BTP3822

BIO PHARMACEUTICS
Drug Metabolism
GROUP MEMBERS:
1) KIRTHANA GANESAN (TD13005)
2) AMIRA NAZIRA BT AZHAR (TD13016)
3) NURUL ISSTTIFA IYAH BT AMINUDDIN (TD10326)
4) SARMALA KRISHNASAMY(TD13008)
 Metabolism or Biotransformation

 breakdown of drugs by living organisms, usually through specialized enzymatic systems.

 xenobiotic metabolism is the set of metabolic pathways that modify the chemical structure of
xenobiotics, which are compounds foreign to an organism's normal biochemistry, such as any
drug or poison.
 the term metabolism is used because products of drug transformation are called metabolites.
 pharmacokinetic process which converts lipid soluble and non-polar compounds to water soluble
and polar compounds so that they are excreted by various processes.
 a series of enzyme- catalysed processes that alters the physiochemical properties of foreign
chemicals (drug/xenobiotics) from those that favour absorption across biological membranes
(lipophilicity) to those favouring elimination in urine or bile (hydrophilicity ).
 as drugs are metabolised their therapeutic effect diminishes.
 First Pass Effect

 biotransformation of drug by liver or gut enzymes before compound reaches

systemic circulation.
 results in lower systemic bioavailability of parent compound, diminished
therapeutic response
 First pass effect may be bypassed if the drug is administered intravenous or
sublingually.
 Role of metabolism process in drug activation

 conversion of an active drug to inactive or less active metabolites called as pharmacological

inactivation.
 exp : phenobarbitone hydoxyphenobarbitone
 conversion of an active drug to equally active metabolites (no change in pharmacological activity).
 diazepam oxazepam
 conversion of an inactive drug (pro-drug) to active metabolite(s) called as pharmacological activation.
 enalapril enalaprilat
 Site/Organ of drug metabolism

 major site of drug metabolism is the liver (microsomal enzyme systems of hepatocytes).
 the primary site for metabolism of almost all drugs because it is relatively rich in a large variety of
metabolising enzymes.
 the main enzymes involved in metabolism belong to the cytochrome P450 group.
 metabolism by organs other than liver (called as extra-hepatic metabolism) is of lesser importance
because lower level of metabolising enzymes is present in such tissues.
 within a given cell, most drug metabolising activity is found in the smooth endoplasmic reticulum
and the cytosol.
 drug metabolism can also occur in mitochondria, nuclear envelope and plasma membrane.
 Drug Metabolising Enzymes

 the drug metabolising enzymes can be broadly divided into two groups: microsomal and non-microsomal
enzymes.
 microsomal enzymes
- enzymes present in the endoplasmic reticulum in liver, kidney, intestinal mucosa and lungs.
- exp: microsomal cytochrome P450, monooxygenase family of enzymes, which oxidize drugs.
- catalyze oxidation, reduction, hydrolysis (phase I reactions) and glucuronide conjugation (phase II reactions)
 non-microsomal enzymes
-enzymes present in cytoplasm, mitochondria of hepatic cells .
-exp: monoamine oxidases (MAO), esterases, amidases, transferases, conjugages .
-catalyse all phase II reactions.
 Drug Toxicity

 toxicity refers to adverse effects of drugs in the body.

 these are the result of excessive pharmacological action of the drug due to
overdosing or prolonged use.
 toxicity may result from extension of the therapeutic effect of the drug.
 drug toxicity, also called adverse drug reaction (ADR) or adverse drug event
(ADE), is defined as the "manifestations of the adverse effects of drugs
administered therapeutically.
 some drug metabolites can be toxic such as those produced from paracetamol
and they can be detoxified by phase II conjugation joining with glutathione.
 Mechanism of drug toxicity

 On-target adverse effects

-result of the drug binding to its intended receptor, but at an inappropriate concentration, with
suboptimal kinetics, or in the incorrect tissue.
 Off-target adverse effects
-caused by the drug binding to a target or receptor for which it was not intended.
 Production of toxic metabolites
-virtually all drug molecules are metabolized by the liver and/or other tissues.
-sometimes metabolism produces a pharmacologically active metabolite, a drug metabolite
can have an adverse effect.
 Detoxification

 specific metabolic pathway, active throughout the human body, that processes unwanted
chemicals for elimination.
 involves a series of enzymatic reactions that neutralize and solubilize toxins, and transport
them to secretory organs (like the liver or kidneys), so that they can be excreted from the
body.
 the liver hepatocytes contain all the necessary enzymes for the detoxification of drugs.
 main enzymes involved belong to the cytochrome P450 group.
 Stages of Metabolism

 metabolism is often divided into two phases of biochemical reaction - phase 1 and phase 2.
 some drugs may undergo just phase 1 or just phase 2 metabolism, but more often, the drug will undergo
phase I and then phase II sequentially.
 Phase I
 involve reduction or hydrolysis of the drug, but the most common biochemical process that occurs is
oxidation.
 oxidation is catalysed by cytochrome P450 enzymes and results in the loss of electrons from the drug
(electrons are negatively charged subatomic particles).
 the drug is now said to be oxidised and after phase 1 reactions, the resulting drug metabolite is still often
chemically active.
 Phase II
 involves conjugation - that is, the attachment of an ionised group to the drug.
 these groups include glutathione, methyl or acetyl groups.
 these metabolic processes usually occur in the hepatocyte cytoplasm.
 the attachment of an ionised group makes the metabolite more water soluble.
 this facilitates excretion as well as decreasing pharmacological activity.
STAGE OF METABOLISM
 Phase 1

 Involve hydrolysis, oxidation and reduction.

 These reaction introduced a functional group (-OH, -NH2, -SH or –COOH)
 Increase in hydrophilicity.
 Reduction in stability by enabling reaction with cellular component.
 Resulting product for phase 1 reaction is susceptible to phase 2 by making xenobiotic highly water
soluble.
Oxidation

 Most important and common metabolic reactions.

 Increase hydrophilicity of xenobiotics by introducing polar functional group such as –
OH.
 Oxidation of xenobiotic is non-specifically catalyzed by a number of enzymes located in
the microsomes.
 It requires molecular oxygen and the reducing agent NADPH to effect reaction (mixed-
function oxidases).
 Mixed-function oxidases consist of three components which are cytochrome P-450,
cytochrome P-450 reductase (NADPH dependent) and phosphatidylcholine.
 The most important component of mixed-function oxidase is cytochrome P-450.
 Steps in oxidation of xenobiotics

 Binding of the substrate (RH) to the oxidized form of the cytochrome

P-450 (𝐹𝑒 3+ ) to form a complex.
 Transfer of one electron from NADPH to the complex by cytochrome P-450
reductase to form reduced (𝐹𝑒 3+ ) P-450 – substrate complex.
 Combination of reduced enzyme-substrate complex with a molecule of
oxygen to form ternary complex.
 Combination of ternary complex with a second electron supplied by NADH
to form a ternary activated oxygen P-450-substrate complex.
 Transfer of one atom of oxygen from the activated oxygen to the substrate Ternary complex
to yield the oxidized product. The other atoms form water. The free oxidized
form of cytochrome P-450 is now ready to attach to yet another molecule
of substrate. Cytochrome P-450 oxidation-reduction cycle
 Oxidative reaction

 Oxidation of aromatic carbon.

 Oxidation of olefin.
 Oxidation of benzylic allylic carbon atom and carbon atom alpha to carbonyl and imines.
 Oxidation of aliphatic carbon atoms.
 Oxidation of alicyclic carbon atoms.
 Oxidation of carbon heteroatom systems:
a) Carbon-nitrogen system
b) Carbon-Sulphur system
c) Carbon-oxygen system
 Oxidation of alcohol, carbonyl and acid function
 Miscellaneous oxidative reaction.
Reduction

 Capable of generating polar functional groups such as hydroxyl and amino which
can undergo further biotransformation or conjugation.
REDUCTIVE REACTION
 Reduction of carbonyl functions (aldehydes/ketones)
 Reduction of alcohols and carbon-carbon double bonds
 Reduction of N-compounds (nitro, azo, and N-oxide)
 Miscellaneous reductive reactions.
 Hydrolysis

 The reaction does not involve change in the state of oxidation of the substrate.
 The reaction results in a large chemical change in the substrate brought about by loss of relatively large fragment
of the molecule.
HYDROLYTIC REACTION
 Hydrolysis of esters and ethers.
 Hydrolysis of amides.
 Hydrolysis of cleavage of non-aromatic heterocycles
 Hydrolytic dehalogenation.
 Miscellaneous hydrolytic reactions.
 Phase II

 involve transfer of suitable endogenous moiety (ex:glucuronic acid,sulphate,glycine) in presence of enzyme

tranferase to drugs/metabolites of phase I reactions having suitable functional groups to form highly polar, readily
excretable and pharmacologically inert conjugates
 Real drug detoxification pathways, because:
1) conjugates/products absolutely free of pharmacological activity
2) conjugates/products highly polar [easily excretable either in bile/urine]
3) tissue-reactive and carcinogenic metabolites [formed from phase I] are rendered harmless by conjugation with
moieties
 Characteristics of moieties (conjugating reagents) :
1) simple endogenous molecules [carbohydrates,proteins,fats]
2) large molecular size
3) strongly polar/ionic in nature
 Characteristics of conjugation reactions:
1) involves initial activation step - (a)drug is activated
(b)conjugating reagent is activated
2) capacity-limited, attributed to – (a)amount conjugating agent
(b)ability to synthesize active nucleotide intermediate
(c)amount of enzyme conjugate transferase
 Molecular weight of conjugation and its route of excretion :
- high mw conjugates (>350) excreted in bile
- low mw conjugates (<250) excreted in urine
 Order of capacities of important conjugation reaction :
Glucuronidation > Amino Acid Conjugation > Sulphation & Gluthathione Conjugation
 Conjugation with Glucuronic Acid
 Also called as glucuronidation
 Most important : (a)readily available source of conjugating moiety
(b)several functional groups can combine easily with D-glucuronic acid
(c)conjugation occurs to a high degree
(d)all mammals able to produce glucuronides
(e)free carboxyl function of glucuronic acid has pKa [3.5-4.0] = ionisable at plasma and urine pH
(f)glucuronidation enzymes in close association with microsomal mixed-function oxidases
(g)can take place in most body tissues
 Occurs in 2 steps: (1)synthesis of activated coenzyme UDPGA from UDPG
(2)transfer of glucuronyl moiety from UDPGA to substrate RXH in presence of enzyme
UDP-glucuronyl tranferase to form conjugate
 Example : Oxygen/O-Glucuronides, Nitrogen/N-Glucuronides, Sulphur/S-Glucuronides, Carbon/C-Glucuronides
 Conjugation with Sulphate Moieties

 Catalysed by nonmicrosomal enzymes

 Occurs less commonly
 Easily saturable, dominant at low substrate concentration
 Occurs in 2 steps: (1)synthesis of activated coenzyme PAPS
(a)initial interaction between sulphate and ATP = yield ATP
(b)activation APS to PAPS
(2)transfer of sulphate group from PAPS to substrate RXH in presence of enzyme
 Functional groups : phenols, alcohols, arylamines, N-hydroxylamines, N-hydroxyamides
 Reaction product : sulphate ester
 Conjugation with Alpha Amino Acids

 Limited availability of amino acid

 Occurs in 2 steps: (1)activation of RCOOH with ATP, and coenzyme A(CoA) form acyl CoA int.
(2)acylation of -amino acid by acyl CoA, with enzyme N-acyl transferase
 Occurs commonly with glycine, glutamine lesser, other amino acids (aspartic acid,
serine,taurine) still uncommon
 Substrate generally acid (aromatic in particular)
 Reaction product : amide
 Example of drugs forming glycine/glutamine conjugates:
-aliphatic acids, alicyclic acids ,aryl acids, arylacetic acids, heterocyclic aryl acids
 Occurs extensively in liver mitochondria, reaction can be used to estimate hepatic function
 Conjugation with Glutathione &
Mercapturic Acid Formation

 Gluatathione: tripeptide with strongly nucleophilic character (presence of –SH (thiol) group)
 Has great affinity for electrophilic substrates
 Different : process does not require initial activation of coenzyme/substrate
 Process :

 Occurs by 2 mechanisms: (1)nucleophilic substitution

(2)nucleophilic addition
 Acetylation

 Basically acylation reaction, similar to conjugation with -amino acids

 Different: substrates are exogenous amines, acylating agent is endogenous acetyl CoA
 Enzyme: nonmicrosomal N-acetyl transferase
 Examples drugs undergoing acylation:
-primary aliphatic amines, primary aromatic amines, sulphonamides, hydrazines/hydrazides
 Methylation

 Different: (1)metabolites formed not polar/water-soluble

(2)metabolites have equal/greater pharmacological activity than parent drug
(3)reaction less importance in metabolism of xenobiotics. More important in biosynthesis
and activation of endogenous amine
 Considered as phase I and phase II reaction
 Occurs in 2 steps: (1)synthesis of activated coenzyme SAM, from L-methionine and ATP
(2)transfer of methyl group from SAM to substrate in presence of
nonmicrosomal enzyme methyl transferase
 Examples substrates undergoing methylation:
- O-methylation: phenols, catechols
- N-methylation: Primary aliphatic amines, secondary alicyclic amines, aromatic heterocycles
- S-methylation
 Miscellaneous Conjugation Reaction

 Conjugation of Cyanide: involves transfer of sulphur atom from thiosulphate to cyanide ion in
presence of enzyme rhodanese to form nucleotides
 Conjugation with Ribose: endogenous purine and pyrimidine bases conjugate with ribose to form
nucleotides
 Conjugation with Taurine: Taurine, β-amino sulphonic acid, conjugates endogenous bile acids to
produce components of bile
 FACTORS AFFECTING
BIOTRANSFORMATION OF DRUG
1) Physicochemical properties of the drug

2) Chemical factors
a. induction of drug metabolising enzymes
b. inhibition of drug metabolising enzymes
c. environmental chemicals

3) Biological factors
a. species differences
b. strain differences
c. sex differences
d. Age
e. Diet
f. altered physiological factors - pregnancy ,hormonal imbalance, disease
states
g. Temporal factors -circadian rhythm ,circannual rhythm
 Physiochemical Properties Of The Drug

 Molecular size and shape

 pKa
 Acidity/basicity
 Lipophilicity
 Steric and electronic characteristics

Molecular size and shape, pKa , acidity/basicity. Lipophilicity and stearic and
electronic characteristics of a drug influence its interaction with the active sites of
enzymes and the biotransformation processes.
 Chemical Factors

 Enzyme induction
 Enzyme inhibition
 Environmental chemicals
 Enzyme induction

 Phenomenon of increased drug metabolising ability of the enzymes by several drugs and chemicals
 2 categories of inducers :
1. phenobarbital type (several drugs & pesticides)
2. Polycylic hydrocarbon type (3-methyl cholanthrene & cigarette smoke)
 Self induction or auto induction: phenomenon in which a drug induces metabolism of other drugs as
well as its own.
Eg:carbamazepine, meprobamate , cyclophosphamide, rifampicin
 Enzyme Inhibition

 Decrease in the drug metabolising ability of an enzyme

 Direct or indirect inhibition
 Direct inhibition :
1. competitive inhibition
2. non-competitive inhibition
3. product inhibition
 Indirect inhibition:
1. repression eg: ethionine,puromycin,actinomycin D,CCl4,carbon
disulphide,disulfiram
2. altered physiology
 Environmental chemicals

 Halogenated pesticides & polycyclic aromatic hydrocarbons

 Organophosphate insecticides & heavy metals
 Temperature, altitude, pressure, atmosphere etc..
Biological factors

species differences

strain differences

sex differences

Age

Diet
 Species differences

 Qualitative & quantitative variation

 In general, drugs are metabolized in man more slowly than in laboratory animal
with the exception of cat.
 Eg: Hexabarbitone rapidly metabolized in mouse(1/2life 19min.) whereas slow in
man (1/2life 360min.).
 Eg: Amphetamine metabolized by oxidative deamination in men while in rat by
aromatic oxidation
 Strain differences

 Drugs behave differently in different individuals due to genetic variations

 A study of inter-subject variability in drug response is called as
pharmacogenetics.
 The inter-subject variations in drug biotransformation may either be
monogenically or polygenically controlled
 Differences observed in the metabolism of a drug among different races is called
as Ethnic variations.
 Sex Differences

 Sex related differences in the rate of metabolism can be attributed to regulation of

such processes by sex hormones since variations among male & female are
generally observed following puberty.
 Eg:
• Women metabolize Benzodiazepines slowly than men
• women on contraceptive pills metabolize a number of drugs at a slower rate.
 Age

 Differences in the drug metabolic rate in different age groups is mainly due
to variations in the enzyme content, enzyme activity & homodynamics
• In neonates: microsomal enzyme system not fully developed.eg:
caffiene
• In infants: same as neonates
• In children: metabolise certain drugs much more rapidly than adults
• In elderly persons: liver size reduced-microsomal enzyme activity is
decreased and hepatic blood flow also declines
 Diet

 The enzyme content and activity is altered by a number of dietary components:

 Low protein diet and high protein
 Protein-carbohydrate ratio
 Fat free diet
 Dietary deficiency of vitamins and minerals
 Grapefruit
 Starvation
 Malnutrition in women
 Alcohol ingestion
ALTERED PHYSIOLOGICAL FACTORS

Pregnancy

Hormonal Imbalance

Disease states
 Altered Physiological Factors

Pregnancy Hormonal imbalance Disease states

• maternal drug • higher level of one
metabolising ability is
reduced-due to high
levels of steroid
hormones
• Ex: the metabolism of
Promazine , Pethidine is
reduced during
pregnancy.
• Liver disease such as
hormone may inhibit the hepatic carcinoma,
activity of few enzymes hepatitis, cirrhosis,
while inducing that of obstructive jaundice
others etc..reduce the hepatic
drug metabolizing ability
& thus increase the half
lives of almost all drugs.
 Temporal factors

 Circadian rhythm : Diurnal variations or variations in the enzyme activity with

light cycle.
 Enzyme activity is maximum during early morning & minimum in late afternoon
which is probably due to high & low serum levels of corticosterone .
 conversion of an active to more active metabolites called as bio activation or toxicological
activation.
 conversion of an inactive to more active toxic metabolites called as lethal synthesis.
 conversion of an inactive drug (pro-drug) to active metabolite(s) called as
pharmacological activation.
 conversion of an active drug to equally active metabolites (no change in pharmacological
activity).
 conversion of an active drug to active metabolites having entirely different
pharmacological activity (change in pharmacological activity).