Methods for the detection of antioxidants which prevent age related diseases: a critical review with particular emphasis on human intervention studies.J Physiol Pharmacol 2005; 56 Suppl 2:49-64JP
It is well documented that reactive oxygen species (ROS) are involved in the aetiology of age related diseases. Over the last decades, strong efforts have been made to identify antioxidants in human foods and numerous promising compounds have been detected which are used for the production of supplements and functional foods. The present paper describes the advantages and limitations of methods which are currently used for the identification of antioxidants. Numerous in vitro methods are available which are easy to perform and largely used in screening trials. However, the results of such tests are only partly relevant for humans as certain active compounds (e.g. those with large molecular configuration) are only poorly absorbed in the gastrointestinal tract and/or may undergo metabolic degradation. Therefore experimental models are required which provide information if protective effects take place in humans under realistic conditions. Over the last years, several methods have been developed which are increasingly used in human intervention trials. The most widely used techniques are chemical determinations of oxidised guanosine in peripheral blood cells or urine and single cell gel electrophoresis (comet) assays with lymphocytes which are based on the measurement of DNA migration in an electric field. By using of DNA-restriction enzymes (formamidopyrimidine DNA glycosylase and endonuclease III) it is possible to monitor the endogenous formation of oxidised purines and pyrimidines; recently also protocols have been developed which enable to monitor alterations in the repair of oxidised DNA. Alternatively, also the frequency of micronucleated cells can be monitored with the cytokinesis block method in peripheral human blood cells before and after intervention with putative antioxidants. To obtain information on alterations of the sensitivity towards oxidative damage, the cells can be treated ex vivo with ROS (H(2)O(2) exposure, radiation). The evaluation of currently available human studies shows that in approximately half of them protective effects of dietary factors towards oxidative DNA-damage were observed. Earlier studies focused predominantly on the effects of vitamins (A, C, E) and carotenoids, more recently also the effects of fruit juices (from grapes, kiwi) and beverages (soy milk, tea, coffee), vegetables (tomato products, berries, Brussels sprouts) and other components of the human diet (coenzyme Q(10), polyunsaturated fatty acids) were investigated. On the basis of the results of these studies it was possible to identify dietary compounds which are highly active (e.g. gallic acid). At present, strong efforts are made to elucidate whether the different parameters of oxidative DNA-damage correlates with life span, cancer and other age related diseases. The new techniques are highly useful tools which provide valuable information if dietary components cause antioxidant effects in humans and can be used to identify individual protective compounds and also to develop nutritional strategies to reduce the adverse health effects of ROS.