PHENOTYPING OF CYP2C19 WITH ENANTIOSPECIFIC HPLC-QUANTIFICATION OF R-MEPHENYTOIN AND S-MEPHENYTOIN AND COMPARISON WITH THE INTRON4 EXON5 G-]A-SPLICE SITE MUTATION/

S-Mephenytoin 4'-hydroxylase (CYP2C19) is a genetically polymorphic cy tochrome P450. A modified method for CYP2C19 phenotyping was evaluated in 174 healthy German volunteers and the results were compared with g enotyping for the intron4/exon5 G-->A splice site mutation (mi) of CYP 2C19, associated with the poor metabolizer (PM) phenotype. A smaller t han usual test-dose of 50 mg (R,S)-mephenytoin was used and urine samp les were collected from 0 to 5 h and from 5 to 8 h after administratio n Trait measurements included the mephenytoin SIR enantiomeric ratio a nd the hydroxylation index (i.e. the molar ratio of 4'-hydroxy-mepheny toin urinary recovery to the administered S-mephenytoin dose). S- and R-mephenytoin were quantified by isocratic: HPLC with a Chiraspher(TM) column and 80% n-hexane and 20% dioxane as the mobile phase. Ah indiv iduals from whom DNA was available (n = 140, including six phenotypica lly identified PMs) were analysed for the mi mutation. The population frequency of this CYP2C19 mutation was 0.15. Four individuals were hom ozygous for m(1) having S/R ratios of 0.9 or greater in both intervals of urine collection. Thus, individuals with an SIR ratio greater than or equal to 0.9 were classified as PMs and seven of all 174 phenotype d individuals were PMs (4%; 95% confidence limits: 1.6-8.1%), Heterozy gous carriers of m(1) (n = 34) had a median SIR ratio (5-8 h urine) of 0.06 compared to 0.01 in individuals without this mutation (n = 102; p = 0.0005, Mann-Whitney U-test). No such gene-dose relation was appar ent with the hydroxylation index. CYP2C9 may be involved in R-mephenyt oin hydroxylation; thus, the Arg(144)/Cys(144)-polymorphism of CYP2C9 was determined (allele frequency of the Cys(144)-variant: 0.13, n = 12 7). A major influence of this polymorphism on the recovery of R-mephen ytoin was not detected. In summary, the proposed HPLC-method for S- an d R-enantiomers of mephenytoin In 5-8 h urine samples reliably discrim inates poor and extensive metabolizers of CYP2C19 as confirmed by geno typing.