Formation of aliphatic amine precursors of N-nitrosodimethylamine after oral administration of choline and choline analogues in the rat.

Trimethylamine and dimethylamine are important precursors of N-nitrosodimethylamine, which is a potent carcinogen in a wide variety of animal species. Choline, a component of the normal human diet, is metabolized by bacteria within the intestine to form trimethylamine and dimethylamine. However, animals on a choline-free diet continue to excrete some trimethylamine and dimethylamine, suggesting that other dietary precursors of these methylamines might exist. To determine whether C-N bond cleavage by the intestinal bacteria is specific to the choline molecule, we measured monomethylamine, dimethylamine, trimethylamine and trimethylamine oxide excretion in rat urine after the administration of compounds that shared structural features with choline. Water, choline, dimethylaminoethanol, diethylaminoethanol, phosphocholine, betaine, carnitine, beta-methylcholine or dimethylaminoethyl chloride were administered by orogastric intubation, and the urine was collected for 24 hr. Administration of choline (15 mmol/kg body weight) resulted in increased urinary excretion of dimethylamine, trimethylamine and trimethylamine oxide (increases of approximately twofold, 500-fold and 50-fold, respectively). Of the administered choline, 12% was converted to trimethylamine or trimethylamine oxide and excreted in the urine within 24 hr. Phosphocholine administration resulted in similar increases in dimethylamine, trimethylamine and trimethylamine oxide excretion by rats. Modification of the ethyl-backbone or quaternary amine end of the choline molecule resulted in marked suppression of methylamine formation. Though administration of some analogues of choline (methylcholine, betaine and carnitine) resulted in the formation of small amounts of trimethylamine or trimethylamine oxide, and the administration of others (dimethylaminoethanol and dimethylaminoethyl chloride) resulted in the formation of some dimethylamine, the amounts formed were minimal compared with the amounts of trimethylamine and trimethylamine oxide formed after choline administration. Thus, of the many components of foods, only choline and its esters are likely to be significant substrates for trimethylamine and dimethylamine formation. How then can we explain the persistence of trimethylamine and dimethylamine excretion observed in choline-deficient rats? We suggest that endogenous (non-bacterial) synthesis of trimethylamine and dimethylamine occurs within some tissue of the rat.

[Assessment of the degree of carcinogenicity of small doses of nitrites]. / Otsenka stepeni kantserogennoi opasnosti malykh doz nitritov.

Chronic experiments on CBA and C57B1 mice and acute experiments on CBA mice established: (a) carcinogenic effect of sodium nitrite given continuously with drinking water (0.1; 1.0 and 10.0 maximum allowable concentration) in combination with morpholine fed with bread, and (b) endogenous synthesis of nitrosomorpholine as a result of simultaneous intragastric administration of same doses of sodium nitrite and morpholine. Also, nitrosomorpholine and N-nitrosodimethylamine synthesis was observed in vitro following addition of low-dose sodium nitrite, morpholine and amidopyrine to human gastric juice. Carcinogenic hazard associated with low-dose nitrite consumption in humans is discussed.

Nitrosation of bromhexine and aminophenazone in gastric juice after high nitrate loading in the diet. An ex-vivo/in-vitro study in humans.

In an ex-vivo study, nitrosation of bromhexine (N-methyl-N-cyclohexyl-(2-amino-3,5-dibromobenzyl)-ammonium hydrochloride) in human gastric fluid under conditions of high dietary nitrate intake was investigated. 26 healthy volunteers received 200 mg of nitrate in a vegetable during a standard breakfast. Nitrite values in saliva were significantly increased 2 to 3 h after nitrate intake. In contrast, nitrite levels in gastric fluid sampled 3 h after nitrate intake remained in the low concentration range of less than or equal to 1.7 micrograms/No2-/ml. Incubation of gastric fluid samples at 37 degrees C with the recommended maximum single oral dose of bromhexine (16 mg/100 ml) or an equimolar concentration of aminophenazone revealed in 4/26 cases formation of N-nitrosomethylcyclohexylamine from bromhexine at barely detectable levels. In contrast, aminophenazone generated N-nitrosodimethylamine at a very high rate, resulting in yields of greater than 50% in several samples. In view of the average daily background exposure to preformed nitrosamines in foods (0.5-1 microgram/capita), according to these results the possible contribution of in-vivo nitrosation of bromhexine corresponds at best to 10% in addition to the daily background exposure and therefore can be regarded as negligible.

[Formation of volatile carcinogenic N-nitroso compounds from drugs under simulated human gastric conditions]. / Bildung flüchtiger kanzerogener N-Nitrosoverbindungen aus Arzneimitteln unter simulierten Bedingungen des menschlichen Magens.

41 commercial drugs approved for peroral application in the GDR, whose active agents contain N,N-dialkylamino groups in their chemical structures, have been investigated under simulated conditions of the human stomach. With the drugs containing aminophenazone, amitriptyline, doxycycline and oxytetracycline as active agents N-nitrosodimethylamine is formed as a result of nitrosation reactions. With the drugs containing clomiphene++, disulfiram, probenecid and a diethylamine-containing liquid hypnoticum, there occurred N-nitrosodiethylamine. In no case N-nitrosodi-n-propylamine or N-nitrosopiperidine were detectable. The isolated active agents amitriptyline, clomiphene++ and probenecid themselves proved not to be nitrosatable. The positive findings with these drugs were caused by not yet identified nitrosatable contaminants of these drugs. The quantitative determination of volatile N-nitroso compounds was done upon steam distillation by means of gas chromatograph and chemiluminescence detector.

Drug-nitrite interactions in human saliva: effects of food constituents on carcinogenic N-nitrosamine formation.

A simple and rapid high-pressure liquid chromatographic assay for monitoring N-nitrosodimethylamine (NDMA) in human saliva was developed. The method was used to study in vitro the effects of common food constituents on NDMA formation in saliva from the interaction of salivary nitrite with aminopyrine and oxytetracycline. Natural phenolic compounds, caffeic acid, and tannic acid, and synthetic additives, erythorbic acid, sorbic acid, propyl gallate, and butylated hydroxytoluene--all inhibited NDMA formation (20-80%). With ascorbic acid, up to 90% inhibition of NDMA synthesis in saliva was observed. In contrast, chlorogenic acid (a phenolic component of coffee) acted as a catalyst (up to 48% increase) of the nitrosamine formation under identical experimental conditions.

In vivo stability of nitrite and nitrosamine formation in the dog stomach: effect of nitrite and amine concentration and of ascorbic acid.

Formation of N-nitrosodimethylamine (NDMA) from the precursors nitrate and dimethylamine has been studied in the chronically fistulated dog stomach. Measurements have also been made as a function of time for nitrite stability and gastric pH, and a nonabsorbable marker has been used to correct for gastric emptying and dilution. In the fasting animal NDMA forms within minutes after addition of the reactants if the pH is below 5. The presence of food in the stomach slows the reaction, and the additional presence of ascorbic acid greatly depresses the amount of NDMA formed. The concentration of nitrite in gastric juice declines rapidly after its introduction, but the concentration of NDMA declines even more rapidly than nitrite. This suggests that NDMA is probably rapidly absorbed directly from the dog stomach, in contrast to earlier results in rodent experiments.

Dimethylnitrosamine demethylase activity in fetal, suckling, and maternal mouse liver and its transplacental and transmammary induction by polychlorinated biphenyls.

Oxidative demethylation of dimethylnitrosamine (DMN) was detectable with liver homogenate preparations from fetal mice (noninbred Swiss) by day 16 of gestation; this activity increased to adult levels by postnatal day 7. Treatment of pregnant mice with a mixture of polychlorinated biphenyls (PCB's), 500 mg Aroclor 1254/kg, on the last day of gestation caused a large elevation in DMN demethylase activity in the livers of their sucklings, relative to the normal ontogenic increase, reaching a maximum at postnatal day 21. Foster nursing experiments showed that this inductive effect was due primarily to lactational delivery of the PCB]s. DMN demethylase activity in the maternal livers was significantly lower than normal during pregnancy. The mothers' livers were also less responsive to DMN demethylase induction by PCB's during the last 5 days of gestation in comparison with the response earlier in pregnancy or just after delivery. In the livers of the mothers, induced activity resulting from treatment with PCB's on day 19 of gestation reached a maximum on postnatal days 4-7 and declined to normal levels by the time of weaning.

Biosynthesis of dimethylnitrosamine in dimethylamine-treated mice after exposure to nitrogen dioxide.

These studies demonstrate the nitrosating potential of NO2 in vivo in ICR mice. Groups of mice were gavaged with 2 mg dimethylamine (DMA) and exposed to NO2 at levels from 0.04 to 44.5 ppm for periods up to 4 hours. Mice were individually frozen and blended to a powder, aliquots of which were homogenized in ice-cold dicholoromethane and 35% aqueous methanol. Concentrates of organic extracts were analyzed or dimethylnitrosamine (DMN) by a Thermal Energy Analyzer with a gas chromatograph interface. Biosynthesis of DMN was dose- and time-dependent with relation to NO2 exposure, reaching a maximum yield of 60-70 ng/mouse (0.0035% DMA conversion) at 2 hours. DMN biosynthesis was inhibited by sodium ascorbate and, more effectively, by ammonium sulfamate.

[Possibility of endogenous dimethylnitrosamine synthesis in rats administered dimethylamine and nitrite with the food]. / Izuchenie vozmozhnosti éndogennogo sinteza dimetilnitrozamina pri vvedenii krysam s pishchei dimetilamina i nitrita.

Concurrent peroral administration of dimethylamine (DMA) and sodium nitrite to rats produced necrosis of liver parenchyma and increased the activity of glutamicoalanine transaminase. Similar changes were recorded after administering dimethylnitrosamine (DMNA) that points to potential synthesis of this carcinogen from the precursors. Prolonged (over 2.5 years) feeding with DMA and nitrite resulted in part of the rats in tumours of the lungs and in other neoplasms. Ascorbic acid that blocks the endogenous synthesis of DMNA interfered with the development of tumour and pretumour lesions that emerged as a result of concurrent feeding with DMA and nitrite.

Carcinogenic N-nitro-dimethylamine from the reaction of the analgesic amidopyrine and nitrite extracted from foodstuffs.

The reaction of the analgesic amidopyrine (100 mg) with nitrite extracted from cured meats and from spinach in varying degrees of spoilage was studied. Unde physiological conditions the carcinogenic dimethylnitrosamine was formed at milligram levels at nitrite concentrations as low as 4 mg (in 175 ml extracted from 100 g boiled ham). The rate of decrease in concentration in the human stomach after ingestion of amidopyrine and of nitrite contained in boiled ham or in a broth from boiled ham was also measured.

In vitro N-nitrosodimethylamine formation by some bacteria.

It was found that human pathogenic bacteria and some species of intestinal bacteria in rats have the ability to catalyze the formation of N-nitrosodimethylamine from dimethylamine and nitrate in vitro.