An HPLC–ESI-MS method for simultaneous determination of fourteen metabolites of promethazine and caffeine and its application to pharmacokinetic study of the combination therapy against motion sickness

The combination therapy, promethazine and caffeine had been proven effective in treating motion sickness and counteracting some possible side effects of using promethazine alone while the mechanism and interaction remained unclear. Therefore, an HPLC–ESI-MS method for simultaneous determination of both drugs, and their metabolites was developed for purpose of pharmacokinetic study. To determine as many metabolites as possible, the influence of parameters such as column, flow rate and pH value of mobile phase, ionization polarity and fragmentation voltage were optimized. Fourteen target analytes were well separated and all of them could be identified and determined in plasma after administration of promethazine and caffeine. The LODs and LOQs were 0.9–6.0 and 2.50–16.0 ng/ml, respectively; the recoveries of three levels of quality control samples were from 86.7% to 102%; the intra-day and inter-day precisions were less than 3% and 9%, separately; and the RSDs of compound stability were all lower than 10% within 24 h after sample preparation. As a pharmacokinetic study of the combination therapy in 30 healthy volunteers, concentration–time curves of the drugs and metabolites were studied. The present method for simultaneous measurement of more than ten metabolites is valuable for the study of mechanism and interaction of the combination therapy.