A novel melatonin metabolite, cyclic 3-hydroxymelatonin: A biomarker of in vivo hydroxyl radical generation

In the current study, we characterized a urinary melatonin metabolite which could provide a safe and effective method to monitor generation of HO(·) in humans. Using mass spectrometry (MS), proton nuclear magnetic resonance (¹H NMR), COSY ¹H NMR analysis, and calculations on the relative thermodynamic stability, a novel melatonin metabolite was identified as cyclic 3-hydroxymelatonin (3-OHM). 3-OHM is the product of the reaction of melatonin with HO(·) which was generated in two different cell-free in vitro systems. Interestingly, this same metabolite, 3-OHM, was also identified in the urine of both rats and humans. A proposed reaction pathway suggests that 3-OHM is the footprint product that results when a melatonin molecule scavenges two HO(·). When rats were challenged with ionizing radiation which results in HO(·) generation, urinary 3-OHM increased dramatically compared to that of controls. These results strongly indicate that the quantity of 3-OHM produced is associated with in vivo HO(·) generation. Since melatonin exists in virtually all animal species and has a wide intracellular distribution and 3-OHM is readily detected noninvasively in urine, we suggest that 3-OHM is a valuable biomarker that can be used to monitor in vivo HO(·) levels in humans and other species. The measurement of urinary 3-OHM as a biomarker of HO(·) generation could provide clinical benefits in the diagnosis and treatment of diseases.