Cyclic-3-hydroxymelatonin (C3HOM), a potent antioxidant, scavenges free radicals and suppresses oxidative reactions.

Cyclic 3-hydroxymelatonin (C3HOM) is an immediate product of melatonin's interaction with reactive oxygen species. Its presence has been detected in mice, rats and humans. In the current study, the antioxidant capacity and reducing power of this molecule have been systematically studied. C3HOM is found to be a more potent antioxidant than melatonin or vitamin C in terms of its ability to scavenge the hydroxyl radical (HO.) and to recover oxidized horseradish peroxidase to its ground state. The antioxidative mechanism of C3HOM is similar to that of the classic antioxidant, vitamin C, rather than to its precursor melatonin. C3HOM effectively prevents the oxidative degradation of cytochrome C induced by hydrogen peroxide (H2O2). It is speculated that some antioxidative activities of melatonin may be mediated by its metabolite, C3HOM. C3HOM prevents mitochondrial cytochrome C injury and, thus, it is likely to inhibit cellular apoptosis induced by the release of oxidized cytochrome C from mitochondria.

Significance and application of melatonin in the regulation of brown adipose tissue metabolism: relation to human obesity.

A worldwide increase in the incidence of obesity indicates the unsuccessful battle against this disorder. Obesity and the associated health problems urgently require effective strategies of treatment. The new discovery that a substantial amount of functional brown adipose tissue (BAT) is retained in adult humans provides a potential target for treatment of human obesity. BAT is active metabolically and disposes of extra energy via generation of heat through uncoupling oxidative phosphorylation in mitochondria. The physiology of BAT is readily regulated by melatonin, which not only increases recruitment of brown adipocytes but also elevates their metabolic activity in mammals. It is speculated that the hypertrophic effect and functional activation of BAT induced by melatonin may likely apply to the human. Thus, melatonin, a naturally occurring substance with no reported toxicity, may serve as a novel approach for treatment of obesity. Conversely, because of the availability of artificial light sources, excessive light exposure after darkness onset in modern societies should be considered a potential contributory factor to human obesity as light at night dramatically reduces endogenous melatonin production. In the current article, the potential associations of melatonin, BAT, obesity and the medical implications are discussed.

Melatonin in relation to the "strong" and "weak" versions of the free radical theory of aging.

That free radicals and the damage they inflict are related to deteriorative cellular and organismal changes associated with aging and also with the development of a variety of age-related diseases is widely debated. There seems to be little doubt that free radical mutilation of essential molecules contributes to these conditions. Numerous investigators, on the basis of their experimental results, have drawn this conclusion. If the free radical theory of aging and disease development has validity, antioxidants could presumably be successfully used to delay the molecular destruction, cellular loss, and organismal death. In the current review we summarize the experimental data related to the utility of melatonin in protecting against reactive oxygen and reactive nitrogen species-induced cellular damage. While the data supporting a role for melatonin in forestalling aging and prolonging life span per se is not compelling, the findings related to melatonin's ability to reduce the severity of a variety of age-related diseases that have as their basis free radical damage is convincing. To date, the bulk of these investigations have been performed in experimental models of diseases in animals. It is now imperative that similar studies be conducted using humans whose quality of life may benefit from treatment with melatonin.

The potential of melatonin in reducing morbidity-mortality after craniocerebral trauma.

Craniocerebral trauma (CCT) is the most frequent cause of morbidity-mortality as a result of an accident. The probable origins and etiologies are multifactorial and include free radical formation and oxidative stress, the suppression of nonspecific resistance, lymphocytopenia (disorder in the adhesion and activation of cells), opportunistic infections, regional macro and microcirculatory alterations, disruptive sleep-wake cycles and toxicity caused by therapeutic agents. These pathogenic factors contribute to the unfavorable development of clinical symptoms as the disease progresses. Melatonin (N-acetyl-5-methoxytryptamine) is an indoleamine endogenously produced in the pineal gland and in other organs and it is protective agent against damage following CCT. Some of the actions of melatonin that support its pharmacological use after CCT include its role as a scavenger of both oxygen and nitrogen-based reactants, stimulation of the activities of a variety of antioxidative enzymes (e.g. superoxide dismutase, glutathione peroxidase, glutathione reductase and catalase), inhibition of pro-inflammatory cytokines and activation-adhesion molecules which consequently reduces lymphocytopenia and infections by opportunistic organisms. The chronobiotic capacity of melatonin may also reset the natural circadian rhythm of sleep and wakefulness. Melatonin reduces the toxicity of the drugs used in the treatment of CCT and increases their efficacy. Finally, melatonin crosses the blood-brain barrier and reduces contusion volume and stabilizes cellular membranes preventing vasospasm and apoptosis of endothelial cells that occurs as a result of CCT.

Melatonin defeats neurally-derived free radicals and reduces the associated neuromorphological and neurobehavioral damage.

Melatonin and its metabolites are potent antioxidants by virtue of their ability to scavenge both oxygen-based and nitrogen-based radicals and intermediates but also as a consequence of their ability to stimulate the activity of antioxidative enzymes. Melatonin also prevents electron leakage from the mitochondrial electron transport chain thereby diminishing free radical generation; this process is referred to as radical avoidance. The fact that melatonin and its metabolites are all efficient radical scavengers indicates that melatonin is a precursor molecule for a variety of intracellular reducing agents. In specific reference to the brain, melatonin also has an advantage over some other antioxidants given that it readily passes through the blood-brain-barrier. This, coupled with the fact that it and its by-products are particularly efficient detoxifiers of reactive species, make these molecules of major importance in protecting the brain from oxidative/nitrosative abuse. This review summarizes the literature on two brain-related situations, i.e., traumatic brain and spinal cord injury and ischemia/reperfusion, and the neurodegenerative disease, amyotrophic lateral sclerosis, where melatonin has been shown to have efficacy in abating neural damage. These, however, are not the only age-associated neurodegenerative states where melatonin has been found to be protective.

Medical implications of melatonin: receptor-mediated and receptor-independent actions.

The functional versatility and diversity of melatonin has exceeded everyone's expectations. The evidence is substantial that melatonin has multiple receptor-mediated and receptor-independent actions. Considering the unexpectedly widespread distribution of cellular membrane receptors as well as the existence of nuclear binding sites/receptors and the fact that some of melatonin's actions are receptor-independent means that melatonin likely functions in every cell with which it comes in contact. This is highlighted by the fact that there are no morpho-physiological barriers to melatonin, e.g., the blood-brain barrier. In addition to its widespread actions, melatonin synthesis occurs in widely diverse tissues with its production not being relegated to the pineal gland. This should not be unexpected given that it is present throughout the animal kingdom including species that lack a pineal gland, e.g., insects, and in single cell organisms. In this review, only a few of melatonin's effects that involve the interaction of the indoleamine with receptors are described. These functions include the control of seasonal reproduction, modulation of sleep processes and influences on bone growth and osteoporosis. Among the actions of melatonin that are likely receptor independent and that are reviewed herein include its ability to neutralize free radicals which leads to a reduction in cataract formation, reducing oxidative stress due to exposure to hyperbaric hyperoxia, ameliorating hyperthyroidism and abating the toxicity of sepsis and septic shock. These actions alone speak to the diversity of beneficial effects of melatonin; however, the review is no way near exhaustive in terms of what melatonin is capable of doing. Because of its ubiquitous benefits, the pharmaceutical industry is developing melatonin analogues which interact with melatonin receptors. Clearly, the intent of the drugs is to take advantage of some of melatonin's numerous beneficial effects.

Melatonin in walnuts: influence on levels of melatonin and total antioxidant capacity of blood.

OBJECTIVE: We investigated whether melatonin is present in walnuts (Juglans regia L.) and, if so, tested whether eating walnuts influences melatonin levels and the total antioxidant status of the blood. METHODS: Melatonin was extracted from walnuts and quantified by high-performance liquid chromatography. After feeding walnuts to rats, serum melatonin concentrations were measured using a radioimmunoassay and the "total antioxidant power" of the serum was estimated by using the trolox equivalent antioxidant capacity and ferric-reducing ability of serum methods. RESULTS: Mean +/- standard error melatonin concentrations were 3.5 +/- 1.0 ng/g of walnut. After food restriction of rats and then feeding them regular chow or walnuts, blood melatonin concentrations in the animals that ate walnuts were increased over those in the rats fed the control diet. Increases in blood melatonin were also accompanied by increases in trolox equivalent antioxidant capacity and ferric-reducing ability of serum values. CONCLUSIONS: Melatonin is present in walnuts and, when eaten, increase blood melatonin concentrations. The increase in blood melatonin levels correlates with an increased antioxidative capacity of this fluid as reflected by augmentation of trolox equivalent antioxidant capacity and ferric-reducing ability of serum values.

Individual and synergistic antioxidative actions of melatonin: studies with vitamin E, vitamin C, glutathione and desferrioxamine (desferoxamine) in rat liver homogenates.

The pharmacological effects of melatonin, vitamin E, vitamin C, glutathione and desferrioxamine (desferoxamine) alone and in combination on iron-induced membrane lipid damage in rat liver homogenates were examined by estimating levels of malondialdehyde and 4-hydroxyalkenals (MDA+4-HDA). Individually, melatonin (2.5-1600 microM), vitamin E (0.5-50 microM), glutathione (100-7000 microM) and desferrioxamine (1-8 microM) inhibited lipid peroxidation in a concentration-dependent manner. Vitamin C had both a pro-oxidative (25-2000 microM) and an antioxidative (2600-5000 microM) effect. The IC50 (concentration that reduces damage by 50%) values were 4, 10, 426, 2290 and 4325 microM for vitamin E, desferrioxamine, melatonin, glutathione and vitamin C, respectively. The synergistic actions of melatonin with vitamin C, vitamin E, and glutathione were systematically investigated. When melatonin was combined with vitamin E, glutathione, or vitamin C, the protective effects against iron-induced lipid peroxidation were dramatically enhanced. Even though melatonin was added at very low concentrations, it still showed synergistic effects with other antioxidants at certain concentrations. Furthermore, melatonin not only reversed the pro-oxidative effects of vitamin C, but its efficacy in reducing lipid peroxidation was improved when it was combined with pro-oxidative concentrations of vitamin C. The results provide new information in terms of the possible pharmacological use of the combination of melatonin and classical antioxidants to treat free radical-related conditions.

Detection and quantification of the antioxidant melatonin in Montmorency and Balaton tart cherries (Prunus cerasus).

The antioxidant melatonin was recently identified in a variety of edible plants and seeds in high concentrations. In plants, as in animals, melatonin is believed to function as a free radical scavenger and possibly in photoperiodism. In this study, melatonin was detected and quantified in fresh-frozen Balaton and Montmorency tart cherries (Prunus cerasus) using high-performance liquid chromatography. Both cherry species contain high levels of melatonin compared to the melatonin concentrations in the blood of mammals. Montmorency cherries (13.46 +/- 1.10 ng/g) contain approximately 6 times more melatonin than do Balaton cherries (2.06 +/- 0.17 ng/g). Neither the orchard of origin nor the time of harvest influenced the amount of melatonin in fresh cherries. The implication of the current findings is that consuming cherries could be an important source of dietary melatonin inasmuch as melatonin is readily absorbed when taken orally. Also, previously published data and the results presented here show that melatonin is not only endogenously produced but also present in the diet.

Melatonin in plants.

Once thought to be exclusively a molecule of the animal kingdom, melatonin has now been found to exist in plants as well. Among a number of actions, melatonin is a direct free radical scavenger and an indirect antioxidant. Melatonin directly detoxifies the hydroxyl radical (OH), hydrogen peroxide, nitric oxide, peroxynitrite anion, peroxynitrous acid, and hypochlorous acid. The products from each of these reactions have been identified in pure chemical systems and in at least one case in vivo; the interaction product of melatonin with the OH, ie., cyclic 3-hydroxymelatonin, is found in the urine of humans and rats. Some of the products that are produced when melatonin detoxifies reactive species are also highly efficient scavengers. As a result, a cascade of scavenging reactions may enhance the antioxidant capacity of melatonin. Additionally, melatonin increases the activity of several antioxidative enzymes, thereby improving its ability to protect macromolecules from oxidative stress. Melatonin is endogenously produced and is also consumed in edible plants. In animal experiments, feeding melatonin-containing foods raised blood levels of the indole. Because physiologic concentrations of melatonin in the blood are known to correlate with the total antioxidant capacity of the serum, consuming food-stuffs containing melatonin may be helpful in lowering oxidative stress.

N1-acetyl-N2-formyl-5-methoxykynuramine, a biogenic amine and melatonin metabolite, functions as a potent antioxidant.

The biogenic amine The biogenic amine N1-acetyl-N2-formyl-5-methoxykynuramine (AFMK) was investigated for its potential antioxidative capacity. AFMK is a metabolite generated through either an enzymatic or a chemical reaction pathway from melatonin. The physiological function of AFMK remains unknown. To our knowledge, this report is the first to document the potent antioxidant action of this biogenic amine. Cyclic voltammetry (CV) shows that AFMK donates two electrons at potentials of 456 mV and 668 mV, and therefore it functions as a reductive force. This function contrasts with all other physiological antioxidants that donate a single electron only when they neutralize free radicals. AFMK reduced 8-hydroxydeoxyguanosine formation induced by the incubation of DNA with oxidants significantly. Lipid peroxidation resulting from free radical damage to rat liver homogenates was also prevented by the addition of AFMK. The inhibitory effects of AFMK on both DNA and lipid damage appear to be dose-response related. In cell culture, AFMK efficiently reduced hippocampal neuronal death induced by either hydrogen peroxide, glutamate, or amyloid b25-35 peptide. AFMK is a naturally occurring molecule with potent free radical scavenging capacity (donating two electrons/molecule) and thus may be a valuable new antioxidant for preventing and treating free radical-related disorders.

Melatonin prevents delta-aminolevulinic acid-induced oxidative DNA damage in the presence of Fe2+.

Delta-aminolevulinic acid (ALA), a heme precursor which accumulates during lead poisoning and acute intermittent porphyria, is reported to cause liver cancer. The carcinogenic mechanisms of ALA may relate to its ability to generate free radicals through metal-catalyzed oxidation which cause oxidative DNA damage. The aim of this study was to compare the efficacy of melatonin, trolox (vitamin E) and mannitol in altering DNA damage induced by ALA. Herein, we found, in the presence of Fe2+, that ALA-induced formation of 8-hydroxydeoxyguanosine in calf thymus DNA was dose and time-dependent. Melatonin, mannitol and trolox, all of which are free radical scavengers, inhibited the formation of 8-hydroxydeoxyguanosine in a concentration-dependent manner. The concentration of each (melatonin, mannitol and trolox) required to reduce DNA damage by 50%, i.e., the IC50, was 0.52, 0.84 and 0.90 mM, respectively.

Biochemical reactivity of melatonin with reactive oxygen and nitrogen species: a review of the evidence.

Melatonin (N-acetyl-5-methoxytryptamine), an endogenously produced indole found throughout the animal kingdom, was recently reported, using a variety of techniques, to be a scavenger of a number of reactive oxygen and reactive nitrogen species both in vitro and in vivo. Initially, melatonin was discovered to directly scavenge the high toxic hydroxyl radical (*OH). The methods used to prove the interaction of melatonin with the *OH included the generation of the radical using Fenton reagents or the ultraviolet photolysis of hydrogen peroxide (H202) with the use of spin-trapping agents, followed by electron spin resonance (ESR) spectroscopy, pulse radiolysis followed by ESR, and several spectrofluorometric and chemical (salicylate trapping in vivo) methodologies. One product of the reaction of melatonin with the *OH was identified as cyclic 3-hydroxymelatonin (3-OHM) using high-performance liquid chromatography with electrochemical (HPLC-EC) detection, electron ionization mass spectrometry (EIMS), proton nuclear magnetic resonance (1H NMR) and COSY 1H NMR. Cyclic 3-OHM appears in the urine of humans and other mammals and in rat urine its concentration increases when melatonin is given exogenously or after an imposed oxidative stress (exposure to ionizing radiation). Urinary cyclic 3-OHM levels are believed to be a biomarker (footprint molecule) of in vivo *OH production and its scavenging by melatonin. Although the data are less complete, besides the *OH, melatonin in cell-free systems has been shown to directly scavenge H2O2, singlet oxygen (1O2) and nitric oxide (NO*), with little or no ability to scavenge the superoxide anion radical (O2*-) In vitro, melatonin also directly detoxifies the peroxynitrite anion (ONOO-) and/or peroxynitrous acid (ONOOH), or the activated form of this molecule, ONOOH*; the product of the latter interaction is proposed to be 6-OHM. How these in vitro findings relate to the in vivo antioxidant actions of melatonin remains to be established. The ability of melatonin to scavenge the lipid peroxyl radical (LOO*) is debated. The weight of the evidence is that melatonin is probably not a classic chain-breaking antioxidant, since its ability to scavenge the LOO* seems weak. Its ability to reduce lipid peroxidation may stem from its function as a preventive antioxidant (scavenging initiating radicals), or yet unidentified actions. In sum, in vitro melatonin acts as a direct free radical scavenger with the ability to detoxify both reactive oxygen and reactive nitrogen species; in vivo, it is an effective pharmacological agent in reducing oxidative damage under conditions in which excessive free radical generation is believed to be involved.

Autoxidation and toxicant-induced oxidation of lipid and DNA in monkey liver: reduction of molecular damage by melatonin.

Melatonin, the main secretory product of the pineal gland, is a free radical scavenger and antioxidant which protects against oxidative damage due to a variety of toxicants. However, there is little information regarding melatonin's antioxidative capacity in tissues of primates. In this study we examined the protective effects of melatonin in monkey liver homogenates against lipid damage that occurred as a result of autoxidation or that induced by exogenous addition of H202 and ferrous iron (Fe2+). Additionally, we tested melatonin's protective effect against oxidative damage to DNA induced by chromium(III) (CrIII) plus H202. The levels of malondialdehyde and 4-hydroxyalkenals were assayed as an index of lipid peroxidation, and the concentrations of 8-hydroxydeoxyguanosine (8-OHdG) as an endpoint of oxidative DNA damage. The increases in malondialdehyde+4-hydroxyalkenals concentrations as a consequence of autoxidation or after the addition of H202 plus Fe2+ to the homogenates were time-dependent. The accumulation of these damaged products due to either auto-oxidative processes or induced by H202 and Fe2+ were significantly reduced by melatonin in a concentration-dependent-manner. The levels of 8-OHdG were elevated in purified monkey liver DNA incubated with a combination of CrCl3 plus H2O2. This rise in oxidatively damaged DNA was prevented by 10 microM concentration of melatonin. Also, melatonin reduced the damage to DNA that was caused by auto-oxidative processes. These findings in monkey liver tissue document the ability of melatonin to protect against oxidative damage to both lipid and DNA in primate tissue, as observed previously in rodent tissue. The findings provide support for the use of melatonin as suitable agent to reduce damage inflicted by free radical species in primates.

High levels of melatonin in the seeds of edible plants: possible function in germ tissue protection.

The seeds of plants represent the anlage of the next generation and are vital to their existence. Melatonin has been identified in the leaves and flowers of plants but not in seeds. In this study, we examined the seeds of 15 edible plants for the presence of melatonin which was extracted using cold ethanol. Melatonin was initially identified by radioimmunoassay and subsequently quantified and confirmed using high performance liquid chromatography. The physiological concentrations of melatonin in the 15 seeds studied ranged from 2 to 200 ng/g dry weight. The highest concentrations of melatonin were observed in white and black mustard seeds. This level of melatonin is much higher than the known physiological concentrations in the blood of many vertebrates. Since the seed, particularly its germ tissue, is highly vulnerable to oxidative stress and damage, we surmise that melatonin, a free radical scavenger, might be present as an important component of its antioxidant defense system. Thus, melatonin in seeds may be essential in protecting germ and reproductive tissues of plants from oxidative damage due to ultraviolet light, drought, extremes in temperature, and environmental chemical pollutants.

Melatonin directly scavenges hydrogen peroxide: a potentially new metabolic pathway of melatonin biotransformation.

A potential new metabolic pathway of melatonin biotransformation is described in this investigation. Melatonin was found to directly scavenge hydrogen peroxide (H(2)O(2)) to form N(1)-acetyl-N(2)-formyl-5-methoxykynuramine and, thereafter this compound could be enzymatically converted to N(1)-acetyl-5-methoxykynuramine by catalase. The structures of these kynuramines were identified using proton nuclear magnetic resonance, carbon nuclear magnetic resonance, and mass spectrometry. This is the first report to reveal a possible physiological association between melatonin, H(2)O(2), catalase, and kynuramines. Melatonin scavenges H(2)O(2) in a concentration-dependent manner. This reaction appears to exhibit two distinguishable phases. In the rapid reaction phase, the interaction between melatonin and H(2)O(2) reaches equilibrium rapidly (within 5 s). The rate constant for this phase was calculated to be 2.3 x 10(6) M(-1)s(-1). Thereafter, the relative equilibrium of melatonin and H(2)O(2) was sustained for roughly 1 h, at which time the content of H(2)O(2) decreased gradually over a several hour period, identified as the slow reaction phase. These observations suggest that melatonin, a ubiquitously distributed small nonenzymatic molecule, might serve to directly detoxify H(2)O(2) in living organisms. H(2)O(2) and melatonin are present in all subcellular compartments; thus, presumably, one important function of melatonin may be complementary in function to catalase and glutathione peroxidase in keeping intracellular H(2)O(2) concentrations at steady-state levels.

Renal toxicity of the carcinogen delta-aminolevulinic acid: antioxidant effects of melatonin.

An increased incidence of cancer in patients suffering from acute intermittent porphyria (AIP) is thought to be related to delta-aminolevulinic acid (ALA) accumulation. Chronic treatment with ALA augmented 8-oxo-7,8-dihydro-2'-deoxyguanosine levels, decreased microsomal and mitochondrial membrane fluidity and increased lipid peroxidation in blood serum. Co-treatment with melatonin completely counteracted the effects of ALA. Melatonin effectively protects DNA and microsomal and mitochondrial membranes in rat kidney from oxidative damage due to ALA. Because of its low toxicity and anticarcinogenic properties, melatonin could be tested as an agent to reduce oxidative damage in patients with AIP.

Melatonin reduces rat hepatic macromolecular damage due to oxidative stress caused by delta-aminolevulinic acid.

Delta-aminolevulinic acid, precursor of heme, accumulates in a number of organs, especially in the liver, of patients with acute intermittent porphyria. The potential protective effect of melatonin against oxidative damage to nuclear DNA and microsomal and mitochondrial membranes in rat liver, caused by delta-aminolevulinic acid, was examined. Changes in 8-hydroxy-2'-deoxyguanosine (8-OHdG) levels, an index of DNA damage, and alterations in membrane fluidity (the inverse of membrane rigidity) and lipid peroxidation in microsomal and mitochondrial membranes, as indices of damage to lipid and protein molecules in membranes, were estimated. Measurements were made in rat liver after a 2 week treatment with delta-aminolevulinic acid (40 mg/kg b.w., every other day). To test the potential protective effects of melatonin, the indole was injected (i.p. 10 mg/kg b.w.) 3 times daily for 2 weeks. 8-OHdG levels and lipid peroxidation in microsomal membranes increased significantly whereas microsomal and mitochondrial membrane fluidity decreased as a consequence of delta-aminolevulinic acid treatment. Melatonin completely counteracted the effects of delta-aminolevulinic acid. Melatonin was highly effective in protecting against oxidative damage to DNA as well as to microsomal and mitochondrial membranes in rat liver and it may be useful as a cotreatment in patients with acute intermittent porphyria.

Protective effects of melatonin against oxidation of guanine bases in DNA and decreased microsomal membrane fluidity in rat liver induced by whole body ionizing radiation.

The aim of the study was to examine the potential protective effect of melatonin against whole body ionizing radiation (800 cGy). Changes in 8-hydroxy-2'-deoxyguanosine (8-OH-dG) levels, an index of DNA damage, and alterations in membrane fluidity (the inverse of membrane rigidity) and lipid peroxidation in microsomal membranes, as indices of damage to lipid and protein molecules in membranes, were estimated. Measurements were made in rat liver, 12 h after their exposure to radiation. To test the potential protective effects of melatonin, the indole was injected (i.p. 50 mg/kg b.w.) at 120, 90, 60 and 30 min prior to radiation exposure. Both 8-OH-dG levels and microsomal membrane rigidity increased significantly 12 h after radiation exposure. Melatonin completely counteracted the effects of ionizing radiation. Changes in 8-OH-dG levels and membrane fluidity are early sensitive parameters of DNA and microsomal membrane damage, respectively, induced by ionizing radiation and our findings document the protective effects of melatonin against ionizing radiation.

Increased levels of oxidatively damaged DNA induced by chromium(III) and H2O2: protection by melatonin and related molecules.

Chromium (Cr) compounds are known occupational and environmental carcinogens. This trace element is found in the workplace primarily in the valence forms Cr(III) and Cr(VI). Cr(III), which was thought originally to be relatively nontoxic, was recently found to be more reactive toward purified DNA than was chromium(VI). Herein, we examined the ability of Cr(III) to induce oxidative DNA damage by measuring the formation of 8-hydroxydeoxyguanosine (8-OH-dG) in purified calf thymus DNA incubated with CrCl3 plus H2O2. In this system we observed that the Cr(III)-induced formation of 8-OH-dG in isolated DNA was both dose- and time-dependent. When melatonin and related molecules, including 6-methoxy-1,2,3,4-tetrahydro-beta-carboline (pinoline), N-acetylserotonin, 6-hydroxymelatonin and indole-3-propionic acid, were co-incubated with CrCl3 plus H2O2, the accumulations of 8-OH-dG in DNA samples were markedly inhibited in a concentration-dependent manner. The concentrations of each indole required to reduce DNA damage by 50%, i.e. the IC50 values, were 0.48, 0.51, 0.88, 1.00 and 3.08 microM for pinoline, melatonin, N-acetylserotonin, 6-hydroxymelatonin and indole-3-propionic acid, respectively. These results suggest that one of the mechanisms by which Cr(III) may induce cancer is via Fenton-type reactions which generate the hydroxyl radical (*OH). The findings also indicate that the protective effects of melatonin and related molecules against Cr(III)-induced carcinogenesis relate to their direct *OH scavenging ability which thereby reduces the formation of the damaged DNA product, 8-OH-dG.