Simple, rapid and selective HPLC methods have been developed for the analysis of amoxycillin, ampicillin, epicillin, mecillinam and propicillin. A Hypersil ODS 5 microm (150x4.6 mm I.D.) column was used with mobile phases containing aqueous phosphate buffers, pH 3-4.6 and either methanol or acetonitrile as the organic modifier. Samples were detected by their optimal UV absorption (210-230 nm). The lower limits of quantitation of the compounds (100 microl injection volume) were 0.1 microg/ml. The assays were linear in the range of 0.1-100 microg/ml with r2 values greater than 0.99. The methods have been applied successfully for the measurement of the flux of the compounds across Caco-2 cells monolayers.