Enhancement of pyruvate production by Torulopsis glabrata using a two-stage oxygen supply control strategy.

The effect of agitation speeds on the performance of producing pyruvate by a multi-vitamin auxotrophic yeast, Torulopsis glabrata, was investigated in batch fermentation. High pyruvate yield on glucose (0.797 g g(-1)) was achieved under high agitation speed (700 rpm), but the glucose consumption rate was rather low (1.14 g l(-1) h(-1)). Glucose consumption was enhanced under low agitation speed (500 rpm), but the pyruvate yield on glucose decreased to 0.483 g g(-1). Glycerol production was observed under low agitation speed and decreased with increasing agitation speed. Based on process analysis and carbon flux distribution calculation, a two-stage oxygen supply control strategy was proposed, in which the agitation speed was controlled at 700 rpm in the first 16 h and then switched to 500 rpm. This was experimentally proven to be successful. Relatively high concentration of pyruvate (69.4 g l(-1)), high pyruvate yield on glucose (0.636 g g(-1)), and high glucose consumption rate (1.95 g l(-1)h(-1)) were achieved by applying this strategy. The productivity (1.24 g l(-1) h(-1)) was improved by 36%, 23% and 31%, respectively, compared with fermentations in which agitation speeds were kept constant at 700 rpm, 600 rpm, and 500 rpm. Experimental results indicate that the difference between the performances for producing pyruvate under a favorable state of oxygen supply (dissolved oxygen concentration >50%) was caused by the different regeneration pathways of NADH generated from glycolysis.

Efficient pyruvate production by a multi-vitamin auxotroph of Torulopsis glabrata: key role and optimization of vitamin levels.

A multi-vitamin auxotroph, Torulopsis glabrata strain WSH-IP303, which can use ammonium chloride as a sole nitrogen source for pyruvate production, was selected. To optimize pyruvate yield and productivity, a simple but useful, orthogonal design method, was used to investigate the relationship between thiamine, nicotinic acid, pyridoxine, biotin, and riboflavin. Thiamine was confirmed to be the most important factor affecting pyruvate production. When the concentration of thiamine was 0.01 mg/l or 0.015 mg/l, glucose consumption was improved by increasing the nicotinic acid concentration. When the concentrations of nicotinic acid, thiamine, pyridoxine, biotin, and riboflavin were 8.0, 0.015, 0.4, 0.04, and 0.1 mg/l, respectively, pyruvate concentration and yield reached 52 g/l and 0.52 g/g, respectively, in a 48-h flask culture. By employing a combination of the optimum vitamin concentrations, a batch culture was conducted in a 2.5-l fermentor with an initial glucose concentration of 112 g/l; and the pyruvate concentration reached 69 g/l after 56 h (yielding 0.62 g/g).

[Effects of metal ions on gamma-poly (glutamic acid) synthesis by Bacillus licheniformis].

In the course of gamma-poly (glutamic acid) gamma-PGA fermentation, metal ions K+, Mg2+, Fe3+, Ca2+, Mo6+, Mn2+, Co2+ and Zn2+ in the medium have certain effects on the synthesis of gamma-poly(glutamic acid). Excess or lack of K+, Mg2+ and Fe3+ results in reduced yield of gamma-PGA. It was found that the gamma-PGA synthesis by Bacillus licheniformis was promoted obviously by Ca2+ and Mo6+. Synthesis and stereochemical composition of gamma-PGA was greatly regulated by Mn2+. gamma-PGA was not produced without Mn2+ in medium, and with the increase of Zn2+ concentration the yield of gamma-PGA and the proportion of D-glutamic acid in the peptide increase. Regulative effect of Co2+ and Zn2+ was almost the same as that of Mn2+, thus the combination of Mn2+, Co2+ cannot enhance gamma-PGA synthesis and affect stereochemical composition of gamma-PGA. Based on the experimental date, an appropriate formulation of metal ions in the medium for gamma-PGA production was obtained.

Biotechnological production of pyruvic acid.

Pyruvic acid is an important organic acid widely used in the chemical and drug, as well as agrochemical, industries. Compared with the chemical method, biotechnological production of pyruvic acid is an alternative approach because of the low cost. An overview of biotechnological production of pyruvate, including direct fermentative production employing eukaryotic and prokaryotic microorganisms, production by a resting cell method and an enzymatic method as well as the recovery of pyruvate, is discussed. A multi-vitamin auxotrophic yeast strain, Torulopsis glabrata. has been used in the commercial production of pyruvate; emphasis is therefore placed on the mechanism and characteristics of pyruvate production by this strain.

[Kinetics for terephthalic acid anaerobic degradation with easily biodegradable organic material co-existence].

The fact of preferential substrate utilization results in a sequence of substrate attack. As typically, the easily biodegradable substrates in TA-containing wastewater are degraded firstly through methane fermentation pathway, and just those intermediate metabolites have been proved to be inhibitors for TA biodegradation. Moreover, TA itself can inhibit the TA biodegradation, too. A kinetic model for the anaerobic digestion of wastewater containing both TA and easily biodegradable pollutants is constructed as q = qmax [formula: see text], The model parameters are estimated with non-linear regression method, the values are as follows: qmax = 1972.0 mgTA/gVSS.d; Ks = 20.2844 gTA/L; Ki,i = 2.041 gCOD/L; Ki,s = 0.0108 gTA/L. The experimental data verification for the model equation is satisfactory. According to the model analysis, a new strategy, a two-step anaerobic system, dealing with this kind of wastewater is suggested.

[Effect of nutritional conditions on the fermentative production of pyruvic acid by Torulopsis glabrata].

The effects of some nutritional conditions, such as peptone concentration, feeding glucose as well as oxygen supply manner and ratio of C/N in batch culture, on the fermentative production of pyruvic acid by Torulopsis glabrata WSH-IP12 were investigated. In shaking-flask culture: (1) peptone of more than 20 g/L inhibited the accumulation of pyruvic acid; (2) production of pyruvic acid was increased from 23.5 g/L to 30.2 g/L by simply feeding glucose. In 5 L jar-fermentor batch culture: (1) high level of dissolved oxygen and (2) increasing the concentration of glucose and peptone proportionally with constant C/N ratio(26:1) improved the production of pyruvic acid. It was also found that, glucose consumption and pyruvic acid production almost stopped under the condition of nitrogen difficiency while recovered by adding peptone and (NH4)2SO4. By using ammonia water instead of potassium hydroxide for the control of pH, the cells kept stronger ability for synthesizing pyruvic acid within the whole process, 57.3 g/L pyruvic acid with the yield of 0.498 g/g was achieved at 55 h of fermentation.

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.

[Studies on fermentation conditions for the accumulation of poly-beta-hydroxybutyrate in Alcaligenes eutrophus].

The results of the cultivation of Alcaligenes eutrophus showed that nitrogen limitation or exhaustion could stimulate the substantial accumulation of PHB. But the exhaustion of nitrogen source in PHB formation period would result in the rapid drop of PHB synthetic rate. Oxygen limitation could also stimulate the formation of PHB, but the content of PHB in the cell was much less than that in case of nitrogen controlled conditions. Obvious influences were observed on PHB fermentation when ammonia water feeding was stopped at different cell growth phases, and better results could be obtained when it was performed at 20 g/L to 30 g/L of residual biomass. Cell dry weight, PHB content and PHB concentration reached 61.9 g/L, 80.5% and 49.0 g/L, respectively under desired conditions.