Abstract
A method of analysis was developed for the measurement of total extractable N-nitroso compounds in biological fluids. The method was based on the selective chemical reduction of N-nitroso compounds to nitric oxide which was subsequently determined with a chemiluminescence analyser. The availability of nitrosatable precursors and nitrosating agents for in vivo N-nitrosation and the occurrence of N-nitroso compounds in vivo were investigated. Nitrosatable precursors were shown to be present in high concentrations in the diet and in biological fluids. The nitrosating agent, nitrite, was shown to be formed from nitrate in the mouth, the amount formed varying considerably between individuals. Raised gastric concentrations of nitrite were shown to be associated with high gastric pH and bacterial colonisation of the stomach. It was demonstrated that nitrite could be transported in the blood, despite its coupled oxidation with oxyhaemoglobin. Total extractable N-nitroso compounds were measured in fasting gastric juice, and were found to be positively correlated with gastric pH. A significant relationship was also demonstrated between raised gastric N-nitroso compound and nitrite concentrations and the growth of nitrate reducing bacteria. Treatment with high doses of vitamin C (4g/day) was found to reduce these gastric N-nitroso compound concentrations. Treatment with the H[2] receptor antagonists Cimetidine and Ranitidine was associated with raised gastric N-nitroso compound concentrations and urinary N-nitrosothiazolidine-4-carboxylic acid concentrations respectively. The N-nitroso compounds present in nitrosated gastric juice were tentatively identified as dipeptide derivatives. Mass spectrometry demonstrated that a major component of nitrosated alanylalanine was N-nitrosoiminodiisopropanoic acid. However, although nitrosated alanylalanine was found to be mutagenic, synthesis of the pure N-nitrosodialkanoic acid demonstrated that this was non-mutagenic. The interrelationships between N-nitroso compound concentration, pH, nitrite concentration, and growth of nitrate reducing bacteria were discussed in relation to the aetiology of human cancer.