Production of cyclodepsipeptides destruxin A and B from Metarhizium anisopliae

Maltose and peptone were the best carbon and nitrogen sources for the production of destruxins from Metarhizium anisopliae. With the addition of 0.1% (w/v) beta-alanine to the basal medium, the yields of cyclodepsipeptides DA and DB were 7.2 and 279 mg/L, respectively, which was 2-fold higher than that of control experiment. Response surface methodology (RSM) was applied to optimize the compositions of maltose, peptone, beta-alanine, and glucose used in a shaker-flask cultivation of M. anisopliae for the production of DA and DB. Estimated optimal compositions for the DA production were maltose 2.58%, peptone 0.72%, beta-alanine 0.02%, and glucose 0.55%. The predicted DA yield was 18.5 mg/L. On the other hand, the optimal compositions for DB production were maltose 2.51%, peptone 0.75%, beta-alanine 0.02%, and glucose 0.43%. A maximum DB yield of 232 mg/L was predicted. These were confirmed by cultivation experiments conducted at the optimized conditions for maximum destruxins production in a shaker-flask. Furthermore, a modest high level of DA (49 mg/L) and DB (268 mg/L) yields were obtained by employing the response surface methodology optimized DB production medium in a no-baffle, stirred-tank fermentor.     

Lovastatin biosynthesis by Aspergillus terreus in a chemically defined medium

Lovastatin is a secondary metabolite produced by Aspergillus terreus. A chemically defined medium was developed in order to investigate the influence of carbon and nitrogen sources on lovastatin biosynthesis. Among several organic and inorganic defined nitrogen sources metabolized by A. terreus, glutamate and histidine gave the highest lovastatin biosynthesis level. For cultures on glucose and glutamate, lovastatin synthesis initiated when glucose consumption levelled off. When A. terreus was grown on lactose, lovastatin production initiated in the presence of residual lactose. Experimental results showed that carbon source starvation is required in addition to relief of glucose repression, while glutamate did not repress biosynthesis. A threefold-higher specific productivity was found with the defined medium on glucose and glutamate, compared to growth on complex medium with glucose, peptonized milk, and yeast extract.     

Lovastatin Biosynthesis by Aspergillus terreus in a Chemically Defined Medium

Lovastatin is a secondary metabolite produced by Aspergillus terreus. A chemically defined medium was developed in order to investigate the influence of carbon and nitrogen sources on lovastatin biosynthesis. Among several organic and inorganic defined nitrogen sources metabolized by A. terreus, glutamate and histidine gave the highest lovastatin biosynthesis level. For cultures on glucose and glutamate, lovastatin synthesis initiated when glucose consumption levelled off. When A. terreus was grown on lactose, lovastatin production initiated in the presence of residual lactose. Experimental results showed that carbon source starvation is required in addition to relief of glucose repression, while glutamate did not repress biosynthesis. A threefold-higher specific productivity was found with the defined medium on glucose and glutamate, compared to growth on complex medium with glucose, peptonized milk, and yeast extract.     

Regulation of heparinase synthesis in Flavobacterium heparinum

The effect of various carbon, nitrogen and sulfur sources on the production of heparinase by Flavobacterium heparinum in defined medium in the presence and absence of heparin as the inducer has been studied. Carbon catabolite repression has been observed in defined medium containing one of several carbon sources including simple sugars, alcohols and organic acids. Fed batch fermentations result in 10 g/l of cells and heparinase titers as high as 100,000 U/l by avoiding carbon catabolite repression. Growth on heparin as a sole carbon source resulted in both a high growth rate of 0.12 h^–1 and a high specific activity of 18 U/mg. Specific heparinase activity was markedly reduced when the end products of heparin catabolism were used as carbon, nitrogen or sulfur sources in defined medium. In defined medium with a low sulfate concentration, of less than 10^–3 M, specific activities as high as 8 U/mg have been observed even in the absence of the normally required inducer, heparin.     

Production of extracellular β-glucosidase by Monascus purpureus on different growth substrates

Various agro-industrial residues in combination with peptone, NH₄Cl and/or soy bran were screened as substrates for extracellular β-glucosidase (BGL) production by Monascus purpureus NRRL1992 on submerged fermentations (SmF). Higher BGL production was achieved when the agro-industrial residues were combined with peptone, and the utilization of NH₄Cl (inorganic nitrogen source) had not supported high enzyme production. The combination between grape waste and peptone was the best for enzyme production, and was selected as the growth substrate for further investigations. The evaluation of the effects of the medium components on enzyme production showed that the influence of peptone was more important than grape waste. The production of extracellular BGL by M. purpureus was inducible and controlled by carbon (glucose) catabolite repression.     

Statistical optimization of lovastatin production by Omphalotus olearius (DC.) singer in submerged fermentation

In this study, culture conditions were optimized to improve lovastatin production by Omphalotus olearius, isolate OBCC 2002, using statistical experimental designs. The Plackett–Burman design was used to select important variables affecting lovastatin production. Accordingly, glucose, peptone, and agitation speed were determined as the variables that have influence on lovastatin production. In a further experiment, these variables were optimized with a Box–Behnken design and applied in a submerged process; this resulted in 12.51 mg/L lovastatin production on a medium containing glucose (10 g/L), peptone (5 g/L), thiamine (1 mg/L), and NaCl (0.4 g/L) under static conditions. This level of lovastatin production is eight times higher than that produced under unoptimized media and growth conditions by Omphalotus olearius. To the best of our knowledge, this is the first attempt to optimize submerged fermentation process for lovastatin production by Omphalotus olearius.     

Influence of cultivation conditions on production of lignocellulolytic enzymes by Ceriporiopsis subvermispora

The aim of this work was to make a survey describing factors that influence the production of extracellular enzymes by white-rot fungus Ceriporiopsis subvermispora responsible for the degradation of lignocellulolytic materials. These factors were: carbon sources (glucose, cellulose, hemicellulose, lignin, maltose and starch), nitrogen sources (ammonium sulphate, potassium nitrate, urea, albumin and peptone), pH, temperature and addition of three different concentrations of Cu²⁺ and Mn²⁺. The cellulase and xylanase activities were similar in medium with different carbon sources and the highest cellulase and xylanase activities were measured in medium with urea and potassium nitrate as nitrogen sources, respectively. The highest laccase activity was observed in medium with lignin and peptone as carbon and nitrogen sources. In other experiments, time course of production of lignocellulolytic enzymes by white-rot fungus C. subvermispora in medium with lignin or glucose as carbon sources was observed.     

Influence of medium design on lovastatin and mevastatin production byAspergillus terreus strains

In order to investigate the influence of medium design on lovastatin and mevastatin production byAspergillus terreus strains, several nitrogen complex sources, such as vegetal flour and peptones of different origin (animal and vegetal) were tested, together with the addition of methionine, an aminoacid that is directly involved in lovastatin biosynthetic pathway. Soybean peptone generally allowed the best lovastatin yields to be achieved (250–280 mg l^?1), particularly in the presence of soybean and peanut flours. For mevastatin, the best results (300–320 mg l^?1) were obtained at 7 days fermentation with modified base medium (CLD), and at 14 days with standard medium (STD), not being possible in this case to associate the best yield with a defined flour and/or peptone. The results show that lovastatin production is influenced by the presence of soybean peptone and by the addition of methionine; instead, the production of mevastatin appears more strictly strain-associated and not directly dependent on the complex ingredients employed.     

Kinetics of α-amylase production in a continuous culture ofBacillus licheniformis

As found during continuous cultivation ofBacillus licheniformis on a semisynthetic medium (glucose or maltose as C source), the specific rate of α-amylase production is proportional to growth rate but is repressed by higher substrate concentrations. Besides glucose or maltose, peptone was also used as an alternative carbon source during cultivation. The specific rate of production of the enzyme on maltose is half that found with glucose.     

Optimization of a liquid medium for beauvericin production in <Emphasis Type="Italic">Fusarium redolens</Emphasis> Dzf2 mycelial culture

Beauvericin (BEA) is a proven and potent antibiotic compound useful for bio-control and a potential antifungal and anticancer agent for human. This study was to evaluate and optimize the nutrient medium for BEA production in mycelial liquid culture of a high BEA-producing fungus Fusarium redolens Dzf2 isolated from a medicinal plant. Among various organic and inorganic carbon and nitrogen sources, glucose and peptone were found the most favorable for the F. redolens Dzf2 mycelial growth and BEA production. Through a Plackett-Burman screening test on a basal medium, glucose, peptone, and medium pH were identified as the significant factors for mycelial growth and BEA production. These factors were optimized through central composite design of experiments and response surface methodology, as 49.0 g/L glucose, 13.0 g/L peptone and pH 6.6, yielding 198 mg/L BEA (versus 156 mg/L in the basal medium). The BEA yield was further increased to 234 mg/L by feeding 10 g/L glucose to the culture during exponential phase. The results show that F. redolens Dzf2 mycelial fermentation is a feasible and promising process for production of BEA.     

Optimization of medium composition for actinorhodin production by Streptomyces coelicolor A3(2) with response surface methodology

Optimization of the fermentation medium for maximization of actinorhodin production by Streptomyces coelicolor A3(2) was carried out. Response surface methodology (RSM) was applied to optimize the medium constituents. A 2⁴ full-factorial central composite design (CCD) was chosen to explain the combined effects of the four medium constituents, viz. sucrose, glucose, yeast extract (YE) and peptone, and to design a minimum number of experiments. The P-values of the coefficients for linear, quadratic and cross-product effect of sucrose and glucose concentration were <0.0001, suggesting that these were critical variables having the greatest effect on the production of actinorhodin in the complex medium. The optimized medium consisting of 339 g/l sucrose, 1 g/l glucose, 1.95 g/l YE and 2.72 g/l peptone predicted 195 mg/l of actinorhodin which was 32% higher than that of the unoptimized medium. The amounts of glucose, YE and peptone required were also reduced with RSM.     

响应面法优化多杀菌素发酵培养基的研究

采用响应面分析方法,对刺糖多孢茵（Saccharopolyspora spinosa）H-2产多杀菌素的发酵培养基进行优化研究。运用单因子试验筛选出葡萄糖和棉籽粉为最适碳源和氮源,通过Plack—ett—Burman设计试验,对影响发酵培养基的8个相关因子进行评估并筛选出具有显著效应的4个因子：葡萄糖、棉籽粉、黄豆饼粉及玉米浆。通过最陡爬坡实验逼近以上4个因子的最大响应区域后,采用Box-Behnken响应面分析法,确定发酵产多杀菌素最佳培养基为葡萄糖64．5g,麦芽糖20g,玉米浆2g,大豆油40g,棉籽粉25g,黄豆饼粉2．4g,蛋白胨25g,CaCO35g,定容至1L,pH7．0。培养基优化后多杀菌素产量由278．1mg／L提高到508．7mg／L,比初始多杀茵素产量提高了1．83倍。     

Sporulation of several biocontrol fungi as affected by carbon and nitrogen sources in a two-stage cultivation system

The development of fungal biopesticides requires the efficient production of large numbers spores or other propagules. The current study used published information concerning carbon concentrations and C:N ratios to evaluate the effects of carbon and nitrogen sources on sporulation of Paecilomyces lilacinus (IPC-P and M-14) and Metarhizium anisopliae (SQZ-1-21 and RS-4-1) in a two-stage cultivation system. For P. lilacinus IPCP, the optimal sporulation medium contained urea as the nitrogen source, dextrin as the carbon source at 1 g/L, a C:N ratio of 5:1, with ZnSO(4)·7H(2)O at 10 mg/L and CaCl(2) at 3 g/L. The optimal sporulation medium for P. lilacinus M-14 contained soy peptone as the nitrogen source and maltose as the carbon source at 2 g/L, a C:N ratio of 10:1, with ZnSO(4)·7H(2)O at 250 mg/L, CuSO(4)·5H(2)O at 10 mg/L, H(3)BO(4) at 5 mg/L, and Na(2)MoO(4)·2H(2)O at 5 mg/L. The optimum sporulation medium for M. anisopliae SQZ-1-21 contained urea as the nitrogen source, sucrose as the carbon source at 16 g/ L, a C:N ratio of 80:1, with ZnSO(4)·7H(2)O at 50 mg/L, CuSO(4)·5H(2)O at 50 mg/L, H(3)BO(4) at 5 mg/L, and MnSO(4)·H(2)O at 10 mg/L. The optimum sporulation medium for M. anisopliae RS-4-1 contained soy peptone as the nitrogen source, sucrose as the carbon source at 4 g/L, a C:N ratio of 5:1, with ZnSO(4)·7H(2)O at 50 mg/L and H(3)BO(4) at 50 mg/L. All sporulation media contained 17 g/L agar. While these results were empirically derived, they provide a first step toward low-cost mass production of these biocontrol agents.     

Use of response surface methodology to optimize culture medium for production of lovastatin by Monascus ruber

Response surface methodology (RSM) was employed to study the effect of culture medium on the production of lovastatin in mixed solid-liquid state (or submerged) cultures by Monascus ruber. The maximal lovastatin yield (131 mg/L, average of three repeats) appeared at the region where the respective concentrations of rice powder, peptone, glycerin, and glucose were around 34.4 g/L, 10.8 g/L, 26.4 ml/L, and 129.2 g/L, respectively. The optimized medium resulted in a significant increase of lovastatin yield, as compared with that obtained by the fermentation of many other M. ruber species.     

Production of statins by filamentous fungi

Several Monascus and Aspergillus strains were screened for statins production. Lovastatin, monacolin J, pravastatin and mevastatin were produced, with higher yields from the A. terreus strains than from Monascus species. Of all the strains investigated M. paxii AM12M, an isolated spontaneous mutant, yielded 127 mg lovastatin/l and 53 mg pravastatin/l at 21 days, and 18 mg pravastatin/l at 16 days employing a whole soybean flour medium; A. terreus BST yielded 230 mg lovastatin/l and 118 mg pravastatin/l at 14 days employing a defatted soybean flour medium. Statins recovery showed that pravastatin was, in both strains, mostly found in both the mycelium and the culture filtrate, while lovastatin remained closely associated (83%) to the A. terreus mycelium or was mainly released into the culture filtrate (64%) of M. paxii culture.     

Effect of nitrogen source and nitrogen concentration on the production of pyruvate by Torulopsis glabrata

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

Production of α-amylase by Myxococcus coralloides D

M.E. FÁREZ-VIDAL, A. FERNANDEZ-VIVAS AND J.M. ARIAS. 1992. Myxococcus coralloides D secreted amylase into a liquid growth medium containing 1% starch. Amylase activity was highest at the end of the exponential growth phase. Of the nitrogen sources tested, the greatest growth and amylase production were obtained with trypticase peptone, casitone, probion L and probion F. When starch was replaced by other carbon sources, amylase production was reduced; trisaccharide produced better results than disaccharide while monosaccharide reduced amylase production to basal levels. Maltose repressed amylase production. Amylase production was greater in stirred flasks, at pH between 6.5 and 7.5, and at temperatures from 28C to 33C. The activity of partially purified M. coralloides D amylase was used to determine the products released from the hydrolysis of starch with thin-layer chromatography, paper chromatography and nuclear magnetic resonance. These products were maltose and glucose and limit dextrins.     

Flocculation onset in Saccharomyces cerevisiae: the role of nutrients

AIMS: To examine the role of the nutrients on the onset of flocculation in an ale-brewing strain, Saccharomyces cerevisiae NCYC 1195. METHODS AND RESULTS: Flocculation was evaluated using the method of Soares, E.V. and Vroman, A. [Journal of Applied Microbiology (2003) 95, 325]. For cells grown in chemically defined medium (yeast nitrogen base with glucose) or in rich medium (containing yeast extract, peptone and fermentable sugars: fructose or maltose), the onset of flocculation occurred after the end of exponential respiro-fermentative phase of growth being coincident with the attainment of the lower level of carbon source in the culture medium. Cells, in exponential respiro-fermentative phase of growth, transferred to a glucose-containing medium without nitrogen source, developed a flocculent phenotype, while these carbon source starved cells, in the presence of all other nutrients that support growth, did not flocculate. In addition, cells in exponential phase of growth, under catabolite repression, when transferred to a medium containing 0.2% (w/v) of fermentable sugar (fructose or maltose) or 2% (v/v) ethanol, showed a rapid triggering of flocculation, while when incubated in 2% (v/v) glycerol did not develop a flocculent phenotype. CONCLUSIONS: The onset of flocculation occurs when a low sugar and/or nitrogen concentration is reached in culture media. The triggering of flocculation is an energetic dependent process influenced by the carbon source metabolism. The presence of external nitrogen source is not necessary for developing a flocculent phenotype. SIGNIFICANCE AND IMPACT OF THE STUDY: This work contributes to the elucidation of the role of nutrients on the onset of flocculation in NewFlo phenotype yeast strains. This information might be useful to the brewing industry, in the control of yeast flocculation, as the time when the onset of flocculation occurs can determine the fermentation performance and the beer quality.     

Effect of rice--glycerol complex medium on the production of Lovastatin by Monascus ruber

Response surface methodology (RSM) was employed to study the effect of the composition of the rice-glycerol complex medium on the production of lovastatin (Lvs) by the ascomyceteMonascus ruber in mixed solid-liquid (or submerged) cultures at 25°C. Four components (rice powder, peptone, glycerol, glucose) were studied to evaluate, the approximate polynomial for all dependent variables, explaining their effects on the production of Lvs. The best composition derived from RSM regression was (in g/L) rice powder 34.4, peptone 10.8, , glucose 129, KNO₃ 8.0, MgSO₄·7H₂O 4.0 and glycerol 36.4 mL/L. With this composition, the Lvs production was 157 mg/L after 10 d of cultivation. In comparison with glycerol and glucose, the rice powder becomes a more suitable carbon source and represents a great potential for the production of Lvs.     

Carbon source dependent somatic embryogenesis and plant regeneration in cotton, Gossypium hirsutum L. cv. SVPR2 through suspension cultures

Highly reproducible and simple protocol for cotton somatic embryogenesis is described here by using different concentrations of maltose, glucose, sucrose and fructose. Maltose (30 g/l) is the best carbon source for embryogenic callus induction and glucose (30 g/l) was suitable for induction, maturation of embryoids and plant regeneration. Creamy white embryogenic calli of hypocotyl explants were formed on medium containing MS basal salts, myo-inositol (100 mg/l), thiamine HCI (0.3 mg/l), picloram (0.3 mg/l), Kin (0.1 mg/l) and maltose (30 g/l). During embryo induction and maturation, accelerated growth was observed in liquid medium containing NH3NO4 (1 g/l), picloram (2.0 mg/l), 2 ip (0.2 mg/l), Kin (0.1 mg/l) and glucose (30 g/l). Before embryoid induction, large clumps of embryogenic tissue were formed. These tissues only produced viable embryoids. Completely matured somatic embryos were germinated successfully on the medium fortified with MS salts, myo-inositol (50 mg/l), thiamine HCl (0.2 mg/l), GA3 (0.2 mg/l), BA (1.0 mg/l) and glucose (30 g/l). Compared with earlier reports, 65% of somatic embryo germination was observed. The abnormal embryo formation was highly reduced by using glucose (30 g/l) compared to other carbon sources. The regenerated plantlets were fertile but smaller in height than the seed derived control plants.