A micromethod for quantitation of debrisoquine and 4-hydroxydebrisoquine in urine by liquid chromatography

Autor(es)

V.A. Pereira ^[1] ; J.O. Auler Jr. ^[2] ; M.J. Carmona ^[3] ; F.H. Mateus ^[4] ; V.L. Lanchote ^[5] ; D.D. Breimer ^[6] ; S.R.C.J. Santos ^[7]

Afiliação do(s) autor(es)

^[1] Universidade de São Paulo - Brasil

^[2] Universidade de São Paulo - Brasil

^[3] Universidade de São Paulo - Brasil

^[4] Universidade de São Paulo - Brasil

^[5] Universidade de São Paulo - Brasil

^[6] Leiden-Amsterdam Center for Drug Research

^[7] Universidade de São Paulo - Brasil

• Tipo de documento: Artigo Científico
• Fonte: Brazilian Journal of Medical and Biological Research; v. 33, n. 5, p. 509-514, 2000-05-00.

Resumo

We describe a new simple, selective and sensitive micromethod based on HPLC and fluorescence detection to measure debrisoquine (D) and 4-hydroxydebrisoquine (4-OHD) in urine for the investigation of xenobiotic metabolism by debrisoquine hydroxylase (CYP2D6). Four hundred µl of urine was required for the analysis of D and 4-OHD. Peaks were eluted at 8.3 min (4-OHD), 14.0 min (D) and 16.6 min for the internal standard, metoprolol (20 µg/ml). The 5-µm CN-reverse-phase column (Shimpack, 250 x 4.6 mm) was eluted with a mobile phase consisting of 0.25 M acetate buffer, pH 5.0, and acetonitrile (9:1, v/v) at 0.7 ml/min with detection at lexcitation = 210 nm and lemission = 290 nm. The method, validated on the basis of measurements of spiked urine, presented 3 ng/ml (D) and 6 ng/ml (4-OHD) sensitivity, 390-6240 ng/ml (D) and 750-12000 ng/ml (4-OHD) linearity, and 5.7/8.2% (D) and 5.3/8.2% (4-OHD) intra/interassay precision. The method was validated using urine of a healthy Caucasian volunteer who received one 10-mg tablet of Declinax®, po, in the morning after an overnight fast. Urine samples (diuresis of 4 or 6 h) were collected from zero to 24 h. The urinary excretion of D and 4-OHD, Fel (0-24 h), i.e., fraction of dose administered and excreted into urine, was 6.4% and 31.9%, respectively. The hydroxylation capacity index reported as metabolic ratio was 0.18 (D/4-OHD) for the person investigated and can be compared to reference limits of >12.5 for poor metabolizers (PM) and <12.5 for extensive metabolizers (EM). In parallel, the recovery ratio (RR), another hydroxylation capacity index, was 0.85 (4-OHD: SD + 4-OHD) versus reference limits of RR <0.12 for PM and RR >0.12 for EM. The healthy volunteer was considered to be an extensive metabolizer on the basis of the debrisoquine test. (AU)

• Processo FAPESP: 97/14466-7 - Teste da debrisoquina na medida da atividade CYP2D6 em pacientes coronarianos submetidos à revascularização do miocárdio com circulação extracorpórea
• Beneficiário: Valeria Adriana Pereira Martins
• Modalidade de apoio: Bolsas no Brasil - Mestrado