Abstract Objectives The purpose of this retrospective study was to investigate the relationship between 25 OH vitamin D (25OHD) deficiency and iron-deficient anaemia (IDA) in the pediatric population. This was aimed to provide a better insight to IDA follow-up and treatment. Methods The data of 120 patients diagnosed with IDA and 125 healthy pediatric patients were analyzed retrospectively. Serum vitamin B 12 , Folate and 25(OH)D levels, between IDA and healthy groups were evaluated. The relationship between vitamins levels and IDA parameters were examined. Logistic regression analysis was used to assess whether 25(OH)D deficiency levels were an independent risk factor for diagnosing IDA. Results In the comparison of vitamins levels between groups, only mean serum 25(OH)D levels were found to be statistically significantly (p=0.000) lower (13.00 ± 2.50 ng/mL) in the group with IDA compared to the healthy group (25.98 ± 3.66 ng/mL). There were strong positive correlations between 25(OH)D deficiency levels and IDA. The deficiency of 25(OH)D levels was not found to be an independent risk factor for IDA (ORs: 0.958, 95%CI: 0.917–1.000). Conclusions Although current results confirm the association between 25(0H)D deficiency and IDA in pediatric patients, they indicate that there was no independent risk factor for IDA.