Acetate production by a coupled syntrophic acetogenesis with homoacetogenesis process: effect of sludge inoculum concentration.

The effect of sludge inoculum concentration on acetate (Ac) production by a novel coupled system was investigated. The coupled system consisted ofa syntrophic acetogenesis phase and a homoacetogenesis phase. The results showed that the inoculum concentration not only affected the acetate production at the syntrophic acetogenesis phase but also that produced at the homoacetogenesis phase. When the inoculum concentration in the syntrophic acetogenesis phase was 4 g l(-1), the acetate yield of the coupled system was found to be about 0.36 g Ac g(-1) glucose. In addition, when the inoculum concentration in the homoacetogenesis phase was 4 g l(-1), a maximum acetate production of 11.2 g l(-1) was obtained in the syntrophic acetogenesis phase and the yield of acetate was 0.39 g Ac g(-1) glucose. Analysis shows that the variation in acetate yield might be attributed to different metabolic pathways and the imbalance between hydrogen production and hydrogen consumption in the coupled system with different inoculum concentrations.

Improved bioconversion of volatile fatty acids from waste activated sludge by pretreatment.

Batch tests were conducted to analyze the influence of various pretreatment methods including thermo-alkaline, thermo-acid, ultrasonic-alkaline, and ultrasonic-acid on the bioconversion of volatile fatty acids (VFAs) from waste activated sludge. Experimental results showed that total VFAs (TVFAs) increased by 68.2% for ultrasonic-alkaline and 59.1% for thermo-alkaline. The TVFAs bioconversion in the case of the thermo-acid or ultrasonic-acid pretreatment, however, was lower than that without pretreatment. The results of VFA distribution showed that acetic acid was the most prevalent product, with a fraction of 35.5 to 57.2% for all cases. The mechanism of VFA production was then investigated. Results revealed that, in the two alkaline-pretreated slurries, soluble COD and soluble BOD/soluble COD significantly increased to yield more soluble substrate for the subsequent fermentation. Soluble proteins comprised the main substance provided that was converted to VFAs. A further investigation indicated that volatile (organic) solids, including insoluble proteins, in the solid phase of the samples pretreated with these two methods were hardly consumed during the fermentation.

Improved glutathione production by gene expression in Escherichia coli.

AIMS: To improve glutathione (GSH) production in Escherichia coli by different genetic constructions containing GSH genes. METHODS AND RESULTS: GSH production was very low in E. coli by the expression of gshI gene. An increase of GSH production was achieved by the expression of both gshI and gshII genes in E. coli. A higher GSH production, namely 34.8 mg g(-1) wet cell weight, was obtained by simultaneous expression of two copies of gshI gene and one copy of gshII gene. CONCLUSIONS: The simultaneous expression of two copies of gshI gene and one copy of gshII gene resulted in a significant increase in GSH production. SIGNIFICANCE AND IMPACT OF THE STUDY: The expression strategy for GSH production described here can be used to increase gene expression and obtain high production rates in other multienzyme reaction systems.

Increasing glycolytic flux in Torulopsis glabrata by redirecting ATP production from oxidative phosphorylation to substrate-level phosphorylation.

AIMS: This study aimed at further increasing the pyruvate productivity of a multi-vitamin auxotrophic yeast Torulopsis glabrata by redirecting ATP production from oxidative phosphorylation to substrate-level phosphorylation. METHODS AND RESULTS: We examined two strategies to decrease the activity of F0F1-ATPase. The strategies were to inhibit F0F1-ATPase activity by addition of oligomycin, or to disrupt F0F1-ATPase by screening neomycin-resistant mutant. The addition of 0.05 mmol l(-1) oligomycin to the culture broth of T. glabrata CCTCC M202019 resulted in a significantly decreased intracellular ATP level (35.7%) and a significantly increased glucose consumption rate (49.7%). A neomycin-resistant mutant N07 was screened and selected after nitrosoguanidine mutagenesis of the parent strain T. glabrata CCTCC M202019. Compared with the parent strain, the F0F1-ATPase activity of the mutant N07 decreased about 65%. As a consequence, intracellular ATP level of the mutant N07 decreased by 24%, which resulted in a decreased growth rate and growth yield. As expected, glucose consumption rate and pyruvate productivity of the mutant N07 increased by 34% and 42.9%, respectively. Consistently, the activities of key glycolytic enzymes of the mutant N07, including phosphofructokinase, pyruvate kinase and glyceraldehyde-3-phosphate dehydrogenase, increased by 63.7%, 28.8% and 14.4%, respectively. In addition, activities of the key enzymes involved in electron transfer chain of the mutant N07 also increased. CONCLUSIONS: Impaired oxidative phosphorylation in T. glabrata leads to a decreased intracellular ATP production, thereby increasing the glycolytic flux. SIGNIFICANCE AND IMPACT OF THE STUDY: The strategy of redirecting ATP production from oxidative phosphorylation to substrate-level phosphorylation provides an alternative approach to enhance the glycolytic flux in eukaryotic micro-organisms.

Redirecting carbon flux in Torulopsis glabrata from pyruvate to alpha-ketoglutaric acid by changing metabolic co-factors.

The nutrition conditions needed to redirect the carbon flux in Torulopsis glabrata, a pyruvate hyper-production yeast, from pyruvate to alpha-ketoglutaric acid (KG) were investigated in a stirred fermentor. A minor amount of KG (1.3 gl(-1)) was produced when NaOH was used to control the pH, while 12 g KG l(-1) was produced when CaCO(3) was used instead. When thiamine and biotin were included in the medium, 13 g KG l(-1) and 68 g pyruvate l(-1) were produced after 48 h when glucose was nearly consumed (approximately 5 gl(-1)). With fermentation continuing for a further 16 h, the concentration of pyruvate decreased to 31 gl(-1), and KG increased to 30 gl(-1). KG thus accumulated at the expense of pyruvate consumption.

[Progress on the biosynthesis of medium-chain-length polyhydroxyalkanoates by microorganisms].

Polyhydroxylkanoates(PHAs) are a class of polyesters produced as reserve materials by a large number of microorganisms under metabolic stress. The most fascinating feature of PHAs is its degradability, and which is supposed to take place of the traditional plastics made from petroleum in the future. PHAs are divided into two classes: short-chain-length PHAs(scl-PHAs) and medium-chain-length PHAs. mcl-PHAs is more welcome owing to its more wide crystallinity and higher extension to break than scl-PHAs, especially when some kind of new functional groups were incorporated into the side chain of the polyester. Since Psedumonas oleovorans is the most typical microorganism to produce mcl-PHAs, here the author summarized how P. oleovorans synthesize the mcl-PHAs and the production of mcl-PHAs by fermentation and give some of the idea about the future research of this field.

[Nutrition condition of hyaluronic acid fermentation with Streptococcus zooepidemicus].

Based on the analysis of metabolic pathway Streptococcous zooepidemicus for hyaluronic acid (HA) synthesis, nucleotide, especially uracil, was considered to be important to cell growth and metabolism. When 0.005 g.L-1 uracil added in the media in which yeast extract as complex nitrogen source, cell growth and HA production were increased by 32% and 34% respectively. From analysis of amino acid in fermentation process, it was show that arginine(Arg) was needed for cell metabolism, and concentration of free Arg maintained at 0 g.L-1 in fermentation process, which was proposed to limit cell growth and HA production. By shake-flask experiment HA concentration reached 0.510 g.L-1 when 0.06 g.L-1 Arg added, in the fermentation with 2.5 L fermentor, when uracil 0.005 g.L-1 and Arg 0.06 g.L-1 were added, the rate of cell growth increased, maximum of specific growth rate, concentration of HA and HA molecular weight reached 0.67 h-1, 5.2 g.L-1 and 2.15 x 10(6) Da from 0.54 h-1, 4.2 g.L-1, 2.0 x 10(6) Da, respectively.

[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.