Determination of cefpodoxime levels in chinchilla middle ear fluid and plasma by high-performance liquid chromatography

Abstract

A high-performance liquid chromatographic method has been developed to determine cefpodoxime levels in chinchilla plasma and middle ear fluid (MEF) to be used in studying otitis media. Cefpodoxime and the internal standard, cefuroxime, were separated on an ODS column (250 × 2.1 mm I.D., 5 μm Hypersil), using a mobile phase of 25 m M acetate buffer (pH 4.3)/15 m M triethylamine—acetonitrile (92.5:7.5, v/v). Following elution of cefpodoxime and the internal standard, at 3.5 and 5.9 min respectively, the acetonitrile concentration was increased to 1:1 (v/v) in a step function to elute endogenous compounds retained on the column. Sample preparation involved protein precipitation with acetonitrile. This fast, efficient protein precipitation procedure together with UV detection allows a quantitation limit of 50 ng/ml with a 50-μl sample size. Recoveries (mean ± S.D., n = 3) at 0.1 μg/ml in MEF were 90.3 ± 2.9% and 88.6 ± 1.2% for cefpodoxime and cefuroxime respectively. Recoveries (mean ± S.D., n = 3) at 0.1 μg/ml in plasma were 72.1 ± 7.3% and 81.1 ± 1.1% for cefpodoxime and cefuroxime respectively. The method was evaluated with biological samples taken from chinchillas with middle ear infections after administering cefpodoxime proxetil.