Budinsky R A, Roberts S M, Coats E A, Adams L, Hess E V
Drug Metab Dispos. 1987 Jan-Feb;15(1):37-43.
A method is described, using HPLC and electrochemical detection, which permits the direct quantitation of procainamide hydroxylamine. Procainamide hydroxylamine was formed from procainamide by hepatic microsomes from both rat and human, with rat microsomes showing higher apparent formation rates. The apparent Km for formation of procainamide hydroxylamine was 0.044 mM for rat liver microsomes, with an apparent Vmax of 2.81 nmol/min/mg of protein. Estimates of Km from three human microsomal samples were 6.29, 2.89, and 6.88 mM. Vmax estimates were 0.31, 0.74, and 0.74 nmol/min/mg of protein, respectively, roughly an order of magnitude less than that observed for the rat. Microsomal formation in both species was inhibited by boiling the microsomes, eliminating NADPH from the incubation system, by preincubation with SKF 525A, cimetidine, or n-octylamine, or by gassing the microsomal incubation mixture with carbon monoxide. These observations suggest that procainamide hydroxylamine formation is cytochrome P-450 mediated. Procainamide hydroxylamine could not be detected in the blood of rats treated with a single dose of procainamide, 100 mg/kg, po. One potential reason for the inability to detect this metabolite in blood is indicated by the rapid disappearance in vitro of procainamide hydroxylamine added to whole blood. Most of this disappearance appears to be due to an interaction with hemoglobin.
本文描述了一种使用高效液相色谱法（HPLC）和电化学检测法直接定量测定普鲁卡因胺羟胺的方法。普鲁卡因胺羟胺由大鼠和人类的肝微粒体作用于普鲁卡因胺而形成，大鼠微粒体的表观形成速率更高。大鼠肝微粒体形成普鲁卡因胺羟胺的表观米氏常数（Km）为0.044 mM，表观最大反应速率（Vmax）为2.81 nmol/分钟/毫克蛋白质。来自三个不同人类微粒体样本的Km估计值分别为6.29、2.89和6.88 mM。Vmax估计值分别为0.31、0.74和0.74 nmol/分钟/毫克蛋白质，大约比大鼠观察值低一个数量级。两种物种的微粒体形成均受到以下因素的抑制：将微粒体煮沸、从孵育系统中去除烟酰胺腺嘌呤二核苷酸磷酸（NADPH）、与SKF 525A、西咪替丁或正辛胺预孵育，或向微粒体孵育混合物中通入一氧化碳。这些观察结果表明，普鲁卡因胺羟胺的形成是由细胞色素P-450介导的。在口服单剂量100 mg/kg普鲁卡因胺的大鼠血液中未检测到普鲁卡因胺羟胺。在血液中无法检测到这种代谢物的一个潜在原因是，添加到全血中的普鲁卡因胺羟胺在体外迅速消失。这种消失大部分似乎是由于与血红蛋白的相互作用。
