DRUG

METABOLISM

The Fate of a Drug

Pharmaceutics

ABSORPTION

DISTRIBUTIONPROTEIN

BOUNDFREE

PLASMA

MOST TISSUES

NONSPECIFIC

BINDING

BIOPHASE

RECEPTOR

BINDING

DOSE

EFFECT

ELIMINATION

RENAL

EXCRETIONMETABOLISM

Pharmacokinetics Pharmacodynamics

Pharmaceutics

ABSORPTION

DISTRIBUTIONPROTEIN

BOUNDFREE

PLASMA

MOST TISSUES

NONSPECIFIC

BINDING

BIOPHASE

RECEPTOR

BINDING

DOSE

EFFECT

ELIMINATION

RENAL

EXCRETIONMETABOLISM

Pharmacokinetics Pharmacodynamics

Pharmaceutics

ABSORPTION

DISTRIBUTIONPROTEIN

BOUNDFREE

PLASMA

MOST TISSUES

NONSPECIFIC

BINDING

BIOPHASE

RECEPTOR

BINDING

DOSE

EFFECT

ELIMINATION

PROTEIN

BOUNDFREE

PLASMA

MOST TISSUES

NONSPECIFIC

BINDING

BIOPHASE

RECEPTOR

BINDING

DOSE

EFFECT

ELIMINATION

RENAL

EXCRETIONMETABOLISM

Pharmacokinetics Pharmacodynamics

Drug at site Drug at site

of administrationof administration

Drug in plasmaDrug in plasma

Drug/metabolitesDrug/metabolites

in urine, feces, bilein urine, feces, bile

Drug/metabolitesDrug/metabolites

in tissuesin tissues

1. Absorption

2. Distribution

4. Elimination

3. Metabolism

BIOTRANSFORMATION

Chemical modification of drugs or foreigncompounds (xenobiotics) in the body.

The apparent function of drug or xenobioticsmetabolism is their transformation into water-soluble derivative which can be easily eliminated viarenal route.

Implications For Drug Metabolism1. Termination of drug action

2. Activation of prodrug

3. Bioactivation and toxication

4. Carcinogenesis

Objective of studying drug metabolism

- medicinal chemists aware of the chemicalprocesses involved in the biotransformation ofdrugs- so design of new more safe drugs.

BIOTRANSFORMATION CAN LEAD TO THE FOLLOWING

Inactivation

Eg: drugs like paracetomol, ibubrufen, chloromphenical etc and their

metabolites are rendered inactive or less active.

Active metabolite from active drug

Activation of inactive drug

Certain drugs are inactive and need activation in the body.

Prodrug gives more bioavailability and less toxic effects.

ACTIVE DRUG ACTIVE METABOLITE

allopurinol alloxanthine

Digitoxin Digoxin

Morphine Morphine 6 glucoronide

Cholrol hydrate trichloroethanol

Prodrug Active metabolite

Levodopa Dopamine

Sulindac Sulfide metabolite

Predinosone Prednisolone

Effect of drug metabolism

Active drug Inactive Drug

Active drugInactive prodrug

Inactive metabolite

Metabolic activation

Lipid Solubility

R-H R-OHChemical hook

Two categories of xenobiotics are not or arehardly subject to metabolic transformations :

1- Hydrophilic compounds (e.g. saccharine, strongacids or bases)

S

NH

O

OO

2- Highly lipophilic polyhalogenated xenobioticssuch as some insecticides e.g. DDT.

Lethal synthesis: a classic example of lethalsynthesis is provided by the metabolic conversionof the nontoxic insecticide parathion into itsoxygenated isostere paraoxon, a potentacetylcholinesterase inhibitor

O2N O-P-OEt

OEt

S

O2N O-P-OEt

OEt

O

Parathion Paraoxon

Fig.2. Metabolic conversion of the nontoxic insecticide parathion to paraoxon

Major Sites of Metabolism

Drug metabolism can occur in every tissue (e.g. gut, lung,kidney)..

The LIVER is the chief organ for drug metabolismbecause:

– The blood flow through the liver is high

– Hepatocytes contain numerous metabolic enzymes

High liver

Medium lung, kidney, intestine

Low skin, testes, placenta, adrenals

Very low nervous system

Hepatic microsomal enzymes(oxidation, conjugation)

Extrahepatic microsomal enzymes(oxidation, conjugation)

Hepatic non-microsomal enzymes (acetylation, sulfation,GSH, alcohol/aldehyde dehydrogenase,hydrolysis, ox/red)

Biotransformation

Nonsynthetic/ phase I/ functionalization rxn’s Synthetic/ phase II/

conjugation

Non-synthetic/ phase I/ functionalization rxn’s

Oxidation

reduction

hydrolysis

cyclization

Decyclization

Oxidation

Reduction

Hydrolysis

• hydroxylationsaromatic, aliphatic, nitrogen

• dealkylations(N-, S-, P)• deaminations• N-, S-, P- oxidations• S-replacements• epoxidations• others

• azo reduction• nitro reduction• disulfide reduction• others

• esters• amides

oxidoreductases

oxidasesmonoamine oxidasesmixed function oxidases

oxidoreductases

reductases

esterases

amidases

peptidases

lipases

PHASE I

Oxidation

Involves addition of oxygen or removal of hydrogen.

Reactions mostly occur in liver by a group of

mono-oxygenases,

cyt P450 reductase

oxygen.

Various oxidation reactions are

Hydroxylation

Oxygenation at C,N or S atoms

N or O dealkylation

Oxidative deamination

Eg: barbiturates, phenothiazines, steroids, paracetomol.

60% of drugs are metabolized primarily by CYPs.

The great majority of these oxidations arecarried out by the haemoprotein cytochrome P-450 which is embedded within the phospholipidenvironment of the microsomes derived from theendoplasmic reticulum of living cells.

1. Oxidation reactions

Cytochrome P450 (CYPs)

• Cytochrome P450 is a family of enzymes locatedin the endoplasmic reticulum of liver.

•When liver is homogenized and biochemically fractionatedthese enzymes are found in the microsomal fraction (smallclosed ER membrane fragments). Thus these enzymes arecalled microsomal enzymes.

Cytochrome P450 enzymes perform many of the mostimportant biotransformation reactions.

These enzymes oxidize a wide variety of compoundsforeign to the body.

There are at least 18 different forms of cytochromeP450 identified in humans, produced by different genes.

Carbon monoxide binds to the reduced Fe(II) heme andabsorbs at 450 nm (origin of enzyme family name).

60% of drugs are metabolized primarily byCYPs.

• Important cytochrome P450 enzymes:

1. CYP3A4. 2. CYP1A2.

3. CYP2C. 3. CYP2D6.• Individual enzymes differ in their substrate specificity

and regulatory properties.

The reaction sequence illustrated by CyP Rrdox Cycle :

OH

CYP450

Fe3+CYP450

Fe3+

CYP450

Fe3+CYP450

Fe3+

CYP450

Fe2+CYP450

Fe2+

CYP450

Fe2+CYP450

Fe2+CYP450

Fe2+CYP450

Fe2+

CYP450

Fe3+CYP450

Fe3+

CYP450

Fe3+CYP450

Fe3+

DRUGDRUG

DRUGDRUG

DRUGDRUGDRUGDRUG

DRUGDRUG

DRUGDRUG

DRUGDRUG

CYP450

reductase

NADPH + H+

e-

O2

O2

e-

O21-

H2O

..:

CYP450 Reaction Sequence

O

NADPH + H+

OH

H+

DRUGDRUG

2. Reduction reactions

Reduction

Involves removal of oxygen or addition of hydrogen.

Reactions mostly occur in liver by a group ofcyt P450 reductase

Eg: Alcohols, Warfarin, Chloramphenicol etc.

It is a major route of metabolism for aromaticnitro and azo compound as well as for a widevariety of aliphatic and aromatic N-oxides whichare reduced to tertiary amines.

Hydrolysis

Cleavage of drug by addition of water.

Ester + water acid + alcohol in presence of enz esterase.

Occurs in liver, intestine , plasma.

Similarly, amides and polypeptides are hydrolysed by amidases and

peptidase enz.

Eg: Aspirin, procaine, oxytocin etc.

Cyclization

Involves formation of ring structure from straight compound.

Eg: Proguanil

De- Cyclization

Involves formation of open structure from ring compound.

Eg: barbiturates, phenytoin etc.

Is a minor pathway.

Synthetic/ phase II/ conjugation

Glucoronideconjugation

Sulphateconjugation

Ribonucleotide/ nucleotide conjugation

Acteylation

Methylation

Glycineconjugation

Glutathione conjugation

Phase II or Conjugation Reactions

– Conjugation reactions link an endogenous moiety(an endocon) to the original drug (if polar functions arealready present) or to the Phase I metabolite.

They are catalyzed by enzymes known as transeferases.

They involve a cofactor which binds to the enzyme in theclose proximity of the substrate and carries the endogenousmolecule or moiety to be transferred.

Forms highly ionized organic acid which is excreted in urineor bile.

Generally conjugation reactions have high energyrequirements.

R SH R S GLU

Glucoronide conjugation

Most important reaction carried out by UDP glucoronsyl transferases

(UGT).

Drugs with hydroxyl and carboxylic acid are easily conjugated with

glucoronic acid.

Glucoronidation increases the mol. Wt of the drug and thus favors excretion

in bile.

Drug excreted in gut by bile is hydrolyzed bacteria and is reabsorped back

and undergo same process, so this enterohepatic circulation prolongs the

action of drugs.

Eg: Aspirin , paracetamol, morphine etc.

Beside xenobiotics, a number of endogenoussubstrates, notably bilirubin and steroids areeliminated as glucuronide conjugates.

In neonates and children, glucuronidationprocesses often are not fully developed. In suchsubjects, drugs and endogenous compounds (e.g.bilirubin) that are normally metabolized byglucuronidation may accumulate and cause serioustoxicity. For example, neonatal hyperbilirubinemia and graybaby syndrome, result from accumulation of toxiclevels of the free chloramphenicol.

Acetylation

Drugs having amino or hydrazine are conjugated by acetyl co-enzyme.

Multiple genes control the N-Acetyl transferases and rate of acetylation shows

genetic polymorphism

Eg: Sulphonamides, hydralazine, clonazepam etc.

Methylation

Methionine and cysteine are methyl donors.

Amines and phenols are methylated.

Eg: Adrenaline, histamine, nicotinic acid etc.

Suphate conjugation

Phenolic compounds and steroids are sulphated by sulfotransferases.

Eg: Cholramphenicol, Adrenal, Methyldopa etc.

Glycine Conjugation

Salicylates and other drugs with carboxylic acid are conjugated with glycine.

Glutathione Conjugation

is a minor pathway.

It also serves to inactivate highly reactive quinone or epoxide intermediate.

Presence of large amount of adducts results in tissue damage.

E.g. Paracetomol

Ribonucleoside / nucleotide synthesis

is a important pathway for activation of drugs in chemotherapy.

Mainly purine and pyramidine drugs are used.

Conjugation with glutathione (GSH)– Glutathione (GSH, -glutamylcysteinylglycine) is athiol-containing tripeptide.– GSH conjugation is an important pathway by whichchemical reactive electrophilic compounds aredetoxified.

– Many serious drug toxicities may be explainablealso in terms of covalent interaction ofmetabolically generated electrophilicintermediates with cellular nucleophiles.

– GSH protects vital cellular constituents againstchemically reactive species by virtue of itsnucleophilic sulfhydryl (SH) group.

Unlike other conjugative phase II reactions, GSHconjugation does not require the initial formation ofan activated enzyme or substrate.

–Two general mechanisms are known for the reactionof compounds with GSH:

1. Nucleophilic displacement at an electron-deficientcarbon or heteroatom.2- Nucleophilic addition to an electron-double bond.

Excretion

Drugs and its metabolites are excreted in

Urine

Faeces E.g. erythromycin, ampicillin, rifampin etc.

Exhaled air e.g. anesthetics, alcohol etc.

Saliva & sweat e.g. lithium, rifampin etc

Milk

Renal excretion

Renal excretion is responsible for excreting almost all drugs.

Net renal excretion (glomerular filtration + tubular secretion)

(or) the amount = - tubular reabsorption

of drug in urine

Glomerular filtration

Glomerular capillaries have pores larger and thus non-

protein bound drug is filtered in the glomerulus, thus gfr

filtration drug depends on plasma binding protein and renal

blood flow (irrespective of lipid soluble and lipid insoluble

drugs).

Glomerular filtration rate (g f r) ~ 120ml/min and declines

progressively after the age of 50 and is low in case of renal

failure.

Tubular reabsorption

Occurs by passive diffusion

Depends on lipid solubility and ionization of drug at

Tubular secretion

This is active transfer of organic acids and bases by two separate class of

transporters OCT &OAT.

In addition, efflux transporter P-GP &MRP2 are located in luminal

membrane of proximal tubules.

Active transport across tubules reduces concentration of its free form in the

tubular vessels and promotes dissociation of protein bound drug which again

is secreted.

Renal excretion

Renal excretion is responsible for excreting almost all drugs.

Net renal excretion (glomerular filtration + tubular secretion)

(or) the amount = − tubular reabsorption

of drug in urine

Kinetics of Elimination

Bioavailability (F), Vol. of distribution (V) and clearance (CL).

Clearance = Rate of elimination / conc. of drug in plasma.

Drug is eliminated by zero and first order reaction.

Half - life

The half -life of a drug is the time taken for its plasma concentration

to be reduced to half its original value .

T1/2 = 0.693 * vd/ CL

Where vd – vol. of distribution

CL – clearance

Factors affecting half – life

As patients age the skeletal muscle mass decreases which could decrease

the vol. of distribution.

In obese person, due to increased adipose tissue higher dose of drug to be

given to reach the target organ.

Some of drug get into the pathologic fluid space like ascites, pleural

membrane causing long term toxicity.

Factors affecting half – life Most common effect on half-life

Effects on volume of distributionAging

decreased

Obesity Increased

Pathologic fluid increased

Effects on clearanceCyt P 450 induction (increased metabolism)

decreased

Cyt P 450 inhbition( decreased metabolism)

Increased

Cardiac failure (decreased clearance) Increased

Hepatic failure (decreased clearance) Increased

Renal failure (decreased clearance) Increased

ENZYME INDUCTION

• Certain drug substrates of CYP, upon repeatedadministration, “induce” or elevate the amountsof specific forms of the enzyme.

• Induction results in accelerated metabolism ofthe drug inducer and any other drugsmetabolized by the induced enzyme.

Classic Enzyme Inducers

• Chronic ethanol

• Phenobarbital

• Tobacco (benzo[a]pyrene)

• PCBs (Poly chlorinated biphenyls), dioxin (environmental)

• Glucocorticoids

ENZYME INHIBITION

• Some drugs are capable of inhibitingmetabolizing enzymes.

• This is usually only important if a patient on astable drug regimen starts taking a second drugmetabolized by the same enzyme.

• Examples: cimetidine, ethanol

Drugs that Inhibit Drug Metabolism by FormingComplexes with CYPs

Amphetamine Itraconazole

Cimetidine Ketoconazole

Dapsone Methadone

2,5-Dimethoxy-4-

methylamphetamine

Methamphetamin

eNortriptyline

Diphenylhydramine SKF 525A

Erythromycin Sulfanilamide

Fenfluramine

Non-nitrogenous Substances that Effect Drug Metabolism by Forming Complexes with CYPs

• Grapefruit juice - CYP 3A4 inhibitor; highlyvariable effects; unknown constituents

• Isosafrole, safrole - CYP1A1, CYP1A2 inhibitor;found in root pear, perfume

• Piperonyl butoxide & alcohol -CYP1A1, CYP1A2inducer; insecticide constituent

Effect of Grapefruit Juice on Felodipine Plasma Concentration

N

CO 2CH3CH3O2C

CH3 CH3

H

H

Cl

Cl

3A4

N

CO 2CH3CH3O2C

CH3 CH3

Cl

Cl

5mg tablet with juice

without

STATE OF HEALTH

• Liver disease

• Kidney disease

• Endocrine diseases

• Reduced blood flow

Optimising Pharmacokinetic Properties

Designing Prodrugs and Bioprecursors

Prodrug is any compound that undergoesbiotransformation prior to exhibiting itspharmacological effects.

Prodrugs can be divided into two classes:

The Carrier-prodrugs :

Result from a temporary linkage of the activemolecule with a transport moiety that isfrequently of lipophilic nature.

Example:

Aspirin for salicylic acid

Prodrugs to mask toxicity and side effects

• Mask groups responsible for toxicity/side effects

• Used when groups are important for activity

Salicylic acid• Analgesic, but causes stomach

ulcers due to phenol group

Aspirin• Phenol masked by ester

• Hydrolysed in body

OH

CO2H O

CO2H

O

H3C

Drug

Barrier to

delivery of

drug

DrugBarrier to

delivery of

drug

Pro-moiety

Pro-moiety Pro-moiety

Drug

Biotransformation

Efficacy

Drug

++

Non-toxic &

rapidly excreted

Practical applications of carrier-prodrug design

To1. Improve of the lipophilicity2. Enhancement of water solubility3. mask toxicity and side effects

4. Increase the duration of Pharmacological effects

5. Mask polar groups

6. target drugs

7. Improvement of patient acceptance8. Increased site-specificity

Prodrugs to increase water solubility

• Often used for i.v. drugs

• Allows higher concentration and smaller dose volume

• May decrease pain at site of injection

Example:

Succinate ester of chloramphenicol (antibiotic)

Succinate ester

O2N

OH

HN

O

O

Cl

ClH

H

O

OHO

Esterase

Chloramphenicol

O2N

OH

HN

O

OH

Cl

ClH

H

Prodrugs to increase water solubility

Example:

Phosphate ester of clindamycin (antibacterial)

• Less painful on injection

CO N

HC

C

Cl

CH3

O

H

HO

OH

OPO32-

SCH3

H

H

H

MeN

H

H

H

H

CH3CH2CH2

Prodrugs to increase water solubility

Example:

Lysine ester of oestrone

• Lysine ester of oestrone is better absorbed orally than oestrone

• Increased water solubility prevents formation of fat globules in gut

• Better interaction with the gut wall

• Hydrolysis in blood releases oestrone and a non toxic amino acid

H2NO O

NH2

OMe

Prodrug

HH

H

H

H2NO OH

NH2

OMe

HO

Lysine Oestrone

H

H HH

+

Increase the duration of Pharmacological effects

Add hydrophobic groups

Example:

Hydrophobic esters of fluphenazine (antipsychotic)

• Given by intramuscular injection• Concentrated in fatty tissue• Slowly released into the blood supply• Rapidly hydrolysed in the blood supply

S

HN CF3

N

N

O (CH2)8CH3

O

fatty ester

Example:

Azathioprine for 6-mercaptopurine

6-Mercaptopurine(suppresses immune response)

• Short lifetime - eliminated too quickly

Azathioprine• Slow conversion to 6-mercaptopurine

• Longer lifetime

Mask polar groups

• Reduces rate of excretion

N

H

SH

NN

N

N

NN

N

S N

N

O2N

Me

H

Improvement of patient acceptance

O2N CH CH CH2

NHCOCHCl2OH

O CO(CH2)14CH3

– Used to reduce solubility of foul tasting orally active drugs

– Less soluble on tongue– Less revolting taste

Example:

Palmitate ester of chloramphenicol (antibiotic)

Palmitate ester

O2N

OH

HN

O

O

Cl

ClH

H

O

Esterase

Chloramphenicol

O2N

OH

HN

O

OH

Cl

ClH

H

Prodrugs used to target drugs

Example:

Hexamine

• Stable and inactive at pH>5

• Stable at blood pH

• Used for urinary infections where pH<5

• Degrades at pH<5 to form formaldehyde (antibacterial agent)

N

N

N

N

Increased site-specificity.

Two targeting possibilities can be considered:Site-directed drug deliverySite-specific drug release

Chemical Delivery Systems: Pharmacologically inactive

molecules that require several steps of chemical and/or

enzymatic conversion to the active drug and enhance drug

delivery to particular organs or sites. Example: Brain-specific

chemical delivery system

The Bioprecursors :

Result from a molecular modification of the active principleitself. This modification generates a new compound, able tobe a substrate for the metabolizing enzymes (often,oxidation and reduction). The metabolite being theexpected active principle.

Example for bioprecursor design based on oxidative bioactivation is the cytotoxic agent cyclophosphamid.

Cyclophosphoramide for phosphoramide mustard

(anticancer agent)

Cyclophosphoramide• Non toxic

• Orally active

Phosphoramide mustard• Alkylating agent

Phosphoramidase

(liver)

O

P

NH O

N

Cl

Cl

HO

P

Cl

Cl

N

OH2N

Sulindac (NSAID) is a representative examplefor bioprecursor bioactivated by reduction.

CH3

CO2H

S

CH3

O

F

CH3

CO2H

S

CH3

F

Reduction

Inactive Active

In vivo

Soft DrugsBiologically active, therapeutically usefulchemical molecules (drugs) characterized by apredictable and controllable in vivo deactivation,after achieving the therapeutic objective, tonontoxic, inactive compounds.

Soft Drugs Examples:

O

O

O

O

N

N

Cl

Cl

Acetylcholine Estrase

OH

OH

O

O

HO

N Cl+

Succinylcholine Chloride Succinic acid Choline Chloride

Hard drugs are "non-metabolizable drugs" ordrugs which are metabolized to biologicallyactive metabolites.

An example of a hard compound is the insecticide DDT(1,1,1-trichloro-2,2-bis(p-chlorophenyl)ethane)