雌二醇偶联碱性磷酸酶工艺改进  

Improvement of Estradiol Coupled Alkaline Phosphatase Process

作　　者：王一恺 WANG Yikai(Shanghai Kehua Bio-Engineering Co.,Ltd.,Shanghai 200233,China)

机构地区：[1]上海科华生物工程股份有限公司,上海200233

出　　处：《生物化工》2022年第2期1-6,11,共7页Biological Chemical Engineering

摘　　要：目的:研究并探索雌二醇偶联碱性磷酸酶的制备工艺,解决原有混合酸酐法制备工艺对人体与环境的毒性、产物不稳定和检测值的相关性偏离问题。方法:采用混合酸酐法及碳二亚胺试剂偶联雌二醇与碱性磷酸酶,在碳二亚胺法中添加N-羟基琥珀酰亚胺(NHS),对比混合酸酐法与碳二亚胺法灵敏度、稳定性及相关性。结果:对比加入NHS后增强反应活性的能力,碳二亚胺法制备的酶结合物稳定性更好、灵敏度高、相关性更好。结论:雌二醇的衍生物采用雌二醇碳3位衍生物(Estradiol3-CME),碳二亚胺试剂(1-(3-二甲氨基丙基)-3-乙基碳二亚胺盐酸盐)(EDC)偶联,再加入微量的NHS作为催化剂,碱性磷酸酶与雌二醇衍生物的反应摩尔比为1∶10,此条件下可以得到活性较好,稳定性较强的酶结合物。Objective: This paper aims to study and explore the preparation process of other estradiol coupled alkaline phosphatase, and improve the preparation process of original mixed anhydride on human body and environment toxicity, the instability of products and correlation deviation of tested values. Methods: Mixed anhydride method and carbodiimide reagent were used to couple estradiol and alkaline phosphatase, and N-hydroxysuccinimide(NHS) was added to carbodiimide method. The sensitivity, stability and correlation of mixed anhydride method and the carbodiimine method were compared. Results: Compared with the ability to enhance the reactivity after adding NHS, its showes that the enzyme conjugate prepared by carbodiimide method had better stability, high sensitivity and better correlation.Conclusion: Estradiol 3-CME, carbodiimide EDC coupling, and NHS as a catalyst are used for estradiol derivatives. The molar ratio of alkaline phosphatase to estradiol derivatives is 1 ∶ 10. In this condition, the enzyme conjugation with good activity and stability can be obtained.

关 键 词：雌二醇 碱性磷酸酶 混合酸酐法 碳二亚胺 化学发光法 

分 类 号：O621.2[理学—有机化学]