The objective of this study is to examine and evaluate whether serum 25(OH)D is associated with disease activity in patients with rheumatoid arthritis (RA). Our results suggested that serum 25(OH)D in RA groups has significant lower level (35.99 ± 12.59 nmol/L) than that in the normal groups (54.35 ± 8.20 nmol/L, P < 0.05). Based on the DAS28, patients with RA were divided into four subgroups, and no differences were found in the four groups (P > 0.05). The 25(OH)D levels in complete remission, low disease activity, middle disease activity, and high disease activity group were 32.86 ± 12.26, 33.97 ± 13.28, 38.41 ± 10.64, and 38.94 ± 13.35 nmol/L, respectively. Based on the serum 25(OH)D levels, patients with RA were divided into inadequate group and normal group, and there were no significant differences in baseline characteristics and disease activity in the two groups. Our results showed that serum 25(OH)D levels in the inadequate group are significantly lower than those in the normal group. However, no correlations were found between 25(OH)D levels and disease activity among 116 patients with RA. The present findings will help to understand the association between 25(OH)D and disease activity of RA.