Dietary Lipid-dependent Activation of the Carcinogen N-2- Fluorenylacetamide in Rats as Monitored by Salmonella ty1

Salmonella typhlmurlum TA1538 has been used as an analyticaltoolto quantifythe abilityof tissuehomogenate supernatantsto activate the carcinogenN-2-fluorenyla cetamide (FAA) and the proximate carcinogen N-hydroxy 2-fluorenylacetamide (N-OH-FAA). Tissue homogenate supernatantswere obtained from male Sprague-Dawley rats fed a basal dit supplementedwith 5 or 20% hoef tallowor cornoil for 6 monthsand given l.p. Injectionsof FAA(10 mg/100 g bodyweight)or of the controlvehicle. After only 21 days of feeding and I injectionof FAAor control, elevated activity of the liver microsomal-free supernatant(5-150) enzyme that converts N-OH-FAAto mutagen was dependent upon the kind and level of dietary fat. Enzymeactivityfrom FAA-treatedrats fed 5% cornoil was 3-fold higherthan ft was in FAA-treatedanimalsfed 5% tallow, whereas in FAA-treated animals fed 20% corn oil enzyme, activitywas 2-fold higherthan it was in rats fed 20% tallow. After6 monthsof feedingand 7 i.p. injectionsof FAAor control vehicle, the mutagen-producingpotential of the liver 5-150 was increased, regardless of the dit. How ever, FAA injections plus 20% corn oil or beef tallow promotedthe greatestenzymeactivity,suggestinga syn ergisticeffct of highfat and FAA.The abilityof 5-150 to activate potentially mutagenic compoundswas 22-fold higher in FAA-treatedrats fed 20% corn oil than in FAA treatedanimalsfed 20%beef tallow. Between21daysand 6 months, hepatic enzyme activity increased 2-fold for rats fed 5% beef tallowor cornoil, 5-foldfor rats fed 20% beef tallow, and 52-fold for animals fed 20% corn oil. Lungand kidney5-150 enzymeactivitieswere assayed with the bacterial system after 6 months.Dietary fat did not affect the activity of the lung, and aithough kidney enzyme activity was increased from 1to 3-fold it was less responsiveto diet than liver. After 21 days and 1 FAA injection, rats had a greater concentrationof urinaryN-OH-FAAglucuronldewhen fed cornoil than whenfed tallow. After6 months,the urineof rats fed 20%cornoil and given 7 inJectionsof FAAexhib ited the greatestamountof urinaryN-OH-FAA. 1 Supported in part by a Grant-in-aid from Continental Oil Company to Charles R. Shaw, the Research Institute of Mechanisms of Carcinogen Action, the Research Institute of Food Science and Nutrition at Texas Tech University, and Eli Lilly and Co. 2 Submitted for partial fulfillment of dissertation, Department of Food and Nutrition, Texas Tech University. 3 To whom requests for reprints should be addressed. Received July 8, 1977; accepted June 2, 1978. INTRODUCTION Since the early experiments of Watson and Mellanby (37), which demonstrated that feeding mice a diet containing 12.5 to 25% butter fat caused an increased incidence of skin tumors produced by tarring, there has been continued interest in the relationship between dietary fat and tumori genesis (7, 14, 34-36). Milleretal. (18, 19, 29) demonstrated in early experiments that the type as well as the level of dietary fat is an important factor in determination of hepatic tumorigenesis induced by p-dimethylaminoazobenzene. These studies (18, 19, 28) showed that rats receiving p dimethylaminoazobenzene in a diet supplemented with 5 or 20% corn oil developed more hepatic tumors than did those on otherwise similar diets that contained a saturated fat or fatty acid. In addition, 1 of these studies (19) showed that increasing the level of corn oil from 5 to 20% substantially increased tumorigenesis. These and many other experi ments indicate that dietary fat increases the risk of certain spontaneous and induced tumors in animals (6, 18, 19, 29, 31) and that dietary fat is highly correlated with certain tumors in human populations (7, 20, 39â€”40). Salmonella typhimurium TA1538(2, 4) has been shown to be useful in the screening of large numbers of mutagens and carcinogens (23, 24). Ninety % of the chemical carcin ogens tested are mutagenic or may be metabolically acti vated to mutagens. Ames et a! (1) have demonstrated that a large group of carcinogens, while having little or no mutagenic activity per se , were converted to active muta gens after incubation with human or rat liver homogenates. One of these compounds, FAA4, is hydroxybated by the mixed-function oxidase system in the microsomal fraction of the cell to N-OH-FAA, which is more carcinogenic than the parent compound (10, 26, 27). In rat liver N-OH-FAA is converted to its sulfate ester (11, 17). This appears to be a major carcinogenic metabobite of N-OH-FAA, since hepatic sulfotransferase activity correlates well with carcinogenic activity in liver (11, 13). In a review of chemical carcinogens, Miller and Miller (25) described other metabolic reactions that convert N-OH-FAA to electrophilic reactants potentially capable of interacting with nucleic acids and/or protein. The ebectrophilic reactants include N-acetoxy-N-2-fbuoren ylacetamide, 2-nitrosofluorene, N-acetoxy-2-fluorenamine and N-O-glucuronate esters of N-OH-FAA. FAA may be further metabolized by a deacetylase to N-hydroxy-2-flu orenamine, the mutagen considered by Stout et al. (32, 33) to be the ultimate carcinogen. 4 The abbreviations used are: FAA, N-2-fluorenylacetamide; N-OH-FAA, Nhydroxy-2-fluorenylacetamide; 5-9, 9,000 x g supernatant of a tissue homog enate;8-150,150,000x g supernatantof a tissuehomogenate. 2836 CANCERRESEARCHVOL. 38 Research. on September 11, 2017. © 1978 American Association for Cancer cancerres.aacrjournals. org Downloaded from