Enantioselective Biotransformation of Prochiral Ketone via ...
Biotransformation _Casarett & Doull's
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Transcript of Biotransformation _Casarett & Doull's
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Biotransforming Enzymes(SYN. Drug metabolizing enzymes, drug detoxication enzymes)
- Two categories of enzyme systems are known toexist in mammals
A. Enzymes with normal endogenoussubstrates
Enzymes that normally occur in the tissues and areresponsible for transformation of normalendogenous chemicals in the tissues.
B. Enzymes having no specific endogenoussubstrates
Enzymes that alter the structure of many foreignchemicals but have no established normal
endogenous substrates.
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Biotransforming Enzymes-Examples
Enzyme Substrate
Cholinesterase
(non-specific)
Acetylcholine
Procaine
Succinylcholine
Monoamine oxidase(MAO)
Epinephrine
Tyramine
Benzylamine
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Biotransforming Enzymes-Examples
Acetylcholine Succinylcholine
Procaine
Substrates for Cholinesterase
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Biotransforming Enzymes-Examples
Epinephrine Tyramine
Benzylamine
Substrates for MAO
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Biotransforming Enzymes
- Two categories of enzyme systems are known toexist in mammals
A. Enzymes with normal endogenoussubstrates - Specific
Enzymes that normally occur in the tissues and areresponsible for transformation of normalendogenous chemicals in the tissues.
B. Enzymes having no specific endogenous
substrates Non-specific
Enzymes that alter the structure of many foreignchemicals but have no established normalendogenous substrates.
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Biotransforming Enzymes-Examples
Enzyme Substrate
Cholinesterase
(non-specific)
Acetylcholine
Procaine
Succinylcholine
Monoamine oxidase(MAO)
Epinephrine &NE
Dopamine
Serotonin
Tyramine
Benzylamine
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Metabolism of Procaine
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Monoamine Oxidase Metabolism of Serotonin
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Biotransforming Enzymes
Basic properties
- Limited in numbers having broad substratespecificities
- Synthesis of some enzymes is triggered by the
presence of the xenobiotic Enzyme induction- In most cases the enzymes are expressed in the
absence of a discernible external stimulus, i.e. theyare expressed constitutively
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Biotransforming Enzymes
Basic properties
- The same enzyme may exert different functions indifferent tissues. e.g. Cyt.P450 enzymes catalyzes thesynthesis of steroid hormones in steroidogenictissues but converts these hormones into theirwater-soluble metabolites to be excreted in the liver
- The structure of a biotransforming enzyme may differamong individuals giving rise to intraspecies variationin the rates of xenobiotic biotransformation.
Pharmacogenetics (The study of the causes,
prevalence, and impact of heritable differences inxenobiotic biotransforming enzymes)
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Phase I and Phase IIBiotransformation
- The reactions catalyzed by xenobiotic biotransformingenzymes generally are divided into two groups:
1. Phase I reactions Non-synthetic reactions2. Phase II reactions Synthetic reactions
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PHASE I METABOLIC PATHWAYS
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Microsomal Oxidation
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Liver Microsomal Enzymes - MFOs
The endoplasmic reticulum (ER) of the liver cells contains a
filamentous-like structures of two types:
- Rough endoplasmic reticulum (RER) and
- Smooth endoplasmic reticulum (SER)
SER contains large proportion of the Drug metabolizing enzymesknown as Mixed Function Oxidases (MFOs).
When the liver cells are ruptured by homogenization the ER
undergoes fragmentation.
The fragments of the SER separated from other parts of the liver
cell by ultra-centrifugation (e.g. at 10,000 g for 10 min) are
commonly known as microsomes.
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Metabolism by Microsomal & Non-microsomal Enzymes
Microsomal Enzymes Non-microsomal enzymes
- Metabolize foreign compounds - Metabolize foreign compounds
- Do not metabolize endogenous
compounds (e.g. phenyl alanine,
tryptophan, etc.)
- Metabolizes endogenous
compounds
- Examples:
Cyt. P450s
Epoxide hydrolase
Flavin-monooxygenases
- Examples:
Alcohol dehydrogenase
aldehyde dehydrogenase
Xanthine oxidase (XO)
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Phase I vs. Phase IIBiotransformation Reactions
Phase I reactions Phase II reactions
A. Oxidation
- Aromatic ring oxidation
- Alicyclic ring oxidation
- Deamination
- Dealkylation
- N-oxidation
- S-oxidation
- Oxidation of Alcohols
- Epoxide hydroxylation
- Hydroxylation of alkyl side chain
A. Conjugation with
- Glucuronic acid
- Sulfate
- Mercaptic acid
- Amino acid
(e.g. Glycine, taurine,
glutamine)
- Glutathione
B. Reduction- Azo reduction
- Nitro reduction
B. Methylation
C. Hydrolysis
- Ester hydrolysis
- Amide hydrolysis
C. Acetylation
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Oxidation Of Drugs By Cytochrome P450
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Oxidation Of Drugs By Cytochrome P450
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Aliphatic Oxidation
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Aromatic Hydroxylation (1)
acetanilid p-hydroxyacetanilid
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Aromatic Hydroxylation (2)
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N-Dealkylation
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O-Dealkylation
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S-Demethylation
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Oxidative Deamination
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S-Oxidation
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N-Oxidation
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N-Hydroxylation
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N-Hydroxylation of AAF
N-Hydroxylation of AAF is the first metabolic step towards
the development of a carcinogenic agent
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Oxidative Dehalogenation
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Desulfuration
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Desulfuration
Ph I Bi t f i E
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Phase I Biotransforming Enzymes
Hydrolysis
Carboxylesterases
Pseudocholinesterases
Paraoxonase
Peptidases
Epoxide Hydrolase
Reduction
Azo Reductase
Nitro Reductase
Carbonyl Reductase
Glutathione Reductase
Enzymes for -
Disulfide reduction
Sulfoxide & N-oxide reduction
Quinone reduction &
Dehalogenation
Phase I Biotransforming Enzymes
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Phase I Biotransforming Enzymes
Oxidation
Alcohol Dehydrogenase
Aldehyde Dehydrogenase
Dihydrodiol Dehydrogenase
Molybdenum Hydroxylases
Xanthine Dehydrogenase & Xanthine Oxidase
Aldehyde Oxidase
Monoamine Oxidase
Cyclooxygenase
Hydrolysis
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Hydrolysis
Carboxylesterases
- glycoproteins present in serum & most tissues.- hydrolyze numerous endogenous lipid compoundsthat may be
a carboxylic acid ester
an amide or
a thioester.
- generate pharmacologically active metabolites fromseveral ester or amide prodrugs.
- additionally may convert xenobiotics to toxic &tumerogenic metabolites.
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Ester Hydrolysis
RCOOR' RCOOH + R'OH
Microsomes and cytosol
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Ester Hydrolysis
Microsomes and cytosol
Enalaprit
Hydrolysis
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Hydrolysis
True - & Pseudocholinesterases
True cholinesterase (Acetylcholinesterase)- present in erythrocyte membranes
Pseudocholinesterases (Butyrylcholinesterase)
- present in serum
Paraoxonase
- present in serum
- hydrolyze phosphoric acid esters
- also known as aryldialkylphosphatase
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Hydrolysis
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y y
Epoxide Hydrolases
- present in virtually all tissues.
- in mammals, there are 5 distinct forms
Microsomal epoxide hydrolase (mEH)
Soluble epoxide hydrolase (sEH)
Cholesterol EH
LTA4 hydrolase
Hepoxilin hydrolase.
- catalyze trans-addition of water to
alkene epoxides
arene oxides.
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Epoxide Hydrolase
Reduction
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- Enzymatic or non-enzymatic
- in vivo reduction of certain metals & xenobioticscontaining:
An aldehyde (-CHO)
Ketone ( )
Quinone
Disulfide (S2-)
O
C
Sulfoxide
N-oxide
Alkene
Azo (-N=N-)
Nitro (-NO2)
- by the interaction with reducing agents(endogenous) viz. glutathione (GSH). FAD, FMN,and NADP.
Reduction
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Azo and Nitro Reduction
- catalyzed by
i. intestinal microflora
ii. Two liver enzymes:
- cytochrome P450 and- NADPH-quinone oxidoreductase
(DT-diaphorase)
- NADPH is required
- inhibited by oxygen
- the anaerobic environment of the lower GIT is well-suited
for such reductions.
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Azo Reduction
RN=NR' RNH2 + R'NH2
Microsomes and cytosol
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Azo Reduction
Microsomes and cytosol
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Nitro Reduction
RNO2 RNH2
Microsomes and cytosol
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Nitro Reduction
Reduction
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Carbonyl Reduction
- substrates: aldehydes and ketones
- metabolites: primary and secondary alcohols
- catalyzed by
i. alcohol dehydrogenaseii. Carbonyl reductases
- monomeric, NADPH-dependent enzymes
- present in blood, cytosolic fraction of varioustissues including liver, and microsomal fraction ofliver
Reduction
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Disulfide Reduction
- substrates: any disulfide compound
- metabolites: thiols
- a two step process
step 1: catalyzed by glutathione reductasestep 2: catalyzed by glutathione-S-transferase
or can occur non-enzymatically.
Reduction
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Sulfoxide & N-oxide reduction
Sulfoxide reduction
(thioredoxin-dependent enzymes)
- in liver and kidney
- substrates: sulfoxides
N-oxide reduction
(NADPH-dependent enzymes,
e.g. CytP450 or NADPH-CytP450 reductase)
- in liver microsomes
- substrates: N-oxides
Reduction
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Quinone reduction
(NADPH-quinone oxidoreductase, a flavoprotein
SYN. DT-diaphorase)
- in cytosol
- substrates: Quinones
- Products: Hydroquinones
Reduction
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Dehalogenation
Three major mechanisms:
1. Reductive dehalogenation
Halogen replaced with a hydrogen
2. Oxidative dehalogenation
Halogen & hydrogen replaced with oxygen
3. Double dehalogenation
2 halogens replaced from two adjacent Cs to form a
C-C double bond
Oxidation
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Alcohol Dehydrogenase (ADH)
- in cytosol (liver, kidney, lungs and in gastric
mucosa)
- Four major classes: Class I-IV
Class Location Substrate
I. ,, & -ADH liver, kidney, lungsand in gastricmucosa
EtOH, other smallaliphatic alcohols
II. -ADH Primarily liver Larger aliphatic &aromatic alcohols
III. -ADH
(Chi-ADH)
liver, kidney, lungsand in gastricmucosa
Long-chain alcohols(pentanol & larger)& aromatic alcohols
IV. - or -ADH Not expressed in
liver*
Retinol
Al h l D h d
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Alcohol Dehydrogenase
CH3CH2OH + NAD+ CH3CHO + NADH + H
+
ethanol acetaldehyde
Oxidation
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Aldehyde Dehydrogenases (ALDHs)
- in mitochondria (liver, kidney, lungs and in gastricmucosa)
- oxidize aldehydes to carboxylic acid
- use NAD
+
as cofactor- also have esterase activity
- may differ in primary amino acid sequences
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Aldehyde Dehydrogenase
CH3CHO + NAD+ CH3COOH + NADH + H
+
acetaldehyde acetate
Oxidation
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Dihydrodiol Dehydrogenases
- forms the aldo-keto reductase super family
- located in cytosol
- chemically NADPH-requiring oxidoreductase
- oxidize various polycyclic aromatic hydrocarbons
Oxidation
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Molybdenum hydroxylases
- also known as molybdozymes
- two major types:
aldehyde oxidase (AO) and
xanthine dehydrogenase/xanthine oxidase (XD/XO)- all are flavoprotein enzymes
- During oxidation, AO and XO are reduced and
then reoxidised by molecular O2
- The oxygen incorporated comes from H2O
(differentiating points between oxidases and
oxigenases).
Oxidation
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Molybdenum hydroxylases
Xanthine dehydrogenase/xanthine oxidase (XD/XO)
XD and XO are two different forms
- these two differs w.r.t. the electron acceptor in the 1st step
of the catalysis: NAD+ for XD and molecular O2 for XO
- Chemical difference:
SH
SH
S
S
oxidation of cysteine residue
XD XO
Oxidation
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Molybdenum hydroxylases
Xanthine dehydrogenase/xanthine oxidase (XD/XO)
Function:
- in vivo conversion of XDXO is important in:
a. ischemia-reperfusion injury
b. alcohol-induced hepatotoxicity and
c. lipopolysaccharide-mediated tissue injury.
- XO contributes to oxidative stress and lipid peroxidation
(by reduction of O2 that leads to the formation of ROS).
X thi O id
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Xanthine Oxidase
Oxidation
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Molybdenum hydroxylases
Aldehyde oxidase
- exists only in oxidase form in cytosol
- transfers electrons to molecular O2 and this can generate
ROS and lead to lipid peroxidation
- plays important role in the catabolism of biogenic amines
and catecholamines.
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Oxidation
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Peroxidase-dependent cooxidation
- a process by which oxidative biotransformation of
xenobiotics by peroxidases couples the reduction of H2O2 &
lipid hydroperoxides to the oxidation of other substrates
cooxidation
- an important peroxidase is prostaglandin H synthetase(PHS) that has two catalytic activities:
a cyclooxigenase (COX) converts arachidonic acid to
prostaglandins (intermediate PGG2) &
a peroxidase converts hydroperoxides (PGG2) to thecorresponding alcohol PGH2.
A hid i id
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Arachidonic acid
PGG2
PGH2
Prostaglandins(PGD2, PGE2, PGF2)
Thromboxane Prostacyclin(PGI2)
X or 2XH
XO or 2X. + H2O
COX
Peroxidase
Oxidation
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Peroxidase-depent cooxidation
- Peroxidases are important in the activation of xenobiotics
to toxic or tumerogenic metabolites, particularly in
extrahepatic tissues that contain low level of cyt. P450.
- In certain cases the oxidation of xenobiotics by
peroxidases involves direct transfer of the peroxide oxygento the xenobiotic for the conversion: X XO
- Alternatively, xenobiotics that can serve as electron
donors, viz. amines, phenols, etc. can also be oxidized to
free radicals during the reduction of a hydroperoxide byperoxidases.
Oxidation
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Peroxidase-depent cooxidation
- In this case the hydroperoxide is still converted to the
corresponding alcohol but the peroxide oxygen is reduced
to H2O instead of being incorporated into the xenobiotic.
- For each molecule of hydroperoxide reduced (a 2 electron
process), 2 molecules of xenobiotic can be oxidized (eachby a 1 electron process).
- Many of the metabolites produced are reactive
electrophiles that can cause tissue damage.
Oxidation
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Peroxidase-depent cooxidation
Role of COX:
- play at least two distinct roles in tumor formation:
1. It may convert certain xenobiotics to DNA-reactive
metabolites and initiate tumor formation.
2. It may promote subsequent tumor growth, perhaps by the
formation of growth-promoting eicosanoids.