Backgrounds <p>Excess folic acid (FA) supplementation intake among women in child-bearing age, which may lead to an increase in the amount of unmetabolized FA in the bloodstream. Whereas, research on the relationship between high-dose FA and gestational diabetes mellitus (GDM) was lacking, and the potential mechanisms were still unclear.</p> Methods and analysis <p>This is an ongoing cohort which is part of the Qingdao Women and Children’s Hospital Health Cohort, China. In total, 4000 pregnant women were recruited in gestational weeks 9–13⁺⁶, and were followed across pregnancy (16 ± 1, 20 ± 1, 24–28 weeks of gestation). Participants would undergo questionnaires investigation, physical measurement, red blood cell folate, serum folate, serum vitamin B12 and methylmalonic acid detection in early pregnancy. Serum liver X receptors α (LXR-α) and sterol regulatory element-binding protein 1 c (SREBP-1c) levels were tested from Down’s syndrome screening at 16 weeks of gestation on the basis of a nested case-control study. GDM was based on 75-g oral glucose tolerance test at 24–28 weeks’ gestation. The levels of red blood cell folate, serum folate, LXR-α/SREBP-1c and visceral fat (VF) depth between pregnant women with GDM and without GDM will be analyzed.</p> Discussion <p>Our findings could help shed light on the role of high-dose FA regulating LXR- α / SREBP-1c to induce VF accumulation in the pathogenesis of GDM. We anticipated that inform clinical recommendations based on VF depth with precise FA supplementation intake.</p> Trial registration <p>The trial is prospectively recorded at the CHiCTR registry (Trial ID: ChiCTR2300070781). Date recorded: 23/04/2023.</p>