Simple quantification of endogenous retinoids in bovine serum by high-performance liquid chromatography – diode-array detection

Autor: Van Merris, K De Wasch, Christian Burvenich, Evelyne Meyer
Rok vydání: 2002
Předmět:
Zdroj: Analytica Chimica Acta. 468:237-244
ISSN: 0003-2670
DOI: 10.1016/s0003-2670(02)00601-3
Popis: A high-performance liquid chromatographic method with diode-array detection (HPLC–DAD) was developed for the simultaneous quantification of endogenous retinoids in bovine serum. The described technique was initially based on an extraction procedure and reversed phase HPLC method designed for human serum, optimised and validated in the bovine. Following denaturation of 2 ml samples, a one-step liquid–liquid extraction was performed. Analytes were separated on a symmetry C 18 column in a total run time of 60 min. The retinoids were monitored at an optimal wavelength of 350 nm as chosen from the retinoid spectra. Retinal was added as internal standard because of its intermediate polarity. Peak-height and peak-area ratios were linear over a concentration range from 1 to 10 ng/ml for 13- cis -, 9- cis -, and all- trans -retinoic acid, 100 to 600 ng/ml for retinol and 10 to 100 ng/ml for retinyl palmitate. Coefficients of variation were acceptable for both within-day and day-to-day assays. To allow the simultaneous analysis of the endogenous retinoids with a wide polarity range, recoveries were compromised. Nevertheless they systematically exceeded 60%. The proposed method is simple, specific and enables fast and accurate routine quantification of acid, alcoholic and esteric forms of retinoids. It was successfully applied to analyse 21 bovine serum samples from a clinical study. Simultaneous assay of endogenous retinoids has not yet been performed in cows.
Databáze: OpenAIRE
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