Background: Jaundice is a common clinical sign in neonatal medicine. Considering the different roles of vitamin D, its lower level may be correlated with neonatal jaundice. Objectives: This study aimed to demonstrate the relationship between serum vitamin D level and neonatal hyperbilirubinemia and to measure its level in their mothers. Subjects and methods: This case-control study was conducted on 90 neonates and their mothers, 60 neonates with hyperbilirubinemia in patients group and 30 healthy neonates age and sex matched in control group. Blood samples from neonates and their mothers were obtained and sent for laboratory estimation of 25-hydroxy vitamin D, calcium, magnesium, phosphorus, alkaline phosphatase and parathyroid hormone. Results: There is highly significant decrease of vitamin D levels among patients than controls, significant negative correlation between vitamin D levels and serum bilirubin in neonates and no significant difference regarding different laboratory parameters among their mothers. Conclusion: Decreased levels of vitamin D were significantly correlated with neonatal indirect hyperbilirubinemia so, low vitamin D can be included among risk factors for neonatal jaundice.