Slides prepared for MBBS Biochemistry lecture. Explains Cytochrome P450 systems, Phase 1 and Phase 2 reactions, drug interactions, and procarcinogen activation

1. Metabolism of
Xenobiotics
Ashikh Seethy
Dept of Biochemistry
Maulana Azad Medical College
New Delhi-110002

2. Introduction
• Xenos = stranger = foreign to life
• Xenobiotics = Foreign chemicals
• E.g. drugs, food additives, pollutants, chemical
carcinogens, insecticides
• Most of these undergo metabolism in human body
• Basic to a rational understanding of:
̴ Pharmacology and therapeutics
̴ Pharmacy
̴ Toxicology
̴ Management of cancer
̴ Drug addiction
• All these areas involve administration of, or
exposure to, xenobiotics

3. Xenobiotic Metabolism
• Major site: Liver
• Purpose:
– Converts lipophilic to hydrophilic compounds
– Facilitates excretion
Hydrophobic xenobiotics can persist in adipose
tissue
• 2 phases
• Phase 1: Hydroxylation
- Cytochrome P450s >> Non-P450 reactions
- Hydroxylation >> Non-hydroxylation
• Phase 2:
Conjugation/ Methylation of compounds produced
in Phase 1

4. First Pass Effect:
• Biotransformation by liver or gut enzymes
before compound reaches systemic circulation
• Results in lower systemic bioavailability of
parent compound
• Examples: Isoniazid, Propanolol
• Certain drugs as administered parenterally [par:
far; enteral: intestine] to bye pass first pass
effect

5. Phase 1 reactions:
• Hydroxylation
• Deamination, dehalogenation, desulfuration
• Epoxidation, Peroxidation
• Reduction
• Hydrolysis
• Effects:
Inactivation of xenobiotic
Inactive  Active compound
Prodrugs, Procarcinogens
• Cytochrome P450 isoforms play a key role

6. Cytochrome P450
• Involved in phase I of the metabolism of
innumerable xenobiotics, including perhaps 50%
of the drugs administered to humans.
• Hemoproteins
• Complex formed between Fe2+ and CO absorbed
light maximally at 450 (447-452) nm
• Involved in the metabolism of many endogenous
compounds, e.g. steroids
• Versatile catalysts – about 60 types of reactions
• Mono-oxygenases
RH + O2 + H+ + NADPH  ROH + H2O + NADP+

7. Nomenclature:

8. Cytochrome P450
• Present in all tissues
• Highest in Liver
• Membrane of smooth Endoplasmic Reticulum
(Microsome)
• In adrenals:
Both microsomal and mitochondrial
Cholesterol and steroid biosynthesis
• Utilizes NADPH
• Broad substrate specificity, thus acting on many
compounds  Different P450s may catalyze
formation of the same product

9. Most isoforms of CYP can be induced or
inhibited
Induction:
• Increased rate of biotransformation due to new
protein synthesis
Increased transcription of mRNA
Stabilization of mRNA
Enzyme stabilization
Effect on translation
• Drug-drug interactions
• Possible subtherapeutic plasma concentrations

10. Drug Interaction
Warfarin
CYP2C9 Phenobarbital
Induction
Inactive
•Co-administration of Warfarin and Phenobarbital
reduces the effect of Warfarin
•The dose of Warfarin must be increased to
achieve therapeutic effect
Procarcinogens
•Ethanol  Induction of CYP2E1
Carcinogens

11. CYP Polymorphisms:
• Certain cyto-P450s exist in polymorphic forms some of which
exhibit low catalytic activity
• CYP2D6:
 Involved in the metabolism of debrisoquin (an
antihypertensive drug) and sparteine (an antiarrhythmic and
oxytocic drug)
 Certain polymorphisms of CYP2D6 cause poor metabolism of
these and a variety of other drugs so that they can
accumulate in the body, resulting in untoward consequences.
• CYP2A6: Metabolism of nicotine to conitine
 Three alleles: A wild type and two null or inactive alleles
 Null alleles Impaired metabolism of nicotine
 These individuals smoke less, presumabl brain
concentrations of nicotine remain elevated longer 
apparently protected against becoming tobacco- dependent
smokers
 ? A novel way to help prevent and to treat smoking.

12. Activation of Procarcinogens by CYP
• CYP1A1: Aromatic hydrocarbon hydroxylases
• Metabolise Polycyclic Aromatic Hydrocarbons
(PAH) present in cigarette smoke
Hydroxylation
• Active carcinogens
• High levels of enzyme in
 Smokers
 Placenta of female smokers  harmful to fetus

13. PHASE II REACTIONS
Reaction Endogenous
Reactant
Enzyme (Location) Types of Substrates
Glucuronid
ation
UDP glucuronic
acid
UDP
glucuronosyltransferase
(microsomes)
Phenols, alcohols, carboxylic
acids, hydroxylamines,
sulfonamides
Glutathione
conjugation
Glutathione GSH-S-transferase
(cytosol microsomes)
Epoxides, arene oxides, nitro
groups, hydroxylamines
Glycine
conjugation
Glycine Acyl-CoA
glycinetransferase
(mitochondria)
Acyl-CoA derivatives of
carboxylic acids
Sulfate
conjugation
Phosphoadenosy
l phosphosulfate
Sulfotransferase (cytosol) Phenols, alcohols, aromatic
amines
Acetylation Acetyl-CoA N-Acetyltransferase
(cytosol)
Sulfonamides, isoniazid,
clonazepam, dapsone,
mescaline
Methylation S-Adenosyl
methionine
Transmethylases (cytosol) Catecholamines, phenols,
amines
Water
conjugation
Water Epoxide hydrolase
(microsomes)
Arene oxides,
disubstituted &
monosubstituted oxiranes
(cytosol) Alkene oxides, fatty acid
epoxides

14. Glucuronidation: Major phase II pathway in
mammals
• By UDP-glucuronyltransferase
• Six forms in human liver
• Forms O-, N-, S-, C- glucuronides
• Conjugates excreted in bile or urine
• Cofactor is UDP-glucuronic acid
• Inducers:
Phenobarbital
Indoles
3-methyl cholanthrene
cigarette smoking
• Substrates:
Morphine, p-nitrophenol, valproic acid, NSAIDS,
bilirubin, steroid hormones

15. Glucuronidation & genetic polymorphisms
• Crigler-Nijar syndrome (severe): inactive
enzyme; severe hyperbilirubinemia; inducers
have no effect
• Gilbert’s syndrome (mild): reduced enzyme
activity; mild hyperbilirubinemia; phenobarbital
increases rate of bilirubin glucuronidation to
normal
• Patients can glucuronidate p-nitrophenol,
morphine, chloroamphenicol

16. Sulfation:
• Sulfotransferases
• Cofactor: 3’-phospho adenosine -5’ -phospho
sulfate (PAPS): Active Sulfate
• Produce highly water-soluble sulfate esters,
eliminated in urine, bile
• Phenols, catechols, amines, hydroxylamines

17. Methylation
• Common, minor pathway which generally
decreases water solubility
• Methyl transferases
• Cofactor: S-adenosylmethionine (SAM)
• -CH3 transfer to O, N, S, C
• Substrates include phenols, catechols, amines,
heavy metals (Hg, As, Se)

18. Acetylation:
• Major route of biotransformation for aromatic
amines, hydrazines
• Generally decreases water solubility
• N-acetyltransferase (NAT)
• Cofactor is Acetyl Coenzyme A
• Humans express two forms- slow/fast
• Substrates include sulfanilamide, isoniazid,
dapsone

19. Acetylation and Polymorphisms
• Rapid and slow acetylators
• Various mutations result in decreased enzyme
activity or stability
• Drug toxicities in slow acetylators:
Nerve damage from dapsone
Peripheral neuropathy due to Isoniazid
Bladder cancer in cigarette smokers due to
increased levels of hydroxylamines

20. Conjugation with Glutathione
• R + GSH R-S-G (R: Electrophilic Xenobiotic)
• Glutathione-S-Transferases
• High levels in Liver cytosol
• Prevent binding of toxic xenobiotics to DNA,
RNA and proteins
• Glutathione conjugates are further metabolized
and excreted in the form of mercapturic acid

21. Phase 1 vs Phase 2
Enzyme Phase I Phase II
Types of reactions Hydrolysis
Oxidation
Reduction
Conjugations
Increase in
hydrophilicity
Small Large
General mechanism Exposes functional
group
Polar compound added
to functional group
Consquences May result in
metabolic activation
Facilitates excretion

22. Reversal of order of the phases
Isoniazid
↓
Acetylation (Phase 2)
↓
Hydrolyzed (Phase 1)
↓
Isonicotinic acid
↓
Excreted

23. Pharmacogenetics:
• Study of contribution of genetic factors to
variation in drug response and toxicity

24. Metabolism of a xenobiotic can result in cell injury,
immunologic damage, or cancer

25. Thank
You