Background: We aimed to determine the level of serum vitamin D and serum ferritin in four groups’ children and adolescents. Methods: This descriptive/analytical study was conducted in 2019 on children and adolescents with thalassemia aged 7-18 years in Ahvaz, Iran. Overall, 160 patients with the target inclusion and exclusion criteria were assigned into four groups (n=40) of people with thalassemia minor, thalassemia major, iron deficiency anemia and healthy individuals. The level of ferritin and vitamin D was assessed. Mann-Whitney test, Wilcoxon test and Spearman's correlation test were used as well as SPSS Statistics V21. Results: The mean age of the participants was 13.07 ± 3.82 yr and 100 people (62.5%) were male. In terms of vitamin D deficiency, 16 people (40%) had severe deficiency in iron deficiency group. The highest level of ferritin was found in thalassemia major group and the lowest in iron deficiency group. Regarding the level of vitamin D and ferritin in the healthy group, there was a statistically significant difference between mean vitamin D and serum ferritin levels (P=0.027). In iron deficiency group, this level of ferritin was observed with mild deficiency (P=0.017). In thalassemia major group, the increase in ferritin was associated with severe vitamin D deficiency (P=0.05. Conclusion: Severe vitamin D is seen in thalassemia major due to the increase in ferritin, but moderate deficiency of this vitamin in healthy people and mild deficiency in iron deficiency anemia group was observed considering the normal levels of ferritin.