玉米芯半纤维素水解液中抑制物对丁醇发酵的影响

Effects of Inhibitors in Corncob Hemicellulose Hydrolysate on Butanol Fermentation

半纤维素水解液抑制物对微生物细胞的毒性限制了其在丁醇发酵中的应用,旨在探讨其对产丁醇共生体系TSH06的抑制作用并为其应用于丁醇发酵奠定基础。通过稀酸水解半纤维素制得水解液,采用P2培养基稀释的半纤维素水解液为底物,分别利用NaOH和氨水调节培养基pH值,结合实时荧光定量PCR方法来研究水解液对产丁醇共生体系TSH06的抑制作用。以NaOH调节pH抑制菌体的生长,氨水调节pH菌体可生长发酵。产丁醇菌株TSH06可以在50%稀释度以下的水解液中发酵生长,并且能够耐受并降解抑制物糠醛与5-羟甲基糠醛,最终丁醇产量达到4.16-5.16 g/L,低于P2培养基中的丁醇产量(8.83 g/L)。稀释水解液中,48 h之后乙酸浓度在3.18-4.16 g/L,远大于P2培养基中的乙酸浓度(低于2 g/L)。相对于P2培养基,在50%水解液中培养的TSH06有机酸生成途径关键基因的基因转录水平明显提高,而有机酸返耗途径以及丁醇生成途径的关键基因的基因转录水平则明显下降。水解液中过多的乙酸抑制了产酸期到产溶剂期的转化,而酸的累积使得菌体在底物被完全消耗之前就趋于衰退死亡,从而造成丁醇产量的降低与底物的不完全利用。

Due to the toxicity of inhibitors in hemicellulose hydrolysate to microorganism,hemicellulose hydrolysate cannot be directlyused as substrate for butanol fermentation.The effects of inhibitors to non-anaerobic butanol-producing symbiotic system TSH06 was studied,aiming at laying the foundation for the butanol fermentation using hydrolysate without detoxification.Hemicellulose hydrolysate of corncob dilutedwith P2 medium was used as substrate in butanol fermentation.The pH of culture media was adjusted to be neutral with Na OH and ammoniarespectively.The inhibition effect of the hydrolysate on TSH06 was studied by q PCR with gene primes of key enzymes.Results showed that theexistence of Na＋ inhibited the growth of TSH06.Furthermore,TSH06 grew and fermented in 50% or lower degrees of diluted hemicellulosehydrolysate,tolerated and degraded the furfural and 5-HMF,and the ultimate butanol concentration reached 4.16-5.16 g/L,lower than thatin P2 medium（butanol concentration of 8.83 g/L）.In diluted hemicellulose hydrolysate,acetic acid concentration remained above 3.18 to4.16 g/L after 48 h,much higher than that in P2 medium（2 g/L）.Compared with P2 medium,the transcription ratio of key genes in theorganic acid production pathway rose significantly while TSH06 in the 50% hemicellulose hydrolysates and declined significantly in the organicacid reflux pathway and butanol production pathway.In conclusion,excessive acetic acid in the hemicellulose hydrolysate might damagecells and block the transition from acidogenesis phase to solventogenesis phase.As the accumulation of acid,bacterial cells tend to declineand death before the substrate is completely consumed,leading to the reduction of butanol production and the substrate is not fully utilized.