GC-MS, LC-MS/MS, Docking and Molecular Dynamics Approaches to Identify Potential SARS-CoV-2 3-Chymotrypsin-Like Protease Inhibitors from Zingiber officinale Roscoe

This study aims to identify and isolate the secondary metabolites of <i>Zingiber officinale</i> using GC-MS, preparative TLC, and LC-MS/MS methods, to evaluate the inhibitory potency on SARS-CoV-2 3 chymotrypsin-like protease enzyme, as well as to study the molecular interaction and stability by using docking and molecular dynamics simulations. GC-MS analysis suggested for the isolation of terpenoids compounds as major compounds on methanol extract of pseudostems and rhizomes. Isolation and LC-MS/MS analysis identified 5-hydro-7, 8, 2'-trimethoxyflavanone (<b>9</b>), (<i>E</i>)-hexadecyl-ferulate (<b>1</b>), isocyperol (<b>2</b>), <i>N-</i>isobutyl-(2<i>E</i>,4<i>E</i>)-octadecadienamide (<b>3</b>), and nootkatone (<b>4</b>) from the rhizome extract, as well as from the leaves extract with the absence of <b>9</b>. Three known steroid compounds, i.e., spinasterone (<b>7</b>), spinasterol (<b>8</b>), and 24-methylcholesta-7-en-3<i>β</i>-on (<b>6</b>), were further identified from the pseudostem extract. Molecular docking showed that steroids compounds <b>7</b>, <b>8</b>, and <b>6</b> have lower predictive binding energies (MMGBSA) than other metabolites with binding energy of -87.91, -78.11, and -68.80 kcal/mole, respectively. Further characterization on the single isolated compound by NMR showed that <b>6</b> was identified and possessed 75% inhibitory activity on SARS-CoV-2 3CL protease enzyme that was slightly different with the positive control GC376 (77%). MD simulations showed the complex stability with compound <b>6</b> during 100 ns simulation time.