Background: Vitamin D deficiency is associated with respiratory problems in neonates. The late preterm or near-term neonates who have been admitted for tachypnea and fully recovered before 12 h are called non-specific respiratory distress syndrome (NRDS) cases. The present study aimed to evaluate the effect of 25-hydroxy vitamin D administration in pregnant women at risk of preterm delivery on the incidence of NRDS in their newborns. Methods: This single-blind clinical trial was performed on mothers and neonates with a gestational age of 32–37 weeks who were referred with labor pains from February 20, 2021 to June 29, 2021 to the Obstetrics and Gynecology Department and intensive care unit of Ghaem Hospital, affiliated to Mashhad University of Medical Sciences, Iran. Within 72 h before premature delivery, a dose of 50,000 units of 25 hydroxyvitamin D was injected intramuscularly to pregnant women in the intervention group. A sample containing 1.5 mL of whole blood was collected from the umbilical cord of the infant and mother to assess the level of 25-hydroxy vitamin D. Results: In the present study, there was a significant difference between the two groups of control and intervention in terms of infant’s weight (p = 001), 1-minute (p = 0.027) and 5-minute Apgar scores (p = 0.001), the incidence of NRDS (p = 0.001), and the maternal age (p = 0.004). However, the results showed no statistically significant difference between the two groups in terms of gender (p = 0.673), type of delivery (p = 0.299), level of 25-hydroxy(OH) vitamin D of the mother (p = 0.053), and infant (p = 0.805). Conclusions: A single injection of vitamin D into the mother prone to preterm birth over 31 weeks of gestation reduces transient respiratory problems in their newborns. Clinical Trial Registration: The study was also registered in the Iranian Clinical Trial Registration Center (IRCT20110807007244N7).