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Phenotyping of N-acetyltransferase type 2 by caffeine from uncontrolled dietary exposure
Alexander Jetter1, Martina Kinzig-Schippers, Michael Illauer
1Department of Pharmacology, Clinical Pharmacology, University of Cologne, Gleueler Strasse 24, 50931 Köln, Germany. alexander.jetter@medizin.uni-koeln.de
Background And Objective:
The standard approach for phenotyping of the human arylamine N-acetyltransferase 2 (NAT2) uses urinary caffeine metabolite ratios after a caffeine test dose taken in after methylxanthine abstinence. We tested whether these standardization measures were still needed when a more sensitive quantification technique was used.
Methods:
A new liquid chromatography/tandem mass spectrometry (LC-MS/MS) method for the quantification of the caffeine metabolites 5-acetylamino-6-formylamino-3-methyluracil (AFMU), 5-acetylamino-6-amino-3-methyluracil (AAMU), 1-methylxanthine (1X), and 1-methylurate (1U) was developed. Urine samples from 77 healthy volunteers collected before and 5-6 h after oral intake of 150-200 mg caffeine were analyzed. The lower limits of quantification were 0.1 microg/ml for caffeine, 1X, 1U, and AFMU, and 0.2 microg/ml for AAMU.
Results:
The urinary NAT2 ratios (AFMU+AAMU) / (AFMU+AAMU+1X+1U) before and after caffeine intake correlated well in 65 volunteers (r(2)=0.827; P< 0.0001). In 12 participants (16%), metabolite concentrations in urine before caffeine intake were below the quantification limit. NAT2 genotyping, done in 41 volunteers for four SNPs, corroborated the phenotyping results.
Conclusion:
NAT2 activity can be determined from a spontaneous urine probe in most subjects by quantification of caffeine metabolites arising from non-standardized dietary caffeine exposure using LC-MS/MS. This may facilitate the phenotyping procedure.
