CYP2D6-Dependent Bufuralol 1-Hydroxylation Assayed by Reversed-Phase lon-Pair High-Performance Liquid Chromatography with Fluorescence Detection

CYP2D6 is an enzyme that is subject to genetic polymorphism
1
. This P450 is absent in individuals with the poor metabolizer CYP2D6 phenotype
2
,
3
), resulting in a substantially reduced capacity to metabolize a large number of clinically useful drugs, including metoprolol, propafenone, haloperrdol, dextromethorphan and codeine (
1
). Quinidine competrtively inhibits CYP2D6, with a K
1
of 3-30 m
M
(
3
–
5
). Because of its preferential inhibition of CYP2D6 (
67
), quinidine is frequently used as an inhibitory chemical probe of this polymorphically expressed P450. Diagnostic catalytic monitors of human hepatic CYP2D6 include debrisoquine 4-hydroxylase (
4
), dextromethorphan Odemethylase (
8
), and bufuralol l’-hydroxylase activities (
2
–
4
). Advantages of using bufuralol as a CYP2D6-selective substrate include the high sensitivity of the assay owing to the highly fluorescent l’-hydroxybufuralol metabohte and the fact that the use of radiolabeled substrate is not required. This chapter describes a modification of a reversed-phase ion-pair high-performance liquid chromatographic (HPLC) assay (
9
) with fluorescence detection for the determmation of bufuralol l’-hydroxylation activity. Methods for other CYP2D6 assays such as debrisoquine 4-hydroxylase (
9
) and dextromethorphan O-demethylase (
9
,
10
) can be found in the cited references.