Oxidative damage is involved in the formation of free radicals, which can cause various diseases. In DNA, this damage is observed primarily at guanine (G) because it is the most easily oxidized base, and one form of oxidative damage is DNA-protein crosslinking. Here, we examined the effectiveness of cherry juice components such as melatonin in preventing oxidative DNA damage. Oxidation was effected by the flash-quench technique, a method used for the selective oxidation of guanine. In the flash quench technique, the photoexcited intercalator, Ru(phen)₂dppz²⁺ [phen = phenanthroline, dppz = dipyridophenazine], gives an electron to the quencher, Co(NH₃)₅Cl²⁺. The oxidized intercalator takes an electron from guanine, creating the guanine radical, which then reacts with protein. In our experiment, samples containing Ru(phen)₂dppz²⁺ , Co(NH₃)₅Cl²⁺, histone protein, pUC19 DNA and water, melatonin (1 mM) were irradiated for 0-5 minutes with blue laser light. In the absence of melatonin, the pUC19 displayed a dramatic intensity loss in the supercoiled band, along with increases in intensity for the nicked band and low-mobility material. In contrast, samples containing melatonin showed little change upon irradiation. When samples were irradiated for 5 minutes with decreasing melatonin concentrations, DNA sustained high protection down to 0.1 mM melatonin and some protection down to 0.001 mM melatonin. In a chloroform extraction assay experiment, catechin (also found in cherries) was seen to inhibit DNA-protein crosslinking. In transient absorption spectroscopy experiments, a positive signal at 510 nm for samples with melatonin was observed, consistent with oxidation of the indole ring to form a melatonin radical. In conclusion, catechin and melatonin inhibit oxidative damage to DNA, and spectroscopic experiments provide evidence for a redox mechanism with melatonin. Future work includes the investigation of other components of cherry juice and the determination of a more complete spectral characterization of the signal for melatonin radical.

Angelica Martinez

Appearances