Abstract:

High costs are the bottlenecks of traditional lactic acid fermentation process using glucose and starch as raw material. Production of lactic acid from raw glycerol, a byproduct of biodiesel, can significantly decrease the costs of fermentation. Therefore, there is a need to overcome the disadvantages of low cell growth rate, biomass of strains, as well as low productive rate and yield of lactic acid. In this work, metabolic evolution of Lactobacillus pentosus R3-8 screened by our group was carried out by adding high concentration raw glycerol and lactic acid. The 60th generation evolved strain using raw glycerol, tolerated 130 g·L^-1 raw glycerol, with 1.23 folds enhancement of biomass. The 50th generation evolved strain using lactic acid can tolerate 20 g·L^-1 lactic acid with 18% increase in biomass. The growth curves of 60th generation evolved strain using raw glycerol cultivated in 5 L bioreactor indicated that the lactic acid concentration, yield and productivity were 45.0 g·L^-1, 0.989 g·g^-1 and 0.47 g·L^-1·h^-1, respectively. Fed-batch cultivation of 60th generation evolved strain using raw glycerol achieved 83.8 g·L^-1 lactic acid, which was two folds of batch cultivation with the same strain.

Key words: metabolic evolution, lactic acid, glycerol, fermentation, metabolism, biochemical engineering

摘要：

以生物柴油产业的副产物粗甘油为底物,可降低乳酸发酵的生产成本。但是,粗甘油发酵生产乳酸存在菌体生长缓慢、菌浓较低、产酸速率和终产物浓度偏低等问题。以实验室筛选的一株戊糖乳杆菌 (Lactobacillus pentosus R3-8)为出发菌株进行代谢进化。通过在培养基中添加高浓度的粗甘油和乳酸,分别进行菌株耐底物和产物抑制的代谢进化。用粗甘油驯化的第60代菌株,可耐受130 g·L^-1的粗甘油,与出发菌株相比,生长速率提高, 且生物量是原始菌株的1.23倍。用乳酸驯化的第50代菌株可耐受20 g·L^-1的乳酸,生物量比初始菌株提升了18%。驯化菌株的5 L发酵罐分批发酵结果显示,以粗甘油驯化至 60 代的菌株的批次发酵水平相对较好,乳酸产量、甘油转化率以及生产强度分别为 45.0 g·L^-1、0.989 g·g^-1和 0.47 g·L^-1·h^-1。以粗甘油驯化至 60 代的菌株进行补料分批发酵,乳酸终浓度为83.8 g·L^-1,比分批发酵提高了近1倍。

关键词: 代谢进化, 乳酸, 甘油, 发酵, 代谢, 生化工程