Determination of retinoic acid in human serum and plasma by high-performance liquid chromatography

All-trans retinoic acid (ATRA) is the main derivative of retinol. It is formed in the intestinal mucosa from retinaldehyde. The aim was to establish a high-performance liquid chromatography method for determination of ATRA in human serum and plasma. Stability of ATRA in serum stored under various conditions was tested as well. ATRA was extracted from serum (plasma) after deproteination with acetonitrile by hexan:ethylacetate mixture (50:50). The extract was then evaporated under nitrogen at room temperature and dissolved in methanol. ATRA was detected after separation on C-18 column at 355 nm. This method uses acitretin as internal standard. Limit of detection was 0.21 mu g/dm(3) and limit of quantification 0.70 mu g/dm(3). The method is linear for ATRA concentration 0.78-25 mu g/dm(3). Based on our results, ATRA has the same concentration in serum and in plasma. Repeatability of the method measured within run is given by variation coefficient 4.31% for serum and 1.86% for plasma. Reproducibility of the method measured between runs is given by variation coefficient 9.55% for serum and 8.00% for plasma. The worst storage conditions were light and room temperature, ATRA decreased by 0.69 mu g/dm(3), i.e., by 39%. Simultaneously, with ATRA, retinol is measured as well. We have introduced a fast, simple, and reliable method for determination of ATRA in human serum and plasma. [GRAPHICS] .