naphthoquinones and 1-2-dihydroxy-1-2-dihydronaphthalene

We consider the cytotoxicity and the protection against oxidative stress for members of the naphthalenediol family and the known antioxidant epigallocatechin gallate (EGCG). Compounds include the 1,2-naphthalenediol (1,2-ND), 1,4-ND, 2,3-ND, 1,8-ND, and 1,4-dipropyl-2,3-naphthalenediol (DPND). The cell line is an adherent clone of rat pheochromocytoma (PC12-AC). Oxidative stress was induced by the peroxyl radical generator AAPH. The relative order of cytotoxicity was 1,4-ND > 1,2-ND > DPND > 2,3-ND > 1,8-ND > EGCG, with EC(50)'s of 15, 40, 160, >250, >250, >>250 muM, respectively. Despite their high toxicity, both 1,4-ND and 1,2-ND showed narrow zones of protective behavior whereas DPND, 2,3-ND and 1,8-ND and especially EGCG showed an extended protective range. The total protection obtained for the combination of cells/oxidative stressor/protective compounds (PC12-AC/AAPH/naphthalenediols) was defined by an integrated measure, the cytoprotective area (CPA). We relate the observed cytotoxicity and CPA to the different electronic structures of the naphthalenediols, characterized by the first and second bond dissociation enthalpies and the pK(a)'s for parent (diol) and semiquinone. Since the 2,3- and 1,8-naphthalenediols do not form quinones, their cytotoxicity is much lower than for the compounds which do. Thus selected members of the naphthalenediol family show promise as antioxidants.

Topics: Animals; Antioxidants; Catechin; Cell Line, Tumor; Cell Survival; Dose-Response Relationship, Drug; Free Radicals; Hot Temperature; Hydrogen-Ion Concentration; Inhibitory Concentration 50; Models, Chemical; Naphthols; Naphthoquinones; Oxidation-Reduction; Oxidative Stress; Quinones; Rats; Tetrazolium Salts; Thiazoles; Time Factors

The oxidation of naphthalene-1,2-dihydrodiol (ND) to o-naphthoquinone (NQ) in the lens is believed to be responsible for the formation of cataracts in naphthalene-fed rats. Studies using either recombinant rat lens (RLAR) or human muscle aldose reductase (HMAR) incubated in vitro with ND in the presence of NAD(P) verified that aldose reductase (EC 1.1.1.21) is the dihydrodiol dehydrogenase that catalyzes the oxidation of ND to NQ. Kinetic studies of Vmax/Km indicated that RLAR catalyzes the NAD-dependent oxidation of ND with an optimal pH of 9.0. The corresponding activity of HMAR was lower than that of rat enzyme. The metabolite produced by the incubation of RLAR with ND in the presence of 2-mercaptoethanol and NAD in 20 mM phosphate buffer, pH 7.5, was isolated by C18 reversed-phase high-performance liquid chromatography. The elution profile showed the formation of a new peak that was identical with a peak generated when NQ was incubated under the same condition. The metabolite in both peaks was identified as 4-(2-hydroxyethylsulfanyl)-1, 2-dihydro-1,2-naphthalenedione (HNQ) by 1H and 13C NMR analyses using homonuclear correlation spectroscopy, heteronuclear multiple quantum coherence, and heteronuclear shift correlations via multiple bond connectivities as well as infrared analysis. HNQ is readily autoxidized to 2,3-dihydro-1-oxa-4-thia-9,10-phenanthrenedione. The stoichiometry of 1:1 between the consumption of ND and the formation of NADH for the formation of HNQ implies that rat lens aldose reductase catalyzes a 2e- oxidation of ND to yield the corresponding ketol, which is autoxidized to NQ.

Topics: Aldehyde Reductase; Animals; Cataract; Cells, Cultured; Disease Models, Animal; Humans; Lens, Crystalline; Naphthols; Naphthoquinones; Oxidation-Reduction; Rats

The effects of aldose reductase inhibitors on lens protein modifications induced by naphthalene-1,2-dihydrodiol were investigated in vitro to confirm the role of aldose reductase on naphthalene cataract formation. HPLC analysis of naphthalene-1, 2-dihydrodiol incubated with aldose reductase and NAD+indicated the formation of a metabolite peak corresponding to 1,2-naphthoquinone. Soluble proteins from rat lenses prepared by gel filtration of crude lens extracts through Sephadex PD-10, incubated with naphthalene-1, 2-dihydrodiol in the presence of NAD+displayed an absorbance ca 450 nm and their spectra were essentially identical to those of 1, 2-naphthoquinone-protein adducts. Similar spectra were also obtained from proteins isolated from the intact rat lens after in vitro incubation in medium containing naphthalene-1,2-dihydrodiol. The spectra obtained from lens proteins incubated with 1, 2-dihydroxynaphthalene were distinct from those of either naphthalene-1,2-dihydrodiol or 1,2-naphthoquinone. Aldose reductase inhibitors possessing either hydantoin or carboxylic acid groups prevented protein modification induced by naphthalene-1, 2-dihydrodiol but not protein modification induced by 1, 2-dihydroxynaphthalene or 1,2-naphthoquinone. Therefore, the metabolite formed from naphthalene-1,2-dihydrodiol by aldose reductase is 1,2-naphthoquinone. Lens proteins modified by naphthalene-1,2-dihydrodiol appear essentially identical to protein adducts formed with 1,2-naphthoquinone and their formation can be prevented by both hydantoin and carboxylic acid containing aldose reductase inhibitors.

Topics: Aldehyde Reductase; Animals; Cataract; Chromatography, High Pressure Liquid; Crystallins; Enzyme Inhibitors; Fluorenes; Hydantoins; Imidazoles; Imidazolidines; Lens, Crystalline; Naphthalenes; Naphthols; Naphthoquinones; Phthalazines; Rats; Spectrophotometry