Untargeted Metabolomics and in Silico Screening of Sisymbrium officinale: Identification of Flavonoid Glycosides as Potential Anti-inflammatory Agents.

Sisymbrium officinale (SO) is a traditional Mediterranean medicinal plant used to treat inflammatory diseases; however, its anti-inflammatory efficacy has not been thoroughly investigated by using metabolomics and in silico approaches. This study aimed to characterize the bioactive constituents of SO and identify potential anti-inflammatory agents. SO was extracted using various solvents: water, methanol, ethanol, acetone, and chloroform. Nuclear magnetic resonance (NMR) spectroscopy provided a rough quantification of various functional groups in SO. Principal component analysis (PCA) of mass spectrometry (MS)-annotated metabolites showed that methanol, ethanol, and acetone extracts formed distinct clusters, whereas water and chloroform were separate. Bioassays─including nitric oxide (NO) inhibition, 2,2'-azino-bis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS), 2,2-diphenyl-1-picrylhydrazyl (DPPH), total phenolic content (TPC), and total flavonoid content (TFC)─confirmed that methanol, ethanol, and acetone extracts exhibited the most potent anti-inflammatory activity compared with those extracted using water and chloroform. Orthogonal partial least-squares (OPLS-DA) discriminant analysis chemometrics (variable importance in projection >1.0, p-value <0.05, and log2(FC) > 1.5) screened 15 SO bioactive candidates belonging to the flavonoid, flavonoid glycoside, cinnamic acid, and phospholipid classes. Further, in silico ADMET analysis and molecular docking of these metabolites with human inducible NO synthase (iNOS, PDB 3E7G), endothelial NOS (eNOS, PDB 6AV7), and neuronal NOS (nNOS, PDB 6CID) receptors identified two flavonoid glycosides─apigetrin and kaempferol 3-O-α-l-arabinopyranoside─as potential anti-inflammatory agents that may act via a competitive inhibition mechanism. These findings provide valuable insight into the therapeutic potential of SO and support further development of its bioactive compounds.