A Capillary Electrophoretic Method for the Analysis of Bupivacaine and Its Metabolites

Abstract

The article studies a capillary electrophoretic (CE) method for the analysis of urinary extracts of the local anesthetic, bupivacaine, and its three main metabolites, desbutylbupi vacaine, 3’-hydroxybupivacaine, and 4’-hydroxybupivacaine, in rat urine. After collection of blank urine, the rats were given a 20 mg/kg intramuscular injection of bupivacaine, and urine was collected for 12 h after dosing. CE analyses were performed using the CAPEL®-205 capillary electrophoresis systems. The data was collected using the Elforan® specialized software. The use of methanol to reduce peak tailing was investigated at different concentrations, but 20% and 30% v/v were proved to be the most optimal at the preparatory stage of the experiment. The resolution of 3’-hydroxybupivacaine and 4’-hydroxybupivacaine was 1.09, 0.98, 0.89 and 0.89 at 15, 40, 70 and 110 s, respectively. The initial resolution (Rs) of desbutylbupi vacaine was achieved with all studied injection periods as Rs = 1.09, 0.97, 0.96 and 0.96 at 15, 40, 70 and 110 s, respectively. Separation efficiencies for 3’- and 4’-hydroxybupivacaine were312×103, 257×103, 196×103 и 169×103μlat injection times of15, 40, 70 and 110 s, respectively. The results showed that the mass of bupivacaine, desbutylbupi vacaine, and 3’- and 4’-hydroxybupivacaine significantly recovered in the rat urine after the dose was administered. The recoveries as a percent of the dose were 0.04, 0.80, 0.15 and 0.05% for desbutylbupi vacaine, bupivacaine, 3’-hydroxybupivacaine, and 4’-hydroxybupivacaine, respectively. Separation of bupivacaine and its metabolites was achieved in 15 min. A particular advantage of this approach over published HPLC methods is that separation of the two hydroxy positional isomers of bupivacaine is possible. A number of unknown peaks were also observed in the electropherograms from the rats dosed with bupivacaine. These did not correspond to any peaks appearing in the blank urine samples. Characterization of these unknown peaks may prove useful for the further understanding of bupivacaine metabolism.
Keywords: Bupivacaine, Capillary electrophoresis, Desbutylbupivacaine, Hydroxybupivacaine, metabolites.