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Obesity refers to the accumulation of excessive fats in the body which could lead to the development of diseases. Obese people have low vitamin D levels for several reasons including larger volume of distribution, vitamin D tightly bound in fatty tissues, reduced absorption, and diets with low vitamin D. Accurately measuring vitamin D metabolites is challenging. The Ultra-High-Performance Liquid Chromatography-Tandem Mass Spectrometry (UHPLC-MS\/MS) method was developed and validated for the analysis of vitamin D metabolites in the serum. Blood samples were collected from 452 subjects which consisted of baseline (vitamin D deficient obese subjects), follow-up (supplemented obese subjects), and healthy volunteers. The vitamin D metabolites were separated adequately by the developed UHPLC-MS\/MS method. Moreover, the validation criteria for the method were within an acceptable range. The baseline, follow-up and even healthy volunteers were deficient in 25OHD3 and 25OHD2. The baseline and healthy subjects had comparable concentration of vitamin D2 and D3. However, healthy subjects had a higher concentration of 25OHD and its epimer compared to the baseline subjects. The vitamin D3 was increased significantly in the follow- up subjects; therefore, the 25OHD3 was increased significantly compared to the baseline as well; however, the increase was insufficient to achieve the optimal range. The UHPLC-MS\/MS method test was applied successfully on estimation of vitamin D metabolites in subjects. This study indicates the significance of taking into account the metabolic and storage effects when evaluating the vitamin D status in obese subjects.<\/jats:p>","DOI":"10.1038\/s41598-024-58154-z","type":"journal-article","created":{"date-parts":[[2024,3,30]],"date-time":"2024-03-30T13:01:58Z","timestamp":1711803718000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":60,"title":["Unraveling the complex interplay between obesity and vitamin D metabolism"],"prefix":"10.1038","volume":"14","author":[{"given":"Bashar","family":"Alzohily","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Asma","family":"AlMenhali","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Salah","family":"Gariballa","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Nayla","family":"Munawar","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Javed","family":"Yasin","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Iltaf","family":"Shah","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2024,3,30]]},"reference":[{"key":"58154_CR1","doi-asserted-by":"publisher","first-page":"341","DOI":"10.1111\/obr.12239","volume":"16","author":"M Pereira-Santos","year":"2015","unstructured":"Pereira-Santos, M., Costa, P. 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