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Objectives: To determine seasonal variations in the vitamin D status of Saudi adolescent girls in Riyadh, Kingdom of Saudi Arabia (KSA) and its effect in biochemical and clinical characteristics. Methods: In this prospective study, a total of 2000 Saudi females aged 12-18 years from different schools in Riyadh, KSA participated and submitted a generalized questionnaire with clinical information. Fasting blood samples were obtained in 1618 subjects for the winter season (December to February) and only 499 subjects returned to submit fasting blood samples for the summer season (June-August). Circulating serum 25(OH)D, parathyroid hormone (PTH), and other biomarkers of bone remodeling were measured during both seasons. Results: Vitamin D deficiency (serum 25(OH)D less than 25 nmol/L) was significantly higher during summer than winter (63.5% versus 40.8%; p less than 0.001). Mean serum PTH was also significantly higher during summer than winter ( p less than 0.01). In all subjects, serum PTH showed a significant inverse association with 25(OH)D at levels below 40 nmol/L (r=-0.21; p less than 0.001). The prevalence of subjects having clinical and metabolic manifestations suggestive of osteomalacia was 2.13% (N=33 out of 1548). Conclusion: Seasonal variations in the vitamin D status of Saudi adolescent females significantly modifies biochemical parameters as response to vitamin D status change. In the meantime, heightened public health awareness should be given to populations at higher risk for vitamin D deficiency. Saudi Med J 2016; Vol. 37 (9): 1002-1008 doi: 10.15537/smj.2016.9.15248 How to cite this article: Sulimani RA, Mohammed AG, Alfadda AA, Alshehri SN, Al-Othman AM, Al-Daghri NM, Hanley DA, Khan AA. Vitamin D deficiency and biochemical variations among urban Saudi adolescent girls according to season. Saudi Med J 2016; 37(9): 1002-1008. doi: 10.15537/smj.2016.9.15248.