A in hydrolysates of hepatic DNA could be the hepatocarcinogen NDHU. A plausible supply of human exposure to NDHU is endogenous nitrosation of dihydrouracil (DHU, 2, Scheme 1). Endogenous nitrosation can take place in humans when salivary nitrite reacts with amines or amides ingested inside the diet plan or present within the physique [15]. This has been clearly demonstrated in studies which showed important increases in N-nitrosoproline in human urine following exposure to nitrite and proline [15]. DHU can be a standard intermediate in mammalian metabolism, formed from uracil (1) by dihydropyrimidine dehydrogenase [16;17]. DHU is present in human plasma [17;18] and urine [16?9]. Levels of DHU excreted inside the urine of healthy humans happen to be reported to range from about 2 ?10 mg/ day. Moreover, DHU is metabolized to ?-ureidopropionic acid (?-UPA, three), that is located at related levels as DHU in urine [16;19], and could also be nitrosated, giving rise to N-?-UPA (5). Therefore, both DHU and ?-UPA could possibly be converted towards the 2carboxyethyldiazonium ion upon nitrosation (Scheme 1). In this study, we tested the hypothesis that 7-CEGua levels in hydrolysates of rat liver DNA could improve upon treatment of rats with NaNO2 plus DHU or ?-UPA. If this hypothesisNIH-PA Author Manuscript NIH-PA Author Manuscript NIH-PA Author ManuscriptChem Biol Interact. Author manuscript; readily available in PMC 2014 October 25.Wang et al.Pagewere right, then endogenous nitrosation of DHU to NDHU, or ?-UPA to N-?-UPA, might be a source of 7-CEGua in human liver DNA, and this process could potentially be involved in liver cancer etiology. We also examined the possibility that 7-CEGua adducts in hydrolysates of hepatic DNA might result from remedy of rats with acrylic acid, which reacts slowly with DNA in vitro, resulting in formation of 7-CEGua [20].NIH-PA Author Manuscript NIH-PA Author Manuscript NIH-PA Author Manuscript2. Components and Methods2.1 Chemical compounds DHU, NaNO2, and acrylic acid had been purchased from Sigma-Aldrich (St. Louis, MO). ?UPA was obtained from Chem-Impex International Inc (Wooddale, IL). two.two Animal study This study was authorized by the University of Minnesota Institutional Animal Care and Use Committee. Male F-344 rats, age six weeks, have been obtained from Charles River Laboratories (Kingston, NY), housed two per cage with Harlan irradiated corncob bedding (Harlan, Indianapolis IN), and allowed to acclimate towards the Research Animal Sources facility, University of Minnesota, for 1 week. The rats were maintained under regular situations (20?4 temperature, 29?two relative humidity and 14/10 light/dark cycle). They have been fed Harlan Teklad 7022 (NIH-07) diet program. Water and diet consumption were measured three times per week.6-Chloro-5H-benzo[a]phenoxazin-5-one structure In Study 1, four rats per group, except as noted, had been treated for 2 weeks as follows: 1, NaNO2 1500 ppm in drinking water; 2, DHU 2480 ppm in powdered diet; 3, NaNO2 1500 ppm + DHU 2480 ppm; four, ?-UPA 2870 ppm in powdered eating plan; five, NaNO2 1500 ppm + ?-UPA 2870 ppm; six, acrylic acid 1565 ppm in drinking water, 6 rats; 7, untreated manage, six rats.2-(4-Nitrophenyl)-2-oxoacetic acid manufacturer The rats were humanely sacrificed by CO2 overdose, and tissues were collected and frozen at -20 until analysis.PMID:23891445 In Study two, four rats per remedy group have been given the compounds for four weeks specifically as in Study 1, except that 1 further group was added having a greater dose of acrylic acid (3130 ppm). There have been 6 rats in the control group. The rats were sacrificed and tissues collected and frozen as in Study 1. 2.three Analysis of rat he.