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The flavonoids, silymarin, myricetin, quercitin, kaempferol, rutin and kaempferol-3-rutinoside have been examined in combination with the food mutagens, 3-amino-1-methyl-5H-pyrido (4,3-b)indole (Trp-P-2) and 2-amino-3-methylimidazo-(4,5-f) quinoline (IQ), in the Comet assay in human lymphocytes from donor A and human sperm from donor B. These compounds alone have been shown to produce positive responses in the Comet assay, as have the food mutagens. However, in combination with the food mutagens, the flavonoids produced antigenotoxic effects since DNA damage was reduced in the Comet assay in lymphocytes and sperm. The assays were performed in the absence of metabolic activation, since when quercetin and kaempferol were examined in blood with metabolic activation, there was little or no difference in response to that obtained in its absence. In the blood, there was an exacerbation or synergy of response at the lowest doses of the flavonoids. In the sperm, with silymarin, myricetin and quercitin, antigenotoxic effects only were observed, but with kaempferol, in general, there were no protective effects. The food mutagen, 2-amino-1-methyl-6-phenylimadazo (4,5-b)pyridine (PhIP), was also examined in addition to Trp-P-2 and IQ in combination with silymarin and myricetin in donors A and C in human lymphocytes only. Similar exacerbation of effects were found at low doses of these flavonoids with antigenotoxic effects at high doses. This was confirmed in the Ames test. There were slightly different profiles in lymphocytes and sperm, but antigenotoxic effects were observed over a similar dose range. This would suggest that effects occur in somatic and germ cells on a one-to-one ratio. These results have implications for man in terms of risk assessment and in the modulation of isolated food constituents.

The main purpose of this study was to determine whether well-known reactive oxygen species (ROS)-generating agents can induce DNA damage in a simple chemical system with or without Fenton reaction components (iron and reducing agents), and to explore whether antioxidants which normally exist in the cellular environment can modify such damage, i.e. to determine chemical reactions of relevance to biological systems. A neutral electrophoresis technique was used to investigate DNA double stranded breaks (DSBs) caused by chemical treatments of lambda-DNA in eppendorf tubes by various ROS-generating compounds and the degree of DNA damage was categorised by analysis of enhanced digital images. Double strand breaks were induced by hydroquinone (HQ), benzoquinone (BQ), benzenetriol (BT), hydrogen peroxide (H2O2), bleomycin (BLM) and sodium ascorbate (Vit C). DNA damage was modulated by various agents including catalase (CAT), superoxide dismutase (SOD), desferoxamine mesylate (DFO), ferrous chloride (FeCl2), reduced glutathione (GSH), trolox, silymarin and myricetin. Individual chemicals (except BLM) at the concentration of 1 mM did not induce large numbers of DSBs without iron [Fe(II) or Fe(III) at 25 microM]. GSH enhanced the damaging effect of HQ, BT and Vit C, did not alter the non-damaging effect of H2O2, but had a small protective effect on BLM. When compared with the non-enzyme protein, bovine serum albumin (BSA), SOD had a protective effect against BT, H2O2 and BLM; in the presence of GSH, SOD diminished the effect of HQ, BQ and Vit C but enhanced the effect of BT, H2O2 and BLM. With both GSH and Fe and compared with BSA, SOD enhanced the effect of HQ, BQ and BLM, ameliorated the effect of H2O2, and did not affect the others. CAT showed a protective effect for almost all examined compounds, but had little effect on BLM. With GSH alone, DFO enhanced the effect of HQ, BQ, H2O2 and ameliorated the effect of BT, BLM and Vit C and trolox was largely protective. With GSH and Fe, DFO was protective for all compounds except doxorubicin (Dox), trolox was protective for all compounds except Dox and BLM, silymarin was protective except that it had little effect on BLM and Dox, but myricetin did not show any protective effect. In conclusion, the results from the present study have further highlighted the adverse potential of reducing agents and redox cycling agents, and also the need for a cautious view of antioxidants.

The effects of the mutagens, cyclophosphamide (CP), ethyl methanesulphonate (EMS), bleomycin (BLM) and the testicular toxin ethylene glycol monomethyl ether (EGME) in bone marrow and testicular cells have been compared in the alkaline COMET assay. Sprague-Dawley rats were administered by gavage with 50, 100 and 150 mg/kg body weight (bw) of CP; 100, 200 and 300 mg/kg bw EMS; 50, 100 and 150 mg/kg bw BLM and 500, 1000 and 1500 mg/kg bw EGME. Effects were examined at week 2 after treatment for CP, EMS BLM and EGME and at weeks 5 and 6 for EGME. Bone marrow cells were removed and separated by aspiration of the femur and testicular cells by decapsulation of the testis, treating with collagenase followed by trypsin. Various statistical methods were used to analyse the data. For CP there was an increase in damage above control values for bone marrow at 50 mg/kg bw which decreased at 100 mg/kg bw, and there was mortality of the animals at 150 mg/kg bw. A similar response was found in the testicular cells. For EMS and BLM, there were only occasional slight increases in damage in bone marrow and testicular cells. Two studies were conducted with EGME. In the first, where effects were examined at week 2 after treatment, there was an increase in damage in bone marrow cells, but a larger response was observed in testicular cells. In the second study where effects were examined at weeks 5 and 6 after treatment, bone marrow and testicular cells were not affected. The overall results showed that damage persisted for 2 weeks after treatment with CP and EGME but not in weeks 5 and 6 for EGME. Various statistical methods were used to analyse the data. Statistically significant responses were produced after treatment with CP and EGME and were dose-related for EGME, but after treatment with EMS and BLM statistical increases were sporadic. These results suggest that the assay is useful for measuring DNA damage and its persistence, and for comparing the sensitivity of different target organs in vivo.