低温混菌发酵生产中链脂肪酸的机理研究及其过程强化

The psychrophilic mixed culture fermentation (PMCF) has a broad market in low temperature area, however, this technology is mainly limited by the low microbial activity and methane supersaturation. Medium chain fatty acids (MCFA, such as caproate and caprylate) are the promising biochemicals. Meanwhile, the tolerance of anaerobes to MCFA is also enhanced at low temperature. Therefore, the production of MCFA provides a new direction for PMCF. For the valorization of organic wastes under psychrophilic conditions, this project intends to produce MCFA in PMCF. Firstly, using ethanol and acetate as the substrates, the effects of temperature and substrates ratio on the MCFA production are demonstrated. The molecular biology technology and enzyme analysis are utilized to reveal the community dynamics and the mechanism of MCFA production under low temperature. Then, the biofilm reactor is constructed to strengthen the production and recovery of MCFA. Then glucose is used as substrate to enrich psychrophilic ethanol-type fermentation bacteria and to produce acetate and ethanol according to the mathematical model. In this two-phases mode, MCFA is produced indirectly from glucose. Finally, the organic wastes such as molasses wastewater are used as substrates. Therefore, the research outcomes will provide the new insight and technology to the removal of organic wastewater and utilization in PMCF.

低温发酵技术在气温较低地区具有广阔的市场，但是存在菌群活性低并且甲烷大量溶解等诸多问题。中链脂肪酸（如：己酸和辛酸）是高附加值的厌氧发酵产品，并且低温条件下厌氧菌对中链脂肪酸的耐受性也会增强。因此，中链脂肪酸的生产为低温发酵提供了新的方向。本项目拟以生产中链脂肪酸为目标，首先以乙醇和乙酸为底物，考察低温和底物比例对中链脂肪酸产率的影响，结合多种分子生物学技术阐明低温生产中链脂肪酸的机理；而后，添加载体构建生物膜反应器，强化生产并回收中链脂肪酸；富集低温乙醇型发酵菌群，结合数学模型调节代谢产物中乙醇和乙酸的含量，构建两段反应器运行模式实现间接生产中链脂肪酸；最后拓展至糖蜜废水等有机废弃物。因此，通过本项目的研究为有机废弃物的低温发酵资源化利用提供新的技术和理论依据。

低温发酵技术在气温较低地区具有广阔的市场，但菌群活性低并且甲烷大量溶解等资源化回收问题限制了它的广泛应用。中链脂肪酸（如：己酸和辛酸）是高附加值的厌氧发酵产品。因此，以低温厌氧发酵过程为研究对象，本项目实现了利用木糖、乙醇和合成气等多种底物生产己酸等中链脂肪酸，最高浓度达到19 g/L。高通量测序分析表明，主要菌群以Bifidobacterium (40-70%) 和 Clostridium_sensu_stricto_12 (14-33%) 为主。利用宏基因组学分析等分解析了不同条件下己酸生产的代谢途径，其中以木糖为底物时主要的代谢途径是FAB路径，而非通常认为的RBO途径。同时，我们构建了低温双室微生物燃料电池系统，并富集得到了高活性的低温（10℃和4℃）产电菌群。产电性能与中温已有报道相似，如最高输出电压均达到550 mV，库伦效率为82.4%，最大功率密度为582 mW/m2。产电菌群以Macellibacteroides fermentans （46.2%）和 Enterococcus lemanii（15.4%）为主。最后，胞外聚合物(EPS)的水解是限制污泥厌氧发酵的主要瓶颈。我们富集并得到了高活性的低温藻酸盐降解菌群，该菌群以拟杆菌属（genus Bacteroides）为核心菌群，能够分泌藻酸盐裂解酶（EC 4.2.2.3）。以剩余污泥为底物时，该菌群降解污泥的代谢产物以乙酸、丙酸和丁酸为主。接种比为5%时，污泥酸化效率提高了约30%。上述工作这拓展了对己酸生产的认识，并为低温有机废物的资源化提供了新的思路。