G. Barja de Quiroga

Simultaneous determination of two antioxidants, uric and ascorbic acid, in animal tissue by high-performance liquid chromatography

A rapid and simple method for the simultaneous analysis of uric and ascorbic acid in extracts of animal tissue is described. The method uses reversed-phase ion-pair chromatography with ultraviolet detection. The technique allows efficient separation of both acids while showing high selectivity, recovery, reproducibility, and sample stability. Calculated levels of both substances in mouse liver tissue were 1.00 +/- 0.05 mumol ascorbic acid/g and 130 +/- 5 nmol uric acid/g.

Relationship between antioxidants, lipid peroxidation and aging

Experiments performed on species as different as flies, rats and frogs are not conclusive about the possibility that antioxidant defenses decrease in old animals. Even when these decreases are found, their physiological meaning is far from clear. Furthermore, a constancy of antioxidant capacity in old age is consistent with the fact that aging is a progressive phenomenon which occurs at a rather constant rate from the mature young to the very old animal, without showing a great acceleration rate in the aged. Nevertheless, experimental results strongly suggest that the maintenance of an appropriate antioxidant/prooxidant balance does have an important role in maintaining health in the aging animal. It is possible that the continuous presence of small amounts of free radicals in the adult tissues of both mature adults and old animals is an important factor in aging (a progressive phenomenon) and susceptibility to disease. Since, similarly to what occurs in procariota, the whole antioxidant system seems to be under homeostatic control in vertebrates, it is imperative to perform comprehensive and detailed studies on the effects of carefully controlled doses of antioxidants on biomarkers of health as well as on the different endogenous cellular antioxidant and prooxidant systems. These studies should have as a final goal the knowledge of which doses of antioxidants are high enough to increase antioxidant protection but low enough to avoid feedback depression of other endogenous antioxidants; this could further improve the health state of humans situated in the middle and last phases of their life span.

Physiological significance of catalase and glutathione peroxidases, and in vivo peroxidation, in selected tissues of the toadDiscoglossus pictus (Amphibia) during acclimation to normobaric hyperoxia

Summary1.Various parameters related to oxidative stress were measured in adultDiscoglossus pictus acclimated for 15 days to either normoxia or hyperoxia (

Effect of natural ageing and antioxidant inhibition on liver antioxidant enzymes, glutathione system, peroxidation, and oxygen consumption inRana perezi

P_{O_2 }

Different levels of hyperoxia reversibly induce catalase activity in amphibian tadpoles

=710 mmHg).2.Total weight of the toads and total and relative wet weight of liver, kidneys, lungs and heart were not changed by hyperoxic acclimation.3.In vivo tissue peroxidation increased in lung, decreased in skeletal muscle, and was not changed in liver, kidney, heart and skin after hyperoxic exposure.4.Hyperoxic acclimation increased catalase activities in the lung, liver, kidney and heart but not in skeletal muscle and skin.5.Liver showed higher GSH-peroxidase activity with cumene-OOH than with H2O2 as substrate, whereas lung, skeletal muscle and skin presented similar GSH-peroxidase activities with both substrates.6.GSH-peroxidase activities did not change between hyperoxic and normoxic animals in liver, lung, skeletal muscle and skin.7.These results show that catalase, not GSH-peroxidase, is the principal H2O2 detoxifying enzyme involved in the adaptation ofD. pictus to hyperoxia.

Lung glutathione reductase induction in aging catalase-depleted frogs correlates with early survival throughout the life span

1. In order to clarify the relative role of catalase (CAT) and glutathione peroxidases (GSH-Px) at normal and high O2 tensions, Rana perezi frogs were chronically treated with aminotriazole (AT), hyperoxia, or both. 2. A 100% survival was observed with both treatments. Hyperoxia increased liver catalase and kidney TBA-RS and decreased GSH-Px. 3. AT caused quantitatively higher alterations than hyperoxia in both organs: CAT was depleted, TBA-RS increased (114% in kidney) and GSH-Px decreased. 4. It is concluded that in Rana perezi (a) CAT, in spite of its much higher KM and Vmax in relation to GSH-Px, is needed to avoid oxidative stress even in normoxia; (b) normoxic tissues have significative amounts of H2O2; (c) GSH-Px does not compensate the lack of CAT.

A comparative study of superoxide dismutase in amphibian tissues

SummaryA study of the physiological role of oxygen free radicals in relation to the ageing process was performed using the liver ofRana perezi, an animal with a moderate rate of oxygen consumption and a life span substantially longer than that of laboratory rodents.Among the five different antioxidant enzymes only superoxide dismutase (SOD) showed an age-dependent decrease. Cytochrome oxidase (COX), glutathione status, in vivo and in vitro liver peroxidation, and metabolic rate did not vary as a function of age.Long-term (2.5 months) treatment with aminotriazole and diethyldithiocarbamate depleted catalase (CAT) activity and did not change both glutathione peroxidases (GPx), COX, reduced (GSH) and oxidized (GSSG) glutathione, or metabolic rate. This treatment resulted in great compensatory increases in SOD (to 250–460% of controls) and glutathione reductase (GR) (to 200%) which are possibly responsible for the lack of increase of in vivo and in vitro liver peroxidation and for the absence of changes in survival rate.The comparison of these results with previous data from other species suggests the possibility that decreases in antioxidant capacity in old age are restricted to animal species with high metabolic rates. Nevertheless, ageing can still be due to the continuous presence of small concentrations of O2 radicals in the tissues throughout life in animals with either high or low metabolic rates, because radical scavenging can not be 100% effective. Compensatory homeostasis among antioxidants seems to be a general phenomenon in different species.

Hyperoxia decreases lung size of amphibian tadpoles without changing GSH-peroxidases or tissue peroxidation

In the lung of Rana perezi no differences as a function of age have been found for any of the five major antioxidant enzymes, reduced (GSH), oxidized (GSSG) or glutathione ratio (GSSG/GSH), oxygen consumption (VO2) and for in vivo or in vitro stimulated tissue peroxidation. This frog shows a moderate rate of oxygen consumption and a life span substantially longer than that of rats and mice. Chronic (2.5 months) catalase depletion in the lung did not affect survival or any additional antioxidant enzyme, GSH, GSSG or in vivo and in vitro lung peroxidation in any age group. Only the GSSG/GSH ratio and the VO2 were elevated in catalase depleted old but not young frogs. After comparison of these results with those obtained in other animal species by other authors we suggest the possibility that decreases in antioxidant capacity in old age be restricted to species with high basal metabolic rates. Nevertheless, scavenging of oxygen radicals can not be 100% effective in any species. Thus, aging can still be due to the continuous presence of small concentrations of O2 radicals in the tissues throughout the life span in animals with either high or low metabolic rates.