Integrated GC-MS and LC-MS Metabolomics Reveals Antibacterial-Associated Chemical Signatures in Piper Peltatum

Piper peltatum is an Amazonian medicinal plant with reported antibacterial properties, but the chemical basis of this activity remains poorly characterized. This study evaluated how extraction solvent and leaf drying influence the antibacterial activity and metabolomic profile of P. peltatum. Fresh and oven-dried leaves were extracted with water or methanol and analyzed by GC-MS and LC-MS/MS, followed by integrated multivariate and differential analyses, metadata-based feature networking, and molecular docking. Antibacterial activity was assessed by MIC and MBC assays. A total of 31 metabolites were putatively annotated at MSI level 2 across both analytical platforms. Only the methanolic extracts showed antibacterial activity: the dried-leaf extract was the most active against Bacillus subtilis (MIC 0.06 mg/mL; MBC 0.12 mg/mL), whereas the fresh-leaf extract was the most active against Staphylococcus aureus (MIC 1.95 mg/mL; MBC > 7.81 mg/mL). Integrated metabolomics clearly separated aqueous from methanolic extracts and distinguished fresh from dried methanolic samples, revealing drying-associated differences in relative metabolomic signals within the methanolic fraction. Docking suggested S. aureus DHFR as a testable target, whereas the FtsZ interdomain-cleft result depended on receptor conformation and was inconclusive. Overall, the results indicate that solvent selection and drying reshape the chemical matrix of P. peltatum and provide candidates for subsequent bioactivity-guided and target-specific validation.