Insights into Chemopreventive Effects of Rosmarinic Acid Against Aflatoxin B1-Induced Genotoxic Effects.

In this study, the chemopreventive effects of rosmarinic acid (RA), a major phenolic acid of the plant L., against the carcinogenic naturally occurring mycotoxin aflatoxin B1 (AFB1) were investigated using both in silico and in vitro approaches. The in silico investigation of the chemical reactions between rosmarinic acid and the carcinogenic metabolite of AFB1, aflatoxin B1 exo-8,9-epoxide (AFBO), was conducted by activation free energies calculations with DFT functionals M11-L and MN12-L, in conjunction with the 6-311++G(d,p) flexible basis set and implicit solvation model density (SMD), according to a newly developed quantum mechanics-based protocol for the evaluation of carcinogen scavenging activity (QM-CSA). Following the computational analyses, the chemoprotective effects of RA were further studied in vitro in human hepatocellular carcinoma HepG2 cells by analyzing its influence on AFB1-induced genotoxicity using a comet assay, γH2AX, and p-H3, while its impact on cell proliferation and cell cycle modulation was assessed using flow cytometry. Our computational results revealed that the activation free energy required for the reaction of RA with AFBO (14.86 kcal/mol) is significantly lower than the activation free energy for the competing reaction of AFBO with guanine (16.88 kcal/mol), which indicates that RA acts as an efficient natural scavenger of AFBO, potentially preventing AFB1-specific DNA adduct formation. The chemoprotective activity of RA was confirmed through in vitro experiments, which demonstrated a statistically significant ( < 0.05) reduction in AFB1-induced single- and double-strand breaks in HepG2 cells exposed to a mixture of AFB1 and RA at non-cytotoxic concentrations. In addition, RA reversed the AFB1-induced reduction in cell proliferation.