Toxicity assessment of carmine and its interaction with calf thymus DNA

Carmine (CRM) is a colouring agent, extensively used in a variety of foods, beverages, cosmetics and pharmaceuticals. A battery of in vitro toxicity tests and chemical interaction studies of CRM with calf thymus DNA (ctDNA) have been conducted. Hemolysis assay clearly revealed the cytotoxic potential of CRM while Allium cepa and seed germination tests for phytotoxicity of CRM were also positive. DNA binding studies withplasmid nicking assay, clearly affirmed the genotoxic potential of CRM. The formation of CRM-ctDNA complex was suggested by UV-visible absorption and fluorescence spectra. The binding constant of CRM-ctDNA complex determined by ITC was found to be 1.72 x 10⁶M^-1. It also revealed the negative ΔH and ΔS values, indicating the role of hydrogen bonds and Van der Waals interactions in the formation of CRM-ctDNA complex. A remarkable (43.4%) displacement of Hoechst as compared to acridine orange (100%) suggests probable disruption of groove which might be possible due to the large size of CRM. Intercalatory mode of CRM binding was further confirmed by significant increase in double helix melting temperature of ctDNA and changes it's in circular dichroism spectra. Molecular docking also supported the notion that DNA binding involved both modes i.e. intercalatory as well as groove binding; moreover, it is consistent with the binding energies determined from ITC. In a nutshell, CRM is significantly cytotoxic to human and plant systems with its remarkable genotoxic potential too, wherein it targets DNA as a partial intercalator and ultimately break its backbone.Communicated by Ramaswamy H. Sarma.