【作者】 张韶瑜；

【导师】 刘昌孝；

【作者基本信息】 天津大学 ， 新药评价学， 2009， 博士

【摘要】 研究目的:痹祺胶囊是应用现代药物制剂理论精制而成的新药,主要用于风湿、类风湿性关节炎,颈椎病、肩周炎等疾病的治疗。具有疗效好、见效快、止痛作用好的特点。目前对于痹祺胶囊组方配伍的科学性及用药安全性尚未见报道。本实验是国家自然科学基金委重点项目(No. 30630075)的重要组成部分,通过体外实验及体内实验两方面全面评价痹祺胶囊配伍组方的基于细胞色素P450 (Cytochrome P450, CYP450)的安全性和科学性。研究方法:采用液相色谱-串联质谱法(LC-MS/MS)建立同时评价CYP1A2、CYP2C9、CYP2C19、CYP2D6和CYP3A酶活性的全新高通量分析方法,同时提取并检测5种特异性探针药物及其相应代谢产物,其中包括非那西丁/扑热息痛、甲苯磺丁脲/4-羟基甲苯磺丁脲/羧基甲苯磺丁脲、4’-羟基美芬妥英、右美沙芬/右啡烷、咪达唑仑/1’-羟基咪达唑仑。采用Gentest高通量重组CYP450酶抑制剂筛选试剂盒分别考察痹祺全药提取物、君药有效成分马钱子碱和士的宁及其与使药甘草的有效成分甘草酸和甘草次酸相结合对以上5种CYP450的体外抑制作用。采用“Cocktail”一点法测定痹祺胶囊配伍组分对Wistar大鼠体内5种CYP450的诱导和(或)抑制作用,并全面分析配伍规律。结果:本实验建立了全新“Cocktail”探针药物组合(非那西丁+甲苯磺丁脲+美芬妥英+右美沙芬+咪达唑仑),并建立了同时检测以上五种探针药物及其相应代谢产物的LC-MS/MS分析方法。马钱子碱和士的宁对5种CYP450基本没有体外抑制作用。马钱子碱和士的宁分别加上甘草酸和甘草次酸后对CYP2C9和CYP2C19可能存在体外抑制作用。大鼠体内实验结果显示,痹祺胶囊组方存在基于CYP450的配伍规律,与空白对照组比较,君药+臣药组、君药+使药组、君药+臣药+佐药组、君药+臣药+使药组、君药+佐药+使药组及痹祺胶囊全药组对CYP1A2具有显著诱导作用;君药+臣药组、君药+臣药+佐药组、君药+臣药+使药组及痹祺胶囊全药组对CYP2C9具有显著诱导作用。结论:本实验所建立的“Cocktail”探针药物组合为首次将此五种探针药物进行组合的全新方法,之前未见报道。痹祺胶囊组方存在基于CYP450较为显著的配伍规律,此规律为进一步探讨君药、臣药、佐药和使药在痹祺胶囊组方中的配伍作用地位,全面综合地分析与评价痹祺胶囊组方配伍的科学性奠定良好基础。更多还原

【Abstract】 Biqi capsulae is a new drug made and refined according to morden pharmaceutical preparation theory. It is for the treatment of rheumatic arthritis, rhermatoid arthritis, cervical spondylopathy, scapulohumeral periarthritis and so on. Biqi capsulae has the advantages of good and rapid healing efficacy and excellent pain relieveing effect. There is no report on the scientificalness and safety of prescription compatibility of Biqi capsulae. This study is an important part of Key Program of National Natural Science Foundation of China (No. 30630075). This study is aimed to investigate and evaluate the scientificalness and safety of Biqi capsulae prescription compatibility on cytochrome P450 (CYP450).A new high-throughput LC-MS/MS method was established to evaluate the in vivo activities of CYP1A2, 2C9, 2C19, 2D6 and 3A in rat plasma and urine. The five-specific probe substrates / metabolites include phenacetin / paracetamol (CYP1A2), tolbutamide / 4-hydroxytolbutamide / carboxytolbutamide (CYP2C9), mephenytoin / 4’-hydroxymephenytoin (CYP2C19), dextromethorphan / dextrorphan (CYP2D6) and midazolam / 1’-hydroxymidazolam (CYP3A). Using High-throughput Inhibitor Screening Kit, we determined the in vitro inhibitory effects of Biqi capsulae extract, active constituents of semen strychni (brucine and strychnine), brucine and strychnine combined with active constituents of Glycyrrhiza uralensis (glycyrrhetic acid and glycyrrhetinic acid) on the above 5 CYPs. Wistar rats were used to analyze and test the potential in vivo induction and (or) inhibition of constituents from Biqi prescription on the above 5 CYPs.A new“Cocktail”has been established, including probes of phenacetin, tolbutamide, mephenytoin, dextromephen and midazolam. A LC-MS/MS analytical method has been established to determine the above 5 probes and their corresponding metabolites. The results showed that brucine and strychnine had no in vitro inhibitory effect on 5 CYPs. It is possible that groups of brucine and strychnine combined with glycyrrhetic acid and glycyrrhetinic acid had in vitro inhibitory effects on CYP2C9 and CYP2C19. The in vivo results showed that Biqi capsulae prescription has compatibility on CYP450s. Compared with blank group, groups of“principal + assistant”,“principal + mediator”,“principal + assistant + complement”,“principal + assistant + mediator”,“principal + complement + mediator”and Biqi capsulae can significantly induce the activity of CYP1A2. Moreover, groups of“principal + assistant”,“principal + assistant + complement”,“principal + assistant + mediator”and Biqi capsulae can significantly induce the activity of CYP2C9.Biqi capsulae prescription has significant compatibility on CYP450s. These results provide important information and establish good foundation for further investigation on scientificalness and safety of Biqi capsulae prescription compatibility.更多还原