Theoretical docking analysis for a multi-scale experimental investigation of biochar filtration of olive mill effluents: Unraveling genotoxicity and polyphenol-DNA interactions

Olive mill wastewater (OMWW) is a major by-product of olive oil production and a serious environmental pollutant due to its high organic and phenolic content, which can harm soil, water, and biodiversity. In this study, we aimed to evaluate the effectiveness of olive-stone-derived biochar as a sustainable filtration method to reduce both phytotoxicity and genotoxicity of OMWW. The raw effluent (pH = 4.8, COD = 65.15 g/L, total phenolics = 6.8 g/L) was filtered using biochar pyrolyzed at 585°C for 90 min. To evaluate the effect of the treatment, a micronucleus assay was performed on Vicia faba root cells to assess genotoxicity, complemented by phytotoxicity tests using radish and lettuce seeds. Five concentrations of treated OMWW (25-100 % v/v), raw OMWW, distilled water (negative control), and maleic hydrazide (positive control) were tested in triplicate. The biochar treatment significantly reduced phytotoxicity, with germination rates exceeding 80 % and germination indices above 100 % in treated samples, indicating improved seed germination and seedling vigor compared to untreated OMWW. Genotoxicity was significantly decreased, with micronucleus frequency dropping from 15.2 ‰ in raw OMWW to below 2 ‰ after biochar filtration. Molecular docking revealed strong binding of key OMWW polyphenols to the DNA double helix (binding energies ranging from -4.39 to -8.96 kcal/mol), with quercetin exhibiting the highest affinity. These interactions likely induce structural alterations in DNA, providing a molecular basis for the genotoxicity observed experimentally. The reduction in genotoxic and phytotoxic effects observed after treatment indicates that biochar likely adsorbs bioactive pollutants, reducing their availability and capacity to induce harmful biological responses. To the best of our knowledge, this is one of the first studies to combine phytotoxicity, genotoxicity, and molecular modeling approaches to assess the detoxification of olive mill wastewater (OMWW). These findings highlight the potential of olive-stone-derived biochar as a low-cost, eco-friendly solution for the sustainable management of olive mill effluents. Future large-scale applications should focus on optimizing the biochar dosage and ensuring the sustainability of the treatment system.