Analysis of hydrogen peroxide in freshly prepared coffees

Coffee has been shown unequivocally to be genotoxic in vitro, but no genotoxicity has been seen in vivo testing. Since the in vitro genotoxicity appears to be dependent on hydrogen peroxide, it is important to know whether hydrogen peroxide is present in prepared coffee and whether it is being formed during the in vitro testing. We have devised a procedure to measure hydrogen peroxide in prepared coffee using disposable reversed-phase columns to decolourize the coffee and retain its catechols. Hydrogen peroxide was assayed in the eluate from the columns by two chromogenic methods: horseradish-peroxidase-mediated oxidation of phenol red and non-enzymatic oxidation of iodide. Three brands of brewed and instant coffee prepared in the manner recommended to the consumer were studied. Although six of the twelve preparations of coffee contained 3-29 microM-hydrogen peroxide, in the other six none could be detected. Sampling, batch, and aging effects may contribute to the variability in the testing, but there is no indication of greater than 100 microM-hydrogen peroxide levels in freshly prepared coffees, as reported in the literature using other methods. Hydrogen peroxide did form slowly in prepared coffee as the beverage became oxygenated, but it formed quickly if the coffee was diluted by addition to an oxygen-containing solution at neutral pH and then incubated at 37 degrees C. These results strongly suggest that adventitious formation of hydrogen peroxide is a confounding factor in the analytical and in vitro genotoxicological testing of coffees.