Vitamin D25-OH and D1-25OH levels were compared with cardiac R2* (1/T2*), left ventricular ejection fraction (LVEF), age, ferritin and liver iron in 24 thalassaemia major patients. Vitamin D25-OH levels were reduced in 13/24 patients while vitamin D1-25OH levels were often elevated. Vitamin D25-OH levels decreased with age (r(2) = 0.48) and with liver iron (r(2) = 0.20). Cardiac R2* was inversely related with the ratio of D25-OH to D1-25OH levels (r(2) = 0.42). LVEF was also proportional to the D25-OH/D1-25OH ratio (r(2) = 0.49). Vitamin D deficiency may be associated with cardiac iron uptake and ventricular dysfunction in thalassaemia major patients.