This case-cohort study examined the association between plasma enterolactone concentration and incidence of colon and rectal cancer in the Diet, Cancer and Health cohort, which enrolled 57,053 participants aged 50-64. Information about diet and lifestyle was obtained by questionnaire, and data on prescriptions of antibiotics were obtained from the Danish Prescription Registry. Cases diagnosed during 5.9 years of follow-up and a randomly selected sample of the cohort had a plasma sample analyzed for enterolactone by time-resolved fluoro-immuno assay. Associations were analyzed by Cox proportional hazards model. A total of 244 colon cancer cases, 137 rectal cancer cases, and 370 sub-cohort members were included in the statistical analyses. For each doubling in enterolactone concentration, we found lower risk of colon cancer among women [IRR (95% CI) = 0.76 (0.60-0.96)] and a tendency toward lower risk of rectal cancer [IRR (95% CI) = 0.83 (0.60-1.14)]. Among men, a doubling in enterolactone tended to be associated with higher risk of colon cancer [IRR (95% CI) = 1.09 (0.89-1.34)] and was associated with statistically significantly higher risk of rectal cancer [IRR (95% CI) = 1.74 (1.25-2.44)]. Exclusion of antibiotics users strengthened the results slightly. In conclusion, with higher enterolactone levels, we found lower risk of colon cancer among women and higher risk of rectal cancer among men.