Antiplasmodial metabolites from Baccharoides anthelmintica seeds: In vitro and in silico profiling.

Ethnopharmacological relevance: Baccharoides anthelmintica (L.) Moench has traditionally been used in ethnomedicine as a general body tonic and for the treatment of malaria and other parasitic infections. Ethnobotanical evidence supporting its antimalarial use has motivated further scientific investigations into its phytochemical composition. However, the specific bioactive constituents responsible for these effects remain poorly characterized. Metabolomic profiling, combined with in vitro and in silico approaches, offers a comprehensive strategy for exploring phytochemical diversity, identifying active compounds, and validating their antimalarial potential. This integrative approach bridges the gap between traditional knowledge and modern drug discovery.

Aim of the study: The purposes of this study were i) to identify the phytochemical composition via metabolomic analysis, ii) to evaluate the in vitro antiplasmodial activity of seed extracts and iii) to perform in silico molecular docking and molecular dynamics simulations to identify and validate potential antimalarial compounds that target key Plasmodium falciparum enzymes.

Methods: The seeds of B. anthelmintica were subjected to Soxhlet extraction using n-hexane, chloroform, and methanol. The antiplasmodial activity of the extracts was evaluated in vitro against P. falciparum strains Pf-K1 (chloroquine-resistant) and Pf-3D7 (chloroquine-sensitive), and cytotoxicity was assessed using VERO cells. Metabolomic profiling was performed using Gas Chromatography-Mass Spectrometry (GC-MS) and Liquid Chromatography-Mass Spectrometry (LC-MS) to identify phytochemical constituents. The identified compounds were further screened by molecular docking against key P. falciparum targets. The top hits were assessed for pharmacokinetics and TargetNet was employed to assess drug-likeness and AdmetSAR was utilized for ADMET predictions. Molecular dynamics (MD) simulations were conducted to validate the stability of the ligand-target complexes.

Results: Among the extracts, the chloroform fraction exhibited the most potent antiplasmodial activity with IC₅₀ values of 1.7 μg/mL (Pf-K1) and 6.2 μg/mL (Pf-3D7), along with low cytotoxicity (CC₅₀: 17.0 μg/mL) against VERO cells. Metabolomic profiling identified a total of 159 unique phytoconstituents, comprising 131 volatile and thermally stable compounds from GC-MS and 28 non-volatile, polar, and thermally labile compounds from LC-MS analyses. In silico screening, identified compound 4,5-dihydro-4,4-undecamethylene-2-phenyl-1,3-oxazin-6-one as a promising lead, exhibiting a strong binding affinity for P. falciparum. G23 complied with Lipinski's rule of five, showed favorable ADMET characteristics, and maintained stable interactions throughout the MD simulation trajectory.

Conclusion: The findings of this study suggest that B. anthelmintica seeds may contain bioactive phytoconstituents with antiplasmodial potential. Among the identified compounds, G23 (4,5-dihydro-4,4-undecamethylene-2-phenyl-1,3-oxazin-6-one) showed promising in silico affinity for key P. falciparum targets and stable binding in molecular dynamics simulations, supporting its potential as a lead candidate for further antimalarial drug development. The antiplasmodial activity of the crude extract provides a stronger basis for these in vitro findings, and the traditional use of B. anthelmintica.