Xenobiotic Biotransformation Basic Concepts

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Section A

Biotransformation: Basic Concepts


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Renal Excretion of Chemicals

Afferent arteriole

Efferent arteriole

Glomerulus

Bowmans capsule

Proximal tubule Water soluble

Lipid soluble

Filtered drugPassive

reabsorption

Excretion and/or further passive reabsorption

Unltered drug

Activesecretion


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Biotransformation of Xenobiotics

Biological basis for xenobiotic metabolism:

! To convert lipid-soluble, non-polar, non-

excretable forms of chemicals to water-soluble, polar forms that are excretable in

bile and urine

Continued


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Biotransformation Reactions

! Phase I Reactions

Enzymatic reactions that addor

exposefunctional groups to xenobioticssuch as -OH, -SH, -NH2

or COOH

Functional groups are analogous tohaving a trailer hitch on a vehicle

Continued


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Biotransformation Reactions

! Phase II Reactions

Enzymatic reactions that result in the

conjugation of large water-soluble,charged (polar)biomolecules to

xenobiotics

For these reactions to occur, afunctional group must be present oneither the parent compound or itsPhase I product


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Foreign Chemical(xenobiotic)

lipophilic not charged

not water soluble poorly excretable

TRUCK

The Truck-Hitch-Trailer Analogy to Xenobiotic Biotransformation

Photo by John Pittman. Creative Commons BY-NC-SA.


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TRUCK HITCH

Phase 1 enzymesadd or expose a

functional group

still lipophilic

possibly reactive

poorly water soluble poorly excretable

catalyzed by P450s




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TRUCKHITCH

Phase 1 enzymesadd or expose a

functional group

still lipophilic

possibly reactive

poorly water soluble poorly excretable

catalyzed by P450s

Phase 2 enzymesconjugate (transfer)

endogenous molecules*to the functional group

TRAILER

not lipophilic

usuallynot reactive

water soluble products excretable

catalyzed by transferases

*sugars, amino acids, sulfates, acetyl groups




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Section B

Biotransformation: Enzymes


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Organ and Cellular Location of

Biotransformation Enzymes! Organs involved in biotransformation

Liver

Lung

Kidney

Intestine

Enterocytes

Gut flora (contribute to entero-hepaticcirculation)

Skin

Gonads


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Biotransformation Enzyme-

Containing Cells in Various Organs

Organ Cell s)

Liver Parenchymal cells (hepatocytes)

Kidney Proximal tubular cells (S3 segment)

Lung Clara cells, Type II alveolar cells

Intestine Mucosa lining cells

Skin Epithelial cellsTestes Seminiferous tubules, Sertoliscells


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Nature of the Xenobiotic

Metabolizing Enzyme System! Phase I metabolism

Small molecular weight changes like

hydroxylation, reduction, hydrolysis, etc.

In general, Phase I metabolismprepares the xenobiotic for subsequent

Phase II reactions


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Cytochrome P450

Characteristics! Can metabolize many xenobiotics (broad

substrate specificity)

!

Can catalyze many types of reactions! Is widely distributed among tissues, and

tissue distribution can be quite varied

Continued


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Multiple Forms of P450Major Mammalian Cytochrome P450 Gene Families

Continued


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Multiple Forms of P450Major Mammalian Cytochrome P450 Gene Families


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Nature of the Xenobiotic

Metabolizing Enzyme System! Phase II metabolism

Involves the complexing or conjugation

of xenobiotics with relatively large andhighly water-soluble adducts to form

glucuronides, sulfates, and glutathioneadducts


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Nature of Systems Involved in

Phase II MetabolismFour primary enzymes:

1.

Glucuronosyltransferaseglucuronic acid

2.

Sulfotransferasesulfate3.

Glutathione-S-transferaseglutathione(GSH)

4.

Acetyltransferaseacetyl


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Phase II Reactions

! Many of the characteristics describedearlier for cytochrome P450 also apply tothese Phase II enzymes

! However, cytochrome P450 is localized incellular membraness, whereas Phase IIenzymes are, for the most part, in the

cytoplasm (water portion) of cells


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Phase II Enzymes: ExamplesGlucuronidation and Sulfation of a Hydroxyl Group

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Glutathione-S-TransferaseStructure of Reduced Glutathione (MW 307)


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Section C

Factors Affecting Biotransformation


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Factors that Affect

Xenobiotic Biotransformation

Likewise, even within a species,including man, there are differences

The basis of such differences is oftengenetic (polymorphisms)


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Examples of Factors that Affect

Xenobiotic Biotransformation! Species, strain, and genetic variation

- Hexobarbital

Aflatoxin B1

Benzo[a]pyrene 7,8 dihydrodiol

Isoniazid

!

Age! Diet

! Exposure to other chemicals


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Species Differences in the Duration ofAction and Metabolism of Hexobarbital

Species Duration of

ction

(Sleeping time, min)

Relative Enzyme

ctivity

(!g hexobarbitolmetabolized/gm/liver/hr)

Mouse 12 8 598 184

Rabbit 49 12 196 28

Rat 90 15 134 51Dog 315 105 36 30

Source: G.P. Quinn, et.al. Species, Strain and Sex Differences in the Metabolism of

Hexobarbital, Aminopyrine and Aniline. Biochem. Pharmacol. 1:152, 1958


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Species, Strain,

and Genetic Variation

Continued


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A B D E F G H I J

1000

2000

3000

4000

C A B D E F G H I JC

His+revertants/mg

Biopsy samples

Metabolism of Aflatoxin B1

and Benzo[a]pyrene

Inter-individual differences in the metabolism of aflatoxin B1 (A)and benzo[a]pyrene 7,8-dihydrodiol (B) to mutagens( assessed

by Ames test) by microsomes from samples of human liver

obtained during abdominal surgery

A B


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The Bimodal Distribution of Patients intoThose who Rapidly Inactivate Isoniazid and

Those who Slowly Metabolize It50

40

30

20

10

0 1 2 3 4 5 6 7 8 9 10 11 12

Plasma Isoniazid ( g/mL)

NumberofPati

ents


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Age as Affecting Xenobiotic

Biotransformation

Schematic representation of the ontogeny ofhepatic drug metabolic activity

Birth Puberty Adult Elderly (>65 yrs)

Age


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Exposure to Other Chemicals


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Section D

Induction of

Biotransformation Enzymes


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Induction of Xenobiotic

Metabolizing Systems1.

Many chemicals can induce the synthesisof the enzymes involved in Phase I and IIxenobiotic metabolism and include

chemicals found in the environment, thediet, and cigarette smoke

2.

Inducers often exhibit specificity for the

enzymes which they induce

Continued


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Depending on the inducer, fairly highdose levels or repeated dosing may berequired; on the other hand, TCDD

(dioxin) is effective as an inducer at1 microgram/kg in some species

Continued


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Studies have demonstrated that acluster of genes referred to as the

Ah locuscontrols the induction of

xenobiotic enzyme activities by polycyclicaromatic compounds and TCDD

Continued


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Such toxic responses as cancer, chemical-induced cataracts, aplastic anemia, andfetal toxicity have been demonstrated to

be affected by this cluster of genes

6.

Evidence exists for theAh locusin man


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Characteristics of the Hepatic Effects ofPhenobarbital and Polycyclic Aromatic Hydrocarbons

haracteristics Phenoba rbital

Polycyclic

Hydrocarbons

Onset of effects 812 hours 36 hours

Time of maximum effect 35 days 2448 hours

Persistence of induction 57 days 512 days

Liver enlargement Marked Slight

Protein synthesis Large increase Small increase

Phospholipid synthesis Marked increase No effect

Liver blood flow Increase No effect

Biliary flow Increase No effectGlucuronidation Increase Small increase

Glutathione conjugation Small increase Small increase

Epoxide hydrolase Increase Small increase

Cytosolic receptor None identified Identified

Continued


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TheAh Receptor

!Ah receptor = Arylhydrocarbon receptor

! Examples = 3-methylcholanthrene

benzo[a]pyrene!Also called TCDD receptoror dioxin

receptor

Continued


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Bioactivation as a Basis for

Chemical Toxicity! One of the possible results of the

interaction of a xenobiotic with enzymesystems is the biotransformation of that

compound to a chemically reactiveintermediate (i.e. Bioactivation)

! The reaction of either this initial reactive

metabolite or secondary reactiveproducts with target molecules bringsabout changes in cellular function (the

Molecular Targets Concept)


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Proposed Relationship Between Biotransformation,Bioactivation, and Toxicity of a Xenobiotic

49Adapted from Casarett & Doulls Toxicology. 4

th

Edition.


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Metabolism and Bioactivationof Benzo[a]pyrene

50


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Chemical Nature ofReactive Intermediates

! ElectrophilesForm covalent (irreversible)bonds with cellular nucleophiles such asGSH, proteins and DNA

! Free RadicalsOdd or unpaired electron

Can act as electrophiles

Can abstract hydrogen from targetmolecules, such as lipids or nucleic acids

Can activate molecular oxygen


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Acetominophen is good exampleof a xenobiotic whose toxicity is

due to bioactivation to anelectrophile


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Relationship between hepatic glutathione levels and

covalent binding of acetaminophen to targetnucleophiles(proteins)

Bioactivation of Acetaminophen

100

80

60

40

20

20 40 60 100 200 400 600 10000

1

2

CovalentB

inding(molecu

les/mgprotein

)

InitialGlutathionein

Liver(%)

Dose of Acetaminophen (mg/kg)

Covalent binding

Glutathione


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Human paraquat exposure canresult in lung toxicity due to its

accumulation in lung cells andredox cycling

! The structure of the herbicide paraquat (A)and the polyamines putrescine (B) and

spermine (C)


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Redox Cycling of Xenobiotics

! Redox cyclingof xenobiotics initially resultsin the formation of a form of active oxygencalled superoxide(O2

)

!

Through a series of non-enzymatic oftenmetal catalyzed, reactions other forms ofreactive oxygen are formed

These include hydrogen peroxide(H2O2), the hydroxyl radical ( OH) andsinglet oxygen (1O2)

Continued


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Redox Cycling of Xenobiotics

! This results in an oxidative stressin cellsand the subsequent modification of criticalbiomolecules leading to cellular toxicity

!

In this situation what is the active formthat causes toxicity?

Documents

Xenobiotic Biotransformation Basic Concepts