Enantioselectivity of debrisoquine 4-hydroxylation in Brazilian Caucasian hypertensive patients phenotyped as extensive metabolizers

Debrisoquine (D), an antihypertensive drug metabolized to 4-hydroxydebrisoq uine (4-OHD) by CYP2D6, is commonly used as an in vivo probe of CYP2D6 acti vity and can be used to phenotype individuals as either extensive (EMs) or poor metabolizers (PMs) of such drugs as beta -adrenergic blockers, tricycl ic antidepressants, and class 1C antiarrnythmics. This report describes rev ersed-phase HPLC systems by which D and 4-OHD or S-(+) and R-(-)-4-OHD in u rine are more selectively quantified without the need for derivatization te chniques. We also studied the urinary excretion of R-(-)- and S-(+)-4-hydro xydebrisoquine in EM hypertensive patients in order to determine weather 4- OHD formation exhibits enantioselectivity. Twelve patients with mild to sev ere essential hypertension were admitted to the study. They received a sing le tablet of Declinax containing 10 mg debrisoquine sulfate. All the urine excreted during the following 8 h was collected. The debrisoquine metabolic ratio (DMR) was calculated as % of dose excreted as D/% of dose excreted a s 4-OHD and the debrisoquine recovery ratio (DRR) was calculated as % of do se excreted as 4-OHD/% of dose excreted as D+4-OHD. Debrisoquine and its me tabolite were determined in urine by HPLC using a reversed-phase Select B L iChrospher column, a mobile phase of 0.25 N acetate buffer, pH 5-acetonitri le (9.1, v/v) and a fluorescence detector. The limit of quantitation was de termined to be 25.0 ng/ml for D and 18.75 ng/ml for 4-OHD. Intra- and inter -day relative standard deviations (RSDs) were less than 10%. All hypertensi ve patients studied showed a DMR of less than 12.6 or a DRR higher than 0.1 2 and were classified as EMs. Direct enantioselective separation on chiral stationary phase involved resolution of S-(+)-4-OHD and R-(-)-4-OHD on a Ch iralcel OD-R column with a mobile phase of 0.125 N sodium perchlorate, pH 5 -acetonitrile-methanol (85:12:3, v/v/v). The quantitation limit of each ena ntiomer was 3.75 ng/ml of urine. Intra- and inter-day RSDs were less than 1 0% for each enantiomer. A high degree of enantioselectivity in the 4-hydrox ylation of D favouring the S-(+) enantiomer was observed, resulting in R-(- )4-OHD not detected in the urine of the EM hypertensive patients studied. ( C) 2000 Elsevier Science B.V. All rights reserved.