Fast isocratic reversed-phase high-performance liquid charomatographic (RP-HPLC) separation of four tetracyclines and flumequine was obtained using a Zorbax SB-C-18 column. Baseline resolution was achieved in 11 min. The peaks were narrow, well separated and without any tails although there was no chelating agents added to the mobile phase. Due to the chaotropic effect, the addition of potassium perchlorate allowed controlling the tetracyclines retention while the retention of flumequine was almost constant. (C) 2004 Elsevier B.V. All rights reserved.