Abstract
The potential of biomarkers to measure overall responses to dietary/environmental toxicants and putative protective factors has been investigated in vitro and in vivo. In stability studies, isolated lymphocytes in phosphate-buffered saline were stable overnight at 4°C if protected from light and stained lymphocytes were stable up to 120 hours in the lysing solution. In the Comet assay, human lymphocytes in vitro showed a concentration-related response to hydrogen peroxide or N-methyl-nitro-N'-nitrosoguanidine. Caffeine did not induce DNA damage even after activation by Aroclor 1254-induced S9 mix but the solvents used to remove it from green tea (chloroform and dichloromethane) did so even without metabolic activation. Dietary carcinogens 2-amino-3-methyl-[4,5-f]quinoline (IQ), Ochratoxin A (OTA) and mushroom and its metabolites were tested for genotoxicity in vitro with human lymphocytes in the Comet assay. A concentration-related response was associated with IQ, OTA, and raw mushroom extract. Baked mushroom extracts were effective only at high concentrations but the effects of agaritine did not reach statistical significance. 4-(Hydroxymethyl)benzenediazonium ion produced considerably more DNA damage than equimolar amounts of agaritine or N'-acetyl-4-(hydroxymethyl)phenyl-hydrazine. Green tea extract in vitro inhibited IQ-induced DNA damage in human lymphocytes using the Comet assay. Green tea flavonoids showed different efficacy against IQ genotoxicity. Caffeine-induced S9 mix and Aroclor 1254-induced S9 mix activated IQ to a similar extent. The DNA synthesis inhibitor aphidicolin failed to increase the sensitivity of the Comet assay. A preliminary in vivo study using male Wistar albino rats showed modulatory effects of green tea extract against IQ (1 mg/kg b.wt) in the Comet assay and TBA test. On examination of the colon no aberrant crypt foci were observed. HPLC profiling showed specific peaks for green tea and IQ metabolites however, the IQ specific peaks were not identified in the IQ+green tea (GT) group. In a more comprehensive study using a higher level of IQ (5 mg/kg b.wt) similar modulatory effects of green tea were observed in the Comet assay and TBA test. The TBA test using HPLC and colorimetric methods demonstrated that although the trends were similar the values were higher with the latter method. HPLC profiling showed chromatographic patterns similar to the trial study however the enzyme digestion identified conjugates especially sulphate conjugates with IQ and IQ+GT group. This in vivo study demonstrates that green tea (2.5% w/v) is capable of reducing IQ-induced genotoxicity and lipid peroxidation when given simultaneously.