This document discusses biotransformation and drug metabolism. It covers topics such as the phases of metabolism, sites of drug metabolism, factors affecting metabolism, and consequences of metabolism such as activation, inactivation, and production of active metabolites. It also discusses concepts like first pass metabolism, enzyme induction, and enzyme inhibition which can impact drug metabolism. Microsomal enzymes like cytochrome P450 play a key role in phase I reactions like oxidation, while phase II involves conjugation reactions adding groups like glucuronic acid to make compounds more water soluble and able to be excreted.

1. Dr. kiran G Piparva
Assistant professor, Pharmacology department,
P.D.U. Government medical college, Rajkot.
12/2/21

2. •Biotransformation
•Enzymes
•Firs pass metabolism
•Microsomal enzyme inhibition
•Microsomal enzyme induction

3. What is biotransformation?
• Chemical alteration of drug in the body.
• Why need of biotransformation???
• So highly polar drugs (Streptomycin, neostigmine) --- Do NOT undergo
metabolism- excreted UNCHANGED form
Non polar (lipid soluble)
compound
Polar ( water soluble )
compound Eliminated by kidney

4. • If lipophilic drugs, or xenobiotics were not metabolized to polar,
readily excretable water-soluble products, they would remain indefinitely
in the body, eliciting their biological effects.
So metabolism is helpful for ………
 Termination of drug action
 Drug elimination- renal /bile/faces
 DETOXIFICATION – Toxin/ xenobiotics converted to non toxins
 Certain drugs pharmacologically inactive and converted to active

5. Site for drug metabolism
• Primary site – Liver
• Others – Kidney, Intestine, Lungs, Plasma
Consequences of
Biotransformation
Inactivation:
Inactive /less
active form-
termination of
action
Active metabolite from active drug
Drug partially converted to one or
more active form
• Drug effect is sumtotal of both
(parent drug+ active metabolite)
Activation of inactive
drug: Prodrug
Inactive drug need
conversion to active form
in body - PRODRUG

6. Prodrug
• Inactive drug- converted in body - active
form
• Advantages: More stable
Better bioavailability
Desirable pharmacokinetic
property
Less side effect/ toxicity
• Examples: Prodrug- Active form
• Levodopa - Dopamine
• Enalapril- Enalaprilat
• Prednisone- Prednisolone
• Dipivefrine -Epinephrine
Active drug active metabolite
• Active drug- Active metabolite
• Digitoxin –digoxin
• Codeine – morphine
• Imipramine- desipramine
• Morphine- morphine 6
glucuronide
• Spironolactone- canrenone

7. Phase 1 Reaction / Non Synthetic Phase
• Functionalization Reaction
• Functional groups (OH,-NH2, -SH,-COOH. -CHO) generated/exposed
• Metabolite: inactive/ active
• Reactions are: Oxidation
Reduction
Hydrolysis
Cyclization
Decyclization

8. Phase 2 Reaction / Synthetic Phase
• Conjugation reactions
• Conjugation of function compound with endogenous substrate to form water
soluble conjugated compound.
Reaction: Glucuronide conjugation
Acetylation
Methylation
Sulfate conjugation
Glycine conjugation
Glutathione conjugation
Ribonucleotide / Ribonucleoside synthesis

9. Phase I / Non Synthetic Reactions
• Oxidation: Addition of O2/ negatively charged radical or removal of H2/
positively charged radical.
• The reaction requires :
Molecular oxygen
Reducing agent NADPH
Mono oxygenase enzymes
• Final step: Involves cytochrome P-450, cytochrome P-450 reductase and
hemoprotein, NADPH, O2.
• In many cases initial insertion of O2 molecule produces short lived highly
unstable intermediates - Epoxides, Superoxide, Quinones.

10. RH + NADPH + O2 + H ________________ ROH + NADP+ + H2O
C,N
• Various oxidation reactions are:
• Hydroxylation
• Oxygenation at C,N or S atoms
• N or 0-dealkylation
• Oxidative deamination
• Majority drugs oxidized by CYTOCHROM “(CYP) :Barbiturate,
propranolol, paracetamol, steroids, benzodiazepines.
• CYP enzymes are susceptible to “ ENZYME INDUCER” and “ ENZYME
INHIBITIORS”.

11. • Monoamine Oxidase (MAO), Diamine Oxidase (DAO)
• Located on Mitochondria/ cytoplasm
• Oxidatively deaminates endogenous substrates including neurotransmitters,
Norepinephrine, Epinephrine, Dopamine, serotonin
• Alcohol & Aldehyde Dehydrogenase
• Ethanol metabolism
• Flavin Monooxygenases: on hepatic ER
• Require flavin adenosine dinucleotide (FAD)
• H2 blocker(Cimetidine, ranitidine), antipsychotic (clozapine), antiemetic (itopride)
• Not undergo INDUCTION/INHIBITON
Non-CYP Drug Oxidations

12. Oxidation by CYP 450
Majority drugs oxidized: Barbiturate,
propranolol, paracetamol, steroids,
benzodiazepines.
CYP enzymes are susceptible to “ ENZYME
INDUCER” and “ ENZYME INHIBITIORS”.
Oxidation by flavin
monooxygenase
Oxidation by mitochondrial/
Cytoplasmic enzyme : MAO, DAO
H2 blocker(Cimetidine, ranitidine),
antipsychotic (clozapine), antiemetic (itopride) -
on hepatic ER
Not undergo INDUCTION/INHIBITON
Neurotransmitters: Adr, NA, alcohol

13. Drug metabolising enzyme
• Microsomal enzyme
• Location: ER in liver, kidney,
intestinal mucosa, lung
• Examples: Monooxygenase
Cytochrome p450
UGTs,
Epoxide hydrolases
• Action: oxidation/
reduction/hydrolysis/glucuronide
conjugation
• Inducible: Drug/ Diet/ agents
• Nonmicrosomal enzymes
• Location: cytoplasm & mitochondria
of hepatic cells and other tissues,
plasma.
• Example: Esterase, Amidases,
flavoprotein oxidases , conjugates
• Action: oxidation/ reduction, many
hydrolytic and all conjugation reactions
except glucuronidation
NOT INDUCIBLE

14. Cytochrome enzyme (CYP)
• Monooxygenase enzyme are heme proteins.
• Major catalyst: in liver, kidney, G.I. tract, skin and lungs: oxidise drug and
endogenous compound.
• Location: Smooth endoplasmic reticulum (ER) in close association with
NADPH-CYP reductase in 10/1 ratio. The reductase serves as the electron
source for the oxidative reaction cycle
• Oxidative reactions require: CYP heme protein, the reductase, NADPH,
phosphatidylcholine and O2 molecular

15. Location of CYP in cell

16. CYP family……
• Multiple CYP gene families have been identified in humans, and the
categorized based on protein sequence homology.
• Families: CYP+ numeral (>40% amino acid sequence homology), eg.
CYP1
• Subfamily: designated by capital letters ; eg. CYP1A
• Subfamily: When more than 1 subfamily has been identified; eg. CYP1A2
• Most of the drug metabolizing enzymes are in CYP 1, 2, & 3 families

17. CYP 1A2
14%
CYP 2C9
14%
CYP 2C19
11%
CYP2
D6
23%
CYP2E
5%
CYP 3A4-5
33%
Relative hepatic content of CYP enzymes and
% drugs metabolized by CYP enzymes
14
CYP3A4 metabolize largest
number of drugs (50%) in liver
Its presence in the GI tract is
responsible for first pass
metabolism - poor bio
availability of many drugs

18. 2) Reduction
• Converse of oxidation –opposite direction (H is added to, or O2 is
removed from a compound)
• E.g. Alcohol, aldehyde, quinone
3) Hydrolysis : liver, intestine, plasma other tissue…
• Cleavage of drug molecule by taking up a molecule of water.
• Ester + water Esterase Acid + Alcohol
• Other hydroxylase (Nonmicrosomal) : Amidase, Polypeptides, epoxide
hydroxylase hydrolyses amide, peptides and epoxide generated by CYP oxygenase
Phase 1 reaction continue…

19. Cyclization
• Formation of ring structure from straight chain compound.
• E.g. Proguanil – cycloguanil
Decyclization
• Opening of ring structure of cyclic drug molecule.
• E.g Phenytoin
Minor
pathway
Phase 1 reaction continue…

20. Phase II reaction: conjugation reaction
• Types of phase II reactions
- Glucuronide conjugation
- Acetylation, Methylation
- Sulfate conjugation,
- Glycine conjugation
- Glutathione conjugation
- Ribonucleoside/ nucleotide synthesis
• Represent terminal inactivation – True detoxification reactions
Drug /metabolite + endogenous substrate = Polar, highly ionized
organic acid, inactive
High molecular weight,

21. 1. Glucuronide Conjugation
• Glucuronidation is the most common conjugative pathway
• Catalysed by UDP- Glucuronosyltransferases (UGTs)
• -OH, -COOH groups are easily conjugated with glucuronic acid
(Glucuronic acid derived from glucose)
• Drug + UDPGA Microsomal Glucuronyl transferase Drug glucuronide + UDP
• Endogenous substances: Bilirubin, steroidal hormone, thyroxin utilise this
pathway.
• Drugs metabolised by this pathway: paracetamol, diazepam

22. Glucuronidation
MW of conjugates
enhances excretion in bilirubin
Drug glucuronide hydrolysed by
bacteria in gut
Liberated drug reabsorbed
follow same fate
Enterohepatic circulation (EHC)
Enterohepatic circulation

23. 2. Acetylation
• Drugs with Amino or Hydrazine groups
• INH,PAS, Hydralazine, Sulphonamides
• R-NH N Acetyltransferase /Acetyl CoA
• Genetic polymorphism (NATs) : Acetylation- Rapid / Slow
R-NHCOCH3
3. Methylation
• Drug with Amine/ Phenols – methylated by methyl transferases (MT)
• Methyl donor: methionine, cysteine
• E.g : Adrenaline, histamine, nicotinic acid, captopril

24. 4. Sulfate Conjugation
• Phenolic compound and steroid – sulfated by sulfotransferases (SULTs)
• E.g. Adrenal and sex steroids
5. Glycine conjugation
• Minor pathway of metabolism
• Drugs having carboxylic acid – conjugated with glycine
• E.g. Salicylate, nicotinic acid

25. • Important pathway for detoxifying chemically
reactive electrophilic compounds.
• Carried by glutathione S transferase (GSTs)
• It serves to inactivate highly reactive
quinone/epoxide intermediates formed during
metabolism of certain drugs. E.g. paracetamol.
6. Glutathione Conjugation Minor pathway

26. • In case of poisoning of drug cases - when excess reactive intermediate
metabolite formed – glutathione supply falls short- toxic adducts are formed
with tissue constitutes resulting in hepatic, renal and other tissue damage.
• E.g. drug: paracetamol
• A severe reduction in GSH content can predispose cells to oxidative
damage
Glutathione Conjugation

27. 7. Ribonucleoside/ nucleotide synthesis..
• Activation of many purine and pyrimidine pathway- cancer chemotherapy
Hofmann elimination
• Inactivation of the drug in the body fluids by spontaneous molecular re
arrangement without the agency of any enzyme
• e.g. Atracurium.

28. Drug with -OH, -COOH groups
Glucuronidation Glucuronosyltransferases (UGTs)
Drug with - Amino or Hydrazine
groups
Drug with Amine/ Phenol groups
Acetylation
Sulfation
Glycine conjugation
Glutathione conjugation
Methylation
N acetyl transferases (NATs)
….UGTs)
Methyltransferases (UGTs)
Sulfoltransferases (UGTs)
Drug with Amine/ Phenol groups
Drug with carboxylic acid
Drug with highly reactive
quinone/epoxide
intermediates
Glutathione S transferases
(UGTs)

29. Phases of Metabolism
Phase 1 Reaction /Non Synthetic Phase
• Functionalization Reaction
• Introductionof functional groups such as
OH,-NH2, -SH,-COOH into the compound to
produce more water soluble compound
• Reactions are:
• Oxidation
• Reduction
• Hydrolysis
• Cyclization
• Decyclization
Phase 2 Reaction/Synthetic Phase
Conjugation reaction : conjugate with
endogenous substances
• Reactions are :
• Glucuronide conjugation
• Acetylation, Methylation
• Sulfate conjugation
• Glycine conjugation
• Glutathione conjugation
• Ribonucleoside/ nucleotide synthesis

31. Factors affecting drug metabolism
• Age
• Gender
• Race
• Nutrition level
• Hereditary or Genetic Factors
• Enzyme Induction
• Enzyme Inhibition
• Miscellaneous Factors Affecting Drug Metabolism
• Most drugs are metabolised by multiple pathway. Stereoisomers of a drug
may be metabolized differently and at different rate.

32. • Both enzymes are deficient in newborn (particularly premature) : take 3
months to reach adult level.
• Enzyme level controlled genetically and influenced by diet, environmental
factors: marked interspecies and interindividual differences –
interindividual variation.

33. First Pass (presystemic) Metabolism
• Metabolism of a drug during its passage from
the site of absorption into the systemic
circulation.
• Site: Liver (major site)
Gut wall
Gut lumen
Skin
Lung
• Bypasses oral route :changing modification of
dosage formulation sublingual , transdermal,
parenteral

34. • Extent of FPM varies from drug to drug – affect bioavailability
• Hepatic drug extraction: (ERliver) Fraction of the absorbed drug
prevented by the liver from reaching systemic circulation.
ER = ____CLliver______
Hepatic blood flow
So systemic bioavailability (F) : fractional absorption (I-ER)

35. Metabolism of drugs with high
hepatic extraction is dependent of
liver blood flow. E.g. propranolol
reduces rate of lignocaine
metabolism by decreasing hepatic
blood flow

36. Inhibition of drug metabolism
• One drug competitively inhibit microsomal enzyme CYP 450 and thereby
inhibit the metabolism of another drug/ endogenous substances (steroid/ bilirubin).
• Sometime drug may inhibit its own metabolism – “Autoinduction” when drug is
substrate to isoenzyme as well inhibit the same isoenzyme.
Clinically drug interaction is NOT much common because………..
• A drug may inhibit one CYP 450 but substrate to another CYP 450
• Different drugs are substrates to different CYP 450 and metabolized by non
saturation kinetics: enzyme is present excess

38. Drug interaction is clinically obvious ONLY when……when both
drugs having
- Affinity for same isoenzyme
- Metabolized by saturation kinetic /
- Or kinetic change from first order to zero order kinetic over the therapeutic
range. (capacity limited metabolism)
- Narrow therapeutic index and chronically administration –long time
- E.g. Boosted anti HIV drug regimen.
• low dose of ritonavir - inhibit CYP3A4 – so lower dose of other anti hiv
drugs (atazanavir, lopinavir, saquinavir)

39. Examples of drugs that inhibit drug metabolizing enzyme:
• Allopurinol
• Omeprazole
• Ketoconazole
• Cimetidine
• Quinidine
• Cigarette smoking

40. • Certain drugs, insecticides and carcinogens - Interact
with DNA- Enhances gene transcription for microsomal
enzyme protein- increase mass of enzyme
• Site :liver and other organ
• Increases metabolism 2-4 folds
• Induction takes 4-14 days to reach its peak and is
maintained till the inducing agent is present and return of
activity after 2-3 wk of withdrawal of inducing agents.
Microsomal Enzyme Induction

41. • Different inducers are relatively selective for certain
cytochrome P-450 enzyme families e.g.
• Phenobarbitone , rifampin, glucocorticoids induce CYP3A4
isoenzymes
• Isoniazid and chronic alcohol consumption induce CYP2E1
• Polycyclic hydrocarbons in cigarette smoking , charcoal boiled meat,
industrial pollutants induces CYP1A

42. • Decreased intensity of action of drug: contraception failure with
rifampicin co administration.
• Increased Intensity of action of drug: that activated by metabolism
• Interfere with adjustment of dose of another drug: ocpills, antiepileptic drugs
• Tolerance-autoinduction : carbamazepine, rifampin- double dose after 2wk
• Faster metabolism of endogenous substances: steroid, bilirubin
• Interference with chronic toxicity testing in animal
• Therapeutic uses of enzyme induction
1. Congenital hemolytic jaundice: phenobarbitone: enhance bilirubin metabolsim
2. Cushing syndrome: phenytoin- enhance degradation of steroid.
3. Chronic poisoning
Consequences of Induction……