The anti-COVID-19 drug Favipiravir: Degradation, Method development, Validation, NMR/LC–MS characterization, and In-vitro safety evaluation

Abstract

It is critical to characterize the degradation products of therapeutic drugs to determine their safety as these degradation products may possess fatal effects on the human physiological system. Favipiravir (FVP), a novel anti-Covid-19 drug, that is recently used all over the world with a great impact on humanity was our target to explore more about its toxicity, the margins of its safety, and its degradants in different degradation conditions. The goal of this study is to identify, characterize, and confirm the structures of FVP oxidative and alkaline breakdown products, as well as to assess their safety utilizing in-vitro SRB cytotoxicity assay on normal human skin fibroblasts (NHSF) cell lines. After oxidative and alkaline degradation of FVP, one degradation product was produced in each condition which was isolated from FVP using flash chromatography, characterized by 1HNMR and LC–MS/MS techniques. A reversed-phase Thermo Fischer Hypersil C18 column (4.6 × 150 mm, 5 m) was used to achieve HPLC chromatographic separation. Acetonitrile-5 mM potassium dihydrogen phosphate (pH 2.5) (50:50, v/v) was employed as the mobile phase, with a flow rate of 1 mL/min. At 332 nm, the column effluent was measured. Over the concentration range of 0.5–100 µg/mL, the calibration curve was linear. The intra-day and inter-day relative standard deviations were less than 2%, and good percentage recoveries were obtained that fulfilled the acceptance criteria of the International Conference on Harmonization (ICH) recommendations. The Plackett–Burman design was used to assess the robustness. Each degradant was isolated single using Flash chromatography and methylene chloride: methanol gradient mobile phase. The chemical structures of the degradation products have been confirmed and compared to the intact FVP using 1H-NMR, and Mass spectroscopy. A postulated mechanism of the degradation process has been depicted and the degradants fragmentation pattern has been portrayed. In addition, the in vitro SRB cytotoxicity assay to evaluate the safety profile of FVP and the degradation end products showed their high safety margin in both conditions with IC50 ˃100 µg/ml with no signs of toxicity upon examination of the treated NHSF cells under the optical microscope

A tris(2-aminoethyl)amine-based zinc complex as a highly water-soluble drug carrier for the anti-COVID-19 drug favipiravir: a joint experimental and theoretical study

“…The NMR spectrum of FAV and its Zn( ii ) complexes both were recorded in a DMSO solution to make the most reliable comparison. The 1 H NMR spectrum image of FAV has been already reported by other researchers, 49–51 and it is the same as that observed in the present work. However, herein we assign the observed doublet at 8.459 ppm to the single hydrogen atom of the favipiravir ring.…”

Fluorimetric Determination of Antiviral (COVID-19) Drug Favipiravir in Bulk and Pharmaceutical Dosage Forms

“…The other methods available for the analysis and quantification of Favipiravir include HPLC using UV detection 7 , HPLC using DAD detection 9 ,HPLC using bioanalytical samples (drug spiked in human plasma) using different extraction methods [10][11][12] . Addition to these, analysis of favipiravir was done by developing method and validating it using FTIR 13 , NMR/LC-MS and invitro safety evaluation 14 . Comparative studies of method developed for analyzing Favipiravir in Pharmaceutical preparations using HPLC and UV were also found 15 .…”

Fluorine-a small magic bullet atom in the drug development: perspective to FDA approved and COVID-19 recommended drugs

“…Recently, fluorinated compounds like nirmatrelvir (PF-07321332) 247 etc. (Brewitz et al 2023;Higashi-Kuwata et al 2023;Abdallah et al 2022;Fang et al 2022).…”