Biopeptide-rich fermented hemp seeds: Boosting anti-inflammatory and immune responses through Lactiplantibacillus plantarum probiotic fermentation.

Cannabis sativa L. (hemp) seeds are increasingly recognized as a promising food source rich in phytochemicals that support inflammatory and immunological reactions. This study investigates whether fermentation with Lactiplantibacillus plantarum can further enhance these functional properties, paving the way for hemp seeds to be developed into potent functional food ingredients. Aqueous, 70 % ethanol, and ethyl acetate extracts from both L. plantarum-fermented (FHS) and unfermented hemp seeds (HS) were evaluated for their anti-inflammatory activities using cell-based assays. The 70 % ethanol extract of FHS exhibited marked inhibitory effects on cytokines, including TNF-α, IL-1β, and IL-10, with fermentation significantly enhancing these effects by 25 %, 39.3 %, and 29.6 %, respectively, compared to the unfermented extracts. Additionally, mRNA expression analysis confirmed the strong immunomodulatory potential of the fermented extracts. Intracellular metabolomic analysis revealed that the 'antifolate resistance', 'nicotine addiction', 'aminoacyl-tRNA biosynthesis', and 'D-amino acid metabolism' are highlighted in the reasons for this enhancement. Furthermore, FHS significantly prolonged the survival of C. elegans exposed to pathogens, with gene expression analysis indicating modulation of the innate immune system via regulation of genes such as gcs-1, lys-1, dbl-1, pmk-1, elt-2, and dod-22. A comprehensive metabolomic and correlation analysis identified five novel bioactive peptides (AAELIGVP, AAVPYPQ, VFPEVAP, DVIGVPLG, PVPKVL) and bioactive acids (indoleacetic acid and homovanillic acid) that were enriched during fermentation, which are strongly linked to the enhanced anti-inflammatory and immunomodulatory effects observed. These findings suggest that L. plantarum-fermented hemp seeds hold significant promise as functional ingredients in anti-inflammatory and immunomodulatory food products, with potential applications in health and wellness industries.