Objective To explore the effects of serum 25-hydroxyl vitamin D3 on insulin resistance and bone metabolism in late pregnancy. Methods A total of 84 women in late pregnancy who received prenatal examination in our hospital from February 2012 to January 2013 were selected as research subjects. They were assigned to diabetes group in late pregnancy and healthy group in late pregnancy according to their physical conditions. TG, LDL-c, HDL-c and FBG levels in the two groups of patients were examined via automatic biochemical analyzer. FINS, serum 25-hy-droxyl vitamin D3, PTH and BALP levels were tested via automated luminescent immunoassay. Correlation analysis was also carried out between serum 25-hydroxyl vitamin and HOMA-IR, PTH and BALP. Results TG, LDL-c and HDL-c were compared between the two groups, and the differences were not statistically significant (P>0.05). FBG and FINS in the diabetes group were (5.83±0.68) mmol/L and (24.93±2.07) U/L respectively; FBG and FINS in the healthy group were (4.96±0.57) mmol/L and (17.67±1.83) U/L respectively. Based on the results above, FBG and FINS in the diabetes group were significantly higher than those in the healthy group, and the differences were statistically significant (P<0.05). 25-hydroxyl vitamin D3, HOMA-IR, PTH and BALP were (24.93±3.23) nmol/L, (0.80±0.08), (183.69±12.34) ng/Land (30.04±10.23) μg/L in diabetes group respectively, while 25-hydroxyl vitamin D3, HOMA-IR, PTH and BALP were (39.74±5.86) nmol/L, (0.43±0.04), (165.24±13.07) ng/L and (24.75±9.68) μg/L in the healthy group respectively. 25-hydroxyl vitamin D3, HOMA-IR, PTH and BALP between the two groups were compared, and the differences were statistically significant (P<0.05). 25-hydroxyl vitamin D3 was negatively correlated with HOMA-IR, PTH and BALP (r=-0.479, P<0.01; r=-0.283, P<0.05; r=-0.354, P<0.05). Conclusion Insufficient serum 25-hydroxyl vitamin D3 in late pregnancy is negatively correlated with insulin resistance and has negative effects on bone metabolism.