Measurement of the concentrations of active agents in drugs using UHPLC (Ultra High Pressure Liquid Chromatography) – and HPLC and UV-detection

Abstract

Introduction: We have been able to routinely measure concentrations of psychotropic drugs in human blood by using HPLC/UV in our laboratory since 2006. Currently, we have a diversity of 52 substances that can be separated and detected by several validated methods. An optimization of this sector is the use of UHPLC (Ultra High Pressure Liquid Chromatography) which allows faster separation of more substances. Methods: We use a reversed phase column and phosphate buffer with methanol and as variable part acetonitrile for the measurement. Substances were measured with a UHPLC/UV method. A protein precipitation step was required for pre-purification of human blood. Results: To measure 21 pharmaceutical drugs by HPLC, the laboratory uses actually four different methods for clinical routine. The UHPLC/UV method could measure 20 of these drugs plus 2 additional compounds. The detection limit was lower than the therapeutic range of all the substances. Therapeutic ranges of pharmaceutical drugs such as haloperidol that have very small concentrations in human blood could be measured by this method. The analytical measurement of 22 substances required 23 minutes. Time for measurement and costs for chemicals could be reduced significantly. Conclusion: UHPLC gives us the opportunity to separate und detect many substances under the same chromatographically conditions. A transfer of the routine analytics of HPLC to an UHPLC reduces costs and needs less time. Besides, this method can be used for routine analysis even though substances have only a very low concentration of active agents.