Biotransformasi

Metabolisme Xenobiotik

Xenobiotic metabolism

PRINSIP DASAR XENOBIOTIK DIBAGI :

1.Golongan OBAT-OBATAN:
anti biotik, anti piretik/analgetik, suplemen, obat
jantung, dll

2. KARSINOGEN:
Zat pewarna makanan, penyedap,
nitrosamin, alkohol, pemanis (sakarin), dll

3. BAHAN KIMIA/ POLUTAN LINGKUNGAN :

Jenis pencemaran ini ada > dari 200.000 jenis

Contoh bahan kimia yang terkandung di dalam rokok

OBAT

DARI MANA
MEREKA DATANG?

KANKER

BAHAN KIMIA

Metabolisme Xenobiotik
Metabolisme Xenobiotik adalah satu set jalur
metabolisme yang memodifikasi struktur kimia dari
senyawa asing ( obat2-an & racun) menjadi senyawa
normal
Jalur ini dikenal juga dengan nama bio transformasi
(biotransformation)
Proses reaksi ini sering menyebabkan detoksifikasi
senyawa racun, tetapi beberapa kasus hasil senyawa
antara (the intermediates in xenobiotic metabolism) dapat
menyebabkan efek toksik.

Absorpsi senyawa xenobiotik

Xenobiotik

Darah

Xenobiotik

Absorpsi
xenobiotik

Xenobiotik
dalam darah

Metabolit

Urina

Distribusi
jaringan lain

Jaringan

Ekskresi
lain (cairan)

Nasib senyawa xenobiotik

Racun/toksik
Metabolit racun /
toksik

Xenobiotik

Metabolit aktif

Aktivitas
berubah

Metabolit inaktif

Aktivitas
meningkat

Metabolik yang
reversibel

Kehilangan
aktivitas
Aktivitas
memanjang

 Xenobiotic metabolism is divided into three phases.

In phase I, enzymes such as cytochrome P450
oxidases introduce reactive or polar groups into
xenobiotics.
These modified compounds are then conjugated to
polar compounds in phase II reactions. These
reactions are catalysed by transferase enzymes such
as glutathione S-transferases.
Finally, in phase III, the conjugated xenobiotics may
be further processed, before being recognised by
efflux transporters and pumped out of cells.

Phase I reactions

Table 6.1

Oxidation
Hydroxylation (addition of -OH group)
N- and O- Dealkylation (removal of -CH side chains)
Deamination (removal of -NH side chains)
Epoxidation (formation of epoxides)
C
Oxygen addition (sulfoxidation, N-oxidation)
Hydrogen removal

epoxide
C

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

See also Chapter 6 of Casarett and Doulls Toxicology

Phases of detoxification
The metabolism of xenobiotics is often
divided into three phases:
modification, conjugation, and
excretion. These reactions act in
concert to detoxify xenobiotics and
remove them from cells.

Sitokrom P450 mitokondria

flavoprotein NADPH-,
adrenodoxin reduktase,
zat besi nonheme-sulfur protein,
adrenodoxin

Contoh lain senyawa yang berfungsi induksibel:

Obat-obatan : barbital, etanol, steroid dan nikotin
Kimia industri :alkohol, khlordan, DDT dan khloroform
Hidrokarbon poliaromatik (polyaromatic hydrocarbon)
:benzopyrenes,dibenzanthrazena

Phase II - conjugation Covalent

"conjugation" to endogenous substances:
In subsequent phase II reactions, these activated xenobiotic
metabolites are conjugated with charged species such as glutathione
(GSH), sulfate, glycine, or glucuronic acid. These reactions are
catalysed by a large group of broad-specificity transferases, which in
combination can metabolise almost any hydrophobic compound that
contains nucleophilic or electrophilic groups.[1] One of the most
important of these groups are the glutathione S-transferases (GSTs).
The addition of large anionic groups (such as GSH) detoxifies
reactive electrophiles and produces more polar metabolites that cannot
diffuse across membranes, and may, therefore, be actively transported.

Biotransformation
Potentially toxic xenobiotic
Detoxification
Inactive metabolite

Relatively harmless
Metabolic
activation
Reactive intermediate

Converting lipophilic to water

soluble compounds
Lipophilic
Xenobiotic

(non-polar)
Phase I - Activation

Reactive intermediate
Phase II - Conjugation
Conjugate

Excretion

Water soluble
(polar)

Biotransformation
Water soluble xenobiotics are easier to
eliminate ( t1/2)
Urine, feces but not exhalation
If within barrier, no out

Multiple enzymes (families)

Constitutively expressed
Inducible
Broad specificity
Polymorphic (allelic variants)
Stereo-isomer specificity: 6-OH in hormones:
CYP2A1
6-OH
CYP3A
6-OH

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
Dioxin/PCBs
Barbiturates

CYP2E1
CYP1A
CYP2B

CYP genes have multiple alleles (2D6 has 53, and 2E1 has
13)

Metabolic enzymes
1.

Microsomal:
1.
2.

2.

Non-microsomal
1.
2.
3.

3.

CYP450 monooxygenases
Flavin monooxygenase

Alcohol dehydrogenase
Aldehyde dehydrogenase
Monoamine and diamine oxidases

Both
1.
2.
3.

Esterases and Amidases

Prostaglandin synthase
Peroxidases

Glucuronidation of phenol

Sulfation of phenol and toluene

Glutathione
-glutamyl-cysteinyl-glycine

Active site of a GST:

GSH conjugation of
acetaminophen

Cooxidation of
acetaminophen
by prostaglandin
endoperoxide
synthetase

Compare to fig. 6-2

Benzene trasformation to
leukemia-causing metabolite

Phase III - further modification

and excretion
After phase II reactions, the xenobiotic conjugates may be further metabolised. A
common example is the processing of glutathione conjugates to acetylcysteine
(mercapturic acid) conjugates.[7] Here, the -glutamate and glycine residues in the
glutathione molecule are removed by Gamma-glutamyl transpeptidase and dipeptidases.
In the final step, the cystine residue in the conjugate is acetylated.
Conjugates and their metabolites can be excreted from cells in phase III of their
metabolism, with the anionic groups acting as affinity tags for a variety of membrane
transporters of the multidrug resistance protein (MRP) family.[8] These proteins are
members of the family of ATP-binding cassette transporters and can catalyse the ATPdependent transport of a huge variety of hydrophobic anions,[9] and thus act to remove
phase II products to the extracellular medium, where they may be further metabolised
or excreted.[10]

Toxication and Detoxication: Must consider both

parent and metabolites:
Metabolism may either decrease or increase toxicity.Toxicants can be
formed in a variety of ways:
A) Parent compound toxic. Metabolites non-toxic.
B) Parent compound non-toxic. Metabolites toxic.

Toxic metabolites formed in situ

Toxic metabolites formed in liver and transported

e.g., Parathion (nerve poison):

ABSORPTION/ELIMINATION:
CHARACTERISTICS OF XENOBIOTICS?
Lipophilic
Penetrate membranes by diffusion
Transported by lipoproteins in blood
HOW ELIMINATE?
Diffusion (limitations?)
Excrete (limitations?)
MAKE IT WATER SOLUBLE!!!!!!

The reactions in these pathways are of particular interest in

medicine as part of drug metabolism and as a factor contributing to
multidrug resistance in infectious diseases and cancer
chemotherapy. The actions of some drugs as substrates or
inhibitors of enzymes involved in xenobiotic metabolism are a
common reason for hazardous drug interactions. These pathways
are also important in environmental science, with the xenobiotic
metabolism of microorganisms determining whether a pollutant
will be broken down during bioremediation, or persist in the
environment. The enzymes of xenobiotic metabolism, particularly
the glutathione S-transferases are also important in agriculture,
since they may produce resistance to pesticides and herbicides.

Permeability barriers and

detoxification
That the exact compounds an organism is exposed to will be largely
unpredictable, and may differ widely over time, is a major
characteristic of xenobiotic toxic stress.[1] The major challenge faced
by xenobiotic detoxification systems is that they must be able to
remove the almost-limitless number of xenobiotic compounds from the
complex mixture of chemicals involved in normal metabolism. The
solution that has evolved to address this problem is an elegant
combination of physical barriers and low-specificity enzymatic
systems.

 All organisms use cell membranes as hydrophobic permeability

barriers to control access to their internal environment. Polar
compounds cannot diffuse across these cell membranes, and the uptake
of useful molecules is mediated through transport proteins that
specifically select substrates from the extracellular mixture. This
selective uptake means that most hydrophilic molecules cannot enter
cells, since they are not recognised by any specific transporters.[2] In
contrast, the diffusion of hydrophobic compounds across these barriers
cannot be controlled, and organisms, therefore, cannot exclude lipidsoluble xenobiotics using membrane barriers.

However, the existence of a permeability barrier means that organisms were able to
evolve detoxification systems that exploit the hydrophobicity common to membranepermeable xenobiotics. These systems therefore solve the specificity problem by
possessing such broad substrate specificities that they metabolise almost any non-polar
compound.[1] Useful metabolites are excluded since they are polar, and in general
contain one or more charged groups.
The detoxification of the reactive by-products of normal metabolism cannot be
achieved by the systems outlined above, because these species are derived from normal
cellular constituents and usually share their polar characteristics. However, since these
compounds are few in number, specific enzymes can recognize and remove them.
Examples of these specific detoxification systems are the glyoxalase system, which
removes the reactive aldehyde methylglyoxal,[3] and the various antioxidant systems
that eliminate reactive oxygen species.[4]

Endogenous toxins
The detoxification of endogenous reactive metabolites such as peroxides and reactive
aldehydes often cannot be achieved by the system described above. This is the result
of these species' being derived from normal cellular constituents and usually sharing
their polar characteristics. However, since these compounds are few in number, it is
possible for enzymatic systems to utilize specific molecular recognition to recognize
and remove them. The similarity of these molecules to useful metabolites therefore
means that different detoxification enzymes are usually required for the metabolism
of each group of endogenous toxins. Examples of these specific detoxification
systems are the glyoxalase system, which acts to dispose of the reactive aldehyde
methylglyoxal, and the various antioxidant systems that remove reactive oxygen
species.

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

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

Phase I reactions

Table 6.1

Oxidation
Hydroxylation (addition of -OH group)
N- and O- Dealkylation (removal of -CH side chains)
Deamination (removal of -NH side chains)
Epoxidation (formation of epoxides)
C
Oxygen addition (sulfoxidation, N-oxidation)
Hydrogen removal

epoxide
C

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

See also Chapter 6 of Casarett and Doulls Toxicology

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, SOD, GPX, CAT in coupled reactions

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
See also p. 40-44 of Casarett and Doulls Toxicology

The CYP-450 reaction cycle

A
G

(B)
C

F
E

Oxidation of vinyl chloride to an

epoxide

Hydrolysis of esters and amides

Hydrolysis of organophosphates

Hydrolysis of epoxides

Stereoselective
hydroxylation

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

Azo- and nitro- reduction

Intestinal flora as part of biotransformation

Ready for elimination

Flora action

Oxidation reactions

Flavin mono-oxygenases
(FMO) catalyzed reactions

Nitrogen compounds

Phase II reactions

Glycoside conjugation - glucuronidation

Sulfate - sulfation
Glutathione (GSH)
Methylation
Acylation
Acetylation
Amino acid conjugation
Deacetylation

Phosphate conjugation