Synthesis of L-asparaginase bySerratia marcescens (Nima)

The production of L-asparaginase by two mutants ofSerratia marcescens grown on 14 different media was studied. The enzyme content increased from trace levels to 2.4 international units per ml when the organisms were grown in glycerol-peptone yeast extract medium. Glucose was the best carbon source under aerobic conditions. The enzyme content increased when L-asparagine was present in the growth medium.     

Alcoholic fermentation of cellulose hydrolysate by Zymomonas mobilis

Summary Growth and ethanol production by three strains of the bacterium Zymomonas mobilis were tested, at 40°C, in a medium containing cellulose hydrolysate (hexose fraction) as carbon source. The thermotolerant mutant C107 exhibited the best growth compared to wild type ZM4 and to the osmotolerant mutant SBE15. When cultivated in media supplemented with various nutrients, growth was only observed in presence of yeast extract (10 g/l) which acted both as a vitamin supplier and pH stabilizer. Using calcium pantothenate instead of yeast extract and sodium acetate to control pH resulted in growth inhibition by the high medium osmolality. Batch fermentation with pH control (KOH addition) showed good growth and ethanol production with the mineral medium.     

Urea enhances cell lysis of Schizosaccharomyces pombe ura4 mutants

Cell lysis is induced in Schizosaccharomyces pombe ?ura4 cells grown in YPD medium, which contains yeast extract, polypeptone, and glucose. To identify the medium components that induce cell lysis, we first tested various kinds of yeast extracts from different suppliers. Cell lysis of ?ura4 cells on YE medium was observed when yeast extracts from OXOID, BD, Oriental, and Difco were used, but not when using yeast extract from Kyokuto. To determine which compounds induced cell lysis, we subjected yeast extract and polypeptone to GC-MS analysis. Ten kinds of compounds were detected in OXOID and BD yeast extracts, but not in Kyokuto yeast extract. Among them was urea, which was also present in polypeptone, and it clearly induced cell lysis. Deletion of the ure2 gene, which is responsible for utilizing urea, abolished the lytic effect of urea. The effect of urea was suppressed by deletion of pub1, and a similar phenotype was observed in the presence of polypeptone. Thus, urea is an inducer of cell lysis in S. pombe ?ura4 cells.     

The induction of mycelial development inAureobasidium pullulans (IMI 45533) by yeast extract

Germ tube and subsequent mycelial development from yeast-like and swollen cells ofAureobasidium pullulans (IMI 45533) was induced by yeast extract in defined liquid medium. This morphogenetic transition was dependent on inoculum size; pH effects were not involved and once mycelial development was induced in the cells it continued even in the absence of yeast extract. The progeny of mycelium and future generations were unaffected by yeast extract. Cessation of germination was not due to any obvious medium changes but appeared to be partly due to the production of a germination inhibitor, which could also be produced by control cells grown in the absence of yeast extract.     

Influence of Zn2+ and Fe3+ on polysaccharide production and mycelium/yeast dimorphism of Aureobasidium pullulans in batch cultivations

Cultivation of Aureobasidium pullulans in medium with a low concentration of yeast extract (0.4 g/l) led to a decrease in the growth rate early in the fermentation as compared to cultivations in medium with high concentration of yeast extract. When this medium was supplemented with zinc and iron the cultivation closely resembled that obtained in medium with high concentration of yeast extract (4.0 g/l). The culture retained a high growth rate throughout the fermentation and the initiation of the mycelial to yeast (M-Y) transition and the exopolysaccharide production was delayed. In a defined medium or in defined medium without iron only a little exopolysaccharide was produced and the yeast fraction of the total biomass at the onset of the stationary phase was 22%–25%. However, cultivation in the defined medium without zinc resulted in a high production of exopolysaccharide and an increased intensity of the M-Y transition, which led to a yeast fraction of 41%.     

II. Evidence of the presence in yeast extract of substances which stimulate the growth of Histoplasma capsulatum and blastomyces dermatitidis similarly to that found in starling manure extract

Summary A medium consisting of agar plus yeast extract contained the necessary metabolites for rapid growth and sporulation ofHistoplasma capsulatum andBlastomyces dermatitidis. H. capsulatum when harvested after 10 or 30 days incubation period from this medium was shown to have a similar number of spores as well as total particle viability for each period of growth.The growth characteristics ofH. capsulatum and four different isolates ofB. dermatitidis on yeast extract medium were similar to that obtained previously using starling (Sturnis vulgaris) manure extract medium. These characteristics are rapid growth consisting of many viable spores and a low ratio of vegetative mycelium.Several isolations ofH. capsulatum from naturally contaminated soil specimens were made using yeast extract medium.From the Communicable Disease Center, Public Health Service, U. S. Department of Health, Education, and Welfare.     

Application of a simple yeast extract from spent brewer's yeast for growth and sporulation of Bacillus thuringiensis subsp. kurstaki: a Physiological Study

The suitability of using a simple brewer's yeast extract (BYE), prepared by autolysis of complete beer slurry, for growth and sporulation of Bacillus thuringiensis kurstaki was studied in baffled shake flasks. In a standard buffered medium with 2.5% (w/v) glucose and 1% (w/v) brewer's yeast extract, growth of B. t. kurstaki resulted in a low biomass production with considerable byproduct formation, including organic acids and a concomitant low medium pH, incomplete glucose utilization and marginal sporulation, whereas growth in the same medium with a commercial laboratory-grade yeast extract (Difco) resulted in a high biomass concentration, complete glucose utilization, relatively low levels of byproducts and complete sporulation (2.6 × 10⁹ spores/ml). When glucose was left out of the medium, however, growth parameters and sporulation were comparable for BYE and commercial yeast extract, but absolute biomass levels and spore counts were low. Iron was subsequently identified as a limiting factor in BYE. After addition of 3 mg iron sulphate/l, biomass formation in BYE-medium more than doubled, low byproduct formation was observed, and complete sporulation occurred (2.8 × 10⁹spores/ml). These data were slightly lower than those obtained in media with commercial yeast extract (3.6 × 10⁹spores/ml), which also benefited, but to a smaller extent, from addition of iron.     

Study of docosahexaenoic acid production by the heterotrophic microalga <Emphasis Type="Italic">Crypthecodinium cohnii</Emphasis> CCMP 316 using carob pulp as a promising carbon source

In this work, carob pulp syrup was used as carbon source in C. cohnii fermentations for docosahexaenoic acid production. In preliminary experiments different carob pulp dilutions supplemented with sea salt were tested. The highest biomass productivity (4 mg/lh) and specific growth rate (0.04/h) were observed at the highest carob pulp dilution (1:10.5 (v/v), corresponding to 8.8 g/l glucose). Ammonium chloride and yeast extract were tested as nitrogen sources using different carob pulp syrup dilutions, supplemented with sea salt as growth medium. The best results were observed for yeast extract as nitrogen source. A C. cohnii fed-batch fermentation was carried out using diluted carob pulp syrup (1:10.5 v/v) supplemented with yeast extract and sea salt. The biomass productivity was 420 mg/lh, and the specific growth rate 0.05/h. Under these conditions the DHA concentration and DHA production volumetric rate attained 1.9 g/l and 18.5 mg/lh respectively after 100.4 h. The easy, clean and safe handling of carob pulp syrup makes this feedstock a promising carbon source for large-scale DHA production from C. cohnii. In this way, this carob industry by-product could be usefully disposed of through microbial production of a high value fermentation product.     

Yeast Extract from Express Five Serum-free Medium contains Factors at about 35 kDa,Essential for Growth of <Emphasis Type="Italic">Trichoplusia ni</Emphasis> Insect Cells

The yeast extract (of unknown origin) present in the commercially available serum-free medium ‘Express Five’ contains factors (‘yeast extract factors’) up to 35 kDa which are essential for growth of Trichoplusia ni insect cells. A yeast extract brand lacking these components could not support growth of T. ni cells. However, cell proliferation was restored by adding chromatographic fractions containing the yeast extract factors. The yeast extract factors were not solely responsible for the growth enhancing effect of yeast extract but some other components, which seem to be generally present in yeast extracts, are also required for T. ni proliferation.     

Utilization of mustard waste isolates for improved production of astaxanthin by Xanthophyllomyces dendrorhous

Astaxanthin production in the wild strain Xanthophyllomyces dendrorhous TISTR 5730 was investigated using different mustard waste media, including mustard waste residue extract (MRE), mustard waste residue hydrolysate (MRH), mustard waste precipitated extract (MPE), and mustard waste precipitated hydrolysate (MPH). The growth of X. dendrorhous and the production of astaxanthin were dependent on the type and initial concentrations of mustard waste media. The MPH medium was the best substrate resulting in yields of biomass and astaxanthin of 19.6 g/L and 25.8 mg/L, respectively, under optimal conditions. MPH medium improved astaxanthin production 11-fold compared to the commonly used commercial yeast malt medium, and 1.3–2.1-fold compared to other mustard waste media.     

Fermentation study for the production of hepatitis B virus pre-S2 antigen by the methylotrophic yeastHansenula polymorpha

Summary Various physico-chemical parameters have been studied in order to improve the production of hepatitis B virus pre-S2 antigen (middle surface antigen) by the methylotrophic yeastHansenula polymorpha. Antigen production was done in two steps: first, production of cells on glycerol (Phase 1), followed by induction of antigen expression with methanol (Phase 2). Dense cultures ofH. polymorpha, equivalent to 35–40 g/l (dry weight), were readily obtained in small fermenters using minimal medium containing glycerol as carbon source. Antigen expression in this minimal medium, after induction with methanol, was however low and never exceeded 1.6 mg/l of culture. Antigen production was greatly enhanced by adding complex organic nitrogen sources along with methanol at induction time; yeast extract was the best of all the sources tested. In shake flasks, antigen production was proportional to yeast extract concentration up to 7% (w/v) yeast extract. it became clear that the nutritional conditions for good antigen expression were different from those for good biomass production. The effects of yeast extract were reproduced in small fermenters: antigen levels reached 8–9 mg/l in medium containing 6% (w/v) yeast extract during induction with methanol. The mechanisms of yeast extract's effects are still unknown but are probably nutritional. The recombinantH. polymorpha strain produced both periplasmic and intracellular antigen. The periplasmic antigen was shown to be present as 20–22-nm particles and was therefore immunogenic. Immunoblotting indicated that part of the pre-S2 antigen was present as a 24-kDa degradation product. These studies have led to a 140-fold increase in volumetric productivity of antigen and to a 4.6-fold increase in specific production.Part of these results have been presented at the Deuxième Congrès de la Société Française de Microbiologie, Strasbourg, France, September 1989.     

Pectin lyase production by Penicillium griseoroseum: Effect of tea extract, caffeine, yeast extract, and pectin

Summary Mycelial growth and production of extracellular pectin lyase by Penicillium griseoroseum at different concentrations of inducers were investigated. The fungus was cultured in mineral medium using sucrose as a carbon source and caffeine, yeast extract, tea extract or pectin as inducers. Caffeine, yeast extract and tea extract in the presence of sucrose, and tea extract alone were capable of inducing pectin lyase in P. griseoroseum, even at low concentrations.     

Studies on Osmophilic Yeasts

Various conditions were studied which gave influences on polyalcohol production by Pichia miso. Pichia miso gave an excellent yield of polyalcohol showing good growth in the vitamin-free medium. Unlike Zygosaccharomyces high concentrations of phosphate such as 2% as KH₂PO₄ in the medium showed no detrimental effect on polyalcohol production. Remarkable reduction in polyalcohol yield was observed in the medium of high concentration of nitrogen sources, among which yeast extract showed the most striking effect. Polyalcohol fermentation was provoked in the medium of 0.1% yeast extract while the metabolic activity distinctly converted to ethanol fermentation when the organisms were incubated in the medium of 4.0% yeast extract. The fact that a large amount of ethanol more than 6% in the medium was produced aerobically by genus Pichia of oxidative dissimilation type seemed to be very interesting and noticeable. For the purpose of industrial production, it was shown that polyalcohol production in jar-fermenter scale was achieved with as good yield as that in shaking flask culture.     

New nonsynthetic medium for Rhizobium culture production from wastes

A nonsynthetic medium was formulated for placement of mannitol fully by saccharified pea husk (Pisum sativum L.) and water hyacinth (Eichhoornia crassipes) with Trichoderma viride QM 9414 and molasses. Yeast extract was Partially replaced by proteolysed pea husk, water hyacinth, and mycelium of T. viride QM 9414 by boiling 4 hr with 5% (v/v) HCl. The rhizobial growth was equal in both standard yeast extract mannitol (YEM) and formulated nonsynthetic media. However, barring Rhizobium phaseoli (urid) E-6, the rhizobial counts in thenon-synthetic medium were higher then the counts in YEM medium. In the fermentor, rhizobial growth was also almost equal to YEM medium. These results indicated that costly ingredients like mannitol and yeast extract can be replaced by hydrolysates of pea husk, water hyacinth, mycelium of T. viride, and molasses.     

Effect of yeast extract concentration on growth ofSchizoccharomyces pombe

Summary The effect of yeast extract on the growth ofSchizosaccharomyces pombe was investigated using a complex-synthetic medium. Batch cultures at low-glucose concentration show that a too low concentration of yeast extract may limit the biomass formation. On this medium kinetics and yields were found to be similar to those obtained on a synthetic-defined medium under both aerobic and anaerobic conditions.     

Enhanced production of poly(3-hydroxybutyrate-co-4-hydroxybutyrate) copolymer and antimicrobial yellow pigmentation from Cupriavidus sp. USMAHM13 with antibiofilm capability

Antibiofilm polymers have the ability to inhibit bacterial biofilm formation, which is known to occur ubiquitously in the environment and pose risks of infection. In this study, production of P(3HB-co-4HB) copolymer and antimicrobial yellow pigment from Cupriavidus sp. USMAHM13 are enhanced through medium optimization. Before the improvement of yellow pigment production, screening for the best additional supplement was performed resulting in high-yield yellow pigmentation using yeast extract with optimum concentration of 2?g/L. Effects of different concentrations of 1,4-butanediol, ammonium acetate, and yeast extract were studied using central composite design. Under optimal conditions, 53?wt% of polyhydroxyalkanoate (PHA) content, 0.35?g/L of pigment concentration, and 5.87?g/L of residual biomass were achieved at 0.56?wt% C of 1,4-butanediol, 1.14?g/L of ammonium acetate, and 2?g/L of yeast extract. Antibiofilm tests revealed that the yellow pigment coated on P(3HB-co-4HB) copolymer had significant effect on the inhibition of bacteria proliferation and colonization from 6?hr onward reaching 100% inhibition by 12?hr, hence effectively inhibiting the biofilm formation.     

Characterization of optimal growth conditions and carotenoid production of strain Rhodotorula mucilaginosa HP isolated from larvae of Pieris rapae

Yeasts have been studied because of their production of a pigment known as carotenoid with potential application in food and feed supplements. A carotenoid‐producing yeast was isolated from the larvae of Pieris rapae, named HP. The strain HP was identified as Rhodotorula mucilaginosa classified by its carbohydrate fermentation pattern and physiological tests. Rhodotorula mucilaginosa HP produces several exogenous enzymes: alkaline phosphatase, esterase, leucine arylamidase, valine arylamidase, acid phosphatase and β‐glucosidase. Using response surface methodology, selected medium components (yeast extract, malt extract, peptone, glucose) were tested to find the optimum conditions for carotenoid production and the growth of R. mucilaginosa HP. Central composite design was used to control the concentrations of medium components. Peptone and glucose had the largest effects on carotenoid production and cell growth of R. mucilaginosa HP, respectively. The estimated optimal growth conditions of R. mucilaginosa HP were: yeast extract 3.23%, malt extract 2.84%, peptone 6.99% and glucose 12.86%. The estimated optimal conditions for carotenoid production were: yeast extract 2.17%, malt extract 2.11%, peptone 5.79% and glucose 12.46%. These results will assist in the formulation of an appropriate culture medium for optimal carotenoid production of R. mucilaginosa HP for commercial use.     

Enhanced Biosynthesis of Sulfide Oxidase by Arthrobacter Species Using Response Surface Methodology

Response surface methodology was used to develop a fermentation medium for the enhanced biosynthesis of a novel sulfide oxidase by Arthrobacter species strain FR‐3. The interactive effect of the medium components – such as glucose as the carbon source, and tryptone and yeast extract as the nitrogen source – was evaluated by a 2³‐factorial central composite statistical design. Glucose and yeast extract were found to be the more influential medium constituents compared to tryptone since they had lower coefficients of linear effect, P‐values (< 0.02). The optimal fermentation medium components for the enhanced production of sulfide oxidase were recorded as glucose (8.98 g/L), tryptone (10.62 g/L) and yeast extract (7.3 g/L). Optimization of the medium constituents increased the experimental enzyme yield by 54 % compared to the unoptimized medium. This is the first report on the overproduction of sulfide oxidase by using response surface methodology.     

Cultivation of the hyperthermophilic archaeonSulfolobus solfataricus in low-salt media

Two low-salt complex media, bactopeptone and desalted yeast extract, were used for high density cultivation of the hyperthermophilic archaeonSulfolobus solfataricus (DSM 1617). Bactopeptone, which has low mineral ion content among various complex media, was good for cell growth in batch cultures; the maximal cell density in bactopeptone was comparable to that in yeast extract. Howver, cell growth was rather poor when bactopeptone was added by the fed-batch procedure. Since several vitamins are deficient in bactopeptone, the effect of vitamins on cell growth was examined. Among the vitamins tested, pyridoxine was found to improve the growth rate ofS. solfataricus. To reduce the growth inhibition caused by mineral ions, yeast extract was dialyzed against distilled water and then fed-batch cultures were carried out using a feed medium containing desalted yeast extract. Although the concentrations of mineral ions in yeast extract were significantly lowered by the dialysis procedure, fed-batch cultivation with desalted yeast extract was unsatisfactory. To examine whether low molecular weight solutes in yeast extract are crucial for cell growth, we investigated the effect of trehalose, a most abundant compatible solute in yeast extract, on the growth pattern. Cell densities were increased and the length of the lag phase was markedly shortened by the presence of trehalose, indicating that trehalose plays an important role in the growth ofS. solfataricus.     

Tea extract as an inexpensive inducer of pectin lyase in Penicillium griseoroseum cultured on sucrose

Extracts of tea, coffee, cocoa, and yeast induced pectin lyase (PL) in Penicillium griseoroseum cultured in a mineral medium with sucrose as the carbon source. PL activity and fungal growth were similar in the treatments with 0.5% tea extract, the highest concentration tested, and 0.03% yeast extract. When tea extract was added singly to the culture medium, P. griseoroseum produced 59% and 17% of the PL activity and mycelial mass, respectively, obtained in a treatment with tea extract and sucrose. These results suggest that the production of the enzyme was not proportional to mycelial growth. No PL was produced in the medium with sucrose and without inducers. The small amounts of pectic substances present in the tea extract could not be responsible for PL induction. PL activity was detected after 12 h of growth in the medium containing sucrose and tea extract added at time zero, and after 48 h of growth when tea extract was added at times 12 and 24 h. Mycelial mass in all treatments was similar after 48 h of incubation. However, the addition of tea extract at time zero increased PL activity by 20–25%. Cyclic AMP at 5 and 10 mM in the culture medium induced 20 and 30%, respectively, of the PL activity obtained with 0.03% yeast extract, suggesting that PL induction brought about by either yeast extract or tea extract might involve the intracellular metabolism of cAMP. Received 22 October 1996/ Accepted in revised form 09 January 1997