以氯化胆碱、磷酸二氢钾、磷酸二氢钠为底物发酵法生产胞磷胆碱的过程中,后期高浓度盐离子导致菌体活力下降甚至衰亡,是限制胞磷胆碱产业化的主要问题。该研究为解决这一问题,首先确定了不同底物对胞磷胆碱发酵的影响。其次为了消除Cl-、K+、Na+对发酵的影响,分别使用电渗析法和离子交换树脂对底物进行预处理,探究二者的分离效率,结果得知离子交换树脂在效率以及成本上均优于电渗析法。因此为了实现胞磷胆碱大规模生产,在此基础上探究不同强度的离子交换树脂,实验确定了最适树脂型号、上样体积、上样浓度、洗脱液浓度、再生溶液类型以及树脂的稳定性,解决了后期高盐离子导致的高渗透压问题,使得以廉价原料高效生产胞磷胆碱成为可能。最后通过5 L发酵罐放大验证,以胞磷胆碱产量、生物量、糖酸转化率为指标,最终胞磷胆碱产量达到了32.1 g/L,糖酸转化率为25.4%,为目前国内发酵法生产胞磷胆碱的最高水平。与直接使用磷酸胆碱作为底物生产胞磷胆碱相比成本降低了85%,也证明了该次实验的可行性,对胞磷胆碱大规模工业化生产有一定的指导意义。
During the production of cytosolic choline by the fermentation method using choline chloride, potassium dihydrogen phosphate, and sodium dihydrogen phosphate as substrates, the high concentration of salt ions in the later stage leads to the decrease of bacterial viability or even the death of the bacteria, which is the main problem limiting the industrialization of cytosolic choline.To solve this problem, the effects of different substrates on cytosolic choline fermentation were first determined.Secondly, to eliminate the effects of Cl-, K+, and Na+ on fermentation, electrodialysis and ion-exchange resin were used to pre-treat the substrates and to investigate the separation efficiency.Results showed that ion-exchange resin was superior to electrodialysis in terms of efficiency and cost.Therefore, to realize the large-scale production of cytosolic choline, different strengths of ion exchange resins were investigated on this basis, and the optimum resin type, sample volume, sample concentration, eluent concentration, regeneration solution type, and stability of the resin were determined through experiments, which solved the high osmolality problem caused by high salt ions in the later stage, and made it possible to efficiently produce cytosolic choline with inexpensive raw materials.Finally, verified by a scale-up of a 5 L fermenter, the final cytosolic choline yield reached 32.1 g/L, and the sugar-acid conversion rate was 25.4%, which was the highest level of cytosolic choline production by fermentation method in China at present, taking the cytosolic choline yield, the biomass and the sugar-acid conversion rate as indicators.Compared with the direct use of choline phosphate as the substrate for the production of cytosine, the cost of cytosine was reduced by 85%, which also proved the feasibility of this experiment, providing some guiding significance for the large-scale industrialized production of cytosine.
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