Abstract Background: Prematurity and its related challenges are one of the major problems for neonatal medicine. Many studies showed relation between prematurity and vitamin D (Serum 25OHD). Aim of Study: This researchwas conducted to assess the prevalence of Serum 25OHD deficiency among preterm infants at birth and to ascertain if lung disease morbidities are con-nected to serum 25OHD status at birth. Patients and Methods: This investigation was prospective cross-sectional research which was carried out at the Neonatal Intensive care unit (NICU) in Kasr Al-Ainy University Hos-pitals, between September 2021 and March 2022, on 80 preterm neonates with a gestation age <34 weeks assessed by the last menstrual cycle or ultrasound who admitted in NICU within 24 hours of life. Serum 25 OHD was assayed by ELISA. Results: Serum 25OHD wasdeficient (<20ng/mL) in 58 cases (63.8%) with serum 25OHD Level, mean ± SD was 6.54±3.2, it's insufficient (20-30ng/ml) in 7 cases (8.8%) with serum 25OHD Level mean ± SD was 23.03±1.58 and it's sufficient (>30ng/ml) in 15 cases (18.8%) with serum 25OHDLevel, mean ± SD was 35.08±2.05. Conclusion: Low 25(OH)D levels are much more common in preterm infants from Egypt. Lack of serum 25OHD is a distinct risk factor for the development of RDS in preterm newborns.