毛果芸香碱对香豆素7-羟化的竞争性抑制作用及其与小鼠CYP 2A5和人CYP 2A6的相互作用。

Competitive inhibition of coumarin 7-hydroxylation by pilocarpine and its interaction with mouse CYP 2A5 and human CYP 2A6.

Kinonen T, Pasanen M, Gynther J, Poso A, Järvinen T, Alhava E, Juvonen R O

Department of Pharmacology and Toxicology, University of Kuopio, Finland.

Br J Pharmacol. 1995 Nov;116(6):2625-30. doi: 10.1111/j.1476-5381.1995.tb17217.x.

Abstract

We have shown earlier that pilocarpine strongly inhibits mouse and human liver coumarin 7-hydroxylase activity of CYP 2A and pentoxyresorufin O-deethylase activity of CYP 2B in vitro. Since pilocarpine, like coumarin, contains a lactone structure we have studied in more detail its inhibitory potency on mouse and human liver coumarin 7-hydroxylation. 2. Pilocarpine was a competitive inhibitor of coumarin 7-hydroxylase in vitro both in mouse and human liver microsomes although it was not a substrate for CYP 2A5. Ki values were similar, 0.52 +/- 0.22 microM in mice and 1.21 +/- 0.51 microM in human liver microsomes. 3. Pilocarpine induced a type II difference spectrum in mouse, human and recombinant CYP 2A5 yeast cell microsomes, with Ka values of 3.7 +/- 1.6, 1.6 +/- 1.1 and 1.5 +/- 0.1 microM, respectively. 4. Increase in pH of the incubation medium from pH 6 to 7.5 increased the potency of inhibition of coumarin 7-hydroxylation by pilocarpine. 5. Superimposition of pilocarpine and coumarin in such a way that their carbonyls, ring oxygens and the H-7' of coumarin and N-3 of pilocarpine overlap yielded a common molecular volume of 82%. 6. The results indicate that pilocarpine is a competitive inhibitor and has a high affinity for mouse CYP 2A5 and human CYP 2A6. In addition the immunotype nitrogen of pilocarpine is coordinated towards the haem iron in these P450s.

摘要

我们先前已表明，毛果芸香碱在体外能强烈抑制小鼠和人肝脏中细胞色素P450 2A的香豆素7-羟化酶活性以及细胞色素P450 2B的戊氧基试卤灵O-脱乙基酶活性。由于毛果芸香碱与香豆素一样含有内酯结构，我们更详细地研究了其对小鼠和人肝脏香豆素7-羟化作用的抑制效力。2. 毛果芸香碱在体外对小鼠和人肝脏微粒体中的香豆素7-羟化酶均为竞争性抑制剂，尽管它不是细胞色素P450 2A5的底物。其抑制常数（Ki值）相似，在小鼠肝脏微粒体中为0.52±0.22微摩尔/升，在人肝脏微粒体中为1.21±0.51微摩尔/升。3. 毛果芸香碱在小鼠、人及重组细胞色素P450 2A5酵母细胞微粒体中诱导出II型差示光谱，其解离常数（Ka值）分别为3.7±1.6、1.6±1.1和1.5±0.1微摩尔/升。4. 将孵育介质的pH从6提高到7.5可增强毛果芸香碱对香豆素7-羟化作用的抑制效力。5. 毛果芸香碱与香豆素以其羰基、环上氧原子以及香豆素的H-7′和毛果芸香碱的N-3相互重叠的方式叠加，得到的共同分子体积为82%。6. 结果表明，毛果芸香碱是一种竞争性抑制剂，对小鼠细胞色素P450 2A5和人细胞色素P450 2A6具有高亲和力。此外，毛果芸香碱的免疫型氮原子与这些细胞色素P450中的血红素铁配位。