氮源及其添加模式对钝齿棒杆菌JDN28-75合成L-精氨酸的影响

氮源是微生物过量合成L-精氨酸的重要营养因子之一,不同氮源对钝齿棒杆菌JDN28-75合成L-精氨酸的影响研究结果表明,硫酸铵为合适的氮源.不同初始硫酸铵浓度对JDN28-75产L-精氨酸的影响研究结果表明,氮源浓度过高或不足,都会使最终L-精氨酸产量有所降低.低浓度的硫酸铵虽然有利于菌体生长,但对L-精氨酸的合成明显不利,同时糖酸转化率也较低;而高浓度的硫酸铵尽管不利于细胞的生长且造成发酵结束时残糖含量过高,却有利于细胞合成L-精氨酸且实际耗糖的糖酸转化率维持在一个较高的水平.初始硫酸铵浓度为60 g/L时,对JDN28-75菌体的生长有明显的抑制作用,最终发酵液中剩余的硫酸铵也较多(大于30 g/L),但高浓度的硫酸铵是L-精氨酸合成所必需的.在上述研究结果的基础上,确定了初始硫酸铵浓度为20 g/L条件下的补氮策略,比较了4种不同的硫酸铵补加模式对产L-精氨酸的影响,结果表明,在总的硫酸铵浓度相同的情况下,采取分批、低浓度添加氮源的方式既可以有效解除发酵前期高浓度硫酸铵对菌体生长的抑制作用,又可以有效维持发酵中后期体系中菌体合成L-精氨酸所需的较高比例的氮源.最后,在5 L全自动发酵罐中采用20 g/L的初始硫酸铵浓度,连续流加25%的氨水来控制发酵体系pH及补加氮源,L-精氨酸的产量可以达到31.7 g/L,较对照组的产酸量(26.0 g/L)提高了21.9%.图4表2参11

Effects of Nitrogen Source and Its Supply Manner on Production of L-arginine by Corynebacterium crenatum JDN28-75

Nitrogen source and its supply strategy plays an important role in microbial over-production of L-arginine.The effect of various nitrogen sources on L-arginine production by Corynebacterium crenatum JDN28-75 was investigated.Ammonium sulfate(AS) was demonstrated to be the optimum nitrogen source.The effect of the initial concentration of AS on L-arginine production was further studied in shaking-flask experiments.In batch fermentation,either deficient or excessive supply of AS would decrease the final titer of L-arginine.Low concentration of AS facilitated cell growth but was not sufficient to achieve a high-level production of L-arginine,while high concentration of AS was beneficial for producing L-arginine but inhibited cell growth and glucose consumption.This observation suggested that an AS feeding strategy might be desirable to further improve the production of L-arginine.Four AS feeding strategies(with the initial AS concentration set at 20 g/L) were proposed and experimentally examined.Batch fermentation with AS fed twice(initial AS concentration 20 g/L,20 g/L AS was fed at 15 h,another 20 g/L AS was fed at 30 h) showed 28% improved L-arginine production as compared to the control,where the initial AS concentration was set at 60 g/L.The improvement was due to that the low initial AS concentration reduced growth inhibition in one aspect,and the continuous supply of low concentration of AS in the later stage of fermentation increased the production of L-arginine in another aspect.The proposed AS feeding strategy was further validated in fermentations performed in a 5-litre jar fermentor with the initial AS concentration set at 20 g/L.Ammonia water(25%) was used to control the pH and serve as a nitrogen source.The titer of L-arginine reached 31.7 g/L,which was 21.9% higher than the control(26.0 g/L).The nitrogen feeding strategy proposed in this study provided an efficient way to improve the microbial production of L-arginine and might be easily scaled-up to larger scale fermentations.Fig 4,Tab 2,Ref 11