Download glutathion-s-Transferase

DRUG METABOLISM
Department of Pharmaceutics
 Drug metabolism or Biotransformation is the major
mechanism for elimination of drug from the body.

It is the process by which the drug is chemically modified
inthe body and the end product of modification is called
METABOLITE

Liver is the major site for metabolism,along with other
organs like kidneys,lungs,blood,skin,saliva,etc;

Drug metabolism involves conversion of one form of drug
to another form………it may be enzymatic or non-enzymatic.
 Generally in liver major metabolism takes place at
microsomes which are present in the endoplasmic reticulum.The
microsomal enzymes are responsible for Metabolising the
compounds.
Hence drug metabolism can be classified as
1. Microsomal
2. Non-microsomal
DRUG LATENTIATION:
when a delayed or prolonged release/response of drug is
required,normally inactive forms or precursors of the drugs can be
commonly used in pharmaceutical practice.so that the active forms will
generates in the body. Such approach is said to be DRUG
LATENTIATION
Ex: chloraamphenicol palmitate,Dichloralphenezone,
Estolates and
steroids.
WHY DRUG METABOLISM/IMPORTANCE OF METABOLISM

As all the chemical substance are not nutrients to the body,whatever the
foreign compounds entering in to the body are called
exogenous compounds or xenoboitic

Generally drugs get excreted based on their lipophilicity, all the drugs
available are not lipophilic,some may be hydrophilic,the hydrophilic
moleculesmay get excreted through urine,but lipophilic molecules
will undergorebsorption from renal tubules in to blood after glomerular
filtration.

So that the drug get accumilated in the body which leads to toxicity.

The metabolic system will makes the lipophilic moeities to make water soluble.

So the drug willl be easily excreted,hence the frug biotransformation isprocess
is also called as DETOXIFICATION PROCESS
PHASE 1
Oxidation(microsomal)
Aromatic hydroxylation
Aliphatic hydroxylation
Epoxidation
N-Dealkylation
O-Dealkylation
S-Dealkylation
Oxidative deamination
N-Oxidation
S-Oxidation
Phosphotionate oxidation
Dehalogenation
Oxidation(non-microsmal)
Alcohol dehydrogenase
Aldehyde oxidation
Xanthine oxidase
Amine oxidase
Aromatases
PHASE 2
Conjugation with sugars
(1) Glucuronidation
O-Glucuronides
N-Glucuronides
(2)Conjugation with other
sugars.
Sulfation
Methylation
Acetylation
Aminoacid conjugation
Glutathione conjugation
Fattyacid conjugation
Condensation reactions
PHASE 1
Reduction
Azo reduction
Nitro reduction
Reduction dehalogenation
Hydrolysis
Ester hydrolysis
Amide hydrolysis
Hydrazine&carbamate
hydrolysis
Hydration
Other minor reactions
PHASE2
OBJECTIVE OF PHASE 1 REACTIONS
1. Increase in hydrophilicity
2. Reduction in stability.
3. Facilitation of conjugation………leads to phase 2
reaction makes the drug hydrophilic.
The compounds which are failed to undergo phase -2
reaction will preceeds to phase-3
to avoid
1. Toxicity to cell components
2. Hydrolysis of conjugates back to reactive species.
3. Inhibition of phase-2 enzymes.
OXIDATION INVOLVING MICROSOMAL MIXED FUNCTION
OXIDASE(MFO’S)
Oxidation of compound r drug is catalysed by many
enzymes located in microsomes.such enzymes requires both molecular oxygen
and reducing agents NADPH to effect reaction.therefore they are refered as
mixed function oxidases or mono oxidases (MFO)
They are located in the endoplasmic reticulum of hepatic cells
performs many functions.The MFO are structuraal enzymes requires
NADPH,O2,CYT P 450,NADPH-CYTP450 reductase enzyme & phospholipids.
CYT P450 is an enzyme responsible for oxidation and
reduction reactions.Chemially it is a heme protien containing iron.
reduction cycle.
The following figure shows the CYT P 450 oxidation-
NON MICROSOMAL OXIDATION
In these type enzymes like alcohol dehydrogenase,aldehyde
dehydrogenase, xanthine oxidase,amine oxidases,aromatase etc;
are included……
REDUCTION
Hepatic microsomes catalyses various reduction reactions and requires
NADPH for this purpose.Azo and nitro reductions is catalysed by CYT P 450
reductase enzyme
HYDROLYSIS
Liver microsomes and ther tissues contain non specific esterases
and Hydrolytic enzymes.
so that they undergo hyrolytic reactions.
HYDRATION
Hydration can be regarded as specialised form of
hydrolysis,where water is added to the compound with out causing the
compound to dissociate in to no.of components.
EX: Epoxides are converted in to di hydrodiols,by the
enzyme epoxide hydratase
PHASE 2 REACTIONS
REACTION
ENZYME
FUNCTIONAL
GROUP
Glucoronidation
UDP-Glucoronyl
transferase
OH,-COOH,-NH2,SH
Glycosidation
UDP-Glucoronyl
transferase
-OH,-COOH,-SH
Sulfation
sulfo Transferase
-OH,-NH2,-SO2NH2
Methylation
Methyl transferase
-OH,NH2.
Acetylation
Methyl transferase
-OH,-NH2,-SO2NH2,COOH.
Aminoacid
conjugation
…………..
-COOH.
Fattyacid
……………
-OH.
Glutathione
conjugation
Gluthion-sTransferase
Epoxide organic
halides
condensation
……………….
various
GLUCORONIDATION

Glucoronidation means conjugation reaction of sugars
with D-Glucoronic acid,
xylose,arabinose and glucose.

Glucoronidation involves the reaction between the high
energy form of conjugating agent with uridine di phosphate
glucornic acid and drugs containing hydroxyl,carboxyl,amine
or thiol groups.

The reaction is mediated by glucoronyl transferase.
GLUTATHIONE CONJUGATION
Glutathione (GSH) is a tripeptide of glutamyl-cysteine-glycine That is
recognized as a protective device with the cell for the Removal of
potentially toxic electrophilic compounds GSH reacts non-enzymatically
and enzymatically via glutathion-s-Transferase, through the nucleophilic
sulfhydral group with Reactive electrophilic oxygen intermediates of
certain drugs Formed during oxidative biotransformation reactions. These
Reactive electrophilic intermediates my react with nucleophilic
macromolecules such as proteins in the cell, leading to a cell Injury and
cellular necrosis
The enzyme catalyzing these reactions are the
glutathione-s-transeferase located in the cytosol of the liver,kidney and gut
of other tissues……..
1. Conjugation of glutathion with the drug molecule through the sulfahydral
Group of cysteine in the presence of glutathione-s-transeferase.
Gly
Glutathione-s-transferase
Drug+GSH
D-S-Cys
Glutamyl
Trans peptidase
Gly
D-S-Cys-NH2
Gln
peptidase
COOH
N-Acetylase
D-S-CH2-CH-NH-CO-CH3
COOH
D-S-CH2-CH-NH2
2. Removal of glutamyl by glutamyl traspeptidase
3.Removal of glycine by peptidase.
4.N- acylation f the cysteine conjugate to form n acetylated cysteine.
SULFATION
•
Conjugation with sulfate is common pathway for metabolism
of phenols,but occur for alcohols,amines and thiols.
•
Sulfo transeferase is the enzyme involved in metabolism of
sulphur
METHYLATION
o
Methylation reactions mainly meant for endogenous substance.
o
High energy intermediate compound S-Adenosyl methionine
(SAM) is
required to form methyl conjugate.
o
Methylation leads to less polar products,which are not easily
excreted from the body.
ACETYLATION

This reaction is common for aromatic amines and sulphonamide which
requires a co-factor acetyl co enzyme A.

It is obtained from the glycolysis pathway .This reaction occurs in kupffer
cells,but not hepatocytes.

It also occurs in reticuloendothelial cells of spleen,lung &gut.

The acetylated product is usually less polar than parent drug.renal toxicity
of sulphonamides can be attributed to precipitation of less polar acetylated
sulphonamides.
AMINOACID CONJGATION
Amino acid conjugation is a special fprm of N- acylation.the usual amino acid
in volved Glycine,glutamine,ornithine,arginine.and taurine Generally ornithine
is the amino Conjugated reaction which predominantly occurs in ureotelic
animals. The conversion of Benzoic acid to hippuric acid and salicylic acid are the
examples of this metabolic pathway.
FATTY ACID CONJUGATION
The fatty acids involved in the conjugation reaction are stearic
acid and palmitic acid.
Microsomal enzymes of liver catalyse these reactions
CONDENSATION REACTION
These are not be enzymatic but purely chemical and have
been found for amines and aldehydes.
FACTORS EFFECTING DRUG METABOLISM
The thereapeutic efficacy,toxicity and biological half life of drug depends
upon the metabolic rate of the particular drug….the following are the
Factors effecting drug metabolism…….
1.Physico chemical properties of drug
2.Chemical factors
Induction of drug metabolising enzymes
Inhibition of drug metabolising enzymes
Environmental chemicals
3.Biological factors
Strain difference
Species diference
Sex difference
Age
Diet
Alterd physiological factors
Pregnancy
Hormonal imbalance
Disease status
Temporal factors
Circadian rhythms
Circannual rhythms
PHYSICO CHEMICAL PROPERTIES OF DRUG
Molecuar size shape,pka,acidity, basicity,lipophilicity,
steric and electronic characteristicsof a drug influence its
interaction with the active sites of the enzyme and
biotrasnformation of the process in which it is subjected
CHEMICAL FACTORS
INDUCTION OF DRUG METBOLISING ENZYMES
The phenomenon of increasing drug metabolic ability of the
enzymes by several drugs and chemicals is called as enzyme
induction,and the agents which bring about such effect are known as
enzyme inducers
PROPERTIES OF ENZYME INDUCERS:
1. They are lipophilic compounds.
2. They are substrate for the inducted enzyme system.
3. They have long elimination half life.
MECHANISM INVOLVED IN ENZYME INDUCTION ARE:
1. Increase in both liver size and liver blood flow.
2. Increase in total &microsomal protein content
3. Increase in stability of enzymes.
4. Increase in synthesis of CYT P 450.
5. Decreased degration of CTY P 450.
6. Proliferation of smooth endo plasmic reticulum.
INDUCERS ARE OF 2 TYPES:
phenobarbital type inducers
poly-cyclic hydro carbon type inducers
phenobarbital type inducers:several drugs and pesticides which incerase
the rate of metaboism of a large no.of drugs.these type can increase the
enzyme activity up to 4 times.
EX: when phenobarbital and dicoumaral are co administered to avoid the
failure of anti-coagulant therapy and the haemorragic crisis.
poly-cyclic hydro carbon type inducers:
such as 3-methyl
cholanthrene &cigaret smoke which stimulates the metabolic rate of few
drugs.
Some drugs like carbamazipine,meprobamate,cyclophosphamide,rifampicin
etc; stimulate their own metabolism……
the phenomena is called auto
induction
INHIBITION IF DRUG METABOLISING ENZYMES
A decrease in the drug metabolising abilitty of an enzyme is called
enzyme inhibition.
1. Direct inhibition
a. Competitive inhibition
b. Non competitive inhibition
c. Product inhibition
2.Indirect inhibition
a.Repression
b. Altered physiology
1a.Competitive inhibition: When structurlly similar compounds compete
for the same site on an enzyme.tis process is reversible,and can be overcome by
high concentration of substrates.
EX:metacholine inhibits metabolism of acetylcholine by competing with it for
cholinesterase….
2a.Non-competitive inhibition:when structurally unrelated agent interacts
with the enzyme and prevents the metabolism of the drug.
EX:lead,mercury,arsenic and orgophosphorous insecticides…..
1c. Product inhibition:when the metabolic product competes with the
substrate for the same enzyme.the phenomenon is called aauto inhibition
EX:certain specific inhibitors like xanthine oxidase inhibitors (allopurinol),
MOA oxidase inhibitors(phenelzine) inhibits the enzyme activity directly.
2a. Repression:it is the decrease in enzyme content due to fall in the rate of
enzyme synthesis.
EX : Carbon tetra chloride,carbon disulphide,disulfiram.
2b.Altered physiology: due to nutritional deficiency or harmonal imbalance
Enyme inhibitors and drugs effected by them
inhibitors
Drugs with decreased
metabolism
MOA inhibitors
Barbiturates,tyramine
Coumarins
Phenytoin
Allopurinol
6-mercaptopurine
PAS
Phenytoin,hexobarbital
ENVIRONMENTAL CHEMICALS
Several environmental influence the drug metabolising ability
 Haalogenated pesticides such as DDT and polycyclic aromatic
Hydrocarbons contained in ciggarate smoke hav enzyme
induction effect.
 organophosphate insecticides and heavy metals such as tin,
Mercury, nickel,cobalt and arsenic inhibits drug metabolic ability
of an enzyme
BIOLOGICAL FACTORS
SPECIES DIFFERENCE:
species difference have been observed in both
Phase -1 and phase-2 reactions.
In phase-1 variations in enzyme and their activity have been observed
Ex: metabolism of amphetamine and ephedrine,in men & rabbit they
are predominantly metabolised by oxidative deamination where as in
Rats the aromatic oxidation is major route.
In phase-2 reactions,the variation is mainly due to complete lack of
Certain conjugating enzymes.
Ex: in pigs, phenol is excreted mainly as glucorunide where as its
sulphares conjugates dominates in rats.
Certain birds utilises ornithine fpr conjugating aromatic acids instead
Of glycine.
AGE
In neonates(up to 2 months),the microsomal enzyme system is
not fully developed and many drugs are bio-transformed slowly
Ex: caffeine has half life of 4 days in sdults,but in neonates it rises
Up to 4 days agent.
 Infants(between 2 months & one year) show almost a similar
profile as neonates in metabolising drugs with improvement in the
capacity as age advances and enzyme activity increases.
 chidren b\w 1 year &12 years and older infants metabolises several
drugs much more rapid than adults as the rate of metabolism
reaches to maximum level.as a result they require more dose than
adults.
 In very elderly persons,liver size is reduced,due to decrease in
microsomal enzyme activity,hepatic blood flow,cardiac output, of
all of which contributes decreased metabolism of drug.

DIET
 Low protein diet decreases and high protein diet increases the metabolising ability
 protein carbohydrate ratio in the diet is also important:a higgh ratio increases the
microsomal mixed function oxidase activity.
fat free diet depresses the CYT P 450 since phospholipids are important contents
of microsomes.
 Dietary deficiency of vitamins & minerals may retard the metabolic activity of
enzymes.
 Grape fruit inhibits metabolism of many drugs and improves their oral
availability.
 starvation results in the decreased amount of glucorunides formed than under
normal conditions .
 Malnutrition results in enhanced metabolism of sex harmones.
 Alcohol ingetion results in short term decrease followed by increase in enzyme
activity
ALTERED PYSIOLOGICAL FACTORS
PREGNANCY: Studies in animals hav e shown the maternal drug metbolising
ability is reduced during later stages of pregnancy.this was due to high
levels of steroid harmones in circulation during pregnancy.
HARMONAL IMBALANCE: Higher levels of one harmone may inhibit the activity
of few enzymes while inducing that of others .adrenolectomy,thyroidectomy,
alloxan-induced diabetes in animals showed impairment in the exzyme
activity with a subsequent fall in the rate of meatabolism.a similar effect was
observed in the pitutary growth harmone.
DISEASE STATUS: Reduction in hepatic drug metabolising ability is apparent
in conditions such as hepatic carcinoma,hepatits,cirrohsis,obstructive
jaundice,etc;.congestive heart failure,myocardial infarction results in decrease
in blood fllow to liver ,impaired metabolism of drugs having high hepatic
extraction ratio.
EX: Propanolol and lidocaine.
TEMPORAL FACTORS
CIRCADIAN RHYTHMS:
Diurnal variations or variations in the
Enzyme activity with light cycle is called as circadian rhythms in
drug metabolism.
It has been observed that the enzyme activity is
maximum during early morning(6-9 a.m) and minimum at late
afternoon(2-5 p.m) which was suggested to correspond with the
high and low serum levels of corticosterone
Study of variations in drug response as influenced by
time is called as chronopharmacology
Time dependent chnge in drug kinetics is known as
chronokinetics
REFERENCES
1. Text book of biopharmaceutics and pharmacokinetics by
v.venkateswarulu, page no135-160.
2. Text book of biopharmaceutics and pharmacokinetics a
treatise, page no139-193.
3. Bio-pharmacokinetics&clinical pharmacokinetics by
milo gibaldi,3rd edition,pageno187-193
4. Encyclopedia of pharmaceutical technology by james
swarbrick,3rd edition,volume-3.
5.
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