Rasmussen B B, Brøsen K
Department of Clinical Pharmacology, Odense University, Denmark.
Ther Drug Monit. 1996 Jun;18(3):254-62. doi: 10.1097/00007691-199606000-00006.
Caffeine metabolism via the 3-demethylation pathway is sequentially catalyzed by cytochrome P4501A2 (CYP1A2), xanthine oxidase, and N-acetyltransferase. The activities of the three enzymes can be estimated from urinary metabolic ratios of four caffeine metabolites, 5-acetylamino-6-formylamino-3-methyluracil (AFMU), 1-methyluric acid (1MU), 1-methylxanthine (1MX), and 1,7-dimethyluric acid (17DMU), after the ingestion of caffeine. A method for quantitation of the four metabolites in human urine has been developed. The method is based on a one-step extraction with ethyl acetate/2-propanol followed by high-performance liquid chromatography with UV detection. The detection limit was 1 microM for AFMU, 1MU, and 1MX and 2 microM for 17DMU. The intraday and interday coefficients of variation were < 3% and < 7%, respectively, and the accuracy was within +/- 3%. The method was employed in a population study of 277 healthy volunteers, each of whom ingested 200 mg caffeine and provided a urine sample approximately 6 h later. The metabolite concentration ranges in the urines were 2.1-327 microM, 4.0-744 microM, 4.9-598 microM, and 6.4-260 microM for AFMU, 1MU, 1MX, and 17DMU, respectively. The CYP1A2 ratio (AFMU + 1MU + 1MX/17DMU) was significantly lower in women than in men, excluding smokers and oral contraceptive users. The CYP1A2 ratio was higher in smokers than in nonsmokers, confirming the induction of CYP1A2 by smoking. In women using oral contraceptives, the CYP1A2 ratio was, as expected, significantly lower than in women not using oral contraceptives. For the N-acetyltransferase ratio (AFMU/1MX) and the xanthine oxidase ratio (1MU/1MX), no differences were seen in terms of sex, smoking habits, or the use of oral contraceptives. All results are in agreement with previous reports on CYP1A2, N-acetyltransferase, and xanthine oxidase activities in humans. Thus, the method is both analytically and biologically reliable for the assessment of CYP1A2, N-acetyltransferase, and xanthine oxidase in humans.
咖啡因通过3-去甲基化途径的代谢依次由细胞色素P4501A2（CYP1A2）、黄嘌呤氧化酶和N-乙酰转移酶催化。摄入咖啡因后，可根据四种咖啡因代谢物5-乙酰氨基-6-甲酰氨基-3-甲基尿嘧啶（AFMU）、1-甲基尿酸（1MU）、1-甲基黄嘌呤（1MX）和1,7-二甲基尿酸（17DMU）的尿代谢比值来估算这三种酶的活性。已开发出一种定量人尿中这四种代谢物的方法。该方法基于用乙酸乙酯/2-丙醇一步萃取，然后进行带紫外检测的高效液相色谱分析。AFMU、1MU和1MX的检测限为1微摩尔/升，17DMU的检测限为2微摩尔/升。日内和日间变异系数分别<3%和<7%，准确度在±3%以内。该方法用于对277名健康志愿者的群体研究，每人摄入200毫克咖啡因，约6小时后提供一份尿样。尿中AFMU、1MU、1MX和17DMU的代谢物浓度范围分别为2.1 - 327微摩尔/升、4.0 - 744微摩尔/升、4.9 - 598微摩尔/升和6.4 - 260微摩尔/升。排除吸烟者和口服避孕药使用者后，女性的CYP1A2比值（AFMU + 1MU + 1MX/17DMU）显著低于男性。吸烟者的CYP1A2比值高于不吸烟者，证实吸烟可诱导CYP1A2。正如预期的那样，使用口服避孕药的女性的CYP1A2比值显著低于未使用口服避孕药的女性。对于N-乙酰转移酶比值（AFMU/1MX）和黄嘌呤氧化酶比值（1MU/1MX），在性别、吸烟习惯或口服避孕药使用方面未观察到差异。所有结果均与先前关于人类CYP1A2、N-乙酰转移酶和黄嘌呤氧化酶活性的报道一致。因此，该方法在分析和生物学上对于评估人类的CYP1A2、N-乙酰转移酶和黄嘌呤氧化酶都是可靠的。
