水-固相磷脂酶D催化反应体系合成磷脂酰甘油
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摘要
构建了高效环保的水-固相磷脂酶D(PLD)催化反应体系用于磷脂酰基转移合成磷脂酰甘油(PG)。通过共价结合曲拉通的纳米二氧化硅载体在纯水相中实现磷脂(PC)吸附,载体上的PC在PLD催化下进行水-固界面磷脂酰基转移合成PG。系统研究了载体表面曲拉通质量摩尔浓度对PC的吸附,当载体表面曲拉通质量摩尔浓度为8.5×10~(-8) mol/g时,PC在载体上的吸附率达98%,PG转化率达90.5%。反应动力学符合PC限制的Michaelis-Menten方程。相比于传统双液相反应体系,水-固反应体系避免了有毒易挥发溶剂的使用;转酯反应温度及pH值操作范围扩大;可实现游离酶与载体的回收再利用,游离酶重复利用7次后PG相对产率仍高达80%,载体使用16次后PC吸附量和PG产率无明显下降。
A highly efficient and environmentally friendly aqueous-solid system for phospholipase D-catalyzed transphosphatidylation was established to produce phosphatidylglycerol(PG). Triton X-100 was covalently bound to the surface of nanosilica, and Triton X-100-modified nanosilica obtained was used for the adsorption of phosphatidylcholine(PC) in the purely aqueous solution. PG was successfully synthesized using the carrier-adsorbed PC under PLD-catalyzed transphosphatidylation. The effect of the content of Triton X-100 on the surface of nanosilica on transphosphatidylation was investigated systematically. The optimal value of Triton X-100 content on the surface of nanosilica was 8.5×10~(-8) mol/g. Results indicated that the highest PC loading and yield of PG reached 98% and 90.5%, respectively. The reaction kinetics conforms to the PC-limited Michaelis-Menten equation. Compared with the traditional liquid-liquid system, the aqueous-solid reaction system avoids the use of toxic and volatile solvents, the tolerance of temperature and pH in our aqueous-solid reaction system were also improved significantly. The recyclability and stability of free PLD solution were studied, the relative yield of PG was still up to 80% in the seventh batch. The carrier was recycled for 16 times, the adsorption capacity of PC and the yield of PG did not obviously decrease.
A highly efficient and environmentally friendly aqueous-solid system for phospholipase D-catalyzed transphosphatidylation was established to produce phosphatidylglycerol(PG). Triton X-100 was covalently bound to the surface of nanosilica, and Triton X-100-modified nanosilica obtained was used for the adsorption of phosphatidylcholine(PC) in the purely aqueous solution. PG was successfully synthesized using the carrier-adsorbed PC under PLD-catalyzed transphosphatidylation. The effect of the content of Triton X-100 on the surface of nanosilica on transphosphatidylation was investigated systematically. The optimal value of Triton X-100 content on the surface of nanosilica was 8.5×10~(-8) mol/g. Results indicated that the highest PC loading and yield of PG reached 98% and 90.5%, respectively. The reaction kinetics conforms to the PC-limited Michaelis-Menten equation. Compared with the traditional liquid-liquid system, the aqueous-solid reaction system avoids the use of toxic and volatile solvents, the tolerance of temperature and pH in our aqueous-solid reaction system were also improved significantly. The recyclability and stability of free PLD solution were studied, the relative yield of PG was still up to 80% in the seventh batch. The carrier was recycled for 16 times, the adsorption capacity of PC and the yield of PG did not obviously decrease.
引文
[1]刘元发,王兴国.大豆磷脂组成与特性[J].粮食与油脂,2000(3):11-15.
[2]李彤,佘绍逸,佘璇瑜,等.肺表面活性物质的临床运用与新生儿肺透明膜病的转归[J].实用医学杂志,2005,21(12):1342-1344.
[3]黄曙光,吴桂芳,程宁.磷脂酰甘油检测对肺透明膜病的诊断意义[J].蚌埠医学院学报,1996(6):379-380.
[4]胡飞,段章群,王淮,等.生物酶法制备磷脂酰丝氨酸的研究进展[J].中国油脂,2012,37(6):54-58.
[5]ZHANG Xiaoli,LI Binglin,WANG Jiao,et al.High-yield and sustainable production of phosphatidylserine in purely aqueous solutions via adsorption of phosphatidylcholine on Triton X-100-modified silica[J].Journal of Agricultural&Food Chemistry,2017,65(49):10767-10774.
[6]LI Binglin,WANG Jiao,ZHANG Xiaoli,et al.An aqueous-solid system for the highly efficient and environmentally friendly transphosphatidylation catalyzed by phospholipase D to produce phosphatidylserine[J].J Agric Food Chem,2016,64(40):7555-7560.
[7]PRIETO M A,VAZQUEZ J A,MURADO M A.A new and general model to describe,characterize,quantify and classify the interactive effects of temperature and p H on the activity of enzymes.[J].Analyst,2015,140(10):3587-602.
[8]RODRIGUES R C,ORTIZ C,BERENGUER Murcia,et al.Modifying enzyme activity and selectivity by immobilization.[J].Chemical Society Reviews,2013,42(15):6290-307.
[9]JUNEJA L R,TANIGUCHI E,SHIMIZUTSUNEOYA-MANE S.Increasing productivity by removing choline in conversion of phosphatidylcholine to phosphatidylserine by phospholipase D[J].Journal of Fermentation&Bioengineering,1992,73(5):357-361.
[2]李彤,佘绍逸,佘璇瑜,等.肺表面活性物质的临床运用与新生儿肺透明膜病的转归[J].实用医学杂志,2005,21(12):1342-1344.
[3]黄曙光,吴桂芳,程宁.磷脂酰甘油检测对肺透明膜病的诊断意义[J].蚌埠医学院学报,1996(6):379-380.
[4]胡飞,段章群,王淮,等.生物酶法制备磷脂酰丝氨酸的研究进展[J].中国油脂,2012,37(6):54-58.
[5]ZHANG Xiaoli,LI Binglin,WANG Jiao,et al.High-yield and sustainable production of phosphatidylserine in purely aqueous solutions via adsorption of phosphatidylcholine on Triton X-100-modified silica[J].Journal of Agricultural&Food Chemistry,2017,65(49):10767-10774.
[6]LI Binglin,WANG Jiao,ZHANG Xiaoli,et al.An aqueous-solid system for the highly efficient and environmentally friendly transphosphatidylation catalyzed by phospholipase D to produce phosphatidylserine[J].J Agric Food Chem,2016,64(40):7555-7560.
[7]PRIETO M A,VAZQUEZ J A,MURADO M A.A new and general model to describe,characterize,quantify and classify the interactive effects of temperature and p H on the activity of enzymes.[J].Analyst,2015,140(10):3587-602.
[8]RODRIGUES R C,ORTIZ C,BERENGUER Murcia,et al.Modifying enzyme activity and selectivity by immobilization.[J].Chemical Society Reviews,2013,42(15):6290-307.
[9]JUNEJA L R,TANIGUCHI E,SHIMIZUTSUNEOYA-MANE S.Increasing productivity by removing choline in conversion of phosphatidylcholine to phosphatidylserine by phospholipase D[J].Journal of Fermentation&Bioengineering,1992,73(5):357-361.