ABSTRACT: OBJECTIVE:To understand the contribution of regional differentials in dietary exposures to regional gradients in health, we examined 20-year trends in the association of US census region of residence with nutritional biomarkers and dietary intakes of American adults. DESIGN:Observational. SETTING:The biomarker and 24 h dietary recall data were from the National Health and Nutrition Examination Surveys (NHANES) conducted during 1988-1994 and 1999-2010. The US census region was operationalized as Northeast, Midwest, South and West. Nutritional biomarker outcomes were serum folate, vitamins B6, B12, C, D and E, and carotenoids; dietary outcomes were intakes of nutrients, food groups and eating patterns. SUBJECTS:US adults, n>8000-40 000 for biomarkers and >43 000 for dietary outcomes. RESULTS:The interactions of survey time period and region were not significant for the examined biomarker and dietary outcomes, indicating similar secular trends among regions. The main effect of region was significant for all nutritional biomarkers except serum vitamin B6, most dietary micronutrients, food groups and eating patterns (P<0·001). The mean serum folate, vitamins B12, C and E, and all carotenoid (except lycopene) biomarker levels, and intakes of dietary fibre, vitamins A, E, C and B6, folate, K, Ca, Mg and Fe, fruits, vegetables and whole grains, were higher in the West and Northeast regions, relative to the South and Midwest regions. CONCLUSIONS:Overall, the regional gradients in dietary exposure, expressed objectively as biomarkers or as self-reported nutrient and food group intakes, paralleled trajectories reported for health outcomes and were remarkably persistent over time.