水相酶法催化制备L-α-甘油磷酸胆碱及其微观特性研究

发布时间：2018-03-04 21:28

  本文选题：磷脂酰胆碱　切入点：甘油磷酸胆碱　出处：《合肥工业大学》2017年硕士论文　论文类型：学位论文

【摘要】：磷脂酰胆碱(PC)及其水解产物L-α-甘油磷酸胆碱(GPC)广泛应用于医药和保健品领域。以磷脂酰胆碱为基础的生物转化产物在医药、食品、化妆等行业有重要用途。本文以富含磷脂酰胆碱的大豆卵磷脂为底物,以磷脂酶催化为例,研究了卵磷脂悬浮液酶催化水解前后的表面化学性质的变化规律及其催化制备甘油磷酸胆碱的工艺,为磷脂酰胆碱的生物转化提供了新的催化体系。论文首先将大豆卵磷脂高速均质为水相的卵磷脂悬浮液,对卵磷脂在水相中的表面化学性质进行了研究。结果表明:当缓冲液pH增加时,颗粒粒径变大,体系的电导率增大,Zeta电势绝对值降低,体系的粘度下降;磷脂酶为生物催化剂催化卵磷脂悬浮液时,卵磷脂悬浮液的Zeta电位出现波动,pH出现略微下降后最终保持在4.70左右,电导率略微上升后保持在9.50mS/cm左右,流体粘度因固含量下降而下降,最终会保持平衡。其次以卵磷脂悬浮液表面化学性质研究结果为基础,构建了大豆卵磷脂的酶法催化制备甘油磷酸胆碱的水相反应体系,并对其催化条件进行了研究。实验结果得到:初始底物浓度为20.0%,反应温度为55℃,初始pH为5.0,酶用量为87.03U/g,当反应进行到第90min时,体系的底物终浓度由20.0%降低至6.5%,此时,加入浓度为25.0%的底物,使得体系的终浓度由6.5%上升至13.0%,继续反应6h,反应的PC转化率达到90%左右。补料调节底物浓度的方式有利于酶催化反应的进行。论文最后研究了水相酶催化反应后的产物分离纯化工艺,该工艺先通过萃取的方法除去脂溶性杂质,萃取时二氯甲烷用量为反应液体积的0.4倍,甲醇的用量为0.5倍。产物经过碱化后再进行脱色。之后,使用溶剂除盐法去除部分乙酸钠,使用的溶剂为无水乙醇,溶解温度为55oC。最后,产物需要通过硅胶柱层析精制,其流动相为9:1的甲醇和30%氨水混合液。本文通过对磷脂酰胆碱水相催化体系的关键特性研究,确定了磷脂酶水相催化体系的pH、粘度、底物含量等参数,并在此体系中制备得到了 GPC产品,研究结果为绿色环保的磷脂酰胆碱生物转化技术提供理论依据。
[Abstract]:Phosphatidylcholine (PCP) and its hydrolysate L- 伪 -glycerophosphatidylcholine (L- 伪 -glycerophosphate choline) are widely used in the field of medicine and health care products. The biotransformation products based on phosphatidylcholine are used in medicine, food, In this paper, soybean lecithin rich in phosphatidylcholine is used as substrate and phospholipase catalysis as an example. The changes of surface chemical properties of lecithin suspension before and after catalytic hydrolysis and the preparation of glycerophosphate choline were studied. It provides a new catalytic system for the biotransformation of phosphatidylcholine. Firstly, soybean lecithin was homogenized into aqueous phosphatidylcholine suspension at high speed. The surface chemical properties of lecithin in aqueous phase were studied. The results showed that the particle size increased with the increase of buffer pH and the absolute value of Zeta potential decreased and the viscosity of the system decreased. When phospholipase was used as a biocatalyst to catalyze lecithin suspension, the Zeta potential of lecithin suspension fluctuated slightly and the pH value decreased slightly, and finally remained around 4.70, and the electrical conductivity increased slightly and remained at 9.50mS / cm or so. The viscosity of the fluid decreases because of the decrease of solid content, which will eventually maintain equilibrium. Secondly, based on the results of the study on the surface chemical properties of lecithin suspension, the aqueous phase reaction system of soybean lecithin catalyzed by enzymatic synthesis of glycerophosphate choline was constructed. The experimental results show that the initial substrate concentration is 20.0, the reaction temperature is 55 鈩，

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