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BiotransformationXenobiotic metabolism
“Essentials of Toxicology”
by Klaassen Curtis D. and Watkins John B Chapter 6
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Biotransformation• Water soluble xenobiotics are easier to
eliminate ( t1/2)– Urine, feces but not exhalation– If within barrier, no out
• Multiple enzymes (families)– Constitutively expressed– Inducible– Broad specificity– Polymorphic (allelic variants)– Stereo-isomer specificity: 6-OH in hormones:
CYP2A1 6-OH CYP3A 6-OH
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Biotransformation
Potentially toxic xenobiotic
Inactive metabolite
Relatively harmless
Reactive intermediate
DetoxificationMetabolic activation
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Converting lipophilic to water soluble compounds
Xenobiotic
Reactive intermediate
Conjugate
Phase I - Activation
Phase II - Conjugation
Excretion
Lipophilic
(non-polar)
Water soluble(polar)
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Phase I
• introduction of functional group
• hydrophilicity increases slightly• may inactivate or activate original compound• major player is CYP or mixed function oxygenase
(MFO) system in conjunction with NAD(P)H• location of reactions is smooth endoplasmic reticulum
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Phase II
• conjugation with endogenous molecules(GSH, glycine, cystein, glucuronic acid)
• hydrophilicity increases substantially• neutralization of active metabolic intermediates• facilitation of elimination • location of reactions is cytoplasm
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Phase I reactions
Oxidation Hydroxylation (addition of -OH group) N- and O- Dealkylation (removal of -CH side chains) Deamination (removal of -NH side chains) Epoxidation (formation of epoxides) Oxygen addition (sulfoxidation, N-oxidation) Hydrogen removal
Reduction Hydrogen addition (unsaturated bonds to saturated) Donor molecules include GSH, FAD, NAD(P)H Oxygen removal
Hydrolysis Splitting of C-N-C (amide) and C-O-C (ester) bonds
OC C
OC
See also Chapter 6 of Casarett and Doull’s “Toxicology”
Table 6.1
epoxide
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Biotransformation
• Activation of xenobiotics is a key element (e.g. benzene, vinyl chloride)– Reactive intermediates include epoxides and free
radical species (unpaired electrons) that are short-lived and hence highly reactive
– Protection is provided by • endogenous antioxidant substances, e.g. GSH• vitamins C and E• antioxidant enzymes, SOD, GPX, CAT in coupled reactions
– Antioxidant molecules are oxidized in the process but have the capacity to regenerate the reduced form from the oxidized - NAD(P)H is a key player
See also p. 40-44 of Casarett and Doull’s “Toxicology”
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Cytochrome P450 (CYP) Mixed Function Oxidases (MFO)
• Located in many tissues but highly in liver ER• Human: 16 gene families• CYP 1,2,3 perform drug metabolism• >48 genes sequenced• Key forms: CYP1A2, CYP2C9, CYP2C19, CYP2D6,
CYP2E1, and CYP3A4• Highly inducible
– Alcohol CYP2E1– Dioxin/PCBs CYP1A– Barbiturates CYP2B
• CYP genes have multiple alleles (2D6 has 53, and 2E1 has 13)
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The CYP-450 reaction cycleA
E
D
C
F
G(B)
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Oxidation of vinyl chloride to an epoxide
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Metabolic enzymes
1. Microsomal:1. CYP450 monooxygenases2. Flavin monooxygenase
2. Non-microsomal1. Alcohol dehydrogenase2. Aldehyde dehydrogenase3. Monoamine and diamine oxidases
3. Both1. Esterases and Amidases2. Prostaglandin synthase 3. Peroxidases
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Cooxidation of acetaminophenby prostaglandin endoperoxide synthetase
Compare to fig. 6-2
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Hydrolysis of esters and amides
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Hydrolysis of organophosphates
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Hydrolysis of epoxides
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Stereoselectivehydroxylation
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Metabolism of benzo(a)pyrene to 9,10 epoxide:Potent mutagen that binds DNA
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Azo- and nitro- reduction
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Flora action
Intestinal flora as part of biotransformation
reabso
rptio
n
Ready for elimination
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Oxidation reactions
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Benzene trasformation to leukemia-causing metabolite
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Flavin mono-oxygenases(FMO) catalyzed reactions
Nitrogen compounds
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Phase II reactions
• Glycoside conjugation - glucuronidation• Sulfate - sulfation• Glutathione (GSH)• Methylation• Acylation
– Acetylation– Amino acid conjugation– Deacetylation
• Phosphate conjugation
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Glucuronidation of phenol
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Sulfation of phenol and toluene
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GSH conjugation of acetaminophen
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Glutathione
-glutamyl-cysteinyl-glycine
Active site of a GST: