A robust method for quantitation of total vitamin D2 and D4 in mushrooms by high performance liquid chromatography with UV detection (HPLC-UV) was developed to analyze mushrooms exposed to UV light. A two-step solid phase extraction (SPE) (silica, carbon black) removed chromatographic interferences typically resolved only with mass spectrometric detection (LC-MS) and allowed quantitation of all vitamin D and pre-D analytes. The vitamin and pre-vitamin forms of D2, D4 and D3 (internal standard), as well as other photoisomers and sterols were resolved. Results for six types of UV-exposed mushrooms were comparable to LC-MS. Screening of ten additional types of UV-exposed mushrooms without the IS confirmed lack of interference with the IS. The limit of quantification (µg/100 g fresh weight) was 0.4 for vitamin D and 0.9 for pre-vitamin D. Mushrooms do not have to be dried, and separatory funnels and large solvent volumes were also eliminated from sample preparation.
Keywords: 22,23-Dihydroergocalciferol; Ergocalciferol; Food analysis; Mushrooms; SPE; Vitamin D.
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