Mutagenic and carcinogenic heterocyclic amines in Chinese cooked foods

Samples of 7 foods commonly eaten in the Northeast of China (i.e. fried and broiled fishes and broiled meat) were tested for mutagenicity on Salmonella typhimurium TA98 with S9 mix. The basic fractions of the samples were mutagenic, inducing 33-2930 revertants/g of cooked food. Fried walleye pollack (a kind of cod fish heated on a stainless steel pan) showed the highest mutagenicity, so attempts were made to isolate mutagens from the basic fraction of this food. The mutagens were purified by treatment with blue cotton and HPLC on a semi-preparative ODS column and analytical cation exchange and ODS columns. 5 mutagens were isolated and identified as 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ), 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx), 2-amino-3,4,8-trimethylimidazo[4,5-f]quinoxaline (4,8-DiMeIQx) and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP). 1 g of fried fish was estimated to contain 0.16 ng of IQ, 0.03 ng of MeIQ, 6.44 ng of MeIQx, 0.10 ng of 4,8-DiMeIQx and 69.2 ng of PhIP. MeIQx and PhIP accounted for 24% and 4.7%, respectively, of the total mutagenicity. The other 3 heterocyclic amines were each responsible for only 0.3-1.2% of the total mutagenicity.     

Formation of 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline in a model system by heating creatinine, glycine and glucose

A mixture of creatinine, glucose and glycine was heated in diethylene glycol containing 14% water for 2 h at 128 degrees C, and the mutagens formed were purified by XAD-2 column chromatography, acid-base partition, Sephadex LH-20 column chromatography, 'blue cotton' treatment and HPLC. Two mutagenic substances were isolated by HPLC. The major mutagen was identified by its UV absorption and mass and NMR spectra as 2-amino-3,8- dimethylimidazo [4,5-f]quinoxaline, which was originally isolated from fried beef. This finding supported the idea that creatinine, amino acids and sugars present in meat are precursors in the formation of the mutagenic imidazoquinoxaline derivative.     

Metabolic activation of food mutagens by liver and lung enzymes from rats, pigs and oxen

Mutagenic activation of the 3 cooked food mutagens 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ) and 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) was compared in liver and lung enzyme preparations from oxen, pigs and rats. Liver preparations from oxen were the most efficient in activating the mutagens, while the rat enzymes were more active than those from pigs. The different cooking mutagens showed different mutagenic potential. MeIQ was the most potent mutagen, followed by IQ and MeIQx in descending order. In oxen, MeIQx was as potent as IQ. The activation with the lung enzymes was 2-3 orders of magnitude lower than with liver. Furthermore, species differences in mutagenic activation with lung enzymes were small compared with liver enzymes. In lung preparations the differences between IQ and MeIQ were small, but in all 3 animal species the mutagenicity of MeIQx was 1 order of magnitude lower than that of the other 2 mutagens.     

Formation of new heterocyclic amine mutagens by heating creatinine, alanine, threonine and glucose.

A mixture of alanine, threonine, creatinine and glucose was heated in diethylene glycol and water (5:1, v/v) for 15 min at 200 degrees C. The mutagens formed were purified by high-performance liquid chromatography using the Ames/Salmonella mutagenic activity to guide the purification. The structures of the purified mutagens were determined using UV absorption, mass and NMR spectrometry. A new mutagenic compound with a mass number of 217 was found and its mass spectrum did not correspond to any known mutagen derived from food. This new compound accounted for 4% of the total mutagenic activity. Other mutagenic compounds were identified as MeIQx (2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline), 4,8-DiMeIQx (2-amino-3,4,8-trimethylimidazo[4,5-f]quinoxaline), and a new mutagen 4,7,8-TriMeIQx (2-amino-3,4,7,8-tetramethylimidazo[4,5-f]quinoxaline) with a mutagenic activity of 73,000 TA98 revertants per microgram. The percentage of the mutagenic activity attributable to MeIQx, 4,8-DiMeIQx and 4,7,8-TriMeIQx was 10%, 70% and 3%, respectively. The yield of MeIQx, 4,8-DiMeIQx and 4,7,8-TriMeIQx was 10, 36 and 6 nmole/mmole creatinine. The formation of TriMeIQx from natural meat components suggests that this new quinoxaline mutagen may be present in cooked foods.     

Mutagenic potency of heterocyclic amines towards Salmonella typhimurium; possible causes of variability in the results observed.

The potent food mutagens and carcinogens 2-amino-3-methylimidazol[4,5-f]quinoline (IQ), 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MEIQ) and 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) are probably the most active bacterial mutagens so far discovered. Important discrepancies were found, however, in the specific mutagenicities published for these compounds. This paper analyzes a number of experimental factors that could explain these differences: purity of the compounds, stability under the experimental conditions employed, solvents used, bacterial toxicity, testing procedure, amount and age of the S9 fraction, dose-effect relationships, day-to-day variability, origin of the compounds investigated or of the bacterial strain and age of the strain culture used. None of these factors was found to play a critical role, when the other experimental conditions were strictly standardized. The in-house testing procedure used probably explains the interlaboratory variations observed.     

Activation of the food mutagens IQ and MeIQ by hepatic S9 fractions derived from various species

The ability of hepatic S9 mixes derived from different rodent species (rat, mouse, Syrian and Chinese hamster) to activate the mutagens 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) and 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ) was investigated using Salmonella typhimurium strain TA98. In general, the mutagenicity of IQ and MeIQ was greatest in the presence of S9 fractions from Swiss albino mice and least from fractions derived from Chinese hamsters. However, treatment of rats or hamsters with Aroclor 1254 had little or no effect on the activation of IQ or MeIQ to mutagens.     

Inhibitory effect of dibenzoylmethane on mutagenicity of food-derived heterocyclic amine mutagens

Dibenzoylmethane (DBM), a structural analogue of curcumin (a bioactive phytochemical present in a widely used spice turmeric) was screened for its inhibitory effect against seven cooked food mutagens (heterocyclic amines): 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ), 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx), 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1), 3-amino-1-methyl-5H-pyrido[4,3-b]indole (Trp-P-2), 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and 2-amino-6-methyldipyrido[1,2-a:3',2'-d]imidazole (Glu-P-1), in both TA98 and TA100 strains of Salmonella typhimurium using Ames Salmonella/reversion assay in the presence of Aroclor1254-induced rat liver S9 homogenate. DBM has been reported to antagonize the mutagenicity of several chemical carcinogens in vitro and has recently been shown to be even more effective than curcumin in suppressing the 7,12-dimethylbenz[a]anthracene (DMBA)-induced mammary tumors in rats. But there are no reports regarding its antimutagenic properties against cooked food mutagens. Results of the present investigations clearly indicate that dibenzoylmethane is a very potent antimutagenic agent, that could effectively inhibit mutagenicity induced by all the tested cooked food mutagens in both the frame shift (TA98) as well as the base pair mutation sensitive (TA100) strains of S. typhimurium. These highly potent inhibitory effects of dibenzoylmethane against heterocyclic amines observed in our preliminary investigations strongly warrant further studies of its efficacy as a cancer chemopreventive agent.     

Formation of mutagens, 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) and 2-amino-3,4,8-trimethylimidazo[4,5-f]quinoxaline (4,8-DiMeIQx), in heated fish meats

Fish meats were heated under conditions close to those used for cooking and processing. The mutagenic activity of the heated fish meats was estimated toward Salmonella typhimurium TA98 with metabolic activation after extraction with boiling water and adsorption to blue cotton. The numbers of His+ revertant colonies/5 g of the meat heat-dried without charring at 220 degrees C for 15 min were about 3000 for bonito, about 1000 for tunny, less than 500 for mackerel, salmon, swordfish, sardine, horse mackerel and cod, and 0 for cuttlefish. The mutagens in the heat-dried bonito meat were purified by thin-layer chromatography (TLC) and high-performance liquid chromatography (HPLC). They were identified as 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) and 2-amino-3,4,8-trimethylimidazo[4,5-f]quinoxaline (4,8-DiMeIQx) by comparison with the authentic specimen with respect to Rf values in TLC, retention times in HPLC, ultraviolet absorption spectra and mass spectra. The contents of MeIQx and 4,8-DiMeIQx in the bonito meat were estimated to be 5.2 and 5.4 ng/g, respectively. The major mutagens produced in the bonito, tunny and mackerel meats heated without charring at 100 degrees C for 48 h and at 220 degrees C for 15 min were found to be MeIQx and 4,8-DiMeIQx. It is interesting to note that the bonito and sardine meats grilled with charring for 15 min contained MeIQx and 4,8-DiMeIQx but higher mutagenicity was observed in the fraction that may contain 2-amino-3-methylimidazo[4,5-f] quinoline (IQ), 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ) and/or 2-aminodipyrido[1,2-a: 3',2'-d]imidazole (Glu-P-2).     

Determination of heterocyclic aromatic amines (HAs) content in samples of household-prepared meat dishes

Aminoazaarene content was investigated in 10 meat samples (including pork, beef, turey and chicken) thermally processed at home according to common recipes used by residents of Upper Silesia region in Poland. The clean-up procedure included tandem solid-phase extraction (SPE) using Extrelut-type columns filled with diatomaceous earth, propylsulphonic acid and chemically bounded phase-C18. Identification and quantitative analysis of HAs fraction was carried out using a HPLC system with DAD-type detector. Separation was achieved using TSK-gel ODS 80-TM column and a mixture of 5% acetonitrile and 95% triethylamine phosphate buffer (pH 3.3) as a mobile phase. The results of qualitative determinations were confirmed by GC-MS method. To achieve this, HAs fractions were derivatized to pentafluoropropionic acid (PFPA) amide derivatives. The summary content of five aminoazaarenes determined in investigated meat samples, i.e. 2-amino-3-methylimidazo [4,5-f]quinoline (IQ), 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx), 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ), 2-amino-3,4,8-trimethylimidazo[4,5-f]quinoxaline (4,8-DiMeIQx), 2-amino-1-methyl-6-phenyl-imidazo[4,5-b]pyridine (PhIP) falls within the range of 1.9-77.4 ng/g of sample. The calculated values of theoretically daily human exposure to five determined HAs were in the range of 0.2-7.7 microg per day per person.     

Inhibition of mutagenicity of food-derived heterocyclic amines by sulforaphane--a constituent of broccoli

Sulforaphane, a constituent of broccoli was investigated for its antimutagenic potential against different classes of cooked food mutagens (heterocyclic amines). These include imidazoazaarenes such as 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ), 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP); pyridoindole derivatives such as 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1) and 3-amino-1-methyl-5H-pyrido[4,3-b]indole (Trp-P-2); and, dipyridoimidazole derivative such as 2-amino-6-methyldipyrido[1,2-a:3',2'-d]imidazole (Glu-P-1). Tests were carried out by Ames Salmonella/reversion assay using Salmonella typhimurium TA98 (frame shift mutation sensitive) and TA100 (base pair mutation sensitive) bacterial strains in the presence of Aroclor 1254-induced rat liver S9. Results of these in vitro antimutagenicity studies strongly suggest that sulforaphane is a potent inhibitor of the mutagenicity induced by imidazoazaarenes such as IQ, MeIQ and MeIQx (approximately 60% inhibition) and moderately active against pyridoindole derivatives such as Trp-P-1 and Trp-P-2 (32-48% inhibition), but ineffective against dipyridoimidazole derivative (Glu-P-1) in TA 100.     

Fecal mutagenicity arising from ingestion of fried ground beef in the human

Fried ground beef has been shown to contain mutagens, and the major mutagenic component has been identified as 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx). Mutagens in feces of 3 adult volunteers were fractionated by treatment of the feces with blue cotton followed by chromatography on a carboxymethyl cellulose column. The chromatographic fraction, corresponding to MeIQx in terms of the position of elution, was examined for mutagenicity in S. typhimurium TA98 with metabolic activation. When meals containing no heated meat were eaten, this fraction of feces showed little or no mutagenicity. On eating fried ground beef, the feces excreted in the next two days showed greatly increased mutagenicity in this fraction. By eating no-meat meal subsequent to the meat meal, the mutagenicity resumed the original low level on the fourth day after the meat meal. The components in the mutagenic fraction were probably metabolites of the mutagens present in cooked meat, since analysis by high pressure liquid chromatography of the mutagenic fraction showed that the active components in the feces were different from the mutagens in cooked meat.     

Inhibitory effect of curcumin and its natural analogues on genotoxicity of heterocyclic amines from cooked food

Curcumin (C) and its natural analogues demethoxycurcumin (dmC) and bisdemethoxycurcumin (bdmC), known for their potent anti-inflammatory, antioxidant, antimutagenic and anticarcinogenic effects, were tested for their possible inhibitory effects against seven cooked food mutagens (heterocyclic amines): 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ), 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx), 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1), 3-amino-1-methyl-5H-pyrido[4,3-b]indole (Trp-P-2), 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) and 2-amino-6-methyldipyrido[1,2-a:3',2'-d]imidazole (Glu-P-1), in both TA98 and TA100 strains of Salmonella typhimurium using Ames Salmonella/reversion assay in the presence of Aroclor induced rat liver S9 homogenate. In the present investigations, curcumin as well as its two natural analogues i.e., dmC and bdmC were found to be highly effective in suppressing genotoxicity of all the tested cooked food mutagens in a dose-dependent manner, in both the frame shift (TA98) as well as base pair mutation sensitive (TA100) strains of S. typhimurium. However, bdmC appeared to be a relatively less active antimutagen compared to C and dmC. More than 80% inhibition of mutagenicity was observed at 200 microg/plate in case of C and dmC in both TA98 and TA100 against all tested cooked food mutagens. Where as, bdmC showed 39-79% inhibition in TA100 and 60-80% inhibition in TA98, at a dose of 200 microg/plate. These findings warrant further biochemical, enzymatic and in vivo investigations in animal models as well as in humans to establish the chemoprotective effect of these agents against mutagenic heterocyclic amines found in cooked food.     

Mutagenicity of food pellets from human diets in The Netherlands

Food pellets from human diets, prepared according to mean consumption figures in The Netherlands, were assessed on mutagenicity and mutagens were identified. Three types of human meals were compared: raw (C), heated (D) and heated with vegetables and fruit (E, a complete meal). In addition 2 animal diets were tested: commercial control diet (A), and a control diet to which vegetables and fruit had been added (B). All human diets contained: 40.6 energy (E)% fat, 13.2 E% protein, 46.2 E% carbohydrate and 5.2% (w/w) fibre. For animal diets these figures were 21.6, 26.0, 52.4 and 10.7% respectively. After extraction samples were tested in the Salmonella-microsome test, tester strains TA1538, TA98 and TA100. Human diets with heated products (D, E) were both clearly mutagenic with approximately 300-500 revertants per gram. Food pellets from animal diets (A, B) displayed no mutagenic activity. HPLC-derived chromatographic fractions of diets D and E showed 3 large mutagenic areas identified as IQ (2-amino-3 methyl-imidazo-[4,5-f]quinoline) and MeIQx (2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline, DiMeIQx (2-amino-3,4,8-trimethylimidazo[4,5-f]quinoxaline and PhIP (2-amino-6-phenylimidazo[4,5-b]pyridine) and other mutagens not completely defined. This mutagen profile was similar to that found previously for fried beef. Mass estimates for these potent mutagens amounted to 15-20 micrograms/kg. Health implications of these findings are discussed. As IQ, MeIOx and DiMeIQx have been found to be weakly carcinogenic in rodents and many other initiating and modulating factors may be present in a complex human diet, a chronic toxicity study is indicated.     

Mutagenic activation of 2-amino-3-methylimidazo[4,5-f]-quinoline (IQ) and 2-amino-3,4-dimethylimidazo[4,5-f]-quinoline (MeIQ) by subcellular fractions and cells isolated from small intestine,kidney and liver of the rat

The mutagenic activity of the pyrolysis products 2-amino-3-methylimidazo[4,5-f]-quinoline and 2-amino-3,4-dimethylimidazo[4,5-f]-quinoline in Salmonella typhimurium TA98 using rat intestinal and renal subcellular fractions as activation systems was approximately 1 and 5 revertants per nmol, respectively. This was 1,000 times less than the activity with a subcellular fraction from rat liver. The mutagenic activity of both compounds was considerably increased using intestinal, renal and hepatic preparations isolated from PCB (Aroclor 1254)-pretreated rats, compared to preparations from control animals. In addition, both compounds displayed a moderate direct-acting mutagenic activity at concentrations above 10^-5 M. Isolated cells from small intestine, kidney and liver incubated in nucleopore chambers were able to convert both compounds into products which mutated bacteria outside the chambers. The concentrations of chemicals required to yield responses of a similar magnitude were approximately 3 orders of magnitude higher in the intestinal and renal systems compared to the hepatic system. The formation of metabolites mutagenic for Salmonella typhimurium by hepatic subcellular and cellular systems was shown to be superior to the respective intestinal and renal systems.Abbreviations AHH arylhydrocarbon hydroxylase - IQ 2-amino-3-methylimidazo[4,5-f]-quinoline - MC 3-methylcholanthrene - MeIQ 2-amino-3,4-dimethylimidazo[4,5-f]-quinoline - PCB polychlorinated biphenyls (Aroclor 1254) - S9 the 9,000 g supernatant tissue fraction - TCDD 2,3,7,8-tetrachlorodibenzo-p-dioxin     

DNA-biding of IQ, Me-IQ and Me-IQx, strong mutagens found in broiled foods

Non-covalent DNA-binding has been studied of 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (Me-IQ) and 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (Me-IQx), strong mutagens found in broiled foods. These mutagens are intercalated into DNA, as found by ultraviolet absorption gel electrophoresis. The binding of IQ is stronger with GC pairs than AT pairs in DNA. The binding constants with calf thymus DNA are 1.6 × 10⁶ (Me-IQ), 0.9 × 10⁶ (IQ) and 0.7 × 10⁶ M⁻¹ (Me-IQx) at pH 6.0. This order of DNA affinity agrees with the order of mutagenicity towards Salmonella typhimurium TA98.     

Comparative genotoxic effects of IQ and MeIQ in Salmonella typhimurium and cultured mammalian cells

The food mutagens 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) and 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ) were studied for their genotoxic potential using hepatocytes isolated from untreated and Aroclor 1254 (PCB) pretreated rats as an activation system. Monolayers of hepatocytes co-incubated with Salmonella typhimurium TA98 activated IQ and MeIQ to bacterial mutagens, with MeIQ being about twice as potent as IQ. The mutagenic activities of IQ and MeIQ were increased by using hepatocytes from PCB-pretreated rats. IQ and MeIQ also caused primary DNA damage in the hepatocytes as determined by increases in the rate of alkaline elution of DNA, as well as increases in DNA-repair synthesis. Furthermore, exposure of V79 cells co-cultured with PCB-pretreated hepatocytes to IQ and MeIQ showed evidence of increased sister-chromatid exchanges and a low and variable increase in the number of 6-thioguanine-resistant mutants. The genotoxic potency of IQ and MeIQ in mammalian cells was low or virtually absent compared to their extreme potency in bacteria. This could be due to a lower capacity of mammalian cells to further metabolize the so-called directly acting bacterial mutagens, formed by a cytochrome P-450 dependent N-hydroxylation, to their ultimate reactive forms.     

Mutagenicity of some heterocyclic amines in Salmonella typhimurium with metabolic activation by human red blood cell cytosol.

Purified human red blood cell cytosol was used to activate the heterocyclic amines 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ), 3-amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1) and 3-amino-1-methyl-5H-pyrido[4,3-b]indole (Trp-P-2) into mutagenic intermediate(s) in the Salmonella test. The liquid preincubation method in the presence of strain TA98 was utilized. In order to understand the mechanism involved in this metabolic activation, some modulators were incorporated in the medium. The results suggest that an oxygenated hemoprotein, probably oxyhemoglobin, is involved in the activation into genotoxic intermediate(s).     

Mutagenic and carcinogenic heterocyclic amines as affected by muscle types/skin and cooking in pan-roasted mackerel

The dependence on muscle types/skin and on degrees of cooking in the formation of heterocyclic amines (HCAs) of pan-roasted mackerel was studied. High levels of 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) were found in very well done skin and ordinary muscle, being 4.2 and 5.3 ng/g, followed by 2-amino-3,4-dimethylimidazo[4,5-f] quinoxaline (MeIQx), being 1.8 and 2.1 ng/g and 2-amino-9H-pyrido[2,3-b]indole (AalphaC), being 1.2 and 2.8 ng/g, respectively. In pan-roasted mackerel, ordinary muscles contributed much more greatly to the formation of HCAs than skins due to its higher HCA contents and composition (76.1%).     

Binding of mutagenic pyrolyzates to fractions of intestinal bacterial cells.

The binding of mutagenic pyrolyzates to cell fractions from some gram-negative intestinal bacteria and to thermally treated bacterial cells was investigated. 3-Amino-1,4-dimethyl-5H-pyrido[4,3-b]indole (Trp-P-1) and 3-amino-1-methyl-5H-pyrido[4,3-b]indole (Trp-P-2) were effectively bound by several of the bacterial cells. The cell wall skeletons of all bacteria effectively bound Trp-P-1 and Trp-P-2. Their cytoplasmic fractions retained Trp-P-1 and Trp-P-2, but to a lesser extent than the cell wall skeletons. 2-Amino-3-methylimidazo[4,5-f]quinoline (IQ) was not found in their cytoplasmic fractions. These cell wall skeletons also bound 2-amino-6-methyldipyrido[1,2-a:3',2'-d]imidazole (Glu-P-1), 2-amino-5-phenylpyridine (Phe-P-1), IQ, 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ), and 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQX). The amount of each mutagen bound differed with the type of mutagen and the bacterial strain used. The outer membrane of Escherichia coli IFO 14249 showed binding of about 123.7 micrograms/mg of Trp-P-2, and its cytoplasmic membrane bound 57.14 micrograms/mg. Trp-P-2 bound to the bacterial cells was extracted with ammonia (5%), methanol, and ethanol but not with water.     

Mutagenic activity and heterocyclic amine carcinogens in commercial pet foods

Twenty-five commercial pet foods were analyzed for mutagenic activity using the Ames/Salmonella test with strain TA98 and added metabolic activation. All but one gave a positive mutagenic response. Fourteen of these samples were analyzed for heterocyclic amine mutagens/carcinogens and all but one contained 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) and 10 of 14 contained 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) as analyzed by HPLC and confirmed by photodiode array peak matching. From these findings it is hypothesized that there is a connection between dietary heterocyclic amines and cancer in animals consuming these foods.