Obesity increases the risk of low 25-hydroxyvitamin D (25(OH)D) concentrations and gestational diabetes (GDM). We explored whether the association between GDM and change in 25(OH)D concentrations measured in the first (7-18 wk) and second (20-27 wk) trimesters of pregnancy is dependent on maternal BMI. The study was a prospective study of 219 women with BMI of ≥30 kg/m2, a history of GDM, or both. The participants were stratified by first-trimester BMI: BMI of <25.0, 25.0-29.9, 30.0-34.9, and ≥35 kg/m2. In the BMI group ≥35 kg/m2, those who did not develop GDM during the follow-up showed higher increase in serum 25(OH)D concentrations compared with women who developed GDM (43.2 vs. 11.5%; P < 0.001). No associations between 25(OH)D concentrations and GDM were observed in other BMI groups. These findings give an important aspect of the role of maternal body size in the association between vitamin D and GDM in high-risk women.