FEMS Microbiol Lett, 1997 Jan 1, 146(1), 161 - 6 The regulatory effect of fermentable sugar levels on the production of leukotoxin by Actinobacillus actinomycetemcomitans; Mizoguchi K et al.; The relationship between sugar availability and RTX (repeats in toxin) cytotoxin (leukotoxin) production in the periodontopathic bacterium, Actinobacillus actinomycetemcomitans, was investigated using a chemostat . A actinomycetemcomitans 301-b produced significant amounts of leukotoxin in anaerobic fructose-limited chemostat cultures at a dilution rate of 0.15 h-1 and at pH 7.0 . When the growth limitation was relieved by pulsing the cultures with 50 or 150 mM fructose (final concentrations), leukotoxin production immediately stopped and the amount of cellular leukotoxin decreased until the culture was returned to fructose-limited conditions . Leukotoxin synthesis was also repressed in the chemostat cultures by pulsing with glucose but not with the non-fermentable sugar analog, alpha-methyl-D-glucoside . Leukotoxin production was also repressed by fructose in chemostat cultures of ATCC 33384, which is generally recognized as a non-leukotoxin-producing or minimally leukotoxic strain. Am J Gastroenterol, 1997 Jan, 92(1), 89 - 94 Influence of a methanogenic flora on the breath H2 and symptom response to ingestion of sorbitol or oat fiber; Kajs TM et al.; BACKGROUND AND OBJECTIVES: We investigated the possibility that a variant of the normal colonic flora, a high concentration of methanogens, influences the host's response to ingestion of nonabsorbable, fermentable materials . METHODS: To better evaluate symptomatic and breath H2 and methane (CH4) responses, subjects were placed on a basal diet (primarily rice and hamburger) that contained minimal amounts of nonabsorbable, fermentable substrate . A breath CH4/H2 ratio of greater or less than 1 on the second day of the basal diet was used to categorize subjects as high (N = 9) or low (N = 25) CH4 producers . After stabilization of the breath gas excretion (day 3 or 4 on the basal diet), the subjects ingested either sorbitol (8.8 g) or oat fiber (10.2 g) . RESULTS: The low CH4 producers had a significantly higher (p < 0.05) breath H2 concentration than the high producers on the basal diet and after ingestion of sorbitol (27.1 +/- 2.7 ppm vs 15.8 +/- 3.6 ppm) or oat fiber (13.1 +/- 0.08 ppm vs 9.6 +/- 1.2 ppm) . Low producers of methane reported significantly increased bloating and cramping after sorbitol ingestion and increased bloating after fiber ingestion, whereas high CH4 producers reported no significant increase in these symptoms . CONCLUSION: The presence of a methanogenic flora is associated with a reduced symptomatic response to ingestion of nonabsorbable, fermentable material in healthy subjects . Manipulation of the normal flora could be of therapeutic value in nonmethanogenic patients with irritable bowel syndrome. Int J Syst Bacteriol, 1997 Jan, 47(1), 191 - 201 Taxonomic study of aromatic-degrading bacteria from deep-terrestrial-subsurface sediments and description of Sphingomonas aromaticivorans sp . nov., Sphingomonas subterranea sp . nov., and Sphingomonas stygia sp . nov; Balkwill DL et al.; Phylogenetic analyses of 16S rRNA gene sequences by distance matrix and parsimony methods indicated that six strains of bacteria isolated from deep saturated Atlantic coastal plain sediments were closely related to the genus Sphingomonas . Five of the strains clustered with, but were distinct from, Sphingomonas capsulata, whereas the sixth strain was most closely related to Blastobacter natatorius . The five strains that clustered with S . capsulata, all of which could degrade aromatic compounds, were gram-negative, non-spore-forming, non-motile, rod-shaped organisms that produced small, yellow colonies on complex media . Their G + C contents ranged from 60.0 to 65.4 mol%, and the predominant isoprenoid quinone was ubiquinone Q-10 . All of the strains were aerobic and catalase positive . Indole, urease, and arginine dihydrolase were not produced . Gelatin was not liquified, and glucose was not fermented . Sphingolipids were present in all strains; 2OH14:0 was the major hydroxy fatty acid, and 18:1 was a major constituent of cellular lipids . Acid was produced oxidatively from pentoses, hexoses, and disaccharides, but not from polyalcohols and indole . All of these characteristics indicate that the five aromatic-degrading strains should be placed in the genus Sphingomonas as currently defined . Phylogenetic analysis of 16S rRNA gene sequences, DNA-DNA reassociation values, BOX-PCR genomic fingerprinting, differences in cellular lipid composition, and differences in physiological traits all indicated that the five strains represent three previously undescribed Sphingomonas species . Therefore, we propose the following new species: Sphingomonas aromaticivorans (type strain, SMCC F199), Sphingomonas subterranea (type strain, SMCC B0478), and Sphingomonas stygia (type strain, SMCC B0712). Int J Syst Bacteriol, 1997 Jan, 47(1), 155 - 9 Schwartzia succinivorans gen . nov., sp . nov., another ruminal bacterium utilizing succinate as the sole energy source; van Gylswyk NO et al.; Four strains of gram-negative, anaerobic, non-spore-forming bacteria that were curved rods which were motile by means of flagella originating from the concave side of the cells and which fermented succinate quantitatively to propionate were isolated from high dilutions of rumen ingesta obtained from cows on pasture . The bacteria were asaccharolytic and not proteolytic and did not ferment amino acids or peptides . Succinate was the only substrate fermented . Rumen fluid together with yeast extract was required for good growth on succinate . Growth on succinate was enhanced in the presence of fumarate . The strains did not grow at 22 degrees C, and growth at 45 degrees C was in all cases less than growth at 39 degrees C . The cellular fatty acid compositions of all four strains were determined . The DNA base composition was about 46 mol% G + C . The complete 16S ribosomal DNA sequence of the type strain (strain S1-1) was determined, and the phylogenetic relationships were analyzed . The most closely related genera were the genera Selenomonas, Zymophilus, and Pectinatus, whereas the recently described succinate-fermenting organism Succiniclasticum ruminis was distantly related . The name proposed for these strains is Schwartzia succinivorans gen . nov., sp . nov.; the type strain is strain S1-1 (= DSM 10502) . These organisms are common inhabitants of the rumina of cows on pasture. Appl Environ Microbiol, 1997 Jan, 63(1), 194 - 200 Effect of 2-bromoethanesulfonic acid and Peptostreptococcus productus ATCC 35244 addition on stimulation of reductive acetogenesis in the ruminal ecosystem by selective inhibition of methanogenesis; Nollet L et al.; Evidence is provided that reductive acetogenesis can be stimulated in ruminal samples during short-term (24-h) incubations when methanogenesis is inhibited selectively . While addition of the reductive acetogen Peptostreptococcus productus ATCC 35244 alone had no significant influence on CH4 and volatile fatty acid (VFA) production in ruminal samples, the addition of this strain together with 2-bromoethanesulfonic acid (BES) (final concentration, 0.01 or 0.03 mM) resulted in stimulation of acetic acid production and H2 consumption . Since acetate production exceeded amounts that could be attributed to reductive acetogenesis, as measured by H2 consumption, it was found that P . productus also fermented C6 units (glucose and fructose) heterotrophically to mainly acetate (> 99% of the total VFA) . Using 14CH3COOH, we concluded that addition of BES and BES plus P . productus did not alter the consumption of acetate in ruminal samples . The addition of P . productus to BES-treated ruminal samples caused supplemental inhibition of CH4 production and stimulation of VFA production, representing a possible energy gain of about 13 to 15%. Appl Environ Microbiol, 1997 Jan, 63(1), 145 - 50 Influence of invertase activity and glycerol synthesis and retention on fermentation of media with a high sugar concentration by Saccharomyces cerevisiae; Myers DK et al.; In the past, the fermentation activity of Saccharomyces cerevisiae in substrates with a high concentration of sucrose (HSuc), such as sweet bread doughs, has been linked inversely to invertase activity of yeast strains . The present work defines the limits of the relationship between invertase activity and fermentation in hyperosmotic HSuc medium . Fourteen polyploid, wild-type strains of S . cerevisiae with different invertase levels gave a similar ranking of fermentation activity in HSuc and in medium in which glucose and fructose replaced sucrose (HGF medium) . Thus, invertase is unlikely to be the most important determinant of fermentation in sweet doughs . Yeasts produce the compatible solute-osmoprotective compound glycerol when exposed to hyperosmotic environments . Under low sugar concentrations (and nonstressing osmotic pressure), there was no correlation between glycerol and fermentation activities . However, there was a strong correlation between the ability of yeasts to ferment in HSuc or HGF medium and their capacity to produce and retain glycerol intracellularly . There was also a strong correlation between intracellular glycerol and fermentation activity of yeasts in a medium in which the nonfermentable sugar alcohol sorbitol replaced most of the sugars (HSor), but the ability to produce and retain glycerol was greater when yeasts were incubated in HGF medium under the same osmotic pressure . The difference between the amounts of glycerol produced and retained in HSor and in HGF media varied with strains . This implies that high fermentable sugar concentrations cause physiological conditions that allow for enhanced glycerol production and retention, the degree of which is strain dependent . In conclusion, one important prerequisite for yeast strains to ferment media with high concentrations of sugar is the ability to synthesize glycerol and especially to retain it. Appl Environ Microbiol, 1997 Jan, 63(1), 128 - 32 Physiological response to anaerobicity of glycerol-3-phosphate dehydrogenase mutants of Saccharomyces cerevisiae; Bjorkqvist S et al.; Mutants of Saccharomyces cerevisiae, in which one or both of the genes encoding the two isoforms of NAD-dependent glycerol-3-phosphate dehydrogenase had been deleted, were studied in aerobic batch cultures and in aerobic-anaerobic step change experiments . The respirofermentative growth rates under aerobic conditions with semisynthetic medium (20 g of glucose per liter) of two single mutants, gpd1 delta and gpd2 delta, and the parental strain (mu = 0.5 h-1) were almost identical, whereas the growth rate of a double mutant, gpd1 delta gpd2 delta, was approximately half that of the parental strain . Upon a step change from aerobic to anaerobic conditions in the exponential growth phase, the specific carbon dioxide evolution rates (CER) of the wild-type strain and the gpd1 delta strain were almost unchanged . The gpd2 delta mutant showed an immediate, large (> 50%) decrease in CER upon a change to anaerobic conditions . However, after about 45 min the CER increased again, although not to the same level as under aerobic conditions . The gpd1 delta gpd2 delta mutant showed a drastic fermentation rate decrease upon a transition to anaerobic conditions . However, the CER values increased to and even exceeded the aerobic levels after the addition of acetoin . High-pressure liquid chromatographic analyses demonstrated that the added acetoin served as an acceptor of reducing equivalents by being reduced to butanediol . The results clearly show the necessity of glycerol formation as a redox sink for S . cerevisiae under anaerobic conditions. Biochim Biophys Acta, 1996 Dec 18, 1277(3), 163 - 200 Bioenergetics of the archaebacterium Sulfolobus; Schafer G; Archaea are forming one of the three kingdoms defining the universal phylogenetic tree of living organisms . Within itself this kingdom is heterogenous regarding the mechanisms for deriving energy from the environment for support of cellular functions . These comprise fermentative and chemolithotrophic pathways as well as light driven and respiratory energy conservation . Due to their extreme growth conditions access to the molecular machineries of energy transduction in archaea can be experimentally limited . Among the aerobic, extreme thermoacidophilic archaea, the genus Sulfolobus has been studied in greater detail than many others and provides a comprehensive picture of bioenergetics on the level of substrate metabolism, formation and utilization of high energy phosphate bonds, and primary energy conservation in respiratory electron transport . A number of novel metabolic reactions as well as unusual structures of respiratory enzyme complexes have been detected . Since their genomic organization and many other primary structures could be determined, these studies shed light on the evolution of various bioenergetic modules . It is the aim of this comprehensive review to bring the different aspects of Sulfolobus bioenergetics into focus as a representative example of, and point of comparison for closely related, aerobic archaea. Biochem J, 1996 Dec 15, 320 ( Pt 3), 723 - 8 Structural elucidation of XR586, a peptaibol-like antibiotic from Acremonium persicinum; Sharman GJ et al.; A novel peptide, XR586, has been isolated from fermentations of Acremonium persicinum (Xenova culture collection number X21488) . The structure of XR586 has been elucidated by means of NMR spectroscopy, electrospray and fast-atom bombardment MS, derivatization and enzymic digestion . It has been shown to be helical by CD measurements . XR586 shows many structural and conformational features in common with peptaibols, particularly the zervamicins . Peptaibol antibiotics are peptides, typically of 15-20 residues, containing a large proportion of alpha-aminoisobutyric acid (Aib) residues . These peptides adopt a helical conformation in solution and display anti-bacterial and toxic properties due to their ability to form pores in membranes . However, while XR586 contains several Aib residues, it lacks a terminal phenylalaninol and terminates in the sequence Phe-Gly . The lack of reduction of the penultimate residue at the C-terminus may indicate that this step is normally at the end of the biosynthetic pathway of peptaibols and occurs with cleavage of Gly . The 1H chemical shift assignments of XR586 are reported in Supplementary Publication SUP 50179 (3 pages), which has been deposited at the British Library Document Supply Centre, Boston Spa, Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem . J . (1996) 313, 9 ("Deposition of data'). FEBS Lett, 1996 Dec 9, 399(1-2), 92 - 4 Re-face stereospecificity at C4 of NAD(P) for alcohol dehydrogenase from Methanogenium organophilum and for (R)-2-hydroxyglutarate dehydrogenase from Acidaminococcus fermentans as determined by 1H-NMR spectroscopy; Berk H et al.; The two diastereotopic protons at C4 of NAD(P)H are seen separately in 1H-NMR spectra . This fact was used to determine the stereospecificity at C4 of NAD(P) for the NADP-dependent alcohol dehydrogenase from Methanogenium organophilum and for the NAD-dependent (R)-2-hydroxyglutarate dehydrogenase from Acidaminococcus fermentans . The reduction of NADP+ with {2H6}ethanol was found to yield (4R)-{4-2H1}NADPH and the oxidation of (4R)-{4-2H1}NADH with 2-oxoglutarate to yield unlabelled {4-1H}NAD+ . These results indicate that both enzymes are Re-face stereospecific at C4 of the pyridine nucleotides. J Med Chem, 1996 Dec 6, 39(25), 5021 - 4 Synthesis and in vitro evaluation of new wortmannin esters: potent inhibitors of phosphatidylinositol 3-kinase; Creemer LC et al.; New C-11 esters of the fermentation product wortmannin have been synthesized, with some of them further derivatized at C-17 . The new esters show greater inhibition of isolated phosphatidylinositol 3-kinase and increased cell cytotoxicity in a rapidly proliferating leukemia cell line, when compared to wortmannin . Reduction of the C-17 ketone caused a slight increase in activity, while acylation of this new alcohol caused severe loss of activity . With their increased activity, the new C-11 esters may be good candidates to explore the in vivo antitumor effects of phosphatidylinositol 3-kinase inhibitors. Gene, 1996 Dec 5, 182(1-2), 101 - 9 A tightly regulated high level expression vector that utilizes a thermosensitive lac repressor: production of the human T cell receptor V beta 5.3 in Escherichia coli; Andrews B et al.; A series of vectors has been constructed to express the human T cell receptor V beta 5.3 under the control of the hybrid trc promoter in Escherichia coli . Transcriptional induction of the trc promoter was achieved chemically by using isopropyl beta-D-thiogalactopyranoside (IPTG) in a bacterial strain that harbors the lacIq gene, or thermally by using the mutant lacIts gene that encodes a temperature-sensitive lac repressor {Bukrinsky et al . (1988) Multicopy expression vector based on temperature-regulated lac repressor: expression of human immunodeficiency virus env gene in Escherichia coli . Gene 70, 415-417} . Several of the plasmids tested also contain the E . coli heat-stable enterotoxin II (STII) signal sequence for protein secretion . In addition, the gene 10 leader sequence from bacteriophage T7 and a minicistron localized upstream of the V beta 5.3 coding sequence were tested for their potential effect on protein production . These elements increased protein yield two-fold when transcription was induced by IPTG, but had no detectable effect on protein yield when transcription was induced thermally . The highest protein yield was obtained when V beta 5.3 was expressed either from plasmid pKB containing the STII signal in strain LJ24, or from plasmid pKBi that lacks the signal sequence, in the protease deficient strain SG21173 (lon, htpR . clp) . Both plasmids contain the lacIts gene, the trc promoter, the two transcription terminators of the rrnB operon, and a tetracycline selection marker . Production of V beta 5.3 using pKBi-V beta 5.3 in strain SG21173 in a 5-liter fermenter under controlled growth conditions yielded over 25 mg V beta 5.3/liter culture . Conversion of the lacIts to the lacIqts gene yielded vector pKBiq-V beta 5.3 which exhibits complete repression of the trc promoter at 30 degrees C . This stringent regulation of the thermally inducible trc promoter, the elimination of IPTG, the inclusion of the tetracycline resistance gene, and the high level of protein yield should render this expression system broadly useful for the high level production of heterologous proteins in E . coli, for both basic research and human therapeutic applications. Arch Latinoam Nutr, 1996 Dec, 44(4 Suppl 1), 36S - 40S Bioavailability of carbohydrates in legumes: digestible and indigestible fractions; Tovar J; Despite their important contribution to seed weight, carbohydrates in pulses have received limited attention . However, experimental evidence accumulated during the last two decades indicate that legumes are rich sources of slowly digestible starch promoting moderate postprandial glycernic and insulinemic responses . Although the reasons for this phenomenon are not completely understood, some intrinsic properties of the starch itself and the microstructure of cotyledon cells appear to determine much of the slow release character . This beneficial feature is rather sensitive to thermal and mechanical processing . A minimum of 10% of the starch occurring in common beans and lentils escapes digestion and absorption in the normal small intestine, and is therefore referred to as "resistant starch" . This material consists mainly of retrograded amylose fractions generated upon cooling of wet-heated pulses . Physically inaccessible starch fractions resulting from cotyledon microstructural properties may also contribute to incomplete digestibility, accounting for up to 40% of the indigestible starch . These indigestible starch fractions are fermented in the large intestine generating gases and volatile fatty acids, compounds that have important influence on the physiology of the colonic mucosa and peripheral metabolism. Arch Latinoam Nutr, 1996 Dec, 44(4 Suppl 1), 31S - 35S Tannins: thermostable pigments which complex dietary proteins and inhibit digestive enzymes; Carmona A; The presence of antinutritional factors in legume seeds and other vegetables has been considered as an expression of the chemical warfare of plants against their predators . As a consequence, the nutritional utilization of these foods has only been possible through the use of a variety of treatments (cooking, fermentation, germination) which increase nutrient bioavailability . Nonetheless, some factors are not destroyed by effect of seed processing, among which stand a family of polymeric polyphenols called tannins . These pigments have the ability to complex and precipitate proteins and inhibit digestive enzymes . This paper describes what has been accomplished in regards to the selection of an appropriate solvent to extract bean polyphenols, the assessment of the most commonly used assay procedures, the purification of bean tannins and the evaluation of their interaction with proteins and digestive enzymes, responsible for their antinutritional effect. Yeast, 1996 Dec, 12(16), 1607 - 33 Pyruvate metabolism in Saccharomyces cerevisiae; Pronk JT et al.; In yeasts, pyruvate is located at a major junction of assimilatory and dissimilatory reactions as well as at the branch-point between respiratory dissimilation of sugars and alcoholic fermentation . This review deals with the enzymology, physiological function and regulation of three key reactions occurring at the pyruvate branch-point in the yeast Saccharomyces cerevisiae: (i) the direct oxidative decarboxylation of pyruvate to acetyl-CoA, catalysed by the pyruvate dehydrogenase complex, (ii) decarboxylation of pyruvate to acetaldehyde, catalysed by pyruvate decarboxylase, and (iii) the anaplerotic carboxylation of pyruvate to oxaloacetate, catalysed by pyruvate carboxylase . Special attention is devoted to physiological studies on S . cerevisiae strains in which structural genes encoding these key enzymes have been inactivated by gene disruption. C R Acad Sci III, 1996 Dec, 319(12), 1071 - 7 The activities of enzymes associated with the intermediary and energy metabolism in hypogean and epigean crustaceans; Hervant F; The activities of 18 enzymes involved in the intermediary and energy metabolism were measured in certain widely-spread peracarid crustaceans: 3 hypogean (Niphargus virei, Niphargus rhenorhodanensis and Stenasellus virei) and 2 epigean (Gammarus fossarum and Asellus aquaticus) ones . The activities of numerous enzymes were correlated with the known metabolic rates of the 5 species . Such rates are reduced in hypogean organisms: levels of enzymatic activity in subterranean species were 1.2 to 8.6 times lower than in epigean species for the main key regulatory enzymes involved in the Krebs cycle and glycolysis (phosphofructokinase, pyruvate kinase, hexokinase and citrate synthetase) . The relative activities of phosphofructokinase, glycogen phosphorylase and hexokinase clearly indicated that glycogen was the main fuel oxidized in both epigean and hypogean organisms . A higher glycogen phosphorylase/hexokinase ratio in hypogean than in epigean crustaceans showed that subterranean species had a greater ability to function anaerobically . The presence of high activities of glutamate-pyruvate transaminase and lactate dehydrogenase in all species (and of malate dehydrogenase and fumarase in hypogean species) was indicative of a coupled fermentation of glycogen and glutamate during anaerobiosis, with lactate and alanine as end-products (as well as succinate in hypogean species) . A low fructose-1,6-bisphosphatase/phosphofructokinase ratio, associated with a low level of phosphoenolpyruvate carboxykinase activity, indicated that the glycolytic pathway was active and that gluconeogenic ability was limited in epigean crustaceans . In contrast, in hypogean species, association of a higher ratio and a high level of phosphoenolpyruvate carboxykinase activity suggested a low glycolytic activity and a high gluconeogenic ability. J Antibiot (Tokyo), 1996 Dec, 49(12), 1212 - 20 TMC-1 A, B, C and D, new antibiotics of the manumycin group produced by Streptomyces sp . Taxonomy, production, isolation, physico-chemical properties, structure elucidation and biological properties; Kohno J et al.; Four new antitumor antibiotics, TMC-1 A, B, C and D were isolated from a fermentation broth of Streptomyces sp . A-230 . Spectroscopic studies have shown that TMC-1 A to D were new members of the manumycin class of antibiotics . These antibiotics showed cytotoxic activities against various tumor cell lines in vitro. J Antibiot (Tokyo), 1996 Dec, 49(12), 1204 - 11 New antitumor substances, FR901463, FR901464 and FR901465 . II . Activities against experimental tumors in mice and mechanism of action; Nakajima H et al.; FR901463, FR901464 and FR901465, novel antitumor substances, were isolated from the fermentation broth of Pseudomonas sp . No . 2663 . Their antitumor activities were examined in three mouse tumor systems and one human tumor system . The three FR compounds prolonged the life of mice bearing murine ascitic tumor P388 leukemia (T/C values were 160%, 145% and 127% for FR901463, FR901464 and FR901465, respectively), and inhibited the growth of a human solid tumor, A549 lung adenocarcinoma, with different effective dose ranges . FR901464 exhibited most prominent effects on these tumor systems among the three FR compounds . FR901464 also inhibited the growth of murine solid tumors, Colon 38 carcinoma and Meth A fibrosarcoma . To address the involvement of transcriptional activation ability of the three FR compounds in the antitumor effect, we selected FR901464 as a candidate compound and investigated cell cycle transition, chromatin status and endogenous gene expression in FR901464-treated tumor cells having elevated transcriptional activity . FR901464 induced characteristic G1 and G2/M phase arrest in the cell cycle and internucleosomal degradation of genomic DNA with the same kinetics as activation of SV40 promoter-dependent cellular transcription in M-8 tumor cells . In contrast to the potent activation of the viral promoter, FR901464 suppressed the transcription of some inducible endogenous genes but not house keeping genes in M-8 cells . These results suggest that FR901464 may induce a dynamic change of chromatin structure, giving rise to strong antitumor activity, and therefore may represent a new type of drug for cancer chemotherapy. J Antibiot (Tokyo), 1996 Dec, 49(12), 1196 - 203 New antitumor substances, FR901463, FR901464 and FR901465 . I . Taxonomy, fermentation, isolation, physico-chemical properties and biological activities; Nakajima H et al.; New antitumor substances, FR901463, FR901464 and FR901465 were isolated from the culture broth of a bacterium of Pseudomonas sp . No.2663 . FR901463, FR901464 and FR901465 remarkably enhanced the transcriptional activity of the promoter of SV40 DNA virus . Further, these compounds exhibited potent antitumor activities against murine and human tumor cell lines in vitro. J Antibiot (Tokyo), 1996 Dec, 49(12), 1189 - 95 Nidulal, a novel inducer of differentiation of human promyelocytic leukemia cells from Nidula candida; Erkel G et al.; Nidulal (1), a novel inducer of differentiation of human HL-60 promyelocytic leukemia cells, was isolated from fermentations of the basidiomycete Nidula candida together with low amounts of niduloic acid (2) . Both compounds are bisabolane sesquiterpenes . Their structures were elucidated by spectroscopic methods . In reporter gene assays nidulal (1) preferentially activated the transcription factor complex AP-1-mediated expression of secreted alkaline phosphatase in COS-7 cells . In addition nidulal (1) and niduloic acid (2) exhibited weak cytotoxic and antibiotic activities. J Dairy Sci, 1996 Dec, 79(12), 2196 - 206 Effect of glucose fermentation on fiber digestion by ruminal microorganisms in vitro; Piwonka EJ et al.; Two in vitro digestion trials were performed to determine whether the negative effect on fiber digestion when pH was maintained at > 6.2 was attributable to glucose alone or to end products of glucose fermentation . In some treatments, glucose was depleted by a previous 6-h incubation; the supernatant from this incubation was used as the buffer source for treatments using the fermented glucose medium . In trial 1, mixed cultures were grown on cellulose, soybean hulls, and corn bran in fresh media with 0 (control) or 25 mM glucose, in media previously fermented for 6 h with 0 (control) or 25 mM glucose, or in fermented control medium plus 25 mM lactic acid . The rate of NDF digestion was decreased with fermented glucose medium but not with fresh glucose medium or lactic acid medium . Concentrations of lactate, propionate, and butyrate did not appear to affect NDF digestion directly . In trial 2, six treatment media were used: control and glucose media that were either fresh or previously fermented for 6 h and fermented control and glucose media treated with a protease . Rate of NDF digestion was slower in cultures with fermented glucose medium that was treated with protease than in fermented control medium without protease . When treated with protease, rate of NDF digestion was not different between the fermented control medium and the fermented glucose medium . Thus, the negative effect on fiber digestion appeared to be attributable partially to a proteinaceous inhibitor that was produced in culture media containing a rapidly fermented sugar. Br J Nutr, 1996 Dec, 76(6), 797 - 808 Substrates available for colonic fermentation from oat, barley and wheat bread diets . A study in ileostomy subjects; Lia A et al.; Nutrients not absorbed in the small bowel will form substrates for microbial growth in the colon which may have implication for the development of colon cancer . The aim of the present study was to investigate whether fibre-rich oat and barley diets increase the excretion of energy-supplying nutrients from the small bowel compared with a low-fibre wheat diet, and whether a possible increase could be related to the beta-glucan content . Nine ileostomy subjects were served four types of bread together with a low-fibre basal diet (12 g dietary fibre/d) . The breads were based on either wheat flour (W diet, 7 g dietary fibre/d), oat bran (OB diet, 29 g dietary fibre/d), the same amount of oat bran with addition of beta-glucanase (EC 3.2.1.4) (OBE diet, 19 g dietary fibre/d) or a fibre-rich barley fraction (B diet, 35 g dietary fibre/d) . An increased ileal excretion of starch was observed with the barley diet but no effect of the oat beta-glucan on starch recovery was found . The NSP + Klason lignin in the ileostomy effluents accounted only for 24, 31, 24 and 35% of the gross energy excretion in the W, OB, OBE and B diet periods respectively . A large part of the dry weight and energy (30, 21, 28 and 27%, in the W, OB, OBE and B diets respectively) in the effluents could not be identified as fat, protein, total starch or NSP + Klason lignin . This unidentified part was probably made up of oligosaccharides, endogenous losses and nutrient complexes . Methods for identifying and analysing these components should be developed and their role as substrates for colonic fermentation and colon cancer development ought to be investigated. J Biochem (Tokyo), 1996 Dec, 120(6), 1055 - 63 Cellular and molecular physiology of Escherichia coli in the adaptation to aerobic environments; Iuchi S et al.; Upon exposure to oxygen, Escherichia coli increases the expression of enzymes essential for aerobic respiration, such as components of the TCA cycle and terminal oxidase complexes . This increase requires the elimination of repression mediated by the Arc regulatory system under anaerobic conditions . Coordinately, the synthesis of enzymes that function in anaerobic processes such as fermentation decreases, partly due to the inactivation of the transcription factor Fnr . E . coli is thus able to adjust the levels of respiratory enzymes to fit its environmental circumstances, and in this case, reduces the production of the less energy efficient fermentation enzymes in favor of the aerobic pathways . In contrast to the advantage in energy production, aerobiosis brings a disadvantage to E . coli: the production of reactive oxygen species (ROS), i.e . superoxide anion radical (O2.-), hydrogen peroxide (H2O2), and hydroxyl radical (.OH) . These byproducts of aerobic respiration damage many biological molecules, including DNA, proteins, and lipids . To alleviate the toxicity of these compounds, E . coli induces the synthesis of protective enzymes, such as Mn-dependent superoxide dismutase (SodA) and catalase I (HP I), and this induction is controlled by the regulatory proteins SoxRS, OxyR, and ArcAB . Thus, ArcAB, Fnr, SoxRS, and OxyR function in concert so that E . coli can optimize its energy production and growth rate . Fnr and SoxRS are cytoplasmic, DNA-binding proteins, and these regulatory systems utilize iron-sulfur clusters as cofactors which may directly sense the redox environment . OxyR is also a cytoplasmic, DNA-binding protein, and appears to respond to redox potential through the oxidation state of a specific cysteine residue . In the ArcAB system (which belongs to the family of two-component regulatory systems), ArcB, a membrane protein, functions as the sensor, and ArcA, a DNA-binding protein, directly controls target gene expression . Under anaerobic conditions, ArcB undergoes autophosphorylation and transphosphorylates ArcA, stimulating ArcA's DNA-binding activity . During aerobic growth, the transphosphorylation of ArcA does not occur . In this signal transduction mechanism, the ArcB C-terminal or "receiver" domain plays a critical role; that is, it stimulates or abolishes the transphosphorylation depending on the metabolic state of the cell, which in turn is influenced by the availability of oxygen . E . coli thus employs at least four global regulatory systems which monitor the cellular oxidative/metabolic conditions, and adjust the expression of more than 70 operons to give the organism a better aerobic life. Appl Microbiol Biotechnol, 1996 Dec, 46(5-6), 653 - 9 Two-step degradation of pyrene by white-rot fungi and soil microorganisms; in der Wiesche C et al.; The effect of soil microorganisms on mineralization of 14C-labelled pyrene by white-rot fungi in solid-state fermentation was investigated . Two strains of white-rot fungi, Dichomitus squalens and a Pleurotus sp., were tested . The fungi were incubated on milled wheat straw contaminated with {14C}pyrene for 15 weeks . CO2 and 14CO2 liberated from the cultures were determined weekly . To study the effect of soil microorganisms on respiration and {14C}pyrene mineralization in different periods of fungal development, the fungal substrate was covered with soil at different times of incubation (after 0, 1, 3, 5, 7, 9 or 11 weeks) . The two fungi showed contrasting ecological behaviour in competition with the soil microflora . Pleurotus sp . was highly resistant to microbial attack and had the ability to penetrate the soil . D . squalens was less competitive and did not colonize the soil . The resistance of the fungus was dependent on the duration of fungal preincubation . Mineralization of {14C}pyrene by mixed cultures of D . squalens and soil microorganisms was higher than by the fungus or the soil microflora alone when soil was added after 3 weeks of incubation or later . With Pleurotus sp., the mineralization of {14C}pyrene was enhanced by the soil microflora irrespective of the time of soil application . With D . squalens, which in pure culture mineralized less {14C}pyrene than did Pleurotus sp., the increase of {14C}pyrene mineralization caused by soil application was higher than with Pleurotus sp. Appl Microbiol Biotechnol, 1996 Dec, 46(5-6), 524 - 32 High volumetric yields of functional dimeric miniantibodies in Escherichia coli, using an optimized expression vector and high-cell-density fermentation under non-limited growth conditions; Horn U et al.; Functional bivalent miniantibodies, directed against the epidermal growth factor receptor, accumulated to more than 3 gl-1 in high-cell-density cultures of Escherichia coli RV308(pHKK) on a pilot scale . The miniantibodies consist of scFv fragments with a C-terminal hinge followed by a helix-turn-helix motif, which homodimerizes in vivo . The improved expression vector pHKK is characterized by the hoklsok suicide system, improving plasmid maintenance, and the inducible lac pl o promoter system with the very strong T7g10 Shine-Dalgarno sequence . The expression unit is flanked by terminators . The prototrophic RV308 cells were cultivated in glucose mineral salt medium and reached a cell density of 145 g dry biomass l-1 after 33 h . After induction, growth continued almost unchanged for a further 4 h with concomitant miniantibody formation . In the fedbatch phase, the concentration of glucose was kept almost constant at the physiological level of approximately 1.5 gl-1, using on-line flow injection analysis for control . Surprisingly, E . coli RV308(pHKK) did not accumulate significant amounts of the metabolic by-product acetate under these unlimited aerobic growth conditions. Arzneimittelforschung, 1996 Dec, 46(12), 1191 - 6 Monitoring the glycosylation pattern of recombinant interferon-omega with high-pH anion-exchange chromatography and capillary electrophoresis; Kopp K et al.; The enzymatic glycosylation of certain asparagine residues in the protein structure has a profound influence on their physico-chemical properties . Many recombinant DNA derived glycoprotein pharmaceuticals for therapeutic use are glycosylated . Their oligosaccharides are important with regard to stability, solubility, in vivo activity and antigenicity . Sophisticated analytical methods that allow a high resolution of synthesized carbohydrate structures are therefore necessary to ensure a constant product quality . To elucidate the robustness of interferon-omega glycosylation regarding process modifications . Chinese Hamster Ovary (CHO)-cells expressing human interferon-omega were cultivated under different fermentation conditions . The most significant glycosylation alterations resulted from the varied parameters such as initial ammonia concentration in the production medium, cultivation mode (adherent versus suspended) or process time . These are detectable with HPAEC (high-pH anion-exchange chromatography) oligosaccharide mapping as well as with capillary electrophoresis. J Nutr, 1996 Dec, 126(12), 3069 - 76 Stable isotope model for assessing production of short chain fatty acids from colon-derived sugar: application in pigs; Kien CL et al.; Sugar reaching the colon because of intestinal maldigestion or malabsorption may be fermented to acetate and other short-chain fatty acids, resulting in stimulation of colonic water absorption and cell proliferation . To explore this phenomenon in more detail, we have developed a stable isotope model for estimating the fraction of colon-derived glucose or lactose that is fermented to acetate, propionate and butyrate . In an initial application of the model, {d3}-acetate and either {1-(13)C}-glucose or {D-1-(13)C}-lactose were infused into the cecum or colon of piglets, and plateau plasma acetate enrichment was monitored in the carotid artery . In acutely anesthetized piglets, the fractions of glucose and lactose fermented to acetate were 17.0 and 20.0%, respectively . In a chronically catheterized piglet, fermentation was higher (34.2%) . When conducted in chronically catheterized animals or via a colostomy or ileostomy in infants, this model may be used to determine how age, previous surgery or antibiotic therapy affects the efficiency of colonic assimilation of carbohydrate. Pharm Acta Helv, 1996 Dec, 71(6), 395 - 403 Quality assurance after process changes of the production of a therapeutic antibody; Brass JM et al.; Process development for the production of a therapeutic humanised antibody is a very complex operation . It involves recombinant genetics, verification of a strong expression system, gene amplification, characterisation of a stable host cell expression system, optimisation and design of the mammalian cell culture fermentation system and development of an efficient recovery process resulting in high yields and product quality . Rapid progress in the field and the wish of some pharmaceutical companies for outsourcing their production are the driving forces for process changes relatively late in the development phase . This literature survey is aimed at identifying the limits of acceptable process changes in up scaling of the fermentation and down stream processing of biopharmaceuticals and defining the demand in production validation to prove product equivalency and identity of the isolated, purified therapeutic antibody. J Anim Sci, 1996 Dec, 74(12), 3112 - 24 A review on environmental impacts of nutritional strategies in ruminants; Tamminga S; Primary (plant), secondary (animal), and tertiary (human) biological systems are driven by energy, either fossil or renewable energy in biomass . Their ratio shifts from about 10:90 in primary, via 25:75 in secondary, to 90:10 in tertiary systems . Energy input in ruminant production is mainly as plants and plant parts from primary production, and the amount needed per unit product (milk, meat) primarily depends on its digestibility . This is high in young, leafy, whole plants, in roots and tubers, and in reproductive organs (whole seeds) or organ parts (by-products) of mature plants . Use of fossil energy per kilogram of DM for primary production ranges from 1 to 3 MJ in forage to over 8 MJ in concentrate feeds, whereas input per kilogram of milk is 1 to 10 MJ . Biomass energy used in ruminant production contains nitrogen (N), phosphorus (P), and potassium (K), but in a ratio rarely balanced to the animals requirements . In secondary systems, energy is partitioned between foods of animal origin and waste . The latter contains OM, N, P, K, and gases (CO2, CH4), which may cause environmental problems . Losses per kilograms of milk vary and are 10 to 45 g for N, 0 to 3 g for P, and 2 to 20 g for K . Environmental impacts of animal production can be reduced by varying the use of inorganic fertilizer and changing the forage to concentrate ratio . Digestibilities can be improved by proper harvest management . Level and ratio of dietary N, P, and K can be adjusted to requirements by selecting proper ingredients, reducing their loss in waste . Limited scope exists to reduce losses in respiration and fermentation gases. J Anim Sci, 1996 Dec, 74(12), 3063 - 75 Physical constraints on voluntary intake of forages by ruminants; Allen MS; Voluntary dry matter intake (VDMI) of forages by ruminants may be limited by distention resulting from restricted flow of digesta through the gastrointestinal tract . An animal's capacity for fill depends on the weight and volume of digesta that causes distention and the flow rate of digesta from the organ in which distention occurs . The reticulorumen is generally regarded as the site in the gastrointestinal tract for which distention limits VDMI with high-fill diets, although evidence suggests that distention of the abomasum may also limit VDMI . Linear decreases in VDMI have been noted with increasing amounts of inert fill inserted into the reticulorumen, but results have not been consistent across several experiments . Reduction in VDMI depends on the extent to which intake is limited by fill before insertion of inert fill; hence animals with high energy requirements consuming relatively low-energy, high-fill diets are affected to the greatest extent . Because NDF generally ferments and passes from the reticulorumen more slowly than other dietary constituents, it has a greater filling effect over time than non-fibrous feed components and has been found to be the best single chemical predictor of VDMI . However, many other factors affect fill, including particle size, chewing frequency and effectiveness, particle fragility, indigestible NDF fraction, rate of fermentation of the potentially digestible NDF, and characteristics of reticular contractions . These factors are only partially accounted for in models that have been developed to predict VDMI . Increased accuracy of prediction of VDMI is expected as models continue to evolve. J Anim Sci, 1996 Dec, 74(12), 3052 - 62 Metabolic constraints on voluntary intake in ruminants; Illius AW et al.; The weak point in all current methods or models of diet formulation is the prediction of intake . The major uncertainty is not in the cases in which physical constraints apply, but in those in which voluntary intake is limited by feedback from metabolic factors . Voluntary intake is, ultimately, a psychological phenomenon, involving the integration of many signals, and reflects the flexibility of biological systems evolved to cope with variability in food supply, composition and animal state . Conditions giving rise to regulatory signals may provide a framework for modeling metabolic constraints on intake . The empirical evidence for metabolic feedback shows that the animal's productive potential, which affects its ability to utilize nutrients, interacts with the balance of absorbed nutrients to regulate intake . The relative importance of the sites where nutrient imbalance occurs (microbial or host animal metabolism) is unclear, as is the relevant time scale (minutes or days) of response . A model of the effects of asynchrony of nutrient supply to ruminal microbes suggests that ammonia and microbial recycling and the contribution of hind-gut fermentation reduce the asynchrony in the balance of nutrients absorbed into the bloodstream . Hitherto, rather little progress has been made in mathematical modeling of the metabolic processes controlling intake . Models that describe the phenomenon in terms of global variables, such as total energy intake, protein supply, and protein synthetic capacity, can simulate the way constraints may operate without requiring or providing a deeper understanding of the metabolic processes involved . Models describing the flux of energy and materials down established metabolic pathways have the potential to explore constraints on intake, but until the problem of parameterizing such models can be overcome, that potential will remain untapped. Lett Appl Microbiol, 1996 Dec, 23(6), 403 - 6 Purification and characterization of glucoamylase produced by Aspergillus niger in solid state fermentation; Selvakumar P et al.; Glucoamylases produced by Aspergillus niger grown on wheat brain in solid cultures were purified . Four different forms, GA I, GA I', GA II and GA III, were found having apparent molecular weights of 112,000, 104,000, and 74,000 and 61,000 Da respectively . The enzymes are glycoproteins with a carbohydrate content of 16%, and optimal activity at 60 degrees C and pH 4.4 . Activity was strongly inhibited by Hg2+ while Mn2+ and Fe2+ were stimulatory . The Km values for the degradation of starch and maltose were 3.5 and 7.8 mg ml-1, respectively. FEMS Microbiol Lett, 1996 Dec 1, 145(2), 189 - 94 Expression in a wine yeast strain of the Aspergillus niger abfB gene; Sanchez-Torres P et al.; A recombinant wine yeast strain has been constructed expressing the gene coding for alpha-L-arabinofuranosidase B from Aspergillus niger under the control of the yeast actin gene promoter . The protein is efficiently secreted by the recombinant yeast, allowing its purification and characterisation . The heterologous alpha-L-arabinofuranosidase B shows similar physico-chemical properties to the native enzyme . The wine produced in microvinification experiments using the recombinant yeast presents the same oenological characteristics as obtained with the untransformed strain . The alpha-L-arabinofuranosidase B protein is detected throughout the fermentation. J Bacteriol, 1996 Dec, 178(23), 6968 - 74 Characterization of the aegA locus of Escherichia coli: control of gene expression in response to anaerobiosis and nitrate; Cavicchioli R et al.; Analysis of the DNA sequence upstream of the narQ gene, which encodes the second nitrate-responsive sensor-transmitter protein in Escherichia coli, revealed an open reading frame (ORF) whose product shows a high degree of similarity to a number of iron-sulfur proteins as well as to the beta subunit of glutamate synthase (gltD) of E . coli . This ORF, located at 53.0 min on the E . coli chromosome, is divergently transcribed and is separated by 206 bp from the narQ gene . Because of the small size of the intergenic region, we reasoned that the genes may be of related function and/or regulated in a similar fashion . An aegA-lacZ gene fusion was constructed and examined in vivo; aegA expression was induced 11-fold by anaerobiosis and repressed 5-fold by nitrate . This control was mediated by the fnr, narX, narQ, and narL gene products . Analysis of an aegA mutant indicated that the aegA gene product is not essential for cell respiration or fermentation or for the utilization of ammonium or the amino acids L-alanine, L-arginine, L-glutamic acid, glycine, and DL-serine as sole nitrogen sources . The ORF was designated aegA to reflect that it is an anaerobically expressed gene . The structural properties of the predicted AegA amino acid sequence and the regulation of aegA are discussed with regard to the possible function of aegA in E . coli. Appl Environ Microbiol, 1996 Dec, 62(12), 4514 - 20 PCR differentiation of commercial yeast strains using intron splice site primers; de Barros Lopes M et al.; The increased use of pure starter cultures in the wine industry has made it necessary to develop a rapid and simple identification system for yeast strains . A method based upon the PCR using oligonucleotide primers that are complementary to intron splice sites has been developed . Since most introns are not essential for gene function, introns have evolved with minimal constraint . By targeting these highly variable sequences, the PCR has proved to be very effective in uncovering polymorphisms in commercial yeast strains . The speed of the method and the ability to analyze many samples in a single day permit the monitoring of specific yeast strains during fermentations . Furthermore, the simplicity of the technique, which does not require the isolation of DNA, makes it accessible to industrial laboratories that have limited molecular expertise and resources. Mol Gen Genet, 1996 Nov 27, 253(1-2), 189 - 97 delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine synthetase is a rate limiting enzyme for penicillin production in Aspergillus nidulans; Kennedy J et al.; The acvA gene from Aspergillus nidulans encoding delta-(L-alpha-aminoadipyl)-L-cysteinyl-D-valine (ACV) synthetase was overexpressed by replacing the wild-type acvA promoter with the ethanol dehydrogenase promoter, alcAp, from A . nidulans . The expression level of alcAp was determined using a strain in which the reporter gene, lacZ, is under the control of alcAp, and was found to be up to 100 times greater than that from the acvA promoter when induced in fermentation conditions . Penicillin yields were found to increase by as much as 30-fold when the acvA gene was overexpressed . Glucose, which strongly represses transcription from alcAp, also repressed penicillin biosynthesis in the overexpression strain . These results prove that ACV synthetase is a rate limiting enzyme for penicillin production in A . nidulans. J Chromatogr A, 1996 Nov 8, 753(1), 73 - 80 Purification of insulin-like growth factor-I and related proteins using underivatized silica; Reifsnyder DH et al.; Adsorption chromatography using underivatized porous glass can be an effective capture step for the purification of recombinant proteins . Classical desorption techniques using chaotropic agents or harsh chemical solvents often result in elution of inactive material and may not be economical at the process scale . More recently, elution schemes have used tetramethylammonium chloride (TMAC) to obtain biologically active material . A TMAC elution was shown to be effective in the initial purification steps for the recovery of recombinant human insulin-like growth factor-I (rhIGF-I) from an Escherichia coli fermentation broth . However, TMAC also elutes other, more hydrophobic, proteins that are difficult to remove in subsequent purification steps . This paper describes the capture of IGF-I from a crude fermentation broth and a more specific elution using a combination of ethanol and NaCl rather than TMAC . This elution also can be used with other proteins including an IGF-I binding protein (BP3) expressed in mammalian cell culture. Rev Invest Clin, 1996 Nov, 48 Suppl, 33 - 43 {Secondary lactase deficiency in children and its epidemiologic implications}; Vega Franco L; The main contributions in the knowledge of secondary lactase deficiency in children are reviewed . We present the clinical features of fermentative diarrhea and the current physiopathological issues, diagnostic procedures, and dietetic treatments, related to this diarrhea, as well as a review of the diseases associated with it . Finally, we discuss the epidemiological implications of the deficiency. Mikrobiol Z, 1996 Nov-Dec, 58(6), 18 - 29 {The binding and absorption by mollicute cells of antisignature analogs of oligodeoxyribonucleotides}; Egorov OV et al.; Processes of absorption and uptake of phosphorothioate, phosphorodithioate and methylphosphonate analogs of oligodeoxynucleotides which are complementary to "signature" 16S rRNA sequences by cells of Acholeplasma laidlawii PG-8 . Mycoplasma fermentans PG-18 and Mycoplasma pneumoniae FH have been investigated . Distinctions in efficiency of absorption by given cells of phosphorothioate and methylphosphonate analogs of oligonucleotides are found out . So, the level of thioates sorption by the cells of A . laidlawii PG-8 exceeded a share of methylphosphonates binding with these cells 5 times as high . A fact of significant accumulation of thio- and dithioate analogs of oligonucleotides by the mollicute cells is established . The intracellular concentration of these compounds in all investigated microorganisms exceeded extracellular one 10(4)-10(5) time . On the basis of obtained experimental data the assumption is made on existence in mollicute cells of two various mechanisms of uptake of analogs of oligonucleotides bearing a negative charge in sugar-phosphate backbone and nonionic oligonucleotide analogs. Tohoku J Exp Med, 1996 Nov, 180(3), 249 - 59 Shift in sodium chloride sources in past 10 years of salt reduction campaign in Japan; Shimbo S et al.; Twenty four-hr total food duplicate samples were collected from nonsmoking house-wives (aged mostly 30 to 60 years) twice at a 10-year interval in winter seasons, once in around 1980 and then in around 1990 in 11 prefectures in Japan . In practice, 342 and 472 samples were obtained in the 1980 and 1990 studies, respectively . Sodium chloride (NaCl) intake via each food item was estimated from the weight of the item in the duplicate . The comparison of 1990 results with 1980 results showed that the total NaCl intake (i.e., NaCl intake via all food items) decreased after a 10-year campaign to lower salt intake . The NaCl/energy ratio however stayed essentially unchanged . Whereas NaCl intake via pickles decreased remarkably and that via miso paste {a fermentation product of soy bean, rice (or wheat) and salt} slightly, the decreases were counteracted by a substantial increase in NaCl intake via soy bean sauce . Meaning of this unexpected counteraction was discussed in relation to the difficulties in the campaign to lower salt intake. J Protein Chem, 1996 Nov, 15(8), 763 - 74 Initial characterization of autoprocessing and active-center mutants of CMV proteinase; Snyder SW et al.; Human cytomegalovirus (CMV) encodes a unique serine proteinase that is required in the maturation of the viral capsid . The CMV proteinase can undergo autocatalytic activation and is subject to proteolytic self-inactivation . Mutant enzyme forms were prepared to eliminate the initial autoprocessing site and thus form an active single-chain protein for structure-function studies . Two mutants of CMV proteinase were cloned and expressed in Escherichia coli . The A143V mutant was a conservative substitution at the first internal cleavage site . The S132A mutant modified one of the triad of residues responsible for catalytic activity . Through the use of computer-controlled high-cell-density fermentations the mutant proteins were expressed in E . coli at approximately 170 mg/L as both soluble (approximately 40% of total) and inclusion-body forms (approximately 60% of total) . The soluble enzyme was purified by standard methods; inclusion-body protein was isolated by standard methods after refolding and solubilization in guanidine or urea . Sedimentation equilibrium and sedimentation velocity analyses reveal that the enzyme undergoes concentration-dependent aggregation . It exhibits a monomer <==> dimer equilibrium (Kd = 1 microM) at low concentrations and remains dimeric at high concentrations (28 mg/ml) . Differential scanning calorimetry data for protein thermal unfolding fit best to a non-two-state model with two components (Tm = 52.3 and 55.3 degrees C) which subsequently aggregate upon unfolding . Analysis of the short-UV circular dichroism spectra of protein forms resulting from expression as soluble molecules (not refolded) reveals that the two mutants have very similar secondary structures which comprise a mixed structural motif of 20% alpha-helix, 26% beta-sheet, and 53% random coil . Though soluble and active (A143V mutant only), CD analysis revealed that protein refolded from inclusion bodies did not exhibit spectra identical to that of protein expressed only in soluble form. Biokhimiia, 1996 Nov, 61(11), 2018 - 39 {The other side of metabolism}; Golubev AG; Since the discovery of enzymic fermentation by Louis Paster, the idea had been widely spread in biochemistry that all molecular interconversions and interactions in living systems are provided by the enzymes . However, recent advances in sensitivity and accuracy of analytical methods lead to isolation and identification of a variety of products that cannot be mapped to a specific biochemical pathway . These products appear in the organism because the chemical characteristics of biomolecules are not limited to the needs of living systems realized through enzyme-catalyzed processes . This review for the first time attempts to summarize such nonenzymic interactions: (i) products of biogenic amine adduct formation to carbonyl-containing compounds in Pictet-Spengler reaction (endogenous neurotoxins which alter mitochondrial functions thus evidently participating in development of age-related cerebral aberrations, i.e., parkinsonism); (ii) products of Schiff base formation with subsequent Amadori rearrangement (nonenzymic glycation of proteins involved in age-related diabetic disorders and atherosclerosis); (iii) products of Michael addition of methylglyoxal, 4-hydroxynonenal, and other products of nonenzymic conversions of carbohydrates and lipids; (iv) products of biomolecule modification with nitrogen- and oxygen-derived free radicals which contribute to cancer and aging . Some applications of these problems in evolutionary and medical biology are reviewed. J Dent Res, 1996 Nov, 75(11), 1885 - 91 Accumulation of fermentable sugars and metabolic acids in food particles that become entrapped on the dentition; Kashket S et al.; Earlier studies (Kashket et al., 1991) showed that particles of high-starch snack foods remained longer on the teeth than those of high-sucrose, low-starch foods . The question arose whether the prolonged presence of food particles enhances cariogenicity . A study was undertaken to measure sugars, starches, and metabolic acids in retained food particles . Subjects consumed portions of different foods, and particles were removed from all bicuspids and first molars at defined times after swallowing . Dry weights, sugars, and short-chain carboxylic acids were determined . High-sucrose foods were cleared rapidly from the teeth, while high-starch foods were retained for up to 20 min . Sucrose, glucose, and fructose persisted in the retained particles . Particles of high-starch foods accumulated maltose and maltotriose, presumably from the breakdown of starch by salivary amylase . At maximum, maltose plus maltotriose constituted 94% of total sugars in particles of potato chips; corresponding values in doughnuts, peanut butter cookies, and salted crackers were 43, 51, and 61%, respectively . Total fermentable sugars in the particles of high-starch foods were similar to those for the high-sucrose confectionery products . Carboxylic acids accumulated within the particles, presumably due to the fermentation of the sugars by entrapped salivary micro-organisms . At maximum (5 to 7 min), acetic, formic, lactic, and propionic acids rose 17-, 30-, 15-, and 1.3-fold, respectively, in doughnuts, and to smaller degrees in potato chips, salted crackers, and chocolate-caramel-peanut bars . In summary, the study demonstrated the persistence of sugars, the progressive accumulation of starch breakdown products, and the fermentation of the accumulated sugars in retained food particles . The findings support the view that high-starch foods contribute to the development of caries lesions. Appl Microbiol Biotechnol, 1996 Nov, 46(4), 353 - 9 Properties of an intracellular beta-glucosidase purified from the cellobiose-fermenting yeast Candida wickerhamii; Skory CD et al.; An intracellular beta-glucosidase was isolated from the cellobiose-fermenting yeast, Candida wickerhamii . Production of the enzyme was stimulated under aerobic growth, with the highest level of production in a medium containing cellobiose as a carbohydrate source . The molecular mass of the purified protein was approximately 94 KDa . It appeared to exist as a dimeric structure with a native molecular mass of about 180 KDa . The optimal pH ranged from 6.0 to 6.5 with p-nitrophenyl beta-D-glucopyranoside (NpGlc) as a substrate . The optimal temperature for short-term (15-min) assays was 35 degrees C, while temperature-stability analysis revealed that the enzyme was labile at temperatures of 28 degrees C and above . Using NpGlc as a substrate, the enzyme was estimated to have a Km of 0.28 mM and a Vmax of 525 mumol product min-1 mg protein-1 . Similar to the extracellular beta-glucosidase produced by C . wickerhamii, this enzyme resisted end-product inhibition by glucose, retaining 58% of its activity at 100 mM glucose . The activity of the enzyme was highest against aryl beta-1,4-glucosides . However, p-nitrophenyl xylopyranoside, lactose, cellobiose, and trehalose also served as substrates for the purified protein . Activity of the enzyme was stimulated by long-chain n-alkanols and inhibited by ethanol, 2-propanol, and 2-butanol . The amino acid sequence, obtained by Edman degradation analysis, suggests that this beta-glucosidase is related to the family-3 glycosyl hydrolases. Phytochemistry, 1996 Nov, 43(4), 791 - 2 11-Hydroxy-4-methyl-2,4,6-dodecatrienoic acid from fermentations of a Mucor species; Lorenzen K et al.; 11-Hydroxy-4-methyl-2,4,6-dodecatrienoic acid was isolated from fermentations of the Mucor species, strain KL 94-92 aq . The compound exhibits cytotoxic activity and the structural elucidation, as well as the biological properties of the new compound, are described. Biotechnol Prog, 1996 Nov-Dec, 12(6), 873 - 6 beta-Lactamase recovery from E . coli cell lysate via two-phase electrophoresis; Oehler RD et al.; beta-Lactamase was recovered from Escherichia coli cell lysate by a novel cell debris removal method using two-phase electrophoresis . The cells were harvested by centrifugation after fermentation, resuspended in a low ionic strength electrophoresis buffer, lysed, and combined with a poly(ethylene glycol)/dextran aqueous two-phase system in the same buffer . The cell lysate was subjected to a 40 V/cm electric field oriented perpendicular to the phase interface for 90 min . Experiments were conducted both with and without a nucleic acid precipitation step using poly(ethylene imine) (PEI) . For PEI-treated lysate at pH 5, the positively charged beta-lactamase was directed to the upper phase, while negatively charged contaminants (including cell debris, nucleic acid/PEI precipitates, and negatively charged proteins) were directed to the lower phase with the applied field . beta-Lactamase yield in the upper phase was 81%, while cell debris and nucleic acids partitioned almost exclusively to the lower phase . For untreated lysate, beta-lactamase did not move in the electric field due to strong interaction with nucleic acids in solution. Biotechnol Prog, 1996 Nov-Dec, 12(6), 744 - 50 Cybernetic model for the growth of Saccharomyces cerevisiae on melibiose; Gadgil CJ et al.; Cybernetic modeling has traditionally been used in the modeling of microbial growth on multiple substrates . In this paper, cybernetic modeling has been applied to serve as a model for growth on substrates such as melibiose, which are disaccharides and enzymatically degrade to a monosaccharide mixture in the fermentation broth . The enzyme alpha-galactosidase has been shown to be strongly induced in the presence of galactose and severely repressed by glucose . In the present model, the relative concentration of alpha-galactosidase has been linked to that of the key enzyme for galactose metabolism . The enzymatic degradation process is placed under the control of the cybernetic variables . The maximum rate of melibiose degradation vm and the Monod parameters for growth of Saccharomyces cerevisiae on pure glucose and galactose were estimated by batch growth experiments . S . cerevisiae growth on melibiose and a mixture of melibiose and glucose under a variety of preculturing conditions was simulated . Depending on the rate of enzymatic degradation (i.e., the value of vm), the cell mass profile for microbial growth on a disaccharide can resemble profiles for growth on a single substrate (melibiose) or can resemble diauxie growth . Experiments indicate that the model is able to accurately predict the cell mass profiles for yeast growth. J Antibiot (Tokyo), 1996 Nov, 49(11), 1096 - 100 Sparoxomycins A1 and A2, new inducers of the flat reversion of NRK cells transformed by temperature sensitive Rous sarcoma virus . I . Taxonomy of the producing organism, fermentation and biological activity; Ubukata M et al.; Streptomyces sparsogenes SN-2325 isolated from a soil sample collected in Hokkaido, was found to produce sparoxomycins A1 and A2, new modulators of proliferation of mammalian cells . Sparoxomycins A1 and A2 reverse the morphology of temperature-sensitive mutant Rous sarcoma virus-infected NRK cells (src(ts)-NRK cells) from the transformed phenotype to the normal phenotype at the permissive temperature. J Antibiot (Tokyo), 1996 Nov, 49(11), 1073 - 8 EI-1511-3, -5 and EI-1625-2, novel interleukin-1 beta converting enzyme inhibitors produced by Streptomyces sp . E-1511 and E-1625 . I . Taxonomy of producing strain, fermentation and isolation; Tanaka T et al.; EI-1511-3, -5 and EI-1625-2, novel interleukin-1 beta converting enzyme (ICE) inhibitors, were isolated from the culture broths of Streptomyces sp . E-1511 and E-1625 . EI-1511-3, -5 and EI-1625-2 selectively inhibited the recombinant human ICE activity with IC50 values of 0.09, 0.38 and 0.2 microM, respectively . Taxonomy, fermentation of the producing strain and isolation of EI-1511-3, -5 and EI-1625-2 are described. J Chromatogr A, 1996 Nov 1, 752(1-2), 111 - 22 Isolation of monoclonal antibodies from cell containing hybridoma broth using a protein A coated adsorbent in expanded beds; Thommes J et al.; A novel expanded bed adsorbent carrying a recombinant protein A ligand was used for the isolation of monoclonal antibodies from cell containing hybridoma fermentation broth . The untreated effluent from a continuous hybridoma cultivation was applied to the stable expanded adsorbent (Streamline rProteinA), which proved to have a high capacity for the MAb studied (14 mg MAb per ml of adsorbent) . A clarified and highly concentrated (up to 50 fold) eluate of high purity was obtained . A scale up of the MAb purification is demonstrated from lab scale (250 mg MAb per purification cycle) to a small pilot scale (2 g MAb per cycle) . Low product concentration in the broth in combination with the high capacity of the adsorbent caused long sample application cycles (10-11 h) . Experimental problems arising from these long cycle times are discussed with regard to a large scale application of the method. Mutagenesis, 1996 Nov, 11(6), 597 - 603 A comparison of the antimutagenic potential of green, black and decaffeinated teas: contribution of flavanols to the antimutagenic effect; Bu-Abbas A et al.; The present study was undertaken to compare the antimutagenic activity of aqueous extracts, at the concentrations used for human consumption, from green, black and decaffeinated black tea . Antimutagenic potential was evaluated against three indirect-acting dietary carcinogens, Glu-P-1, benzo(a)pyrene and nitrosopyrrolidine . All three types of tea gave rise to strong and concentration-dependent suppression of the mutagenicity of the three premutagens in the presence of an activation system . No major difference in the antimutagenic potential of the three types of tea could be discerned . Black tea, decaffeinated black tea and, to a lesser extent, green tea also antagonized the mutagenicity of the direct-acting mutagen 9-aminoacridine . All three types of tea inhibited markedly the NADPH-dependent reduction of cytochrome c and the O-dealkylations of ethoxy-, methoxy- and, to a much lesser extent, pentoxy-resorufin . When the microsomal metabolism was terminated, after the metabolic activation of the premutagens, incorporation of the aqueous tea extracts into the activation system caused a concentration-dependent suppression of mutagenic response . No significant difference in the antimutagenic activity of the three types of tea in this system was evident . Bearing in mind the much higher concentration of flavanols in green tea compared with the black teas, it may be concluded either that these compounds are unlikely to be the major tea components responsible for the antimutagenic, and possibly anticarcinogenic, properties of tea or that their fermentation products are similarly active. Br J Nutr, 1996 Nov, 76(5), 701 - 9 Microbial amino acid synthesis and utilization in rats: the role of coprophagy; Torrallardona D et al.; Four rats were housed in cages with mesh floors; another four rats were housed in tubular anticoprophagy cages, in which they could not turn round to reach their own faeces . Both groups were fed for 6 d on a low-protein diet containing fermentable carbohydrates and 15NH4Cl . At the end of the experiment the rats were killed and their carcasses were homogenized, lysine was isolated by ion-exchange chromatography and its 15N enrichment measured by isotope-ratio mass spectrometry . The 15N enrichment in the lysine of the microbial fraction of faeces and the total amount of lysine in the body were also determined in order to estimate the amount of microbial lysine absorbed . The 15N enrichment in body lysine of non-coprophagic rats was not different from that previously measured in rats given unlabelled NH4Cl, but in coprophagic rats it was significantly higher . The daily absorption of microbial lysine by the coprophagic rats accounted for 20.7 (SE 2.55) mg/kg body weight0-75, but was only 0.5 (SE 1.04) mg/kg body weight0-75 for the non-coprophagic rats . This value was not significantly different from zero . The utilization of microbial amino acids via coprophagy resulted in a higher weight gain (adjusted for intake) in the coprophagic group (15.5 g/6 d) than in the non-coprophagic rats (3.1 g/6 d) . It was concluded that, in rats, the utilization of microbial lysine occurred exclusively via coprophagy. Br J Nutr, 1996 Nov, 76(5), 689 - 700 Microbial amino acid synthesis and utilization in rats: incorporation of 15N from 15NH4Cl into lysine in the tissues of germ-free and conventional rats; Torrallardona D et al.; The absorption of lysine synthesised by the gastrointestinal microflora was estimated by comparing the 15N incorporated into body lysine in four germ-free (15N-GF) and four conventional (15N-CV) rats . They were fed for 10 d on a protein-free diet containing fermentable carbohydrates and 15NH4Cl; another four conventional rats (control), fed on the same diet but with unlabelled NH4Cl, were used to estimate the natural abundance of 15N . The eviscerated carcass of each rat was homogenized and a sample hydrolysed . Lysine was isolated by ion-exchange chromatography and its 15N enrichment was measured by isotoperatio mass spectrometry . The 15N-CV rats significantly incorporated 15N into their body lysine . The 15N-GF rats had a statistically significant, although small, incorporation of 15N into their body lysine, probably arising from a measurement artifact . It was concluded, therefore, that all {15N}lysine was of microbial origin . The total lysine content in the body and the 15N enrichment of lysine in the microbial fraction of the faeces of the 15N-CV rats were also determined . The amount of microbial lysine absorbed by the 15N-CV rats was estimated by dividing the total amount of {15N}lysine in the body by the enrichment of microbial lysine . It was estimated that the daily absorption of microbial lysine by the conventional rats was 21.3 (SE 2.04) mg/kg body weight0.75. Can J Microbiol, 1996 Nov, 42(11), 1100 - 3 Ochratoxin A: an antiinsectan metabolite from the sclerotia of Aspergillus carbonarius NRRL 369; Wicklow DT et al.; Ochratoxin A, a known mycotoxin with demonstrated toxicity to insects, has been isolated from the sclerotia of the fungus Aspergillus carbonarius NRRL 369 . The sclerotia, harvested from a solid substrate fermentation of corn kernels at 28 degrees C, produced quantities of ochratoxin A exceeding 50 ppm/g dry weight of sclerotia . Evidence is presented that ochratoxin A accounts for the activity of the methanol extract against larvae of the detritivorous beetle Carpophilus hemipterus (Nitidulidae) (75% reduction in feeding rate) and corn ear worm Helicoverpa zea (50% mortality with 99% reduction in weight gain among surviving larvae) when incorporated into a pinto bean diet at levels less than those occurring naturally in the sclerotia. Int J Food Sci Nutr, 1996 Nov, 47(6), 455 - 68 Nutrient composition and nutritional importance of green leaves and wild food resources in an agricultural district, Koutiala, in southern Mali; Nordeide MB et al.; This paper discusses the nutrient composition and the nutritional importance of green leaves and wild gathered foods in an area with surplus food production in Mali . In this West African country, there is little information about the nutrient composition and the nutritional quality of foods in general, and of wild gathered foods in particular . Food frequency was collected in two cross-sectional surveys . Focus group discussions with women in the area were used to collect information about seasonality, availability and preparation of various foods . Selected food samples were collected for chemical analysis of nutrient composition . The food samples of green leaves (Adansonia digitata, Amaranthus viridis, Tamarindus indica, Allium cepa), seeds and flour (Parkia biglobosa) and fruits (Tamarindus indica) were analysed for water, energy, fat, protein, minerals, amino acids and carotenoids . Availability and use of the foods varied with seasons . In the rainy season, wild gathered foods (e.g . A . digitata) were used as much as fresh cultivated foods (e.g., A . viridis and A . cepa) . The wild food resources were more frequently used in rural than in urban areas, with A . digitata as the dominating green leaves . Green leaves were rich in energy, protein and minerals (calcium, iron) . Leaves of A . viridis were, in particular, rich in beta-carotene (3290 micrograms/100 g) . Chemical score in dried green leaves varied from 47 (A . cepa) to 81 (A . digitata), with lysine as the first limiting amino acid . P . biglobosa fermented seeds, with 35% fat and 37% protein were a complementary source of lysine in the diet . Based on the seasonality, the frequency of use and the nutrient contents of selected green leaves and wild gathered foods in Koutiala district, it is concluded that these traditional and locally produced foods are valuable and important nutrient contributors in the diet both in rural and urban areas, but most important in rural areas. Arch Microbiol, 1996 Nov, 166(5), 350 - 6 Sodium ion-dependent hydrogen production in Acidaminococcus fermentans; Hartel U et al.; Acidaminococcus fermentans is able to ferment glutamate to ammonia, CO2, acetate, butyrate, and H2 . The molecular hydrogen (approximately 10 kPa; E' = -385 mV) stems from NADH generated in the 3-hydroxybutyryl-CoA dehydrogenase reaction (E degrees ' = -240 mV) of the hydroxyglutarate pathway . In contrast to growing cells, which require at least 5 mM Na+, a Na+-dependence of the H2-formation was observed with washed cells . Whereas the optimal glutamate fermentation rate was achieved already at 1 mM Na+, H2 formation commenced only at > 10 mM Na+ and reached maximum rates at 100 mM Na+ . The acetate/butyrate ratio thereby increased from 2.0 at 1 mM Na+ to 3.0 at 100 mM Na+ . A hydrogenase and an NADH dehydrogenase, both of which were detected in membrane fractions, are components of a model in which electrons, generated by NADH oxidation inside of the cytoplasmic membrane, reduce protons outside of the cytoplasmic membrane . The entire process can be driven by decarboxylation of glutaconyl-CoA, which consumes the protons released by NADH oxidation inside the cell . Hydrogen production commences exactly at those Na+ concentrations at which the electrogenic H+/Na+-antiporter glutaconyl-CoA decarboxylase is converted into a Na+/Na+ exchanger. Arch Microbiol, 1996 Nov, 166(5), 342 - 9 Fermentation of trans-aconitate via citrate, oxaloacetate, and pyruvate by Acidaminococcus fermentans; Hartel U et al.; Growing cells of Acidaminococcus fermentans (DSM 20731 and ATCC 25085) fermented trans-aconitate via citrate, oxaloacetate, and pyruvate to approximately 2 CO2, 1.8 acetate, 0.1 butyrate and 0.9 H2 . The carbon and electron recoveries were close to 100% . On citrate no growth was observed and washed cells were unable to ferment this tricarboxylate . In cell-free extracts, however, citrate as well as trans-aconitate were readily fermented to CO2 and acetate . Under these conditions, also cis-aconitate, oxaloacetate, and pyruvate were formed, whereas butyrate and intermediates of glutamate fermentation, 2-oxoglutatrate and glutaconate, could not be detected . Citrate Si-lyase, a Mg2+-dependent oxaloacetate decarboxylase, and pyruvate synthase were present in quantities that corresponded to the growth rate of the organism. J Anim Sci, 1996 Nov, 74(11), 2860 - 70 Protein levels in beef cattle finishing diets: industry application, university research, and systems results; Galyean ML; Consulting nutritionists were surveyed to determine current formulation and management practices for finishing beef cattle . Among the six consultants surveyed, percentage of CP in finishing diets ranged from 12.5 to 14.4%, with urea levels ranging from .5 to 1.5% of DM . Finishing diets were based primarily on highly processed, rapidly fermented grains (steam-flaked and high-moisture grain), with roughage levels ranging from 3 to 11% of DM . All six consultants considered feed bunk management to be a critical factor affecting feed intake and performance; five of the six consultants used aggressive implant programs based on estrogen + trenbolone acetate . Recent university research was reviewed with respect to CP level and source in finishing diets . Finishing cattle managed on aggressive implant programs seem to respond to higher levels of CP better than would be expected from the factorial calculation approach . Moreover, improvements in performance noted in recent research seemed to be more consistent when supplemental CP was derived from ruminally degraded vs undegraded sources . Calculation of protein requirements with a metabolizable protein (MP) system yielded estimates of protein needs by finishing cattle that agreed more closely with current industry practices than did calculation based on the factorial method . The difference between the MP system and the factorial method was primarily a result of accounting for microbial N needs in the MP system . Reasons for production responses to CP levels that are greater than those calculated by the factorial method include increased accretion of protein by rapidly growing, implanted cattle, particularly during the initial phase of the finishing period, alleviation of a microbial N deficiency, and ruminal and systemic effects of ruminally degraded N on acid-base balance of beef cattle fed rapidly fermented, high-grain diets . Reasons for production responses to supplemental CP need further research. J Nutr, 1996 Nov, 126(11), 2920 - 33 The incidence of swine dysentery in pigs can be reduced by feeding diets that limit the amount of fermentable substrate entering the large intestine; Pluske JR et al.; Two experiments were conducted to test the hypothesis that feeding diets which limit the amount of fermentable substrate entering the large intestine would protect pigs against experimental infection with Serpulina hyodysenteriae, the causative agent of swine dysentery . Experiment 1 examined the effect of grain processing (hammer milling vs . steam flaking) and grain type (barley, groats, corn, sorghum and wheat) on indices of fermentation in the large intestine and the incidence of swine dysentery . Experiment 2 examined the role of five diets, steam-flaked corn, steam-flaked sorghum, hammer-milled wheat, extruded wheat and cooked white rice, on these same measures . All diets contained an animal protein supplement and no antibiotics . Pigs fed diets based on steam-flaked corn and steam-flaked sorghum had a lower incidence of disease (11-33%) than pigs fed diets based on other grains (75-100%) . Pigs fed the diet based on cooked white rice were fully protected against swine dysentery . Both the soluble non-starch polysaccharide (NSP) concentration and the total NSP concentration of the diets explained a significant proportion of the variation in swine dysentery (R2 = 0.56, P = 0.016, and R2 = 0.71, P = 0.002, respectively), such that pigs eating diets containing <1.0 g/100 g soluble NSP showed reduced disease . However, pigs fed corn, sorghum and steam-flaked sorghum (Experiment 2), which contained only 0.4-0.5 g/100 g soluble NSP, still had a high incidence of disease (>50%) . This was attributable to a higher level of resistant starch present in these grains . These data provide evidence that the expression of swine dysentery is associated with an increased concentration of fermentable substrate entering the large intestine. J Nutr, 1996 Nov, 126(11), 2790 - 7 Serum acetate:propionate ratio is related to serum cholesterol in men but not women; Wolever TM et al.; Acetic and propionic acids, produced by colonic fermentation of unabsorbed carbohydrates, may influence systemic lipid metabolism . To determine whether the ratio of the concentrations of acetate to propionate in peripheral serum of fasting humans was related to serum cholesterol, we studied 62 men {age 45 +/- 17 y (mean +/- SD), range 19-74 y; body mass index 25.0 +/- 2.8 kg/m2} and 69 women {43 +/- 18 y, (range, 18-77 y); body mass index 23.0 +/- 3.1 kg/m2} with normal serum lipid concentrations . The concentrations of serum acetate, propionate and butyrate (means +/- SD) were similar in men (98 +/- 33, 3.8 +/- 1.5 and 2.3 +/- 1.5 micromol/L, respectively) and women (92 +/- 38, 3.9 +/- 1.9 and 2.3 +/- 1.6 micromol/L) . There were significant positive relationships between the serum acetate:propionate ratio and total cholesterol (r = 0.466, P = 0.0002) and LDL cholesterol (r = 0.384, P = 0.0023) in men, but in women the relationships were not significant (R = 0.174, P = 0.15 and r = 0.135, P = 0.27, respectively) . The relationships in men remained significant after adjustment for age and body mass index . These data support the hypothesis that, at least in men, colonic short-chain fatty acids influence systemic lipid metabolism . The relationships among the factors influencing colonic short-chain fatty acid production, the enterohepatic circulation of endogenous estrogens, dietary phytoestrogens and blood lipids in women, however, need further clarification. Int J Food Microbiol, 1996 Nov, 33(1), 85 - 102 Moulds in food spoilage; Filtenborg O et al.; There is an increasing knowledge and understanding of the role played by moulds in food spoilage . Especially the discovery of mycotoxin production in foods has highlighted the importance of moulds in food quality . It is, however, only within the last 5-10 years that major progresses have been made towards the prevention of spoilage caused by moulds . This is due to recent international agreements on taxonomy and analytical methods for foodborne moulds, which has led to the discovery, that a specific, very limited funga (= mycobiota) is responsible for the spoilage of each kind of food . This is called the associated or critical funga and has been shown to consist of less than ten species . In this paper the associated funga is described for the following foods: citrus and pomaceous fruit, potato and yam tubers, onions, rye, wheat, rye bread, cheese and fermented sausage and whenever possible the selective principle of the food is discussed . In the description only references which are using the new taxonomy and mycological methods have been used . The individual fungas are very different from each other, which again means that the potential appearance of a specific mycotoxin is restricted to a limited number of foods . The important mycotoxin pattern of each food is described including toxins which originate from 'carry over' . For some foods examples are also given on spoilage of sensoric properties due to moulds . Finally, preventive action against the growth of the associated funga is described for some of the foods and it is concluded that optimization of the prevention and control of moulds in foods must be based on knowledge of the associated funga. Am J Clin Nutr, 1996 Nov, 64(5), 700 - 5 In vivo lactose digestion in preterm infants; Kien CL et al.; In vitro studies of intestinal lactase activity and breath-hydrogen studies have suggested that the capacity for lactose digestion in preterm infants is less than the usual intake . To explore this question using an in vivo approach, we determined the fraction of dietary lactose hydrolyzed to glucose (and galactose) in 14 preterm infants with a gestational age of 26-31 wk at the time of birth but a postconceptional age of 31-37 wk at the time of study . The percentage of lactose digested was estimated after 6-h, primed, constant gastric infusions of {1-(13)C}glucose and D-{-1-(13)C}lactose on alternate days . A coefficient of lactose fermentation was derived from the rates of pulmonary excretion of hydrogen and carbon dioxide . Mean (+/- SD) lactose digestion was 79 +/- 26% . There was a significant inverse rank (r = -0.799, P < 0.01) and linear (r = -0.587, P < 0.05) correlation between this variable and postconceptional age . The percentage of lactose fermented averaged 35 +/- 27%. Appl Environ Microbiol, 1996 Nov, 62(11), 4036 - 8 Aflatoxin and cyclopiazonic acid production by a sclerotium-producing Aspergillus tamarii strain; Goto T et al.; The production of aflatoxins B1 and B2 by Aspergillus tamarii (subgenus Circumdati section Flavi) is reported for the first time . The fungus was isolated from soil collected from a tea (Camellia sinensis) field in Miyazaki Prefecture, Japan . Three single-spore cultures, NRRL 25517, NRRL 25518, and NRRL 25519, were derived from subcultures of the original isolate 19 (MZ2) . Each of these single-spore cultures of A . tamarii produced aflatoxins B1 and B2 and cyclopiazonic acid, as well as black, pear-shaped sclerotia . The demonstration of aflatoxin production by A . tamarii is examined in connection with A . tamarii phylogenetic relationships, chemical ecology, and potential use in food fermentations. Appl Environ Microbiol, 1996 Nov, 62(11), 4009 - 13 Culturing enterohemorrhagic Escherichia coli in the presence and absence of glucose as a simple means of evaluating the acid tolerance of stationary-phase cells; Buchanan RL et al.; Prior growth of seven enterohemorrhagic and one nonenterohemorrhagic strains of Escherichia coli in tryptic soy broth with (TSB+G) and without (TSB-G) 1% glucose was evaluated for its effect on acid tolerance . The final pHs of 18-h TSB+G and TSB-G cultures were 4.6 to 5.2 and 6.9 to 7.0, respectively . Cells were then transferred to brain heart infusion broth adjusted to pH 2.5 or 3.0 with HCl, incubated at 37 degrees C for up to 7 h, and assayed periodically for viable populations with brain heart infusion and MacConkey agars . All enterohemorrhagic strains were acid resistant (< 0.5 log decline after 7 h) when initially cultured in TSB+G, but substantial differences in acid tolerance were observed among strains cultured in TSB-G (log declines ranged from < 0.3 to > 3.8) . The results indicated that prior growth in a medium with and without a fermentable carbohydrate is a convenient way to studying the induction of acid tolerance, that acid inactivation is preceded by a period of acid injury, and that pH-independent and pH-dependent stationary-phase acid tolerance phenotypes may exist among strains of enterohemorrhagic E . coli. Mol Gen Genet, 1996 Oct 28, 252(6), 700 - 8 SLS1, a new Saccharomyces cerevisiae gene involved in mitochondrial metabolism, isolated as a syntheticlethal in association with an SSM4 deletion; Rouillard JM et al.; SSM4 was isolated as a suppressor of rna14-1, a mutant involved in nuclear mRNA maturation . In order to isolate genes interacting with SSM4, we have searched for mutants that are syntheticlethal in association with an SSM4 deletion . Among the mutants obtained, one, named sls1-1, shows a pet- phenotype . We have cloned and sequenced this gene . It encodes a protein with a calculated molecular mass of 73 kDa . This protein contains a mitochondrial targeting presequence but does not show homology with other known proteins . Deletion of SLS1 does not affect cell viability on glucose but is lethal on a non-fermentable medium . The Sls1p protein does not appear to be involved in mitochondrial DNA replication, transcription, or in RNA splicing maturation or stability . We have also tagged this protein and localized it in mitochondria . Treatment with alkaline carbonate does not extract this protein from mitochondria, suggesting strongly that it is a mitochondrial integral membrane protein . Thus, the SLS1 gene, encodes a mitochondrial integral membrane protein and is paradoxically synlethal in association with a deletion of the SSM4 gene, which encodes an integral nuclear membrane protein. Hum Gene Ther, 1996 Oct 20, 7(16), 1971 - 80 High-yield production of pBR322-derived plasmids intended for human gene therapy by employing a temperature-controllable point mutation; Lahijani R et al.; Production of large quantities of highly purified plasmid DNA is essential for gene therapy . A low-copy-number pBR322-derived plasmid (VCL1005) was converted to a high-copy-number plasmid (VCL1005G/A) by incorporating a G-->A mutation that affects initiation of DNA replication from the ColE1 origin of replication . Because the phenotypic effect of this mutation is enhanced at an elevated temperature, a further increase in yield was achieved by changing the growth temperature from 37 degrees C to 42 degrees C at mid-log phase during batch and fed-batch fermentation . The combined effect of the single base-pair change and the elevated growth temperature produced an overall yield of 2.2 grams of plasmid DNA available for recovery from a 10-liter fed-batch fermentation compared to 0.03 grams from a 10-liter batch fermentation, a 70-fold increase in yield . The plasmid DNA isolated from this process contained lower levels of RNA and chromosomal DNA contaminants, simplifying downstream processing. Eur J Biochem, 1996 Oct 15, 241(2), 633 - 43 Differential requirement of the yeast sugar kinases for sugar sensing in establishing the catabolite-repressed state; De Winde JH et al.; Addition of rapidly fermentable sugars to cells of the yeast Saccharomyces cerevisiae grown on nonfermentable carbon sources causes a variety of short-term and long-term regulatory effects, leading to an adaptation to fermentative metabolism . One important feature of this metabolic switch is the occurrence of extensive transcriptional repression of a large group of genes . We have investigated transcriptional regulation of the SUC2 gene encoding repressible invertase, and of HXK1, HXK2 and GLK1 encoding the three known yeast hexose kinases during transition from derepressed to repressed growth conditions . Comparing yeast strains that express various combinations of the hexose kinase genes, we have determined the importance of each of these kinases for establishing the catabolite-repressed state . We show that catabolite repression involves two distinct mechanisms . An initial rapid response is mediated through any kinase, including Glk1, which is able to phosphorylate the available sugar . In contrast, long-term repression specifically requires Hxk2 on glucose and either Hxk1 or Hxk2 on fructose . Both HXK1 and GLK1 are repressed upon addition of glucose or fructose . However, fructose repression of Hxk1 is only transient, which is in line with its preference for fructose as substrate and its requirement for long-term fructose repression . In addition, expression of HXK1 and GLK1 is regulated through cAMP-dependent protein kinase . These results indicate that sugar sensing and establishment of catabolite repression are controlled by an interregulatory network, involving all three yeast sugar kinases and the Ras-cAMP pathway. Eur J Biochem, 1996 Oct 15, 241(2), 468 - 75 The Escherichia coli-derived Fab fragment of the IgM/kappa antibody IN-1 recognizes and neutralizes myelin-associated inhibitors of neurite growth; Bandtlow C et al.; A recombinant Fab fragment was prepared from the monoclonal IgM/kappa antibody IN-1, which neutralizes central nervous system myelin-associated neurite growth inhibitors both in vitro and in vivo . The variable domain gene sequences were amplified and cloned after cDNA synthesis from the hybridoma RNA . After insertion into the tet promoter vector pASK85, which provided the constant domains of class IgG1/kappa, equipped with a His6 tag, large amounts of the Fab fragment were produced in Escherichia coli by medium cell density fermentation . The Fab fragment was purified to homogeneity by immobilized metal-affinity chromatography and its biochemical activity was compared with the original IN-1 antibody . In an assay for neurite outgrowth and fibroblast spreading, the Fab fragment showed a similar neutralizing effect on inhibitory substrate properties of central nervous system myelin as the unpurified IgM, although an approximately tenfold higher concentration was necessary . Immunoprecipitation experiments revealed a more selective antigen-binding behaviour for the Fab fragment . The Fab fragment was also successfully applied for antigen detection in immunohistochemical analyses . Therefore, the recombinant Fab fragment of IN-1 shows full functionality in vitro and appears to be well suited for replacing the monoclonal IgM in investigations on fiber tract regeneration in vivo. Biochem Biophys Res Commun, 1996 Oct 3, 227(1), 53 - 8 Modeling interpretation of microbe metabolism detected by nuclear magnetic resonance; Bastianoni S et al.; A new modeling approach is discussed for the analysis of microbial dynamics . It is based on structurally nonlinear compartmental models and on the dynamics of the substrate and product . Experimental data were acquired by NMR spectroscopy which is a noninvasive technique . This combined approach was tested with the fermentation of glucose to ethanol by Saccharomyces cerevisiae . The model was fitted with the experimental data to obtain the values of the parameters of the model . Similar processes can be analyzed and compared using this approach. Z Lebensm Unters Forsch, 1996 Oct, 203(4), 379 - 84 Analysis of free amino acids in green coffee beans . II . Changes of the amino acid content in arabica coffees in connection with post-harvest model treatment; Arnold U et al.; To investigate amino acid changes in green coffee beans in the post-harvest period, amino acid concentrations were determined in green beans and after modelled drying, fermentation and storage . After the drying at alternating temperatures up to maximally 40 degrees C, considerable changes in the concentrations of individual amino acids were identified . At the beginning of the storage period, significant changes in concentration were found to a minor extent . Under the condition of drying, it was mainly the concentration of glutamic acid that changed considerably . There was an increase in all the samples by 500 mg/kg dry matter on average, which corresponds to an increase of about 50% of the original value . In contrast, the concentration of aspartic acid in most of the samples decreased clearly due to drying . For the predominant part of the coffee samples, there was a significant increase in the hydrophobic amino acids Val, Phe, Ile and Leu . Changes of the quantities of other amino acids were non-uniform and only insignificant . Constant drying at 80 degrees C for most of the amino acids brought about only minor concentration changes compared to those values obtained at 40 degrees C . Modelled fermentation had no significant effect on the concentrations of the individual amino acids . After a 4-week storage of dried beans, amino acid concentrations did not change further . It is very possible that different post-harvest treatment parameters may influence the amount of aroma precursor compounds in the coffee beans. Lett Appl Microbiol, 1996 Oct, 23(4), 266 - 8 Detection of maltose fermentation genes in the baking yeast strains of Saccharomyces cerevisiae; Oda Y et al.; The presence of any one of the five unlinked MAL loci (MAL1, MAL2, MAL3, MAL4 and MAL6) confers the ability to ferment maltose on the yeast Saccharomyces cerevisiae . Each locus is composed of three genes encoding maltose permease, alpha-glucosidase and MAL activator . Chromosomal DNA of seven representative baking strains has been separated by pulse-field gel electrophoresis and probed with three genes in MAL6 locus . The DNA bands to which all of the three MAL-derived probes simultaneously hybridized were chromosome VII carrying MAL1 in all of the strains tested, chromosome XI carrying MAL4 in six strains, chromosome III carrying MAL2 in three strains and chromosomes II and VIII carrying MAL3 and MAL6, respectively, in the one strain . The number of MAL loci in baking strains was comparable to those of brewing strains. Lett Appl Microbiol, 1996 Oct, 23(4), 231 - 3 Rapid detection and isolation of DNA-binding compounds from Streptomyces xanthochromogenes; Holmalahti J et al.; The culture medium of Streptomyces xanthochromogenes JH903 was found to show selective activity against DNA-repair-deficient Escherichia coli CM871 strain . In this report we describe a simple method to locate and isolate DNA binding compounds from the fermentation broth . The method is based on the retention of DNA-reacting compounds in cellulose complexed with DNA, and purification of these compounds with thin-layer chromatography . Screening of microbial metabolites from chloroform extracts of fermentation media resulted in detection of five genotoxic fractions. J Antibiot (Tokyo), 1996 Oct, 49(10), 998 - 1005 Halymecins, new antimicroalgal substances produced by fungi isolated from marine algae; Chen C et al.; Novel antimicroalgal substances halymecins A (1), B (2) and C (3) were isolated from the fermentation broth of a Fusarium sp . and halymecins D (4) and E (5) from an Acremonium sp . The structures of these halymecins, Fig . 1, were determined based on extensive 2D NMR studies as well as mass spectral data . These chemical structures are conjugates of di- and trihydroxydecanoic acid . Halymecin A showed antimicroalgal activity against Skeletonema costatum. J Antibiot (Tokyo), 1996 Oct, 49(10), 990 - 7 Harziphilone and fleephilone, two new HIV REV/RRE binding inhibitors produced by Trichoderma harzianum; Qian-Cutrone J et al.; During the screening of the natural products for their ability to inhibit the binding of REV (regulation of virion expression) protein to {33P} labeled RRE (REV responsive element) RNA, two novel fungal metabolites, harziphilone and fleephilone, were isolated from the butanol-methanol (1:1) extract of the fermentation broth of Trichoderma harzianum by bioassay guided fractionation . The structures of these two new compounds were established by spectroscopic methods . Harziphilone and fleephilone showed inhibitory activity against the binding of REV-protein to RRE RNA with IC50 values of 2.0 microM and 7.6 microM, respectively . However both compounds did not protect CEM-SS cells from acute HIV infection at concentration levels up to 200 micrograms/ml using an XTT dye reduction assay . In addition, harziphilone demonstrated cytotoxicity at 38 microM against the murine tumor cell line M-109. J Antibiot (Tokyo), 1996 Oct, 49(10), 985 - 9 A new isopatulin derivative pintulin produced by Penicillium vulpinum F-4148 taxonomy, isolation, physico-chemical properties, structure and biological properties; Mikami A et al.; During our screening program of natural products from fungal metabolites for drugs effective against tumor cell lines, we discovered a new isopatulin derivative, pintulin, from the fermentation broth of Penicillium vulpinum F-4148 . Pintulin shows weak activity against tumor cell lines, compared to that of adriamycin. J Antibiot (Tokyo), 1996 Oct, 49(10), 974 - 9 Studies on new antitumor antibiotics, leptofuranins A, B, C and D.I . Taxonomy, fermentation, isolation and biological activities; Hayakawa Y et al.; The retinoblastoma protein (pRB) is inactivated in a wide variety of human cancers . In the course of our screening for antitumor antibiotics by using pRB-inactivated cells, an actinomycete identified as Streptomyces tanashiensis was found to produce four new active substances, leptofuranins A, B, C and D . The leptofuranins arrested the growth of normal cells and induced apoptotic cell death against tumor cells and cells transformed with the adenovirus E1A gene. Appl Environ Microbiol, 1996 Oct, 62(10), 3655 - 61 Measurement of minimum substrate concentration (Smin) in a recycling fermentor and its prediction from the kinetic parameters of Pseudomonas strain B13 from batch and chemostat cultures; Tros ME et al.; The minimum substrate concentration required for growth, Smin, was measured for Pseudomonas sp . strain B13 with 3-chlorobenzoate (3CB) and acetate in a recycling fermentor . The substrates were provided alone or in a mixture . Smin values predicted with kinetic parameters from resting-cell batches and chemostat cultures differed clearly from the values measured in the recycling fermentor . When 3CB and acetate were fed as single substrates, the measured Smin values were higher than the individual Smin values in the mixture . The Smin in the mixture reflected the relative energy contributions of the two substrates in the fermentor feed . The energy-based maintenance coefficients during zero growth in the recycling fermentor were comparable for all influent compositions (mean +/- standard deviation, 0.34 +/- 0.07 J mg {dry weight}-1 h-1) . Maintenance coefficient values for acetate were significantly higher in chemostat experiments than in recycling-fermentor experiments . 3CB maintenance coefficients were comparable in both experimental systems . The parameters for 3CB consumption kinetics varied remarkably with the experimental growth conditions in batch, chemostat, and recycling-fermentor environments . The results demonstrate that the determination of kinetic parameters in the laboratory for prediction of microbial activity in complex natural systems should be done under conditions which best mimic the system under consideration. J Clin Pathol, 1996 Oct, 49(10), 824 - 8 Mycoplasma fermentans, but not M penetrans, detected by PCR assays in synovium from patients with rheumatoid arthritis and other rheumatic disorders; Schaeverbeke T et al.; AIM/BACKGROUND: Mycoplasmas, especially Mycoplasma fermentans, were suggested more than 20 years ago as a possible cause of rheumatoid arthritis but this hypothesis was never substantiated . In view of the superior sensitivity of the polymerase chain reaction (PCR) assay over culture, the aim was to use this method to seek M fermentans and M penetrans in synovial samples from patients with various arthritides . METHODS: Synovial fluid samples (n = 154) and synovial biopsy specimens (n = 20) from 133 patients with various rheumatic disorders were stored at -80 degrees C for between one and 40 months . Aliquots (500 microliters) of the synovial fluid samples were centrifuged and the deposit, and also the synovial biopsy specimens (approximately 1 g) were placed in lysis buffer with proteinase K for DNA extraction . The DNA was tested by using a semi-nested PCR assay for M fermentans and a single-round PCR for M penetrans . RESULTS: M fermentans was detected in the joints of eight (21%) of 38 patients with rheumatoid arthritis, two (20%) of 10 patients with spondyloarthropathy with peripheral arthritis, one (20%) of five patients with psoriatic arthritis, and four (13%) of 31 patients with unclassified arthritis . M fermentans was not found in the joints of the seven patients with reactive arthritis, the 29 with osteoarthritis or post-traumatic hydrarthrosis, the nine with gouty arthritis, nor the four with chronic juvenile arthritis . M penetrans was not detected in any sample . CONCLUSIONS: These findings show that the presence of M fermentans in the joint is associated with inflammatory rheumatic disorders of unknown cause, including rheumatoid arthritis . However, whether this organism triggers or perpetuates disease of behaves as a passenger remains conjectural. Protein Eng, 1996 Oct, 9(10), 921 - 6 Differentiation between proteolytic activation and autocatalytic conversion of human prothrombin . Activation of recombinant human prothrombin and recombinant D419N-prothrombin by snake venoms from Echis carinatus and Oxyuranus scutellatus; Fischer BE et al.; Recombinant human prothrombin (r-prothrombin) and recombinant mutant prothrombin with active site Asp419 substituted by Asn (D419N-prothrombin) were expressed in recombinant CHO cells, isolated and purified from the fermentation supernatant . The r-Prothrombin and D419N-prothrombin were digested by both Echis carinatus venom and Oxyuranus scutellatus venom . Prior to, during and after activation, generation of thrombin activity and the proteolytic degradation of the prothrombin polypeptide chain were analysed . Owing to the recombinant preparation and inactivity of D419N-prothrombin and its activation products, the proteolytic action of E.carinatus and O.scutellatus venoms could be studied without addition of thrombin inhibitor, without interference from autocatalytic digestion of prothrombin and in the absence of any other blood coagulation protease . The comparison between the activation of r-prothrombin and D419N-prothrombin by snake venoms permitted differentiation between proteolytic activation and autocatalytic conversion of prothrombin . Incubation of D419N-prothrombin with E.carinatus venom resulted in the generation of stable D419N-meizothrombin by hydrolysis of the peptide bond Arg320-Ile321 . By contrast, O.scutellatus venom exhibited activity towards peptide bonds Arg320-Ile321 and Arg271-Thr272 and lower activity towards peptide bond Arg155-Ser156, thus converting D419-prothrombin into D419N-thrombin and also liberating Fragment-1, Fragment-2 and Fragment-1/2 activation peptide . Activation of r-prothrombin by E.carinatus and O.scutellatus venoms demonstrated the autocatalytic potential of prothrombin-derived molecules and indicated that meizothrombin hydrolysed the cleavage between Fragment-2 and thrombin A-chain in the meizothrombin molecule, but not in prothrombin, preferentially at position Arg284-Thr285 . By contrast, both meizothrombin and thrombin exhibited no detectable activity towards peptide bond Arg320-Ile321 between thrombin A- and B-chain, although this site exhibits the optimum sequence for thrombin cleavage. Yeast, 1996 Oct, 12(13), 1367 - 75 Sequence and analysis of an aldose (xylose) reductase gene from the xylose-fermenting yeast Pachysolen tannophilus; Bolen PL et al.; A xylose reductase gene was isolated from the xylose-fermenting yeast Pachysolen tannophilus as a cDNA clone by selecting clones that hybridized specifically to xylose-inducible messenger RNA . Use of the cDNA clone as a probe in Northern hybridizations identified a xylose-inducible mRNA species large enough to encode a 36 kDa xylose reductase protein known to be produced by this yeast . A corresponding genomic clone was isolated as a 3 kb EcoRI fragment that specifically hybridized to the cDNA clone . The sequence of the cDNA and the largest open reading frame of the genomic clone are identical . The predicted translation product exhibits: (1) significant sequence identity with a previously published N-terminal amino acid sequence from purified P . tannophilus xylose (aldose) reductase protein exhibiting NADH/NADPH-dependent activities (aldose reductase, EC 1.1.1.21); (2) identity with a protein composed of 317 amino acid residues with a calculated molecular mass of 36.2 kDa, equivalent to that reported for purified P . tannophilus xylose reductase; and (3) considerable sequence similarity to, and features of, a superfamily of oxidoreductases. J Dairy Sci, 1996 Oct, 79(10), 1774 - 80 Effect of the degradation of organic matter and crude protein on ruminal fermentation in dairy cows; Arieli A et al.; The potential of the dacron bag technique to assess fluctuations in ruminal metabolites was studied using 40 Israeli-Friesian dairy cows assigned to an experiment with a 2 x 2 factorial design . Diets contained a low (62%) or high (65%) percentage of ruminally degradable CP and a low (55%) or high (59%) percentage of ruminally degradable OM . Metabolites were monitored before feeding and at 3 and 6 h postfeeding . Before feeding, total VFA and propionate were higher, and acetate and pH were lower, in diets containing a high percentage of ruminally degradable OM than in diets containing a low percentage of degradable OM . By 3 H postfeeding, acetate, butyrate and pH were lower, and propionate was higher, in the diets containing a high percentage or ruminally degradable OM than in the diets containing a low percentage of ruminally degradable OM . By 6 h postfeeding, propionate was higher, and acetate was lower, in diets containing a high percentage of ruminally degradable OM than in diets containing a low percentage of ruminally degradable OM . In the diets with a high percentage of ruminally degradable OM, before feeding and by 3 h postfeeding, ammonia concentrations were higher and lower, respectively, relative to the diets containing a low percentage of degradable OM . Milk yield and composition and DMI were similar among treatments . The correlation was good between the degradability data obtained by the dacron bag technique and the meal-induced variations in ruminal metabolites . The lack of a positive yield response to controlled fluctuations in ruminal metabolites may be related to surplus CP intake. J Dairy Sci, 1996 Oct, 79(10), 1767 - 73 Effects of an enzyme additive on composition of corn silage ensiled at various stages of maturity; Sheperd AC et al.; Corn forage at the milk, soft dough, and black layer stages of maturity (22, 28, and 44% DM, respectively) was treated with an enzyme additive at 1, 10, or 100 times the recommended dose and ensiled in l