Lactarius deliciosus Extract from Green Microwave-Assisted Eutectic Solvent Extraction as a Therapeutic Candidate Against Colon Cancer

<i>Lactarius deliciosus</i> is a widely distributed edible mushroom valued as a functional food due to its rich content of nutrients, phenolic compounds, and flavonoids, which contribute to its strong antioxidant and antimicrobial properties. The present study aimed to optimize a green microwave-assisted extraction method for maximal recovery of bioactive phenolic compounds from <i>Lactarius deliciosus</i> extract (LDE) and to evaluate its antioxidant, antiproliferative, antimetastatic, and anti-inflammatory effects on human colon carcinoma (Caco-2) cells. The study demonstrated that solvent polarity and composition critically influence the recovery of antioxidant biomolecules, identifying water and NaDES 1 (glycerol/glycine/water) as the most efficient and sustainable solvents for microwave-assisted extraction at 225 °C. The LDE showed high levels of phenolic compounds-particularly 4-hydroxybenzoic and vanillic acids-indicating potent antioxidant potential and possible anticancer efficacy. The results revealed that the LDE significantly reduced colony formation and cell adhesion in a dose-dependent manner, leading to nearly complete inhibition of clonogenic survival at the IC₅₀ concentration and a marked increase in cell death among non-adherent colon cancer cells. In addition, LDE inhibited the proliferation of Caco-2 cells by inducing G0/G1 cell cycle arrest and apoptosis, associated with altered mitochondrial potential and increased caspase-3 activity. The LDE modified the redox balance of the cell by decreasing the ROS levels and exerts anti-inflammatory effects through significant downregulation of <i>NOS2</i> expression, without adversely affecting the intestinal barrier. The study concludes that LDE bioactive compounds show strong promise as anticancer and functional ingredients, demonstrating antioxidant, antiproliferative, antimetastatic, and anti-inflammatory effects.