Microcolumn high-performance liquid chromatographic assay for doxycycline in isolated alveolar macrophages

A procedure for the isolation of doxycycline from the alveolar macrophages is described. Due to the minimal amount of the sample, it was necessary to employ microcolumn HPLC. A great advantage of the methodology developed in the present paper consists in the direct injection of an aqueous supernatan t over disintegrated macrophages without the use of liquid-liquid or solid- liquid extraction. Another advantage of the method is the fact that the inj ection of an aqueous solution on a microcolumn with a lipophilic stationary phase makes the analyzed substances concentrate, which increases the sensi tivity of assay. Separation was performed on a Separon SGX C-18 microcolumn , 150x1 mm, detection at 254 nm by UV, 1 mu l flow cell. Rabbits were treat ed intravenously with Vibramycin inj, sic. in a dose of 13 mg/kg body weigh t doxycycline. A procedure for washing macrophages from rabbit lungs was de veloped. Dry matter of macrophages ranged from 5.9 to 11.4 mg. In thr sampl es of three rabbits the levels of doxycycline ranged from 110 to 270 ng per 1 mg of macrophages dry matter. (C) 1999 Elsevier Science B.V. All rights reserved.