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Biomedical subjects

A A Woodall

Publications and source records attributed to A A Woodall.

7 recordsLinked to original sources
PubMed1998

DNA oxidation matters: the HPLC-electrochemical detection assay of 8-oxo-deoxyguanosine and 8-oxo-guanine.

Oxidative DNA damage is important in aging and the degenerative diseases of aging such as cancer. Estimates commonly rely on measurements of 8-oxo-2'-deoxyguanosine (oxo8dG), an adduct that occurs in DNA and is also excreted in urine after DNA repair. Here we examine difficulties inherent in the analysis of oxo8dG, identify sources of artifacts, and provide solutions to some of the common methodological problems. A frequent criticism has been that phenol in DNA extraction solutions artificially increases the measured level of oxo8dG. We found that phenol extraction of DNA contributes a real but minor increase in the level of oxo8dG when compared, under equivalent conditions, with a successful nonphenol method. A more significant reduction in the baseline level was achieved with a modification of the recently introduced chaotropic NaI method, reducing our estimate of the level of steady-state oxidative adducts by an order of magnitude to 24,000 adducts per cell in young rats and 66,000 adducts per cell in old rats. Of several alternative methods tested, the use of this chaotropic technique of DNA isolation by using NaI produced the lowest and least variable oxo8dG values. In further studies we show that human urinary 8-oxo-guanine (oxo8Gua) excretion is not affected by the administration of allopurinol, suggesting that, unlike some methylated adducts, oxo8Gua is not derived enzymatically from xanthine oxidase. Lastly, we discuss remaining uncertainties inherent both in steady-state oxo8dG measurements and in estimates of endogenous oxidation ("hit rates") based on urinary excretion of oxo8dG and oxo8Gua.

8-Hydroxy-2'-Deoxyguanosine↗
PubMed1997

Carotenoids and protection of phospholipids in solution or in liposomes against oxidation by peroxyl radicals: relationship between carotenoid structure and protective ability.

The ability of carotenoids to protect egg-yolk phosphatidylcholine (EYPC) lipids against oxidation by peroxyl radicals generated from azo-initiators was studied. In homogeneous organic solution, all the carotenoids tested ameliorated lipid peroxidation by AMVN, but none was as effective as alpha-tocopherol. Beta-ring carotenoids showed a correlation between protective effect and rate of carotenoid destruction. Beta,beta-Carotene and zeaxanthin, which react with peroxyl radicals at similar rates, gave a similar degree of protection in organic solution. The reactivity and protective ability of the 4,4'-diketocarotenoids, astaxanthin and canthaxanthin was less. Carotenoids incorporated into ordered membrane systems (EYPC liposomes) displayed different protective efficacies. Zeaxanthin and beta-cryptoxanthin were more effective than beta,beta-carotene against oxidation initiated in the aqueous and lipid phases. Astaxanthin and canthaxanthin afforded less protection to the liposomal lipids. Lycopene was destroyed most rapidly but was least effective as an antioxidant. Located in the hydrophobic inner core of the bilayer, the hydrocarbons lycopene and beta,beta-carotene would not be in a position to readily intercept free-radicals entering the membrane from the aqueous phase. Carotenoids with polar end groups span the bilayer with their end groups located near the hydrophobic-hydrophillic interface where free-radical attack from AAPH first occurs. Hydrogen abstraction from C-4 may be one of the mechanisms of carotenoid antioxidant activity in this system. The chemical reactivity of a carotenoid is not the only factor that determines its ability to protect membranes against oxidation. The position and orientation of the carotenoid in the bilayer is also of importance.

Azo Compounds↗
PubMed1997

Oxidation of carotenoids by free radicals: relationship between structure and reactivity.

The relationship between structure and reactivity is reported for a collection of carotenoids in solution reacted with oxidants generated by a modified Fenton process or with peroxyl radicals generated via the azo-initiators AMVN and AIBN. The initial rates of oxidation were in the order: lycopene > beta,beta-carotene, zeaxanthin > echinenone, isozeaxanthin > astaxanthin, canthaxanthin. The oxidative degradation caused rapid bleaching, due to disruption and breakdown of the polyene chromophore. A number of reaction mechanisms are likely to be involved. Isozeaxanthin, canthaxanthin and astaxanthin, in which the C-4 and C-4' positions are occupied by functional groups, react more slowly than beta,beta-carotene and zeaxanthin, in which this position is free. Products such as the 4-methoxy (or 4-ethoxy) and 4,4'-dimethoxy (or 4,4'-diethoxy) derivatives were isolated from reactions of beta,beta-carotene with peroxyl radicals in the presence of methanol or ethanol. Electron density calculations suggest that the different reactivities cannot be attributed solely to differences in electron distribution along the polyene chain of the different chromophores, which would alter the susceptibility to free-radical addition to the conjugated double-bond system. Other reactions must therefore be considered, including hydrogen abstraction from positions allylic to the polyene chain (C-4 of beta,beta-carotene and its derivatives, and of lycopene). Lycopene, lutein and zeaxanthin all reacted rapidly with oxidising agents, so these dietary carotenoids must also be considered as potential antioxidants.

Carotenoids↗
PubMed1997

gamma-tocopherol traps mutagenic electrophiles such as NO(X) and complements alpha-tocopherol: physiological implications.

Peroxynitrite, a powerful mutagenic oxidant and nitrating species, is formed by the near diffusion-limited reaction of .NO and O2.- during activation of phagocytes. Chronic inflammation induced by phagocytes is a major contributor to cancer and other degenerative diseases. We examined how gamma-tocopherol (gammaT), the principal form of vitamin E in the United States diet, and alpha-tocopherol (alphaT), the major form in supplements, protect against peroxynitrite-induced lipid oxidation. Lipid hydroperoxide formation in liposomes (but not isolated low-density lipoprotein) exposed to peroxynitrite or the .NO and O2.- generator SIN-1 (3-morpholinosydnonimine) was inhibited more effectively by gammaT than alphaT. More importantly, nitration of gammaT at the nucleophilic 5-position, which proceeded in both liposomes and human low density lipoprotein at yields of approximately 50% and approximately 75%, respectively, was not affected by the presence of alphaT. These results suggest that despite alphaT's action as an antioxidant gammaT is required to effectively remove the peroxynitrite-derived nitrating species. We postulate that gammaT acts in vivo as a trap for membrane-soluble electrophilic nitrogen oxides and other electrophilic mutagens, forming stable carbon-centered adducts through the nucleophilic 5-position, which is blocked in alphaT. Because large doses of dietary alphaT displace gammaT in plasma and other tissues, the current wisdom of vitamin E supplementation with primarily alphaT should be reconsidered.

Antioxidants↗
PubMed1996

Dietary supplementation with carotenoids: effects on alpha-tocopherol levels and susceptibility of tissues to oxidative stress.

The ability of dietary supplementation with carotenoids to protect chick tissues against oxidative stress in vitro was examined. Male Leghorn chicks were fed on diets supplemented (100 mg supplement/kg diet) with either beta-carotene, zeaxanthin (beta,beta-carotene-3,3'-diol), canthaxanthin (beta,beta-carotene-4,4'-dione) or alpha-tocopherol, or on a control diet, from 1 d old until 37 d of age. Tissues (liver, heart, skeletal muscle and plasma) were removed and assayed for total carotenoids and alpha-tocopherol content and portions subjected to oxidative stress by incubation of homogenates with cumene hydroperoxide and FeSo4. Animals receiving zeaxanthin and canthaxanthin had significantly greater carotenoid concentrations in liver, heart, muscle and plasma compared with untreated controls (P < 0.05); animals fed on diets supplemented with beta-carotene, or alpha-tocopherol did not have significantly different tissue carotenoid contents compared with untreated controls. alpha-Tocopherol supplementation elevated alpha-tocopherol levels in all tissues examined (P < 0.05). Supplementation with carotenoids did not affect tissue alpha-tocopherol levels, but beta-carotene lowered plasma alpha-tocopherol levels by 50% (P < 0.05). Incubation of plasma or tissue homogenates with oxidant stressors induced lipid peroxidation (production of thiobarbituric-acid reactive substances) in all tissues. Animals given alpha-tocopherol, beta-carotene or zeaxanthin had a reduced susceptibility to oxidant stress in liver compared with unsupplemented controls (P < 0.05), and alpha-tocopherol-supplemented animals had reduced susceptibility in skeletal muscle compared with tocopherol-supplemented animals had reduced susceptibility in skeletal muscle compared with unsupplemented controls (P < 0.05). Canthaxanthin supplementation did not influence the susceptibility to oxidant stress in any tissue examined. These results suggest that zeaxanthin, a carotenoid present in animal and human diets, may have significant activity as an antioxidant against oxidative stress in tissues.

Animals↗