Biochemical Properties, Antioxidant Activity, and In Vitro Ruminal Fermentation of Four Medicinal Plant Species Grown in Northwestern Tunisia

This study assesses the phytochemicals and bioactivity of four plant species: Polypodium vulgare, Chamaemelum nobile, Ocimum forsskaolii Benth, and Lavandula stoechas. Plant chemical composition was determined. Antioxidant activity was assessed using the DPPH assay, and HPLC identified key phenolic compounds. In vitro ruminal fermentation trials evaluated gas production, metabolizable energy (ME), organic matter digestibility (OMd), and the production of volatile fatty acids (VFAs), which were subsequently identified using GC–MS. Significant variations (p ≤ 0.05) were observed among species. C. nobile and O. forsskaolii exhibited the highest total nitrogen contents (3.39 ± 0.42% and 3.20 ± 0.69%, respectively). All species contained high levels of insoluble dietary fiber, with L. stoechas showing the highest neutral detergent fiber (62.39%). C. nobile also recorded the highest polyphenol and flavonoid levels (73.88 ± 0.79 mg GAE/g DW and 27.85 ± 0.54 mg QE/g DW, respectively), along with strong antioxidant activity (IC50 = 0.38 mg/mL). HPLC identified catechol in P. vulgare, ferulic acid in C. nobile, chlorogenic acid in O. forsskaolii, and apigenin in L. stoechas as major compounds. For in vitro fermentation, the highest gas production was recorded at D20, accompanied by increases in ME, OMd, and VFA production. GC-MS analysis revealed that the distribution of total VFAs among acetate, propionate, butyrate, and branched-chain fatty acids varied in a clear dose-dependent manner, closely associated with the acetate-to-propionate (A/P) ratio, which in turn influenced CH4 production. These findings highlight the potential of plant extracts rich in dietary fiber and bioactive compounds to offer a sustainable alternative to conventional feed additives, enhancing ruminal fermentation, energy efficiency, and livestock performance.