重组大肠杆菌全细胞合成D-苯基乳酸  

Biocatalytic production of D-phenyllactic acid by using whole cells of recombinant Escherichia coli

作　　者：鲍志伟 苏晓 杨柳婷 陈耀 罗希 付永前[1] 孙小龙[1] BAO Zhiwei;SU Xiao;YANG Liuting;CHEN Yao;LUO Xi;FU Yongqian;SUN Xiaolong(Institute of Biomass Resources,Taizhou University,Taizhou 318000,China)

机构地区：[1]台州学院生物质资源研究所,浙江台州318000

出　　处：《食品与发酵工业》2019年第1期49-53,共5页Food and Fermentation Industries

基　　金：国家自然科学基金(21007044;21106091);台州市课题(1701hb01);浙江省大学生科技创新计划暨新苗人才计划(2018R436026)

摘　　要：苯基乳酸(phenyllactic acid,PLA)是一种重要的有机酸,广泛应用于食品、医药、化妆品等领域。通过构建D-乳酸脱氢酶重组大肠杆菌BL21(DE3)/p ET28a-ldh D利用全细胞催化合成D-苯基乳酸。对重组大肠杆菌的诱导及转化条件进行了优化,最终确定了最佳的诱导条件:OD600=0. 8时,添加IPTG至终浓度为0. 2 mmol/L,在25℃下诱导4 h;最佳的转化条件:pH 7. 0磷酸缓冲液,13 g/L苯丙酮酸钠,5 g/L葡萄糖,30 g/L菌体(干重),37℃,200 r/min,转化2 h。在最优的条件下,苯基乳酸的产物浓度达到5. 2 g/L,产率为40%。该文还探索了苯丙酮酸钠与葡萄糖分批添加对苯基乳酸合成的影响,并初步得到了最佳工艺,经3 h转化,苯基乳酸产物质量浓度和产率分别提高到7. 38 g/L和52. 7%,显示了较好的应用前景,也为后续酶的改造奠定了基础。Phenyllactic acid is an important organic acid that has been widely used in foods,pharmaceuticals,cosmetics and other fields.D-phenyllactic acid was catalytically synthesized by whole cells through constructing recombinant Escherichia coli BL21 (DE3)/pET28a-ldh D.The induction and transformation conditions were optimized and determined as follows:IPTG was added to a final concentration of 0.2mmol/L when OD 600 reached 0.8 and cells were induced at 25℃ for 4h.The optimal transformation conditions were as follows:using pH=7.0 phosphate buffer,13g/L sodium phenylpyruvate,5g/L glucose,30g/L cells (dry weight),at 37℃ and 200r/min for 2h.Under the optimal conditions,phenyllactic acid accumulated up to 5.2g/L and the yield was 40%.Effects of fed-batch process using sodium pyruvate and glucoase on phenyllactic acid synthesis were explored,and the best processing scheme was obtained.After 3h of conversion,the concentration of D-phenyllactic and its yield increased to 7.38g/L and 52.7%,respectively.This study showed that the process of whole cell transformation for D-phenyllactic acid synthesis had good application prospects,which laid a foundation for subsequent enzymatic modification.

关 键 词：D-苯基乳酸 重组大肠杆菌 全细胞转化 D-乳酸脱氢酶 苯丙酮酸钠 

分 类 号：TQ921[轻工技术与工程—发酵工程]