Absence of PhIP adducts, p53 and Apc mutations, in rats fed a cooked beef diet containing a high level of heterocyclic amines

Abstract

Meat cooked at high temperatures contains mutagens and carcinogens known as heterocyclic amines (HCA). Cooking temperature and time determine the amount of HCA produced. The present study examined the DNA of liver, colon, and stomach from rats fed a high level of HCA for 27 weeks. Male Sprague‐Dawley rats were fed a high‐fat AIN‐76A‐based diet containing 60% by weight cooked beef containing a high level of HCA, especially 2‐amino‐l‐methyl‐6‐phenylimidazo[4,5‐b]pyridine (PhIP, 72 ng/g cooked beef), the most abundant HCA in cooked meat products. At the end of 27 weeks the rats were terminated, and small portions of liver, colon, and stomach were quick‐frozen in liquid nitrogen. The DNA was isolated from the thawed tissue by phenol‐chloroform extraction, and the genomic DNA was analyzed for the presence of PhIP adducts by ³²P‐postla‐beling analysis. The DNA was also used in polymerase chain reactions to amplify the rat p53 and Apc genes, then direct dye‐terminator DNA sequencing was carried out. Results showed no PhIP adducts in any tissue. In addition, no signature p53 or Apc gene mutations were seen in colon or stomach DNA. These results indicate that the high level of HCA present in a diet of well‐cooked meat does not cause 1) persistent PhIP adducts similar to those produced by feeding pure PhIP at high doses or 2) p53 and Ape gene mutations in nontumor tissue.