1. Biotransformation Xenobiotic metabolism
“Essentials of Toxicology”

2. Biotransformation
Biotransformation means chemical alteration of chemicals such as nutrients, amino acids, toxins, xenobiotics or drugs in the body. It is also needed to render nonpolar compounds polar so that they are not reabsorbed in renal tubules and are excreted.
Biotransformation may results into-
Active Inactive form
Active Active or toxic form
Inactive Active form
Unexcretable Excretable form

3. Biotransformation Results of biotransformation more potent active
TOXIC
less potent
active
active
Drug
or Poison
biotransformed Drug or Poison
inactive
inactive
In general -
nonsynthetic precede synthetic reactions
nonsynthetic reactions can produce active metabolites
synthetic reactions produce inactive metabolites

4. Biotransformation

5. Biotransformation Why is Biotransformation necessary?
• Most drugs are excreted by the kidneys
• For renal excretion drugs should:
– have small molecular mass
– be polar in nature
– should be fully ionized at body pH
• Most drugs are complex and do not have these properties and thus have to be broken down to simpler products.

6. Biotransformation Pharmacologically active organic molecules tend to-
Ctd…
Pharmacologically active organic molecules tend to-
Be highly lipophilic & remain unionized or partially ionized at physiologic PH.
• Thus readily pass across biological barriers –membranes
• Strongly bound to plasma proteins
Such substances are not readily filtered at the glomerulus.
Their lipophilicity also facilitates to be reabsorbed through lipophilic renal tubular membranes.
• This property also stops them from getting eliminated
• They have to be converted to simpler hydrophilic compounds so that they are eliminated and their action is terminated.

7. Biotransformation
Ctd…
• Biotransformation can also result in bioactivation, which involves the production of reactive metabolites that are
more toxic, mutagenic, or carcinogenic than their parent compound(s).
• Drugs may converted to-
– less toxic materials
– more toxic materials
– materials with different type of effect or toxicity
Beside these, biotransformation is called a biochemical defense mechanism as it handles different xenobiotics, drugs, toxicants, body wastes (hemoglobin) or other unwanted substances to those we get exposed.

8. Biotransformation Where do biotransformations occur?
Liver is the principal organ of drug metabolism although every tissue has some ability to metabolize drugs.
Other tissues that display considerable activity include the GIT, the lungs, the skin, and the kidneys.
Following oral administration, many drugs (e.g. isoproterenol,morphine) absorbed intact from the small intestine and transported first via the portal system to the liver, where they undergo extensive metabolism ( first-pass metabolism).
Some orally administered drugs (e.g.clonazepam, chlorpromazine) are extensively metabolized in the intestine than in the liver.

9. Biotransformation Ctd…
Thus intestinal metabolism may contribute to the overall first-pass effect.
First pass effects may so greatly limit the bioavailability of orally administered drugs.
The lower gut harbors intestinal microorganisms that are capable of many biotransformation reactions.
Although drug biotransformation in vivo can occur by spontaneous, noncatalyzed chemical reactions, the vast majority are catalyzed by specific cellular enzymes.At the cellular level, these enzymes may be located in the –
i) Endoplasmic reticulum ii) mitochondria iii) cytosol iii) lysosomes iv) even the nuclear envelope or v) plasma membrane.

10. Biotransformation
Water soluble xenobiotics are easier to eliminate in urine, feces but not exhalation as t1/2 is low.
Lipophilic barbiturates such as thiopental & phenobarbital would have half-lives greater than 100 years if they were not converted to water-soluble compounds.
Multiple enzymes (families)
Constitutively expressed
Inducible
Broad specificity
Polymorphic
Stereo-isomer specificity

11. Biotransformation Relatively harmless Potentially toxic xenobiotic
Metabolic activation
Detoxification
Inactive metabolite
Reactive intermediate

12. Converting lipophilic to water soluble compounds
(non-polar)
Xenobiotic
Phase I - Activation
Reactive intermediate
Phase II - Conjugation
Conjugate
Water soluble
(polar)
Excretion

13. Phase I introduction of functional group
hydrophilicity increases slightly
may inactivate or activate original compound
major player is CYP or mixed function oxygenase (MFO) system in conjunction with NAD(P)H
location of reactions is smooth endoplasmic reticulum

14. Phase II conjugation with endogenous molecules
(GSH, glycine, cystein, glucuronic acid)
hydrophilicity increases substantially
neutralization of active metabolic intermediates
facilitation of elimination
location of reactions is cytoplasm

15. Phase I reactions epoxide Oxidation Reduction Hydrolysis C O
Hydroxylation (addition of -OH group)
N- and O- Dealkylation (removal of -CH side chains)
Deamination (removal of -NH side chains)
Epoxidation (formation of epoxides)
Oxygen addition (sulfoxidation, N-oxidation)
Hydrogen removal
Reduction
Hydrogen addition (unsaturated bonds to saturated)
Donor molecules include GSH, FAD, NAD(P)H
Oxygen removal
Hydrolysis
Splitting of C-N-C (amide) and C-O-C (ester) bonds
epoxide O
C C C O

16. Biotransformation Activation of xenobiotics is a key element
(e.g. benzene, vinyl chloride)
Reactive intermediates include epoxides and free radical species (unpaired electrons) that are short-lived and hence highly reactive
Protection is provided by
endogenous antioxidant substances, e.g. GSH
vitamins C and E
antioxidant enzymes
Antioxidant molecules are oxidized in the process but have the capacity to regenerate the reduced form from the oxidized - NAD(P)H is a key player

17. Cytochrome P450 (CYP) Mixed Function Oxidases (MFO)
Located in many tissues but highly in liver ER
Human: 16 gene families
CYP 1,2,3 perform drug metabolism
>48 genes sequenced
Key forms: CYP1A2, CYP2C9, CYP2C19, CYP2D6, CYP2E1, and CYP3A4
Highly inducible
Alcohol CYP2E1
Barbiturates CYP2B

18. Proportion of drugs metabolized by different CYPs
CYPs are the major enzymes involved in drug metabolism, accounting for ~75% of the total metabolism.Most drugs undergo deactivation by CYPs, either directly or by facilitated excretion from the body. Also, many substances are bioactivated by CYPs to form their active compounds.
Proportion of drugs metabolized by different CYPs

19. Figure CYP450 Reaction SequenceOH
DRUG
DRUG
CYP450
Fe3+
CYP450
Fe3+
CYP450
Fe3+
DRUG
DRUG
NADPH + H+ OH e-
CYP450
reductase
CYP450
Fe3+
CYP450
Fe2+
NADPH + H+
DRUG
DRUG : .. O H+ e-
H2O O2
CYP450
Fe2+
CYP450
Fe2+
DRUG
DRUG
O21- O2

20. Oxidation of vinyl chloride to an epoxide

21. Metabolic enzymes Microsomal: Non-microsomal Both
CYP450 monooxygenases
Flavin monooxygenase
Non-microsomal
Alcohol dehydrogenase
Aldehyde dehydrogenase
Monoamine and diamine oxidases
Both
Esterases and Amidases
Prostaglandin synthase
Peroxidases

22. Cooxidation of acetaminophen
by prostaglandin endoperoxide synthetase

23. Hydrolysis of esters and amides

24. Hydrolysis of organophosphates

28. Hydrolysis of epoxides

29. Stereoselective
hydroxylation

30. Metabolism of benzo(a)pyrene to 9,10 epoxide:
Potent mutagen that binds DNA