OBJECTIVE To determine if patients with rheumatoid arthritis (RA) are at risk of vitamin D deficiency and whether the levels of vitamin D are correlated with clinical parameters in RA. METHODS A total of 280 treatment-naïve RA patients, and 140 age- and sex-matched healthy volunteers were enrolled. Serum levels of 1,25-dihydroxycholecalciferol (1,25(OH)2D3), the active form of vitamin D, were measured by enzyme-linked immunosorbent assay (ELISA). Concentrations of 1,25(OH)2D3 less than 25 ng/mL were defined as insufficient. Linear regression was performed to evaluate correlations as (modifying and) confounding factors were controlled. RESULTS The levels of serum 1,25(OH)2D3 in RA individuals (12.24 ± 6.68 ng/ml) were significantly lower than in healthy controls (21.08 ± 7.14 ng/ml; p < 0.05). An inverse association was found between the levels of 1,25(OH)2D3 and ESR in obese and overweight individuals with RA (βobese = -0.385, βoverweight = -0.395, both p < 0.05), but not in normal and underweight subjects. A significant negative association between levels of 1,25(OH)2D3 and DAS28 score (β = -0.164, p = 0.018) was observed. The levels of 1,25(OH)2D3 were associated moderately and inversely with the absolute numbers of Th-17 (β = -0.158, p = 0.019) and positively with those of CD4+ regulatory T (Treg) cell (β = 0.146, p = 0.025). The levels of 1,25(OH)2D3 in anti-cyclic citrullinated peptide (anti-CCP)-positive patients with RA were lower than in the anti-CCP-negative RA patients (10.86 ng/ml versus 15.98 ng/ml; t = -3.08, p < 0.01). CONCLUSIONS A significant association was observed between levels of vitamin D and parameters of disease, including body mass index (BMI), DAS28, Th17 cell counts, Treg cell counts, and presence of anti-CCP antibody in RA patients.