BIOTRANSFORMATION

KATHMANDU UNIVERSITY
DEPARTMENT OF PHARMACY

PRESENTED BY: PHR. BIBEK SINGH MAHAT,

ROLL NO. 07, M. PHARM. (INDUSTRIAL),
MAY 2010
 INTRODUCTION
| Biotransformation
is the process whereby a
substance is changed from one chemical to
another (transformed) by a chemical reaction

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within the body.

| Biotransformation is vital to survival in that it

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transforms absorbed nutrients (food, oxygen,
etc.) into substances required for normal body
f ti
functions.

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 INTRODUCTION
| Biotransformation also serves as an important
defense mechanism by converting toxic xenobiotics
and body wastes into less harmful substances and/or
substances that can be excreted from the body.

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| Lipophilic to
toxicants
icants are hard
h d ffor the
th body
b d to
t
eliminate
| Most lipophilic toxicants can be transformed into

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hydrophilic metabolites
| Hydrophilic
y p chemicals are easier for the bodyy to
eliminate.

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 INTRODUCTION
| When biotransformation results in metabolites of
lower toxicity, the process is known as
detoxification.

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| In many cases, however, the metabolites are more
t i than
toxic th the
th parentt substance.
bt This
Thi is
i known
k as
bio-activation.
| Occasionally,
Occasionally biotransformation can produce an

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unusually reactive metabolite that may interact
with cellular macromolecules (e.g., DNA).

| Thiscan lead to very serious health effects, for

example,
l cancer or birth
i defects.
f 4
 XENOBIOTIC
| A xenobiotic is a chemical which is found in an
organism but which is not normally produced or
expected to be present in it. It can also cover
substances which are present in much higher

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concentrations than are usual.
| The body removes xenobiotics by xenobiotic
metabolism. This consists of the deactivation and the

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secretion of xenobiotics, and happens mostly in the
liver.
| Hepatic enzymes are responsible for the metabolism of
xenobiotics by first activating them and then
conjugating the active secondary metabolite followed by
excretion in bile or urine. E.g.hepatic microsomal 5

cytochrome P450.
 BIO-CHEMICAL REACTIONS
| Bio-Chemical reactions are continually taking
place in the body……………………..
p y
• They are a normal aspect of life,

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• pparticipating
p g in the buildingg upp of new tissue,,
• tearing down of old tissue,
• conversion of food to energy,
gy,

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• disposal of waste materials,
• and elimination of toxic xenobiotics.

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 BIO-CHEMICAL REACTIONS
ƒ Enzymes are the catalysts for nearly all
biochemical reactions in the body.y
ƒ Without these enzymes, essential

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biotransformation reactions would take place
slowly or not at all, causing major health
problems.

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ƒ it is the drug or chemical transforming enzymes
that hold the key to xenobiotic transformation.

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 BIOTRANSFORMING ENZYMES

| Most biotransforming enzymes are high molecular

weight proteins, composed of chains of amino acids
linked together by peptide bonds.

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| When a substrate fits into the enzyme's structure,
an enzyme-substrate complex can be formed.
| This allows the enzyme to react with the substrate

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with
i h the
h result
l that
h two different
diff products
d are
formed.
| If th
the substrate
b t t does
d nott fit into
i t the
th enzyme, no
complex will be formed and thus no reaction can
occur.
occur 8
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BIOTRANSFORMING ENZYMES
 SPECIFICITY OF BIOTRANSFORMING
ENZYMES

| there are three main types of specificity:

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 CATEGORIZATION OF THE
BIOTRANSFORMATION REACTIONS
| They are usually classified as Phase I and Phase II
reactions.
| Phase I reactions are generally reactions which modify
the chemical by adding a functional structure to "fit"

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into the Phase II enzyme for conjugation (joined
together) with another substance.
| Phase II reactions consist of those enzymatic reactions

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that
h conjugate
j the modified
ifi xenobiotic
i i with
i another
substance.
| The conjugated products are larger molecules than the
substrate and generally polar in nature (water-soluble).
Thus, they can be readily excreted from the body. They
also
l have
h poor ability
bilit to
t cross cell
ll membranes.
b 11
 CATEGORIZATION OF THE
BIOTRANSFORMATION REACTIONS
| In some cases, the xenobiotic already has a functional
group that can be conjugated and can be biotransformed
b a Phase
by Ph II reaction
ti without
ith t going
i through
th h a Phase
Ph I
reaction. Examples:-

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| Phenol can be directly conjugated ;

| Benzene requires both Phase I and Phase II reactions.

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Phase I – Reactions Nonsynthetic Reactions
•hydroxylations
aromatic, aliphatic, nitrogen
OXIDATION •dealkylations(N-, S-, P)
deaminations
•deaminations
•N-, S-, P- oxidations oxidoreductases
•S-replacements oxidases
•epoxidations monoamine oxidases
mixed function oxidases
•others
th

•azo reduction oxidoreductases

REDUCTION •nitro reduction
reductases
•disulfide reduction
•others

esterases
•esters
amidases
HYDROLYSIS •amides
peptidases
•hemiacetals,acetal
s, h
hemiketals,
ik t l ketals
k t l lipases
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hydrolases
 OXIDATION
| Oxidation is a chemical reaction in which
a substrate loses electrons.

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EXAMPLES OF OXIDIZING REACTIONS.

¾ Alcohol dehydrogenation
¾ Ald h d d
Aldehyde dehydrogenation
h d ti
¾ Alkyl/acyclic hydroxylation

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¾ Aromatic hydroxylation
¾ Deamination
¾ Desulfuration

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¾ N-hydroxylation
¾ N-oxidation
¾ Sulphoxidation

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 REDUCTION
| Reduction is a chemical reaction in which the substrate
gains electrons.
| Reductions
R d ti are mostt likely
lik l to
t occur with
ith xenobiotics
bi ti ini
which oxygen content is low.

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| Reductions can occur across nitrogen
nitrogen-nitrogen
nitrogen double
bonds (azo reduction) or on nitro groups (NO2).

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EXAMPLES OF REDUCING REACTIONS.

¾ Azo reduction
¾ Dehalogenation
Disulfide reduction

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¾

¾ Nit o reduction
Nitro ed ction
¾ N-oxide reduction
¾ Sulfoxide reduction

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 HYDROLYSIS
| Hydrolysisis a chemical reaction in which the
addition of water splits the toxicant into two
fragments or smaller molecules.

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| Thehydroxyl group (OH-) is incorporated into
one fragment and the hydrogen atom is

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i
incorporated
t d into
i t the
th other.
th

| Larger
L chemicals
h i l suchh as esters,
t amines,
i
hydrazines, and carbamates are generally
biotransformed by hydrolysis.
hydrolysis 18
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HYDROLYSIS
 SUMMARY OF
PHASE -I BIOTRANSFORMATION

| Toxicants are converted to metabolites that

are sufficiently
ffi i l ionized,
i i d or hydrophilic,
h d hili to be
b
either eliminated from the body without further

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biotransformation or converted to an
intermediate metabolite that is ready for Phase
II biotransformation.

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| The intermediates from Phase I transformations
mayy be p
pharmacologically
g y more effective and
in many cases more toxic than the parent
xenobiotic.
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 SUMMARY OF
PHASE -I BIOTRANSFORMATION

| Theprincipal reaction of drug/toxin metabolism

is OXIDATION.

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| The enzymes responsible are oxido-reductases;
called mixed-function oxidases.

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| Mostprominent and important among these is
the
h cytochrome
h P450 system consists
i off Cyt
C P
450 and Cyt P 450 reductase
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STRUCTURE OF CYTOCHROME P450 SYSTEM

cytochrome P 450

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 CYP450 Reaction Sequence

OH

DRUG DRUG
CYP450
Fe3+

CYP450 CYP450
incorporation of
Fe3+ Fe3+ oxygen
DRUG DRUG peroxide
NADPH + H+ dioxygen
etc.
OH e-
CYP450
reductase
CYP450
Fe3+ CYP450
NADPH + H+ Fe2+
2
DRUG
DRUG
O H+
e-
H 2O
CYP450 CYP450 O2
Fe2+ Fe2+
DRUG DRUG

O21- O2
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IMPORTANCE & QUANTITY OF CYP450 SYSTEM

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 bibekmahatsingh@gm
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P450 3A
CYTOCHROME
PHASE -II BIOTRANSFORMATION

| A xenobiotic that has undergone a Phase I reaction is

now a new intermediate metabolite that contains a
reactive chemical group, e.g., hydroxyl (-OH), amino

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((-NH2),
NH2), and carboxyl ((-COOH).
COOH).

| Manyy of these intermediate metabolites do not

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possess sufficient hydrophilicity to permit
elimination from the body.

| These metabolites must undergo additional

biotransformation as a Phase II reaction. 26
 PHASE -II BIOTRANSFORMATION

| The primary Phase II reactions are:

Most Important :

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o Glucuronide conjugation

o Sulfate conjugation

Others :

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o Acetylation

o Amino acid conjugation

j g
o Glutathione conjugation

o Methylation
y
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 GLUCURONIDE CONJUGATION

| One of the most important and common Phase II

reactions.

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| Molecules added directly to the toxicant or its
phase I metabolite is glucuronic acid, a
molecule derived from glucose, a common
carbohydrate (sugar) that is the primary source

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of energy for cells.
| The sites of glucuronidation reactions are
substrates having an oxygen, nitrogen, or sulfur
bond.
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 GLUCURONIDE CONJUGATION

| This includes a wide array of xenobiotics as well as

endogenous substances, such as bilirubin, steroid
hormones and thyroid hormones.

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| It is a high-capacity
high capacity pathway for xenobiotic
conjugation.
| It usually decreases toxicity, although there are

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some notable exceptions, for example, the
production of carcinogenic substances.
| They are generally quite hydrophilic and are
excreted by the kidney or bile.
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 GLUCURONIDE CONJUGATION

| The glucuronide conjugation of aniline is

ill
illustrated
d below:-
b l

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 GLUCURONIDE CONJUGATION

Glucuronide formation H CO2H

OH
gluconic acid HO
OH HO
HO OPO32-
H OH
OH COOH H H
HO

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OH UTP
OH
O OH
PPi
OH COOH
HO H CO2H
O OH UDP
HO
OH
glucose OH
HO
HO O

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H OH
HO
H H
glucuronic acid OH

UDP
O
OH O
H CO2H UDP H CO2H OH
COH HO HO C
+
HO HO
HO O HO O
H OH H OH
O
salicylic acid H H H H 31
a glucuronide
UDP-glucuronide
derivative
 SULFATE CONJUGATION

| Sulfate conjugation is another important Phase

II reaction that occurs with many xenobiotics
xenobiotics.
| In general, sulfation decreases the toxicity of

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xenobiotics.
xenobiotics
| The highly polar sulfate conjugates are readily
secreted in the urine.
urine

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| In general, sulfation is a low-capacity pathway
for xenobiotic conjugation.
| Often glucuronidation or sulfation can conjugate
the same xenobiotics.
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 SULFATE CONJUGATION
sulfate ester formation

NH2

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HO + N N O

N N O SO
O O
Sulfates are carried as O
O S O PO
phosphoadenosine- O
phosphosulfate derivatives O y
The enzymes catalyzing
y g

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O
(PAPS) - a high energy form. this type of reaction are
HO O called sulfotransferases.
O P O

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 BIOTRANSFORMATION SITES

| Liver is the primary biotransforming organ due

to its large size and high concentration of
biotransforming enzymes

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| Within the liver cell, the primary subcellular
components that contain the transforming

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enzymes are the microsomes (small vesicles) of
the endoplasmic
p reticulum and the soluble
fraction of the cytoplasm (cytosol).

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BIOTRANSFORMATION SITES

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Liver SER

smooth
endoplasmic
reticulum

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 MODIFIERS OF BIOTRANSFORMATION

| Age
| Genetic variability in biotransforming capability

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 MODIFIERS OF BIOTRANSFORMATION

| Poor nutrition
| Enzyme inhibition and enzyme induction
can be caused by prior or simultaneous exposure

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to xenobiotics. In some situations exposure to a
substance will inhibit the biotransformation
capacity for another chemical due to inhibition of
specific
p enzymes
y

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| Examples of enzyme inducers are alcohol,
isoniazid, polycyclic halogenated aromatic
h d
hydrocarbons
b ((e.g., di
dioxin),
i ) phenobarbital,
h b bit l
and cigarette smoke

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 MODIFIERS OF BIOTRANSFORMATION

| Dose level
| At low doses,
doses a xenobiotic may follow a
biotransformation pathway that detoxifies the

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substance.
| However, if the amount of xenobiotic exceeds the
specific enzyme capacity, the biotransformation
pathway is "saturated"
saturated . In that case,
case it is possible that

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the level of parent toxin builds up.
| In other cases, the xenobiotic may enter a different
biotransformation pathway that may result in the
production of a toxic metabolite.
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 REFERENCES
| National Library of Medicine; Emily Monosson ; 2008
"Biotransformation”.
| Encyclopedia of Earth; Eds. Cutler J. Cleveland, Washington,
D.C.: Environmental Information Coalition, National Council

bibekmahatsingh@gm
for Science and the
Environment<http://www.eoearth.org/article/Biotransformation
>Diaz E (editor). (2008). Microbial
| Biodegradation:
g Genomics and Molecular Biology
gy ((1st ed.

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ed.). Caister Academic Press. ISBN 978-1-904455-17-
2.http://www.horizonpress.com/biod.
| www. wikipedia,
p , the free encyclopedia
y p

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THANK - YOU