Cigarette smoking has been associated with increased risk of hepatocellular carcinoma (HCC) in some epidemiological studies. Cytochrome P450 1A1 (CYP1A1) is involved in the biotransformation of tobacco-derived polycyclic aromatic hydrocarbons (PAHs) into carcinogenic metabolites. The aim of this study was to determine whether CYP1A1 polymorphisms were related to HCC risk among chronic hepatitis B virus (HBV) carriers. Genotypic variants of CYP1A1 were determined using polymerase chain reaction in 81 incident cases of HCC and 409 controls nested in a cohort study of 4841 male chronic HBV carriers. No overall association between CYP1A1 genotypes and HCC was observed. The presence of the Mspl (odds ratio (OR) 3.15, P = 0.0196) or Ile-Val (OR 1.99, P = 0.0855) variant allele of CYP1A1 increased HCC risk among smokers, but posed no increased risk among non-smokers. The smoking-related HCC risk was most pronounced among those who had a susceptible allele of the CYP1A1 and a deficient genotype of glutathione S-transferase M1, which detoxifies PAH electrophilic metabolites produced by CYP1A1. In the absence of the Ile-Val variant allele, the Mspl polymorphism was still associated with smoking-related HCC. This study suggests that tobacco-derived PAHs play a role in HCC risk among chronic HBV carriers, and CYP1A1 polymorphism is an important modulator of the hepatocarcinogenic effect of PAHs. The Mspl and Ile-Val polymorphisms of CYP1A1 may have different mechanisms for increasing susceptibility to smoking-related HCC.