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Biotransformation of chlorzoxazone by hepatic microsomes from humans and ten other mammalian species.
Abstract: The 6-hydroxylation of chlorzoxazone (CLZ) is currently being used in both in vivo and in vitro studies to quantify cytochrome P450 2E1 (CYP2E1) activity in humans. Comparatively little is known with regard to the biotransformation of this drug in other species. The NADPH-dependent biotransformation of CLZ was therefore studied using hepatic microsomes derived from humans and ten other mammalian species. In all species, 6-hydroxychlorzoxazone (6OH-CLZ) was the only metabolic product that could be identified by HPLC with ultraviolet detection. Enzyme kinetic analysis was used to characterize this CLZ 6-hydroxylase activity. Although the majority of kinetic data conformed to a single-enzyme Michaelis-Menten model, a two-enzyme (high and low affinity) model was required for four species (ferret, monkey, pig, and rat). Apparent K(m) values for the high-affinity component ranged from 12 microM (pig) to 95 microM (rabbit). The rank order of Vmax/K(m), an index of intrinsic clearance, was: mouse > horse > monkey > rabbit > cow > ferret > pig > human 1 > rat > human 2 > cat > dog. Diethyldithiocarbamate (DDC), a CYP2E1 inhibitor in humans, was a potent mechanism-based inhibitor of 6OH-CLZ formation in microsomes from all species examined. Preincubation of microsomes for 15 min in the presence of DDC and NADPH significantly enhanced the maximum degree of inhibition but had no effect on inhibitor potency. Inhibitor concentrations at 50% of maximum inhibition (IC50max) for DDC with preincubation ranged from 9 microM (human) to 45 microM (cow). In conclusion, DDC-sensitive CLZ 6-hydroxylation was identified as the principal NADPH-dependent pathway for chlorzoxazone metabolism in liver microsomes from humans and ten other mammalian species. These data indicate a species-conserved mechanism for the oxidative biotransformation of chlorzoxazone.
Publication Date: 1997-04-01 PubMed ID: 9113344DOI: 10.1002/(sici)1099-081x(199704)18:3<213::aid-bdd15>3.0.co;2-0Google Scholar: Lookup The Equine Research Bank provides access to a large database of publicly available scientific literature. Inclusion in the Research Bank does not imply endorsement of study methods or findings by Mad Barn.
- Journal Article
- Research Support
- U.S. Gov't
- P.H.S.
Summary
This research summary has been generated with artificial intelligence and may contain errors and omissions. Refer to the original study to confirm details provided. Submit correction.
This research investigates how the drug chlorzoxazone (CLZ) is metabolized in humans and ten other mammal species. The study reveals the metabolic pathway of how the drug is processed by these animals, and this is the principal NADPH-dependent method for CLZ metabolism.
Understanding the Study
The research conducted is focused on:
- The biotransformation of chlorzoxazone (a muscle relaxant drug), a process through which the drug is metabolized or chemically altered by the body.
- Specifically, the study looked at the hepatic microsomes (very small vesicles originated from the endoplasmic reticulum in the liver cells), which play a key role in drug metabolism in eleven mammalian species, including humans.
- The researchers were interested in the activity of the enzyme cytochrome P450 2E1 (CYP2E1), which results in 6-hydroxylation (addition of an hydroxyl group) of CLZ.
Findings of the Research
Various discoveries were made through the research:
- In all species studied, 6-hydroxychlorzoxazone (6OH-CLZ) was found to be the only metabolic product that could be identified. This indicates that the 6-hydroxylation pathway is predominant in the metabolism of CLZ.
- Using enzyme kinetic analysis, it was noticed that while most of the kinetic data conformed to a single-enzyme Michaelis-Menten model, there were four species (ferret, monkey, pig, and rat) where a two-enzyme model was needed to explain the findings.
- The apparent K(m) values for the high-affinity component ranged from 12 microM (pig) to 95 microM (rabbit), revealing variability in enzymes’ affinity for the substrate among species.
- The rank order of Vmax/K(m), which is an index of intrinsic clearance, was different between species, indicating different metabolic rates and levels of enzyme efficiency across species.
- Diethyldithiocarbamate (DDC), a CYP2E1 inhibitor, displayed significant inhibitory impact on 6OH-CLZ formation across all species, suggesting its potential role in modulating CLZ metabolism in the future.
- Preincubation of microsomes with DDC and NADPH resulted in amplification of the maximum degree of inhibition, but had no effect on the potency of the inhibitor, thus providing clues for further research and potential therapeutic applications.
Conclusion
In conclusion, this research confirms that DDC-sensitive CLZ 6-hydroxylation is the principal NADPH-dependent pathway for chlorzoxazone metabolism in humans and other mammalian species. This indicates that there is a species-conserved mechanism for the oxidative biotransformation of chlorzoxazone.
Cite This Article
APA
Court MH, Von Moltke LL, Shader RI, Greenblatt DJ.
(1997).
Biotransformation of chlorzoxazone by hepatic microsomes from humans and ten other mammalian species.
Biopharm Drug Dispos, 18(3), 213-226.
https://doi.org/10.1002/(sici)1099-081x(199704)18:3<213::aid-bdd15>3.0.co;2-0 Publication
Researcher Affiliations
- Department of Pharmacology and Experimental Therapeutics, Tufts University School of Medicine, Boston, MA 02111, USA. MCOURT@OPAL.TUFTS.EDU
MeSH Terms
- Aged
- Animals
- Antidotes / pharmacology
- Biotransformation
- Cats
- Cattle
- Chlorzoxazone / metabolism
- Chlorzoxazone / pharmacokinetics
- Cytochrome P-450 CYP2E1 / metabolism
- Ditiocarb / pharmacology
- Dogs
- Enzyme Inhibitors / pharmacology
- Female
- Ferrets
- Horses
- Humans
- In Vitro Techniques
- Macaca fascicularis
- Mice
- Microsomes, Liver / drug effects
- Microsomes, Liver / enzymology
- Middle Aged
- Muscle Relaxants, Central / metabolism
- Muscle Relaxants, Central / pharmacokinetics
- NADP / pharmacology
- Rabbits
- Rats
- Species Specificity
- Swine
Grant Funding
- K01-RR-00104 / NCRR NIH HHS
- MH-19924 / NIMH NIH HHS
- MH-34223 / NIMH NIH HHS
Citations
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- Martinez SE, Andresen MC, Zhu Z, Papageorgiou I, Court MH. Pharmacogenomics of poor drug metabolism in Greyhounds: Cytochrome P450 (CYP) 2B11 genetic variation, breed distribution, and functional characterization. Sci Rep 2020 Jan 9;10(1):69.
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