发酵类脂溶性药物中残留DNA检测关键影响因素——溶剂的筛选与确定  

Critical factors for measuring the residual DNA in fermented lipid-soluble drugs--solvents choosing and optimising

作　　者：王琰[1] 张培培[1] 崔生辉[1] 姚尚辰[1] 胡昌勤[1] 许明哲[1] 张彦民[2] WANG Yan;ZHANG Pei-pei;CUI Sheng-hui;YAO Shang-chen;HU Chang-qin;XU Ming-zhe;ZHANG Yan-min(National Institutes for Food and Drug Control,Beijing 102629,China;Shool of Pharmacy,Xi’an Jiaotong University,Xi’an 710061,China)

机构地区：[1]中国食品药品检定研究院,北京102629 [2]西安交通大学药学院,西安710061

出　　处：《药物分析杂志》2020年第4期651-658,共8页Chinese Journal of Pharmaceutical Analysis

基　　金：国家“重大新药创制”科技重大专项资助项目(2017ZX09101001-007)。

摘　　要：目的:考察不同溶剂对发酵类脂溶性药物中残留DNA测定准确性的影响并确定最佳溶剂。方法:以脂溶性药物-环孢素为例,通过研究DNA检测全过程中各步骤溶解因素对检测结果的影响,从供试品溶液制备、供试品溶液与DNA试剂盒试剂相溶性及DNA溶液稳定性3个方面入手,对不同种类的溶剂进行考察和筛选,并根据荧光染色法的原理,采用Qubit■DNA检测试剂盒对环孢素供试品溶液中残留DNA进行测定,探讨溶剂选择的适用性和合理性。此外,应用筛选的溶剂对另一种脂溶性药物——两性霉素B中残留DNA进行了测定,进一步验证可靠性。结果:0.1 mol·L^-1盐酸-N,N-二甲基甲酰胺(1∶4)、0.1 mol·L^-1醋酸-N,N-二甲基甲酰胺(1∶4)和0.1 mol·L^-1柠檬酸-N,N-二甲基甲酰胺(1∶4)3种混合溶剂对脂溶性药物均有较好溶解性并能与DNA试剂较好相溶;进一步的DNA稳定性研究结果显示,0.1 mol·L^-1柠檬酸-N,N-二甲基甲酰胺(1∶4)对DNA结构与性质不产生影响,稳定性最好。结论:采用0.1 mol·L^-1柠檬酸-N,N-二甲基甲酰胺(1∶4)作溶剂,不仅对环孢素和两性霉素B具有较好的溶解性,而且对DNA不产生影响,稳定性较高,可实现终产品中残留DNA的准确测定。本文研究结果为脂溶性药物中残留DNA检测溶剂的选择提供了一定科学参考。Objective:To investigate the influence of solvents on the accuracy of measurement of residual DNA in lipid-soluble drugs and then determine the suitable solvents.Methods:In this paper,we took a case study on-cyclosporine,a liposoluble drug,investigating various kinds of solvents with focus on test sample solution,inter-compatibility of test sample solution with Qubit■working solution in DNA kit,and DNA stability in solvents as well.Residual DNA in cyclosporine samples was measured using Qubit■DNA kit based on fluorescence staining method.The applicability and rationality of solvent selection was also discussed.In addition,the selected solvent was also used for the determination of residual DNA in amphotericin B for the purpose of reliability verification.Results:0.1 mol·L^-1 hydrochloric acid-DMF(1∶4),0.1 mol·L^-1 acetic acid-DMF(1∶4)and 0.1 mol·L^-1 citric acid-DMF(1∶4)had better solubility for cyclosporine and better inter-compatibility with Qubit■working solution.However,further investigation showed that it had no effect on the structure and properties of DNA and presented the best stability when it took 0.1 mol·L^-1 citric acid-DMF(1∶4)as the solvent.Conclusion:Not only does 0.1 mol·L^-1 citric acid-DMF(1∶4)show good solubility for lipid-soluble drugs,but it also has high stability for DNA,which is an appropriate solvent for more accurate determination of residual DNA in fermented lipidsoluble drugs.

关 键 词：发酵类脂溶性药物 环孢素 两性霉素B 残留DNA 测定影响因素 溶剂筛选 质量控制 

分 类 号：R917[医药卫生—药物分析学]