Valine production from hydrocarbon byMicrococcus varians

A bacterium isolated from Assam (India) soil was found to accumulateL-valine in the growth medium and was identified asMicrococcus varians. The strain grew and accumulated valine in a purely synthetic medium, but supplementation with either casamino acids or yeast extract or with both, improved the yield. The entire fermentation period could be divided into a growth phase and a production (phase which could be prolonged by adjustment of pH to neutral range. Among the different hydrocarbon and nitrogen sources tested straight run gas-oil and ammonium sulphate, respectively, were found most suitable. Antibiotics inhibited growth but stimulated extracellular valine accumulation. Vitamins stimulated growth and valine yield and an inoculum level of 10 % was found to be optimal. The yield ofL-valine under optimal conditions was 2.95 g/L.     

Production of l-glutamic acid by aBacillus sP

A strain ofBacillus cereus var.mycoides isolated from Burdwan soil producesl-glutamate in the medium. The strain is able to grow and produce in a synthetic medium but supplementation with casamino acid or yeast extract improves the yield. Maintenance of pH of the fermentation medium near neutrality prolongs the active growth period and improves the yield. Glucose and ammonium nitrate were found to be most suitable carbon and nitrogen sources, respectively. Cane sugar molasses (as a substitute for glucose) significantly stimulated the growth but glutamate production was less. Various B vitamins stimulate the growth and glutamate yield. The yield of glutamate under optimal condition is 5.2 g/l.     

Lysine production from hydrocarbon by micrococcus species

A bacterium isolated from Assam (India) soil was found to accumulate L-lysine in the mineral salt-hydrocarbon medium and identified to be a strain of Micrococcus luteus. The strain is able to grow and accumulate l-lysine in a purely synthetic medium, but supplementation of the synthetic medium with casamino acid or yeast extract or both, improves the yield. The entire fermentation period can be divided into a growth phase and a production phase, which can be prolonged by adjustment of the pH to the neutral range. Among the different hydrocarbon and nitrogen sources tested straight run gas oil (47percnt;) and ammonium sulphate (0.4%), respectively, were found most suitable. Erythromycin at 1 μg/ml level inhibited growth bu¸t stimulated lysine excretion. An inoculum level of 10% (v/v) of the medium was optimal for lysine production. The yield of lysine under optimal conditions was found to be 3.25 g per litre of the medium. Lysine has been isolated in crystalline form from the fermented broth by ion exchange resin chromatography and found to be pure sample of L-isomer.     

Influence of antibiotics and trace salts on lysine production by arthrobacter globiformis

A hydrocarbon utilizing strain of Arthrobacter globiformis Lb isolated from local soil has been found to yield lysine 3.4 g l^?1, keeping the medium optimal for pH, C- and N-sources. Addition of antibiotics and micronutrients to that optimal media stimulated cell growth and enhanced lysine yield.     

Production of glutamic acid byArthrobacter globiformis: Influence of cultural conditions

Arthrobacter globiformis isolated from Burdwan soil excretes glutamic acid in a glucose mineral salt medium with suboptimal level of biotin. Glutamate begins to accumulate in the medium from the mid-exponential phase of growth and its excretion could be prolonged by adjustment of pH to the neutral range. Among the different carbon and nitrogen sources tested glucose (8%) and ammonium nitrate (0.53%), respectively, were found to be most suitable. Molasses could not be used as a substitute for glucose even if antibiotics or Tween 80 are incorporated in the medium. Bacitracin (1 μg/mL) stimulated glutamate excretion. Atemperature of 28°C and an inoculum dose of 4% were optimal for production. Under optimal conditions, in the flask culture the isolate excreted 16.1 g glutamic acid per litre in 120 h. Glutamic acid isolated from the fermented broth was found to be purel-diastereomer.     

Regulation of differentiation of the dimorphic fungusPaecilomyces viridis by nitrogen sources,antibiotics and metabolic inhibitors

The effect of nitrogen and carbon sources, vitamins, antibiotics and metabolic inhibitors on growth and differentiation ofPaecilomyces viridis was investigated. Sodium nitrate,l-asparagine,l-proline and peptone were found to be suitable nitrogen sources for mycelial growth (M) in a synthetic medium with glucose.Paecilomyces viridis could also grow slowly in a synthetic medium containing benzylpenicillin or bacitracin as the only nitrogen sources and very slowly even in a medium with polymyxin as the nitrogen source. Ammonium salts, area,l-arginine,d, l-aspartic acid andl,-serine were found to support intensive sporulation. Partially yeast-like growth (Y) was facilitated by NaNO₂, (NH₄)₂SO₄, NH₄NO₃, urea,d, l-alanine,l-arginine,d, l-aspartic acid,l-cysteine,l-glutamic acid andl-serine. Partially yeastlike growth could be observed in a medium with peptone and at an initial pH of 2. The following compounds appear as suitable carbon sources for mycelial growth:d-glucose,d-galactose,d-mannose, maltose, sucrose, chitin andd-mannitol. No changes in morphology could be detected on any of the 25 used carbon sources in a synthetic medium with NaNO₃. Yeast-like growth was induced by the antibiotics azalomycin F, cyanein (brefeldin A), griseofulvin and monorden (radicicol). After removal of the antibiotics, mycelial growth was restored. Sporulation was stimulated by chloramphenicol, 2-deoxy-d-glucose, furancarboxylic acid and stipitatic acid. Deformation of phialides was observed after treatment with actinomycin D, amphotericin B, boromycin, citrinin, cycloheximide, cytochalasin D, fungicidin and scopathricin. Microcyclic conidiation or growth of phialides directly from conidia were induced by cycloheximide, desertomycin, ethidium bromide and 5-fluorouracil.     

Production ofl-phenylalanine by double auxotrophic mutants ofArthrobacter globiformis: Effect of temperature,trace salts and inoculum dose

Three tryptophan-plus-tyrosine double auxotrophic mutants were isolated from a biotin-requiring glutamate-producingArthrobacter globiformis. The mutants were found to producel-phenylalanine in a mineral salt medium. Further improvement ofl-phenylalanine production was achieved by isolation of mutants resistant to β-2-thienylalanine from these double auxotrophs. Temperature of 30 °C and a 4% inoculum dose were found to be optimum for phenylalanine production. Addition of some trace salts does not enhance phenylalanine yield. Under optimal cultural conditions one mutant yielded 6.8 g phenylalanine per L medium in flask culture.     

Nutritional requirements ofXanthomonas phaseoli var.fuscans

As found by Starr (1946),l-glutamic acid is necessary for the growth ofXanthomonas phaseoli var.fuscans. According to our results, the growth is stimulated byl-asparagine in the presence ofl-glutamic acid;l-asparagine itself, however, does not serve as a source of carbon and nitrogen.Xanthomonas phaseoli var.fuscans grew well in a medium containing tryptone. Some peptides of the acidic fraction isolated from tryptone affected the growth as much as tryptone itself. Vitamins and plant growth substances did not affect the growth of the bacteria; proteins appeared to be a poor carbon and nitrogen source. On substituting glucose in a glutamic acid-containing medium with another saccharide, the growth of the bacteria was found equal or better in media containing mannose, sucrose, fructose, maltose or starch. The bacteria grew less satisfactorily in media containing galactose and cellobiose as compared with media containing glucose.     

Production of valine by a Bacillus sp.

A bacterium isolated from Burdwan (India) soil was found to accumulate L-valine in the growth medium and identified to be a strain of Bacillus subtilis. The strain is able to grow and accumulate valine in a purely synthetic medium, but supplementation of the synthetic medium with either Casamino acids or yeast extract or with both, significantly improves the yield. The entire fermentation period can be divided into a growth phase and a production phase, which can be prolonged by adjustment of pH to the neutral range. Among the different carbon and nitrogen sources tested glucose at 8.5% and L-glutamic acid at 0.8%, respectively, were found most suitable. Cane sugar molasses tested as a substitute for glucose significantly stimulated growth but valine production was less. Different vitamins tested stimulated growth and valine yield and an inoculum level of 10% (v/v) of the medium was found to be optimal. The yield of valine under optimal conditions was found to be 4.53 g per litre of the medium. Valine has been isolated in crystalline form from the fermented broth by ion exchange resin chromatography and found to be a pure sample of the L-isomer.     

Influence of B-vitamins and trace elements on lysine production by micrococcus varians 2Fa

A hydrocarbon utilizing strain of Micrococcus varians 2Fa isolated from local soil has been found to yield lysine 2.6 g l^?1. Addition of B-vitamins and trace elements to the optimal media, has been found to stimulate growth and enhance lysine yield.     

Biological active metabolite cyclo (l-Trp-l-Phe) produced by South China Sea sponge Holoxea sp. associated fungus Aspergillus versicolor strain TS08

Sponge-associated fungi represent the single most prolific source of novel natural products from marine fungi. Cyclo (l-Trp-l-Phe) exhibits biological functions such as plant growth regulation, moderate cytotoxicity and thus has the application potential in pharmaceutical and agricultural biotechnologies. In this study, a fungal strain TS08 was isolated from sponge Holoxea sp. in the South China Sea and identified as A. versicolor according to its 18S rRNA gene and morphological, physiological, and biochemical characteristics. Meanwhile, cyclo (l-Trp-l-Phe) was found to be produced by A. versicolor strain TS08 mainly in the exponential growth phase. The highest yield of cyclo (l-Trp-l-Phe), 13.24 mg/g (per crude extract of EtOAc), 2.51% of cell dry weigh, was obtained on the tenth day of the fungal cultivation. It was the first time to find the biological active cyclo (l-Trp-l-Phe) in sponge-associated microorganism.     

Production of 3-indoleacetic acid by aRhizobium sp. fromMimosa pudica

ARhizobium species isolated from the root nodules of the sensitive plant,Mimosa pudica, produced 60 mg/L of 3-indoleacetic acid (IAA) froml-tryptophan in culture. The production of IAA started simultaneously with the growth and had no different growth or production phase. The stationary phase of growth was reached after 55 h, but the production of IAA increased gradually up to 80 h, and then remained constant. The IAA production could be promoted in the culture medium up to 365% by supplementing the medium with maltose, CuSO₄ and Triton X-100.     

Regulation of lysine and dipicolinic acid biosynthesis in Bacillus brevis ATCC 10068: significance of derepression of the enzymes during the change from vegetative growth to sporulation

Lysine biosynthetic pathway enzymes of Bacillus brevis ATCC 1068 were studied as a function of stage of development (growth and sporulation). The synthesis of aspartic-2-eemialdehyde dehydrogenase (ASA-dehydrogenase), dihydrodipicolinate synthase (DHDPA-synthase), DHPA-reductase and diaminopimelate decarboxylase (DAP-decarboxylase) was found not to be co-regulated, since lysine was not a co-repressor for these enzymes. Unlike the aspartokinase isoenzymes, the other enzymes of the lysine pathway were not derepressed in thiosine-resistant, lysine-excreting mutants. Thus, the aspartokinase isoenzymes were the key enzymes during growth and regulation of lysine biosynthesis through restriction of l-ASA synthesis via feedback control by lysine on the aspartokinases was therefore suggested.In contrast to other Bacillus species, the levels of the lysine biosynthetic pathway enzymes of strain ATCC 10068 were not derepressed during the change from vegetative growth to sporulation. Two control mechanisms, enabling the observed preferential channelling of carbon for the synthesis of spore-specific diaminopimelic acid (DAP) and dipicolinic acid (DPA) were a) loss of DAP-decarboxylase, b) inhibition of DHDPA-reductase by DPA. Increase in the level of the DAP pool during sporulation, as a consequence of the loss of DAP-decarboxylase, and its relevance to the non-enzymatic formation of DPA has been discussed.Abbreviations l-ASA l-aspartic-2-semialdehyde - DAP diaminopimelic acid - DPA dipicolinic acid - DHDPA dihydrodipicolinate - AGM aspargine-glycerol medium - PY peptone-yeast extract - NB+NSM nutrient broth plus nutrient sporulation medium     

Influence of amino acids on the biosynthesis of cyclosporin A by Tolypocladium inflatum

 An indigenously isolated strain of Tolypocladium inflatum, when grown as a suspension culture in semi-synthetic and synthetic media, produced cyclosporin A. Biosynthesis of this well-known immunosuppressive agent was found to be influenced heavily by the external addition of the amino acid constituents of the molecule. In synthetic media, L-leucine and L-valine were found to act as strong inducers of drug production. L-Valine increased the specific production of cyclosporin A by 75% in semi-synthetic medium and by ten times in synthetic medium compared to an unsupplemented control culture. D-Valine had no stimulating effect on the production. The presence of amino acids in the exponential growth phase ensured optimal production, as was indicated in the experiment in which L-valine was added at different times; 4 g/l was the optimum concentration of exogenous L-valine. On the other hand, exogenous sarcosine and L-methionine tended to diminish drug production. Received: 23 October 1995/Received revision: 23 January 1996/Accepted: 29 January 1996     

Enzymatic production of <Emphasis Type="SmallCaps">l</Emphasis>-amino acids from the corresponding <Emphasis Type="SmallCaps">dl</Emphasis>-5-substituted hydantoins by <Emphasis Type="Italic">Bacillus fordii</Emphasis> MH602

Bacillus fordii MH602 was newly screened from soil at 45 °C and exhibited high activities of hydantoinase and carbamoylase, efficiently yielding l-amino acids including phenylalanine, phenylglycine and tryptophan with the bioconversion yield of 60–100% from the corresponding dl-5-substituted hydantoins. Hydantoinase activity was found to be cell-associated and inducible. The optimal inducer was dl-5-methylhydantoin with concentration of 0.014 mol L⁻¹ and added to the fermentation medium in the exponential phase of growth. In the production of optically pure amino acids from dl-5-benylhydantoin, the optimal temperature and pH of this reaction were 45–50 °C and 7.5 respectively. The hydantoinase was non-stereoselective, while carmbamoylase was l-selective. The hydantoinase activity was not subject to substrate inhibition, or product inhibition by ammonia. In addition, The activities of both enzymes from crude extract of the strain were thermostable; the hydantoinase and carbamoylase retained about 90% and 60% activity after 6 h at 50 °C, respectively. Since reaction at higher temperature is advantageous for enhancement of solubility and for racemization of dl-5-substituted hydantoins, the relative paucity of l-selective hydantoinase systems, together with the high level of hydantoinase and carbamoylase activity and unusual substrate selectivity of the strain MH602, suggest that it has significant potential applications.     

Construction of an arginine-producing strain of Serratia marcescens

Summary In Serratia marcescens Sr41, l-canavanine was demonstrated to be a weak cell growth inhibitor in minimal medium containing glucose as the sole carbon source. The inhibition of cell growth was enhanced by changing the carbon source from glucose to l-glutamic acid. An arginine regulatory mutant (i.e., argR mutant) in which formation of l-arginine biosynthetic enzymes was genetically derepressed was isolated by selecting for l-canavanine resistance on the glutamate medium. Furthermore, an l-arginine-producing strain was constructed by introducing the mutation leading to feedback-resistant N-acetylglutamate synthase into the argR mutant. The resulting transductant produced about 40 g/l of l-arginine, whereas the wild strain produced no l-arginine and the argR mutant only 3 g/l.     

Effects of fumarate,l-malate,and anAspergillus oryzae fermentation extract ond-lactate Utilization by the ruminal bacteriumSelenomonas ruminantium

Growth ofSelenomonas ruminantium HD4 in medium that contained 21mm d-lactate was stimulated to varying degrees by 10mm l-malate, 10mm fumarate, and 2% (v/v)Aspergillus oryzae fermentation extract (Amaferm). Amaferm treatment caused the greatest growth stimulation. Initial uptake rates (30s) and long-term uptake rates (30 min) ofd-lactate by whole cells ofS. ruminantium were increased in the presence of 10mm l-malate. Amaferm (25 l/ml) also stimulated long-term uptake rates ofd-lactate, whereas fumarate had no effect. Initial uptake ofd-lactate was depressed in the presence of fumarate or Amaferm. When eitherl-malate, fumarate, or Amaferm was included in thed-lactate growth medium, a homosuccinate fermentation resulted and an inverse relationship was observed between growth (protein synthesis) and succinate production. Recent research demonstrated that Amaferm containsl-malate, and this dicarboxylic acid may be involved in stimulatingd-lactate utilization byS. ruminantium.     

Histidine decarboxylase production by Lactobacillus hilgardii: Effect of organic acids

Histidine decarboxylase production from Lactobacillus hilgardii 5w, isolated from wine, was inhibited by the presence of l-malic acid in the basal culture medium. The inhibition was related to l-malic acid concentration. The maximal production of the enzyme at 12 h of culture incubated at 30°C was inhibited 71% by 2 g/L l-malic acid and 47% by 0.5 g/L. In these conditions l-malic acid consumption was 16% and 20% respectively. The addition of 300 mg/L citric acid to the basal medium stimulated the enzyme production from 9 to 45 nmoles/min/mg dry weight, and the increase was correlated with citric acid concentration. When different concentrations of l-malic acid were added to the basal medium plus 200 mg/L citric acid, reversion of stimulation was observed, achieving the maximum at a concentration of 2 g/L. In this case, citric acid comsumption was not modified, whereas L-malic acid utilization was higher.     

Culture growth and IAA production by a microbial diazotropic symbiont of stem-nodules of the legumeAeschynomene aspera

The stem nodules of the legumeAeschynomene aspera contain indoleacetic acid and a high amount of tryptophan. TheAzorhizobium caulinodans isolated from the stem nodules of the leguminous emergent hydrophyte produced a high amount of IAA (14.8 mg/L) inl-tryptophan-supplemented basal medium. The IAA yields paralleled the culture growth rate and increased up to 52 h. No separate growth and production phase was observed. The IAA production was increased 344% when the medium was supplemented withl-tryptophan, sucrose, FeSO₄·7H₂O, NaNO₃, ascorbic acid and sodium dodecyl sulfate. The possible role of the IAA production in the legume-bacterium symbiosis is discussed.     

Potential of solid state fermentation for production of L(+)-lactic acid by Rhizopus oryzae

Production of l(+)-lactic acid by Rhizopus oryzae NRRL 395 was studied in solid medium on sugar-cane bagasse impregnated with a nutrient solution containing glucose and CaCO₃. A comparative study was undertaken in submerged and solid-state cultures. The optimal concentrations in glucose were 120 g/l in liquid culture and 180 g/l in solid-state fermentation corresponding to production of l(+)-lactic acid of 93.8 and 137.0 g/l, respectively. The productivity was 1.38 g/l per hour in liquid medium and 1.43 g/l per hour in solid medium. However, the fermentation yield was about 77% whatever the medium. These figures are significant for l(+)-lactic acid production.