Abstract Background Despite the popular use of vitamin supplements and several prospective cohort studies investigating their effect on cancer incidence and cardiovascular disease (CVD), scientific data supporting their benefits remain controversial. Inconsistent results may be partly explained by the fact that use of supplements is an inconsistent behavior in individuals. We examined whether vitamin supplement use patterns affect cancer and CVD risk in a population-based cohort study in Japan. Methods A total of 28,903 men and 33,726 women in the Japan Public Health Center-based Prospective Study cohort, who answered questions about vitamin supplement use in the first survey from 1990-1994 and the second survey from 1995-1998, were categorized into four groups (never use, past use, recent use, and consistent use) and followed to the end of 2006 for cancer and 2005 for CVD. Sex-specific hazard ratios (HRs) and 95% confidence intervals (95% CIs) were used to describe the relative risks of cancer and CVD associated with vitamin supplement use. Results During follow-up, 4501 cancer and 1858 CVD cases were identified. Multivariate adjusted analysis revealed no association of any pattern of vitamin supplement use with the risk of cancer and CVD in men. In women, consistent use was associated with lower risk of CVD (HR 0.60, 95% CI 0.41-0.89), whereas past (HR 1.17, 95% CI 1.02-1.33) and recent use (HR 1.24, 95% CI 1.01-1.52) were associated with higher risk of cancer. Conclusions To our knowledge, this is the first prospective cohort study to examine simultaneously the associations between vitamin supplement use patterns and risk of cancer and CVD. This prospective cohort study demonstrated that vitamin supplement use has little effect on the risk of cancer or CVD in men. In women, however, consistent vitamin supplement use might reduce the risk of CVD. Elevated risk of cancer associated with past and recent use of vitamin supplements in women may be partly explained by preexisting diseases or unhealthy background, but we could not totally control for this in our study.