基于UHPLC-Q/Orbitrap-MS/MS鉴定丹葛酚酮胶囊体内代谢产物研究

Study on metabolites in vivo of Dangefentong Capsules based on UHPLC-Q/Orbitrap-MS/MS

丹葛酚酮胶囊是基于丹参葛根对药,以丹参酚酸、丹参酮和葛根黄酮为药物成分,治疗糖尿病周围神经病变的组分中药新药。探究丹葛酚酮胶囊体内化学成分及代谢产物,对阐明药物药效物质基础及作用机制具有重要意义。该研究利用超高效液相色谱-四极杆/静电场轨道阱质谱(UHPLC-Q/Orbitrap-MS/MS),以Eclipse plus C(2.1 mm×50 mm,1.8μm)为色谱柱,0.1%甲酸水溶液(A)-0.1%甲酸乙腈溶液(B)为流动相进行梯度洗脱,采用加热电喷雾离子源(HESI),扫描方式为正、负离子模式,质量扫描范围m/z 100~1000,快速分析Beagle犬灌胃丹葛酚酮胶囊后体内代谢产物和生物转化途径。以雄性Beagle犬为实验对象,灌胃丹葛酚酮胶囊,收集血浆、尿液、粪便样品,采用高分辨质谱结合对照品和文献分析鉴定体内原型成分和代谢产物。结果显示,共鉴定出107个化学成分,丹葛酚酮胶囊原型成分58个,包括42个丹参成分和16个葛根成分,代谢产物49个,包括21个丹参代谢产物和28个葛根代谢产物。代谢产物主要来自丹参酚酸类、丹参酮类和葛根黄酮类成分甲基化、羟基化、硫酸化、葡萄糖醛酸化等。该实验快速分析灌胃丹葛酚酮胶囊后Beagle犬体内化学成分,为进一步阐明丹葛酚酮胶囊药效物质基础及作用机制提供参考。

Dangefentong Capsules is a new traditional Chinese medicine preparation for the treatment of diabetic peripheral neuropathy.It is based on the Salviae Miltiorrhizae Radix et Rhizoma-Puerariae Lobatae Radix herb pair with salvianolic acids,tanshinones and pueraria flavonoids as main components.Studying the chemical composition in vivo of Dangefentong Capsules and its metabolites is of great significance for making clear its pharmacodynamic material basis and the action mechanism.The UHPLC-Q/Orbitrap-MS/MS was applied to rapidly analyze the metabolites and metabolic pathways of Dangefentong Capsules in Beagle dogs after gavage.Eclipse plus Ccolumn(2.1 mm×50 mm,1.8μm)was used,and gradient elution was performed with 0.1%formic acid aqueous solution(A)-formic acid acetonitrile solution(B).A heated electrospray ion source(HESI)was employed.The scanning mode was set as the positive and negative ion mode,and the mass scanning range was m/z 100-1000.The plasma,urine and feces samples were collected after male Beagle dogs were administered with Dangefentong Capsules.The prototype components and metabolites were identified by UHPLC-Q/Orbitrap-MS/MS analysis combined with reference substances and references.The results showed that 107 chemical components were identified,including 58 prototype components and 49 metabolites.The identified prototype components included 42 components from Salviae Miltiorrhizae Radix et Rhizoma and 16 components from Puerariae Lobatae Radix.The metabolites consist of 21 and 28 metabolites of Salviae Miltiorrhizae Radix et Rhizoma and Puerariae Lobatae Radix,respectively.They are mainly derived from the methylation,hydroxylation,sulfation and glucuronidation of salvianolic acids,tanshinones and pueraria flavonoids.This research rapi-dly analyzes the chemical components in vivo of Beagle dogs administered with Dangefentong Capsules,laying a basis for illustrating the pharmacodynamic material basis and mechanism of Dangefentong Capsules.