CAPILLARY GAS-CHROMATOGRAPHY OF HEXADECYLPHOSPHOCHOLINE IN CACO-2T CELLS AND CELL-CULTURE MEDIA

The gas-chromatographic determination of hexadecylphosphocholine (HePC ), an experimental antitumor agent of the alkyllysophospholipid group, in Caco-2T cell culture and cell culture media is described. The Caco -2T cells were treated with HePC at a concentration of 40 mu g/ml (9.8 mu M) and the uptake of the drug into the cells (calculated per milli gram protein) was measured after 48 h culture (37 degrees C, 10% CO2). Also, a reversibility test for another 48 h was carried out in which the retention of the drug was measured. The toxicity of HePC on Caco-2 T cells in viability assays was determined. Before the capillary gas-c hromatographic determination, sample cleanup was performed by solid-ph ase extraction (SPE) on a weak cation-exchange column of the CBA (carb oxylic acid) type. For quantitation, racemic -O-octadecyl-2-O-methylgl ycero-3-phosphorylcholine (ET-18-OMe) was added as internal standard, followed by derivatization with trimethylsilylbromide. The results sho wed that HePC taken up by the cells during 48 h of treatment was still detectable 48 h after removal of the drug from the medium. (C) 1995 A cademic Press,Inc.