Numerous cross-sectional studies demonstrate an inverse association between plasma 25-hydroxyvitamin D [25(OH)D] and blood pressure or hypertension. Prospective data, however, are limited. Among 1484 women aged 32 to 52 years who did not have hypertension at baseline, we prospectively analyzed the association between plasma levels of 25(OH)D and the odds of incident hypertension using a nested case-control study design. We matched cases and controls on age, race, and month of blood collection and further adjusted for body mass index, physical activity, family history of hypertension, oral contraceptive use, and plasma levels of parathyroid hormone, calcium, phosphorous, creatinine, and uric acid. Median plasma 25(OH)D levels were lower in the cases (25.6 ng/mL) than in the controls (27.3 ng/mL; P<0.001). Women in the lowest compared with highest quartile of plasma 25(OH)D had an adjusted odds ratio for incident hypertension of 1.66 (95% CI: 1.11 to 2.48; P for trend=0.01). Compared with women with sufficient levels, those with vitamin D deficiency (<30 ng/mL; 65.7% of the study population) had a multivariable odds ratio of 1.47 (95% CI: 1.10 to 1.97). Plasma 25(OH)D levels are inversely and independently associated with the risk of developing hypertension.