Column packed with heterosurface adsorbent for direct chromatographic analysis of pharmaceuticals in biological fluids

The application of a short column 20×4,0 mm ID packed with restricted access media (RAM) adsorbent for direct determination of theophylline and caffeine in blood plasma is demonstrated. The feature of RAM-adsorbent is connected with absence of the retention of bulky proteins from human blood, while low molecular mass pharmaceutical are retained according to their hydrophobicity. The use of such column allows the determination of caffeine and theophylline in human plasma after filtration without the unwanted time and labour consuming sample preparation procedures, such as precipitation of proteins. RAM-sorbent can be used in eluent pH range between 4,5–7,5, and the range of acceptable concentration of organic modifier in eluent is limited only by the required eluent strength. Special precautions should be undertaken to prevent biodegradation of RAM-adsorbent during prolonged use of the column with eluent containing a small (less than 10 %) concentration of acetonitrile or methanol. Total sample loading of blood plasma up to 50 mg in terms of dry substance can be applied to the column, after which some column regeneration is required. The retention of caffeine and theophylline have not changes after regeneration: the retention factor for theophylline k = 4,0 ± 0,1 and caffeine k = 8,5 ± 0,4 with 0,02 М Na₂HPO₄ (pH = 6,0) – acetonitrile (90:10) as eluent.