The document discusses drug metabolism, which is the process of modifying drugs to facilitate their excretion from the body. It highlights the role of various enzymes in modifying drug structures, the significance of liver and intestines in metabolism, and the distinction between phase I and phase II metabolic processes. Understanding drug metabolism is crucial for grasping pharmacological effects, drug interactions, and overall drug efficacy.

1. An introduction to Medicinal
chemistry
Lecture 6 & 7

2. Drugs Metabolism
• Metabolism is the process of preparing foreign chemicals for removal from the body.
• Metabolism aims at changing the structural configuration in order to promote
xenobiotic excretion.
• Drugs are usually lipophilic with ability for renal reabsorption in the renal tubules.
• A drug can be excreted easily if it is more polar.
• Various enzymes systems thus aim at altering the water solubilities to promote
excretion.

3. • Refers to enzyme-mediated biotransformations (detoxication) that alter the
pharmacological activity of both endogenous and exogenous compounds.
• metabolic reactions have also been described as detoxification processes.
• A metabolite is any intermediate or product resulting from metabolism.
• the metabolites have terminal activity, altered toxicities .
Drug
Metabolic enzymes
Metabolite

4. • Drugs undergo a variety of chemical changes in the animal organism
by enzymes of the liver, intestine, kidney, lung, plasma and other
tissues.
• Many enzymes take place in such biotransformations; oxidase,
hydrolase, lipase, synthetase, dehydrogenase, …etc
Drug
Metabolic enzymes
Metabolite

5. The importance of studying drug
metabolism:
• Understanding the pharmacological and toxicological
activity of drugs.
• The importance of shortening the drug’s duration of action.
• The complications of drug-drug interactions mainly
depends on the induction or inhibition of metabolic
enzymes

6. 6
Sites of Drug metabolism
 Drugs metabolism can occur in most tissues and
organs of the body, e.g. liver, kidney, gut, blood,
plasma.
 The liver is probably the most efficient site for
phase I metabolism.
 The kidney and gut wall are important sites for
phase II metabolic pathways.

7. Sites of drug biotransformation.
The Liver
• Is the most important organ in metabolism and
detoxification of endo- and exogenous cpds.
• It is well perfused.
• Very rich in metabolising enzymes (most of them!)
• Orally administered drugs are usually susceptible to
the first pass metabolism.
• May be significant and result in reduced oral
bioavailability.

8. Sites of drug biotransformation.
The Intestines
• Important site for extra hepatic metabolism of orally administered drugs.
• Contain CYP 3A4 isozyme  drug metabolism.
• Also contains p-glycoprotein  Drug extrusion to GIT
• Esterases are important for metabolism of ester prodrugs.
• Bacteria micro flora also produce azo and nitro reductases for activation of
prodrugs. E.g. sulfasalazine
• Intestinal enzyme is also responsible for hydrolysis of glucuronide
conjugates that are circulated in bile e.g. digoxin.

9. Sites of drug biotransformation.
Other Organs
• Kidneys
• Lungs
• Adrenal Glands
• Brains
• Placenta
• Brain
• Skin

10. Metabolism may result in:
• Pharmacologically inactive drug (detoxification effect).
Inactive or inert metabolite.
• Pharmacologically active drug (bioactivation…prodrug
approach). Active metabolite.
• Change the pharmacological activity (toxic effect). Toxic
metabolite.

12. Drug metabolism
• Can be divided into two distinct categories:
• Phase-I:
Reactions which introduce or unmask hydrophilic groups in the drug
structure (functionalisations).
• Phase-II:
Reactions which conjugate the drug or its phase-I metabolite with a
hydrophilic, endogenous species (conjugation reactions).

13. 13
Phase I metabolism: (Functionalization)
• Most often the products of phase I metabolism
are more polar than the drug molecule.
This can be achieved inside the body by:
1. Oxidations: Hydroxylation, alcohol oxidation
2. Reductions: aldehydes and ketones, nitro groups
reduction.
3. Hydrolyses: esters and amides.
4. Elimination reactions: deamination, dealkylation, and
dehalogenation.
Phase I vs. Phase II

14. Phase I/ Functionalization Reactions
• Include oxidative, reductive and hydrolytic biotransformation reactions.
• Introduce polar groups into the xenobiotic to produce a more water soluble
molecule.
• COOH
• OH
• NH2
• SH
• The products may not be sufficiently hydrophilic but are suitable precursors for
phase II/ Conjugation.

15. Phase I/ Functionalization Reactions
Modalities:
a) Direct introduction of the functional group.
• E.g. aromatic or aliphatic hydroxylation reactions
b) Modifying or ‘Unmasking’ of existing polar functional groups.
Existing functional Group New functionality
Ketones, aldehydes
(Reduction)
Alcohols
Alcohols(Oxidation) Carboxylic Acids
Esters, Amide (Hydrolysis) COOH, NH2, OH
Azo, Nitro (Reduction) NH2
N-, O-, S- (O-dealkylation) NH2, OH, SH

16. 16
• Phase II metabolism: (Conjugation)
a very polar, highly hydrophilic biomolecule (conjugating
agent) is added to the drug or its metabolite to make it
water soluble.
• the known conjugating agents are:
glucuronic acid, amino acids, sulfate, or glutathione.
• Most often these conjugates are biologically inactive and
are then excreted in urine to remove the drug from the
body.
Phase I vs. Phase II

17. Phase-II metabolism
• Involves the following conjugation reactions that are catalyzed by
transferase enzymes:
• Glucuronidation.
• Sulfation.
• Amino acid conjugation.
• Methylation.
• Acetylation.

19. Phase-I reactions
• Two general types of enzyme systems take part in these reactions:
• Microsomal Mixed Function Oxidases (MFOs)
• Flavoprotein, NADPH-monooxygenase
• Cytochrome P450
• Non-cytochrome oxidizing enzymes.
• Xanthine oxidase
• Alcohol/aldehyde dehydrogenase

20. Role of Cyp450 monoxygenases.
• They are membrane bound proteins.
• Have an approximate molecular weight of 50 kD, and contain a haeme moiety.
• There are about 30 human cytochrome P450 enzymes.
• Six, CYP1A2 CYP2C9, CYP2C19, CYP2D6, CYP2E1 and CYP3A4 are the metabolising enzymes.
• each member catalyzes the biotransformation of a unique group of drugs
• They are involved in:
• The metabolism of many drugs and dietary substances.
• Synthesis of steroid hormones and other extracellular lipid signalling molecules.

21. Role of Cyp450 monoxygenases.
Nomenclature
CYP-ARABIC NUMBER-CAPITAL LETTER ARABIC NUMBER
CYP 3A4
CYP: cytochrome P-450 enzymes
Arabic Number : Family (CYP 1, CYP 3….)
40% identical amino acid sequence
Capital Letter: Sub Family
(CYP 1A, CYP 2C, CYP 3A ….)
55% identical amino acid
sequence
Arabic Number: Individual
Enzyme (CYP 1A2, CYP 2C9,
CYP 3A4 ….)
>90% identical amino acid
sequence

22. CYP Biotransformations
• Chemically diverse small molecules are
converted, generally to more polar compounds
• Reactions include:
• Aliphatic hydroxylation, aromatic hydroxylation
• Dealkylation (N-,O-, S-)
• N-oxidation, S-oxidation
• Deamination
• Dehalogenation

23. Oxidation.
• Are by far, the most common and important in drug metabolism.
• Normally the first step of drug metabolism
• Mainly carried out by mixed function oxidases.

25. 25
Common metabolic oxidations:
According to the chemical nature of the drug
molecule:
1.1) Aromatic hydroxylations.
1.2) Aliphatic hydroxylations.
1.3) Epoxidation of alkenes.
1.4) Oxidation of alcohols to aldehydes.
1.5) Oxidation of aldehydes to carboxylic acids
1.6) Oxidation of sulfide to sulfoxides to sulfones
1.7) Oxidation of imines to imine oxides
1.8) Dealkylation on heteroatoms
Phase I Metabolism

26. Oxidation of Aromatic Moieties
• Aromatic hydroxylation refers to mixed function
oxidation of aromatic cpds (arenes).
• They corresponding end products being arenols.
• Proceed initially through an epoxide intermediate
known as an “arene oxide”
• This undergoes spontaneous rearrangement to form
an arenol in most cases.
• Arene oxides are important in metabolic,
toxicological and formation of arenols.

27. 27
• Many drugs possess aromatic rings.
• Hydroxylation of these rings is a major route of drug
metabolism.
• Hydroxylation occurs most readily on rings that are
electron-rich, i.e those that have electron-donating
groups (EDG) directly attached to the ring (OH, OCH3,
NH2, alkyl groups).
• The presence of electron-withdrawing groups (EWG)
on the ring usually inhibits hydroxylation on that ring.
Examples of such groups include halides (F, Cl, Br, I),
nitro, carbonyls, sulfoxides and sulfones.
• Aromatic hydroxylation most often occurs at the para
position to the EDG.
1.1 Aromatic Hydroxylation

28. Aromatic hydroxylation
• The least substituted aromatic ring will be favorably
oxidized, especially at the least hindered carbon atom
• The activated ring will be better oxidized (the ring
bearing an electron donating group)

29. 29
S
N
CH2CH2CH2 N(CH3)2
Cl
Electron-withdrawing
group
Electron-donating
groups
Hydroxylation on this
ring is favored
Hydroxylation on this
ring is inhibited
• While the positions ortho- to the substituent are also
electron-rich, steric crowding usually inhibits
hydroxylation at these positions.
Aromatic Hydroxylation

30. • The least substituted aromatic ring will be favorably
oxidized, especially at the least hindered carbon atom
N
N
Cl
Meclizine
Antiemetic agent
N
N
Cl
OH
Major metabolite

31. • The activated ring will be better oxidized (the ring
bearing an electron donating group)
H
N
H
N
N
Cl
Cl
Clonidine
No aromatic hydroxylation

32. Oxidative Phase-I involving cytochrome P-450
enzymes:
• Aromatic hydroxylation:

33. • Aromatic epoxidation:
Benzo[a]pyrene
O
NH
N
N
N
N
O
Covalently bound
Deoxyguanosine adduct
(systemic toxicity)
O
HO
OH
Deoxyribose
NH
N
N
N
N
O
Deoxyribose
DNA
It is Adenine (base) present in DNA

34. Mixed function oxidase enzyme (MFO)
1.2 Metabolic Oxidation of Alkenes
NH
H
N
CH3CH2CH2CH
O O
O
H3C
MFO
NH
H
N
CH3CH2CH2CH
O O
O
H3C
O
NH
H
N
CH3CH2CH2CH
O O
O
H3C
HO
HO
Secobarbital
Epoxide
hydrolase
Not Observed Isolated metabolite
N
O
NH2
Carbamazipine
N
O
NH2
O
N
O
NH2
HO
OH
HN
N
H
O
O O
Secobarbital
HN
N
H
O
O O
OH
HO
Secodiol

35. 35
1.3 Metabolic Oxidation of Alkyl Groups
• Aliphatic carbons are also subject to metabolic oxidation.
• The product of these oxidation reactions is an alcohol.
• The carbon undergoing metabolism must have at least one
attached hydrogen.
• These oxidations can be categorized as:
1) α-Oxidations:
Oxidation of aliphatic carbons that are adjacent to a functional
group.
The result of these oxidations is that an oxygen atom is inserted
into a C-H bond to give a C-OH group.
CH2 R
O
C
R CH2 R CH2 R
S
O
O
R CH2 R
O2N CH2 R C CH2 R
N

36. 36
N
N
O
H3C
Cl
MFO
NH
H
N
O
O
O
CH3CH2CH2CH
H3C
MFO
N
N
O
H3C
Cl
OH
NH
H
N
O
O
O
CH3CH2CH2CH
H3C
OH
-carbon
-carbon
Metabolic Oxidation of Alkyl Groups

37. 37
CO2H
CH3CH2CH2
CH3CH2CH2
MFO
CO2H
CH3CH2CH2
CH2CH2CH2
HO
CO2H
CH3CH2CH2
CH3CHCH2
HO
Valproic Acid
-carbon
(-1)-carbon
+
Product of -oxidation Product of (-1)-oxidation
2) ω-oxidation
Oxidation of aliphatic carbons at or near the end of a
chain of aliphatic carbons.
ω-oxidation: oxidation occurs at the terminal carbon of
the chain.
ω-1 oxidation: occurs on the carbon preceding the
terminal C-atom (penultimate carbon
oxidation).

38. Oxidation of benzylic carbons
• The carbons directly attached to aromatic rings are oxidized to
aldehydes and carboxylic acids via alcohols.
• Read about oxidation of olefins
R R
R

39. 39
1.4 Metabolic Oxidation of Alcohols
• The enzyme alcohol dehydrogenase catalyzes the oxidation of alcohols to
carbonyl compounds.
• Primary alcohols are oxidized to aldehydes and secondary alcohols are oxidized
to ketones.
• Aldehydes are highly reactive and undergo rapid metabolic oxidation to
carboxylic acids.
• Tertiary alcohols and ketones cannot be further oxidized?.

40. 40
CH3
CO2H
HOH2C
CH3
CH3
CO2H
H3C
CH3
MFO
MFO
CH3
CO2H
H3C
CH3
HO
CH3
CO2H
HO2C
CH3
CH3
CO2H
OHC
CH3
Ibuprofen
Product of (-1)-oxidation
Product of -oxidation
Alcohol
dehydrogenase
Aldehyde (not observed)
Aldehyde dehydrogenase
Major metabolite
Metabolic Oxidation of Alcohols
* Mixed function oxidase enzyme (MFO)

41. Other phase-I metabolic enzymes
• Alcohol dehydrogenase and aldehyde dehydrogenase:
R
OH
R
O
H R
O
OH
Alcohol
dehydrogenase
Aldehyde
dehydrogenase
R
O
R
No oxidation

43. 43
1.5 Metabolic Oxidation of Sulfides and Sulfoxides
• Sulfides can be metabolically oxidized by MFOs to sulfoxides and to sulfones.
• Sulfoxides can be oxidized to sulfones.
• Sulfones, in which sulfur is already in its highest oxidation state cannot be
oxidized any further.
S
N Cl
CH2CH2CH2 NMe2
S
N Cl
CH2CH2CH2 NMe2
O
S
N Cl
CH2CH2CH2 NMe2
O O
MFO
MFO
MFO

45. C-N Metabolism

46. 46
1.6 Metabolic Oxidation at sp2
Nitrogen
• The MFO system is also capable of oxidizing sp2
nitrogen as found in imines and certain aromatic
heterocycles such as pyridine and quinoline.
N
MFO
N
N
Cl
O
H3C
MFO
N
O
N
N
Cl
O
H3C
O

48. Oxidative Phase-I involving cytochrome
P-450 enzymes:
• N-oxidation:
• Mostly for primary and secondary amines as well as
aromatic amines:
• This gives N-oxide that will be rapidly converted to
hydroxylamines.

49. R NH2 R N
H
OH
R NO2
NH2
H
N
OH
N
O
proteins and nucleic acids
NO2
N
OH
Protein
Oxidize Fe+2
in
hemoglobin to Fe+3
(methemoglobin or ferrhemoglobin)
this form is no longer capable to transport oxygen
(methemoglobinemia toxicity)
N
O

 

50. 50
1.7 Metabolic Dealkylation
• Many drugs contain a heteroatom such as nitrogen,
oxygen, or sulfur that is attached to an alkyl group.
• When such drugs undergo metabolism these alkyl
groups may be removed in a process called N-(or O- or
S-)-dealkylation.
S
N
CH2CH2CH2 N
CH3
CH3
S
N
CH2CH2CH2 N
CH3
H
-CH2O
-CH2O
S
N
CH2CH2CH2 N
H
H

51. N-Dealkylation
(1O
/ 2O
Amines )
In the case of primary or secondary amines,
dealkylation of an alkyl group starts at the carbon
adjacent to the nitrogen;

52. 52
• Primary amines may undergo deamination.
• If the carbon on the other side of the nitrogen contains a
hydrogen, the C-N bond can be cleaved to give an
aldehyde or ketone. This process is called oxidative
deamination.
S
N
CH2CH2CH2 N
H
H
-NH3
S
N
CH2CH2CH
O
S
N
CH2CH2CO2H
MFO
Note: Aldehydes are usually rapidly
oxidized to acids via the MFO system.

53. N-Deamination.
(3O
Amines )
• In the case of tertiary amines, with hydroxylation of the nitrogen.
• The intermediate products are labile and break up into the
dealkylated amine and aldehyde.

54. • N-dealkylation:
HN R
OH
NH2
R
O
H

55. N-dealkylation
Cl
N
N
C6H6
Cl
N
NH
C6H6
N-t-butylnorchlorcyclizine norchlorcyclizine
Br
N
Br
NH2
Br
O
OH
Brompheniramine

56. Oxidative Phase-I involving cytochrome P-450
enzymes:
• Oxidative deamination:

57. O-Dealkylation
• O-Dealkylation of drugs possessing C—O bond involves hydroxylation of α-carbon
to form an unstable hemiacetal or hemiketal intermediates.
• These intermediates spontaneously cleave to form alcohol and carbonyl
compound.

58. • O-dealkylation:
O
N
O
OH
CH3
O
N
HO
OH
Oxidative demethylation
Codeine Morphine

59. S-Dealkylation
• S-Dealkylation involves oxidative cleavage of alkyl carbon-sulfur
bonds.

60. Other phase-I metabolism
• Sulfoxidation: by flavin monooxygenase
N
S
N
Chlorpromazine
N
S
N
O
N
S
NH2
O
N
S
O
OH
O
Deaminated metabolite Dealkylated metabolite

61. Reductive Reactions
• Drugs containing carbonyl, nitro, and azo groups are metabolized by
reduction to alcohols and amines respectively.
• The reduced compounds are conjugated and eliminated from the
body.
Read about the reduction of chloral hydrate

62. 2. Metabolic Reductions
• Functional groups that most typically undergo
metabolic reduction include:
– Ketones - Nitro groups - Azo groups
• Less commonly, aldehydes and sulfoxides can be
reduced.

63. 63
2.1 Metabolic Reduction of Carbonyl Groups
• Of all of the various functional groups that contain a
carbonyl unit?, only ketones and aldehydes usually
undergo metabolic reduction to alcohols.
O
C
R H
O
C
R OH
OH
C
R H
H
Aldehyde
Primary Alcohol
Carboxylic Acid
Aldehyde
dehydrogenase
Aldo-reductase
Major
Minor
Metabolic Oxidation
Product
Metabolic
Reduction
Product

64. 64
• Ketones however, are resistant to further oxidation. They
are readily reduced metabolically to secondary alcohols.
O
C
R R' OH
C
R R'
H
Ketone
Secondary Alcohol
Keto-reductase
Major
Metabolic
Reduction
Product
X
Further oxidation of
ketones is not possible

65. N
O
HO
OH
O
H
Naloxone
CH2 CH CH2
keto-reductase
N
OH
HO
OH
O
H
CH2 CH CH2

66. 66
O
C
H
N
O2N
N NH
O
O
Dantrolene
Nitro-reductase
O
C
H
N
H2N
N NH
O
O
2.2 Metabolic Reduction of Nitro Groups
Aminodantrolene

67. 67
2.3 Metabolic Reduction of Azo Groups
• Bacteria within the intestine have azo-reductases that can reduce azo
compounds to amines.
N
N
OH
CO2H
HO
CO2H
Intestines
(bacterial
Azo-reductase)
NH2
H2N
OH
CO2H
HO
CO2H
+
Olsalazine
5-Aminosalicylic Acid
(Active metabolite)

68. Other phase-I metabolism
• Azoreduction:
N
N
COOH
OH
S
H
N
N
O
O
Azulfidine
NH2
S
H
N
N
O
O
H2N
COOH
OH
P-aminosalicylic acid
Sulfapyridine
Antibacterial action Anti-inflammatory action

69. 69
2.4 Metabolic Reduction of Sulfoxides
• As with aldehydes, sulfoxides are most commonly
oxidized to sulfones by metabolism. Occasionally they
may undergo metabolic reduction to sulfides.
O
S
R'
R
O
S
R'
R
O
S
R'
R
Oxidation
Most
Common
Sulfone
Less
Common
Reduction
Sulfide

70. 70
H
S
O
H3C
CH2CO2H
F
H
S
H3C
CH2CO2H
F
CH3 CH3
Sulindac
(Pro-drug form)
Active Metabolite
Metabolic
reduction
Sulfo-reductase

71. 71
3. Hydrolysis
• Hydrolysis is the process of breaking bonds by the
addition of water.
• Functional groups that are most often metabolized by
hydrolysis include esters (and lactones) and amides
(and lactams).
• Hydrolysis of esters ALWAYS results in two products –
a carboxylic acid and an alcohol.
O
C
OR'
R
H2O
catalyst
O
C
OH
R
+ R'OH

72. 72
• Hydrolysis of amides ALWAYS results in two products –
a carboxylic acid and an amine.
O
C
NR'2
R
H2O
catalyst
O
C
OH
R
+ R'2NH

73. Hydrolytic phase-I metabolism
• Hydrolysis is also observed for a wide variety of drugs.
• The enzymes involved in hydrolysis are esterases, amidases, and proteases.
• These reactions generate hydroxyl or amine groups, which are suitable for phase II conjugation.
• By non-specific esterase and amidase enzymes that present in plasma, gut, liver and kidney.
• It has a beneficial role in most of prodrugs that after hydrolysis inside the body release the
active form of the drug.

74. Ester vs. Amide bond
Ester bond is relatively weaker than amide bond, it will be rapidly
hydrolyzed by esterase enzyme
R O
O
R N
H
O
The reactivity of ester and amide bond depend on how much the carbonyl carbon is
electropositive
Nitrogen atom is less electronegative than oxygen, so it will be waeker electron withdrawing
atom
therefore, the crabonyl carbon attached to oxygen atom will be more electropositive, and
more reactive toward nucleophilic attack of water molecule during hydrolysis.

75. Nucleophilic attack of hydroxide anion on
ester and amide
R O
O
R N
H
O
OH OH

76. Example
Procaine
Short acting local anesthetic
Procainamide
Long acting antiarrhythmic
T1/2 = 2.5-4.5 hr
T1/2 = 40-84 second

77. Hydrolytic phase-I metabolism
• Examples of prodrugs activated by hydrolytic
enzymes:
• Dipivefrine: is a di-tertbutylcarboxy ester of adrenaline….
More lipophilic… better penetration through the corneal
membrane….then will be hydrolyzed to give the active form
(adrenaline)

78. Why Dipifevrine has been prepared?
• Adrenaline is a polar drug….difficult access into the ocular
cavity.
• Adrenaline has a generalized adrenergic effect…. Many side
effects such as increase blood pressure, heart rate and
bronchodilation.
• Dipifevrine is more lipophilic, better penetration… localized
effect.
HO
HO
H
N
OH
Adrenaline

79. General notes regarding phase-I
metabolism
• Hydrolysis normally catalyzed by carboxylesterases:
• Cholinesterase…. Hydrolyzes choline-like esters (such as
succinylcholine), procaine and acetylsalicylic acid.
• Arylcarboxyesterase.
• Liver carboxyesterase
O
O
O
O
N
N
Syccinylcholine
OH
O
OH
O
N
HO
N
COOC2H5
HN
COO

80. HN
COOCH3
HN
COO
Zwitter ionic
Polar
Easily excreted

81. General notes regarding phase-I
metabolism
• Esters that are sterically hindered are hydrolyzed
more slowly and may be appeared unchanged in
urine:
OH
O
O
N
Atropine
50% excreted unchanged in urine

82. OH
O
O
N
Atropine

83. General notes regarding phase-I
metabolism
• Amides are more stable to hydrolysis than
esters….large fraction of amide containing drugs are
normally excreted unchanged.
H2N
N
H
O
N
Procainamide
60% excreted unchanged in urine

84. H2N
O
O
N
Procaine
H
N
O
N
Lidocaine
Procaine has a short duration
of anesthesia
lidocaine has a long duration
of anesthesia

85. CH3
CH3
H
N
O
N
Lidocaine
Nucleophilic attack of HO-

86. Phase II/ Conjugation Reactions
Overview
• Attach small, polar and ionisable, endogenous, functionalities to
• handles of phase metabolites.
• Parent compounds with existing suitable functional groups.
• Functionalities include:
• Glucuronic acid
• Sulphate
• Glycine
• Amino Acids

87. Phase II/ Conjugation Reactions
Over View
• The conjugated metabolites are:
• Easily excreted.
• Generally devoid of pharmacological activity.
• Generally devoid of toxicities.
• Some phase II reactions may attenuate or terminate biological activity.
• Methylation.
• Acetylation
• Glutathione (GSH) conjugation protects the body against toxic metabolites.

88. 88
Phase II Metabolism
Conjugation Reactions
During phase II metabolism either the parent
Compound or its phase I metabolites are
converted into more water soluble entities that
can then be excreted
Common classes of phase II metabolisms include
:
1) Glucuronide conjugates
2) Sulfate conjugates
3) Glycine and glutamate conjugates
4) Glutathione conjugates
5) Acetylation
6) Methylation

89. 89
1
)
Glucuronide Conjugates
 The most common phase II
metabolites
 Functional groups susceptible to glucuronidation include: alcohols and phenols,
carboxylic acids, amines, thiols
O
HO2C
HO
HO OH
OH
-Glucuronic Acid
O
N
H
H
HO
CH3
OH O
N
H
H
CH3
OH
O
O
HO2C
HO
HO
OH
UDPGA
Morphine
UDPGA: Uridine-5-diphospho-α-D-glucuronic acid
Activated Coenzyme acts as carrier for the glucuronic

90. Glucuronidation
• Glucuronidation involves conjugation of metabolite or drug molecule with
glucuronic acid.
• In these reactions glucuronic acid molecule is transferred to the substrate from a
cofactor.
• Glucuronides are generally inactive and are rapidly excreted into the urine and bile.
• Molecules that undergo Glucuronidation are associated with:
• phenolic hydroxyl,
• Alcoholic hydroxyl, and
• carboxylic acid groups.

91. 9
1
2
)
Sulfate Conjugates
 Sulfate conjugates are formed mainly from phenols,
although they can also be formed from alcohols and
aromatic amines.
 There is less available sulfate in the body than there is
glucuronic acid.
 The coenzyme, that acts as sulfate carrier is: 3’-
phosphoadenosine-5’-phospho-sulfate (PAPS) is
responsible for transferring a sulfate to a suitable substrate.

92. Sulfation
• Sulfate conjugation involves transfer of a sulphate molecule from a
cofactor (to the substrate (metabolite or drug moiety) by the enzymes
(sulfotransferases).
• Substrate molecules include:
• Alcoholic hydroxyl,
• phenolic hydroxyl and
• aromatic amine groups.

93. 93
3
)
Glycine and Glutamine Conjugates
 Glycine and glutamine form conjugates with
carboxylic acids.
 The products are carboxamides, and are more water-
soluble than the carboxylic acids.
O
C
R N
H
CO2H
R'
R' = H (glycine)
R' = CH2CH2CONH2 (glutamine)
O
C
R OH
Carboxyamides
H2N
CO2H
R'
ATP
HSCoA
CO2H
O
H3C O
CO2H
OH
C
OH
O
N
H
CO2H
Acetylsalicylic Acid
(Aspirin)
Salicylic Acid Glycine Conjugate
Hydrolysis
(Phase I) Phase II

94. 4
)
Glutathione Conjugates
 Glutathione (GSH) conjugation is a means by which
the body can detoxify reactive electrophilic species.
 Glutathione possesses a nucleophilic thiol (SH)
group that can react with electrophiles before they
react with other electrophiles such as those that
belong to critical components, including proteins
and DNA.
H
N
H
O
NH2
CO2H
HS
H
NH
O
Glutathione
CO2H
Glutamic acid
Cysteine
Glycine
Nucleophile

95. 95
H
N
H
O
NH2
CO2H
HS
H
NH
O
CO2H
H
N
H
O
NH2
CO2H
S
H
NH
O
CO2H
E
Electrophile (E)
GST
1
)
The reaction of glutathione with
electrophiles is catalyzed by the
enzyme Glutathione S-transferase
(GST)
.
Glutathione
conjugate

96. Glutathione conjugation
• For electrophilic drugs and metabolites (Detoxication):
G-SH
O
R R
SG
H
H H H
OH
G-SH O OH
SG
HS
N
H
O COOH
HN
H O
COOH
NH2
Glutathione (tripeptide: glutamate+cysteine+glycine)

97. Detoxication by glutathione adduct
formation
OH
O
O
N
O GSH
OH
O
O
N
GS
HO
[O]
O
Arene oxide
OH
SG

98. Glutathione conjugation
• Toxicity of aromatic compounds came from the
formation of arene oxide during the metabolism that
will be attacked by endogenous nucleophile such as
proteins, DNA or RNA.

99. Glutathione conjugation
O
O
H
H
NIH
Shift
OH
H2O
Or epoxide hydrolase
OH
OH
GSH
OH
SG
DNA, RNA, proteins
OH
M

100. 100
5
)
Acetylation
 Primary amines undergo acetylation of the amino group to
give acetamides.
 Primary amines are often formed from phase I reduction of
aromatic nitro groups.
NH2
acetyl-CoA
N-acetyltransferase
NH
CH3
O
HN
N
NH2
HN
N
NH
CH3
O
Histamine
(aliphatic amine)
H2N CO2H HN CO2H
H3C
O
p-Aminobenzoic Acid
(aromatic amine)

101. 101
6
)
Methylation
 Methylation is a relatively minor phase II metabolic pathway.
 Among the groups that undergo this reaction are phenols, catechols (ortho-
dihydroxyaromatic compounds), amines, and thiols.
 Methylation does not increase water solubility, but it does usually render the
metabolite biologically inactive.
 S-Adenosylmethionine (SAM) acts as the methylating agent and the reaction is
catalyzed by various methyltransferases.
CH3
NH2
CO2H
HO
HO
S-(-)--Methyldopa
CH3
NH2
CO2H
MeO
HO
COMT
SAM
catechol COMT = catechol O-methyl transferase

102. FACTORS AFFECTING DRUG METABOLISM
1. Age-related differences:
 Newborn suffer from underdevelopment or deficiency of oxidative and conjugative enzymes leading to
reduction of metabolic capacity
o oxidative (cytochrome P-450) metabolism of tolbutamide is reduced in newborn leading to increased t ½
(40 hours Vs 8 hours in adults).
o Glucuronyltransferase activity reduction leading to the reduction in chloramphenicol conjugation with
glucuronic causing the accumulation of toxic levels of the drug causing gray baby syndrome.
 Elderly patients show evidence of inefficient drug metabolism due to compromised liver function
2. Sex differences:
 Diazepam has an average half-life of 41.9 hours in females but only 32.5 hours in males.
 Women have a significantly lower concentration of alcohol dehydrogenase and so do not metabolise alcohol so
rapidly as men.
102

103. FACTORS AFFECTING DRUG METABOLISM
3. Genetic Factors:
 Variations in the genetic codes of individuals can result in the absence of enzymes, low concentrations of
enzymes or the formation of enzymes with reduced activity.
 Acetylation shows rate differences in human
 Rapid acetylators undergo fast elimination leading to inadequate therapeutic response to the drug.
 Slow acetylators are subject to toxicity due to accumulation of the drug.
o For example, the antituberculous drug isoniazid (acetylation). Slow acetylation is found in 75% of
Caucasians and Negroes but in only 10 % of Japanese and Eskimos.
103

104. FACTORS AFFECTING DRUG METABOLISM
4. Species differences:
 Strain differences in metabolism may also take place e.g. oxidative deamination or aromatic hydroxylation are
two metabolic pathways of amphetamine.
• In human, guinea pig and rabbit oxidative deamination predominates.
• In rats aromatic hydroxylation is the predominates.
 There is also some difference due to presence or absence of the particular transferase enzyme e.g. cats lack
glucuronyl transferase while pigs lack sulfotransferase enzyme and thus unable to conjugate phenols .
 A dose of 50 mg/kg of body mass of hexobarbitone will anaesthetise humans for several hours but the same
dose will only anaesthetise mice for a few minutes.
104

105. 5.Enzymes
Enzyme induction:
• Coadministration of warfarine (anticoagulant) ((largely eliminated through liver metabolism)) and the hypnotic
phenobarbitone (efficient enzyme enducer) leads to marked decrease in anticoagulant activity of warfarine.
Enzyme inhibition
• Phenylbutazone stereoselectively inhibits the metabolism of the more potent (S)(-)- enantiomer of warfarine.
This explains the increased hypoprothrombinemia and hemorrhaging in patients taking both warfarine and
phenylbutazone.
6. Miscellaneous factors affecting drug metabolism:
o Dietary factors.
o Physiologic state of the liver (Hepatic cancer, cirrhosis, hepatitis).
o Pregnancy, hormonal disturbances (thyroxin, steroids).
o Environmental factors: Cigarette smoke produces polynuclear aromatic hydrocarbons. CYP1A2 metabolises the
polynuclear aromatic hydrocarbons to carcinogens responsible for lung and colon cancer.
FACTORS AFFECTING DRUG METABOLISM
105