Lautala P, Kivimaa M, Salomies H, Elovaara E, Taskinen J
Department of Pharmacy, University of Helsinki, Finland.
Pharm Res. 1997 Oct;14(10):1444-8. doi: 10.1023/a:1012133008134.
Nitrocatechol COMT inhibitors are a new class of bioactive compounds, for which glucuronidation is the most important metabolic pathway. The objective was to characterize the enzyme kinetics of nitrocatechol glucuronidation to improve the understanding and predicting of the pharmacokinetic behavior of this class of compounds.
The glucuronidation kinetics of seven nitrocatechols and 4-nitrophenol, the reference substrate for phenol UDP-glucuronosyltransferase activity, was measured in liver microsomes from creosote-treated rats and determined by non-linear fitting of the experimental data to the Michaelis-Menten equation. A new method that combined densitometric and radioactivity measurement of the glucuronides separated by HPTLC was developed for the quantification.
Apparent K(m) values for the nitrocatechols varied greatly depending on substitution pattern being comparable with 4-nitrophenol (0.11 mM) only in the case of 4-nitrocatechol (0.19 mM). Simple nitrocatechols showed two-fold Vmax values compared with 4-nitrophenol (68.6 nmol min-1 mg-1), while all disubstituted catechols exhibited much lower glucuronidation rate. Vmax/K(m) values were about 10 times higher for monosubstituted catechols compared to disubstituted ones. The kinetic parameters for COMT inhibitors were in the following order: K(m) nitecapone > > entacapone > tolcapone; Vmax nitecapone > entacapone > tolcapone; Vmax/K(m) tolcapone > nitecapone > entacapone.
Nitrocatechols can in principle be good substrates of UGTs. However, substituents may have a remarkable effect on the enzyme kinetic parameters. The different behaviour of nitecapone compared to the other COMT inhibitors may be due to its hydrophilic 5-substituent. The longer elimination half-life of tolcapone in vivo compared to entacapone could not be explained by glucuronidation kinetics in vitro.
硝基儿茶酚儿茶酚-O-甲基转移酶（COMT）抑制剂是一类新型生物活性化合物，其葡萄糖醛酸化是最重要的代谢途径。目的是表征硝基儿茶酚葡萄糖醛酸化的酶动力学，以增进对这类化合物药代动力学行为的理解和预测。
在杂酚油处理大鼠的肝微粒体中测量了七种硝基儿茶酚和4-硝基苯酚（苯酚UDP-葡萄糖醛酸基转移酶活性的参考底物）的葡萄糖醛酸化动力学，并通过将实验数据非线性拟合到米氏方程来确定。开发了一种结合密度测定法和放射性测量法的新方法，用于定量由高效薄层层析（HPTLC）分离的葡萄糖醛酸苷。
硝基儿茶酚的表观米氏常数（K(m)）值因取代模式而异，仅在4-硝基儿茶酚（0.19 mM）的情况下与4-硝基苯酚（0.11 mM）相当。简单的硝基儿茶酚与4-硝基苯酚（68.6 nmol·min⁻¹·mg⁻¹）相比，显示出两倍的最大反应速度（Vmax）值，而所有二取代儿茶酚的葡萄糖醛酸化速率均低得多。单取代儿茶酚的Vmax/K(m)值比二取代儿茶酚高约10倍。COMT抑制剂的动力学参数顺序如下：K(m) 硝苯地平>>恩他卡朋>托卡朋；Vmax 硝苯地平>恩他卡朋>托卡朋；Vmax/K(m) 托卡朋>硝苯地平>恩他卡朋。
硝基儿茶酚原则上可以是尿苷二磷酸葡萄糖醛酸基转移酶（UGTs）的良好底物。然而，取代基可能对酶动力学参数有显著影响。硝苯地平与其他COMT抑制剂相比的不同行为可能归因于其亲水性的5-取代基。托卡朋在体内的消除半衰期比恩他卡朋长，这无法用体外葡萄糖醛酸化动力学来解释。
