose concentrations, moreover no positive effect on biomass and enzyme production was found . Lower values caused not only decrease of ethanol but also decrease of biomass formation (from 1.69 g h(-1) to 0.81 g h(-1)) and enzyme overall productivity (from 2.2 U h(-1) to 0.63 U h(-1)) . Successfully performed cultivation with three precultures predicted scale-up possibility of feedback RQ-controlled NdrGal-T production with S . cerevisiae BT150 from lab to pilot-scale fermentor.

Enzyme Microb Technol, 2000 Jul 1, 27(1-2), 127 - 133
Biological detoxification of coffee husk by filamentous fungi using a solid state fermentation system; Brand D et al.; Studies were carried out on detoxification of coffee husk in solid state fermentation using three different strains of Rhizopus, Phanerochaete, and Aspergillus sp . Fungal strains were selected by their ability to grow on a coffee husk extract-agar medium . Using R . arrizus LPB-79, the best results on the degradation of caffeine (87%) and tannins (65%) were obtained with pH 6.0 and moisture 60% in 6 days . When P . chrysosporium BK was used, maximum degradation of caffeine and tannins were 70.8 and 45%, respectively, with coffee husk having 65% moisture and pH 5.5 in 14 days . The Aspergillus strain, isolated from the coffee husk, showed best biomass formation on coffee husk extract-agar medium . Optimization assays were conducted using factorial design, and surface response experiments with Aspergillus sp . The best detoxification rates achieved were 92% for caffeine and 65% for tannins . The results showed good prospects of using these fungal strains, in particular Aspergillus sp., for the detoxification of coffee husk.

Enzyme Microb Technol, 2000 Jul 1, 27(1-2), 74 - 82
Production, properties and application to nonaqueous enzymatic catalysis of lipase from a newly isolated Pseudomonas strain; Gaoa X et al.; A potent bacterium for lipase production was isolated from soil and identified as Pseudomonas species . It produced lipase constitutively . A mutant of this strain with a lipase productivity 3.25-fold higher was obtained by treatment with ultraviolet (UV) and nitrosoguanidine (NTG) . Its fermentation condition was optimized to a lipase yield of 87.5 U/ml . The lipase had maximum activity at pH 9.0 and 45 degrees C . It was stable at pHs from 7.0 to 11.0 and below 60 degrees C . The effects of metal ions, surfactants and bile salts were also studied . The lipase was 1,3-specific . In organic solvents, the thermal stability of the lipase was significantly enhanced . Its optimum temperature was also slightly increased . The optimum water activity was found between 0.5 and 0.6 . The lipase was successfully applied in organic phase to catalyze the glycerolysis of palm oil for monoglyceride (MG) production, and the enantioselective esterification of (R,S)-2-octanol . The enantioselectivity of the lipase could be enhanced substantially by treatment with an amphipathic.

Enzyme Microb Technol, 2000 Jun 1, 26(9-10), 826 - 833
Mathematical modeling of citric acid production by repeated batch culture; Arzumanov TE et al.; A mathematical model has been created for the process of citric acid biosynthesis by yeast (mutant strain Yarrowia lipolytica) cultivated by the repeated batch (RB) method on ethanol under conditions of nitrogen limitation . The model accounts for cell growth as a function of nitrogen concentration in the culture liquid; nitrogen uptake by growing cells; citric acid production; pH control in the fermentor by means of NaOH addition; and changes in system volume . The model represents a system of five nonlinear differential equations . Experimental measurements of cell concentration, citric acid concentration, and cultivation broth volume were used with the least squares method to determine the values of eight model parameters . The parameter values obtained were consistent with literature data and general concepts of cell growth and citric acid biosynthesis . The model has been used to predict optimum RB culture conditions.

Enzyme Microb Technol, 2000 Jun 1, 26(9-10), 812 - 818
From gene to product in yeast: production of fungal cutinase; Verripsab T et al.; In the mid-1970s, information technology and recombinant DNA technology were considered as the breakthrough technologies of the final quarter of the 20th century . Now, about 25 years later, information technology has penetrated deeply into our society and nearly everyone uses this technology . Compared to the formidable success of information technology, the progress in the commercialization of recombinant DNA technology is moderate, even when taking into account that all that is related to the technological application of biological sciences needs extensive safety testing . However, there are signs that the speed of this commercialization will increase in the first decade of the 21st century . Moreover, new breakthroughs in our understanding of the complete genetic make up of eukaryotes will contribute to this increase in speed . An important aspect of the commercialization of this technology is the development of cells as factories for the production of valuable and/or useful molecules . Lower eukaryotes, such as yeasts and molds, are the most promising candidates to become the factories of the future, but at present these factories still contains a lot of process lines that may be superfluous under the well controlled conditions in fermentors . On the other hand, the speed and yield of these cellular production lines can be increased by eliminating the rate-determining steps of these process lines . In this contribution to the European Union symposium from Cell to Factory, some steps in the improvement of S . cerevisiae as cell factories for (heterologous) hydrophobic molecules are presented.

Enzyme Microb Technol, 2000 Jun 1, 26(9-10), 801 - 807
Accelerated prediction of recombinant protein production in Saccharomyces cerevisiae by using rapid monitoring techniques; Brown R et al.; The use of a stopped-flow analyser for the monitoring of the production of a secreted recombinant protein, a wild-type cutinase, from S . cerevisiae CEN.PK111-32D pUR7320 is described . Induction is through use of a galactose promoter, and the monitoring facility is used to record the formation of the cutinase and cell density with optical density measurements . A range of induction conditions was studied with a view to using the monitoring to predict the likely level of cutinase formation . Results achieved within 4 to 5 h of induction were of sufficient quality to allow the use of simple modelling relating cutinase formation and cell production to predict likely final specific activities of the product . The utility of such monitoring and prediction is discussed with regards to improved process confidence and definition during fermentation production.

Enzyme Microb Technol, 2000 Jun 1, 26(9-10), 743 - 747
The influence of hexoses addition on the fermentation of d-xylose in Debaryomyces hansenii under continuous cultivation; Tavares JM et al.; The effect of hexoses (glucose and galactose) addition to the feed xylose mineral medium of Debaryomyces hansenii chemostat cultures grown at a constant dilution rate of 0.055 h(-1) was studied . Xylitol was the major product detected amongst all tested conditions . The maximal values for xylitol yield and volumetric productivity (0.56 gg(-1) xylose and 0.21 gl(-1)h(-1), respectively) were obtained for a glucose/xylose feeding ratio of 10%, showing that the addition of small amounts of glucose, but not galactose, enhanced the xylitol production . A xylitol yield increase of 30%, compared with the sole xylose-containing feed medium, was observed . It was found that the oxygen requirement for D . hansenii growth is lower under glucose compared with xylose . Ethanol and glycerol were only produced for glucose/xylose feeding ratio above 30% . The byproducts accumulation was correlated with glucose metabolism, because a direct relationship between the increase of ethanol (and glycerol) concentration and the increase of glucose in the feed medium was found.

Enzyme Microb Technol, 2000 Jun 1, 26(9-10), 724 - 736
Regulation of fermentative capacity and levels of glycolytic enzymes in chemostat cultures of Saccharomyces cerevisiae; van Hoek P et al.; Regulation of fermentative capacity was studied in chemostat cultures of two Saccharomyces cerevisiae strains: the laboratory strain CEN.PK113-7D and the industrial bakers' yeast strain DS28911 . The two strains were cultivated at a fixed dilution rate of 0.10 h(-1) under various nutrient limitation regimes: aerobic and anaerobic glucose limitation, aerobic and anaerobic nitrogen limitation on glucose, and aerobic ethanol limitation . Also the effect of specific growth rate on fermentative capacity was compared in glucose-limited, aerobic cultures grown at dilution rates between 0.05 h(-1) and 0.40 h(-1) . Biomass yields and metabolite formation patterns were identical for the two strains under all cultivation conditions tested . However, the way in which environmental conditions affected fermentative capacity (assayed off-line as ethanol production rate under anaerobic conditions) differed for the two strains . A different regulation of fermentative capacity in the two strains was also evident from the levels of the glycolytic enzymes, as determined by in vitro enzyme assays . With the exception of phosphofructokinase and pyruvate decarboxylase in the industrial strain, no clear-cut correlation between the activities of glycolytic enzymes and the fermentative capacity was found . These results emphasise the need for controlled cultivation conditions in studies on metabolic regulation in S . cerevisiae and demonstrate that conclusions from physiological studies cannot necessarily be extrapolated from one S . cerevisiae strain to the other.

Enzyme Microb Technol, 2000 Jun 1, 26(9-10), 706 - 714
An interlaboratory comparison of physiological and genetic properties of four Saccharomyces cerevisiae strains; van Dijken JP et al.; To select a Saccharomyces cerevisiae reference strain amenable to experimental techniques used in (molecular) genetic, physiological and biochemical engineering research, a variety of properties were studied in four diploid, prototrophic laboratory strains . The following parameters were investigated: 1) maximum specific growth rate in shake-flask cultures; 2) biomass yields on glucose during growth on defined media in batch cultures and steady-state chemostat cultures under controlled conditions with respect to pH and dissolved oxygen concentration; 3) the critical specific growth rate above which aerobic fermentation becomes apparent in glucose-limited accelerostat cultures; 4) sporulation and mating efficiency; and 5) transformation efficiency via the lithium-acetate, bicine, and electroporation methods . On the basis of physiological as well as genetic properties, strains from the CEN.PK family were selected as a platform for cell-factory research on the stoichiometry and kinetics of growth and product formation.

Enzyme Microb Technol, 2000 Jun 1, 26(9-10), 671 - 677
Yield improvement of heterologous peptides expressed in yps1-disrupted Saccharomyces cerevisiae strains; Egel-Mitani M et al.; Heterologous protein expression levels in Saccharomyces cerevisiae fermentations are highly dependent on the susceptibility to endogenous yeast proteases . Small peptides, such as glucagon and glucagon-like-peptides (GLP-1 and GLP-2), featuring an open structure are particularly accessible for proteolytic degradation during fermentation . Therefore, homogeneous products cannot be obtained . The most sensitive residues are found at basic amino acid residues in the peptide sequence . These heterologous peptides are degraded mainly by the YPS1-encoded aspartic protease, yapsin1, when produced in the yeast . In this article, distinct degradation products were analyzed by HPLC and mass spectrometry, and high yield of the heterologous peptide production has been achieved by the disruption of the YPS1 gene (previously called YAP3) . By this technique, high yield continuous fermentation of glucagon in S . cerevisiae is now possible.

Biotechnol Bioeng, 2000 Aug 20, 69(4), 429 - 39
Studies on the interaction of fermentation and microfiltration operations: erythromycin recovery from Saccharopolyspora erythraea fermentation broths; Davies JL et al.; Changes in fermentation media not only affect the performance of the fermentation itself (with regard to the kinetics of biomass and product formation and the yields obtained) but also the initial product-recovery operations downstream of the fermentor . In this work, microfiltration experiments to remove Saccharopolyspora erythraea biomass from fermentation broth and to recover erythromycin were carried out using two fundamentally different media; a soluble complex medium (SCM) and an oil-based process medium (OBM) . Small-scale batch fermentations of 14-L working volume were carried out in triplicate using both media . Broth samples were taken from each fermentation at regular intervals from the end of the exponential-growth phase onwards . These were then processed using a Minitan II (acrylic), tangential crossflow-filtration module, fitted with a single 60 cm(2) Durapore hydrophilic 0.2 microm membrane, operated in concentration mode . The OBM fermentations produced higher titers of erythromycin but required longer fermentation times due to increased lag phases and slower maximum-growth rates . The OBM also increased the loading on the membrane; at maximum product titers residual oil concentrations of 3 g . L(-1), antifoam concentrations of 2 g . L(-1) and flour concentrations estimated at approximately 10 g/L(-1) were typical . It was found that both the permeate flux and erythromycin transmission were affected by the choice of medium . The OBM had significantly lower values for both parameters (12.8 Lm(-2) h(-1) and 89.6% respectively) than the SCM (35.9 Lm(-2) h(-1) and 96.7% respectively) when the fermentations were harvested at maximum erythromycin titers . Transmission of erythromycin stayed approximately constant as a function of fermentation time for both media, however, for the OBM the permeate flux decreased with time which correlated with an increase in broth viscosity . The relatively poor microfiltration performance of the OBM medium was, however, offset by the higher titers of erythromycin that were achieved during the fermentation . The filtration characteristics of the SCM broth did not show any correlation with either broth viscosity or fermentation time . Image-analysis data suggested that there was a correlation between hyphal morphology (main hyphal length) and permeate flux (no such correlation was found for the OBM broth) . Moreover, it has been shown for the OBM broth that the residual flour had a profound effect on the microfiltration characteristics . The influence of the residual flour was greater than that imposed by the morphology and concentration of the biomass . The understanding of the factors governing the interaction of the fermentation and microfiltration operations obtained in this work provides a first step towards optimization of the overall process sequence .

Biotechnol Bioeng, 2000 Aug 20, 69(4), 377 - 84
Data model for the elimination of matrix effects in enzyme-based flow-injection systems; Becker TM et al.; This contribution presents a new conceptional enzyme-based flow injection analysis (FIA) system for the process and quality control of food processing and biotechnological systems . It provides the determination of different analytes in distinct process media on the base of a common experimental set-up . In contrast to known comparable systems, analysis is performed without the commonly used sample preparation and dilution steps . Instead, the adaptation to the necessary measurement range is realized by optimization of intrinsic system parameters . The central principle of the work presented is the elimination of occurring interferences by the heterogeneous matrix of the process sample . Based on a particular injection mode, the application of dehydrogenases as indicator enzymes and a specially developed data model using cognitive methods, cross sensitivities of the detector as well as disturbed reaction rates of the enzymes could be almost completely compensated . Two applications are presented, the analysis of ethanol in non-alcoholic beer and the online determination of D-/L-lactate during a lactic acid fermentation, which reveal the advantage of the developed system .

Yeast, 2000 Jun 15, 16(8), 675 - 729
Tailoring wine yeast for the new millennium: novel approaches to the ancient art of winemaking; Pretorius IS; Yeasts are predominant in the ancient and complex process of winemaking . In spontaneous fermentations, there is a progressive growth pattern of indigenous yeasts, with the final stages invariably being dominated by the alcohol-tolerant strains of Saccharomyces cerevisiae . This species is universally known as the 'wine yeast' and is widely preferred for initiating wine fermentations . The primary role of wine yeast is to catalyze the rapid, complete and efficient conversion of grape sugars to ethanol, carbon dioxide and other minor, but important, metabolites without the development of off-flavours . However, due to the demanding nature of modern winemaking practices and sophisticated wine markets, there is an ever-growing quest for specialized wine yeast strains possessing a wide range of optimized, improved or novel oenological properties . This review highlights the wealth of untapped indigenous yeasts with oenological potential, the complexity of wine yeasts' genetic features and the genetic techniques often used in strain development . The current status of genetically improved wine yeasts and potential targets for further strain development are outlined . In light of the limited knowledge of industrial wine yeasts' complex genomes and the daunting challenges to comply with strict statutory regulations and consumer demands regarding the future use of genetically modified strains, this review cautions against unrealistic expectations over the short term . However, the staggering potential advantages of improved wine yeasts to both the winemaker and consumer in the third millennium are pointed out .

Biotechnol Bioeng, 2000 Aug 5, 69(3), 339 - 43
Optimized synthesis of L-sorbose by C(5)-dehydrogenation of D-sorbitol with Gluconobacter oxydans; De Wulf P et al.; The optimization of L-sorbose synthesis by regiospecific dehydrogenation of D-sorbitol using Gluconobacter oxydans is reported . The current L-sorbose production processes that are based on G . oxydans and other bacterial strains are suboptimal as to yield and rate of L-sorbose synthesis . One reason for these problems is the toxicity that is induced by the substrate D-sorbitol when used in concentrations of >10% (w/v) . This phenomenon significantly limits the potentials of L-sorbose production from an industrial point of view . The goal of this study was to develop a fast production process that yields L-sorbose in stoichiometric amounts starting from D-sorbitol concentrations that exceed 10% (w/v) . A gradual improvement of the inoculum build-up procedure, culture medium composition, and process parameters ultimately led to a theoretically maximal L-sorbose productivity (200 g L(-1) of L-sorbose from 200 g L(-1) of D-sorbitol in 28 h of fermentation) using a Gluconobacter oxydans mutant strain that was selected under conditions of substrate inhibition . Because the D-sorbitol/L&HYPHEN;sorbose bioconversion is used to mass-produce vitamin C, the procedure reported here will contribute to a more efficient and more economic synthesis of vitamin C .

Biotechnol Bioeng, 2000 Aug 5, 69(3), 312 - 22
Determination of yeast glycogen content by individual cell spectroscopy using image analysis; Cahill G et al.; A rapid technique has been developed to determine the glycogen content of yeast on an individual cell basis using a combination of image analysis technology and staining of yeast cells with an I(2):KI solution . Changes in mean cellular glycogen content during alcoholic fermentation have been reported using this technique . The glycogen content of stored brewer's yeast is heterogeneous compared to freshly propagated yeast which have a more uniform distribution of glycogen . Analysis of the distribution of yeast glycogen during fermentation indicates that a fraction of yeast cells do not dissimilate glycogen . Therefore, conventional analysis of the mean glycogen content of yeast used to inoculate fermentations is of limited use, unless information regarding the proportion of cells which utilize glycogen is known . Analysis of the distribution of glycogen within a yeast population can serve as a useful indicator of yeast quality .

Biotechnol Bioeng, 2000 Aug 5, 69(3), 292 - 300
Growth and sporulation stoichiometry and kinetics of Coniothyrium minitans on agar media; Ooijkaas LP et al.; Coniothyrium minitans was cultivated on agar media with different concentrations of starch, urea, and trace elements . By means of elemental balances, the stoichiometry of growth and sporulation was established . C . minitans produced byproducts on all media, especially in the medium with high urea concentrations, where 30% of the starch was converted into byproducts . Simple empirical models were used to describe the kinetics of growth, sporulation, CO(2) production, and substrate consumption on all media . Total biomass and mycelium could be described reasonably well with the logistic law . Starch, urea, and oxygen consumption and CO(2) production could be described as a function of total biomass by the linear-growth model of Pirt . There were almost no differences between media for the estimates of yield coefficients and maintenance coefficients . Only at high urea concentrations were maintenance coefficients much higher . Similar to substrate consumption and CO(2) production, the kinetics of sporulation could be described as a function of mycelium production with the linear-growth model . It is shown that sporulation of C . minitans is growth-associated . Based on kinetics, the process costs for producing spores are roughly calculated . In addition, it is shown that fermentor costs represent the majority of production costs .

Biotechnol Bioeng, 2000 Jul 20, 69(2), 213 - 21
Study of drop and bubble sizes in a simulated mycelial fermentation broth of up to four phases; Galindo E et al.; The mean sizes and size distributions of air bubbles and viscous castor oil drops were studied in a salt-rich aqueous solution (medium), first separately, and then simultaneously as a three-phase system . The dispersion was created in a 150-mm-diameter stirred tank equipped with a Rushton turbine, and the sizes were measured using an advanced video technique . Trichoderma harzianum biomass was added in some experiments to study the effect of a solid phase under unaerated and aerated conditions to give either three-or four-phase systems . In all cases, the different dispersed phases could be clearly seen . Such photoimages have never been obtained previously . For the three phases, air-oil-medium, aeration caused a drastic increase in Sauter mean drop diameter, which was greater than could be accounted for by the reduction in energy dissipation on aeration . Also, as in the unaerated case, larger drops were observed as the oil content increased . On the other hand, mean bubble sizes were significantly reduced with increasing oil phase up to 15% with bubbles inside many of the viscous drops . With the introduction of fungal biomass of increasing concentration (0.5 to 5 g L(-1)) under unaerated conditions, the Sauter mean drop diameter decreased . Finally, in the four-phase system (oil {10%}-medium-air-biomass) as found in many fermentations, all the phases (plus bubbles in drops) could clearly be seen and, as the biomass increased, a decrease in both the bubble and the drop mean diameters was found . The reduction in size of bubbles (and therefore increase in interfacial area) as the oil and bio- mass concentration increased provides a possible explanation as to why the addition of an oil phase has been reported to enhance oxygen transfer during many fermentations .

Biotechnol Bioeng, 2000 Jul 20, 69(2), 119 - 28
Influence of scale-up on the quality of recombinant human growth hormone; Bylund F et al.; The aerobic fed-batch production of recombinant human growth hormone (rhGH) by Escherichia coli was studied . The goal was to determine the production and protein degradation pattern of this product during fed-batch cultivation and to what extent scale differences depend on the presence of a fed-batch glucose feed zone . Results of laboratory bench-scale, scale-down (SDR), and industrial pilot-scale (3-m(3)) reactor production were compared . In addition to the parameters of product yield and quality, also cell yield, respiration, overflow, mixed acid fermentation, glucose concentration, and cell lysis were studied and compared . The results show that oxygen limitation following glucose overflow was the critical parameter and not the glucose overflow itself . This was verified by the pattern of byproduct formation where formate was the dominating factor and not acetic acid . A correlation between the accumulation of formate, the degree of heterogeneity, and cell lysis was also visualized when recombinant protein was expressed . The production pattern could be mimicked in the SDR reactor for all parameters, except for product quantity and quality, where 30% fewer rhGH-degraded forms were present and where about 80% higher total yield was achieved, resulting in 10% greater accumulation of properly formed rhGH monomer .

J Cardiopulm Rehabil, 2000 May-Jun, 20(3), 143 - 6
Dietary trans fatty acid; Lichtenstein AH; Trans fatty acids are unsaturated fatty acids that contain at least one double bond in the trans configuration . In the diet they occur at relatively low levels in meat and dairy products as a by-product of fermentation in ruminant animals or in hydrogenated fats as a consequence of the hydrogenation process . In general, dietary hydrogenated fat/trans fatty acids have been reported to increase LDL cholesterol levels relative to oil in the natural state or cis fatty acids . In contrast, dietary hydrogenated fat/trans fatty acids have been reported have to have little effect or decrease HDL cholesterol levels, the later observation restricted to relatively high intakes of trans fatty acids . These two effects result in higher, therefore less favorable, total or LDL cholesterol/HDL cholesterol ratios . Significant increases in Lp(a) levels have been reported after consumption of diets relatively high in trans fatty acids compared with either unsaturated or saturated fatty acids . However, the magnitude of the change is for the most part small and the physiological significance of this observation has yet to be resolved . Data related to the mechanism by which hydrogenated fat/trans fatty acids alter serum lipid levels and other risk factors for cardiovascular disease are in the nascent stages . At this time it would appear prudent that public health recommendations should be aimed at encouraging the moderate consumption of products low in saturated fat or minimally hydrogenated . Trans fatty acids intake should not be stressed at the expense of saturated fat but should augment it.

Antimicrob Agents Chemother, 2000 Jul, 44(7), 1809 - 17
Genetic localization and molecular characterization of the nonS gene required for macrotetrolide biosynthesis in Streptomyces griseus DSM40695; Smith WC et al.; The macrotetrolides are a family of cyclic polyethers derived from tetramerization, in a stereospecific fashion, of the enantiomeric nonactic acid (NA) and its homologs . Isotope labeling experiments established that NA is of polyketide origin, and biochemical investigations demonstrated that 2-methyl-6,8-dihydroxynon-2E-enoic acid can be converted into NA by a cell-free preparation from Streptomyces lividans that expresses nonS . These results lead to the hypothesis that macrotetrolide biosynthesis involves a pair of enantiospecific polyketide pathways . In this work, a 55-kb contiguous DNA region was cloned from Streptomyces griseus DSM40695, a 6.3-kb fragment of which was sequenced to reveal five open reading frames, including the previously reported nonR and nonS genes . Inactivation of nonS in vivo completely abolished macrotetrolide production . Complementation of the nonS mutant by the expression of nonS in trans fully restored its macrotetrolide production ability, with a distribution of individual macrotetrolides similar to that for the wild-type producer . In contrast, fermentation of the nonS mutant in the presence of exogenous (+/-)-NA resulted in the production of nonactin, monactin, and dinactin but not in the production of trinactin and tetranactin . These results prove the direct involvement of nonS in macrotetrolide biosynthesis . The difference in macrotetrolide production between in vivo complementation of the nonS mutant by the plasmid-borne nonS gene and fermentation of the nonS mutant in the presence of exogenously added (+/-)-NA suggests that NonS catalyzes the formation of (-)-NA and its homologs, supporting the existence of a pair of enantiospecific polyketide pathways for macrotetrolide biosynthesis in S . griseus . The latter should provide a model that can be used to study the mechanism by which polyketide synthase controls stereochemistry during polyketide biosynthesis.

Int J Food Microbiol, 2000 May 25, 56(1), 105 - 9
Ochratoxin formation in Aspergillus ochraceus with particular reference to spoilage of coffee; Mantle PG et al.; Production of ochratoxin on media by eight isolates of Aspergillus ochraceus from coffee or its processing environment in India, Indonesia, Kenya, and Brazil, and seven Brazilian isolates from other commodities, has been compared with yields in shaken fermentation on shredded wheat and coffee (Coffea arabica) . Shredded wheat most consistently allowed expression of biosynthesis of ochratoxins A and B in yields up to 3.5% of the dry product . Culture on artificial media was an unreliable predictor of ochratoxin yield on both shredded wheat and coffee . Coffee was a relatively poor substrate for ochratoxin production particularly when sterilised . Notably, two Asian coffee isolates produced 400 mg kg(-1) ochratoxin A on unsterilised ground green coffee, showing this to be a preferred substrate for further experimentation . The study focused on isolates of A . ochraceus, which from evidence of culture on media would not be expected to be suitable fungi for future studies to establish both the fact of spoilage of coffee by A . ochraceus and the dynamics of ochratoxin formation by isolates of this species.

Curr Microbiol, 2000 Aug, 41(2), 89 - 95
Influence of yeast flocculation on the rate of Jerusalem artichoke extract fermentation; Schorr-Galindo S et al.; Variations in residual sugar composition have been observed during Jerusalem artichoke extract fermentations by using Saccharomyces diastaticus NCYC 625, a flocculating yeast strain . In batch cultures, these differences were due to the inulin polymer size distribution of the extracts: measurements of enzymatic activities on different polymerized substrates have shown that the hydrolysis and fermentation yield decreased when the fructose/glucose ratio of the extract increased . Inulin hydrolysis appeared to be the limiting factor of the fermentation rate . A comparison of continuous and batch cultures with the same extract showed that fermentability differences were related to the structure and size of the yeast flocs . This led to an hydrolysis selectivity of the inulin polymers according to their size: the chemostat culture in which the floc average size was larger gave longer chained residual sugars.

J S Afr Vet Assoc, 1999 Dec, 70(4), 167 - 71
Laminitis and dermatitis in heifers associated with excessive carbohydrate intake: skin lesions and biochemical findings; Yeruham I et al.; The effects of a sudden addition of a large quantity of readily fermentable carbohydrate to the feed ration of pregnant heifers are described . Clinical and pathological changes caused by the resulting disease were confined to the digits and skin . The 4 acutely affected heifers were reluctant to get up or move (group II) . They tended to lie down or stand with feet bunched together and the back arched, often shifting weight from limb to limb . They walked stiffly with great tenderness and pain in the digits . Extreme pain was noticed when the digits were examined . In 4 of 8 heifers, separation of the sole at the heel, with leakage of exudate, and under-running of the sole were observed . Necrotic dermatitis of the legs, alopecia and hyperkeratosis of the tail were noticed in all 8 heifers . Skin lesions appeared simultaneously . Four of the heifers (group I) recovered, and the other 4 (group II) were sent to slaughter . No post mortem examination was performed . The biochemical findings revealed a significantly higher concentration of total serum globulins and sodium, and increased activity, in CK, LDH and AST . A significantly decreasing pattern was noted in blood urea concentration, cholesterol, triglycerides, albumin and calcium . No significant differences among the various groups were found in the activities of amylase, GGT, and concentration of creatinine, total bilirubin, inorganic phosphorus, magnesium and potassium.

Appl Microbiol Biotechnol, 2000 May, 53(5), 610 - 2
Study on the production of 6-pentyl-alpha-pyrone using two methods of fermentation; Kalyani A et al.; The lactone 6-pentyl-alpha-pyrone has a characteristic coconut aroma and is produced by Trichoderma species . A study on the fermentative production of 6-pentyl-alpha-pyrone in both surface and submerged conditions by Trichoderma harzianum was carried out . Maximum concentrations of 455 mg/l and 167 mg/l after 96 h and 48 h of fermentation in surface and submerged conditions, respectively, were obtained without using any additional recovery operations . The resultant yields are higher than those previously reported in the literature, which may be attributable to strain characteristics in combination with the choice of fermentation conditions employed in the present study . Enough scope exists for further improvement in the yields by optimizing the cultural and nutritional parameters.

Appl Microbiol Biotechnol, 2000 May, 53(5), 601 - 4
Biotransformation of methyl ent-17-hydroxy-16beta-kauran-19-oate by Rhizopus stolonifer; Vieira HS et al.; Methyl ent-17-hydroxy-16beta-kauran-19-oate was fed to a 2-day-old culture of the fungus Rhizopus stolonifer, fermenting at room temperature (25 degrees C) in an orbital shaker (21) . After 11 days, both broth and mycelia were extracted with ethyl acetate . Two novel compounds were isolated from this experiment: methyl ent-9alpha,17-dihydroxy-16beta-kauran-19-oate and methyl ent-7alpha,17-dihydroxy-16beta-kauran-19-oate . Their structures were fully confirmed by spectroscopic methods.

Appl Microbiol Biotechnol, 2000 May, 53(5), 568 - 74
Overexpression of the OLE1 gene enhances ethanol fermentation by Saccharomyces cerevisiae; Kajiwara S et al.; The fermentation characteristics of Saccharomyces cerevisiae strains which overexpress a constitutive OLE1 gene were studied to clarify the relationship between the fatty acid composition of this yeast and its ethanol productivity . The growth yield and ethanol productivity of these strains in the medium containing 15% dextrose at 10 degrees C were greater than those of the control strains under both aerobic and anaerobic conditions but this difference was not observed under other culture conditions . During repeated-batch fermentation, moreover, the growth yield and ethanol productivity of the wild-type S . cerevisiae increased gradually and then were similar to those of the OLE1-overexpressing transformant in the last batch fermentation . However, the unsaturated fatty acid content (77.6%) of the wild-type cells was lower than that (86.2%) of the OLE1-recombinant cells . These results suggested that other phenomena caused by the overexpression of the OLE1 gene, rather than high unsaturated fatty acid content, are essential to ethanol fermentation by this yeast.

Exp Brain Res, 2000 May, 132(2), 203 - 15
Pavlovian aversive and appetitive odor conditioning in humans: subjective, peripheral, and electrocortical changes; Hermann C et al.; Differential Pavlovian conditioning of aversive and appetitive odors was examined in 30 male healthy subjects . The appetitive conditioning group (n=15) received a pleasant odor (vanilla), the aversive conditioning group (n=15) an unpleasant odor (fermented yeast) as unconditioned stimulus . Slides of two different neutral faces that were easy to discriminate served as conditioned stimuli (CS) . An EEG was recorded from nine electrodes . Electromyographic activity was measured bilaterally from the m . corrugator supercilii and m . zygomaticus . The startle response was obtained from the m . orbicularis oculi . Finally, heart rate and skin conductance response were assessed . The subjective data and the skin conductance response revealed successful differential aversive conditioning . By contrast, the pleasant odor failed to produce appetitive odor conditioning . The conditioned and unconditioned response of the corrugator muscles confirm previous reports on the m . corrugator being strongly involved in the expression of negative affect . Contrary to previous findings, magnitude of the startle reflex was not found to be modulated depending on the valence of the CS . Central psychophysiological parameters showed little change during differential conditioning . The presence of subjective-evaluative conditioning and contingency awareness without significant changes in cortical and cardiovascular correlates might be due to extremely localized cortical processing of conditioned olfactory cues or primarily subcortical processing . The latter interpretation is strengthened by the presence of differential conditioning in non-voluntary responses such as the corrugator muscles.

J Bacteriol, 2000 Jun, 182(12), 3423 - 8
Purification and characterization of a membrane-bound hydrogenase from the hyperthermophilic archaeon Pyrococcus furiosus; Sapra R et al.; Highly washed membrane preparations from cells of the hyperthermophilic archaeon Pyrococcus furiosus contain high hydrogenase activity (9.4 micromol of H(2) evolved/mg at 80 degrees C) using reduced methyl viologen as the electron donor . The enzyme was solubilized with n-dodecyl-beta-D-maltoside and purified by multistep chromatography in the presence of Triton X-100 . The purified preparation contained two major proteins (alpha and beta) in an approximate 1:1 ratio with a minimum molecular mass near 65 kDa and contained approximately 1 Ni and 4 Fe atoms/mol . The reduced enzyme gave rise to an electron paramagnetic resonance signal typical of the so-called Ni-C center of mesophilic NiFe-hydrogenases . Neither highly washed membranes nor the purified enzyme used NAD(P)(H) or P . furiosus ferredoxin as an electron carrier, nor did either catalyze the reduction of elemental sulfur with H(2) as the electron donor . Using N-terminal amino acid sequence information, the genes proposed to encode the alpha and beta subunits were located in the genome database within a putative 14-gene operon (termed mbh) . The deduced sequences of the two subunits (Mbh 11 and 12) were distinctly different from those of the four subunits that comprise each of the two cytoplasmic NiFe-hydrogenases of P . furiosus and show that the alpha subunit contains the NiFe-catalytic site . Six of the open reading frames (ORFs) in the operon, including those encoding the alpha and beta subunits, show high sequence similarity (>30% identity) with proteins associated with the membrane-bound NiFe-hydrogenase complexes from Methanosarcina barkeri, Escherichia coli, and Rhodospirillum rubrum . The remaining eight ORFs encode small (<19-kDa) hypothetical proteins . These data suggest that P . furiosus, which was thought to be solely a fermentative organism, may contain a previously unrecognized respiratory system in which H(2) metabolism is coupled to energy conservation.

Zh Mikrobiol Epidemiol Immunobiol, 1999 May-Jun, (3), 6 - 9
{The development of a process for culturing Bordetella pertussis immobilized on a polyurethane carrier}; Miriasova LV et al.; The processes of the cultivation of Bordetella pertussis, immobilized on polyurethane carrier in a fermenter, were carried out and studied . Acellular pertussis preparations were produced from the culture fluid obtained in the batch and multi-cycle cultivation processes with immobilized cells, as well as in the process with interrupted fermentation (for confirming the possibility of the preservation of cell viability) . The content of protein and B . pertussis toxin in these preparations, as well as their leukocytes-stimulating and hemagglutinating activity, did not differ from similar characteristics of preparations obtained from culture fluid in homogeneous cultivation.

Curr Opin Microbiol, 2000 Jun, 3(3), 248 - 51
Environmentally directed mutations and their impact on industrial biotransformation and fermentation processes; Zelder O et al.; Microbial adaptation plays an important role in the selection of improved strains for biotechnological processes and for the maintenance and stability of the selected production strains . Most of the knowledge about adaptation processes and environmentally directed mutations originates from environmental microbiology and from studies on biological evolution . The increasing information on the molecular mechanisms of adapted mutations and on the development of methods frequently used in environmental and evolutionary microbiology, such as the selection in semi-continuous cultures or chemostats, can be used as input and tools for the improvement of industrial production organisms.

Curr Opin Microbiol, 2000 Jun, 3(3), 263 - 9
Microecology of the termite gut: structure and function on a microscale; Brune A et al.; Long considered simply as anoxic fermentors, termite guts are in fact axially and radially structured environments with physicochemically distinct microhabitats . Recent developments in termite gut microecology, which combined traditional and modern techniques, have focused on the spatial organization of important microbial populations and their in situ activities, and have significantly furthered our understanding of functional interactions within highly structured microenvironments.

Appl Biochem Biotechnol, 2000 Apr, 87(1), 37 - 55
Continuous production of antibiotics in an airlift fermentor utilizing a transverse magnetic field; Al-Qodah Z; In this study, a series of experiments was conducted to demonstrate the feasibility of continuous production of penicillin antibiotic using a three-phase magneto airlift fermentor with immobilized Penicillium chrysogenum . The fermentation processes were carried out in a 2.4-L external loop airlift utilizing a transverse magnetic field . It was found that the application of the magnetic field to a bed of ferromagnetic beads affects both the hydrodynamics of the reactor and the rate of the bioconversion process occurring inside it . One hundred hours after startup, the maximum penicillin concentration increased 48% as the magnetic field intensity increased from 0 to 35 mT, owing to the increased residence time of the substrate in the riser and the positive effect of the magnetic field on the effective fluid-solid interfacial area . In addition, the detached biomass concentration in the liquid phase was found to be only 5% of the immobilized biomass, owing to low shear levels and the absence of friction among the solid-phase particles.

Appl Biochem Biotechnol, 2000 Apr, 87(1), 1 - 15
Expression and high-level secretion of Trichoderma reesei endoglucanase I in Yarrowia lipolytica; Park CS et al.; The endoglucanase I (EGI) from fungus Trichoderma reesei was cloned, expressed, and secreted from Yarrowia lipolytica using the XPR2 promoter . The signal sequence of EGI transferred from T . reesei was efficiently processed in the Y . lipolytica secretory pathway and directed the secretion of active EGI into the culture medium . However, the recombinant EGI produced from YLCSIn strain was hyperglycosylated and significantly larger than the native enzyme produced by the parent strain . The expression of EGI using XPR2 preproregion has caused secretion of modified proteins that still retained cellulase activity . This resulted from imprecise processing of the N-terminus of recombinant protein . While the batch culture produced 5 mg EGI/L from YLCSIn strain, the EGI yield was increased approx 20-fold when the fed-batch fermentation process strategy in combination with the high-cell density cultivation technique was employed . These results showed that the Y . lipolytica is a useful host organism for production of a large amount of large size heterologous proteins, especially when used in combination with high-cell density and fed-batch culture techniques.

Cancer Biother Radiopharm, 1999 Aug, 14(4), 277 - 89
MSC, a new benzoquinone-containing natural product with antimetastatic effect; Hidvegi M et al.; An orally applicable fermentation product of wheat germ containing 0.04% substituted benzoquinone (MSC) has been invented by Hungarian chemists under the trade name of AVEMAR . Oral administration (3 g/kg body weight) of MSC enhances blastic transformation of splenic lymphocytes in mice . The same treatment shortens the survival time of skin grafts in a co-isogenic mouse skin transplantation model, pointing to the immune-reconstructive effect of MSC . A highly significant antimetastatic effect of MSC has been observed in three metastasis models (3LL-HH, B16, HCR-25) . The antimetastatic effect of MSC--besides the immune-reconstitution--may also be due to its cell adhesion inhibitory, cell proliferation inhibitory, apoptosis enhancing, and antioxidant characteristics, also observed in our in vitro experiments . It is even more noteworthy that combined treatment with MSC and one of the following antineoplastic agents (5-FU and DTIC)--both in wide use in every day clinical practice--exhibited a significantly enhanced antimetastatic effect in appropriate metastasis models (established from C38 mouse colon carcinoma and B16 mouse melanoma respectively) as compared to the effect elicited by any component of these therapeutic compositions (MSC + 5-FU and MSC + DTIC) administered alone . The results show that the fermented wheat germ extract (MSC) has more than an additive effect and synergistically enhanced the metastasis inhibitory effect of both antineoplastic agents studied till now . It is also worthy of mention that the synchronous treatment with MSC profoundly decreased the toxic side effects of the applied antineoplastic agents (decreased weight loss etc) . Based on the biological effects of MSC--shown to be non-toxic by subacute toxicology studies--this product may be used as an adjuvant in the therapy of malignant neoplasia and other diseases caused by or following immune-deficiency.

Appl Biochem Biotechnol, 2000 Spring, 84-86, 1137 - 45
Lipase production by Penicillium restrictum using solid waste of industrial babassu oil production as substrate; Palma MB et al.; Lipase, protease, and amylase production by Penicillium restrictum in solid-state fermentation was investigated . The basal medium was an industrial waste of babassu oil (Orbignya oleifera) production . It was enriched with peptone, olive oil, and Tween-80 . The supplementation positively influenced both enzyme production and fungal growth . Media enriched with Tween-80 provided the highest protease activity (8.6 U/g), whereas those enriched with peptone and olive oil led to the highest lipase (27.8 U/g) and amylase (31.8 U/g) activities, respectively.

Appl Biochem Biotechnol, 2000 Spring, 84-86, 1049 - 61
Recovery of volatile products from dilute high-fouling process streams; Madson PW et al.; As biomass hydrolysis and fermentation technologies approach commercial viability, advancements in product recovery technologies will be required . For cases in which fermentation products are more volatile than water, recovery by distillation is often the technology of choice . Distillation technologies that will allow the economic recovery of dilute volatile products from streams containing a variety of impurities have been developed and commercially demonstrated . Distillation tower and tray designs, along with specialized heat-exchanger designs, allowing for extended processing intervals on solutions containing lignocellulosic residues, organic acids, and inorganic salts concentrations > 100 g/L are in commercial operation . In the case of ethanol, distillation energy consumption efficiencies for processing solutions containing < 40 g/L of desired product can approach demonstrated energy consumption efficiencies for solutions containing concentrations > 120 g/L . These proprietary technologies have been applied individually at commercial scale, and designs have been developed that incorporate the combined technologies with only a marginal increase in capital investment compared to traditional methods.

Appl Biochem Biotechnol, 2000 Spring, 84-86, 859 - 74
Improvements in titer, productivity, and yield using Solka-floc for cellulase production; Hayward TK et al.; Researchers studying cellulase enzymes for the economical production of fuel ethanol envision cellulose as the carbon source . However, submerged Trichoderma reesei cultures grown on cellulose exhibit high run-to-run variability . Thus, an investigation of 30 batch cellulase production experiments was instrumental in determining fermentation conditions that improved enzyme titers, yields, and productivities . Eighteen of the 30 batch experiments experienced minimal process upsets and were classified into eight groups based on agitation rate, gas sparge rate, and the use of oxygen supplementation . Comparing corn steep liquor with yeast extract/peptone also tested the effect of different sources of nitrogen in the media . Average 7-d enzyme titers were doubled from 4 to 8 FPU/mL primarily by increasing aeration.

Appl Biochem Biotechnol, 2000 Spring, 84-86, 801 - 8
Inhibition of microbial xylitol production by acetic acid and its relation with fermentative parameters; Morita TA et al.; Precipitated sugarcane bagasse hemicellulosic hydrolysate containing acetic acid was fermented by Candida guilliermondii FTI20037 under different operational conditions (pH 4.0 and 7.0, three aeration rates) . At pH 7.0 and kLa of 10 (0.75 vvm) and 22.5/h (3.0 vvm) the acetic acid had not been consumed until the end of the fermentations, whereas at the same pH and kLa of 35/h (4.5 vvm) the acid was rapidly consumed and acetic acid inhibition was not important . On the other hand, fermentations at an initial pH of 4.0 and kLa of 22.5 and 35/h required less time for the acid uptake than fermentations at kLa of 10/h . The acetic acid assimilation by the yeast indicates the ability of this strain to ferment in partially detoxified medium, making possible the utilization of the sugarcane bagasse hydrolysate in this bio-process . The effects on xylitol yield and production are reported.

Appl Biochem Biotechnol, 2000 Spring, 84-86, 779 - 89
Mycelial pellet formation by Rhizopus oryzae ATCC 20344; Zhou Y et al.; Factors in a cultivation medium affecting fungal growth morphology and fumaric acid production by Rhizopus oryzae ATCC 20344 were investigated . These factors included the initial pH value and trace metals such as zinc, magnesium, iron, and manganese in the cultivation medium . It was found that a significant change in the growth morphology of R . oryzae ATCC 20344 occurs when the initial pH value is varied . A lower initial pH value in the cultivation medium was inhibitory to fungal growth, and fast growth in the cultivation medium at a higher initial pH value promoted the formation of large pellets or filamentous forms . Trace metals in the cultivation media also had significant effects on pellet formation and fumaric acid fermentation.

Appl Biochem Biotechnol, 2000 Spring, 84-86, 751 - 9
Effects of initial pH on biological synthesis of xylitol using xylose-rich hydrolysate; Morita TA et al.; Sugarcane bagasse, an agricultural residue plentiful in Brazil, was utilized for xylitol production by a biotechnological process . A medium fermentation prepared with this xylose-rich biomass at an oxygen transfer volumetric coefficient of 10/h1 and different initial pH values was inoculated with cells of Candida guilliermondii FTI 20037 . The maximum values of xylitol and cell volumetric productivities (Qp = 0.56 g/{L.h} and Qx = 0.11 g/{g.h}), xylitol yield factor (YP/S = 0.79 g/g), and xylose uptake rate (qs = 0.197 g/{g.h}) were attained at pH 7.0 without further pH control . The results show that the yeast performance was influenced by the pH, an important bioengineering parameter in this fermentation process.

Appl Biochem Biotechnol, 2000 Spring, 84-86, 743 - 9
Solid-state fermentation with Aspergillus niger for cellobiase production; Tsao GT et al.; Aspergillus niger NRRL3 was cultivated in a moist wheat bran and ground corncob solid medium supplemented with inorganic minerals for the production of cellobiase (beta-1,4-glucosidase, EC 3.2.1.21) . With this method, A . niger NRRL3 was able to produce a high concentration of cellobiase (215 IU/g of solid substrate) after 96 h of incubation . Temperature and moisture content affected final cellobiase titers . The best conditions for cellobiase production from solid substrate by A . niger NRRL3 were determined to be 70% moisture and 35 degrees C.

Appl Biochem Biotechnol, 2000 Spring, 84-86, 679 - 91
Cellulase production of Trichoderma reesei Rut C 30 using steam-pretreated spruce . Hydrolytic potential of cellulases on different substrates; Szengyel Z et al.; Various techniques are available for the conversion of lignocellulosics to fuel ethanol . During the last decade processes based on enzymatic hydrolysis of cellulose have been investigated more extensively, showing good yield on both hardwood and softwood . The cellulase production of a filamentous fungi, Trichoderma reesei Rut C 30, was examined on carbon sources obtained after steam pretreatment of spruce . These materials were washed fibrous steam-pretreated spruce (SPS), and hemicellulose hydrolysate . The hemicellulose hydrolysate contained, besides water-soluble carbohydrates, lignin and sugar degradation products, which were formed during the pretreatment and proved to be inhibitory to microorganisms . Experiments were performed in a 4-L laboratory fermentor . The hydrolytic capacity of the produced enzyme solutions was compared with two commercially available enzyme preparations, Celluclast and Iogen Cellulase, on SPS, washed SPS, and Solka Floc cellulose powder . There was no significant difference among the different enzymes produced by T . reesei Rut C 30 . However, the conversion of cellulose using these enzymes was higher than that obtained with Iogen or Celluclast cellulases using steam-pretreated spruce as substrate.

Appl Biochem Biotechnol, 2000 Spring, 84-86, 643 - 54
Study of biocatalyst to produce ethanol from starch . Coimmobilization of glucoamylase and yeast in gel; Giordano RL et al.; This article presents a detailed study on the conditions for achieving a stable biocatalyst to be used in the production of ethanol from starch . Different pellets were used depending on which characteristic of the biocatalyst was being studied: (a) Saccharomyces cerevisiae entrapped in pectin or calcium alginate gel particles; (b) silica containing immobilized glucoamylase entrapped in pectin gel particles; or (c) pectin gel particles, with the silica-enzyme derivative and yeast coimmobilized . The influence of several variables on the mechanical resistance of the particle, on the viability of the microorganism, and on the rate of substrate hydrolysis was studied with biocatalyst . The best conditions found were 6% pectin gel, 2-mm particle diameter, and cure in 0.2M CaCl2.2H2O/60 mM acetate buffer, pH 4.2, for gel preparation; and 6.0 g/L of CaCl2.2H2O in the fermentation medium . Biocatalyst (c) was successfully tested for the production of ethanol from liquefied manioc flour syrup.

Appl Biochem Biotechnol, 2000 Spring, 84-86, 633 - 41
Effect of the oxygen transfer coefficient on xylitol production from sugarcane bagasse hydrolysate by continuous stirred-tank reactor fermentation; Martinez EA et al.; The effect of the oxygen transfer coefficient on the production of xylitol by bioconversion of xylose present in sugarcane bagasse hemicellulosic hydrolysate using the yeast Candida guilliermondii was investigated . Continuous cultivation was carried out in a 1.25-L fermentor at 30 degrees C, pH 5.5, 300 rpm, and a dilution rate of 0.03/h, using oxygen transfer coefficients of 10, 20, and 30/h . The results showed that the microbial xylitol production (11 g/L) increased by 108% with the decrease in the oxygen volumetric transfer coefficient from 30 to 20/h . The maximum values of xylitol productivity (0.7 g/{L.h}) and yield (0.58 g/g) were obtained akLa 20/h.

Appl Biochem Biotechnol, 2000 Spring, 84-86, 617 - 32
Influence of lignocellulose-derived aromatic compounds on oxygen-limited growth and ethanolic fermentation by Saccharomyces cerevisiae; Larsson S et al.; Phenolic compounds released and generated during hydrolysis inhibit fermentation of lignocellulose hydrolysates to ethanol by Saccharomyces cerevisiae . A wide variety of aromatic compounds form from lignin, which is partially degraded during acid hydrolysis of the lignocellulosic raw material . Aromatic compounds may also form as a result of sugar degradation and are present in wood as extractives . The influence of hydroxy-methoxy-benzaldehydes, diphenols/quinones, and phenylpropane derivatives on S . cerevisiae cell growth and ethanol formation was assayed using a defined medium and oxygen-limited conditions . The inhibition effected by the hydroxy-methoxy-benzaldehydes was highly dependent on the positions of the substituents . A major difference in inhibition by the oxidized and reduced form of a diphenol/quinone was observed, the oxidized form being the more inhibitory . The phenylpropane derivatives were examined with respect to difference in toxicity depending on the oxidation-reduction state of the gamma-carbon, the presence and position of unsaturated bonds in the aliphatic side chain, and the number and identity of hydroxyl and methoxyl substituents . Transformations of aromatic compounds occurring during the fermentation included aldehyde reduction, quinone reduction, and double bond saturation . Aromatic alcohols were detected as products of reductions of the corresponding aldehydes, namely hydroxy-methoxy-benzaldehydes and coniferyl aldehyde . High molecular mass compounds and the corresponding diphenol were detected as products of quinone reduction . Together with coniferyl alcohol, dihydroconiferyl alcohol was identified as a major transformation product of coniferyl aldehyde.

Appl Biochem Biotechnol, 2000 Spring, 84-86, 577 - 93
Simulated dynamics and control of an extractive alcoholic fermentation; Costa AC et al.; In this study, we investigated the dynamics of a computer simulation of a continuous alcoholic fermentation process combined with a flash column under vacuum . The alcohol was partially extracted in order to maintain its concentration at about 40 kg/m3 in the fermentor . The mathematical model of the fermentation was developed for industrial conditions and considers the effect of the temperature on the kinetic parameters . The performance of the dynamic matrix control algorithm, single input single output and multiple input multiple output, for the control of the extractive process was studied . The concepts of factorial design were used in a simulation study to determine the best control structures for the process.

Appl Biochem Biotechnol, 2000 Spring, 84-86, 561 - 76
Two-stage dilute-acid pretreatment of softwoods; Nguyen QA et al.; Whole tree chips obtained from softwood forest thinnings were pretreated via single- and two-stage dilute-sulfuric acid pretreatment . Whole-tree chips were impregnated with dilute sulfuric acid and steam treated in a 4-L steam explosion reactor . In single-stage pretreatment, wood chips were treated using a wide range of severity . In two-stage pretreatment, the first stage was carried out at low severity to maximize hemicellulose recovery . Solubilized sugars were recovered from the first-stage prehydrolysate by washing with water . In the second stage, water-insoluble solids from first-stage prehydrolysate were impregnated with dilute sulfuric acid, then steam treated at more severe conditions to hydrolyze a portion of the remaining cellulose to glucose and to improve the enzyme digestibility . The total sugar yields obtained after enzymatic hydrolysis of two-stage dilute acid-pretreated samples were compared with sugar yields from single-stage pretreatment . The overall sugar yield from two-stage dilute-acid pretreatment was approx 10% higher, and the net enzyme requirement was reduced by about 50% . Simultaneous saccharification and fermentation using an adapted Saccharomyces cerevisiae yeast strain further improved cellulose conversion yield and lowered the enzyme requirement.

Appl Biochem Biotechnol, 2000 Spring, 84-86, 505 - 24
Repeated solid-phase fermentation and extraction for enzyme production; Tsao GT et al.; Solid-phase fermentation has been found to have a much higher productivity than the popular liquid submerged fermentation in producing cellulase enzymes . The highest reported productivity in the literature for cellulases by Trichoderma cultures in submerged fermentation is 158 filter paper units (FPU)/(h.L) of fermenting liquid . From preliminary experiments of solid-phase fermentation in 1000-mL flasks, a productivity of 234 FPU of cellulases/(h.L) of solid-bed volume was obtained . When two novel techniques--pressure pulsation and repeated extraction--were applied, a productivity of 806 FPU/(h.L) was achieved . The same techniques also greatly enhanced the productivity of other enzymes by fungal cultures in solid-phase fermentation.

Appl Biochem Biotechnol, 2000 Spring, 84-86, 371 - 80
Effects of environmental conditions on xylose reductase and xylitol dehydrogenase production by Candida guilliermondii; Sene L et al.; The effects of environmental conditions, namely initial pH (2.5-7.0) and temperature (25 and 35 degrees C), on xylose reductase and xylitol dehydrogenase levels, as well as on xylitol production, were evaluated . Although the fermentative parameter values increased with an increase in pH and temperature (the maximum Yp/s and Qp were 0.75 g/g and 0.95 g/{L.h}, respectively, both attained at pH 6.0, 35 degrees C), the highest xylose reductase activities (nearly 900 IU/mg of protein) were observed at an initial pH varying from 4.0 to 6.0 . Xylitol dehydrogenase was favored by an increase in both initial pH and temperature of the medium . The highest xylitol dehydrogenase specific activity was attained at pH 6.5 and 35 degrees C (577 IU/mg of protein).

Appl Biochem Biotechnol, 2000 Spring, 84-86, 237 - 45
Enzyme production of Trichoderma reesei Rut C-30 on various lignocellulosic substrates; Shin CS et al.; Economical production of cellulase enzyme is key for feasible bio-ethanol production from lignocellulosics using an enzyme-based process . On-site cellulase production can be more feasible with the process of separate hydrolysis and fermentation (SHF) than with simultaneous saccharification and fermentation, since the cost of enzyme is more important and a variety of substrates are available for the SHF process . Cellulase production using various biomass substrates available for SHF, including paper sludge, pretreated wood (steam exploded), and their hydrolysis residues, was investigated in shake flasks and a fermenter for their productivities and titers . Among the newspaper sludge, office paper sludge, and steam-exploded woods treated in various ways, the steam-exploded wood showed the best properties for substrate in cellulase production . The best titer of 4.29 IU/mL was obtained using exploded wood of 2% (w/v) slurry in the shake flask, and the titer with the same substrate was duplicated to about 4.30 IU/mL in a 3.7-L fermenter . Also, the yield of enzyme reached 215 IU/g of substrate or 363 IU/g of cellulose . Despite various pretreatment attempts, newspaper and office paper substrate was inferior to the exploded-wood substrate for cellulase production . However, hydrolysis residues of papers showed quite promising results . The hydrolysis residue of office paper produced 2.48 IU/mL of cellulase in 7 d . Hence, the utilization of hydrolysis residues for cellulase production will be further investigated in the future.

Appl Biochem Biotechnol, 2000 Spring, 84-86, 81 - 96
Cellulose and hemicellulose hydrolysis models for application to current and novel pretreatment processes; Jacobsen SE et al.; Acids catalyze the hydrolysis of cellulose and hemicellulose to produce sugars that organisms can ferment to ethanol and other products . However, advanced low- and no-acid technologies are critical if we are to reduce bio-ethanol costs to be competitive as a pure fuel . We believe carbohydrate oligomers play a key role in explaining the performance of such hydrolysis processes and that kinetic models would help us understand their role . Various investigations have developed reaction rate expressions based on an Arrhenius temperature dependence that is first order in substrate concentration and close to first order in acid concentration . In this article, we evaluate these existing hydrolysis models with the goal of providing a foundation for a unified model that can predict performance of both current and novel pretreatment process configurations.

Appl Biochem Biotechnol, 2000 Spring, 84-86, 39 - 50
Fourier transform infrared quantification of sugars in pretreated biomass liquors; Tucker MP et al.; The process of converting renewable lignocellulosic biomass to ethanol requires a number of steps, and pretreatment is one of the most important . Pretreatment usually involves a hydrolysis of the easily hydrolyzed hemi-cellulosic component of biomass using some form of thermal/chemical/mechanical action that results in a product that can be further hydrolyzed by cellulase enzymes (the cellulosic portion) . The sugars produced can then be fermented to ethanol by fermentative microorganisms . If the pretreatment step is not severe enough, the resultant residue is not as easily hydrolyzed by the cellulase enzyme . More severe pretreatment conditions result in the production of degradation products that are toxic to the fermentative microorganism . In this article, we report the quantitative analysis of glucose, mannose, xylose, and acetic acid using Fourier transform infrared (FTIR) spectroscopy on liquors from dilute-acid-pretreated soft-wood and hard-wood slurries . Comparison of FTIR and high-performance liquid chromatography quantitative analyses of these liquors are reported . Recent developments in infrared probe technology has enabled the rapid quantification of these sugars by FTIR spectroscopy in the batch reactor during optimization of the pretreatment conditions, or interfaced to the computer controlling a continuous reactor for on-line monitoring and control.

J Appl Microbiol, 2000 Jun, 88(6), 1065 - 71
Influence of phenolic compounds on the physiology of Oenococcus oeni from wine; Reguant C et al.; This study shows that the growth of Oenococcus oeni CECT 4100 in a synthetic medium is affected by phenolic compounds in different ways, depending on their type and concentration . Generally they have no effects at low concentrations, but hydroxycinnamic acids are inhibitory at high concentrations . Malolactic fermentation was stimulated in the presence of catechin and quercetin, but increasingly delayed with increasing amounts of p-coumaric acid . Gallic acid appeared to delay or inhibit the formation of acetic acid from citric acid . This could lead to a better control of malolactic fermentation and suppress the increase in volatile acidity, which is undesirable in the wine-making process.

J Nat Prod, 2000 May, 63(5), 602 - 4
Cytonic acids A and B: novel tridepside inhibitors of hCMV protease from the endophytic fungus Cytonaema species; Guo B et al.; Two novel human cytomegalovirus protease inhibitors, cytonic acids A (1) and B (2), have been isolated from the solid-state fermentation of the endophytic fungi Cytonaema sp . Their structures as p-tridepside isomers were elucidated by MS and NMR methods.

Int J Syst Evol Microbiol, 2000 May, 50 Pt 3, 1125 - 31
Acholeplasma vituli sp . nov., from bovine serum and cell cultures; Angulo AF et al.; Organisms isolated from commercial foetal bovine serum and from cell culture lines containing such serum supplements were found to consist of non-helical, non-motile, pleomorphic coccoid forms . One strain (FC 097-2T) cultivated directly from foetal bovine serum was selected for characterization . In ultrastructural examination, individual round cells lacked cell wall structures and cells varied in size, with a mean diameter of about 700 nm . However, variable numbers of cells were filterable through membranes of 300 nm . Optimum growth occurred between 30 and 37 degrees C . The organism fermented glucose, fructose and mannose, but did not hydrolyse arginine . The strain was insensitive to 500 U penicillin ml(-1) and was capable of growing in the absence of serum or cholesterol . The organism was serologically distinct from all 13 currently described species in the genus Acholeplasma and from other sterol-requiring species in the genus Mycoplasma, using growth inhibition, immunoperoxidase and immunofluorescence tests . Strain FC 097-2T was found to have a DNA G+C composition between 37.6 +/- 1 mol% and 38.3 +/- 1 mol% . The genome size was determined to be 2095 kbp . The 16S rDNA sequence of strain FC 097-2T was compared to 16S rDNA sequences of other mollicutes in nucleotide databases . No deposited sequence was found to be identical; the closest relatives were several members of the genus Acholeplasma . On the basis of these findings and other similarities to acholeplasmas in morphology and growth, the absence of a sterol requirement for growth, and similar genomic characteristics, the organism was assigned to the genus Acholeplasma . Strain FC 097-2T is designated the type strain (ATCC 700667T) of a new species, Acholeplasma vituli.

Int J Syst Evol Microbiol, 2000 May, 50 Pt 3, 1035 - 42
Reductively debrominating strains of Propionigenium maris from burrows of bromophenol-producing marine infauna; Watson J et al.; Two novel strains of Propionigenium maris able to reductively debrominate 2,4,6-tribromophenol (TBP) to monobromophenols were isolated from marine hemichordate and polychaete burrows . These two strains, DSL-1 and ML-1, were anaerobic, non-motile rods that stained Gram-negative and required 0.05% yeast extract for growth . Strain DSL-1 fermented pyruvate and succinate to predominantly butyrate and strain ML-1 fermented glucose and succinate primarily to propionate . No inorganic terminal electron acceptors were identified . The pH and temperature optima for growth were 7.6 and 30 degrees C for strain DSL-1 and 7.0 and 32 degrees C for strain ML-1, respectively; doubling times for strains DSL-1 and ML-1 were 0.32 h and 0.30 h, respectively . Both strains required 2-3% (w/v) NaCl for optimal growth . Morphological and physiological features, as well as the results of 16S rDNA sequence analysis, showed these to be new strains of Propionigenium maris . Because they differ from the P . maris type strain (DSM 9537T) in a number of respects, including their ability to rapidly debrominate di- and tribromophenols, and in their specific habitats, the species description is amended to include these ecologically important properties.

J Biol Regul Homeost Agents, 2000 Apr-Jun, 14(2), 83 - 92
Antibody production by molecular farming in plants; Fischer R et al.; "Molecular farming" is the production of pharmaceutical proteins in transgenic plants and has great potential for the production of therapeutic anti-cancer antibodies and recombinant therapeutic proteins . Plants make fully functional recombinant human or animal antibodies . Cultivating transgenic plants on an agricultural scale will produce almost unlimited supplies of recombinant proteins for uses in medicine . Combinatorial library technology is a key tool for the generation and optimisation of therapeutic antibodies ahead of their expression in plants . Optimised antibody expression can be rapidly verified using transient expression assays in plants before creation of transgenic suspension cells or plant lines . Subcellular targeting signals that increase expression levels and optimise protein stability can be identified and exploited using transient expression to create high expresser plant lines . When high expresser lines have been selected, the final step is the development of efficient purification methods to retrieve functional antibody . Antibody production on an industrial scale is then possible using plant suspension cell culture in fermenters, or by the propagation of stably transformed plant lines in the field . Recombinant proteins can be produced either in whole plants or in seeds and tubers, which can be used for the long-term storage of both the protein and its production system . The review will discuss these developments and how we are moving toward the molecular farming of therapeutic antibodies becoming an economic and clinical reality.

Bioseparation, 2000, 9(1), 43 - 53
A new method for yeast recovery in batch ethanol fermentations: filter aid filtration followed by separation of yeast from filter aid using hydrocyclones; da Matta VM et al.; In the Melle-Boinot process for alcohol production, centrifuges are normally used for yeast recovery at the end of a batch fermentation . Centrifuges are expensive equipment and represent an impressive part of the equipment costs in alcohol industries . In the present work, an alternative method for yeast recovery using less expensive equipment was studied . Instead of using centrifuges, yeast was separated from the fermented broth by filter aid filtration, followed by separation of yeast from the filter aid using hydrocyclones . A stainless steel plate-and-frame filter of filtration area 1.14 m2 and two 30 mm hydrocyclones, which followed the Bradley and Rietema recommended proportions, were used in this work . The filter aid was perlite . Tests of direct separation of yeast from the fermented broth using the Bradley hydrocyclone proved to be completely unfeasible, since the maximal reduced total efficiency obtained was only 1% . When the hydrocyclones were used to separate perlite from the resuspended filtration cake, the perlite total separation efficiency obtained in the underflow was as high as 95% when using the Bradley hydrocyclone with an underflow diameter of 3 mm . To show the feasibility of the proposed new method of yeast recovery, a complete cycle of experiments, which included fermentation, yeast separation, and new fermentation using the recycled cells, was performed with good results.

Bioseparation, 2000, 9(1), 1 - 6
Anion exchange purification of plasmid DNA using expanded bed adsorption; Ferreira GN et al.; Recent developments in gene therapy with non-viral vectors and DNA vaccination have increased the demand for large amounts of pharmaceutical-grade plasmid DNA . The high viscosity of process streams is of major concern in the purification of plasmids, since it can cause high back pressures in column operations, thus limiting the throughput . In order to avoid these high back pressures, expanded bed anion exchange chromatography was evaluated as an alternative to fixed bed chromatography . A Streamline 25 column filled with 100 ml of Streamline QXL media, was equilibrated with 0.5 M NaCl in TE (10 mM Tris, 1 mM EDTA, pH = 8.0) buffer at an upward flow of 300 cmh-1, E . coli lysates (obtained from up to 3 liters of fermentation broth) were injected in the column . After washing out the unbound material, the media was allowed to sediment and the plasmid was eluted with 1 M NaCl in TE buffer at a downward flow of 120 cmh-1 . Purification factors of 36 +/- 1 fold, 26 +/- 0.4 plasmid purity, and close to 100% yields were obtained when less than one settled column volume of plasmid feed was injected . However, both recovery yield and purity abruptly decreased when larger amounts were processed-values of 35 +/- 2 and 5 +/- 0.7 were obtained for the recovery yield and purity, respectively, when 250 ml of feedstock were processed . In these cases, gel clogging and expansion collapse were observed . The processing of larger volumes, thus larger plasmid quantities, was only possible by performing an isopropanol precipitation step prior to the chromatographic step . This step led to an enhancement of the purification step.

Vet Hum Toxicol, 2000 Jun, 42(3), 133 - 6
Effects of pearl millet (Pennisetum typhoides), and fermented and processed fermented millet on Nubian goats; Abdel Gadir WS et al.; The effects of Pearl millet, fermented millet, or processed fermented millet were investigated in 15 goats assigned as untreated controls, Pearl millet-fed (166.6g/animal/d), fermented millet-fed (166.6 g/animal/d or 83.3g/animal/d), and processed fermented millet-fed (166.6g/ animal/d) . Pearl millet caused goiter within 72-108 d, and changes in thyroid follicles were correlated with clinical manifestations and alterations in serum and tissue iodine and selenium concentrations . Neither fermented nor processed fermented millet produced goiter, but their toxicity was characterized by ruminal bloat, dyspnea, diarrhea, ataxia, and enterohepatonephrotoxicity with death within 5-15 d . Organ lesions correlated with changes in serum enzymes and other serum constituents, hematology, and tissue and serum iodine and selenium concentrations.

Am J Clin Nutr, 2000 Jun, 71(6 Suppl), 1682S - 7S; discussion 1688S-90S
Prebiotics and probiotics: are they functional foods?
Roberfroid MB.
A probiotic is a viable microbial dietary supplement that beneficially affects the host through its effects in the intestinal tract . Probiotics are widely used to prepare fermented dairy products such as yogurt or freeze-dried cultures . In the future, they may also be found in fermented vegetables and meats . Several health-related effects associated with the intake of probiotics, including alleviation of lactose intolerance and immune enhancement, have been reported in human studies . Some evidence suggests a role for probiotics in reducing the risk of rotavirus-induced diarrhea and colon cancer . Prebiotics are nondigestible food ingredients that benefit the host by selectively stimulating the growth or activity of one or a limited number of bacteria in the colon . Work with prebiotics has been limited, and only studies involving the inulin-type fructans have generated sufficient data for thorough evaluation regarding their possible use as functional food ingredients . At present, claims about reduction of disease risk are only tentative and further research is needed . Among the claims are constipation relief, suppression of diarrhea, and reduction of the risks of osteoporosis, atherosclerotic cardiovascular disease associated with dyslipidemia and insulin resistance, obesity, and possibly type 2 diabetes . The combination of probiotics and prebiotics in a synbiotic has not been studied . This combination might improve the survival of the bacteria crossing the upper part of the gastrointestinal tract, thereby enhancing their effects in the large bowel . In addition, their effects might be additive or even synergistic.

Adv Drug Deliv Rev, 1997 Dec 8, 28(3), 323 - 334
Rumen-stable delivery systems; Papas A et al.; Ruminants have a distinct digestive system which serves a unique symbiotic relationship between the host animal and predominantly anaerobic rumen bacteria and protozoa . Rumen fermentation can be both beneficial by enabling utilization of cellulose and non-protein nitrogen and detrimental by reducing the nutritive value of some carbohydrates, high biological value proteins and by hydrogenating unsaturated lipids . In addition it can also result in the modification and inactivation of many pharmacologically active ingredients administered to the host animal via the oral route . The advances in ruminant nutrition and health demand a rumen-stable delivery system which can deliver the active ingredient post-ruminally while simultaneously meet efficacy, safety and cost criteria . In contrast to drug delivery systems for humans, the demand for low-cost has hindered the development of effective rumen-stable delivery systems . Historically, heat and chemical treatment of feed components, low solubility analogues or lipid-based formulations have been used to achieve some degree of rumen-stability, and products have been developed accordingly . Recently, a polymeric pH-dependent rumen-stable delivery system has been developed and commercialized . The rationale of this delivery system is based on the pH difference between ruminal and abomasal fluids . The delivery system is composed of a basic polymer, a hydrophobic substance and a pigment material . It can be applied as a coating to solid particles via a common encapsulation method such as air-suspension coating . In the future, the delivery system could be used to deliver micronutrients and pharmaceuticals post-ruminally to ruminant animals . A further possible application of the delivery system is that it could also be combined with other controlled delivery devices/systems in order to enhance slow release or to achieve targeted delivery needs for ruminants . This paper discusses the rumen protection and the abomasal release mechanism of the polymeric coating . It also reviews other rumen stable delivery systems and methods for evaluating their in vitro and in vivo performance.

Adv Drug Deliv Rev, 1997 Dec 8, 28(3), 303 - 322
Intraruminal devices; Cardinal JR; Intraruminal boluses are drug delivery devices designed to provide long term delivery of drug to the reticulo-rumen compartments of ruminant animals . This cavity is the site of the bacterial fermentation and breakdown of foodstuffs and represents an ideal site for the long term delivery of drugs and nutrients to these grazing animals . Often such animals are 'turned out' in the spring to spend much of the spring, summer and autumn seasons grazing open range . As such, the administration of drugs during the grazing season is complicated by the need to first retrieve the animals prior to drug administration . Long term delivery of drug circumvents this problem providing for decreased costs and improved efficiencies for the farmer . Numerous types of delivery devices have been developed to meet this need . This review summarizes the various approaches that have been developed, provides information on the critical design features and reviews the relevant anatomy and physiology of these animals . Some comments are provided regarding needed future developments for this class of drug delivery system.

Orig Life Evol Biosph, 2000 Feb, 30(1), 33 - 43
Sugars as the optimal biosynthetic carbon substrate of aqueous life throughout the universe; Weber AL; Our previous analysis of the energetics of metabolism showed that both the biosynthesis of amino acids and lipids from sugars, and the fermentation of organic substrates, were energetically driven by electron transfer reactions resulting in carbon redox disproportionation (Weber, 1997) . Redox disproportionation--the spontaneous (energetically favorable) direction of carbon group transformation in biosynthesis--is brought about and driven by the energetically downhill transfer of electron pairs from more oxidized carbon groups (with lower half-cell reduction potentials) to more reduced carbon groups (with higher half-cell reduction potentials) . In this report, we compare the redox and kinetic properties of carbon groups in order to evaluate the relative biosynthetic capability of organic substrates, and to identify the optimal biosubstrate . This analysis revealed that sugars (monocarbonyl alditols) are the optimal biosynthetic substrate because they contain the maximum number of biosynthetically useful high energy electrons/carbon atom while still containing a single carbonyl group needed to kinetically facilitate their conversion to useful biosynthetic intermediates . This conclusion applies to aqueous life throughout the Universe because it is based on invariant aqueous carbon chemistry--primarily, the universal reduction potentials of carbon groups.

Biomed Sci Instrum, 2000, 36, 227 - 31
Modeling and optimisation of D-fructose isomerisation using a fluorosensor; Abarna V et al.; D-Fructose was isomerized in a 1.5 litre Tokyo Rikakikai fermentor using Sweetzyme T at 40 degree Celsius at five different feed concentrations ranging from 0.01 to 1.0 M . The stirrer speed was maintained at 100 rpm . The progress of the reaction was monitored using a Dr . Ingold Fluorosensor . The fluorescence voltage was recorded as a function of time and the maximum noted . It was observed that the maximum fluorescence voltage increased as concentration increased from 0.01 to 0.04 and thereafter decreased indicating an optimum of 0.04 M . The fluorescence voltage time data fitted a first order model with an error of less than 0.8 percent.

CMAJ, 2000 May 16, 162(10), 1409 - 13
Illness outbreak associated with Escherichia coli O157:H7 in Genoa salami . E . coli O157:H7 Working Group; Williams RC et al.; BACKGROUND: An outbreak of Escherichia coli O157:H7 infection was identified in the spring of 1998, with a 7-fold increase in the number of laboratory-confirmed E . coli O157:H7 cases in southern Ontario . This prompted an intensive investigation by local, provincial and federal public health officials . METHODS: Case interviews of 25 people from southern Ontario were conducted using a broad food history and environmental exposure survey . Laboratory investigations involved both case and food sampling . Specimens of foods sold locally and reportedly consumed by those affected were tested . Common suppliers of suspected foods were identified by cross-referencing suppliers' lists with stores frequented by those who fell ill . A case-control study involving 25 cases and 49 age-matched controls was conducted . This was followed by a comprehensive environmental investigation of the meat processing plant identified as the source of the E . coli . RESULTS: Thirty-nine outbreak-related cases occurred between April 3 and June 2, 1998 . Of the 36 case specimens tested all were positive for E . coli O157:H7 . The case-control study identified Genoa salami as the most probable (odds ratio 8 {confidence interval 2-35}) source of the outbreak . Samples of Genoa salami produced by the most commonly identified supplier later tested positive for E . coli O157:H7, and the pathogen matched the same pulsed-field gel electrophoresis pattern and phage type of the case specimens . INTERPRETATION: Our investigation, which led to a national recall of the brand of dry fermented Genoa salami identified as the source of the outbreak, supports an adherence to stringent manufacturing requirements for fermented meat products . A review of the Canadian standards for fermented meat processing and the effectiveness of their implementation is warranted.

Boll Chim Farm, 2000 Jan-Feb, 139(1), 8 - 13
{Cleaning validation of the prefermentation-fermentation group for production of biomedical products}; Lencioni E et al.; The fermentation-group is largely employed to produce pharmaceuticals by means of biotechnology techniques . The operative functions of these plants are surely kept under control by the validation process, while its cleaning is less investigated . The aim of this work is the cleaning validation of a fermentation-group . Furthermore, Standard Operative Procedures (SOPs) have been validated relating to the production necessities and to the allowed safety limits.

J Nutr, 2000 Jun, 130(6), 1572 - 7
Chronic consumption of short-chain fructooligosaccharides does not affect basal hepatic glucose production or insulin resistance in type 2 diabetics; Luo J et al.; Short-chain fructooligosaccharides (FOS) are prebiotics, which escape digestion in the small intestine and are fermented by the colonic microflora into short-chain fatty acids . Recently, we found that the daily consumption of 20 g FOS decreased basal hepatic glucose production in healthy subjects without any effect on insulin-stimulated glucose metabolism . In this study, we evaluated the effects of the chronic ingestion of FOS on plasma lipid and glucose concentrations, hepatic glucose production and insulin resistance in type 2 diabetics . Type 2 diabetic volunteers (n = 10; 6 men, 4 women) received either 20 g/d FOS or sucrose for 4 wk in a double-blind crossover design . FOS did not modify fasting plasma glucose and insulin concentrations or basal hepatic glucose production . The plasma glucose response to a fixed exogenous insulin bolus did not differ at the end of the two periods . Erythrocyte insulin binding also did not differ . Serum triacylglycerol, total and HDL cholesterol, free fatty acid, apolipoproteins A1 and B and lipoprotein (a) concentrations were not modified by the chronic ingestion of FOS . We conclude that 4 wk of 20 g/d of FOS had no effect on glucose and lipid metabolism in type 2 diabetics.

Acta Biochim Pol, 1999, 46(4), 991 - 1000
The access of metabolites into yeast mitochondria in the presence and absence of the voltage dependent anion selective channel (YVDAC1); Kmita H et al.; Since yeast Saccharomyces cerevisiae mutants depleted of the voltage dependent anion selective channel (YVDAC1) are still able to grow on a non-fermentable carbon source, a functional transport system in the outer mitochondrial membrane must exist to support the access of metabolites into mitochondria . It was assumed that the properties of the system could be inferred from the differences in the results observed between wild type and mutant mitochondria since no crucial differences in this respect between the two types of mitoplasts were observed . YVDAC1-depleted mitochondria displayed a highly reduced permeability of the outer membrane, which was reflected in increased values of K0.5(NADH) for respiration and K0.5(ADP) for triggering phosphorylating state as well as in delayed action of carboxyatractylate (CATR) in inhibition of phosphorylating state . The parameters were chosen to express the accessibility of the applied species to the intermembrane space . The passage of the molecules through the outer membrane depleted of YVDAC1 could be partially improved in the presence of bivalent cations (Mg2+, Ca2+), as in their presence lower values of the calculated parameters were obtained . The restrictions imposed on the transport of molecules through the YVDAC1-depleted outer membrane resulted in a competition between them for the access to the intermembrane space as measured by changes in parameters observed for a given species in the presence of another one . The competition was stronger in the absence of Mg2+ and depended on charge and size of transported molecules, as the strongest competitor was CATR and the weakest one--NADH . Thus, it can be concluded that the transport system functioning in the absence of YVDAC1 is modulated by bivalent cations and charge as well as size of transported molecules . Since an increased level of respiration due to the dissipation of delta psi causes an increase of K0.5(NADH) in both wild type and YVDAC1-depleted mitochondria it is concluded that a common property of YVDAC1 and the system functioning in YVDAC1-depleted mitochondria seems to be the dependence of the capacity on the level of mitochondrial respiration.

J Antimicrob Chemother, 2000 Apr, 45 Suppl 1, 29 - 33
Comparative potency of gemifloxacin, new quinolones, macrolides, tetracycline and clindamycin against Mycoplasma spp; Duffy LB et al.; This study determined the comparative in vitro potency of the new investigative quinolone gemifloxacin (SB-265805) using low-passaged clinical isolates and type strains of mycoplasma commonly found in the human respiratory and urogenital tracts . Organisms studied were Mycoplasma pneumoniae, Mycoplasma hominis, Mycoplasma fermentans, Mycoplasma genitalium, Mycoplasma penetrans and Ureaplasma urealyticum, obtained from different geographical regions . Comparator drugs were levofloxacin, trovafloxacin, grepafloxacin, azithromycin, clarithromycin, tetracycline and clindamycin . MICs were determined using a microbroth dilution method . The overall range of MICs of gemifloxacin was < or =0.008-0.125 mg/L for different Mycoplasma spp . and < or =0.008-0.5 mg/L for Ureaplasma spp . Depending on the species tested, gemifloxacin showed variable results when compared with the macrolides . However, gemifloxacin was as potent as, or more potent than, tetracycline, clindamycin and the other quinolones investigated.

J Dairy Sci, 2000 May, 83(5), 1042 - 51
Milk production during the complete lactation of dairy cows fed diets containing different amounts of protein; Wu Z et al.; Milk production response to four different amounts of protein supplementation was measured in a complete lactation study utilizing 58 multiparous Holstein cows treated with bovine somatotropin . The four treatments were as follows (the first number is the dietary crude protein content (% of dry matter) during the first 16 wk of lactation and the second number is the protein content for wk 17 to 44 of lactation): 15.4-16.0, 17.4-16.0, 17.4-17.9, and 19.3-17.9 . Diets were formulated to maximize the supply of amino acids to the intestine . High moisture ear corn was finely ground to promote ruminal fermentation and microbial protein synthesis, and roasted soybeans and expeller processed soybean meal were used as the protein supplements to provide relatively high amounts of rumen-undegradable protein . Alfalfa silage and corn silage (3:2) provided the forage . Milk production for the 308-d lactation for each of the treatment groups was 10,056, 10,831, 11,095, and 11,132 kg . Cows of this production level fed diets similar to those used in this experiment benefit from dietary protein of approximately 17.5% during the first 30 wk of lactation . A reduction in dietary protein to 16% can be made around wk 30 of lactation . This amount of dietary protein should, with prevailing feed prices, be compatible with maximum profit and a moderate amount of nitrogen excretion to the environment.

J Dairy Sci, 2000 May, 83(5), 984 - 95
Duration of regrowth of ryegrass (Lolium perenne) effects on grazing behavior, intake, rumen fill, and fermentation of lactating dairy cows; Chilibroste P et al.; The relative importance of duration of sward regrowth and fill and fermentation in the rumen on the control of grazing time and intake rate during the first grazing session of the day was studied . Four lactating dairy cows were allowed to graze ryegrass (Lolium perenne) swards, with five different regrowth periods after mowing (6, 9, 16, 22, and 30 d) . The cows were allowed to graze until they stopped voluntarily (cessation of grazing activity for at least 15 min) . Before and after grazing the rumen contents were evacuated, weighed, sampled, and returned to the animals . Samples of rumen liquid were taken immediately before rumen evacuation and approximately 30, 60, 120, and 240 min after the grazing session was finished . Grazing time and intake rate did not follow a significant trend with period of regrowth . Bite rate did not change significantly with duration of regrowth with cows exhibiting high rates of biting for all the sward conditions . Rumen pools sizes of dry matter, neutral detergent fiber, and volatile fatty acids (VFA), measured after grazing, increased significantly with days of sward regrowth, even though the changes over days were small . Concentration of VFA followed a significant quadratic trend with a maximum concentration observed at approximately 110 min after cessation of grazing . In this study, rumen fill, VFA (either total or major components), ammonia, pH, and osmotic pressure as individual variables were not correlated with grazing time or dry matter intake.

Br Poult Sci, 2000 Mar, 41(1), 75 - 82
Effect of food enzymes on utilisation of lupin carbohydrates by broilers; Kocher A et al.; 1 . The effects of 3 commercial enzyme products on the nutritive value of 2 lupin species were investigated with the emphasis on changes in composition of non-starch polysaccharides (NSPs) along the digestive tract . Enzyme A contained primarily cellulase, beta-glucanase and xylanase activities, enzyme B primarily hemicellulase, pentosanase and xylanase activities, and enzyme C primarily hemicellulase, pectinase and beta-glucanase activities . 2 . The enzymes were added to semi-purified diets based on sorghum and casein containing 35% whole seed lupins (Lupinus angustifolius cv Gungurru or Lupinus albus cv Kiev mutant) . Control diets contained no lupins . 3 . Food conversion ratio (FCR), excreta moisture content and apparent metabolisable energy (AME) were affected by lupin species but not by enzyme supplementation . 4 . In diets with L . angustifolius, enzyme C significantly increased digesta viscosity and increased the concentration of soluble NSPs in all sections of the intestine . 5 . Digestibility of protein and NSPs in the ileum and microbial fermentation in the ileum and caeca were not affected by adding enzymes to diets containing L . angustifolius . 6 . Enzyme addition to diets with L . albus did not affect digesta viscosity nor concentration of soluble NSPs but caused a significantly (P<0.05) reduced concentration of insoluble NSP in the ileum . 7 . Enzyme addition to L . albus significantly (P<0.05) increased NSP digestibility in the ileum but had no effects on protein digestibility and fermentation in the ileum and caeca.

Mol Cell Biochem, 2000 Feb, 205(1-2), 149 - 61
Butyrate inhibits proliferation-induced proliferating cell nuclear antigen expression (PCNA) in rat vascular smooth muscle cells; Ranganna K et al.; Arterial injury-induced vascular smooth muscle cell (VSMC) proliferation in intima is the important etiologic factor in vascular proliferative disorders such as atherosclerosis, hypertension and restenosis after balloon angioplasty . Butyrate, a naturally occurring short chain fatty acid, is produced by bacterial fermentation of dietary fiber and by mammary glands of certain mammals . Studies have shown that butyrate at millimolar concentrations, which are physiological, induces growth arrest, differentiation and apoptosis . We examined the effect of physiological concentrations of butyrate on rat VSMC proliferation and proliferation-induced PCNA expression to determine anti-atherogenic potential of butyrate . Butyrate concentrations, closer to physiological range, exhibited antiproliferative effects on both serum-induced proliferation of serum-starved quiescent VSMCs and actively proliferating non-confluent VSMCs . Treatment of serum-starved quiescent VSMCs with 1-8 mmol/l concentration of butyrate caused a concentration-dependent decrease in serum-induced VSMC proliferation and cell proliferation-associated increase in total cellular proteins and RNA levels . Similarly, exposure of actively growing VSMCs to 5 mmol/l butyrate resulted in the inhibition of cell proliferation and proliferation-induced increase in cellular proteins and RNA levels . Furthermore, cellular morphology was significantly altered . Analysis of cell cycle regulatory proteins indicated that levels of PCNA, an excellent marker for cell proliferation, was significantly altered by butyrate both in actively proliferating and serum-induced quiescent VSMCs . These observations suggest that butyrate exhibits potential antiatherogenic capability by inhibiting VSMC proliferation and proliferation-associated increase in PCNA expression and thus merits further investigations regarding therapeutic significance of butyrate in vascular proliferative disorders.

Mol Gen Genet, 2000 Apr, 263(3), 411 - 22
Protein phosphatase type-1 regulatory subunits Reg1p and Reg2p act as signal transducers in the glucose-induced inactivation of maltose permease in Saccharomyces cerevisiae; Jiang H et al.; The REG1 gene encodes a regulatory subunit of the type-1 protein phosphatase (PP1) G1c7 in Saccharomyces cerevisiae, which directs the catalytic subunit to substrates involved in glucose repression . Loss of REG1 relieves glucose repression of many genes, including the MAL structural genes that encode the maltose fermentation enzymes . In this report, we explore the role of Reglp and its homolog Reg2p in glucose-induced inactivation of maltose permease . Glucose stimulates the proteolysis of maltose permease and very rapid loss of maltose transport activity - more rapid than can be explained by loss of the permease protein alone . In a reg1delta strain we observe a significantly reduced rate of glucose-induced proteolysis of maltose permease, and the rapid loss of maltose transport activity does not occur . Instead, surprisingly, the slow rate of proteolysis of maltose permease is accompanied by an increase in maltose transport activity . Loss of Reg2p modestly reduces the rates of both glucose-induced proteolysis of maltose permease and inactivation of maltose transport activity . Overexpression of Reg2p in a reg1delta strain suppresses the effect on maltose permease proteolysis and partially restores the inactivation of maltose transport activity, but does not affect the insensitivity of MAL gene expression to repression by glucose observed in this strain . Thus, protein phosphatase type-1 (Glc7p-Reglp and Glc7p-Reg2p) plays a role in transduction of the glucose signal during glucose-induced proteolysis of maltose permease, but only Glc7p-Reglp is involved in glucose-induced inactivation of maltose transport activity and glucose repression of MAL gene expression . Overexpression of REG1 partially restores proteolysis of maltose permease in a grr1delta strain, which lacks glucose signaling, but does not rescue rapid inactivation of maltose transport activity or sensitivity to glucose repression . A model for the role of Reglp and Reg2p in glucose signaling pathways is discussed . We also uncovered a previously unrecognized G2/M delay in the grr1delta but not the reg1delta strains, and this delay is suppressed by REG1 overexpression . The G1/S delay seen in grr1delta mutants is slightly suppressed as well, but REG1 overexpression does not suppress other grr1delta phenotypes such as insensitivity to glucose repression.

Biotechnol Bioeng, 2000 Jul 5, 69(1), 83 - 90
Aqueous two-phase systems containing urea: influence of protein structure on protein partitioning; Ramsch C et al.; During recombinant E . coli fermentation with high-expression levels inclusion bodies are often formed . Aqueous two-phase systems have been successfully used in the presence of urea for the initial recovery step of inclusion bodies from E . coli . Basic studies of the complex interactions are lacking . For a systematic study of protein partitioning in the presence of urea we selected T4-lysozyme mutants with different thermal stability as a model . The stabilization of these variants by phase components was investigated measuring the fluorescence emission of tryptophan residues in the protein . Protein structure was stabilized at pH 7 in the order of S0(4)(2-) >> PEG = Dextran > H(2)O . The conformation of proteins was shown to have a strong influence on the partitioning in aqueous two-phase systems . Tryptophan and its homologuous di- and tripeptdides were partitioned in similar phase systems to normalize for contribution from hydrophobic interactions .

Biotechnol Bioeng, 2000 Jul 5, 69(1), 66 - 73
Reactive extraction of penicillin G in hollow-fiber and hollow-fiber fabric modules; Yang C et al.; Penicillin G (Pen G) can be rapidly extracted in hollow-fiber liquid-liquid contactors using N-lauryl-N-trialkylmethylamine (Amberlite LA-2) as the extractant . n-Butylacetate is much better than decanol as a diluent for such an extraction, although decanol can give a partition coefficient four times larger . The overall mass transfer coefficient found is a function of aqueous flow on the lumen side of the fiber, and is less dependent on shell-side flow . In backextraction, the overall mass transfer coefficient determined is only one tenth that of the forward extraction, primarily because the hydrophobic hollow fibers used have a high mass transfer resistance under these conditions . The mass transfer experiments show that hollow-fiber extraction of Pen G is competitive with centrifugal extraction . The prospects for extraction of other fermentation products with hollow fibers can be estimated based on the present study .

J Agric Food Chem, 2000 May, 48(5), 1971 - 6
Optimization of simultaneous saccharification and fermentation for the production of ethanol from lignocellulosic biomass; Hari Krishna S et al.; Simultaneous saccharification and fermentation (SSF) of alkaline hydrogen peroxide pretreated Antigonum leptopus (Linn) leaves to ethanol was optimized using cellulase from Trichoderma reesei QM-9414 (Celluclast from Novo) and Saccharomyces cerevisiae NRRL-Y-132 cells . Response surface methodology (RSM) and a three-level four-variable design were employed to evaluate the effects of SSF process variables such as cellulase concentration (20-100 FPU/g of substrate), substrate concentration (5-15% w/v), incubation time (24-72 h), and temperature (35-45 degrees C) on ethanol production efficiency . Cellulase and substrate concentrations were found to be the most significant variables . The optimum conditions arrived at are as follows: cellulase = 100 FPU/g of substrate, substrate = 15% (w/v), incubation time = 57.2 h, and temperature = 38.5 degrees C . At these conditions, the predicted ethanol yield was 3.02% (w/v) and the actual experimental value was 3.0% (w/v).

J Agric Food Chem, 2000 May, 48(5), 1892 - 7
Development and testing of seven new synthetic coyote attractants; Kimball BA et al.; Available evidence indicates that effective coyote attractants are blends of volatile substances . Typically, attractants are a combination of biological substances such as fermented glandular materials, urines, and rotted meats . Although effective, these attractants have several distinct disadvantages . Among these is the possibility that they are unnecessarily complex and variable and, thus, difficult to replicate from one batch to the next . Although attractants containing a few reagent grade materials are available, the chemicals selected and their concentrations are not derived from actual attractants . For this reason, commercially available coyote attractants were analyzed with the intention of developing relatively simple synthetic alternatives . Purge and trap headspace analysis with gas chromatography/mass selective detection was employed to identify the volatile components of known conventional and synthetic attractants . All identified compounds were grouped according to chemical functionality, and one compound from each functional group was chosen to represent the group . Using only these representative compounds, seven synthetic attractants were formulated . Bioassays with captive coyotes (Canis latrans) were conducted to compare behavioral responses elicited by the seven new attractants, a currently available synthetic attractant, and a control . The results indicated that the attractants elicited significantly different behavioral profiles.

J Agric Food Chem, 2000 May, 48(5), 1865 - 71
Influence of halogen salts on the production of the ochratoxins by Aspergillus ochraceus Wilh; Stander MA et al.; The first report of the biological production of bromo ochratoxin B by Aspergillus ochraceus Wilh . is presented as well as a study of the influence of potassium bromide, potassium iodide, potassium fluoride, and potassium chloride on the production of ochratoxin A and ochratoxin B . Potassium fluoride and potassium iodide inhibited the growth of the fungus, whereas potassium chloride substantially stimulated the production of ochratoxin A in shaken solid substrate fermentation on whole wheat or shredded wheat, generally giving a high yield of ochratoxins . Increasing levels of potassium bromide led to a decline in ochratoxin A production and an increase in bromo-ochratoxin B, ochratoxin B, and 4-hydroxy ochratoxin B . Nevertheless, A . ochraceus was much less versatile in the bromo analogues than other fungi, which produce metabolites containing chlorine . Analysis included aminopropyl solid-phase extraction column cleanup followed by quantitative analysis on reversed-phase HPLC using fluorescence detection and employing N-(5-chloro-2-hydroxybenzoyl)phenylalanine as an internal standard.

J Agric Food Chem, 2000 May, 48(5), 1803 - 9
Volatile flavor components in red fermented soybean (Glycine max) curds; Chung HY; Red fermented soybean (Glycine max) curds (FSC) were extracted with a simultaneous steam distillation and extraction (SDE) apparatus and analyzed by gas chromatography-mass spectrometry (GC-MS) and further evaluated by gas chromatography-mass spectrometry-flame ionization detector-olfactometry (GC-MS-FID-O) . A combined total of 89 compounds were identified from three samples (A-C) . The numbers of components identified (samples A-C) were 87, 85, and 81, respectively, which were divided into 10 compound classes . The quantity of 48 compounds was statistically significant (p < 0.05) . Sample B generally had the lowest level, whereas sample C had the higher level of each common component among the samples . GC-MS-FID-O analyses showed that the majority of the fruity, diacetyl, cantaloupe-like odors were eluted early during the run (RI < 1100), followed by the meaty flavor, rosy, and prune-like aromas (RI > 1100) . Ethyl 2-methylpropanoate, 2,3-butanedione, ethyl butanoate, ethyl 2-methylbutanoate, 3-(methylthio)propanal, benzeneacetaldehyde, and ethyl 3-phenylpropionate were found to be important common components contributing to the characteristic aroma of red FSCs.

J Agric Food Chem, 2000 May, 48(5), 1789 - 98
Influence of a Saccharomyces cerevisiae selected strain in the volatile composition of rosé wines . Evolution during fermentation; Fraile P et al.; There has been considerable controversy about the use of selected pure strains in wine fermentation . For that reason it is important to determine the influence of this vinification technique in the composition of wine because it arises from the type of yeast and the subsequent evolution during fermentation . This study researches the volatile composition of rose wines from the Garnacha must, inoculated with one selected NA33 strain of Saccharomyces cerevisiae . The inoculated yeast did not predominate in all of the samples . These samples showed a behavior intermediate between those of the control and samples in which NA33 did predominate . The greatest concentration of higher alcohols was in the control wine, and its evolution was similar in all fermentations . The esters formed at the end of the fermentation and their concentrations were higher in the control than in the inoculated samples . In the control, acids were produced above all, in the first half of fermentation, and decreased from then onward . In the sample in which the yeast predominated, the synthesis occurred later and to a lesser extent than in the control.

J Agric Food Chem, 2000 May, 48(5), 1727 - 33
Fermentation of a bacterial cellulose/xylan composite by mixed ruminal microflora: implications for the role of polysaccharide matrix interactions in plant cell wall biodegradability; Weimer PJ et al.; Growth of the cellulose-synthesizing bacterium Acetobacter xylinum ATCC 53524 in media supplemented with 5% (w/v) glucose and 0.2% (w/v) of a water-soluble, nearly linear xylan from tobacco stalks resulted in the synthesis of a highly crystalline composite having a xylose/glucose ratio ranging from 0.06 to 0.24 . The digestion of one composite (88% cellulose/12% xylan) by mixed ruminal microflora displayed kinetics of gas production similar to those of an unassociated mixture of the two components added in a xylan/cellulose ratio similar to that of the composite . The data suggest that intimate association of xylan and cellulose, as is typically found in secondary plant cell walls, does not inhibit the rate of digestion of the component polysaccharides.

J Antibiot (Tokyo), 2000 Mar, 53(3), 256 - 61
Biological activity of guanacastepene, a novel diterpenoid antibiotic produced by an unidentified fungus CR115; Singh MP et al.; Fermentation extracts of culture CR115, an unknown plant endophyte originally isolated from Costa Rica, were found to be active against antibiotic-resistant bacteria . The metabolite responsible for activity was identified as a novel diterpenoid antibiotic guanacastepene (mol . wt . 374.47 and mol . formula C22H30O5) . Mechanistic studies done in an E . coli imp strain suggested membrane damage as the primary mode of bactericidal action . This compound also lysed human RBCs and caused leakage of intracellular potassium from E . coli imp.

J Antibiot (Tokyo), 2000 Mar, 53(3), 227 - 32
Arisostatins A and B, new members of tetrocarcin class of antibiotics from Micromonospora sp . TP-A0316 . I . Taxonomy, fermentation, isolation and biological properties; Furumai T et al.; Arisostatins A and B, new members of tetrocarcin class of antibiotics were isolated from the culture broth of an actinomycete strain . The producing strain, TP-A0316, was identified as Micromonospora sp . Arisostatins were obtained from the culture fluid by solvent extraction and chromatographic purification . They showed antibiotic activity against Gram-positive bacteria and antitumor activity.

Small Rumin, Res. . 2000 Jul 1, 37(1-2), 115 - 124
Comparison of mineral and cholesterol composition of different commercial goat milk products manufactured in USA; Park YW; Concentrations of 12 major and trace minerals and cholesterol in commercial goat fluid milk, evaporated, powdered, yogurt, and cheese products manufactured in the US were evaluated for compositional differences . Minerals were determined by an Inductively Coupled Argon Plasma Emission Spectroscopy (ICAP), while cholesterol was analyzed using colorimetric and gas chromatographic (GC) methods . Mean total solids content (%) of fluid milk, evaporated milk, powdered milk, yogurt, plain soft and Monterey Jack cheeses were: 11.3, 20.9, 94.1, 11.5, 32.5, and 57.7, respectively . Mean calcium and phosphorus contents (ppm, dry basis) of the corresponding products were: 103, 125; 440, 393; 7715, 7471; 161, 144; 691, 1105; 3492, 3067, respectively . The respective iron and zinc contents (ppm) of the corresponding products were: 0.062, 0.349; 1.518, 1.635; 3.33, 30.21, 0.117, 0.338; 7.16, 3.64; 8.86, 3.81 . The levels of potassium (K) in cheeses were lowest among all the products including fluid goat milk, suggesting that a significant amount of K was lost during cheese manufacturing processes . Levels of all trace minerals were higher in yogurt and cheeses than in fluid milk . The levels of trace minerals in cheeses were greater than those in yogurt products . Iron and aluminum contents of certain goat milk products were considerably higher than normal, possibly due to contamination of minerals from manufacturing utensils and product metal cans . Sulphur contents of fermented products were significantly greater than those of fluid milk, which may be accountable for the microbial synthesis of sulfur containing proteins during the manufacturing processes of the products . Cholesterol contents (mg/100g, wet basis) of fluid, evaporated, powdered goat milk and Monterey Jack cheese determined by GC method were: 11.0, 24.9, 119.5 and 91.7, respectively . Cholesterol contents of the goat milk products analyzed by colorimetric method were substantially greater than those by GC method.

Z Naturforsch {C}, 2000 Mar-Apr, 55(3-4), 213 - 7
Use of immobilized Candida cells on xylitol production from sugarcane bagasse; de Carvalho W et al.; In this study we used the yeast Candida guilliermondii FTI 20037 immobilized by entrapment in Ca-alginate beads (2.5-3 mm diameter) for xylitol production from concentrated sugarcane bagasse hemicellulosic hydrolysate in a repeated batch system . The fermentation runs were carried out in 125- and 250-ml Erlenmeyer flasks placed in an orbital shaker at 30 degrees C and 200 rpm during 72 h, keeping constant the proportion between work volume and flask total volume . According to the results, cell viability was substantially high (98%) in all fermentative cycles . The values of parameters xylitol yield and volumetric productivity increased significantly with the reutilization of the immobilized biocatalysts . The highest values of xylitol final concentration (11.05 g/l), yield factor (0.47 g/g) and volumetric productivity (0.22 g/lh) were obtained in 250-ml Erlenmeyer flasks containing 80 ml of medium plus 20 ml of immobilized biocatalysts . The support used in this study (Ca-alginate) presented stability in the experimental conditions used . The results show that the use of immobilized cells is a promising approach for increasing the xylitol production rates.

FEMS Microbiol Ecol, 2000 Apr 1, 32(2), 121 - 128
Mathematical estimations of hyper-ammonia producing ruminal bacteria and evidence for bacterial antagonism that decreases ruminal ammonia production(1); Rychlik JL et al.; Mixed ruminal bacteria (MRB) from cattle fed hay produced ammonia from protein hydrolysate twice as fast as MRB from cattle fed mostly grain, and a mathematical model indicated that cattle fed hay had approximately four-fold more hyper ammonia-producing ruminal bacteria (HAB) . HAB had a high maximum velocity of ammonia production (V(max)) and low substrate affinity (high K(m)), but simulations indicated that only large changes in V(max) or K(m) would cause a large deviation in HAB numbers . Some carbohydrate-fermenting ruminal bacteria produced ammonia at a slow rate (CB-LA), but many of the isolates had almost no activity (CB-NA) . The model indicated that the ratio of CB-LA to CB-NA had little impact on HAB numbers . Validations based on predicted ratios of HAB, CB-LA and CB-NA over-predicted the specific activity of ammonia production by MRB, but co-culture incubations indicated that washed MRB from cattle fed grain could inhibit HAB . Because autoclaved MRB had virtually no effect on HAB and the incubations were always carried out at pH 7.0, the inhibition was not simply a chemical effect (e.g . low pH).

Biochem, Eng . J. . 2000 Jun 1, 5(2), 129 - 136
Comparison of biotin production by recombinant Sphingomonas sp . under various agitation conditions; Saito I I et al.; Biotin production by fermentation of recombinant Sphingomonas sp./pSP304 was investigated . A complex medium containing 60g/l of glycerol and 30g/l of yeast extract was suitable for biotin production . Biotin was produced in the late logarithmic or stationary phase after glycerol starvation . The optimum pH value for biotin production was 7.0 . When the dissolved oxygen concentration (DO) was controlled at a constant level, the biotin concentration produced after 120h was significantly lower than that obtained in a test tube culture . Therefore, a batchwise jar-fermentor culture with a constant agitation speed and without DO control was conducted for investigating the effect of agitation conditions on biotin production . Six types of impeller were tested: turbine-blade type, turbo-lift type, rotating mesh type (EGSTAR((R))), screw with draft tube type, Maxblend((R))type, and anchor type . With some impellers, agitation speed was also changed . Both the maximum cell concentration and biotin production varied depending on agitation conditions . Relatively high cell concentrations were attained with four of the impeller types, turbine-blade type, rotating mesh type, Maxblend((R)) type, and anchor type . Among these impellers, the turbine-blade impeller with sintered sparger was suitable for biotin production . After 120h, the cell concentration reached an OD(660) of 43 and a biotin concentration of 66mg/l was obtained, which was comparable with the results from the test tube culture . Morphological variation was also observed depending on the agitation conditions: oval-shaped, rod-shaped, and elongated-shaped cells . Biotin production was relatively high in slightly long rod-shape cells but low in elongated cells . The difference in morphology appeared to depend on the shear stress . It was found that biotin production was strongly correlated with cell length and the oxygen transfer coefficient (k(L)a); cell lengths in the range 4-7microm and k(L)a values in the range 1.5-2.0/min were found to be suitable for biotin production in jar-fermentor culture.

Biochem, Eng . J. . 2000 Jun 1, 5(2), 109 - 114
Agitation induced mycelial fragmentation of Aspergillus oryzae and Penicillium chrysogenum; Amanullah A et al.; Given the impact of mycelial morphology on fermentation performance, it is important to understand the factors that influence it, including agitation-induced fragmentation . The successful application of the energy dissipation/circulation function (EDC) to correlate fragmentation of Penicillium chrysogenum with agitation intensity and with different impeller types {5} has already been demonstrated . The EDC function takes into account the specific energy dissipation rate in the impeller swept volume and the frequency of mycelial circulation through that volume . In order to explore whether the EDC function can be used more generally to correlate fragmentation of different filamentous species, the present study extended the concept to agitation-induced, off-line fragmentation of Aspergillus oryzae grown in chemostat culture . The work shows that at EDC values off-line greater than that in the chemostat, fragmentation with different impellers can be correlated with the EDC . For EDC values less than those used in the chemostat, fragmentation did not occur . The earlier results of Justen et al . {5} with Penicillium chrysogenum are also reconsidered and found to behave similarly.

Arch Microbiol, 2000 Apr, 173(4), 269 - 77
Rhodoferax antarcticus sp . nov., a moderately psychrophilic purple nonsulfur bacterium isolated from an Antarctic microbial mat; Madigan MT et al.; A new species of purple nonsulfur bacteria isolated from an Antarctic microbial mat is described . The organism, designated strain ANT.BR, was mildly psychrophilic, growing optimally at 15-18 degrees C with a growth temperature range of 0-25 degrees C . Cells of strain ANT.BR were highly motile curved rods and spirals, contained bacteriochlorophyll a, and showed a multicomponent in vivo absorption spectrum . A specific phylogenetic relationship was observed between strain ANT.BR and the purple bacterium Rhodoferax fermentans FR2T, and the two organisms shared several physiological and other phenotypic properties, with the notable exception of growth temperature optimum . Tests of genomic DNA hybridization, however, showed Rfx . fermentans FR2T and strain ANT.BR to be genetically distinct bacteria . Because of its unique set of properties, especially its requirement for low growth temperatures, we propose to recognize strain ANT.BR as a new species of the genus Rhodoferax, Rhodoferax antarcticus, named for its known habitat, the Antarctic.

J Org Chem, 2000 Feb 25, 65(4), 990 - 5
Structures of TMC-95A-D: novel proteasome inhibitors from Apiospora montagnei sacc . TC 1093; Kohno J et al.; Four novel proteasome inhibitors, TMC-95A-D (1-4) have been isolated from the fermentation broth of Apiospora montagnei Sacc . TC 1093, isolated from a soil sample . All of the molecular formulas of 1-4 were established as C(33)H(38)N(6)O(10) by high-resolution FAB-MS . Their planar structures were determined on the basis of extensive analyses of 1D and 2D NMR, and degradation studies . Compounds 1-4 have the same planar structures to each other, and are unique highly modified cyclic peptides containing L-tyrosine, L-aspargine, highly oxidized L-tryptophan, (Z)-1-propenylamine, and 3-methyl-2-oxopentanoic acid units . The absolute configuration at C-11 and C-36 of 1-4 was determined based on chiral TLC and HPLC analyses of their chemical degradation products . The ROESY analysis along with (1)H-(1)H coupling constants clarified the absolute stereochemistry at C-6, -7, -8, and -14 of the cyclic moieties . These studies revealed the relationships of 1-4 to be diastereomers at C-7 and C-36.

J Chromatogr Sci, 2000 May, 38(5), 207 - 10
Elimination of amino acid interferences in the chiral ligand-exchange chromatographic analysis of lactic acid enantiomers in wine; Buglass AJ et al.; Chiral ligand-exchange liquid chromatography is used to identify and quantitate lactic acid enantiomers in wines that have or have not undergone malolactic fermentation . The stationary phase is (R)-penicillamine, which is bound lipophilically to a C18 bonded silica matrix . The mobile phase is 1mM copper sulfate, and the detection mode is ultraviolet . Serious interference from (S)-aspartic acid and other amino acids is eliminated by the use of propanesulfonic acid-type cation exchange solid-phase extraction cartridges prior to chromatographic analysis . Lactic acid enantiomers in wine are quantitated in the range of 10 to 500 mg/L . The detection limit is 3 mg/L . The method is also successful in the determination of lactic acid enantiomers in certain beers (e.g., lambic beers), kim-chi, sauerkraut, and various yogurts.

J Am Diet Assoc, 2000 May, 100(5), 524 - 8; quiz 529-30
Improved lactose digestion and intolerance among African-American adolescent girls fed a dairy-rich diet; Pribila BA et al.; OBJECTIVE: To determine whether African-American adolescent girls who were fed a dairy-rich diet for 21 days could adapt to lactose, experiencing an overall improvement in lactose tolerance as well as a decrease in hydrogen gas production . DESIGN: Twenty-one-day dietary intervention study . SUBJECTS/SETTING: Seventeen of 21 African-American girls (aged 11 to 15 years) enrolled in a calcium metabolism study chose to participate in the lactose tolerance study . Subjects were screened for any diseases, conditions, or medications that might alter calcium metabolism or colonic fermentation . Subjects were housed in a fraternity on the Purdue University, West Lafayette, Ind, campus, and were supervised 24 hours a day . INTERVENTION: Subjects consumed a dairy-based diet averaging 1,200 mg calcium and 33 g lactose per day for 21 days . Lactose digestion was assessed by an 8-hour breath hydrogen test on days 1 and 21, and symptoms of intolerance (abdominal pain, bloating, flatulence, and diarrhea) were evaluated hourly on a ranked scale during the breath hydrogen tests and once each evening during the 21-day feeding period . MAIN OUTCOME MEASURES: A comparison of breath hydrogen production and gastrointestinal symptoms at the beginning and end of the study . STATISTICAL ANALYSES PERFORMED: The Wilcoxon signed ranks test was used to compare the area under the curve for the 2 breath hydrogen tests . Spearman's p test for trend was used to determine whether there was a change in symptoms . All statistical analyses were 2-tailed and significance was set at P = .05 . RESULTS: Fourteen of the 17 subjects had lactose maldigestion . Breath hydrogen excretion decreased significantly (P < .03) from the beginning (148.3 +/- 27.0 ppm x hours) to the end (100.7 +/- 19.3 ppm x hours) of the 21-day period . Gastrointestinal symptoms were negligible during both the breath hydrogen tests as were symptoms during the 21-day period . APPLICATIONS/CONCLUSIONS: The diet was well tolerated by the subjects . Furthermore, the decrease in breath hydrogen suggests colonic adaptation to the high-lactose diet . The results indicate that lactose maldigestion should not be a restricting factor in developing adequate calcium diets for this population . The existence of lactose maldigestion does not result in lactose intolerance in this population when it is fed a dairy-rich diet.

EMBO J, 2000 May 15, 19(10), 2161 - 7
Ubc8p functions in catabolite degradation of fructose-1, 6-bisphosphatase in yeast; Schule T et al.; The key gluconeogenic enzyme fructose-1,6-bisphosphatase (FBPase) is synthesized when cells of the yeast Saccharomyces cerevisiae are grown on a non-fermentable carbon source . After shifting the cells to glucose-containing medium, in a process called catabolite degradation, FBPase is selectively and rapidly broken down . We have isolated gid mutants, which are defective in this glucose-induced degradation process . When complementing the defect in catabolite degradation of FBPase in gid3-1 mutant cells with a yeast genomic library, we identified the GID3 gene and found it to be identical to UBC8 encoding the ubiquitin-conjugating enzyme Ubc8p . The in vivo function of Ubc8p (Gid3p) has remained a mystery so far . Here we demonstrate the involvement of Ubc8p in the glucose-induced ubiquitylation of FBPase as a prerequisite for catabolite degradation of the enzyme via the proteasome . Like FBPase, Ubc8p is found in the cytoplasmic fraction of the cell . We demonstrate cytoplasmic degradation of FBPase.

J Commun Dis, 1998 Dec, 30(4), 263 - 5
Colicinogenicity and enteropathogenicity of Escherichia coli associated with gastroenteritis in man; Chakraborty D et al.; Sixty strains of Escherichia coli, were isolated and identified from 94 samples of infantile diarrhoea and traveller's diarrhoea . Eleven strains were found to be colicinogenic and colicin 'H' was found to be predominant in conjunction with other colicins . Five strains were enteropathogenic of which 055 and one untypable strain were highly enteropathogenic . A positive correlation could be established between haemolytic character, dulcitol fermentation and the virulence of the strain involved.

Yeast, 2000 May, 16(7), 641 - 50
The phosphatidylcholine to phosphatidylethanolamine ratio of Saccharomyces cerevisiae varies with the growth phase; Janssen MJ et al.; This study compares the effect of the growth phase on the phospholipid composition and the activity of several phospholipid biosynthetic enzymes in a wild-type yeast grown in fermentable (glucose) and non-fermentable (lactate) semi-synthetic and complete synthetic media . Several distinct differences as well as similarities were found . The cellular phosphatidylcholine: phosphatidylethanolamine (PC:PE) ratio was found to vary with the growth phase, with increases in PC levels at the expense of PE during the transition to stationary phase . The variation was most pronounced in semi-synthetic lactate medium, which is routinely used for the isolation of mitochondria, where the PC:PE ratio changed from 0.9 to 2.2 during this transition . Similar growth phase-dependent changes in PC and PE content were observed in isolated organelles such as mitochondria, mitochondria-associated membranes and microsomes . Phosphatidylinositol (PI) levels were much higher in cells grown on lactate compared to cells grown on glucose (20% vs . 5-10%) . Irrespective of the medium, PI levels increased upon entering stationary phase . The activities of the phospholipid biosynthetic enzymes phosphatidylserine synthase and the phospholipid-N-methyltransferases were found to be maximal at the end of logarithmic growth and to decrease upon entering stationary phase in all media . Cells grown on lactate displayed a significantly higher phospholipid:protein ratio than cells grown on glucose . The results are discussed in terms of regulation of phospholipid biosynthesis and membrane biogenesis in response to growth phase and carbon source .

J Antibiot (Tokyo), 2000 Feb, 53(2), 163 - 70
Adxanthromycins A and B, new inhibitors of ICAM-1/LFA-1 mediated cell adhesion molecule from Streptomyces sp . NA-148 . II . Physico-chemical properties and structure elucidation; Takahashi S et al.; Adxanthromycins A and B are new inhibitors of ICAM-1/LFA-1 mediated cell adhesion molecule isolated from the fermentation broth of Streptomyces sp . NA-148 . The molecular formula of adxanthromycins A and B were determined as C42H40O17 and C48H50O22, respectively by FAB-MS and NMR spectral analyses, and the structures of both compounds were elucidated to be a dimeric anthrone peroxide skeleton containing alpha-D-galactose by various NMR spectral analyses and chemical degradation.

J Antibiot (Tokyo), 2000 Feb, 53(2), 123 - 30
A new antimitotic substance, FR182877 . I . Taxonomy, fermentation, isolation, physico-chemical properties and biological activities; Sato B et al.; Using the characteristic morphological changes of mammalian cells, we screened novel antimitotic substances and found that a strain of Streptomyces sp . No.9885 produced FR182877 . This substance was isolated from the culture broth by ethyl acetate extraction, silica gel column chromatography and ODS column chromatography . Structural studies on FR182877 suggested that it had a unique hexacyclic structure encompassing its highly strained double bond . FR182877 exhibited potent antitumor activities against murine ascitic tumor and solid tumor in vivo.

J Antibiot (Tokyo), 2000 Feb, 53(2), 110 - 3
CD45 tyrosine phosphatase inhibitory components from Aspergillus niger; Alvi KA et al.; Two inhibitors of CD45 tyrosine phosphatase, dihydrocarolic acid (1) and penitricin D (2), were isolated from a fermentation broth of the fungus Aspergillus niger and purified by HSCCC (high speed countercurrent chromatography) followed by HPLC . The structures were determined by NMR . The inhibitory activities of both compounds were specific to tyrosine phosphatases.

J Antibiot (Tokyo), 2000 Feb, 53(2), 105 - 9
TMC-95A, B, C, and D, novel proteasome inhibitors produced by Apiospora montagnei Sacc . TC 1093 . Taxonomy, production, isolation, and biological activities; Koguchi Y et al.; In our course of screening for novel proteasome inhibitors, TMC-95A and its diastereomers, TMC-95B to D, were isolated from the fermentation broth of Apiospora montagnei Sacc . TC 1093 . TMC-95A inhibited the chymotrypsin-like (ChT-L), trypsin-like (T-L), and peptidylglutamyl-peptide hydrolyzing (PGPH) activities of 20S proteasome with IC50 values of 5.4nM, 200nM, and 60nM, respectively . TMC-95B inhibited these activities to the same extent as TMC-95A, while the inhibitory activities of TMC-95C and D were 20 to 150 times weaker than that of TMC-95A and B . TMC-95A did not inhibit m-calpain, cathepsin L, and trypsin at 30 microM, suggesting its high selectivity for proteasome . Taxonomy of the producing strain is also described.

Planta, 2000 Apr, 210(5), 823 - 30
Isolation and characterisation of cell wall polysaccharides from cocoa (Theobroma cacao L.) beans; Redgwell RJ et al.; Cell wall material (CWM) was prepared from sun-dried cocoa (Theobroma cacao L.) bean cotyledons before and after fermentation . The monosaccharide composition of the CWM was identical for unfermented and fermented beans . Polysaccharides of the CWM were solubilised by sequential extraction with 0.05 M trans-1,2-diaminocyclohexane-N,N,N',N'-tetraacetic acid (CDTA), 0.05 M Na2CO3, and 1 M, 4 M and 8 M KOH . The non-cellulosic sugar composition for each fraction was similar for unfermented and fermented samples, indicating that fermentation caused no significant modification of the structural features of individual cell wall polysaccharides . Pectic polysaccharides accounted for 60% of the cell wall polysaccharides but only small amounts could be solubilised in solutions of CDTA, Na2CO3, and 1 M and 4 M KOH . The bulk of the pectic polysaccharides were solubilised in 8 M KOH and were characterised by a rhamnogalacturonan backbone heavily substituted with side-chains of 5-linked arabinose and 4-linked galactose . Linkage analysis indicated the presence of additional acidic polysaccharides, including a xylogalacturonan and a glucuronoxylan . Cellulose, xyloglucan and a galactoglucomannan accounted for 28%, 8% and 3% of the cell wall polysaccharides, respectively . It is concluded that the types and structural features of cell wall polysaccharides in cocoa beans resemble those found in the parenchymatous tissue of many fruits and vegetables rather than those reported for many seed storage polysaccharides.

Biosci Biotechnol Biochem, 2000 Mar, 64(3), 620 - 2
Novel fungal metabolites, demethylsorbicillin and oxosorbicillinol, isolated from Trichoderma sp . USF-2690; Abe N et al.; The novel compounds, demethylsorbicillin (1) and oxosorbicillinol (2), were isolated from a fermentation broth of Trichoderma sp . USF-2690 . The structures of these compounds, which were determined from spectroscopic evidence, suggest the possibility that methylation at C-6 and oxidation at C-1 and C-6 of sorbicillin were controlled in the early polyketide stage before the formation of oxidized sorbicillin dimers . In a 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical-scavenging assay, 2 gave an ED50 value of 87.7 microM.

Biosci Biotechnol Biochem, 2000 Mar, 64(3), 613 - 6
Increased intestinal calcium absorption from the ingestion of a phosphorylated guar gum hydrolysate independent of cecal fermentation in rats; Watanabe O et al.; The Apparent calcium absorption was increased in rats fed on P-GGH and GGH . However, this increase in calcium absorption from GGH feeding was cancelled by a cecectomy, whereas the corresponding increase from P-GGH feeding was not . The change in femoral calcium content was similar to that in calcium absorption . The calcium solubility in the ileum was increased in those rats fed on P-GGH . We conclude that cecal fermentation did not contribute to the increased calcium absorption by the rats fed on P-GGH.

Appl Microbiol Biotechnol, 2000 Apr, 53(4), 410 - 4
His-tagged tryparedoxin peroxidase of Trypanosoma cruzi as a tool for drug screening; Guerrero SA et al.; Tryparedoxin peroxidase has recently been identified as a constituent of the complex peroxidase system in the trypanosomatid Crithidia fasciculata {Nogoceke E, Gommel DU, Kiess M, Kalisz HM, Flohe L (1997) Biol Chem 378: 827-836} . In trypanosomatids, hydroperoxides are reduced at the expense of NADPH by means of a cascade of three oxidoreductases: the flavoprotein trypanothione reductase, tryparedoxin and tryparedoxin peroxidase . Inhibitors of these enzymes are presumed to be trypanocidal drugs . Here, we present the heterologous expression of a putative tryparedoxin peroxidase gene of Trypanosoma cruzi (accession no AJ012101) as an N-terminally His-tagged protein (TcH6TXNPx) . The product was purified with a high yield (8.75 mg from 11 fermentation broth of A(600)2.1) from the cytosolic fraction of sonified Escherichia coli BL21(DE3){pET22b( + )/TcH6TXNPx} by metal-chelating chromatography . TcH6TXNPx proved to be fully active when tested with heterologous tryparedoxins of C . fasciculata (His-tagged TXN1H6 and TXN2H6) . TcH6TXNPx displayed ping-pong kinetics with a k(cat) of 1.7 s(-1) and limiting Km values of 51.8 microM and 1.7 microM for t-butyl hydroperoxide and CfTXN2H6, respectively.

Free Radic Biol Med, 2000 Mar 15, 28(6), 999 - 1004
Dimerumic acid as an antioxidant of the mold, Monascus anka; Aniya Y et al.; We previously reported that the mold Monascus anka, traditionally used for fermentation of food, showed antioxidant and hepatoprotective actions against chemically induced liver injuries . In the present study, the antioxidant component of M . anka was isolated and identified . The antioxidant was elucidated to be dimerumic acid . DPPH (1,1-diphenyl-2-picrylhydrazyl) radical was significantly scavenged by the antioxidant whereas hydroxyl radical and superoxide anion were moderately scavenged . When the antioxidant (12 mg/kg) was given to mice prior to carbon tetrachloride (CCl(4), 20 microl/kg, ip) treatment, the CCl(4)-induced liver toxicity in mice seen in an elevation of serum aspartate aminotransferase and alanine aminotransferase activities was depressed, suggesting the hepatoprotective action of the antioxidant . The liver microsomal glutathione S-transferase activity, which is known to be activated by oxidative stress or active metabolites, was increased by CCl(4) treatment and the increase was also depressed by pretreatment with the mold antioxidant . Thus these data confirmed that the dimerumic acid isolated from M . anka is the potential antioxidant and protective against CCl(4)-induced liver injury.

J Nutr, 2000 May, 130(5S Suppl), 1378S - 83S
Dietary factors influencing zinc absorption; Lonnerdal B; Marginal zinc deficiency and suboptimal zinc status have been recognized in many groups of the population in both less developed and industrialized countries . Although the cause in some cases may be inadequate dietary intake of zinc, inhibitors of zinc absorption are most likely the most common causative factor . Phytate, which is present in staple foods like cereals, corn and rice, has a strong negative effect on zinc absorption from composite meals . Inositol hexaphosphates and pentaphosphates are the phytate forms that exert these negative effects, whereas the lower phosphates have no or little effect on zinc absorption . The removal or reduction of phytate by enzyme (phytase) treatment, precipitation methods, germination, fermentation or plant breeding/genetic engineering markedly improves zinc absorption . Iron can have a negative effect on zinc absorption, if given together in a supplement, whereas no effect is observed when the same amounts are present in a meal as fortificants . Cadmium, which is increasing in the environment, also inhibits zinc absorption . The amount of protein in a meal has a positive effect on zinc absorption, but individual proteins may act differently; e.g., casein has a modest inhibitory effect of zinc absorption compared with other protein sources . Amino acids, such as histidine and methionine, and other low-molecular-weight ions, such as EDTA and organic acids (e.g., citrate), are known to have a positive effect on zinc absorption and have been used for zinc supplements . Knowledge about dietary factors that inhibit zinc absorption and about ways to overcome or remove these factors is essential when designing strategies to improve the zinc nutrition of vulnerable groups.

Biotechnol Bioeng, 2000 Jun 20, 68(6), 681 - 8
A vortex-bowl disk atomizer system for the production of alginate beads in a 1500-liter fermentor; Champagne CP et al.; Using a model system, a concept for the immobilization of microbial cultures within alginate beads directly in a 1500-L fermentor with a height to diameter ratio of 1.85 is described . The system is comprised of a 60-cm diameter bowl fixed to the top of an agitation shaft, where calcium-ion-rich media is continuously recirculated from the bulk solution to the bowl . The rotation of the shaft and bowl creates a climbing film (vortex) of solution . An atomizing disk centrally recessed within the bowl sprays an alginate solution into the climbing film where the droplets harden into beads . The effect of heat treatment on the alginate solution on resulting bead properties was examined . The sterilization operation did not appear to have a major effect on the alginate bead mechanical properties of firmness and elasticity which was much more a function of alginate concentration . Beads of various sizes were produced by the unit . The system was characterized by the dimensionless numbers Reomega = (omega x rho x D(2))/mu and ReQ = (Q x rho)/(mu x D) . At Reomega and ReQ values less than 500 and 0.15, respectively, the mechanism was direct drop . Parent droplets followed by satellite droplets were observed . When either the flow rate or speed was increased, filaments formed predominantly, which was unwanted in this system because filament breakdown into smaller droplets does not occur due to the proximity of the disk to the climbing film in the bowl . This system could be applied to the immobilization of microorganisms, as well as plant or animal cell cultures, and for other sizes or fermentors . The overflow from the bowl carries the gellified beads into the bulk solution where immobilized cells could act upon the fermentation media .

Biotechnol Bioeng, 2000 Jun 20, 68(6), 602 - 18
Application of metabolic flux analysis for the identification of metabolic bottlenecks in the biosynthesis of penicillin-G; van Gulik WM et al.; A detailed stoichiometric model was developed for growth and penicillin-G production in Penicillium chrysogenum . From an a priori metabolic flux analysis using this model it appeared that penicillin production requires significant changes in fluxes through the primary metabolic pathways . This is brought about by the biosynthesis of carbon precursors for the beta-lactan nucleus and an increased demand for NADPH, mainly for sulfate reduction . As a result, significant changes in flux partitioning occur around four principal nodes in primary metabolism . These are located at: (1) glucose-6-phosphate; (2) 3-phosphoglycerate; (3) mitochondrial pyruvate; and (4) mitochondrial isocitrate . These nodes should be regarded as potential bottlenecks for increased productivity . The flexibility of these principal nodes was investigated by experimental manipulation of the fluxes through the central metabolic pathways using a high-producing strain of P . chrysogenum . Metabolic fluxes were manipulated through growth of the cells on different substrates in carbon-limited chemostat culture . Metabolic flux analysis, based on measured input and output fluxes, was used to calculate the fluxes around the principal nodes . It was found that, for growth on glucose, ethanol, and acetate, the flux partitioning around these nodes differed significantly . However, this had hardly any effect on penicillin productivity, showing that primary carbon metabolism is not likely to contain potential bottlenecks . Further experiments were performed to manipulate the total metabolic demand for the cofactor nicotinamide adenine dinucleotide phosphate (NADPH) . NADPH demand was increased stepwise by cultivating the cells on glucose or xylose as the carbon source combined with either ammonia or nitrate as the nitrogen source, which resulted in a stepwise decrease of penicillin production . This clearly shows that, in penicillin fermentation, possible limitations in primary metabolism reside in the supply/regeneration of cofactors (NADPH) rather than in the supply of carbon precursors .

J Chromatogr B Biomed Sci Appl, 2000 Mar 31, 740(1), 17 - 33
New strategy for the design of ligands for the purification of pharmaceutical proteins by affinity chromatography; Sproule K et al.; A new approach for the identification of ligands for the purification of pharmaceutical proteins by affinity chromatography is described . The technique involves four steps . Selection of an appropriate site on the target protein, design of a complementary ligand compatible with the three-dimensional structure of the site, construction of a limited solid-phase combinatorial library of near-neighbour ligands and solution synthesis of the hit ligand, immobilisation, optimisation and application of the adsorbent for the purification of the target protein . This strategy is exemplified by the purification of a recombinant human insulin precursor (MI3) from a crude fermentation broth of Saccharomyces cerevisiae.

Biotechnol Bioeng, 2000 Jun 5, 68(5), 524 - 30
Effect of alcohol compounds found in hemicellulose hydrolysate on the growth and fermentation of ethanologenic Escherichia coli; Zaldivar J et al.; Lignocellulose can be readily hydrolyzed into a mixture of sugars using dilute mineral acids . During hydrolysis, a variety of inhibitors are also produced which include aromatic alcohols from lignin and furfuryl alcohol from pentose destruction . Seven compounds were investigated individually and in binary combinations (catechol, coniferyl alcohol, furfuryl alcohol, guaiacol, hydroquinone, methylcatechol, and vanillyl alcohol) . Aromatic alcohols and furfuryl alcohol inhibited ethanol production from xylose in batch fermentations primarily by inhibiting the growth of Escherichia coli LY01, the biocatalyst . The toxicities of these compounds were directly related to their hydrophobicity . Methylcatechol was the most toxic compound tested (MIC = 1.5 g/L) . In binary combination, the extent of growth inhibition was roughly additive for most compounds tested . However, combinations with furfuryl alcohol and furfural (furaldehyde) appear synergistic in toxicity . When compared individually, alcohol components which are formed during hemicellulose hydrolysis are less toxic for growth than the aldehydes and organic acids either on a weight basis or a molar basis .

J Agric Food Chem, 1999 Jun, 47(6), 2496 - 500
Enhanced organically bound selenium yeast production by fed-batch fermentation; Demirci A et al.; This study describes a fed-batch fermentation protocol for enhanced production of Saccharomyces cerevisiae containing organically bound selenium . Two levels of sodium selenate concentrations were applied as either a single dose or continuous addition . Fermentations with high sodium selenate (63.2 g/L in cane molasses feeding medium) demonstrated 24 g/L of biomass with 1382 microg of selenium/g of dry biomass for single-dose addition and 40 g/L of biomass with 1491 microg of selenium/g of dry biomass for continuous addition . Low selenium concentration (31.6 g/L in cane molasses feeding medium) demonstrated higher biomass concentration with higher selenium level; 37 g/L of biomass with 2846 microg of selenium/g of dry biomass and 45 g/L of biomass with 2495 microg of selenium/g of dry biomass for single-dose and continuous addition, respectively . Also, two adapted S . cerevisiae strains were evaluated in fed-batch fermentation . A single dose of low concentration demonstrated >3000 microg of selenium/g of dry biomass, but biomass concentration was lower (< or =32 g/L) for these adapted strains.

J Agric Food Chem, 1999 Jun, 47(6), 2491 - 5
Production of organically bound selenium yeast by continuous fermentation; Demirci A et al.; This paper describes a protocol for incorporation of sodium selenite or sodium selenate into Saccharomyces cerevisiae biomass by continuous fermentation in a medium with minimal sulfur and methionine concentrations . Selenium incorporation was followed by atomic absorption analysis and methylene blue reduction time (MBRT) . Continuous fermentation at 0.2 h(-1) dilution rate and sodium selenite addition gradient up to 0.69 g/L of Na(2)SeO(3) yielded 1 . 89 g/L of biomass with 1904 microg of selenium/g of dry biomass . However, MBRT was 0.1 min, which indicated that the majority of selenium was in the inorganic form . On the other hand, continuous fermentation at 0.2 h(-1) dilution rate and sodium selenate gradient up to 0.28 g/L of Na(2)SeO(4) yielded 0.76 g/L of dry biomass with 687 microg of selenium/g of dry biomass, and MBRT was 26 min, which indicated a high concentration of organically bound selenium . Overall, the results indicate a Se/S ratio of 3.9:1 and a dry biomass/Se ratio of 5.5:1 as optimal for continuous production of organically bound selenium.

J Agric Food Chem, 1999 Jun, 47(6), 2455 - 8
Potential environmental impact of effluents from the artichoke (Cynara scolymus L.) byproduct ensiling process using additives; Megias MD et al.; Three treatments have been tested on canned artichoke byproduct after 50 days of ensilage: formic acid at 20% in doses of 2 mL . kg(-)(1) (FA), cane sugar molasses at 50 g.kg(-)(1) (M), and sodium chloride at 30 g.kg(-)(1) (SC) . A fourth batch acted as a control group (C) . The nutritive value, fermentation characteristics, environmental pollution effect, and total volume of effluents released have been studied . The highest nutritive value recorded was with SC silage . The use of the additives did not significantly improve the fermentation stability of the silage, but the total production of effluents in each treatment-52.7 (FA), 46.9 (M), and 55.2 (SC)-was significantly lower (P < 0.01) than that of the control group (70.1 L.Tm(-)(1)) . The chemical oxygen demand (COD), 117300 mg of O(2).L(-)(1), and the conductivity, 46.4 microOmega(-)(1) . cm(-)(1), were significantly higher (P < 0.01) in M and SC, respectively, than in the other group.

J Agric Food Chem, 1999 Jun, 47(6), 2421 - 8
Fate of xanthohumol and related prenylflavonoids from hops to beer; Stevens JF et al.; The fate of three prenylated flavonoids of the chalcone type, xanthohumol, desmethylxanthohumol, and 3'-geranylchalconaringenin, was monitored with LC/MS-MS from hops (Humulus lupulus L.) to beer in two brewing trials . The three prenylchalcones were largely converted into their isomeric flavanones, isoxanthohumol, prenylnaringenins, and geranylnaringenins, respectively, in the boiling wort . Losses of prenylflavonoids were due to incomplete extraction from the hops into the wort (13-25%), adsorption to insoluble malt proteins (18-26%), and adsorption to yeast cells (11-32%) during fermentation . The overall yield of xanthohumol, after lagering of the beer and largely in the form of isoxanthohumol, amounted to 22-30% of the hops' xanthohumol . About 10% of the hops' desmethylxanthohumol, completely converted into prenylnaringenins, remained in the beers . 3'-Geranylchalconaringenin behaved similarly to desmethylxanthohumol . Solubility experiments indicated that (1) malt carbohydrates form soluble complexes with xanthohumol and isoxanthohumol and (2) solubility does not dictate the isoxanthohumol levels of finished beers.

Curr Genet, 2000 Mar, 37(3), 175 - 82
Glutathione, but not transcription factor Yap1, is required for carbon source-dependent resistance to oxidative stress in Saccharomyces cerevisiae; Maris AF et al.; Resistance of haploid yeast to hydrogen peroxide and to tert-butylhydroperoxide strongly increases when 4% glucose is replaced by glycerol or ethanol as the carbon source of the complex medium . Using a GSH1-promoter-lacZ-fusion reporter construct we could demonstrate that GSH1 is one of the genes that are up-regulated during the shift from fermentative to oxidative metabolism . A gsh1 mutant did not exhibit respiratory growth resistance to H2O2, whereas it was only slightly impaired in acquiring resistance against t-BOOH in the same experimental conditions . An isogenic deltayap1 mutant, although more sensitive to oxidative stress than the wild-type (WT), could increase resistance to both peroxides by a similar factor as observed for the WT when shifted from 4% glucose to a non-fermentable carbon source . This indicates that in this case induction of resistance to oxidative stress is independent from Yap1 and from the Yap1-mediated stress response via the STRE motif.

Enzyme Microb Technol, 2000 May 1, 26(8), 602 - 607
Influence of yeast immobilization on fermentation and aldehyde reduction during the production of alcohol-free beer; van Iersel MF et al.; Production of alcohol-free beer by limited fermentation is optimally performed in a packed-bed reactor . This highly controllable system combines short contact times between yeast and wort with the reduction of off-flavors to concentrations below threshold values . In the present study, the influence of immobilization of yeast to DEAE-cellulose on sugar fermentation and aldehyde reduction was monitored . Immobilized cells showed higher activities of hexokinase and pyruvate decarboxylase compared to cells grown in batch culture . In addition, a higher glucose flux was observed, with enhanced excretion of main fermentation products, indicating a reduction in the flux of sugar used for biomass production . ADH activity was higher in immobilized cells compared to that in suspended cells . However, during prolonged production a decrease was observed in NAD-specific ADH activity, whereas NADP-specific activity increased in the immobilized cells . The shifts in enzyme activities and glucose flux correlate with a higher in vivo reduction capacity of the immobilized cells.

Enzyme Microb Technol, 2000 May 1, 26(8), 582 - 592
A systematic approach to the large-scale production of protein crystals; Lee TS et al.; Crystallization has recently emerged as a suitable process for the manufacture of biocatalysts in the form of cross-linked enzyme crystals (CLECs) or for the recovery of proteins from fermentation broths . In both instances it is essential to define conditions which control crystal size and habit, and that yield a reliable recovery of the active protein . Experiments to define the crystallization conditions usually depend on a factorial design (either incomplete or sparse matrix) or reverse screening techniques . In this work, we describe a simple procedure that allows the effect of three factors, for example protein concentration, precipitant concentration and pH, to be varied simultaneously and smoothly over a wide range . The results are mapped onto a simple triangular diagram where a 'window of crystallization' is immediately apparent, and that conveniently describes variations either in the crystal features, such as their yield, size, and habit, or in the recovery of biological activity . The approach is illustrated with two enzymes, yeast alcohol dehydrogenase (ADH I) and Candida rugosa lipase . For ADH the formation of two crystal habits (rod and hexagonal) could be controlled as a function of pH (6.5-10) and temperature (4-25 degrees C) . At pH 7, in 10 to 16% w/v polyethylene glycol (PEG) 4000, only rod-shaped crystals formed whereas at pH 8, in 10 to 14% w/v PEG, only hexagonal crystals existed . For both enzymes, catalyst recovery was greatest at high crystallization agent concentrations and low protein concentration . For ADH, the greatest activity recovery was 87% whereas for the lipase crystals, by using 45% v/v 2-methyl-2,4-pentanediol (MPD) as the crystallization agent, a crystal recovery of 250 crystals per microl was obtained . For the lipase system, the use of crystal seeding was also shown to increase the crystal recovery by up to a factor of four . From the crystallization windows, the original conditions based on literature precedent (35% v/v MPD, 1 mM CaCl(2), 1.8 mg protein/ml) were altered (47.5% v/v MPD, 2 mM CaCl(2), 3 mg protein/ml) . This led to an improved recovery of the lipase under conditions that scale reliably from 0.5 ml to 500 ml with no change in size, shape or recovery of the crystals themselves . Finally, these crystals were crosslinked with 5% v/v glutaraldehyde and mass and activity balances were calculated for the entire process of CLEC production . Up to 35% of the lipase activity present in the crude solid was finally recovered in the lipase CLECs after propan-2-ol fractionation, crystallization, and crosslinking.

J Appl Microbiol, 2000 May, 88(5), 746 - 55
The stress response is repressed during fermentation in brewery strains of yeast; Brosnan MP et al.; Yeast cells encounter a variety of environmental stresses during brewing and must respond to ensure cell survival . Cells can respond to stress by inducing a Heat Shock Response in which heat shock proteins (Hsps) are synthesized . In laboratory strains of Saccharomyces cerevisiae, the heat shock protein, Hsp104, plays a major role in the acquisition of tolerance to a variety of stresses such as heat, ethanol and sodium arsenite, and as such acts as an excellent stress indicator . The induction of Hsp104 in bottom-and top-fermenting brewery strains was examined when grown under laboratory and industrial fermentation conditions, and it was found that each brewing strain exhibits its own unique pattern of Hsp104 expression . During industrial fermentations, brewery strains are capable of mounting a stress response at the early stages of fermentation . However, as the fermentation proceeds, the response is repressed . The results suggest that conditions experienced in industrial brewing prevent the activation of the stress response . This study increases our understanding of alterations in gene expression patterns during the brewing process, and yields information that will aid in the definition of best practice in yeast management.

J Dairy Sci, 2000 Apr, 83(4), 776 - 87
Effects of Isotricha, Dasytricha, Entodinium, and total fauna on ruminal fermentation and duodenal flow in wethers fed different diets; Ivan M et al.; The objective was to measure rumen fermentation and duodenal flow of amino acids and nonammonia N components in five groups of five ruminally and duodenally cannulated wethers that were fauna-free, or inoculated with the ciliate protozoa genera of Isotricha, Dasytricha, Entodinium, or a normal population (total) of fauna . They were used in two 25-d periods and fed a haycrop-based diet in the first period and a corn silage-based diet in the second period . Feces, duodenal digesta, and rumen contents were sampled in each period and analyzed . The number of Entodinium in wethers containing the Entodinium monofauna was higher than the total protozoa numbers, including Entodinium, in wethers containing total fauna population . The type of diet or fauna did not affect total volatile fatty acid concentrations in rumen fluid . The ammonia N concentration in rumen fluid was higher in wethers containing total fauna (25 mg/100 ml) than in fauna-free wethers fed the two diets (18 and 12 mg/ 100 ml) . In comparison with the respective fauna-free wethers, the concentration of ammonia in wethers containing Entodinium was higher when fed the corn silage diet, but not different when fed the haycrop diet . Ruminal presence of total fauna or Entodinium decreased the nonammonia N by 16 and 17%, and total amino acid flows from the stomach by 20 and 19%, respectively . Flow of bacteria N was decreased in wethers fed the two diets when Entodinium or total fauna were present in the rumen . The presence of Isotricha resulted in higher flow of bacteria N in wethers fed the haycrop diet, but the presence of Dasytricha resulted in higher bacteria N flow in wethers fed the corn silage diet . Entodinium was the most detrimental of ciliate protozoa species concerning protein nutrition of the host ruminant.

J Dairy Sci, 2000 Apr, 83(4), 756 - 64
Rumen fermentation in lactating cows selected for milk fat content fed two forage to concentrate ratios with hay or silage; Murphy M et al.; Sixteen multiparous cows, including eight rumen fistulated cows, were used in a 4x4 Latin square experiment designed to study dietary effects on rumen and blood parameters and milk production in cows differing in genetic capacity for milk fat content . Diets contained forage to concentrate ratios of 50:50 or 30:70 with either grass hay or silage as the forage . Ruminal fermentation was characterized by a high molar percentage of butyrate, 14 to 17% . Forage to concentrate ratio affected most rumen parameters, with the exception of the molar percentage of propionate (18 to 19%) . The silage had a higher fiber degradation rate compared with hay . Compared to hay diets, silage diets had higher ruminal outflow rates, lower acetate:propionate ratios, and greater milk production with no differences in milk composition . Cows selected for low milk fat had higher molar percentages of propionate in the rumen . The low milk fat cows had higher milk production than cows selected for high milk fat but did not differ in milk fat yield . Cows fed the 30:70 diets had higher plasma insulin concentrations in response to a glucose challenge . The low milk fat cows had lower basal concentrations of insulin and lower insulin responses to a glucose challenge . Small changes in nutrient metabolism and supply were sufficient to influence milk production.

J Dairy Sci, 2000 Apr, 83(4), 746 - 55
Sites of digestion and bacterial protein synthesis in dairy heifers fed fresh oats with or without corn or barley grain; Garcia SC et al.; Six Holstein-Friesian heifers fitted with ruminal, duodenal, and ileal cannulas were used in a replicated 3x3 Latin square to study the effects of partial replacement {1:1 dry matter (DM) basis} of fresh winter oats (WO) by ground corn (C) or barley (B) on digestion and bacterial protein synthesis . Supplemented diets contained 24% starch, and all diets were fed indoors at 2.5% of body weight (DM basis) . Ruminal and total tract digestibilities of organic matter and neutral detergent fiber were similar for all treatments . Ruminal and total tract starch digestibility was similar for C and B diets . Nitrogen intake was greater for WO than for supplemented diets . However, duodenal flows of nonammonia N and bacterial N did not differ among treatments . The efficiency of bacterial protein synthesis was similar for the three diets, suggesting that the fermentation of high quality fresh forage DM provided sufficient energy for the microorganisms in the rumen . Ruminal ammonia-N concentration was greater for WO than for supplemented diets, and for C than for B diet . Ruminal fluid pH and concentrations of total volatile fatty acid were not different among diets, but concentration of acetate was higher and that of propionate was lower for WO than for supplemented diets and for C than for B diet . Supplementation of WO with barley rather than with corn decreased C2:C3 ratio without affecting fiber digestion . Supplements increased N utilization relative to N intake but did not increase duodenal nonammonia N flow.

Int J Food Microbiol, 2000 Apr 10, 55(1-3), 285 - 90
Modifications to methods for the enumeration and detection of injured Escherichia coli O157:H7 in foods; Blackburn CW et al.; Reliable methods are required for the detection and enumeration of potentially injured E . coli O157 in food in the presence of outnumbering competing bacteria . Selective agents can prevent or inhibit the recovery and subsequent multiplication of injured cells and direct inoculation, either into selective enrichment broths or onto selective agar plates is still used in many methods for E . coli O157 detection and enumeration . When compared with tryptone soya agar (TSA), sorbitol MacConkey agar (SMAC) was shown to underestimate the concentration of viable E . coli O157:H7 subjected to low pH and high NaCl concentration . Using a resuscitation stage on TSA followed by membrane transfer to SMAC improved recovery to levels obtained on TSA . The membrane method was used to monitor the numbers of artificially contaminated E . coli O157:H7 during the fermentation of a meat product and demonstrated better survival when compared to counts on SMAC . Six rapid methods for the detection of E . coli O157 in food (BAX E . coli O157, Reveal 8 E . coli O157-H7 screening test, VIP EHEC, VIDAS E . coli O157 (ECO), EHEC-Tek and Tecra E . coli O157 visual immunoassay), were evaluated using beetburgers, parsley and fermented meat artificially contaminated with injured cells . Methods using direct selective enrichment, with or without an elevated incubation temperature gave false-negative results . The incorporation of a non-selective pre-enrichment medium improved the detection rates of these assays by up to ten fold.

Int J Food Microbiol, 2000 Apr 10, 55(1-3), 259 - 62
Identification of genes responsible for improved cryoresistance in fermenting yeast cells; Tanghe A et al.; Using repetitive freezing and thawing, different mutant industrial Saccharomyces cerevisiae strains with increased freeze resistance have been isolated . To get a better insight in the mechanisms responsible for this elevated resistance and to give us the opportunity to modify other strains so that they become more suitable for use in frozen dough preparations, we applied the microarray technology in order to identify genes that are differentially expressed in a freeze-resistant mutant when compared to a freeze-sensitive industrial yeast strain.

Int J Food Microbiol, 2000 Apr 10, 55(1-3), 103 - 7
A specific mutation in Saccharomyces cerevisiae adenylate cyclase, Cyr1K176M, eliminates glucose- and acidification-induced cAMP signalling and delays glucose-induced loss of stress resistance; Dumortier F et al.; The cAMP-protein kinase A (PKA) pathway in the yeast Saccharomyces cerevisiae plays a major role in the control of metabolism, proliferation and stress resistance . Derepressed cells show a rapid increase in the cAMP level (within 1 min) after addition of glucose or after intracellular acidification . A specific mutation in adenylate cyclase, the enzyme that catalyzes the synthesis in cAMP, largely prevents both cAMP responses . The responsible mutation was originally called lcr1 (for lack of cAMP responses); lcr1 was later identified as allelic with CYR1/CDC35 . The mutation was introduced into the CYR1 gene of a W303-1A wild type strain, which resulted in a large decrease in cAMP signalling . Furthermore, there was a strong reduction in GTP/Mg2+-stimulated but not in Mn2+-stimulated adenylate cyclase activity in isolated plasma membranes, which is consistent with the absence of signalling through adenylate cyclase in vivo . Glucose-induced activation of trehalase was reduced and mobilization of trehalose and glycogen and loss of stress resistance were delayed in the lcr1 mutant . Because of the absence of cAMP signalling during exponential growth on glucose, it was concluded that glucose-induced cAMP signalling is restricted to the transition from gluconeogenic/respiratory to fermentative growth . Activation of the PKA pathway is mediated by a G protein (either Ras1/Ras2 or Gpa2) . Constitutive activation of the pathway by Ras2val19 or Gpa2val132 has a negative effect on glycogen and trehalose accumulation and heat shock survival . The lcr1 mutation partially suppresses this effect indicating that the target sites of the two G-proteins on adenylate cyclase might have at least a part in common.

Hepatogastroenterology, 2000 Mar-Apr, 47(32), 393 - 5
First clinical data of a natural immunomodulator in colorectal cancer; Jakab F et al.; BACKGROUND/AIMS: MSC (trade-name AVEMAR) is a per os applicable complex of multiple, biologically active molecules obtained from fermented wheat-germ extract . Preclinical studies suggest potent anti-metastatic activity and it has a favorable toxicity profile . It has been aimed in a pilot-scale, phase II clinical study to document whether or not MSC as a support to surgery or plus chemotherapy adds any therapeutic benefit compared to the same combination without MSC in colorectal cancer . METHODOLOGY: From 1998 to June 1999, 18 control patients and 12 consecutive colorectal cancer patients respectively, were enrolled into this study . All patients underwent curative surgery . The control group (18 patients) received no other therapy or adjuvant chemotherapy alone . The MSC group (12 patients) received MSC alone or plus adjuvant chemotherapy . Until now, the median follow-up has been 9 months . RESULTS: Interim data of the study document that in the MSC group no new metastases, neither hepatic nor other, have occurred, so far . On the contrary, several new metastases have developed in the control group . CONCLUSIONS: Orally administered MSC is a potent candidate to be regarded as a supportive therapy to surgery or plus chemotherapy for colorectal cancer patients.

Ukr Biokhim Zh, 1999 Jul-Aug, 71(4), 45 - 9
{Effect of aromatic compounds and Mn2+ on the ligninolytic enzyme complex from the fungus Pleurotus floridae (FRIES) Kummer--a white-rot wood fungus}; Dombrovs'ka OM et al.; The influence of aromatic compounds and Mn ions on activities of ligninolityc enzymes from white-rot fungus Pleurotus floridae has been studied . The specific inducers: vanillic acid and vanillyl alcohol--for activity of manganese-dependent peroxidase; vanillyl alcohol--for activity of cellobiose: quinone oxidoreductase during submerged, fermentation of Pleurotus floridae in Kirk's medium have been revealed . The inducers of laccase activity among studied aromatic compounds have not been revealed . The influence of Mn2+ in concentration range 0.4-68.4 mM on activities of ligninolytic enzymes of submerged culture of fungus P . floridae has been studied . Concentration of Mn ions 32.4 mM was optimal for manganese-dependent peroxidase activity.

Yeast, 2000 Apr, 16(6), 539 - 45
Identification by functional analysis of the gene encoding alpha-isopropylmalate synthase II (LEU9) in Saccharomyces cerevisiae; Casalone E et al.; The function of the open reading frame (ORF) YOR108w of Saccharomyces cerevisiae has been analysed . The deletion of this ORF from chromosome XV did not give an identifiable phenotype . A mutant in which both ORF YOR108w and LEU4 gene have been deleted proved to be leucine auxotrophic and alpha-isopropylmalate synthase (alpha-IPMS)-negative . This mutant recovered alpha-IPMS activity and a Leu(+) phenotype when transformed with a plasmid copy of YOR108w . These data and the sequence homology indicated that YOR108w is the structural gene for alpha-IPMS II, responsible for the residual alpha-IPMS activity found in a leu4Delta strain . The leu4Delta strain appeared to be very sensitive to the leucine analogue trifluoroleucine . In the absence of leucine, its growth was not much impaired in glucose but more on non-fermentable carbon sources .

Appl Environ Microbiol, 2000 May, 66(5), 2057 - 61
Mitotic recombination and genetic changes in Saccharomyces cerevisiae during wine fermentation; Puig S et al.; Natural strains of Saccharomyces cerevisiae are prototrophic homothallic yeasts that sporulate poorly, are often heterozygous, and may be aneuploid . This genomic constitution may confer selective advantages in some environments . Different mechanisms of recombination, such as meiosis or mitotic rearrangement of chromosomes, have been proposed for wine strains . We studied the stability of the URA3 locus of a URA3/ura3 wine yeast in consecutive grape must fermentations . ura3/ura3 homozygotes were detected at a rate of 1 x 10(-5) to 3 x 10(-5) per generation, and mitotic rearrangements for chromosomes VIII and XII appeared after 30 mitotic divisions . We used the karyotype as a meiotic marker and determined that sporulation was not involved in this process . Thus, we propose a hypothesis for the genome changes in wine yeasts during vinification . This putative mechanism involves mitotic recombination between homologous sequences and does not necessarily imply meiosis.

Biochim Biophys Acta, 2000 Jan 31, 1490(1-2), 33 - 42
Differential regulation of glutaredoxin gene expression in response to stress conditions in the yeast Saccharomyces cerevisiae; Grant CM et al.; Glutaredoxins are small heat-stable proteins that are active as glutathione-dependent oxidoreductases and are encoded by two genes, designated GRX1 and GRX2, in the yeast Saccharomyces cerevisiae . We report here that the expression of both genes is induced in response to various stress conditions including oxidative, osmotic, and heat stress and in response to stationary phase growth and growth on non-fermentable carbon sources . Furthermore, both genes are activated by the high-osmolarity glycerol pathway and negatively regulated by the Ras-protein kinase A pathway via stress-responsive STRE elements . GRX1 contains a single STRE element and is induced to significantly higher levels compared to GRX2 following heat and osmotic shock . GRX2 contains two STRE elements, and is rapidly induced in response to reactive oxygen species and upon entry into stationary phase growth . Thus, these data support the idea that the two glutaredoxin isoforms in yeast play distinct roles during normal cellular growth and in response to stress conditions.

J Nat Prod, 2000 Apr, 63(4), 489 - 91
Biotransformation of testosterone and pregnenolone catalyzed by the fungus Botrytis cinerea; Farooq A et al.; Testosterone (1), a male sex hormone, and pregnenolone (2), a precursor of many steroidal hormones, were oxidized by fermentation with the fungus Botrytis cinerea . The fermentation of 1 yielded 7beta,17beta-dihydroxyandrostan-3-one (3) (73%) in a yield comparable to chemical transformations . Fermentation of 2 by the same fungus afforded a major metabolite 3beta,11alpha, 16beta-trihydroxypregn-5-en-20-one (4) (39%) along with a minor metabolite 11alpha,16beta-dihydroxypregn-4-ene-3,20-dione (5) (6%) . The metabolites are characterized by detailed physical and spectroscopic studies.

J Anim Sci, 2000 Apr, 78(4), 1067 - 77
Influence of abomasal infusion of high levels of lysine or methionine, or both, on ruminal fermentation, eating behavior, and performance of lactating dairy cows; Robinson PH et al.; Four multiparous late-lactation Holstein cows were fed a basal ration designed to be co-limiting in intestinally absorbable supplies of methionine and lysine . Cows were supplemented with no amino acids, lysine by abomasal infusion to 140% of the calculated intestinally absorbable requirement, methionine by abomasal infusion to 140% of requirement, or both amino acids in a 4 x 4 Latin square design with 28-d periods . Unsupplemented cows consumed 23.8 kg/d of dry matter and produced 36.9 kg/d of milk containing 3.70% fat, 3.22% protein, and 4.82% lactose . Cows ate less dry matter and produced less milk and milk lactose, and tended (P = .06 or .08) to produce less milk protein when abomasally infused with methionine alone or together with lysine . Infusion of lysine alone resulted in production values numerically between those of unsupplemented cows and those cows supplemented with methionine alone or together with lysine . Evaluation of the results with two metabolic models of dairy cows indicated that performance of unsupplemented cows may have been limited by delivery of metabolizable or digestible protein, or intestinally absorbable lysine, isoleucine, or histidine, depending on the metabolic model used to evaluate animal performance . Regardless, results are consistent with those using nonruminant species, which have shown that imbalanced profiles of intestinally absorbable amino acids are associated with reduced dry matter intake and animal performance . Results also show that negative effects on performance of lactating dairy cows can occur if methionine is supplied at levels substantially in excess of calculated intestinally absorbable requirements, either alone or together with lysine.

J Bacteriol, 2000 May, 182(10), 2823 - 30
In vivo analysis of the mechanisms for oxidation of cytosolic NADH by Saccharomyces cerevisiae mitochondria; Overkamp KM et al.; During respiratory glucose dissimilation, eukaryotes produce cytosolic NADH via glycolysis . This NADH has to be reoxidized outside the mitochondria, because the mitochondrial inner membrane is impermeable to NADH . In Saccharomyces cerevisiae, this may involve external NADH dehydrogenases (Nde1p or Nde2p) and/or a glycerol-3-phosphate shuttle consisting of soluble (Gpd1p or Gpd2p) and membrane-bound (Gut2p) glycerol-3-phosphate dehydrogenases . This study addresses the physiological relevance of these mechanisms and the possible involvement of alternative routes for mitochondrial oxidation of cytosolic NADH . Aerobic, glucose-limited chemostat cultures of a gut2Delta mutant exhibited fully respiratory growth at low specific growth rates . Alcoholic fermentation set in at the same specific growth rate as in wild-type cultures (0.3 h(-1)) . Apparently, the glycerol-3-phosphate shuttle is not essential for respiratory glucose dissimilation . An nde1Delta nde2Delta mutant already produced glycerol at specific growth rates of 0.10 h(-1) and above, indicating a requirement for external NADH dehydrogenase to sustain fully respiratory growth . An nde1Delta nde2Delta gut2Delta mutant produced even larger amounts of glycerol at specific growth rates ranging from 0.05 to 0.15 h(-1) . Apparently, even at a low glycolytic flux, alternative mechanisms could not fully replace the external NADH dehydrogenases and glycerol-3-phosphate shuttle . However, at low dilution rates, the nde1Delta nde2Delta gut2Delta mutant did not produce ethanol . Since glycerol production could not account for all glycolytic NADH, another NADH-oxidizing system has to be present . Two alternative mechanisms for reoxidizing cytosolic NADH are discussed: (i) cytosolic production of ethanol followed by its intramitochondrial oxidation and (ii) a redox shuttle linking cytosolic NADH oxidation to the internal NADH dehydrogenase.

Prikl Biokhim Mikrobiol, 2000 Mar-Apr, 36(2), 169 - 72
{Transformation of lignins from grape solids during alcoholic fermentation}; Bezhuashvili MG et al.; Conversion of lignins contained in solid parts of Rkatsiteli grapes (crests, seeds, and skin) during alcoholic fermentation by wine yeast in Reader's medium was studied . Various species of wine yeast were used: Saccharomyces oviformis, S . vini Kakhuri 42, S . chodati Teliani 79, and S . uvarum Tsinandali 77 . We found that lignins from solid parts of grapes are partially decomposed during alcoholic fermentation, which releases low-molecular-weight aromatic compounds into the medium . A peculiar feature of lignin decomposition during alcoholic fermentation is the formation of reduction products.

Prikl Biokhim Mikrobiol, 2000 Mar-Apr, 36(2), 164 - 8
{Furane compounds formed upon heating highly-sugared fermented media}; Pisarintskii AF et al.; 5-Methylfurfural diethyl acetal, 5-methylfuran acid ester, and 4-hydroxypentanoic acid ethyl ester were first identified in high-sacchariferous corn media subjected to alcoholic fermentation by Mucor X-I culture and in aged sweet wines by means of GLC, GLC-MS, and TLC, as well as by UV and IR spectroscopic examinations . These substances are products of enzymatic esterification of sugars followed by their dehydration by heat treatment or long seasoning of grape wines.

Genome Res, 2000 Apr, 10(4), 403 - 9
Evolution and variation of the yeast (Saccharomyces) genome; Mortimer RK; In this review we describe the role of the yeast Saccharomyces in the development of human societies including the use of this organism in the making of wine, bread, beer, and distilled beverages . We also discuss the tremendous diversity of yeast found in natural (i.e., noninoculated) wine fermentations and the scientific uses of yeast over the past 60 years . In conclusion, we present ideas on the model of "genome renewal" and the use of this model to explain the mode by which yeast has evolved and how diversity can be generated.

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