A Simple, Sensitive, and Greener HPLC-DAD Method for the Simultaneous Analysis of Two Novel Orexin Receptor...

The orexin receptor antagonist (ORA) is one of the new psychopharmacological agents used in the treatment of insomnia. There are currently no documented greener high-performance liquid chromatography-diode array detector (HPLC-DAD) methods for the analysis of ORA antagonists, lemborexant (LMB) and suvorexant (SUV) simultaneously. Therefore, in this study, a simple, sensitive, and greener HPLC-DAD method has been developed for the simultaneous quantitative analysis of LMB and SUV in bulk and laboratory-prepared mixture. The developed method was validated for numerous validation parameters and evaluated for greenness. The C₁₈ Waters Spherisorb CN (4.6 × 250 mm²; 5 μm) column was used for the chromatographic separation. The mobile phase composition was ethanol: 10 mM KH₂PO₄ buffer in a ratio of (60:40 v/v). The DAD detection was performed at 253 nm using a Waters DAD detector. The greenness was evaluated using the analytical Eco-Scale (AES), ChlorTox, and analytical GREEnness (AGREE) techniques. The calibration curves showed excellent linearity for LMB and SUV between the concentration range of 125-5000 ng/mL and 250-10,000 ng/mL, respectively. In addition, the proposed HPLC-DAD method was accurate, precise, robust, highly sensitive, and greener. AES, ChlorTox, and AGREE scales were predicted by the HPLC-DAD method to be 91, 1.14 g, and 0.79, respectively, showing an excellent greenness profile. The greener HPLC-DAD method was successfully used to analyze both medicines quantitatively in bulk and laboratory-prepared synthetic mixtures. The findings of this study indicated that the proposed HPLC-DAD method may be consistently applied to evaluate LMB and SUV in bulk and dosage forms.