In order to improve the fermentation intensity of α-ketoglutaric acid (α-KG) with Yarrowia lipolytica,a high α-KG production strain 1-C6 screened previously was treated by iterative mutagenesis,and the fermentation conditions for α-KG production with the final obtained mutant were systematically optimized.Firstly,based on the previous established high-throughput screening method,a high-yield mutant 4-E2 was obtained by iterative mutagenesis with ethyl methane sulfonate,nitrosoguanidine and ultraviolet light.Compared with the starting strain 1-C6,the production of α-KG was enhanced by 56.7%.Then,effects of agitation speed,ventilation ratio,addition of sodium acetate and CaCO3 on the accumulation of α-KG with mutant 4-E2 were investigated in 3 L fermenter.It was found that a two-stage ventilation ratio combined with relative higher rotating speed was more suitable for α-KG production,and the optimum addition of sodium acetate and CaCO3 were 6 g/L and 10 g/L,respectively.On this basis,a strategy of constant feeding strategy was further determined after exploring different strategies of fed-batch fermentation.Under the optimal conditions,the production of α-KG reached 75.86 g/L,which was 44.4% higher than that of the batch fermentation.The results could provide references for the industrialization of α-KG fermentation.
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