Toxic oxygen free radicals have been implicated as important pathologic mediators in many clinical disorders. We discuss the chemistry of oxygen radical production and the roles of iron and of various antioxidants as well as the diseases that have received active attention in oxy-radical research. Particular attention is focused on cigarette smoke oxidants, ischemia-reperfusion-induced radical production, carcinogenesis, and aging. Such research may well provide a firm foundation for therapeutic breakthroughs.

Oxidative damage to DNA is caused by reactive by-products of normal metabolism, as well as by radiation. Oxidized DNA bases excised by DNA repair enzymes and excreted in urine were measured in four different species to determine the relation between specific metabolic rate (ml of 02 consumed per gram of body weight per hr) and oxidative DNA damage. An average of 6.04 nmol of thymine glycol per kg/day and 2.58 nmol of thymidine glycol per kg/day were found in mouse urine and 1.12 nmol of thymine glycol per kg/day and 0.95 nmol of thymidine glycol per kg/day were found in monkey urine. On a body weight basis, mice excrete 18 times more thymine glycol plus thymidine glycol than do humans, and monkeys excrete 4 times more thymine glycol plus thymidine glycol than do humans. When results among mice, rats, monkeys, and humans are compared, specific metabolic rate correlates highly with oxidative DNA damage. These findings are consistent with the theory that free radicalinduced DNA damage may play a central role in the aging process.efforts focused on one DNA oxidation product, the thymine glycol lesion (5,6-dihydro-5,6-dihydroxythymine) because this lesion has suitable properties of repair, product excretion, stability (for recovery and measurement of its repair products), and traceability of the origin of the repair products to cellular DNA. Background levels of thymine glycol and thymidine glycol in the human were determined, and a total oxidative damage rate to DNA was estimated as ==300 hits per cell per day in human for just these two oxidized bases (8,9). Hydroxymethyluracil, another known oxidative DNA damage product, was also assayed and accounted for another 700 hits per cell per day (8, 9). Because many known oxidative DNA damage products have been described, we suspect that the total damage rate is thousands of hits per cell per day. The rat has a damage rate about 15 times that of the human (8, 9). In this paper, we report the background levels of thymine glycol and thymidine glycol in mouse and monkey and compare the levels in all four species to specific metabolic rate and life span.Free radicals that are formed in the body as a consequence of aerobic metabolism can produce oxidative damage to macromolecules in somatic cells (1)(2)(3)(4)(5). This type of damage may be an important factor in aging and age-dependent diseases such as cancer and heart disease (6). Circumstantial evidence implicating free radicals in aging includes the impressive inverse correlation between the specific metabolic rate (ml of 02 consumed per gram of body weight per hr of a given species and the life span of that species (for review, see ref. 7). One explanation for this inverse correlation is that smaller animals, with higher metabolic rates, consume greater quantities of oxygen (on a body-weight basis) and so produce harmful free-radical by-products at a higher rate, leading to a higher rate of damage to critical cellular targets. The higher rate of damage to cells might then cause a higher rate of cellular aging.Many...

Thymine glycol is a DNA damage product of ionizing radiation and other oxidative mutagens. In an attempt to find a noninvasive assay for oxidative DNA damage in individuals, we have developed an HPLC assay for free thymine glycol and thymidine glycol in urine. Our results indicate that humans excrete about 32 nmol of the two glycols per day. Rats, which have a higher specific metabolic rate and a shorter life span, excrete about 15 times more thymine glycol plus thymidine glycol per kg of body weight than' do humans.' We present evidence that thymine glycol and thymidine glycol are likely to be derived from repair of oxidized DNA, rather than from alternative sources such as the diet or bacterial flora. This noninvasive assay of DNA oxidation products may allow the direct testing of current theories which relate oxidative metabolism to the processes of aging and cancer in man.Oxygen radicals and other reactive oxygen species are generated in vivo as a consequence of normal metabolism (1-4). The oxidation of certain cellular components by these oxygen species could contribute to aging and age-dependent diseases such as cancer. Oxygen radicals have been shown to produce base damage and strand breaks in DNA (1, 3), as well as to initiate the process of lipid peroxidation (1, 3). The latter results in the formation of lipid hydroperoxides, which, in the presence of cellular iron-containing compounds, can also break down to yield oxygen radicals (1).Numerous defense mechanisms within the cell have evolved to limit the levels of reactive oxygen species and the damage they induce (1-5). Among the cellular defenses are the enzymes superoxide dismutase, catalase, and glutathione peroxidase (1-4), as well as antioxidants such as (3-carotene (1-4), the tocopherols (6), and uric acid (7). However, low levels of reactive oxygen species can escape these cellular defenses (8) and produce damage to DNA, protein, and unsaturated fats. One product that is formed in DNA in vitro as a consequence of chemical oxidation or ionizing radiation (an oxidative mutagen) is thymine glycol (5,6-dihydroxydihydrothymine) (9,10). Mammalian DNA repair systems are known to excise this lesion from DNA in vitro (5,(11)(12)(13)(14)(15)(16)(17). We report here the identification, in both human and rat urine, of the cis isomer of thymine glycol (5,6-dihydroxydihydrothymine) and its deoxyribonucleoside thymidine glycol (5,6-dihydroxydihydrothymidine) and we propose that these bases are derived from repair of oxidized DNA.MATERIALS AND METHODS GBq), were oxidized to the cis-glycols by potassium permanganate (9). After removal of the unreacted permanganate with Na2S205, the supernatant was adjusted to 1 M NH4OAc (pH 8.8) and loaded onto a 0.5 x 2 cm boronate affinity column (Affi-Gel 601, Bio-Rad), equilibrated with the same buffer. The column was washed with 1 M NH4OAc (pH 8.8) (buffer A), the cis-glycol fraction was removed by elution with 0.1 M HOAc, and the eluate was concentrated by lyophilization.[3H]Thymine glycol and [3H]thymidine glycol ...

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We have demonstrated that ammonia is oxidized to nitrate in the rat. Male Sprague-Dawley rats gavaged with 1000 mumol N-15-ammonium chloride each day for 5 days were found to excrete low, but significant amounts of excess N-15-nitrate in their urines on the five days of treatment and on the five subsequent days. We recovered a total of 0.28 +/- 0.03 mumol excess N-15-nitrate (mean +/- SE) per rat, which indicates that ammonia is converted to nitrate in a yield of approximately 0.0080%. The oxidation of N-15-labeled glycine and L-glutamic acid to N-15-nitrate could not be detected. N-15-Hydroxylamine was oxidized in the rat to N-15-nitrate in a yield of 4.7%, which shows that hydroxylamine is a possible intermediate in the ammonia oxidation process. Injection of rats with Arochlor 1254, an inducer of several isozymes of cytochrome P-450, did not significantly affect the rate of endogeneous nitrate synthesis. Carbon tetrachloride, which causes hepatic lipid peroxidation, produced a small but significant increase in nitrate synthesis. We confirmed the observation that a bacterial endotoxin can greatly stimulate nitrate synthesis, and we showed that concurrent treatment with superoxide dismutase does not modify the effect of the endotoxin. An in vitro chemical model system was used to demonstrate that oxidation of ammonia to nitrate by the hydroxyl radical at physiological pH is chemically feasible. Our results are consistent with the hypothesis that ammonia is oxidized to nitrate in vivo by a non-enzymatic process which involves active oxygen species such as the hydroxyl radical. We estimate that a 215 g rat produces 3.0 mumol of nitrate per day via ammonia oxidation.