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Background The deficiency of vitamin D3 (VD) is a universal health issue, its role in different kind of diseases is being studied recently. However, its role in thyroid diseases is not well established yet. This study aims to determine the impact of VD in the pathogenesis of hypothyroidism and hyperthyroidism. Method. Three hundred Iraqi females with age ranged between 30 and 55 years participate in this research, 100 of them were hypothyroid patients, 100 females were hyperthyroid patients and the other 100 females were healthy volunteers served as controls. Thyroid hormones, VD, liver function parameters, and kidney function parameters were determined using different analysis techniques. Results. The levels of VD were significantly decreased in both hypothyroid and hyperthyroid patients (19.644 ± 10.524 for hypothyroid patients and 22.712 ± 11.249 for hyperthyroid patients vs. 30.880 ± 2.587 for controls, p <0.0001). Liver function parameters were within the normal ranges in all the patients. Creatinine and uric acid were within the normal ranges, while urea was significantly increased and out of the normal clinical range in both hypothyroid and hyperthyroid patients (39.560 ± 9.912 for hypothyroidism patients and 42.460 ±7.171 for hyperthyroid patients vs. 26.920 ± 5.033 for controls, p <0.0001). Conclusion. Vitamin D3 and kidney function tests must be included in the differential detection of thyroid diseases. Still, further investigations are needed to understand the underlying mechanism by which VD affects thyroid hormone regulation.