We aimed to explore the possible influence of CYP2C9 (*2, *3 and IVS8-109 A > T), CYP2C19 (*2, *3 and *17) and ABCB1 (1236C > T, 2677G > A/T and 3435C > T) on phenytoin (PHT) plasma concentrations in 64 Mexican Mestizo (MM) patients with epilepsy currently treated with PHT in mono- (n = 25) and polytherapy (n = 39). Genotype and allele frequencies of these variants were also estimated in 300 MM healthy volunteers. Linear regression models were used to assess associations between the dependent variables (PHT plasma concentration and dose-corrected PHT concentration) with independent variables (CYP2C9, CYP2C19 and ABCB1 genotypes, ABCB1 haplotypes, age, sex, weight, and polytherapy). In multivariate models, CYP2C9 IVS8-109 T was significantly associated with higher PHT plasma concentrations (t(64) = 2.27; P = 0.03). Moreover, this allele was more frequent in the supratherapeutic group as compared with the subtherapeutic group (0.13 versus 0.03, respectively; P = 0.05, Fisher's exact test). Results suggest that CYP2C9 IVS8-109 T allele may decrease CYP2C9 enzymatic activity on PHT. More research is needed to confirm findings.