Download Metabolism Phase-I

‫‪Drug metabolism‬‬
‫د‪ .‬محمد نورالدين محمود‬
Brief introduction
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The body treats drugs as foreign substances and has methods of getting rid of such chemical
invaders.
If the drug is polar, it will be quickly excreted by the kidneys. However, non-polar drugs are not easily
excreted and the purpose of drug metabolism is to convert such compounds into more polar
molecules that can be easily excreted.
Change the structure of drug to more polar molecule usually affects the drug interaction with specific
and non-specific receptors available in the tissues.
The metabolism is carried out by two sets of reactions (i.e. phase-I and phase-II).
Both sets of reactions can also be regioselective and stereoselective. This means that metabolic
enzymes can distinguish between identical functional groups or alkyl groups located at different
parts of the molecule (regioselectivity), as well as between different stereoisomers of chiral
molecules (stereoselectivity).
Brief introduction
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Drug metabolism or biotransformations are the chemical reactions that are responsible for the
conversion of drugs into other products (metabolites) within the body before or after they have
reached their sites of action (i.e. wherever drug is in free form).
Drug
metabolism or
biotransformations
Metabolites
It is thought that biotransformation of molecule is intended
1. Directly to increase molecule polarity  increase molecular excretion
2. Indirectly affects molecular interaction with specific and non-specific receptors  molecular activity
Since metabolism involve enzymesubstrate interaction, the properties of
specificity is involved i.e. enzymes can
distinguish between identical functional
groups or alkyl groups located at different
parts of the molecule (regioselectivity), as
well as between different stereoisomers of
chiral molecules (stereoselectivity).
The outcomes of metabolism
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Metabolism of a compound is the chemical modification of the compound molecular structure using
enzymes (biotransformation)
Therefore, changing the molecular structure of a drug by metabolism will have effects on dynamical
behavior of drug interaction with receptors and –consequently- the kinetical behavior of the
molecule.
Therefore, changes in drug molecular structure may affect:
1.
2.
3.
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Drug interaction with target receptor (pharmacological effect)
Drug interaction with cell membranes and active transporters (elimination)
Drug interaction with specific and non-specific receptors in tissue (tissue distribution and half life)
Brief introduction
Distribution
(Site of action - target
cells, tissues, receptors)
Absorption
Highly
hydrophilic
(Oral, Topical,
IV, IM, SC, IP,
Inhalation)
Elimination
(Urine, other
excretory
fluids)
Metabolism
(Chemical change of
drug
metabolites)
Produce pharmacologically
toxic metabolite
Produce pharmacologically
inactive metabolite
Produce pharmacologically
less-active metabolite
Produce pharmacologically
more-active metabolite
Activate an inactive drug
(PRODRUG)
Oral absorption
Hydrophilic
Lipophilic
Bile excretion
Renal elimination
Hydrophilic
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If the drug is polar enough, part of it
will be directly excreted
If the drug already containing polar
functional group, it can be directly
conjugated (phase-II)
If the drug after functionaliztion
(Phase-I) becomes polar enough, it
will be directly excreted.
Otherwise, the drug need to be
functionalized, conjugated then
excreted
DRUG
PHASE-I
PHASE-II
EXCRETION
What does metabolism do to drug molecule?
Less active
Receptor
Inactive
(prodrug)
Unwanted
receptor
Active
What does metabolism do to drug molecule?
A) Biotransformation may produce a pharmacologically inactive metabolite which is readily excreted.
Hydrolysis of
 procaine (local anesthetic drug) by esterases or
 procainamide (antiarrhythmic agent) by amidases
What does metabolism do to drug molecule? (Cont.)
B) Biotransformation may produce a pharmacologically less active metabolite.
Cytochrome P450 oxidative N-dealkylation of
propranolol to nor-propranolol : anti-hypertensive drug
morphine to nor-morphine : opiate analgesic drug
What does metabolism do to drug molecule? (Cont.)
C) Biotransformation may produce a pharmacologically more active metabolite.
 Cytochrome P450 oxidative O-dealkylation of codeine to morphine
 Esterase hydrolysis of diamorphine to morphine
What does metabolism do to drug molecule? (Cont.)
D) Biotransformation may activate an inactive drug (Prodrug).
Hydrocortisone: a steroid hormone
Hydroxycamptothecin: anticancer agent
Both drugs has low aqueous solubility so
are linked to hydrophilic group to make it
water soluble suitable for injection
What does metabolism do to drug molecule? (Cont.)
E) Biotransformation may activate an pharmacologically toxic metabolites.
 Thalidomide is an anti-nausea and sedative
drug that was introduced in the late 1950s
to be used as a sleeping pill, and was quickly
discovered to help pregnant women with
the effects of morning sickness.
 It was sold until 1962, when it was
withdrawn after being found to be a
teratogen, which caused many different
forms of birth defects.
 More than 20,000 children in 46 countries
were born with deformities such as
phocomelia as a consequence of
thalidomide use.
Racemisation of R-thalidomide to S-thalidomide by isomerases
Significance of Drug Metabolism
Knowing drug metabolism helps us to:
A. Know the rate of drug metabolism: which controls the duration and intensity of the action of
many drugs by controlling the amount of the drug reaching its target site.
B. Know the products of drug metabolism: which controls the activity of the drugs whether
being inactivated (detoxified) or activated (as in prodrugs).
C. Know the competitive and uncompetitive interaction during metabolism: which controls
drug-drug interactions
D. Know the possible products of drug metabolism: which need to be documented for newly
discovered drugs.
Drug metabolism has great importance in medicinal chemistry because it influences the
deactivation, activation, detoxification and toxification of the vast majority of drugs.
Sites of metabolism
 Liver
 Primary site!
 Highly perfused organ
 Rich in enzymes
 Acts on endogenous and exogenous compounds
 First pass effect!
 Extrahepatic metabolism sites
 Intestinal wall
 Sulfate conjugation
 Esterase and lipases - important in prodrug metabolism
i.e. β-glucuronidase enzymes – hydrolyze glucuronides for reabsorption (Enterohepatic
recirculation)
 Bacterial flora
 Reduction of Aromatic nitro and azo compounds the biotransformations that they
 Lungs, kidney, placenta, brain, skin, adrenal glands carry out are often more substrate selective and more
limited to particular types of reaction (e.g., oxidation,
 Limited ability and largely unknown

glucuronidation).
Sites of Drug Metabolism Reactions
The primary site for drug metabolism is liver
Others are- Kidney, intestine, lungs, plasma
Types of Drug Metabolism Reactions
 Drugs metabolism reactions can be divided into two distinct
categories or phases:
1. Phase – I Reactions
2. Phase – II Reactions
1. Phase – I or Functionalization Reactions
2. Phase – II or
A. OXIDATION REACTIONS
A. Glucuronic acid conjugation.
Oxidation of aromatic moieties.
Oxidation of olefins.
Oxidation at benzylic, allylic carbon atoms, and
carbon atoms α to carbonyl or imines.
Oxidation at aliphatic and alicyclic carbon atoms.
Oxidation involving carbon-hetero atom systems.
Carbon-nitrogen systems (N-dealkylation, oxidative
deamination, N-oxide formation, N-hydroxylation).
Carbon-oxygen systems (O-dealkylation).
Carbon-sulfur systems (S-dealkylation,
S-oxidation, desulfuration).
Oxidation of alcohols and aldehydes.
Other miscellaneous oxidative reactions.
B. REDUCTION REACTIONS
Reduction of aldehydes and ketones.
Reduction of nitro and azo compounds.
Other miscellaneous reduction reactions.
C. HYDROLYSIS REACTIONS
Hydrolysis of esters and amides.
Hydration of epoxides and arene oxides.
Conjugation Reactions
B. Sulfate conjugation.
C. Acetylation.
D. Methylation.
E. Conjugation with glycine, glutamine, and other a.a.
F. Glutathione or mercapturic acid conjugation.
Phase – I Reactions
 Reactions which introduce or unmask a polar functional group (e.g., OH, COOH,
NH2, SH) into the molecule to produce a more water soluble compound.
 The compound now either be polar enough to be excreted or may undergo phase-II
reactions
 In this step drugs undergoes functionalization reaction of oxidation, reduction or
hydrolysis.
 Phase-I oxidation reactions are catalyzed by the superfamilies of
 cytochrome P450s (CYPs),
 flavin-containing monooxygenases (FMO),
 epoxide hydrolyses (EHs).
Phase – II Reactions
 Reactions which attach/conjugate polar or hydrophilic endogenous compounds to the
functional groups of phase-I metabolites or parent compounds that already have suitable
functional groups to form water soluble conjugated products, thereby facilitating drug
elimination.
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 Reactions which conjugate polar or hydrophilic endogenous compounds to the drugs or its
metabolites to form water soluble conjugated products, thereby facilitating drug
elimination.
 Conjugated metabolites have increased molecular weight, improved water solubility and
are generally devoid of any pharmacological activity and toxicity in human.
 Phase-II reactions are catalyzed by several superfamilies of
 UDP-glucuronosyltransferases (UGT),
 Sulfonyltransferases (SULT),
 N-acetyltransferases (NAT), and
 Methyltransferases (MT).
Differences between phase-I and phase-II reactions
PHASE – I REACTIONS
PHASE – II REACTIONS
1 Functionalization reactions
Conjugation reactions
2 Metabolites may or may not have
increased molecular weight
Metabolites always have increased molecular
weight
3 Metabolites may undergo Phase-II
conjugation reaction
Metabolites never undergo Phase-I
4 Metabolites may be pharmacologically
less active or inactive
Metabolites are generally pharmacologically
inactive
5 May produce toxic metabolites
Generally does not produce toxic metabolites
Functionalization reaction
6 It may activate an inactive drug (prodrug) It does not activate an inactive drug
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The reactions in Phase-I are carried by enzymes which can accept foreign compound (whether drug
or other molecule) as substrate
Therefore, if drug can bind to enzyme it will be metabolized otherwise it will not.
The drug may have affinity to more than single enzyme, thus the drug will have more than single
metabolite.
Note the products may be
different if the sequence of
metabolism is different
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The enzymes belong to Phase-I can:
- Add polar functional groups to a wide variety of drugs.
- Unmask polar functional groups which might already be present in a drug. E.g. demethylate a methyl ether
to reveal a more polar hydroxyl group.
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Once the polar functional group has been added, the overall drug is more polar and water soluble,
and is more likely to be excreted when it passes through the kidneys.
The enzymes involved are mainly non-specific enzymes which are present in liver (E.g. cytochrome
P450 enzymes) or gut wall, plasma and other tissues (e.g. estrases, amidases)
The drug may have different affinities to different enzymes. If the affinity is higher toward enzyme-1
than enzyme-2, the drug metabolite-1 will usually be higher than metabolite-2
𝐾𝑑1
In addition, each type of enzyme is available in different
+
copies with minor differences in structure
that lead to minor differences in substrate specificity
E.g. enzyme-1 is available in isoforms A, B and C.
+
𝐾𝑑2
Affected drug functional groups during phase-I
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Some functional groups of drugs are most prone to oxidation, e.g.
1.
2.
3.
4.
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Some are prone to reduction e.g.
1.
2.
3.
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N-methyl
Aromatic rings
Terminal alkyl chain
Least hindered alicyclic rings
Nitro
Azo
Carbonyl
Some are prone to hydrolysis
1.
2.
Ester
Amide
Important enzymes involved in phase-I
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Two general types of enzyme systems take part in these reactions:
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Microsomal Mixed Function Oxidases (MFOs)
• Flavoprotein, NADPH-monooxygenase
• Cytochrome P450
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Non-cytochrome oxidizing enzymes.
• Xanthine oxidase
• Alcohol/aldehyde dehydrogenase
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Reductases
Estrases, amidases
Different metabolites are obtained from different orientations of drug in enzyme
binding site.
Drug
Drug Bind with
Orinetation-1 and 𝐾𝑑1
Metabolite-1
Drug
Drug
Drug Bind with
Orinetation-2 and 𝐾𝑑2
Drug
Drug
Drug
Q) How a drug can
have different
metabolites, even if
it is bind to a single
enzyme?
Metabolite-2
Drug cannot bind with Orinetation-3
Thus gives no metabolite at this site
A) The drug binds to
the enzyme through
different
orientations
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The reactions catalysed by cytochrome P450 enzymes can involve the oxidation of carbon, nitrogen,
phosphorus, sulphur, and other atoms.
 The oxidation reactions are easily taking place on nitrogen, phosphorus and sulfur atoms
 The oxidation reactions are easily taking place on carbon atoms if the carbon atom is either exposed (i.e.
easily accessible to the enzyme) or activated.
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For example, methyl substituents on the carbon skeleton of a drug are often easily accessible and are
oxidized to form alcohols, which may be oxidized further to carboxylic acids. In the case of longer
chain substituents, the terminal carbon and the penultimate
Carbon are the most exposed carbons in the chain, and are both susceptible to oxidation. If an
aliphatic ring is present, the most exposed region is the part most likely to be oxidized.
Nomenclature of CYPs
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CPY – Arabic Number – Capital Letter - Arabic Number
There are at least 33 different cytochrome P450 (CYP) enzymes can be classified into
Families(1-4), subfamilies (many) and members (many).
Most drugs in current use are metabolized by five primary CYP enzymes (CYP3A, CYP2D6, CYP2C9,
CYP1A2, and CYP2E1).
The isozyme CYP3A4 is particularly important in drug metabolism and is responsible for the
metabolism of most drugs.
Family
CYP3A4
Subfamily
Member no.
Classifications of CYPs
RELATIVE HEPATIC CONTENT
OF CYP ENZYMES
CYP2D6
2%
% DRUGS METABOLIZED
BY CYP ENZYMES
CYP2E1
7%
CYP 2C19
11%
CYP 2C9
14%
CYP2D6
23%
CYP 2C
17%
OTHER
36%
CYP 1A2
12%
CYP 3A4-5
26%
CYP 1A2
14%
CYP 3A4-5
33%
CYP2E1
5%
Important CYPs
 CYP3A4 has a large binding pocket and can accept large, bulky molecules that are
neutral. It catalyses N-dealkylations; aliphatic benzylic hydroxylations; aromatic
hydroxylations; epoxidations, oxidation of atoms near the end of a molecule.
 CYP2D6 prefers less bulky, basic molecules and catalyses O-demethylations; Ndealkylations; oxidation of atoms near the middle or end of a molecule.
 CYP2C9 has a strong preference for substrates with acidic functional groups. It
catalyses O-demethylations; aliphatic benzylic hydroxylations; aromatic
hydroxylations; oxidation of atoms near the middle or end of a molecule.
cytochrome P450 in the oxidation of xenobiotics.
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The enzymes that constitute the cytochrome P450 family are the most important metabolic enzymes
and are located in liver cells.
They are haemoproteins (containing haem and iron) and they catalyse a reaction that splits
molecular oxygen, such that one of the oxygen atoms is introduced into the drug and the other ends
up in water. As a result, they belong to a general class of enzymes called the monooxygenases .
cofactors
Simplified depiction of the proposed
activated oxygen–cytochrome P450-substrate
complex. Note the simplified apoprotein
portion and the heme (protoporphyrin IX)
portion or cytochrome P450 and the
proximity of the substrate R-H undergoing
oxidation.
Proposed catalytic reaction cycle involving cytochrome P450 in the
oxidation of xenobiotics.
H
OH
H
The other oxygen is still
bound to Fe, thus can be
use to oxidize the substrate
H
H
H
One of the activated
oxygen forms water
H
H
The Fe is now getting
oxidized and give e to
O2 to make it activated
H
The Fe is now reduced
thus can either bind
CO or O2
Drug requirements for oxidation reactions
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For any compound to undergo oxidative metabolism it needs
1.
2.
3.
The atom should bear hydrogen atom: to be replaced with OH
The atom need to be exposed to catalytic binding site of the enzyme
The atom need to be electron rich to attract the activated oxygen.
Presence of H atom
Electron rich
e
feed
-
Nitrogen
Oxygen
Sulfur
Pi system
C H
Exposed
A. Oxidation of Aromatic Moieties
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Aromatic compounds (arenes) undergoes aromatic
hydroxylation through an epoxide intermediate called
an “arene oxide” to their corresponding phenolic
metabolites (arenols).
R
R
R
O
Arene
O
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In most of the drugs,
hydroxylation occurs at
para position
O
C2H5
HN
O
Arene oxide
O
Phenobarbital
O
Arenol
C2H5
HN
N
H
OH
OH
N
H
O
A. Oxidation of Aromatic Moieties (Cont.)
 Effect of substituents on aromatic hydroxylation:
 Electron donating group activate (electron-rich) the aromatic ring towards hydroxylation
 Electron withdrawing group deactivate the aromatic ring towards hydroxylation
Compounds with two aromatic rings, hydroxylation
occurs preferentially in the more electron-rich ring.
Cl
Cl
H
N
N
H
Cl
Very less aromatic hydroxylation
N
H
Clonidine hydrochloride
(Anti-hypertensive drug)
CCOOH
No report on aromatic hydroxylation
SO2N(CH2CH2CH3)2
Probeniside
(Uricosuric agent)
A. Oxidation of Aromatic Moieties (Cont.)
• Electron donating group (EDG)
- Electronegative atoms: O > N > C sp2
( –O > NR2 > NH2 > OH > OR > OCOR > CH2=cH2 or
benezne)
• Electron withdrawing group (EWG) is any group which
have:
1. Pi system than can take electrons toward the pi resonance
(-SO3H>-COOH>-COOR).
2. Positive charge that can take electrons
(–N+R3>-N+H3)
3. Or both 1 and 2
(- N + O2-)
4. Halogens
Q) Halogens are
electronegative atoms, but
why they are not electron
donators (being poor e
withdrawals)?
A) Halogens attract electron
and not share it, thus induce
positive charge in the
connected atom.
A. Oxidation of Aromatic Moieties (Cont.)
Non-toxic
Non-toxic
toxic
Other possible
fates for arene
oxide, some of
them lead to
toxic effect
A. Oxidation of Aromatic Moieties (Cont.)
 Rearrangement of arene oxide through intramolecular hydride (deuteride) migration called the “NIH
Shift”.
 Because of an isotope effect on cleavage of the C-D bond, the proton is preferentially removed.
 Competing pathway to NIH shift is simple loss of a proton or deuterium from the cation intermediate
The more stabilizing the R group is the
more deprotonation that occurs
when R is electron withdrawing only
0-30% of the product retains
deuterium;
when R is electron donating, 40-54%
retention of D is found)
A. Oxidation of Aromatic Moieties (Cont.)
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Environmental polutants, such as polychlorinated biphenyls (PCBs) and 2,3,7,8-tetrachlorodibenzo-pdioxin (TCDD), have attracted considerable public concern over their toxicity and health hazards.
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These compounds appear to be resistant to aromatic oxidation because of the numerous
electronegative chlorine atoms in their aromatic rings (which causes inactivation).
Polycyclic aromatic hydrocarbons are ubiquitous environmental contaminants that are formed from
auto emission, refuse burning, industrial processes, cigarette smoke, and other combustion
processes.
E. g. Benzo[α]pyrene, a potent carcinogenic agent.
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A. Oxidation of Aromatic Moieties (Cont.)
 Aromatic hydroxylation of benzo[α]pyrene, can occur at a number of positions.
 The hydroxylated product can covalently binds to DNA. Therefore, certain arene oxides of
benzo[α]pyrene (e.g. 4,5-oxide, 7,8-oxide, 9,10-oxide) appear to display some mutagenic and
tumorigenic activity.
Although the formed arene
oxides and epoxides improves
hydrophilicity
and
renal
excretions, the molecules are so
reactive that can covalently
linked to nucleophilic groups in
DNA and proteins thus damages
them.
unless been eliminated
conjugation (Phase II)
by
1. Oxidation of Aromatic Moieties (Examples)
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Some metabolites are active e.g. the parahydroxylated metabolite of
phenylbutazone, oxyphenbutazone, is
pharmacologically active and has been
marketed itself as an anti-inflammatory
agent (Tandearil)
Examples of drugs and xenobiotics that undergo aromatic hydroxylation
in humans. Arrow indicates site of aromatic hydroxylation
B. Oxidation of Olefins
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The metabolic oxidation of olefinic carbon-carbon double bonds leads to the corresponding epoxide
(or oxirane) in a manner similar to aromatic oxidation.
Olefinic epoxides are more stable than arene oxides (Therefore, less toxic than arene oxide).
Olefinic epoxides are susceptible to enzymatic hydration by epoxide hydrase to form trans-1,2dihydrodiols, in a manner similar to arene oxides.
Frequently, the
Or may conjugate to macromolecules  toxic effect
epoxides formed from
the biotransformation
of an olefinic
compound are minor
products, because of
their further conversion
to the corresponding
1,2-diols.
Why the oxidation on
alefinic carbone of
aclofenac is easier than
on aromatic carbone?
B. Oxidation of Olefins (Cont.)
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The conjugation of styrene to macromolecules
after being oxidized is the cause of their
toxicity.
Conjugate to nucleic acids and
proteins which leads to toxic
effects
B. Oxidation of Olefins (Cont.)
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The safest pathway to get rid (detoxify) epoxide and arene oxide is by glutathione adduct formation.
Epoxide
C. Oxidation At Benzylic Carbon Atoms
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Carbon atom attached to aromatic rings (benzylic position) are susceptible to oxidation, forming the
corresponding alcohol (or carbinol) metabolites.
D. Oxidation At Allylic Carbon Atoms
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Microsomal hydroxylation at allylic carbon atoms is commonly observed in drug metabolism.
E. Oxidation At Carbon Atoms α to Carbonyls and Imines
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Microsomal hydroxylation at Carbon Atoms α to Carbonyls and Imines is also observed in drug
metabolism.
F. Oxidation At Aliphatic and Alicyclic Carbon Atoms
• Alkyl or aliphatic carbon centers are subject to mixed function oxidation.
1. ω oxidation: oxidation at terminal methyl group.
2. ω – 1 oxidation: oxidation at penultimate carbon atom (i.e. next to last carbon).
3. Activated carbon atom, that is next to sp , sp2 carbons
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The initial alcohol metabolites formed from these ω and ω – 1 oxidations are susceptible to further
oxidation to yield aldehyde, ketones, or carboxylic acids.
Alternatively, alcohol metabolites may undergo glucuronide conjugation.
trans-4-hdyroxyacetohexamide is the
major metabolite due to good exposure of
C4 and stereoselectivity of enzyme
G. Oxidation Involving CARBON-HETEROATOM Systems
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Metabolic oxidation of
 Carbon-Nitrogen
 Carbon-Oxygen
 Carbon-Sulfur
• Involves two basic types of biotransformation process occurs for Carbon-heteroatom oxidations:
1. Hydroxylation of the α-carbon atom attached directly to the heteroatom (N, O, S) and
decomposition with cleavage of carbon-hetero atom bond (dealkylation) . E.g. N-, O-, Sdealkylation, deamination reactions.
2.
Hydroxylation or oxidation of the heteroatoms (N, S, only).
G. Oxidation Involving CARBON-HETEROATOM Systems (Cont.)
• Oxidation of Tertiary Aliphatic and Alicyclic Amines:
1. N-dealkylation: oxidative removal of alkyl groups
2. Deamination
3. N-oxidation
G1) Oxidation Involving CARBON-NITROGEN
• Oxidation of Tertiary Aliphatic and Alicyclic Amines:
1. N-dealkylation: oxidative removal of alkyl groups
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Small alkyl groups (methyl, ethyl, isopropyl) removed rapidly and preferentially.
The first alkyl group of tertiary amine is removed more rapidly than second alkyl group.
Bisdealkylation of aliphatic tertiary amine to corresponding primary amine occurs very slowly.
N-dealkylation of t-butyl group is not possible by the carbinolamine pathway because α-carbon
hydroxylation can not occur.
G1) Oxidation Involving CARBON-NITROGEN (cont.)
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Examples of oxidative N-dealkylation,
which followed by oxidative deamination
which may be followed by oxidation
Several consecutive oxidative
dealklation is taking place on terminal
CH3 groups since the main carbon
contains NO hydrogen
G1) Oxidation Involving CARBON-NITROGEN (cont.)
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Alicyclic tertiary amine often generate lactam metabolites by α-carbon hydroxylation reaction.
G1) Oxidation Involving CARBON-NITROGEN (cont.)
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Some secondary alicyclic amines like tertiary amines are metabolised to their corresponding lactam
derivatives.
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Endogenous primary amines (e.g. dopamine, norepinephrine, tryptamine, and serotonin) and
xenobiotics are metabolised via oxidative deamination by a specialized family of enzymes called
monoamine oxidases (MAOs).
MAO is a flavin (FAD)-dependent enzyme found in two forms, MAO-A & MAO-B.
MAO-A & MAO-B have about 70% amino acid sequence homology.
MAO enzymes are located on the outer membrane of mitochondria.
MAOs are mostly found in LIVER and intestinal mucosa.
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G1) Oxidation Involving CARBON-NITROGEN (cont.)
• Oxidation of Secondary Primary Amines:
2. Oxidative deamination
• N-dealkylation of secondary amines proceeds via carbinolamine pathway (similar to tertiary amines)
and gives rise to primary amine metabolite.
• Primary amine metabolites are susceptible to oxidative deamination following the process similar to
N-dealkylation.
In general, dealkylation of secondary amines is believed to occur before oxidative
deamination.
In some cases, direct deamination of the secondary amine is also possible.
α α
•
N-dealkylation and
Oxidative
deamination
•
If α-carbon
hydroxylation can
not occur, then
oxidative
deamination is not
possible.
α
G1) Oxidation Involving CARBON-NITROGEN (cont.)
3.
N-oxidation:
–
–
–
–
Mostly for primary and secondary amines as well as aromatic amines
Primary amines will be converted to hydroxylamines, nitrone then nitrogen dioxide
Secondary amines will be converted to hydroxylamines then nitrone.
N-oxidation of secondary amines occurs much less than oxidative dealkylation and deamination
Toxic
metabolites
G1) Oxidation Involving CARBON-NITROGEN (cont.)
Although N-oxidation is less
common, it will be the major
metabolite for compounds
which have no hydrogen
atom on Cα
G1) Oxidation Involving CARBON-NITROGEN (cont.)
G2) Oxidation Involving CARBON-OXYGEN
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Several drugs containing ether group undergo oxidative O-dealkylation.
The biotransformation involves an initial α-carbon hydroxylation to form a either hemiacetal or a
hemiketal, which undergoes spontaneous carbon-oxygen bond cleavage to yield the dealkylated
oxygen species (phenol or alcohol) and a carbon moiety (aldehyde or ketone).
G2) Oxidation Alcohols and Aldehydes (least important)
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Many oxidative processess (e.g. benzylic, allylic, alicyclic or aliphatic hydroxylation) generate alcohol or
carbinol metabolites as intermediate products. Alcohols can also be oxidized further :Primary alcohols
– The carbon is well exposed here
– Oxidation of primary alcohol generates aldehydes.
– Oxidation of aldehydes generates carboxylic acid derivatives.
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Secondary alcohols
-
The carbon is NOT well exposed here
Oxidation of secondary alcohol to ketones is NOT often important as it reduces back to secondary alcohol.
Secondary alcohol group being more polar and functionalized, is more likely to be conjugated than the ketone
moiety.
H1) Oxidation Involving CARBON-SULFUR
•
Several drugs containing CARBON-SULFUR functional group are susceptible to
1.
2.
3.
S-dealkylation
Desulfuration
S-oxidation
• The first two process involve oxidative carbon-sulfur bond cleavage.
1. S-dealkylation: Similar to N- and O-dealkylation.
H2-H3) Oxidation Involving CARBON-SULFUR (cont.)
2.
Desulfuration :
Oxidative conversion of
carbon-sulfur double
bonds (C=S) (thiono) to
the corresponding
carbon-oxygen double
bond (C=O) is called
desulfuration.
3. S-oxidation :
• S-oxidation yields the
corresponding sulfoxide
derivatives.
• Sulfoxide
drugs/metabolites may
further oxidised to
sulfones (-SO2-)
Case study for oxidation reactions
•
•
2-acetylaminoflurorene is wellknown hepatocarcinogenic, it
undergoes an N-hydroxylation
reaction catalyzed by CYP to form
the corresponding N-hydroxy
metabolite (also called a
hydroxamic acid)
Hydroxamic acid undergoes
conjugation to form O-sulfate
ester, which ionizes to generate
the electrophilic nitrenium
species.
Case study for oxidation reactions
•
Chlorpormazine undergoes S-oxidation, oxidative N-dealkylation and oxidative deamination
Case study for oxidation reactions
• Applications for oxidation favorable sites:
1. The least substituted aromatic ring will be favorably oxidized, especially at the least hindered
carbon atom
Deactivated ring
(Rare metabolite)
e rich and exposed
(Major metabolite)
Some steric hindrance
(minor metabolite)
Reduction
Reductive Reactions
•
Reductive process play an important role in the metabolism of many compounds containing
carbonyl, nitro and azo groups
- Carbonyl reductions
Generate
alcohols
Conjugated
- Nitro and Azo reductions
Generate
amines
Conjugated
Phase-I
O-conjugates
N-conjugates
Phase-II
1. Reduction of Aldehydes and Ketones
 Aldehydes reduces to primary alcohols.
 Ketones reduces to secondary alcohols.
 Reactions mediated by
Aldo-Keto reductase enzymes
 Bioreduction of ketones often leads to the creation of an asymetric centre and thereby, two
possible stereoisomeric alcohols.
 One of the stereoisomer may preferentially form predominantly over other stereoisomer and
thus shows product stereo selectivity in drug metabolism.
2. Reduction of Nitro and Azo Compounds
 Bioreduction of aromatic nitro and azo compounds leads to aromatic primary amine
metabolites.
 Aromatic nitro compounds are reduced initially to the
nitroso and hydroxylamine
intermediates that subsequently further reduced to amine
+ O
Ar N
Ar N O
Ar NHOH
O
Nitro
Hydroxylamine
Nitroso
Ar NH2
Amine
hydrazo intermediate (-NH-NH-) that subsequently cleaved
reductively to yield the corresponding amines.
 Azo reduction proceed via
Ar N N Ar'
Azo
Ar NHNH Ar'
Hydrazo
H2N Ar + H2N Ar'
Amine
Case study for reduction reactions
•
Bacterial reductases play a
role
in
enterohepatic
recirculation of nitro or
azo containing drugs.
•
Sufasalazine is an azo
containing prodrug which
is
activated
to
sulfanilamine by intestinal
bacteria
Hydrolysis
Hydrolytic Reactions
•
•
Hydrolysis of Esters and Amides is catalyzed by widely distributed hydrolytic enzymes.
Esters  alcohols, phenols and carboxylic acids
- Non-specific esterases (liver, plasma, kidney, and intestine)
- Plasma pseudocholinesterases also participate
•
•
Amides amines and carboxylic acids
- Liver microsomal amidases, esterases and deacylases
- Hydrolysis of esters is major metabolic pathway for ester drugs
Hydrolytic Reactions (Cont.)
•
Aspirin is hydrolyzed
to salicylic acid by
plasma estrases to
releas salicylic acid
which have antiinflammatory effect.
 Hydrolysis of
procaine (local
anesthetic drug) by
esterases is faster
(t0.5=1 min) than
 Hydrolysis of
procainamide
(antiarrhythmic
agent) by amidases
(t0.5=4 hours)
COOH
O
O
COOH
OH
OH
CH3
Asprin
(Acetylsalicyclic acid)
O
CH3
Salicyclic Acid
Acetic Acid
Hydrolytic Reactions esters bond is weaker than amide bond
- 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 weaker electron withdrawing atom
- Therefore, the carbonyl carbon attached to oxygen atom will be more electropositive, and more
reactive toward nucleophilic attack of water molecule during hydrolysis.
Case study for hydrolytic reactions
•
Dipivefrine is a prodrug of adrenaline, which is used to treat glaucoma.
•
Dipivefrine: is a di-tertbutylcarboxy ester of adrenaline, thus it is more lipophilic and can better penetration
through the corneal membrane then hydrolyzed to give the active form (adrenaline)
General notes for phase-I reactions
•
Hydrolysis normally catalyzed by
carboxylesterases:
– Cholinesterase…. Hydrolyzes cholinelike esters (such as succinylcholine),
procaine and acetylsalicylic acid.
– Arylcarboxyesterase.
– Liver carboxyesterase
Zwitterion can be
easily excreted
General notes for phase-I reactions (Cont.)
•
Esters that are sterically hindered are hydrolyzed more slowly and may be appeared unchanged in
urine
Steric hindrance
for estrases
•
Amides are more stable to hydrolysis than esters….large fraction of amide containing drugs are
normally excreted unchanged.
Bioactivation of omeprazole (case study)
•
•
•
Proton-pump are present in parietal cells to exchange of K+ with H+
Proton-pump inhibitors are bioactivated next to the parietal cells (i.e. At highly acidic environment)
Omeprazole (a proton-pump inhibitor) is activated by
– Protonation at benzimidazole ring followed by attachment of pyridine nitrogen to form another ring
– The new ring is opened and loses a water molecule to generate sulfonamide
– The sulfonamide is highly susceptible to nucleophilic attach by SH of cysteine residue of the proton pump
which leads to pump damage.