A validated eco-friendly HPLC-FLD for analysis of the first approved antiviral remdesivir with other potential add-on therapies for COVID-19 in human plasma and pharmaceuticals

It is crucial to have a reliable and sensitive method for separating common drugs used in SARS-CoV-2 pneumonia treatment protocols for ongoing treatment and upcoming investigations. This study presents an HPLC-FLD approach to analyze three co-administered medicines - remdesivir (RDV), hydroxychloroquine sulphate (HCQ), and levofloxacin hemihydrate (LVX) - in their pure forms, pharmaceutical preparations, and spiked human plasma. The HPLC-FLD analysis was conducted using a Symmetry® C18 column (100 mm × 4.6 mm ID, 3.5 μm particle size) at 40 °C, with (A) an aqueous mixture of 0.02 M phosphate buffer and 0.2% heptane-1-sulphonic acid sodium solutions (50 : 50) adjusted to pH 3, (B) acetonitrile, and (C) methanol as the mobile phase. The injection volume was 10 μL, and the flow rate was 1.5 mL min −1 . The detection was done using a multi-wavelength excitation and emission fluorescence detector, with individual optimization for each drug. The drug separation time was less than 10 minutes, and the method showed sensitive and wide linearity ranges for all medicines, with r 2 values of more than 0.999. The impact of the mobile phase pH and flow rate on suitability parameters (retention time and number of theoretical plates) was studied. The method was found to be environmentally friendly based on GAPI and AGREE metrics. The validity of the method was evaluated following ICH and FDA guidelines. An HPLC-FLD method was proposed to determine remdesivir, hydroxychloroquine, and levofloxacin in their pure form and in human plasma, with high sensitivity and a wide linear range. Validation was done as per ICH and FDA guidelines, and the method greenness was evaluated.