Antioxidant activity of 4-methylcoumarin compounds

Polyphenolic coumarins are known to act as antioxidants in biological systems, but it is difficult to distinguish their antioxidant activity from the many other effects they produce in cells. We have determined the radical scavenging capacities of 22 structurally related natural and synthetic 4-methylcoumarins, by measuring their reaction with radicals galvinoxyl and 2,2-diphenyl-1-picrylhydrazyl using electron paramagnetic resonance spectroscopy. Efficient antioxidant activity of 4-methylcoumarins in cells was verified using the DCF fluorescent probe assay for determination of intracellular reactive oxygen species levels. As expected the o-dihydroxysubstituted coumarins were found to be excellent radical scavengers and better than the m-dihydroxysubstituted or monohydroxysubstituted analogs, but surprisingly the corresponding o-diacetoxy derivatives also turned out to be good scavengers, even in the absence of an esterase. Another unexpected result was that the antioxidant efficiency of 4-methylcoumarins could be modulated by introducing an ethoxycarbonylethyl substituent at the C-3 position; this effect cannot be explained by simple electron donating/withdrawing properties. Coumarin concentrations of 10 µM or less were used in all experiments, corresponding to the levels relevant for therapeutic purposes. Considering that 4-methylcoumarins, in contrast to many other coumarins, are not metabolised to toxic epoxide intermediates, these results indicate new promising strategies for the design of non-toxic antioxidant coumarin-based drugs.