A novel UPLC–ESI-MS/MS method for the quantitation of disulfiram, its role in stabilized plasma and its application

• Disulfiram has therapeutic application in cancer.
• Solid phase extraction was utilized in the separation of disulfiram.
• Determination of disulfiram was firstly conducted by UPLC–ESI-MS/MS method.
• The method possesses a LLOQ of 0.6 ng/mL, much lower than previous methods.
• The method with stabilizing agent added showed better reproducibility.

Disulfiram (DSF) has been used to treat alcoholism for many years and it has been suggested to play a key role in combatting many kinds of tumors. However, disulfiram has complex pharmacokinetics and is rapidly eliminated which limits its use as a tumor treatment. Therefore, a rapid and sensitive analytical method based on ultra performance liquid chromatography coupled to electrospray ionization-tandem mass spectrometry (UPLC–ESI-MS/MS) was developed and validated for the determination of disulfiram in rat plasma. Blood samples were pre-stabilized with a stabilizing agent and then plasma was obtained and subjected to solid phase extraction (SPE), and chromatographed on a Phenomenex Kinetex® XB C18 column with gradient elution using a mobile phase consisting of acetonitrile–water (containing 0.1% formic acid and 1 mM ammonium acetate) at a flow rate of 0.2 mL/min for 3 min. Multiple reactions monitoring in positive mode was carried out with disulfiram at 296.95/115.94 and diphenhydramine (internal standard, IS) at 256.14/167.02 over a linear range from 0.6 to 1200 ng/mL. The extraction recovery of disulfiram for different concentrations ranged from 75.7% to 78.3%. The intra- and inter-day precision was less than 8.93% and 12.39%, respectively, and the accuracy was within ±7.75%. The validated method was successfully applied to a pharmacokinetic study of disulfiram in rat plasma after oral administration of a dose of 180 mg/kg.