This study presents an innovative approach to the sustainable valorization of industrial sweet cherry (Prunus avium L.) waste from the Vignola Region, Italy, transforming what is typically discarded into a high-value bioactive resource. Unlike conventional extractions, our hydroethanolic extract (VCE) was obtained from the entire cherry waste, including the pericarp, pulp, and stone, as generated by industrial processing. This full-fruit extraction strategy represents a novel and efficient use of agricultural by-products, aligning with circular bioeconomy principles. Sweet cherries are known for their phenolic richness, and spectrophotometric assays (TPC, TFC, reducing power, DPPH, and ABTS) confirmed the extract’s antioxidant capacity. In vitro studies using RAW 264.7 macrophages revealed no cytotoxic effects (MTT assay), along with significant anti-inflammatory activity, evidenced by reduced ROS and NO production and downregulation of iNOS and COX-2. Western blotting showed inhibition of NF-κB nuclear translocation and MAPK pathway signaling. Additionally, agarose gel electrophoresis showed protection against oxidative DNA damage. UPLC-MS/MS analysis identified sakuranetin, aequinetin, and dihydrowogonin as the most representative compounds in VCE. Molecular docking simulations revealed strong and specific binding affinities of these compounds to NF-κB p65 and key MAPK targets. These findings highlight whole sweet cherry waste—including the pit—as a potent and sustainable source of bioactive compounds with promising nutraceutical and pharmaceutical applications.

Background: The accumulation of agri-food waste is a major environmental and economic challenge and converting these by-products into bioactive compounds fits within the circular bioeconomy. This study aimed to evaluate the antimicrobial potential of extracts derived from Cannabis sativa L. leaves (CSE), Crocus sativus tepals (CST), and Prunus avium L. cherry waste (VCE) against four key bacterial species (Staphylococcus aureus, Bacillus subtilis, Escherichia coli, and Pseudomonas aeruginosa). Methods: Minimum inhibitory concentration (MIC) assays were performed to assess antibacterial activity, while a bioinformatic pipeline was implemented to explore possible molecular targets. Full-proteome multiple sequence alignments across the bacterial strains were used to identify conserved, strain-specific proteins, and molecular docking simulations were applied to predict binding interactions between the most abundant compounds in the extracts and their targets. Results: CSE and CST demonstrated bacteriostatic activity against S. aureus and B. subtilis (MIC = 15.6 mg/mL), while VCE showed selective activity against B. subtilis (MIC = 31.5 mg/mL). CodY was identified as a putative molecular target for CSE and CST, and ChaA for VCE. Docking results supported the possibility of spontaneous binding between abundant extract constituents and the predicted targets, with high binding affinities triggering a strong interaction network with target sensing residues. Conclusions: This study demonstrates the antimicrobial activity of these agri-food wastes and introduces a comprehensive in vitro and in silico workflow to support the bioactivity of these agri-food wastes and repurpose them for innovative, eco-sustainable applications in the biotechnology field and beyond.