Further studies on apparent total N-nitroso compound excretion in mouse feces.

A65

Introduction: We report here further studies on factors affecting fecal apparent total N-nitroso compounds (ANC) in mice. This project concerns the hypothesis that N-nitroso compounds (NOC) in the colon contents are a significant cause of human colon cancer. Our previous research demonstrated that feeding beef and hot dogs increased fecal NOC excretion in mice and that a purified nitrosated ANC precursor in hot dogs was directly mutagenic in the Ames test and contained a compound provisionally identified as N-glucosyl glycine. In other experiments on factors affecting fecal ANC output in mice, we reported that feeding NaNO₂ (1 or 2 g/L drinking water) increased fecal ANC output and 1% hemin fed with NaNO₂ increased fecal ANC two-fold compared to feeding nitrite alone. We now report additional experiments on factors affecting fecal ANC output in mice. Methods: Male 6-week-old Swiss mice were generally fed AIN-93G semipurified diet and tap water, and were treated in various ways for 7 days. Control groups were included in each experiment. Each group contained 4-27 (mean, 9) mice. During the last 24 h feces was collected from individual mice. Total 24-h feces for each mouse was dried at 0^oC to constant weight, and extracted with water. Each extract was analyzed in duplicate for total NOC as before. Results: Treatments and results in nmol ANC/g, mean ± SE, were: (a) For 1000, 250 and 64 mg NaNO₂/L drinking water; 21 ± 2, 9 ± 2, and 7 ± 1 (7.0, 2.3 and 3.5 x untreated controls); (b) for 2 g NaNO₂/L water plus 250, 125 and 64 mg hemin/kg diet, 233 ± 49, 135 ± 19 and 138 ± 27 (4.6, 2.2 and 2.3 x control with 2 g NaNO₂/L alone); (c) for 1 g NaNO₂/L water plus 23 g Na ascorbate/kg diet, 17 ± 4 (0.59 x control with 1 g NaNO₂/L alone); (d) for 1 g NaNO₂/L water plus 400 mg omeprazole/kg diet, 3 ± 1 (0.20 x control with 1 g NaNO₂/L alone); (e) for 1 g NaNO₂/L water plus 18% hot dogs in diet, 59 ± 4 (2.6 x control with hot dog alone). Discussion: We confirmed the finding that nitrite alone increased fecal NOC output. As little as 64 mg NaNO₂/L significantly increased fecal ANC level 3.5 times. Hemin increased fecal NOC output when fed with nitrite. As little as 64 mg hemin/kg diet significantly increased fecal NOC levels. The finding for hemin supports findings by others that heme or hemin increases fecal ANC levels in humans and promotes colon carcinogenesis in rats. The findings that NOC output was lowered when ascorbate or omeprazole was fed with nitrite support our view that nitrite produces ANC by an acid-catalyzed reaction in the stomach and that the resulting ANC pass down the gut to the feces. Support: National Cancer Institute RO3 grant CA-11753 and core grant P30-CA-36727.