Squamous cell carcinoma of the oesophagus is one of the most common cancers among black males in South Africa. Genetic polymorphism in the cytochrome P450 2E1 (CYP2E1) gene, coding for one of the main enzymes involved in the bioactivation of tobacco- and alcohol-related substances, was investigated for its role in the development of oesophageal cancer. Three single nucleotide polymorphisms -1053C--> T, -1293G--> A (both give rise to CYP2E1*5 ) and 7632T--> A ( CYP2E1*6 ) in the 5'-untranslated region of CYP2E1 were investigated in 189 patients and 198 control individuals in South Africa. The mutant variants occurred between patients and controls at frequencies of 1% and 2% (-1053C--> T), 1% and 3% (-1293G--> A) and 18% and 7% (7632T--> A), respectively. In comparing patients with controls, the heterozygous CYP2E1*6 genotype was associated with increased risk of the development of squamous cell carcinoma of the oesophagus (odds ratio, 5.90; p < 0.001) after adjusting for age, sex, smoking and alcohol consumption. In contrast, we did not find a significant association between CYP2E1*5 and oesophageal cancer. Six novel mutations, -1371G--> A, -1359C--> G, -1342C--> G, -1261T--> del, -1189T--> C and -1103C--> G, were identified by DNA sequence analysis of the CYP2E1 promoter region. In summary, our case-control study of oesophageal cancer revealed an elevated risk associated with the CYP2E1*6 allele in South Africans.