超高效液相色谱-高分辨质谱分析大鼠肝微粒体中厚朴酚与和厚朴酚的体外代谢
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摘要
采用CYP450亚型酶特异性抑制剂,结合超高效液相色谱-高分辨质谱(UPLC-HRMS)对厚朴活性成分(厚朴酚、和厚朴酚)在肝微粒体中的体外代谢产物及主导代谢的细胞色素P450(CYP450)亚型酶进行分析与鉴定。特异性抑制剂实验显示CYP2E1亚型酶可能是主导厚朴酚苯环羟基化代谢产物的亚型酶,CYP1A2和CYP2C是主导厚朴酚氧化代谢的亚型酶。CYP2C是将和厚朴酚转化为氧化代谢产物的主要的CYP450亚型酶,CYP1A2和CYP2E1可能是负责将和厚朴酚转化为其羟基化和氢化代谢产物的主要的CYP450亚型酶,CYP2B6亚型酶很可能用于指导和厚朴酚苯环羟基化代谢产物。不同CYP450亚型酶在厚朴酚、和厚朴酚与大鼠肝微粒体体外代谢的实验中,呈现出对氧化反应和羟基化反应不同的催化活性,提示在临床用药中应注意药物-药物、药物-酶之间的相互作用。
CYP450 subtype enzyme specific inhibitor,combined with ultra-high performance liquid chromatography-high resolution mass spectrometry(UPLC-HRMS) was used for the analysis and identification of in-vitro metabolites of Magnolia active ingredients(magnolol,honokiol) in liver microsomes and the predominantly metabolized cytochrome P450(CYP450) enzyme subtype.Specific inhibitor experimental results showed that CYP2 E1 was likely to be the major enzyme responsible for the transformation of magnolol into its benzene ring hydroxylation metabolite in RLMs.CYP1 A2 and CYP2 C appeared to be the major enzymes responsible for the oxidation of magnolol to its metabolite in RLMs.The results indicated that CYP2 C was the main CYP450 isoenzymes transforming honokiol into its oxidated metabolites in RLMs.CYP1 A2 and 2 E1 were likely to be the major enzymes responsible for the transformation of honokiol into its hydroxylated and hydrogenated metabolites in RLMs.The CYP2 B6 subtype enzyme was likely to be used to convert honokiol to benzene ring hydroxylated metabolites.This study demonstrated that different cytochrome P450(CYP450) enzyme isoforms showed different activities in the in-vitro metabolism of magnolol and honokiol and rat liver microsomes,suggested that we should pay attention to drug-drug,drug-enzyme interactions in clinical medications.
CYP450 subtype enzyme specific inhibitor,combined with ultra-high performance liquid chromatography-high resolution mass spectrometry(UPLC-HRMS) was used for the analysis and identification of in-vitro metabolites of Magnolia active ingredients(magnolol,honokiol) in liver microsomes and the predominantly metabolized cytochrome P450(CYP450) enzyme subtype.Specific inhibitor experimental results showed that CYP2 E1 was likely to be the major enzyme responsible for the transformation of magnolol into its benzene ring hydroxylation metabolite in RLMs.CYP1 A2 and CYP2 C appeared to be the major enzymes responsible for the oxidation of magnolol to its metabolite in RLMs.The results indicated that CYP2 C was the main CYP450 isoenzymes transforming honokiol into its oxidated metabolites in RLMs.CYP1 A2 and 2 E1 were likely to be the major enzymes responsible for the transformation of honokiol into its hydroxylated and hydrogenated metabolites in RLMs.The CYP2 B6 subtype enzyme was likely to be used to convert honokiol to benzene ring hydroxylated metabolites.This study demonstrated that different cytochrome P450(CYP450) enzyme isoforms showed different activities in the in-vitro metabolism of magnolol and honokiol and rat liver microsomes,suggested that we should pay attention to drug-drug,drug-enzyme interactions in clinical medications.
引文
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[3] WU J,ZHONG R L,WANG D W,et al.Chinese Journal of Experimental Traditional Medical Formulae(武洁,钟荣玲,王大为,等.中国实验方剂学杂志),2014,20(23):110.
[4] Chinese Pharmacopoeia Commisstion.Pharmacopoeia of People's Republic of China(Ⅰ).Beijing:China Medical Science Press(国家药典委员会.中华人民共和国药典(I部).北京:中国医药科技出版社,2015:539,553,610.
[5] Eun Jung Kwak,Young Soon Lee,Eun Mi Choi.Mediators of Inflammation,2012,(1):829650.
[6] Akiko Muroyama,Aya Fujita,Cheng Lv.Parkinson's Disease,2012,(4):985157.
[7] Zhao C ,Liu Z Q.Biochimie,2011,93(10):1755.
[8] WANG J J.Experimental Study on Effect and Mechanism of Honokiol on Type 2 Diabetes Mellitus.Wuhan:Hubei University(王俊俊.和厚朴酚抗2型糖尿病作用与机制实验研究.武汉:湖北大学),2016.
[9] Lan K H,Wang Y W,Lee W P,et al.Liver Int,2012,32(6):989.
[10] Zuo G Y,Zhang X J ,Han J.BMC Complementary & Alternative Medicine ,2015,15(1):425.
[11] Bjornsson T D,Callaghan J T,Einolf H J,et al.Journal of Clinical Pharmacology,2003,43(5):443.
[12] Weaver R,Graham K S,Beattie I G,et al.Drug Metabolism and Disposition:the Biological Fate of Chemicals,2003,31(7):955.
[13] Kim K B,Seo K A,Yoon Y J,et al.Xenobiotica,2008,38(9):1165.
[14] Yao H T ,Chang Y W ,Lan S J.Life Sciences,2006,79(26):2432.
[15] Li Shizhe,Liu Shu,Pi Zifeng.Journal of Pharmaceutical and Biomedical Analysis,2007,138:289.