Background <p>This study aims to investigate the association of pre-conception vitamin D levels on adverse pregnancy outcomes in women undergoing in vitro fertilization with fresh embryo transfer.</p> Methods <p>This was a retrospective cohort study using archived serum 25-hydroxyvitamin D measured in the pre-conception period before ovarian stimulation in patients undergoing in vitro fertilization with fresh autologous embryo transfer. A total of 306 women were included and adverse pregnancy outcomes in their resulting pregnancy were recorded. Patients who were vitamin D deficient (&lt; 20ng/ml) were compared with those who were non-deficient (≥20ng/ml) and analysed for any association with adverse pregnancy outcomes.</p> Results <p>A total of 16/306 (5.3%) patients had hypertensive disorders of pregnancy (gestational hypertension and/or pre-eclampsia). The adjusted odds ratio for hypertensive disorders of pregnancy using vitamin D deficiency as a reference was 0.190 (95% CI 0.042–0.852) (<i>p</i> = 0.030). Other pregnancy complications were not significantly different with regards to pre-conception vitamin D status.</p> Conclusions <p>Pre-conception vitamin D deficiency is associated with an increased risk of hypertensive disorders in pregnancy in women undergoing in vitro fertilization with fresh embryo transfer.</p> Trial registration <p>HKUCTR 2361 (9th March 2018).</p>