Rodent metabolism studies

In rodent metabolism studies both parent compound and metabolite were detected in feces and metabolites were detected in urine using GC/LRMS. HRGC/LRMS has also been used successfully in determination of CDDs in rat feces (Abraham et al. 1989a). Adequate comparisons of sensitivity, accuracy, and precision cannot be made because of the lack of these data for several methods and the differences in the media and analytes for the available data. 

Among the requirements for registration of pesticides in the United States, the 1978 guidelines proposed by the U.S. Environmental Protection Agency (3) list general metabolism studies "in at least one mammalian species, preferably the laboratory rat. .." Although similar tests have been conducted on other terrestrial species with increasing frequency, the small rodents have remained the principal source of metabolism data from intact animals. 

According to the EPA guidelines mentioned above, metabolism studies (in rodents) have the following purposes ... 

In summary, although there have been contradictory reports in related literature, as well as discrepancies in data from humans and rodents, recent studies point to a higher nicotine metabolism in females than males. 

Chronic inhalation studies in rodents found no increase in tumors. The lARC has determined that there is evidence suggesting lack of carcinogenicity of DMF in animals. Metabolic studies of DMF show quantitative differences in human and rodent pathways, suggesting that rodent studies may not be indicative of human results. ... 

The two most common drug metabolism studies are mass balance and tissue distribution. Mass balance studies are usually conducted in both the rodent and the nonrodent species used for toxicology evaluations, whereas tissue distri-... 

In a study by Orzechowski et al. (1995), hepatocytes from adult male Wistar rats and NMRI mice were incubated for 1 hour with 0.5 mM 14C-benzene, and the supernatant analyzed for metabolites. Formation of sulfate conjugates of benzene, hydroquinone, and 1,2,4-benzenetriol was also studied in a separate experiment. Mouse hepatocytes produced two metabolites (1,2,4-trihydroxybenzene sulfate and hydroquinone sulfate) that were not found in rat hepatocyte incubations. These sulfate metabolites were found in incubations including benzene, or the metabolites themselves, hydroquinone and 1,2,4-benzenetriol. Mouse hepatocytes were almost three times more effective in metabolizing benzene, compared to rat hepatocytes. This difference was accounted for in the formation of hydroquinone, hydroquinone sulfate, and 1,2,4-trihydroxybenzene sulfate. These in vitro experiments indicate there are both quantitative and qualitative differences in rodent metabolism of benzene. 

It is conceivable that some excipients may not require the standard 2-year, two rodent species carcinogenicity studies. Such excipients include those that are not absorbed (or are rapidly metabolized and/or rapidly excreted), that do not exhibit toxicity in 90-day studies, and those that are negative for genotoxi-city. This is the approach taken by the IPEC-Americas Safety Committee and one of the reasons that the 1996 peer-reviewed journal publication indicates that the conduct of rodent carcinogenicity studies is conditional. The carcinogenicity studies that are conditional are the traditional 50 animals/sex/group rodent studies conducted for 18 or 24 months or variations thereof. The decision to make these tests conditional was also... 

In vitro metabolic studies with rodent and human liver microsomal prepara- tions have established that MPTP undergoes both oxidative N-demethylation and C-6 (allylic) oxidation in reactions that are -nicotinamide adenine dinucleotide phosphate (NADPH) dependent and therefore likely to be cytochrome P-450 catalyzed (Weissman et al. 1985 Ottoboni et al. 1990). Although the latter transformation can lead to the toxic pyridinium metabolite MPP, the cytochrome P450-catalyzed pathway is unlikely to contribute significantly to the neurotoxicity of MPTP. As mentioned above, liver aldehyde oxidase diverts the inter-mediate dihydropyridinium metabolite away from pyridinium ion formation by catalyzing the conversion of structure 40 to the nontoxic lactim structure 41. Further-more, even if formed in the periphery, the polar pyridinium metabolite would have limited access to the central nervous system (CNS). The low... 

The metabolic pathways leading to the production of these urinary pyridinium metabolites are likely to be mediated by one or more forms of liver cytochrome P450. In vitro metabolic studies with rodent (Igarashi et al., unpublished results) and human (Usuki et al., submitted) microsomal preparations have demonstrated the NADPH-dependent oxidation of both HP and HPTP to HPP. Ongoing studies in the authors laboratory have shown that HPP and related pyridinium metabolites are present in brain tissues obtained from C57 black mice that had been treated with HPTP (Van der Schyf et al. 1994). Additionally, results obtained from intra-cerebral microdialysis, mitochondrial respiration, and rat embryonic mesencephalic cell culture studies suggest that HPP possesses MPP type neurotoxic properties (Rollema et al. 1992, 1994 Bloomquist et al. 1994). 

The two most common drug metabolism studies are mass balance and tissue distribution. Mass balance studies are usually conducted in both the rodent and the nonrodent species used for toxicology evaluations, whereas tissue distribution is performed only in the rodent. For mass balance, a radio-labeled compound is administered to the test species and urine, feces, and, if necessary, expired air are collected at intervals and counted for total radioactivity. Commonly used intervals are 0-4, 4-8, 8-12, 12-24, and then daily, up to 168 hours or until more than 95% of the administered dose has been excreted. Depending on the pharmacokinetic profile of the candidate, other collection intervals can be selected to give a better picture of the excretion profile. For tissue distribution, a radiolabeled compound is administered to the test species, and after predefined times, usually 2, 4, 8, 24, and 48 hours, the test species... 

In studies with both hamster and human lung short-term cultures, major metabolites of BaP included the 9,10-dihydrodiol, the 7,8-dihydrodiol, and the sulfate conjugate of 3-hydroxy-BaP (78, 80, 81). Glucuronides of 3-hydroxy-BaP and the 7,8- and 4,5-dihydrodiols were also detected. The 7,8-dihydrodiol was active in binding to DNA of cultured hamster lung and cultured human bronchus (81, 503). Metabolic studies with the (-)-7,8-dihydrodiol in cultured human bronchus indicated that it was converted to the 7,8-dihydrodiol-9,10-epoxide (isomer I) which bound to DNA to give the major adduct 5 (Fig. 4) (503). The results of these studies indicate similarities in the metabolic activation of BaP in rodents and man. 

During digestion and transfer across the small intestine, and in the liver, flavanols are rapidly metabolized in phase I and phase II biotransformations to various 0-sulfated, <9-glucuronidated and (7-methylated forms. Various UGT isozymes, expressed in the intestine and in the Uver, have been identified to catalyze the glucuronidation of flavonoids. In humans consuming cocoa, plasma levels of non-methylated epicatechins such as epicatechin-7-sulfate and methylated metabolites such as 3 -0-methylepicatechin have been reported to occur in micromolar concentrations within 1 h after intake. Metabolic studies have confirmed the presence of these conjugates in the plasma and urine of rodents and humans, as well as in the bile and brain of rats. It has been reported that colonic microflora can break flavonoids flavan structure to form simple phenolics and rig-flssion metabolites that may be physiologically relevant. 

Metabolic studies with progesterone-2l-C in rodents showed that approximately 10% of the radioactivity was expired as CO2. About 80% of the radioactivity was in the combined excreta and the majority of this in the feces. In bile fistula animals the radioactivity was in the bile this suggests that most of the fecal activity was introduced into the intestines of the normal animals by way of the bile. However, one must again be reminded of species variation, and it is apparent that much work remains to be done in this important area. 

In vivo deuterium ( H) MRS has been used to characterize amino acid metabolism, body iron content, brain and kidney metabolism and body fat utilization rates in rodents. These studies rely on the use of deuterium labelling or the existence of natural-abundance deuterium in water or lipids. For example, deuterium-labelled methionine was used to confirm the dominant contribution of the glycine/ sarcosine shuttle to the metabolism of excess methionine, while deuterium-labelled glucose was used to show that systemic glucose level influences brain... 

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