Simultaneous HPLC-UV quantification of donepezil and rivastigmine in human plasma using an SBA-15/graphene oxide nanocomposite based DLLME dSPE approach
Abstract
This study aimed to develop and validate a sensitive, environmentally friendly analytical method for the simultaneous quantification of donepezil and rivastigmine in human plasma to support therapeutic drug monitoring (TDM). The approach combined a novel dispersive solid-phase extraction (dSPE) utilizing a purposely synthesized SBA-15/graphene oxide (GO) nanocomposite characterized by FT-IR, SEM, and TGA with dispersive liquid-liquid microextraction (DLLME), followed by HPLC-UV detection. Critical parameters affecting extraction efficiency, including sorbent amount, desorption/disperser solvent, sample pH, equilibration time, and ionic strength, were systematically optimized, and the method was validated according to ICH M10 and FDA (2018) guidelines. The optimized SBA-15/GO nanocomposite displayed a highly ordered mesoporous structure with an average pore diameter of 66.77 nm, with optimal extraction achieved using 0.001 g of sorbent, acetonitrile as the desorption/disperser solvent, a sample pH of 6.0, a 30-minute equilibration time, and no salt addition. The method exhibited excellent linearity over 0.15–200 µg/mL for donepezil and 0.01–50 µg/mL for rivastigmine (R² = 0.998 for both), with limits of detection of 0.04 and 0.002 µg/mL, respectively. Absolute extraction recoveries reached 83% for donepezil and 79% for rivastigmine, with enrichment factors of 10, while intra- and inter-assay precision (RSD) remained below 7.8% and 8.2%, respectively, and accuracy (RE) ranged from –6.0% to +7.1%. Clinical applicability was assessed by analyzing plasma samples from six medicated patients receiving either donepezil (5 or 10 mg/day) or rivastigmine (1.5 or 3 mg/day), with results confirming dose-proportional concentrations and demonstrating the method's suitability for routine TDM. Moreover, the procedure is rapid, with a total preparation time of less than 45 minutes, consumes minimal organic solvent (under 2 mL per sample), and adheres strictly to green analytical chemistry principles.