Quantitative analysis of retinoids in biological fluids by high-performance liquid chromatography using column switching. II. Simultaneous determination of etretinate, acitretin and 13-cis-acitretin in plasma.
An automated gradient high-performance liquid chromatographic method for the determination of etretinate, acitretin and 13-cis-acitretin in plasma was developed, using a column-switching technique. After protein precipitation with ethanol, 0.5 ml of the supernatant was injected onto a precolumn (17 mm x 4.6 mm I.D.), filled with 37-53 microns C18 Corasil. Polar plasma components were washed out using 1% ammonium acetate and 1% acetic acid-acetonitrile (8:2, v/v); the retained retinoids were then transferred to the analytical column (125 mm x 4 mm I.D., filled with 5-microns ODS material) in the backflush mode, separated by gradient elution and detected at 360 nm by UV detection. The limit of quantification was 2 ng/ml and the inter-assay precision in the concentration range 20-1000 ng/ml was between 0.9 and 4.0% for all three compounds. To optimize the recovery for etretinate (greater than 60%), protein was precipitated from plasma with ethanol before injection, instead of direct injection of plasma samples, and a mobile phase containing 20% acetonitrile, instead of pure water or buffer, was used.
Wyss
R
R
Preclinical Development, F. Hoffmann-La Roche & Co., Ltd., Basle, Switzerland.
Bucheli
F
F
D017255:Acitretin; D000818:Animals; D002851:Chromatography, High Pressure Liquid; D005050:Etretinate / Q000097:blood*; D006801:Humans; D007202:Indicators and Reagents; D051379:Mice; D051381:Rats; D012176:Retinoids / Q000097:blood*; D012996:Solutions; D013696:Temperature; D014212:Tretinoin / Q000031:analogs & derivatives* / Q000097:blood
D007202:Indicators and Reagents; D012176:Retinoids; D012996:Solutions; D014212:Tretinoin; D005050:Etretinate; D017255:Acitretin