Inhibition of the anaerobic acetate degradation by formate

Summary Granular sludge from an UASB reactor fed with VFA showed a very low affinity for formate which provide little support to the theory of interspecies formate transfer. It is shown that formate can inhibit acetate degradation by anaerobic sludge.     

Toward homosuccinate fermentation: metabolic engineering of Corynebacterium glutamicum for anaerobic production of succinate from glucose and formate

Previous studies have demonstrated the capability of Corynebacterium glutamicum for anaerobic succinate production from glucose under nongrowing conditions. In this work, we have addressed two shortfalls of this process, the formation of significant amounts of by-products and the limitation of the yield by the redox balance. To eliminate acetate formation, a derivative of the type strain ATCC 13032 (strain BOL-1), which lacked all known pathways for acetate and lactate synthesis (Δcat Δpqo Δpta-ackA ΔldhA), was constructed. Chromosomal integration of the pyruvate carboxylase gene pyc(P458S) into BOL-1 resulted in strain BOL-2, which catalyzed fast succinate production from glucose with a yield of 1 mol/mol and showed only little acetate formation. In order to provide additional reducing equivalents derived from the cosubstrate formate, the fdh gene from Mycobacterium vaccae, coding for an NAD(+)-coupled formate dehydrogenase (FDH), was chromosomally integrated into BOL-2, leading to strain BOL-3. In an anaerobic batch process with strain BOL-3, a 20% higher succinate yield from glucose was obtained in the presence of formate. A temporary metabolic blockage of strain BOL-3 was prevented by plasmid-borne overexpression of the glyceraldehyde 3-phosphate dehydrogenase gene gapA. In an anaerobic fed-batch process with glucose and formate, strain BOL-3/pAN6-gap accumulated 1,134 mM succinate in 53 h with an average succinate production rate of 1.59 mmol per g cells (dry weight) (cdw) per h. The succinate yield of 1.67 mol/mol glucose is one of the highest currently described for anaerobic succinate producers and was accompanied by a very low level of by-products (0.10 mol/mol glucose).     

发酵产丁二酸过程中废弃细胞的循环利用

对厌氧发酵产丁二酸后的废弃细胞进行破壁处理,考察了以细胞水解液作为有机氮源重新用于丁二酸发酵的可行性。比较了超声破碎、盐溶、酶解3种方法破碎细胞获得的水解液作为氮源发酵产丁二酸的效果,结果表明酶解制得的细胞水解液效果最佳。以总氮含量为1.11g/L的酶解液(相当于10g/L酵母膏)作为氮源发酵,丁二酸产量可达42.0g/L,继续增大酶解液用量对耗糖、产酸能力没有显著提高。将细胞酶解液与5g/L酵母膏联用发酵36h后,丁二酸产量达75.5g/L,且丁二酸生产强度为2.10g/(L·h),比使用10g/L酵母膏时提高了66.7%。因此,厌氧发酵产丁二酸结束后的废弃细胞酶解液可以替代原培养基中50%的酵母膏用于发酵。     

Citrate catabolism and production of acetate and succinate by Lactobacillus helveticus ATCC 15807

The citrate metabolism of Lactobacillus helveticus ATCC 15807 was studied under controlled-pH fermentations at pH 4.5 and pH 6.2. The micro-organism was able to co-metabolize citrate and lactose at both pH from the beginning of growth, which enhanced the rate of lactose consumption and lactic acid production, compared with cultures without citrate. The effect of citrate on cell growth was dependent on the balance between the ratio of dissociated to non-dissociated forms of the acetic acid produced and the extra ATP gained by the cells, both facts related to the citrate metabolism. The citrate catabolism determined a change in the fermentation pattern of L. helveticus ATCC 15807 from homolactic to a mixed-acid profile, regardless of the external pH. Within this new fermentation pattern, acetate was the major product formed (13–20 mM), followed by succinate (2.4–3.7 mM), while acetoine, dyacetile or butanediol were not detected. The mixed-acid profile displayed by L. helveticus ATCC 15807 was linked to NADH₂ oxidase activity rather than the acetate kinase enzyme.     

Lactic acid production from sugarcane bagasse hydrolysates by Lactobacillus pentosus: Integrating xylose and glucose fermentation

Lactic acid, traditionally obtained through fermentation process, presents numerous applications in different industrial segments, including production of biodegradable polylactic acid (PLA). Development of low cost substrate fermentations could improve economic viability of lactic acid production, through the use of agricultural residues as lignocellulosic biomass. Studies regarding the use of sugarcane bagasse hydrolysates for lactic acid production by Lactobacillus spp. are reported. First, five strains of Lactobacillus spp. were investigated for one that had the ability to consume xylose efficiently. Subsequently, biomass fractionation was performed by dilute acid and alkaline pretreatments, and the hemicellulose hydrolysate (HH) fermentability by the selected strain was carried out in bioreactor. Maximum lactic acid concentration and productivity achieved in HH batch were 42.5 g/L and 1.02 g/L h, respectively. Hydrolyses of partially delignified cellulignin (PDCL) by two different enzymatic cocktails were compared. Finally, fermentation of HH and PDCL hydrolysate together was carried out in bioreactor in a hybrid process: saccharification and co-fermentation with an initial enzymatic hydrolysis. The high fermentability of these process herein developed was demonstrated by the total consumption of xylose and glucose by Lactobacillus pentosus, reaching at 65.0 g/L of lactic acid, 0.93 g/g of yield, and 1.01 g/L h of productivity. © 2018 American Institute of Chemical Engineers Biotechnol. Prog., 35: e2718, 2019     

戊糖乳杆菌发酵产乳酸及高产菌株诱变选育

戊糖乳杆菌(Lactobacillus pentosus)是能利用木质纤维素水解液发酵产乳酸的潜力菌株,发酵条件优化与高产菌株的选育是提高乳酸产量的重要手段。通过单因素试验、Plackett-Burman设计与响应面试验,对戊糖乳杆菌ATCC 8041产乳酸的发酵培养基及发酵条件进行了优化。结果表明,该菌株发酵培养基的最佳组合为葡萄糖93.11 g/L、酵母浸粉5.19 g/L、碳酸钙29.43 g/L、蛋白胨10.00 g/L、Na2HPO4·12H2O 5.00 g/L、Mg SO4 0.20 g/L、Mn SO4 50 mg/L;最佳发酵条件为37℃、p H6.5、接种量6%、装液量80%。在此优化条件下,该菌株发酵产乳酸为54.12 g/L。进一步以戊糖乳杆菌ATCC 8041为出发菌株,通过原生质体进行紫外诱变,经多重筛选,最终获得一株遗传稳定性好的高产乳酸突变株,命名为戊糖乳杆菌Lactic UVC-02,由中国典型培养物保藏中心保存,注册号为CCTCC M 2013209。该突变株Lactic UVC-02经葡萄糖发酵,乳酸产量达64.17 g/L,比出发菌株ATCC 8041(54.12g/L)提高18.6%。     

Fermentation of citrate by Lactobacillus plantarum in the presence of a yeast under acid conditions

Summary At pH 3.6, Lactobacillus plantarum is unable to grow on citrate or to ferment it in the absence of another carbon source such as glucose. In a defined medium containing glucose and citrate, with a higher concentration of the former than the latter, as in many fermented alcoholic beverages, L. plantarum will first ferment the sugar. The production of lactate from glucose degradation increases the acidity of the medium and inhibits the fermentation of citrate. In co-culture with Saccharomyces cerevisiae, part of the glucose is fermented by the yeast, partly avoiding the pH drop and the inhibition of citrate fermentation by L. plantarum. Fermentation was still possible at pH values around 3.0. Offprint requests to: C. Kennes     

Metabolic activities of Listeria monocytogenes in the presence of sodium propionate, acetate, lactate and citrate

The effects of sodium propionate, acetate, lactate and citrate on cell proliferation, glucose and oxygen consumption, and ATP production in Listeria monocytogenes were investigated in growing and resting cells. Media pH was 6.7-6.8. Growth inhibition increased while glucose consumption continued in the presence of ≥ 1% propionate, ≥ 3% acetate and ≥ 5% lactate in broth during incubation at 35°C, indicating that glucose consumption was uncoupled from cell proliferation. Acetate and propionate were the most effective antilisterials, whereas citrate (5%) was only slightly inhibitory. Of the four salts, only lactate supported growth, oxygen consumption and ATP production. While concentrations of 1 and 5% propionate, acetate and citrate did not have an effect on oxygen consumption, they inhibited ATP production. ATP production in the presence of the four salts was consistently lower at pH 6.0 than at neutral pH. Lactate served as an alternative energy source for L. monocytogenes in the absence of glucose but became toxic to the organism in the presence of the carbohydrate.     

Characterisation of autolytic enzymes in Lactobacillus pentosus

AIMS: To characterize autolysis and autolytic system of the lactic acid bacterium Lactobacillus pentosus. METHODS AND RESULTS: Autolysis of nine Lact. pentosus strains was evaluated in buffer solution. Their peptidoglycan hydrolase profiles were examined by renaturing SDS-PAGE and revealed two major activity bands at 58 and 112 kDa. Specificity analysis indicated the presence of at least two different types of peptidoglycan hydrolase activities in Lact. pentosus 1091. CONCLUSIONS: Autolysis of Lact. pentosus was shown to be strain dependent and involvement of at least two different autolysins was evidenced. SIGNIFICANCE AND IMPACT OF THE STUDY: The autolytic system of Lact. pentosus was characterized for the first time and the data obtained could be used in the selection of strains of technological interest.     

Production of hydrogen and methane from potatoes by two-phase anaerobic fermentation

A two-phase anaerobic process to produce hydrogen and methane from potatoes was investigated. In the first phase, hydrogen was produced using heat-shocked sludge. About 12h lag-phase vanished, hydrogen yield increased from 200.4 ml/g-TVS to 217.5 ml/g-TVS and the maximum specific hydrogen production rate also increased from 703.4 ml/g-VSS d to 800.5 ml/g-VSS d when improved substrate was used, in which Cl(-) was substituted for SO(4)(2-). Better performances of 271.2 ml-H(2)/g-TVS and 944.7 ml-H(2)/g-VSS d were achieved when potatoes were pretreated by alpha amylase and glucoamylase. In the second phase, methane was produced from the residual of the first phase using methanogens. The maximum additional methane yield was 157.9 ml/g-TVS and the maximum specific methane production rate was 102.7 ml/g-VSS d. The results showed that the energy efficiency increased from about 20% (hydrogen production process) to about 60%, which indicated the energy efficiency can be improved by combined hydrogen and methane production process.     

CO2-dependent fermentation of phenol to acetate, butyrate and benzoate by an anaerobic, pasteurised culture

Fermentative degradation of phenol was studied using a non-methanogenic, pasteurised enrichment culture containing two morphologically different bacteria. Phenol was fermented to benzoate, acetate and butyrate and their relative occurrence depended on the concentration of hydrogen. Proportionately more benzoate was formed with high initial levels of H2. The influence of PH2 on the fermentation pattern was studied both in dense cell suspensions and in growing cultures by addition of hydrogen. An increase in growth yield (OD578) was observed, compared to controls, as a consequence of phenol degradation; however, the increase was less in H2-amended treatments, in which most of the phenol ended up as benzoate. The degradation of phenol in the dense cell suspension experiments was dependent on CO2. Benzoate was not degraded when added as a substrate to the growing culture. This is, to our knowledge, the first report concerning the fermentative degradation of phenol to nonaromatic products.     

戊糖乳杆菌31-1菌株产细菌素发酵条件优化

对戊糖乳杆菌31-1产细菌素的条件进行了优化,分别研究了培养温度,培养基起始pH值,培养基碳源、氮源,刺激因子等因素对细菌素产量的影响。组合因素优化结果得到最佳培养基与培养条件为：乳糖30g、胰胨15g、豆胨20g、牛肉膏30g、蛋白胨20g、吐温801mL、磷酸氢二钾2g、乙酸钠5g、柠檬酸铵2g、硫酸镁0．58g、硫酸锰0．25g,蒸馏水定容至1000mL,30℃培养24h,培养起始pH为6．5。在此条件下培养细菌素效价可达到640AU／mL,与起始培养基相比细菌素产量提高了8倍。