LC determination of cephalosporins in in vitro rat intestinal sac absorption model

Cefotaxime sodium (CX) and Ceftazidime pentahydrate (CZ) are peptidomimetic cephalosporins (CPS) which exist as zwitterionic compounds at physiological pH and because of this reason they are not absorbed appreciably on peroral administration. The permeability of these compounds can be increased transiently by altering membrane characteristics of absorptive epithelium by use of sorption promoters (SPs). In present work a simple validated HPLC method utilizing isocratic mobile phase and having short retention times for CX and CZ is developed which can be used to monitor their concentrations in Kreb's Ringer Bicarbonate (KRB) solution in in vitro intestinal sac absorption model. The same was utilized to determine apparent permeability coefficients and absorption profiles of CPS by modified Wilson–Wiseman method. The CPS were analysed by the reverse phase HPLC method using Shim-pack C18 column. The mobile phase used was of isocratic composition with phosphate buffer (pH 7.0, 3.5 g/l of KH2PO4 dissolved in 0.03 M Na2HPO4 · 2H2O) and methanol in proportion 85:15 for CZ and 70:30 for CX. The flow rate was 1ml/min and quantitative determinations were carried out at 254 nm at 25 °C. The method was found specific because none of the proposed SPs, components of KRB and intestinal sac artefacts interfered with the drug peaks. The drug concentration versus area under peak relationship was found to be linear in concentration range of 0.25–20.0 μg/ml. The recovery studies, intraday variation, interday variation and interanalyst variation were within statistical limits. The limit of detection (LOD) was 95.0 and 100.0 ng/ml for CZ and CX, respectively. The limit of Quantitation (LOQ) was 240.0 and 250.0 ng/ml for CZ and CX, respectively. The proposed method was found to be rapid and selective and hence applied for continuous monitoring of CPS in in vitro intestinal sac absorption studies.