drug metabolism/ biotransformation/ pharmacokinetics

1. BIOTRANSFORMATION
(Metabolism of drugs)
Dr. Arun kumar. R
PGY1
Department of Pharmacology
SRM MCH & RC

2. OUTLINE
➢ Introduction
➢ Phases of metabolism
➢ Phase 1 metabolism
➢ Cytochrome P family
➢ Phase 2 metabolism
➢ First pass metabolism
➢ Microsomal drug induction
➢ Drug inhibition
➢ Factors affecting biotransformation
➢ Biotransformation in drug development
➢ Take home message

3. Clinical vignette
Q. A 68 year old woman is brought to the emergency for treatment of chest
pain. She is currently taking aspirin and clopidogrel daily, as well as omeprazole for
heartburn. What can be the most likely contributor to her symptoms?

4. Biotransformation
Chemical alteration of drug in the body that converts non polar or lipid soluble compounds
to polar or lipid insoluble compounds
Why does metabolism occur ?
The many therapeutic agents that are lipophilic, hydrophobic do not pass readily into the
aqueous environment of the urine, in the absence of metabolism, they accumulate in fat and
cellular phospholipid bilayers
Xenobiotics - substances foreign to the body, can be from natural or artificial sources.

5. The metabolism of drugs and other xenobiotics into more hydrophilic metabolites is essential for
termination of their biological, pharmacological activity and can easily be eliminated from the
body through the urine or the bile
Consequences:
• Inactivation - Formation of an inactive metabolite from an active drug, that is readily
excreted from the body (Ex: Phenobarbitone Hydroxyphenobarbitone)
• Active metabolite from an active drug (Ex: Diazepam Oxazepam)
• Activation of inactive drug (Generate biologically active metabolites, Ex: Levodopa)
• Toxic to non toxic metabolite

6. Sites :
Liver, GI tract, Kidneys, Lungs, Plasma
Liver: Major site (Eg: Morphine, Prazosin,
Paracetamol, Lidocaine, Nitroglycerine)
GIT: Insulin, Catecholamines, Salbutamol,
Clonazepam, Chlorpromazine
Lung: Prostanoids
Plasma: Succinylcholine, Adenosine
Placenta: Alcohol

7. 1. Phase 1 reactions/ Non-synthetic [oxidation/ reduction/ hydrolysis,
CYPs]
Introduction of functional groups such as (–OH, –COOH, –O–, or NH2) and converts parent
drug to more polar metabolite.
1. Phase 2 reactions/ Synthetic [conjugation]
Conjugation reactions in which a covalent linkage is formed between a functional group
on the parent compound or phase 1 metabolite and an endogenous substrate like glucuronic acid,
sulfate, acetate or an amino acid.
Biotransformation reactions

8. Q. Which of the following reactions represents Phase II of drug
metabolism ?
a) Amidation
b) Hydrolysis
c) Oxidation
d) Reduction
e) Sulfation

9. Phase 1 reactions/
Non-synthetic
Phase 2 reactions/
Synthetic
Biotransformation
● Oxidation
● Reduction
● Hydrolysis
● Cyclization
● Decyclization
● Glucuronide conjugation
● Acetylation
● Methylation
● Sulphate conjugation
● Glycine conjugation
● Glutathione conjugation
● Ribonucleoside or
nucleotide synthesis

10. Phase I Phase II
oxidation/ reduction/
hydrolysis
Drugs directly enter Phase
II metabolism
Conjugation drug is
usually inactive
After Phase I the drug
may be activated,
inactivated or unchanged
DRUG/
XENOBIOTICS
Conjugation products

11. Biotransformation reactions
Bertram G. Katzung , Todd W. Vanderah ,Basic & Clinical Pharmacology 15th Edition

12. Metabolising enzymes
Goodman & Gilman's the pharmacological basis of therapeutics

13. Drug metabolising enzymes : Microsomal enzymes
Site: These enzymes are present in the smooth endoplasmic reticulum of the liver, lungs,
kidney and intestinal mucosa.
Principal enzymes:
Cytochrome P - 450
MFOs (mixed function oxidases)
Microsomal enzymes are concerned with Phase 1 reactions - oxidation, reduction, hydrolysis and
glucuronyl conjugation of Phase 2

14. Goodman & Gilman's the pharmacological basis of therapeutics

15. Phase 1 or Non synthetic reactions
Oxidation:
Addition of oxygen (negatively charged radical) or removal of hydrogen (positively charged
radical)
Reactions are carried out by a group of monooxygenases in the liver
Important oxidation reactions are hydroxylation, oxygenation at C, N or S atoms, oxidative
deamination etc.,
Microsomal oxidations - those catalysed by microsomal enzymes (CYP dependent)

16. Multiple CYP gene families have been identified in humans and categorized based on protein sequence
homology
The cytochrome P-450 isoenzymes are identified and grouped into more than 20.
Families - 1,2,3…. (CYP1)
Most of the drug metabolising enzymes are in CYP 1,2 & 3 families.
Subfamilies are identified as A, B & C… (CYP1A)
An additional numeral when isoform is identified (CYP1A2)
Italics indicate gene (CYP1A2); regular font is enzyme(CYP1A2)
Isoenzyme families important in humans are CYP1, CYP2, CYP3
Cytochrome P family

17. Cytochrome enzymes
Goodman & Gilman's the pharmacological basis of therapeutics

18. Carryout biotransformation of largest number (30- 50%) of drugs. In addition to liver,
these isoforms are expressed in intestine (responsible for first pass metabolism at this site)
and kidney as well.
Example:
Inhibition of CYP3A4 by erythromycin, ketoconazole etc.,
Rifampicin, phenytoin, carbamazepine are inducers of CYP3A4
CYP3A4/5

19. Cytochromes : Metabolism of drugs
Tripathi, K. D. (2018). Essentials of medical pharmacology (8th edition)

20. Tripathi, K. D. (2018). Essentials of medical pharmacology (8th edition)

21. Monoamine oxidase (MAO), Diamine oxidase (DAO)
MAO (mitochondrial) oxidatively deaminates endogenous substrates including
neurotransmitters.
Dopamine, Serotonin, norepinephrine, epinephrine
Alcohol & Aldehyde dehydrogenase
Non-specific enzymes found in soluble fraction of liver
Ethanol metabolism
Flavin monooxygenases
These enzymes are involved in metabolism of drugs like ranitidine
Non CYP drug oxidations:

22. This reaction is the converse of oxidation and involves cytochrome P-450 enzymes working
in the opposite direction.
Alcohols, aldehydes, quinones are reduced. Drugs primarily reduced are chloralhydrate,
chloramphenicol, halothane, warfarin
Reduction process catalysed by microsomal enzymes
Reduction

23. Site: Hydrolysis occurs in liver, intestines, plasma and other tissues.
Microsomal hydrolysis:
Hydrolytic reactions are rarely microsomal, except for
Example:
1. Pethidine to Pethidinie acid (meperidinie)
2. Lidocaine hydrolysis
Non -microsomal hydrolysis:
Enzymes involved are esterases, amidases.
Example:
1.Procaine to PABA by plasma choline esterases
2.Atropine to tropic acid
3.Hydrolysis of penicillin-G & procainamide by amidases
Hydrolysis

24. Cyclization
Formation of a ring structure from a straight chain compound
Ex: Proguanil
Decyclization
Opening up of ring structure of the cyclic drug molecule
Ex: Barbiturates, Phenytoin
Cyclization and Decyclization

25. Conjugation of the drug or its phase I metabolite with an endogenous substrate
To form a polar, highly ionized organic acid, which is easily excreted in urine or bile
Conjugation reactions have high energy requirement and are generally faster than phase 1
reactions
Phase 2 reactions/ Synthetic [conjugation]

26. • Carried out by a group of UDP-glucuronosyl transferases (UGTs)
• Compounds with a phenolic, alcoholic, hydroxyl or carboxylic acid group are easily
conjugated with glucuronic acid
Example:
• Chloramphenicol, aspirin, paracetamol, diazepam, lorazepam, morphine
• Other than drugs, endogenous substrates like bilirubin, steroidal hormones and thyroxine
utilize this and gets excreted in the bile
Drug glucuronides excreted in bile can be hydrolysed by bacteria in the gut- the liberated drug
is reabsorbed and undergoes the same fate. This enterohepatic cycling of the drug prolongs its
action.
Example: oral contraceptives.
Glucuronide conjugation: [Microsomal conjugation]

27. Non –microsomal conjugation
N-Acetyl conjugation
Site: Cytoplasm
Catalysed by N-Acetyl transferase using acetyl coenzyme-A
Important in sulfonamide metabolism because acetyl-sulfonamides are less soluble than
the parent compound and may cause renal toxicity due to precipitation in kidney.
Example : Isoniazid, Dapsone, Sulfonamides, Histamine

28. Sulfate conjugation
Site: Cytoplasm
Major pathway for phenols, also occurs for alcohol, amines
Catalysed by Sulfotransferases using PAPS (3-phosphoadenosine-5-phosphosulfate)
Infants and young children have predominating O-sulfate conjugation
Example : Aspirin, Paracetamol, Corticosteroids, Methyldopa, Chloramphenicol

29. Non–microsomal conjugation : Minor metabolic pathways
Amino acid conjugation:
Site: mitochondria
• ATP dependent acid: CoA ligase forms CoA-amino acid conjugates which then react with
drugs by N-Acetylation
• Drugs like Nicotinic acid, Aspirin, Benzoic acid, Glycine, Arginine
Methyl conjugation:
Site: cytoplasm
• Drugs like Dopamine, Epinephrine, Histamine are catalysed by enzyme transmethylase
with S-adenosine methionine

30. Minor metabolic pathways - contd..
Glutathione conjugation:
Glutathione is a protective factor for removal of potentially toxic compounds
Site: cytoplasm
Drugs like ethacrynic acid, sulfobromophthalein, Paracetamol are catalysed by enzyme g-
glutamyl transpeptidase - the products can be further acetylated to N-acetylcysteine conjugate.
Example: Paracetamol
Ribosides and Riboside Phosphates:
Purines & pyrimidines, used as antimetabolites in chemotherapy, form their active metabolites
by forming ribonucleosides and ribonucleotides.

31. Phase II reactions
• Some drugs are directly metabolised by phase II reactions, before phase I
• Same drugs can be metabolised by different enzymes simultaneously
Ex: Amitriptyline metabolised by CYP2D6, CYP2C9, CYP2C19
• Same CYP can metabolise different drugs simultaneously
Ex: CYP3A4 can bind metabolize diazepam and testosterone simultaneously
Isoniazid
Hydrolysis
(phase I)
N-acetyl
conjugate
(phase II)

32. Spontaneously metabolised in plasma through molecular rearrangement
No enzyme action involved
Example: Atracurium (skeletal muscle relaxant)
Non-enzymatic Biotransformation
HOFMANN ELIMINATION

33. First Pass Metabolism/ Pre - systemic elimination/ First pass effect
Metabolism of a drug during its passage from the site of absorption into the systemic
circulation. Occurs for drugs that are taken orally.
• Significant amount of drug is inactivated before reaching systemic circulation
• Certain amount of drug is absorbed as it passes through GIT wall and portal circulation.
• Decreased bioavailability of the drug
• Diminished therapeutic effect
• Bypassed by parenteral administration of drug - intravenous and sublingual.

34. Attributes of drugs with high first pass metabolism
• Oral dose is considerably higher than sublingual or parenteral dose
• Marked individual variation in the oral dose due to differences in the extent of first pass
metabolism
• Oral bioavailability of a drug is increased if another drug is competing with it
• Oral bioavailability is apparently increased in patients with severe liver disease
Example: Propranolol
Principles of Pharmacology by H. L. Sharma and K. K. Sharma

35. Microsomal enzyme induction
• A number of drugs, many carcinogenic chemicals and insecticides on repeated
administration increase the synthesis of microsomal enzyme protein i.e increase
the rate of metabolism.
• As this requires synthesis of proteins, induction takes 4 - 14 days to reach its peak
and equal time to dissipate after the enzyme inducer is stopped.

36. Different inducers are selective for certain forms of cytochrome P-450 enzyme
CYP3A is induced by - Antiepileptic drugs : Phenobarbitone, Rifampicin and
Glucocorticoids.
CYP2E1 is induced by - Isoniazid, Chronic alcohol consumption
CYP1A is induced by - Cigarette smoking, Industrial pollutants,
Charcoal broiled meat

37. ENZYME INDUCTION
Principles of Pharmacology by H. L. Sharma and K. K. Sharma

38. Clinical relevance of enzyme induction
Decreased intensity of therapeutic effect of the drug.
Example: Failure of Oral contraceptive pills, Warfarin if enzyme inducers like
phenytoin or rifampicin are used concomitantly. Smoking (polycyclic aromatic
hydrocarbons) and theophylline.
Increased intensity of therapeutic effect of the drug.
Example: Hepatotoxicity from paracetamol in alcoholics

39. Tolerance - autoinduction
Example: Barbiturates, Carbamazepine enhance their own metabolism
leading to tolerance.
Interfere with adjustment of dose of another drug and side effects like
osteomalacia.
Therapeutic benefits:
The knowledge of enzyme induction can also be used for therapeutic benefits
Example: To treat neonatal jaundice - Phenobarbitone

40. Inhibition of metabolism
One drug may competitively inhibit the metabolism of another drug - if it utilizes
same enzyme
Unlike induction it is a rapid process and usually reversible
A drug may inhibit one isoenzyme while being itself a substrate for another
isoenzyme.
Example: Quinidine - metabolised by CYP3A4 but inhibits CYP2D6
Inhibition of drug metabolism occurs in a dose related manner and can lead to
toxicity of the object drug
Blood flow limited metabolism
Ex: Propranolol reduces rate of lignocaine metabolism by decreasing hepatic blood

41. ENZYME INHIBITION
Example:
● Unexpected nausea/ vomiting,
tremors in Theophylline with
Erythromycin.
● QT prolongation-Torsades de
pointes in Terfenadine with
Ketoconazole
● L-dopa and Carbidopa - prevents
peripheral decarboxylation
● Alcohol - Disulfiram (aldehyde
dehydrogenase inhibitor) -
aversion of alcohol
Rang and Dale's pharmacology (Ninth edition)

42. Factors affecting biotransformation
Biological factors : age, race, diet
(neonates-glucuronyl transferase-grey baby syndrome-chloramphenicol)
Concurrent use of drugs: Enzyme Induction & Inhibition
Pathological status : Hepatitis, Hypothyroidism
Pollutant exposure from environment or industry : pesticides, charcoal broiled meat
Genetic polymorphism : Succinyl choline (Atypical pseudocholinesterase) - prolonged
apnea due to retartded metabolism

43. Role of metabolism in drug discovery
Two key elements in new drug development, efficacy and safety depend on drug
biotransformation.
Computer based (in silico) computation - Metabolomics
In vitro - Human liver S9 fractions
In vitro studies can give information about
● Metabolite stability
● Metabolite profile
● Metabolite identification
● CYP induction/ inhibition
● Drug-Drug interaction studies
● CYP isoform identification

45. Recap
Biotransformation aims to convert water insoluble drugs/ substances to water soluble
form and is excreted through kidney/ bile
Phase I reactions - addition of functional groups changes the biological property of the
xenobiotic
CYP 450 is the principal enzyme of phase I reactions
Phase II reactions - phase I metabolite is conjugated to increase the water solubility
Biotransformation can determine the efficacy and toxicity of a drug by controlling its
biological t1/2

46. Recap
Different factors influence the xenobiotic metabolism
CYP inducers and inhibitors are the most important cause for drug-drug interactions
Prediction of metabolism and ADRs by knowledge of biotransformation with modern In
vitro and In silico methods are emerging as the most important in the new drug
development process.

47. Reference
Goodman, Louis S., Alfred Gilman, Laurence L. Brunton, John S. Lazlo, and Keith L.
Parker. 2017. Goodman & Gilman's the pharmacological basis of therapeutics. New York:
McGraw-Hill.
Ritter, J. (2020). Rang and Dale's pharmacology (Ninth edition) Edinburgh: Elsevier.
Tripathi, K. D. (2018). Essentials of medical pharmacology (8th edition). Jaypee Brothers
Medical.
Bertram G. Katzung , Todd W. Vanderah ,Basic & Clinical Pharmacology 15th Edition,
McGraw-Hill Lange.
Principles of Pharmacology by H. L. Sharma and K. K. Sharma; Paras Medical Publishers,
New Delhi.2018.
Illustrated reviews : Pharmacology by Karen Whalen, Lippincott, Wolters Kluwer

48. Thank
you