Analyte stability during the total testing process: studies of vitamins A, D and E by LC-MS/MS

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There are limited evidence based studies demonstrating the stability of fat-soluble vitamins (FSV) measured in blood. This study aimed to examine the effects of light, temperature and time on vitamins A, D and E throughout the total testing process.


Four experiments were conducted. Three investigated the sample matrix, of whole blood, serum and the extracted sample, against the variables of temperature and light; and the fourth experiment investigated the sample during the extraction process against the variable of light. All samples were analysed via our simultaneous FSV method using liquid chromatography-tandem mass spectrometry technology. The allowable clinical percentage change was calculated based on biological variation and desirable method imprecision for each analyte. The total change limit was ±7.3% for 25-OH-vitamin D3, ±11.8% for retinol and ±10.8% for α-tocopherol.


Vitamins D and E were stable in the investigated conditions (concentration changes <4%) in the pre-analytical and analytical stages. Vitamin A showed photosensitivity in times >48 h with concentration changes of −6.8% (blood) and −6.5% (serum), both are within the allowable clinical percentage change. By contrast, the extracted retinol sample demonstrated a concentration change of −18.4% after 48 h of light exposure. However, vitamin A in the serum and extracted solution was stable for one month when stored at −20°C.


Blood samples for vitamins D and E analyses can be processed in normal laboratory conditions of lighting and temperature. The required conditions for vitamin A analysis are similar when performed within 48 h. For longer-term storage, serum and vitamin A extracts should be stored at −20°C.

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