硫酸锌(ZnSO4)对米根霉发酵产乳酸的影响

Effect of ZnSO4 on L-lactic acid production by Rhizopus oryzae

摘要:

摘要:【目的】为了了解无机盐与米根霉L-乳酸代谢之间的关系，提高米根霉菌株RLC41-6发酵产L-乳酸的产率与质量，研究了ZnSO4浓度与菌株乳酸代谢和细胞内乳酸脱氢酶活性的关系。【方法】在米根霉培养基中加入不同浓度ZnSO4，经过36℃培养36 h后，应用HPLC-反相色谱法测定产物中的L-乳酸含量，并利用活性PAGE分析法测定细胞内乳酸脱氢酶的活性和组成。【结果】实验结果显示，ZnSO4对除LDH1之外的其它几条同工酶都有促进作用，尤其对LDH4，LDH5作用明显，当ZnSO4浓度大于0.02%时，LDH4，LDH5达到最大水平，同时高浓度的锌离子在体外抑制了LDH的活性。当ZnSO4浓度为0.02%时LDH酶活达到最大200 U/mL，HPLC图谱表明，此时发酵产物的只有L-乳酸，且产量达到最大137g/L，乳酸转化率为91%。【结论】Zn+会影响米根霉的乳酸代谢过程，并导致发酵过程中产物类型的变化，合适浓度的ZnSO4在米根霉代谢产乳酸的过程中，提高了乳酸脱氢酶LDH的表达，抑制丙酮酸进入苹果酸和富马酸途径，从而有利于提高葡萄糖到乳酸的代谢。

关键词:关键词:L-乳酸，米根霉，HPLC，乳酸脱氢酶，ZnSO4

Abstract:

Abstract:［Objective］To improve the yield and quality of L-lactic acid by Rhizopus oryzae，we aim to understand the relationship between inorganic salts utilization and the L-lactic acid metabolism of the strain RLC41-6，through systematic analysis of the effects of zinc ion concentration on the production of L-lactic acid and the Lactic Dehydrogenase (LDH) activity．［Methods］ Rhizopus oryzae was cultured at 36℃ for 36h with different quantity of ZnSO4 in fermentation medium．The fermentation products were monitored by reversed-phase high performance liquid chromatography (RPHPLC)，LDH isoenzyme composition in the cell was analysed by non-denatured polyacrylamide gel electrophoresis (PAGE)．［Results］Our results showed that the concentration of ZnSO4 in medium could modulate the expression of LDH isoenzyme except LDH1，especially stimulated the expression of LDH4 and LDH5．When initial concentration of ZnSO4 is above 0.02%，the LDH4 and LDH5 reached the highest level．However，the activity of LDH was inhibited by higher concentration zinc ion in extracellular environment． When ZnSO4 concentration is 0.02%，LDH activity reaches its maximum 200 U/mL，the HPLC assay showed only L-lactic in the fermentation products (137 g/L)，while the conversion rate of glucose to lactic acid is 91%．［Conclusion］Zinc ion can regulate the metabolic processes of Rhizopus oryzae and modulate the types of the final fermentation products. An optimal concentration of ZnSO4 can not only facilitate the LDH expression but also prevent pyruvate from transformation into the malic acid and fumaric acid during the metabolism process，thereby enhance the metabolism of glucose to lactic acid of Rhizopus oryzae．

Key words:Keywords: L-lactic acid，Rhizopus oryzae，HPLC，Lactic dehydrogenase，ZnSO4