Metabolism of Ethyl Carbamate by Pulmonary Cytochrome P450 and Carboxylesterase Isozymes: Involvement of CYP2E1 and Hydrolase A

Forkert, Poh‐Gek; Lee, Raymond P.

doi:10.1006/taap.1997.8233

Cited by 35 publications

(26 citation statements)

References 23 publications

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“…As shown in Fig. 1, earlier studies suggested that metabolism of urethane occurs via two major pathways (Yamamoto et al, 1990;Page and Carlson, 1994;Forkert and Lee, 1997;Lee et al, 1998). The first pathway is thought to entail oxidative metabolism of urethane catalyzed by cytochromes P450, leading to the formation of vinyl carbamate (VC) (Fig.…”

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confidence: 81%

“…[ethyl-1,2-3 H] or [ethyl-1-14 C]urethane to 12-day-old and adult male mice (Ribovich et al, 1982). Generally, the current hypothesis states that while esterase is the primary enzyme responsible for urethane metabolism, CYP2E1 is responsible for urethane activation (Yamamoto et al, 1990;Forkert and Lee, 1997;Lee et al, 1998). In an attempt to address this hypothesis and assess the metabolic basis of urethane toxicity and carcinogenicity, the present work is designed to more directly characterize the enzymes responsible for urethane metabolism using CYP2E1-null versus wild-type mice.…”

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confidence: 99%

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Cytochrome P450 2E1 (CYP2E1) Is the Principal Enzyme Responsible for Urethane Metabolism: Comparative Studies Using CYP2E1-Null and Wild-Type Mice

Hoffler¹,

El‐Masri²,

Ghanayem³

2003

J Pharmacol Exp Ther

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Urethane ([carbonyl-14 C]ethyl carbamate) is a fermentation byproduct in alcoholic beverages and foods and is classified as reasonably anticipated to be a human carcinogen. Early studies indicated that while CYP2E1 is involved, esterases are the primary enzymes responsible for urethane metabolism. Using CYP2E1-null (KO) mice, current studies were undertaken to elucidate CYP2E1's contribution to urethane metabolism. [Carbonyl-14 C]urethane was administered by gavage to male CYP2E1-null and wild-type mice at 10 or 100 mg/kg and its metabolism and disposition were investigated. CO 2 was confirmed as the main metabolite of urethane. Significant inhibition of urethane metabolism to CO 2 occurred in CYP2E1-null versus wild-type mice. Pharmacokinetic modeling of 14 CO 2 exhalation data revealed that CYP2E1 is responsible for approximately 96% of urethane metabolism to CO 2 in wild-type mice. The contributions of other enzymes to urethane metabolism merely account for the remaining 4%. The half-life of urethane in wild-type and CYP2E1-null mice was estimated at 0.8 and 22 h, respectively. Additionally, the concentration of urethane-derived radioactivity in blood and tissues was dose-dependent and significantly higher in CYP2E1-null mice. High-performance liquid chromatography analysis showed only urethane in the plasma and liver extracts of CYP2E1-null mice. Because the lack of CYP2E1 did not completely inhibit urethane metabolism, the disposition of 10 mg/kg urethane was compared in mice pretreated with the P450 inhibitor, 1-aminobenzotriazole or the esterase inhibitor, paraoxon. Unlike paraoxon, 1-aminobenzotriazole resulted in significant inhibition of urethane metabolism to CO 2 in both genotypes. In conclusion, this work demonstrated that CYP2E1, not esterase, is the principal enzyme responsible for urethane metabolism.

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confidence: 81%

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confidence: 99%

Cytochrome P450 2E1 (CYP2E1) Is the Principal Enzyme Responsible for Urethane Metabolism: Comparative Studies Using CYP2E1-Null and Wild-Type Mice

Hoffler¹,

El‐Masri²,

Ghanayem³

2003

J Pharmacol Exp Ther

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“…Enzyme Assays ---Microsomal ER O-deethylase activity as an index of CYP1A1 was measured by the method of Sinjari et al 17) Microsomal PN Odeethylase activity as an index of CYP1A2 was assayed according to the method of Sesardic et al 18) Microsomal and cytosolic PNA hydrolase activities as an index of carboxylesterase were determined by the method of Forkert and Lee. 19) Cytosolic sulfotransferase activities toward 4-OH-PL were measured according to a published HPLC method. 20) In the 4-OH-PL sulfation assay, activities were calculated using peak height ratios of 4-OH-PL sulfate to 4-OH-BTL as an internal standard because of the lack of an authentic standard.…”

Section: Methodsmentioning

confidence: 99%

Effects of Pretreatment of Hep G2 Cells with .BETA.-Naphthoflavone on Cytotoxicity of Propranolol and its Active Metabolite 4-Hydroxypropranolol

Miyano

Motoyama

Fukuoka

et al. 2003

JOURNAL OF HEALTH SCIENCE

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Cytotoxicities of propranolol (PL) and its active metabolite, 4-hydroxypropranolol (4-OH-PL), were examined in a human hepatoma cell line, Hep G2. Hep G2 cells were cultured in the presence of β-naphthoflavone (BNF, 25 or 50 µM), lansoprazole (LPZ, 25 or 50 µM) or 0.5% dimethylsulfoxide (vehicle) for 48 hr. The cells were harvested, and microsomal and cytosolic fractions were prepared by differential centrifugation methods. Various enzyme activities were determined as follows: microsomal 7-ethoxyresorufin (ER) O-deethylation as a CYP1A1 index, microsomal phenacetin (PN) O-deethylation as a CYP1A2 index, microsomal and cytosolic p-nitrophenyl acetate (NPA) hydrolysis as a carboxylesterase index and cytosolic 4-OH-PL sulfation as a sulfotransferase index. The pretreatment of Hep G2 cells with LPZ or BNF increased microsomal ER O-deethylase activities, and the potency of BNF was much higher than that of LPZ. Cytosolic 4-OH-PL sulfation was also elevated by the pretreatment with BNF but not with LPZ. Microsomal PN O-deethylase activity was not detectable in either the control or BNFpretreated group under the conditions used. Microsomal and cytosolic NPA hydrolase activities were similar between the control and the BNF-pretreated groups. Cytotoxicities of PL and 4-OH-PL were attenuated in BNFpretreated Hep G2 cells compared to non-pretreated Hep G2 cells. These results suggest that increased activities of microsomal CYP1A1 and cytosolic sulfotransferases by pretreatment with BNF may contribute to the attenuating the cytotoxicity of PL and 4-OH-PL in Hep G2 cells, at least in part.

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“…It induced DNA adducts in mouse liver and lung (Fernando et al, 1996). Forkert and Lee (1997) demonstrated that urethane metabolism in lung microsomes is mediated by CYP2E1 and the carboxylesterase isozyme hydrolase A. Using the standard induction procedures, the level of CYP2E1 in rat liver is actually suppressed and this may account for the negative findings with these substances in the Ames test and other in vitro tests (Burke et al, 1994).…”

Section: Occurence Of Unique In Vivo Positivesmentioning

confidence: 99%

Scientific opinion on genotoxicity testing strategies applicable to food and feed safety assessment

2011

EFS2

296

136

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The Scientific Committee reviewed the current state-of-the-science on genotoxicity testing and provided a commentary and recommendations on genotoxicity testing strategies. A step-wise approach is recommended for the generation and evaluation of data on genotoxic potential, beginning with a basic battery of in vitro tests, comprising a bacterial reverse mutation assay and an in vitro micronucleus assay. Consideration should be given to whether specific features of the test substance might require substitution of one or more of the recommended in vitro tests by other in vitro or in vivo tests in the basic battery. In the event of negative in vitro results, it can be concluded that the substance has no genotoxic potential. In case of inconclusive, contradictory or equivocal results, it may be appropriate to conduct further testing in vitro. In case of positive in vitro results, review of the available relevant data on the test substance and, where necessary, an appropriate in vivo study to assess whether the genotoxic potential observed in vitro is expressed in vivo is recommended. Suitable in vivo tests are the mammalian erythrocyte micronucleus test, transgenic rodent assay, and Comet assay. The approach to in vivo testing should be step-wise. If the first in vivo test is positive, no further testing is necessary and the substance should be considered as an in vivo genotoxin. If the test is negative, it may be possible to conclude that the substance is not an in vivo genotoxin. However, in some cases, a second in vivo test may be necessary (e.g. if the first test is negative but more than one endpoint in the in vitro tests are positive, an in vivo test on a second endpoint may be necessary). The combination of assessing different endpoints in different tissues in the same animal in vivo should also be considered.

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Metabolism of Ethyl Carbamate by Pulmonary Cytochrome P450 and Carboxylesterase Isozymes: Involvement of CYP2E1 and Hydrolase A

Cited by 35 publications

References 23 publications

Cytochrome P450 2E1 (CYP2E1) Is the Principal Enzyme Responsible for Urethane Metabolism: Comparative Studies Using CYP2E1-Null and Wild-Type Mice

Cytochrome P450 2E1 (CYP2E1) Is the Principal Enzyme Responsible for Urethane Metabolism: Comparative Studies Using CYP2E1-Null and Wild-Type Mice

Effects of Pretreatment of Hep G2 Cells with .BETA.-Naphthoflavone on Cytotoxicity of Propranolol and its Active Metabolite 4-Hydroxypropranolol

Scientific opinion on genotoxicity testing strategies applicable to food and feed safety assessment

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