Absolute Rate Constants for the Quenching of Reactive Excited States by Melanin and Related 5,6-Dihydroxyindole Metabolites: Implications for Their Antioxidant Activity

Abstract

The triplet-excited state of benzophenone and the singlet-excited state of 2,3-diazabicyclo[2.2.2]oct-2-ene (Fluorazophore-P) have been employed as kinetic probes to obtain information on the antioxidant activity of the skin and eye pigment melanin and its biogenetic precursors 5,6-dihydroxyindole (DHI) and 5,6-dihydroxyindole-2-carboxylic acid (DHICA). The excited states were generated by the laser-flash photolysis technique and their reaction kinetics was examined by time-resolved transient absorption or fluorescence spectroscopy, respectively. The reaction between triplet benzophenone and DHI produced with unit efficiency the corresponding 6O-centered semiquinone radical, which was characterized by its characteristic transient absorption. The quenching rate constants for DHI (3.1-8.4 x 10(9) M-1 s-1) and DHICA (3.3-5.5 x 10(9) M-1 s-1) were near the diffusion-controlled limit, indicating excellent antioxidant properties. Kinetic solvent effects were observed. The reactivity of synthetic melanin, assessed through the quenching rate constant of Fluorazophore-P and normalized to the number of monomer units, was more than one order of magnitude lower (2.7 x 10(8) M-1 s-1) than that of its precursors. The trend of the quenching rate constants, i.e. DHI > DHICA approximately alpha-tocopherol > melanin, along with the preferential solubility of DHICA in aqueous environments, serves to account for several experimental results from biochemical studies on the inhibition of lipid peroxidation by these natural antioxidants.

Chemical and Structural Disorder in Eumelanins: A Possible Explanation for Broadband Absorbance

“…Eumelanin is the more prevalent melanin pigment and has long been thought to be synthesized in vitro from tyrosine via the Raper-Mason enzymatic pathway (3,4). To date, biochemical and biophysical research has primarily focused on understanding the basic photochemistry, photobiology and photophysics of these important substances, and has been stimulated by the key role that melanins play in photoprotection and their potential involvement in the development of melanoma skin cancer (2). In addition to this biologically directed research, melanins have also attracted interest from molecular biophysicists (5)(6)(7), quantum chemists (8)(9)(10)(11)(12)(13)(14)(15), and more recently, the functional materials and condensed matter physics communities (16) due to their rather unique physio-chemical properties.…”

Study on the binding of 2,3-diazabicyclo[2.2.2]oct-2-ene with bovine serum albumin by fluorescence spectroscopy

“…Therefore, fluorescence quenching can be considered as a sensitive technique to monitor protein fluorescent probe interactions. Critical literature survey reveals that attempts have not been made so far to investigate the mechanism of interaction of azoalkane [2,3- The DBO fluorophore has already been extensively used as a fluorescent probe to study its inter and intramolecular fluorescence quenching by biomolecular compounds such as tryptophan and tyrosine [9][10][11], the nucleobase guanine [12,13], and antioxidants [14][15][16]. More recently, DBO has also been employed as a fluorescence energy acceptor [17][18][19][20], e.g., to investigate fluorescence energy transfer from the amino acid Trp as donor in peptides [17][18][19][20].…”

Photobleaching of retinal pigment epithelium melanosomes reduces their ability to inhibit iron‐induced peroxidation of lipids

“…There is another way melanin can protect pigmented cells against oxidative damage: by acting as an antioxidant, neutralizing oxidizing radicals and other reactive oxygen species that may be generated via chemical and photochemical reactions. In model systems of different complexity, synthetic and natural melanins have been shown to act as efficient scavengers of reactive free radicals, quenchers of singlet oxygen and excited triplet states of certain photosensitizing dye molecules, and inhibitors of lipid peroxidation (Sarna et al, 1986;Sealy et al, 1984;Ostrovsky et al, 1987;Scalia et al, 1990;Korytowski et al, 1995;Zareba et al, 1995;Krol and Liebler, 1998;Rozanowska et al, 1999;Zhang et al, 2000;Ye et al, 2003). Results of an intriguing study recently published by Seagle et al (2005a) indicated an inverse correlation between blue-light-induced apoptosis of human RPE cells in vitro and their melanin content.…”