Previous research has reported reduced serum 25-hydroxyvitamin D levels in patients with chronic kidney disease (CKD), although the relationship between vitamin D status and insulin resistance (IR) in patients with CKD has not been examined in the general population. We examined the association that kidney function, based on glomerular filtration rate (eGFR) estimated from serum creatinine, has with serum levels of 25-hydroxyvitamin D and components of the metabolic syndrome among 14 679 participants in the Third National Health and Nutrition Examination Survey (NHANES III). In this analysis, adjusted mean serum 25-hydroxyvitamin D was significantly lower only in the participants with a severe (15-29 ml/min/1.73 m(2)) decrease in eGFR compared to those with normal kidney function (61.6 vs 73.3 nmol/l, P=0.0063). Serum 25-hydroxyvitamnin D (P=0.0018) and level of kidney function (P=0.0003) were inversely associated, independent of each other, with homeostasis model assessment of insulin resistance (HOMA-IR), adjusting for confounders. Participants with high 25-hydroxyvitamin D levels (>81 nmol/l) had lower HOMA-IR. We conclude that 25-hydroxyvitamin D deficiency is not as prevalent in the US general population with decreased eGFR as previously reported in patients with CKD; and that vitamin D and kidney function have independent inverse associations with IR.