Metabolism Of Heterocyclic Aromatic Amines Research Articles

The Human Fecal Microbiota Metabolizes Foodborne Heterocyclic Aromatic Amines by Reuterin Conjugation and Further Transformations.

Heterocyclic aromatic amines (HAAs) are process-induced food contaminants with high mutagenic and/or carcinogenic potential. Although the human gut microbiota is known to affect the metabolism of dietary constituents, its impact on HAA metabolism and toxicity has been little studied. Here, the glycerol-dependent metabolism of seven foodborne HAAs (AαC, Trp-P-1, harman, norharman, PhIP, MeIQx, and MeIQ) by the human fecal microbiota is investigated. As analyzed by HPLC-DAD/FLD, the extent of conversion is strongly dependent on glycerol supplementation and HAA structure. AαC (60-100%) and the 2-aminoimidazoazarenes (up to 58%) are especially prone to microbial conversion. Based on high-resolution MS and/or NMR spectroscopy data, 70 fecal metabolites are identified in total, mainly formed by chemical reactions with one or two molecules of microbially derived reuterin. Moreover, it has been demonstrated that the human fecal microbiota can further transform reuterin adducts by reduction and/or hydroxylation reactions. Upon isolation, some reuterin-induced HAA metabolites appear to be partially unstable, complicating structural identification. The formation of microbial metabolites needs to be incorporated into risk assessment considerations for HAAs in human health. In this study, several HAA metabolites, mainly reuterin-dependent, are identified in vitro, providing the basis for future human studies investigating microbial HAA metabolism.

Metabolism of heterocyclic aromatic amines (HAA) by Lactobacillus reuteri DSM 20016

Metabolism of heterocyclic aromatic amines (HAA) by Lactobacillus reuteri DSM 20016

Metabolism and toxicology of heterocyclic aromatic amines when consumed in diet: Influence of the genetic susceptibility to develop human cancer. A review

There is not sufficient scientific evidence to support the hypothesis that human cancer risk is specifically due to the intake of heterocyclic aromatic amines (HAAs) in diet. Epidemiological evidence appears to imply two main factors in the HAAs carcinogenicity. These factors are the very high frequency of consumption of red meats, and very darkly browned meats from cooking, which are HAAs-containing foods. The present review focuses on the fact that the cancer risk is notably enhanced when certain genotypes related to HAAs metabolism are present. Thus, genetic predisposition seems to be the main factor in cancer development related to HAAs, and possibly the co-presence of other mutagenic compounds in diet.

Interspecies metabolism of heterocyclic aromatic amines and the uncertainties in extrapolation of animal toxicity data for human risk assessment

Heterocyclic aromatic amines (HAAs) are potent bacterial mutagens that are formed in cooked meats, tobacco smokes condensate, and diesel exhaust. Many HAAs are carcinogenic in experimental animal models. Because of their wide-spread occurrence in the diet and environment, HAAs may contribute to some common types of human cancers. The extrapolation of animal toxicity data on HAAs to asses human health risk has many uncertainties, which can lead to tenuous risk assessment estimates. Perhaps the most critical and variable parameters in interspecies extrapolation are the effects of dose, species differences in catalytic activities of xenobiotic metabolism enzymes (XMEs), human XME polymorphisms that lead to interindividual differences in carcinogen metabolism and dietary constituents that may either augment or diminish the carcinogenic potency of these genotoxins. The impact of these parameters on the metabolism and toxicological properties of HAAS and uncertainties in extrapolation of animal toxicity data for human risk assessment are presented in this article.

The role of genetic polymorphisms in metabolism of carcinogenic heterocyclic aromatic amines.

More than twenty heterocyclic aromatic amines (HAAs) have been identified in grilled meats, fish, poultry, and tobacco smoke condensate. HAAs are carcinogens and induce tumors at multiple sites in experimental laboratory animals. Because of the widespread occurrence of HAAs in foods, these chemicals may contribute to the etiology of several common human cancers that are associated with frequent consumption of grilled meats including colon, rectum, prostate, and breast. HAAs require metabolism in order to exert their genotoxic effects. Metabolic activation occurs by N-hydroxylation, a reaction catalyzed by cytochromes p450 (CYP). Some N-hydroxy-HAA metabolites may directly react with DNA, but further metabolism by N-acetyltransferases (NATs) or sulfotransferases (SULTs) may occur to form highly reactive N-acetoxy or N-sulfonyloxy esters that readily react with DNA bases. The N-acetoxy ester of the HAA 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) is detoxified by glutathione S-transferases (GSTs), which catalyze the reduction of the reactive intermediate back to the parent amine. Some HAAs also undergo detoxification through conjugation reactions with the phase II enzymes such as UDP-glucuronosyltransferases (UGTs) or SULTs to form stable, polar products that are readily eliminated. All of these xenobiotic metabolism enzyme systems (XMEs) display common genetic polymorphisms, which may affect protein expression, protein stability, catalytic activity, and thus, the biological potency of these procarcinogens. In this review, the roles of common genetic polymorphisms of XMEs involved in HAA metabolism and cancer risk are discussed.

Metabolism of heterocyclic aromatic amines by human hepatocytes and cytochrome P4501A2

The metabolism of 2-amino-3,8-dimethylimidazo[4,5- f]quinoxaline (MeIQx) and 2-amino-1-methyl-6-phenylimidazo[4,5- b]pyridine (PhIP) was investigated in primary human and rat hepatocytes. The genotoxic metabolites 2-(hydroxyamino)-3,8-dimethylimidazo[4,5- f]quinoxaline (HONH-MeIQx) and 2-(hydroxyamino)-1-methyl-6-phenylimidazo[4,5- b]pyridine (HONH-PhIP), which are formed by cytochrome P4501A2 (CYP1A2), were detected as stable N 2-glucuronide and N 2- and N 3-glucuronide conjugates, respectively. These products accounted for as much as 10% of the amount of MeIQx and 60% of PhIP added to human hepatocytes. Significantly lower amounts of these products were formed in rat hepatocytes. The phase II conjugates N 2-(3,8-dimethylimidazo[4,5- f]quinoxalin-2-yl-sulfamic acid (MeIQx- N 2-SO 3H) and N 2-(β-1-glucosiduronyl)-2-amino-3,8-dimethylimidazo[4,5- f]quinoxaline (MeIQx- N 2-Gl), as well as the 7-oxo derivatives of MeIQx and N-desmethyl-MeIQx, 2-amino-3,8-dimethyl-6-hydro-7 H-imidazo[4,5- f]quinoxalin-7-one (7-oxo-MeIQx), and 2-amino-6-hydro-8-methyl-7 H-imidazo[4,5- f]quinoxalin-7-one ( N-desmethyl-7-oxo-MeIQx) were also identified. A novel CYP1A2-derived metabolite was characterized as 2-amino-3-methylimidazo[4,5- f]quinoxaline-8-carboxylic acid (IQx-8-COOH) and was the predominant metabolite formed in human hepatocytes exposed to MeIQx at levels approaching human exposure. Unlike human hepatocytes, rat cell preparations, even following pretreatment with the potent CYP1A1/CYP1A2 inducer 3-methylcholanthrene (3-MC) did not produce IQx-8-COOH but did catalyze the formation of 2-amino-3,8-dimethyl-5-hydroxyimidazo[4,5- f]quinoxaline (5-HO-MeIQx) as a major CYP-mediated detoxication product. In the case of PhIP, direct glucuronidation of the N 2 and N 3 positions also occurred in human and rat hepatocytes. Glucuronide and sulfate conjugates of 2-amino-4′-hydroxy-1-methyl-6-phenylimidazo[4,5- b]pyridine (4′-HO-PhIP) were detected as relatively minor metabolites in human hepatocytes but were the major products formed in rat hepatocytes, accounting for up to 50% of the metabolism. Rat CYP1A2, but not the human ortholog, significantly contributes to 4′-hydroxylation of PhIP. Important differences exist between human and rat liver enzymes in catalytic activity and regioselectivity of MeIQx and PhIP metabolism. Some human hepatocyte preparations are more active at transforming MeIQx and PhIP to a genotoxic species than rat hepatocytes pretreated with potent inducer 3-MC. These pronounced interspecies differences in metabolism of MeIQx and PhIP may affect the biological activity of these mutagens and must be considered when assessing human health risk.

Effects of dairy products on heterocyclic aromatic amine-induced rat colon carcinogenesis.

Heterocyclic aromatic amines (HAA) are initiating agents of colon carcinogenesis in animals and are suspected in the aetiology of human colon cancer. In the context of prevention, it seems interesting to test possible protective compounds, such as fermented milk, against HAA food carcinogens. Male F344 rats were used in a model of HAA-induced colon carcinogenesis. The HAA, 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ) and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) (ratio 1:1:1) were administered in food for a 7 week induction period, with a cumulative dose of 250 mg of the HAA, per kg body weight. Four different diets were given to four rat groups: supplemented with 20% water, 30% non-fermented milk, 30% Bifidobacterium animalis DN-173 010 fermented milk and 30% Streptococcus thermophilus DN-001 158 fermented milk. Fecal mutagenicity was quantified during the induction period. At the end of the treatment, DNA lesion levels were determined in the liver and colon using the number of 8-oxo-7,8-dihydro-2'desoxyguanosine (8-oxodGuo) oxidized bases, "3D Test" and comet assay. The metabolic activity of hepatic and colon cytochrome P450 (CYP450) 1A1 and 1A2 was also evaluated. Aberrant colon crypts were scored, 8 weeks after the last HAA treatment. The results showed that dairy products decreased the incidence of aberrant crypts in rats: 66% inhibition with the milk-supplemented diet, 96% inhibition with the B.animalis fermented milk-supplemented diet and 93% inhibition with the S.thermophilus fermented milk-supplemented diet. Intermediate biomarkers showed that there was a decrease in HAA metabolism, fecal mutagenicity and colon DNA lesions. These results demonstrate the early protective effect of milk in the carcinogenesis process. This effect being more pronounced in the case of milk fermented by lactic acid bacteria.

HETEROCYCLIC AROMATIC AMINE METABOLISM, DNA ADDUCT FORMATION, MUTAGENESIS, AND CARCINOGENESIS

Heterocyclic aromatic amines (HAAs) are carcinogenic compounds formed in meats, fish, and poultry prepared under common household cooking practices. Some HAAs are also formed in tobacco smoke condensate. Because of the widespread occurrence of HAAs in these daily staples, health concerns have been raised regarding the potential role of HAAs in the etiology of some human cancers associated with frequent consumption of these products. In this review, the metabolism of HAAs to biologically active metabolites that bind to DNA and provoke mutations and cancer in various biological systems is discussed. Some of the current analytical and molecular methods that are used to measure biomarkers of HAA exposure and genetic damage in experimental animal models and humans are also presented. These biochemical data combined may help to better assess the role that HAAs may have in the development of some common forms of human cancers.

Interspecies differences in metabolism of heterocyclic aromatic amines by rat and human P450 1A2

The catalytic efficiences of cytochrome P450 (P450)-mediated N-oxidation of the 2-amino-3,8-dimethylimidazo[4,5- f]quinoxaline (MeIQx) and 2-amino-1-methyl-6-phenylimidazo[4,5- b]pyridine (PhIP) by recombinant human P450 1A2 were 10–19-fold higher than rat P450 1A2, while methoxyresorufin O-demethylation activity was comparable for both P450s. Similar findings were observed with rat and human liver microsomal samples. Interspecies differences in P450 enzyme expression and catalytic activities may be significant and must be considered when assessing human health risk.

Heterocyclic aromatic amines in human urine following a fried meat meal

In a search for suitable biomarkers for human dietary exposure to heterocyclic aromatic amines (HAAs), we have investigated the concentrations of three common fried food mutagens in food and urine after consumption of a fried meat meal. In this connection we developed a method for the determination of HAAs and have investigated the common fried red meat HAAs 2-amino-l-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), 2-amino-3,8-dimethylimidazo[4,5- b]quinoxaline (MeIQx) and 2-amino-3,8-dimethylimidazo[4,5-ƒ]quinoxaline (DiMeIQx). Eight volunteers participated in the study, each consuming a meal of fried minced beef patties (295 g), boiled potatoes, and a green salad. Urine was collected for two 12-hr periods prior to and following the meal. HAAs were determined in cooked meat and in untreated and acid hydrolysed urine by a series of liquid/liquid extractions, followed by Blue cotton adsorption and finally by a novel derivatization technique for gas chromatography-mass spectrometry (GC-MS). The primary amino groups were derivatized by acylation with heptafluorobutyric acid anhydride, and the resulting amide methylated using diazomethane. Phenolic hydroxyl groups were also methylated by this procedure, making it possible to detect hydroxylated HAAs, possible metabolites or constituents of the fried meat. 4′-Hydroxy-PhIP 2-amino-l-methyl-6-(4-hydroxyphenyl)imidazo[4,5- b]pyridine (4′-OH-PhIP) was indeed found in meat as well as in urine. The contents of PhIP, MeIQx and DiMeIQx in meat were 4.0 ± 2.6, 3.5 ± 0.9 and 0.3 ± 0.1 ng g −1 (mean ± SD, n = 4), from which the mean amounts ingested were calculated to be 1180, 1030 and 90 ng, respectively. Total amounts of HAAs in the 0-24-hr post-meal untreated urine (and percent of ingested dose) were 6–23 ng PhIP (0.5–2%) and 10-63 ng MelQx (1-6%). In hydrolysed urine, the levels of HAAs were higher, totalling 24–100 ng PhIP (2–8.5%) and 133–329 ng of MeIQx (13–32%). DiMeIQx was below detection limit in all urine samples. Judged from our study, there were rather large inter-individual variations in the amounts of excreted HAAs, possibly caused by variations in the activities of enzymes taking part in HAA metabolism.

Interplay between heterocyclic amines in cooked meat and metabolic phenotype in the etiology of colon cancer.

Although the etiology of colon cancer remains uncertain, an increasing body of epidemiologic evidence indicates that red meat consumption is an important risk factor. The cooking of red meat produces a class of potent experimental carcinogens, the heterocyclic aromatic amines (HAA). These induce cancers in several different sites, including the colon, in rats and mice. Other epidemiologic studies indicate that an individual's genetically determined metabolic phenotype (polymorphisms for N-acetyltransferase and N-hydroxylase) modulates the risk of colon cancer. Both N-acetyltransferase and N-hydroxylase are involved in the metabolism of HAA. An increased risk of colon cancer has been observed in rapid acetylators in four of five studies; further, in two of these the association was found only in meat eaters. The latter observation supports the hypothesis that HAA are involved in colon carcinogenesis. Considerable progress has been made in the study of the molecular pathogenesis of colon cancer, which typically entails the cumulation of several genetic events (mutations and deletions) in oncogenes and tumor suppressor genes. It would now be a crucial contribution to elucidating the causation of colon cancer to show that such mutations are induced in human colonic mucosa by food-borne heterocyclic aromatic amines.

Species differences in metabolism of heterocyclic aromatic amines, human exposure, and biomonitoring.

Heterocyclic aromatic amines (HAAs) are animal carcinogens and suspected human carcinogens which are formed in cooked foods at the low parts per billion level. HAAs in cooked meats were purified by either immunoaffinity chromatography or solid phase tandem extraction, which allowed for the simultaneous analysis of 11 HAAs by HPLC. The metabolism of two prominent HAAs, 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) and 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), was investigated in animal models and in vitro with human tissues to develop strategies for human biomonitoring. MeIQx and IQ are rapidly absorbed from the gastrointestinal tract of rodents and transformed into several detoxification products which are excreted in urine and feces. Metabolites result from cytochrome P450-mediated ring oxidation at the C-5 position followed by conjugation to sulfate or beta-glucuronic acid. Other major metabolites include the phase II conjugates, N2-glucuronide and N2-sulfamate. A metastable N2-glucuronide conjugate of the genotoxic metabolite of N-hydroxy-MeIQx was also detected in urine and bile. The binding of both carcinogens to blood proteins was low and suggests that human biomonitoring through protein adducts may be difficult. These metabolic pathways exist in nonhuman primates and several of these pathways also occur in vitro with human liver. The urinary excretion of MeIQx in seven human subjects following consumption of cooked beef or fish ranged between 2 and 22 ng in 12 hr when determined by negative ion chemical ionization GC-MS. After acid hydrolysis of urine, the amount of MeIQx increased 4- to 10-fold in 6 of the 7 subjects.(ABSTRACT TRUNCATED AT 250 WORDS)

Species Differences in Metabolism of Heterocyclic Aromatic Amines, Human Exposure, and Biomonitoring

Species Differences in Metabolism of Heterocyclic Aromatic Amines, Human Exposure, and Biomonitoring

Mammalian cell mutagenicity and metabolism of heterocyclic aromatic amines

Heterocyclic aromatic amines are bacterial mutagens which also induce DNA damage in mammalian cells. Damage has been demonstrated using a number of endpoints, including gene mutation, chromosome aberrations, sister-chromatid exchange, DNA-strand breaks, DNA repair and oncogene activation. Although the responses in mammalian cells are weak when compared to bacterial mutagenicity, heterocyclic aromatic amines are rodent carcinogens. Metabolic N-oxidation by cytochrome P450 is an initial activation step with subsequent transformation of the N-hydroxy metabolites to the ultimate mutagenic species by O-acetyltransferase or sulfotransferase. Major routes of detoxification include cytochrome P450-mediated ring oxidation followed by conjugation to glucuronic or sulfuric acid. Direct conjugation to the exocyclic amine group also occurs. Major reactions include N-glucuronidation and sulfamate formation.

Prostaglandin H synthase catalyzed metabolism of heterocyclic aromatic amines of the IQ-type and their activation to mutagens

Prostaglandin H synthase catalyzed metabolism of heterocyclic aromatic amines of the IQ-type and their activation to mutagens

Mutagenicity and carcinogenicity of extracts from airborne particles

In a search for suitable biomarkers for human dietary exposure to heterocyclic aromatic amines (HAAs), we have investigated the concentrations of three common fried food mutagens in food and urine after consumption of a fried meat meal. In this connection we developed a method for the determination of HAAs and have investigated the common fried red meat HAAs 2-amino-l-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), 2-amino-3,8-dimethylimidazo[4,5-b]quinoxaline (MeIQx) and 2-amino-3,8-dimethylimidazo[4,5-ƒ]quinoxaline (DiMeIQx). Eight volunteers participated in the study, each consuming a meal of fried minced beef patties (295 g), boiled potatoes, and a green salad. Urine was collected for two 12-hr periods prior to and following the meal. HAAs were determined in cooked meat and in untreated and acid hydrolysed urine by a series of liquid/liquid extractions, followed by Blue cotton adsorption and finally by a novel derivatization technique for gas chromatography-mass spectrometry (GC-MS). The primary amino groups were derivatized by acylation with heptafluorobutyric acid anhydride, and the resulting amide methylated using diazomethane. Phenolic hydroxyl groups were also methylated by this procedure, making it possible to detect hydroxylated HAAs, possible metabolites or constituents of the fried meat. 4′-Hydroxy-PhIP 2-amino-l-methyl-6-(4-hydroxyphenyl)imidazo[4,5-b]pyridine (4′-OH-PhIP) was indeed found in meat as well as in urine. The contents of PhIP, MeIQx and DiMeIQx in meat were 4.0 ± 2.6, 3.5 ± 0.9 and 0.3 ± 0.1 ng g−1 (mean ± SD, n = 4), from which the mean amounts ingested were calculated to be 1180, 1030 and 90 ng, respectively. Total amounts of HAAs in the 0-24-hr post-meal untreated urine (and percent of ingested dose) were 6–23 ng PhIP (0.5–2%) and 10-63 ng MelQx (1-6%). In hydrolysed urine, the levels of HAAs were higher, totalling 24–100 ng PhIP (2–8.5%) and 133–329 ng of MeIQx (13–32%). DiMeIQx was below detection limit in all urine samples. Judged from our study, there were rather large inter-individual variations in the amounts of excreted HAAs, possibly caused by variations in the activities of enzymes taking part in HAA metabolism.