Phase I/II metabolism

Drug metabolism
Drugs can undergo one of several pathways into the body:
• spontaneously changed into other compounds → atracurium into laudanosine;
• excreted unchanged → 50% benzylpenicillin excreted into urine;
• metabolised by enzymes to different compounds to facilitate excretion

Sites of drug metabolism (taken p/o):

• Liver/hepatocytes
• The smooth endoplasmic reticulum of the

hepatocyte is the principal site of metabolism

https://resources.wfsahq.org/atotw/the-role-of-the-liver-in-drug-metabolism/
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Phase 1 – General Information
• Modification of the substrate 🡪 functionalization
• Introducing a reactive/polar group

After Phase 1 the molecule should be …

• hydrophilic enough to be excreted via urine
• able to react in phase 2

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Phase 1 – Enzymes and Reactions
Monoamine • Aliphatic hydroxylation
oxidase (MAO)
• Epoxidation/hydroxylation of
Flavin containing
aromatic rings
Monooxygenase Cytochrom
(FMO) P450 (CYP) • O-, N-, S-dealkylation
• Epoxidation of alkenes and
alkines
Epoxidhydrolase
(EH)
Cyclooxygenase
(COX)
• Desulfurization
…
Dehydrogenase

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W. Wätjen, E. Fritsche, Fremdstoffmetabolismus, 2010
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M.D. Coleman, Human Drug Metabolism, 2020
CYP-Reactions
• Aliphatic hydroxylation

• Epoxidation/hydroxylation
of aromatic rings

• O-, N-, S-dealkylation

• Epoxidation of alkenes and alkines

• Desulfurization
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Toxicology lecture by Prof. Dr. Marko, 2022
CYP monooxygenase– Heme group and mechanism

O2 + NADPH + H+ + RH → NADP+ + H2O + ROH

http://www.chemgapedia.de/vsengine/vlu/vsc/de/ch/11/toxikologie/kap_1/vlu/stoffwechsel.vlu/7
M.D. Coleman, Human Drug Metabolism, 2020 Page/vsc/de/ch/11/toxikologie/kap_1/kap1_5/kap15_41.vscml.html
Phase 1 – Activation or Toxification?

• Aflatoxin B1

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https://www.researchgate.net/figure/Metabolites-and-enzymes-involved-in-aflatoxin-B1-AFB1-metabolism-in-the-turkey-liver_fig3_282893492
Phase 1 – Activation or Toxification?
• Codeine

CYP2D6

Codeine Morphine

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https://fb.cuni.cz/file/5635/FB2012A0008.pdf
Phase 2 – General Informations
• Phase 2 enzymes form a conjugate of the substrate
• Facilitate the elimination of drugs (water solubility)
• Inactivation of potentially toxic compounds
• Covalently bound
• Dependency on co-factors
• Acceptor sites for the conjugation: O-, N-, S-

Goodman and Gilman's The Pharmacological Basis of

Therapeutics, 11th Edition;

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 Phase 2
enzymes -
localization
• Found in most tissue in
the body: highest levels in
the GI-tract (liver, small
and large intestine)
• All of the enzymes are
located in the cytosol,
except UDP-
Glucuronosyl-
transferase (membrane of http://what-when-how.com/human-drug-metabolism/conjugation-
ER) and-transport-processes-conjugation-and-transport-processes-
human-drug-metabolism-part-1/
Camille Georges Wermuth, The Practice of medicinal chemistry, 3rd edition
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Phase 2 - Reactions
Glucuronidation
• UDP-Glucosyl-Transferase (UGT)
• Transfer of glucuronic acid
• Cofactor: UDP-glucuronic acid
https://www.ciimar.up.pt/Arcopol2_2016/pa
• Linkage through: alcoholic and phenolic hydroxyl ge_15.htm

groups, carboxyl-, sulfuryl-, carbonyl-

groups or amines
• Hydrophilicity ↑, molecular weight ↑
Goodman and Gilman's The Pharmacological Basis of Therapeutics, 11th edition
Camille Georges Wermuth, The Practice of medicinal chemistry, 3rd edition
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 Phase 2 - Reactions
Enzyme TransferredGr Cosubstrate / Linkagethrou
oups Cofactor gh:
Sulfation Sulfotransferase Sulfates 3´- Hydroxylgrou Chemically reactive
(SULT) phopshoadenosi ps ofaromatic metabolites
ne-5´- and
phosphosulfate aliphaticcomp
(PAPS) ounds
Glutathioneconju Glutathione-S- Glutathione Glutathione (Tri- Thioether Reaction with reactive
gation Transferase (GST) peptide) electrophiles

N-Acetylation N- Acetyl groups Acetyl-CoA Aromatic Hydrophilicity ↓

acetytransferase( amines or
NAT) hydrazines
Methylation Methyltransferas Methylgroups S-adenosyl- Different
e (MT) methionine Heteroatoms
(SAM)

Goodman and Gilman's The Pharmacological Basis of Therapeutics, 11th edition

https://drughunter.com/resource/phase-ii-drug-metabolism/
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 Pro & Con of metabolic stability
Advantages
• Increased bioavailability (longer
t1/2)
• Lower variability in drug levels
• Diminishing the number of active
metabolites
Disadvantage:
• Elimination of „soft spots“
within the molecule may result
in a change of binding and
orientation

10.1016/S1359-6446(04)03280-5
https://doi.org/10.1016/j.apsb.2018.04.003

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 Blocking metabolically labile groups (1)

• removing or blocking the sites of metabolic soft spots

• sites potentially labile towards oxidation e.g. benzylic or allylic positions

• can be blocked:
• through the introduction of a halogen atom onto carbon atom
• or by replacing the benzylic CH2 with an isostere

• Example:
• replacing a methyl group with an acetylenyl moiety and p-fluoro substitution on the phenyl ring
led to an improvement in bioavailability

10.1016/S1359-6446(04)03280-5

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 Blocking metabolically labile groups (2)

10.1016/S1359-6446(04)03280-5

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Modification of metabolically labile groups (1)

• not only oxidations but other types of metabolism can affect functional groups
• amidases and esterases (present in liver) → hydrolyse amides and esters
respectively
• Phase II metabolism can be used as a target to improve stability
• Replacing a labile ester linkage with an amide group adds stability with respect
to esterase activity

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 Modification of metabolically labile groups (2)

ASPIRIN

10.1016/S1359-6446(04)03280-5
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Factors affecting phase 1/2
1. Biological factors:
• Age: drugs are metabolised more slowly in the neonate and elderly compared
• Diet: low protein intake; fat free diet; vitamin deficiency; starvation
• Sex difference: male > female have greater drug metabolizing capacity;
• Pathological conditions: liver disease, reduced hepatic blood flow in heart failure

1. Drug interactions:

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Factors affecting phase 1/2
Genetic polymorphism: affects plasma cholinesterase, acetylation enzymes and certain
variants of the CYP enzymes

Example: the depolarising muscle relaxant, suxamethonium & plasma cholinesterase

deficiency
• significantly slower metabolism of the drug → prolonged neuromuscular blockade (longer post-op recovery
period in ICU; need for mechanical ventilation; prolonged muscle weakness)

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Thank you for your
attention!