ABSTRACT: Placental CYP27B1 may contribute to circulating maternal calcitriol concentrations across gestation, but determinants of CYP27B1 and CYP24A1 expression in term human placental tissue are not well established.We hypothesized that higher CYP27B1 protein expression would be associated with increased maternal calcitriol during gestation and that CYP27B1 expression would be impacted by substrate availability.This was a prospective, longitudinal study.The study was completed in an urban, prenatal clinic located in Rochester, New York.The study was undertaken in a cohort of 70 pregnant adolescents (?18 y of age) and their term neonates.There was no intervention.Protein and mRNA expressions of CYP27B1, CYP24A1, and vitamin D receptor were measured in term placental tissue and related to 25-hydroxyvitamin D [25(OH)D], 1,25-dihydroxyvitamin D, PTH, serum total calcium, IL-6, leptin, and osteoprotegerin measured in maternal serum at midgestation and delivery and in umbilical cord serum at birth.Placental CYP27B1 protein expression was significantly positively associated with maternal 25(OH)D at both midgestation (n = 68, P = .009) and delivery (n=67, P = .006). Maternal serum 1,25-dihydroxyvitamin D concentrations at midgestation were positively correlated with term placental CYP27B1 mRNA expression (n = 49, P = .002). Significant positive associations were evident between placental CYP27B1 and CYP24A1 protein expression (P = .001, n = 70). Maternal PTH concentrations at midgestation or delivery did not significantly impact placental protein or transcript level of either enzyme. Variability in placental CYP27B1 protein expression was best captured by a model that included maternal midgestation 25(OH)D concentration, placental vitamin D receptor protein expression, and maternal midgestation IL-6 concentrations (P = .002, n = 60, R(2) = 0.22).Higher maternal 25(OH)D during pregnancy was associated with significantly higher placental protein expression of CYP27B1 at term supportive of a link between substrate availability and placental production of calcitriol.