基于UHPLC-Q-Orbitrap HRMS技术的肾康注射液在大鼠体内代谢产物的鉴定及代谢途径分析

Metabolites identification and metabolic pathway analysis of Shenkang Injection in rats based on UHPLC-Q-Orbitrap HRMS

目的 采用UHPLC-Q-Orbitrap HRMS技术鉴定肾康注射液(Shenkang Injection,SKI)在大鼠体内的代谢产物,并提出可能的代谢途径。方法 SD大鼠经尾iv SKI后,收集其血浆、尿液和粪便样本。以0.1%甲酸水溶液(A)-乙腈(B)为流动相,梯度洗脱,Waters BEH C18柱进行色谱分离;使用Q Exactive HRMS和加热电喷雾离子源(HESI)进行正、负离子模式下的质谱扫描分析。根据“提取-化合物筛选-鉴定-验证”的大鼠体内代谢产物的鉴定策略对质谱数据进行分析。结果在大鼠血浆、尿液和粪便中共鉴定出121种外源化合物,包括29种原型化合物和92种代谢产物。这些代谢产物主要经历的生物转化有羟基化、还原和去饱和等Ⅰ相代谢反应,以及甲基化、乙酰化、磺酸化、葡萄糖醛酸化、甘氨酸结合、半胱氨酸结合和牛磺酸结合等Ⅱ相代谢反应。结论 为SKI的体内药理学和作用机制的进一步研究提供了物质基础,所提出的鉴定策略为中药的体内药效物质研究提供了一种系统、全面、可靠的方法。

Objective To identify the metabolites of Shenkang Injection(肾康注射液,SKI)in rats by UHPLC-Q-Orbitrap HRMS and propose the possible metabolic pathway.Methods The plasma,urine and feces samples were collected after SD rats were administrated with SKI by injecting the tail vein.The Waters BEH C18 column was used to separate the samples with 0.1%formic acid solution(A)-acetonitrile(B)as mobile phase for gradient elution.The mass spectrometry scanning was carried out in positive and negative ion modes by Q Exactive HRMS and heated electrospray ion source(HESI).The mass spectrometry data were analyzed according to proposed strategy of“extraction-compound screening-identity-validation”for the identification of metabolites in rats.Results A total of 121 xenobiotics were identified in rat plasma,urine and feces,including 29 prototype compounds and 92 metabolites.The main biotransformations of these metabolites refer to phase I metabolic reactions such as hydroxylation,reduction and desaturation,and phase II metabolic reactions such as methylation,acetylation,sulfation,glucuronidation,glycine conjugation,cysteine conjugation and taurine conjugation.Conclusion This study provides a material basis for further study of pharmacology and mechanism of action of SKI in vivo.The proposed identification strategy provides a systematic,comprehensive and reliable method for the study of pharmacodynamic substances of traditional Chinese medicine in vivo.