CYP2S1 is a recently discovered member of the cytochrome P450 (CYP) gene superfamily. Interestingly, even though the DNA sequence identifies it as the sole member of the new CYP2S family, CYP2S1 exhibits many features typical to CYP1 family members, e.g. dioxin-inducibility mediated by the aryl hydrocarbon receptor (AHR) and the aryl hydrocarbon receptor nuclear translocator (ARNT). In addition, CYP2S1 metabolises some aromatic hydrocarbons as well as cellular substances. These characteristics, together with a wide extrahepatic tissue distribution, suggest that CYP2S1 may have an important role in both exogenous and endogenous metabolism. This is the first study characterising CYP2S1 alleles and naming them with the recommended CYP allele nomenclature. We used denaturing gradient gel electrophoresis (DGGE) and direct sequencing to investigate genetic variation of CYP2S1 in 100 male Finnish Caucasians. Those exons in which variation was found were examined in subsequent 100 subjects. The coding region of all of the nine exons, as well as a 449 bp fragment of the proximal promoter region, was analysed. This systematic investigation revealed eight single nucleotide polymorphisms (SNPs), which comprise nine different variant alleles (haplotypes), in addition to the wild-type allele. Seven of the SNPs occurred in the protein-coding areas and one in the proximal 3' untranslated region (3'UTR). Two of these sequence variations (10347C > T and 13106C > T) result in non-conservative amino acid substitutions, i.e. Arg380Cys and Pro466Leu, respectively. The respective allelic variants, CYP2S1*2 ([10347C > T]) and CYP2S1*3 (13106C > T; 13255A > G]), occurred in our study population at frequencies of 0.50 and 3.75%, respectively. The most common of the variant alleles was CYP2S1*1H (23.8%), harbouring a 13255A > G substitution located in the 3'UTR.