超高效液相色谱-串联质谱法研究文多灵在大鼠肝微粒体中的体外代谢产物及途径  

Metabolites and metabolic pathways of vindoline in rat liver microsomal investigated by using ultra-high performance liquid chromatography/quadrupole time-of-flight mass spectrometry

作　　者：张玉倩 李德强[1] 解伟伟 张晓炜[1] 生晓娜[1] 靳怡然[1] 王巧 张国华[1] ZHANG Yu-qian;LI De-qiang;XIE Wei-wei;ZHANG Xiao-wei;SHENG Xiao-na;MU Xi-yan;JIN Yi-ran;WANG Qiao;ZHANG Guo-hua(Department of Pharmacy,Hebei Medical University Second Hospital,Shijiazhuang 050017,Hebei Province,China;Department of Pharmaceutical Analysis,School of Pharmacy,Hebei Medical University,Shijiazhuang 050017,Hebei Province,China)

机构地区：[1]河北医科大学第二医院药学部,河北石家庄050017 [2]河北医科大学药学院药物分析教研室,河北石家庄050017

出　　处：《中国临床药理学杂志》2020年第20期3355-3358,共4页The Chinese Journal of Clinical Pharmacology

基　　金：河北省人才培养工程基金资助项目(A201902012)。

摘　　要：目的建立超高效液相色谱-串联质谱法(UHPLC-MS/MS)法测定文多灵在大鼠肝微粒体中的代谢物及参与体外代谢的CYP450酶。方法大鼠肝微粒体样品采用乙酸乙酯萃取,色谱柱为Acquity UPLC@BEH C18(2.1×100 mm,1.7μm),柱温:40℃,流动相为3 mmol·L^-1乙酸铵缓冲溶液-乙腈进行梯度洗脱,流速为0.3 mL·min^-1,质谱采用电喷雾离子源,正离子检测方式,来鉴定在大鼠肝微粒体中文多灵的代谢产物。并利用α-萘黄酮,盐酸苯海拉明,磺胺苯吡唑,奎那定,二乙基二硫代氨基甲酸胺和酮康唑分别作为CYP1A2,CYP2B,CYP2C11,CYP2D1,CYP2E1,CYP3A的特异性抑制剂,确定介导文多灵代谢的CYP450酶亚型。结果文多灵在CYP450孵育体系中共检测22个新增产物峰其中发现5种新的体外代谢产物;CYP1A2,CYP2B,CYP2C11,CYP2D1,CYP2E1分别参与了17,7,6,8,3种文多灵的体外代谢产物的生成,而CYP3A参与了所有代谢产物的生成。结论该方法快速,灵敏,准确的鉴定大鼠体内文多灵的代谢产物,同时确定了参与文多灵代谢的酶亚型。Objective To establish an ultra performance liquid chromatography-tandem mass spectrometry(UPLC-MS/MS)method for identifying the metabolites of Vindoline(VDL)in rat liver microsomes,and determining the related metabolic CYP450 isoforms of rat liver microsomes.Methods Rat liver microsomes samples were extracted from rat liver microsomes with liquid-liquid by extraction ethyl acetate,then the supernatant was separated on Acquity UPLC@BEH C18(2.1×100 mm,1.7μm),and eluted by acetonitrile-water(containing 3 mmol·L^-1 of ammonium acetate)with a gradient elution.The flow rate was 0.3 mL·min^-1.Detection of metabolites of VDL in rat liver microsomes were achieved using positive ion electrospray ionization.In addition,in order to determining the related metabolic CYP450 isoforms of VDL in rat liver microsomes,α-naphthoflavone,diphenhydramine hydrochloride,sulfaphenazole,quinine,diethyldithiocarbamate and ketoconazole were applied as specific inhibitors for CYP1A2,CYP2B,CYP2C11,CYP2D1,CYP2E1 and CYP3A.Results The ion chromatograms of blank and control samples were compared,the 22 new product peaks were detected initially and 5 new ones were found;CYP1A2,CYP2B,CYP2C11,CYP2D1,and CYP2E1 are involved in the production of 17,7,6,8,and 3 metabolites of VDL,respectively,while CYP3A is involved in the production of all metabolites.Conclusion The established method in this study was simple,rapid,specific,and suitable for fast,sensitive,and accurate identifying metabolites of VDL in rats,and determining the enzyme subtypes involved in VDL metabolism.

关 键 词：文多灵 代谢 细胞色素450 超高效液相色谱-串联质谱法 特异性抑制剂探针 

分 类 号：R97[医药卫生—药品]