Development of an LC–MS/MS method for simultaneous determination of memantine and donepezil in rat plasma and its application to pharmacokinetic study

• First report on simultaneous quantification of memantine and donepezil in plasma.
• Sensitive method (0.2 ng/mL), low sample volume (50 μL) and short run time (3 min).
• Application to pharmacokinetics and drug interaction studies in rats.

Recently, a fixed dose combination (FDC) of memantine (MM) and donepezil (DPZ) has been approved for the treatment of Alzheimer’s disease (AD). In the present work, a liquid chromatography–tandem mass spectrometry (LC–MS/MS) method for the simultaneous determination of MM and DPZ was developed and validated in rat plasma over the linearity range of 0.2–400 ng/mL using amantadine (AM) as an internal standard. Both the analytes and IS were extracted using one step liquid–liquid extraction procedure. The analytes were separated on C18 reversed phase column with mobile phase consisting of a mixture of methanol and 10 mM ammonium acetate, pH 5 (92:8 v/v) at a flow rate of 0.7 mL/min. The detection of the analytes was done on triple quadrupole mass spectrometer operated in positive electrospray ionization mode (ESI) and quantified using multiple reaction monitoring (MRM). The method was fully validated in terms of linearity, accuracy, precision, recovery, matrix effect, dilution integrity, carry-over effect and stability. The within- and between-run precisions were <10% and accuracy was all within ±10%. The mean recovery of MM and DPZ was found to be greater than 80%. The % RSD value at higher as well as lower concentration was well within the acceptable range (±15%) in all the stability experiments. The method was successfully applied to the oral pharmacokinetics and drug–drug interaction study of MM and DPZ in male Sprague Dawley (SD) rats.