Xenobiotic metabolism Flashcards
What is the definition of biotransformation?
The process where the body attempts to convert lipophilic substances into more polar metabolites that can be excreted quicker
What is the primary result of biotransformation?
1.The parent molecule is transformed into a more polar metabolite, often by the addition of ionizable functional groups
2. Molecular weight and size are often increased
3. The excretion is facilitated, and hence elimination of the compound from the tissue
What are the consequences of metabolism?
- The biological half-life is decreased
- The duration of exposure is reduced
- Accumulation of the compound in the body is avoided
- The duration of the biological activity may be affected.
- Detoxification –> Less toxic/inactivation
- Toxification –> Can sometimes lead to more toxic compounds or another toxic effect
What characterize enzymes involved in metabolism?
Enzymes involved in metabolism are flexible and their substrate specificity is generally broad
What is the location of enzymes in biotransformation?
- Many are found in the smooth endoplasmic reticulum (SER)
- Some in the cytosol
- Few in other organelles e.g. mitochondria
What are the main types of phase 1 reactions?
1) Oxidation
2) Reduction
3) Hydrolysis
Which types of phase 2 reactions are there?
1) Glucuronidation
2) Sulfation
3) Glutathione conjugation
4) Conjugation with amino acids
5) Methylation
6) Acetylation
What is the purpose of phase 1 reactions
Alteration of the original molecule by exposing or introducing a functional group (-OH, -NH2, -SH, -COOH)
What is the purpose of phase 2 reactions
Conjugation of a polar group to the functional group –> Increases hydrophilicity (except for methylation and acetylation) –> facilitates excretion
Which enzymes are usually included in Phase 1 drug metabolism?
- Cytochrome P450s (CYP450 or CYPs)
- Alcohol dehydrogenase (ADH)
- Aldehyd dehydrogenase (ALDH)
- Esterases/amidases (hydrolases)
- Amine oxidase (MAO, FMO)
- Xanthine oxidase (XO) – a pyrine
- Alkylhydrazine oxidase
Where can phase 1 enzymes generally be found?
ER:
- CYPs
- FMO
- ADH/ALDH
- Epoxide hydrolase
- (Esterases/ amidases)
Mitochondria:
- MAO A/B
- ADH/ALDH
Cytosol:
- Esterases/ amidases
- ADH/ALDH
- Epoxide hydrolase
Which microsomal oxidation processes are involved in phase 1 reactions?
- Aromatichydroxylation o Aliphatichydroxylation o Alicyclic
- Heterocyclic
- N-,S- and O-dealkylation o N-oxidation
- N-hydroxylation
- S-oxidation
- Desulfaration
- Deamination
- Dehalogenation
Which non-microsomal oxidation processes are involved in phase 1 reactions?
1) Amine oxidation
2) Alcohol and aldehyde oxidation
3) Peroxidases
Describe the catalytic cycle of the cytochrome P-450 monooxygenase system
- Addition of substrate to enzyme
- Donation of an electron
- Addition of oxygen and rearrangement
- Donation of a second electron and loss of water
What enzymes are involved in amine oxidations and what are their main characteristics?
Monoamine oxidases (MAO):
- Located in mitochondria
- Oxidative deamination of primary, secondary and tertiary amines
Diamine oxidases:
- Located in the liver and other tissue
- Soluble enzyme
- Is mainly involved in the metabolism of endogenous compounds such as the aliphatic diamine putrescine
Alcohol dehydrogenase:
- Is a cytosolic enzyme (soluble fraction)
- Located in the liver and also kidney and lungs
- Coenzyme NAD (sometimes NADP - slower)
Aldehyde dehydrogenase:
- Located in the mitochondria (ALDH2)
Other enzymes:
Molybdenum hydroxylases
Xanthine oxidase
Aldehyde oxidase
Peroxidases
- Are dependent on co-oxidation
What enzymes are involved in reduction
Enzymes who catalyzes reduction:
* Gut bacterial azo- and nitro-reductase
* Cytochrome P-450 NADPH reductase –> works in low conditions of oxygen
* FAD alone (is not an enzyme)
(CYPs are also capable of reduction)
Reactions are dependent on:
- NAD(P)H
Inhibited by:
- O2
What enzymes are involved in hydrolysis
1) Esterases
2) Amidases
3) Epoxide hydrolases
4) Peptidases
(CYPs are also capable of hydrolysis)
Types of esterases
Class A)
A-esterases: Hydrolyze organophosphates (arylesterases, paraoxonase). These are not inhibited by phosphate triesters.
Class B)
B-esterases: Inhibited by organophosphates e.g. paraoxon (include carboxylesterases, cholinesterases, arylamidases)
Class C)
C-esterases: Are acetylesterases. These are not inhibited by paraoxon and the preferred substrates are acetyl esters.
What are the steps and the product of glucuronidation (glucuronic acid conjugation)?
- Conjugation with UDP-Glucuronic acid catalyzed by UDP-glucuronosyl transferase (UGT)
2) Glucuronic acid is afterwards converted into hydroxyl, carboxyl, nitrogen and sulfur
Which enzyme catalyze sulfation
Sulfotransferase (SULT)
- with the cofactor 3’-phosphoadenosyl-5-phosphosulfate (PAPS):
What are the product of sulfation?
Highly water soluble sulfate esters
How can PAPS be depleted
When large amount of foreign compound conjugated with sulfate (e.g. paracetamol) is administered
Which enzyme catalyze glutathione conjugation
Glutathione-S-transferases (GST)
Describe the structure of glutathione
Glutathione is a tripeptide composed of:
1) Glutamat
2) Cysteine
3) Glycine
Why is glutathione significant?
1) It has a reactive SH group
2) It is protected from protease digestion because the bond between the cysteine and glutamate is NOT a peptide bond
3) It is present in relatively high concentrations
How does glutathione detoxify?
It has a nucleophilic thiol group, and it can detoxify substances in one of three ways:
1. Conjugation catalyzed by a GSH transferase
2. Chemical reaction with a reactive metabolite to form a conjugate
3. Donation of a proton or hydrogen atom to reactive metabolites or free radicals, respectively
Acetylation is the major route of biotransformation for which compounds?
Sulfonamides, aromatic amines, hydrazines and hydrazides
Which enzymes catalyse acetylation reactions?
N-acetyltransferases:
- NAT1; is in many tissues incl. red blood cells
- NAT2; mostly in Kupffer cells in the liver but is also active in ER cells, in the spleen gut and lung
What is the purpose of acetylation?
To inactivate the reactive compound.
Prevents further metabolization by CYP to more toxic metabolites
(not like the conjugation reactions where the purpose is to make the compound more hydrophilic)
Which phase 2 reactions can paracetamol undergo?
Paracetamol can undergo:
1) Glucuronidation
2) Sulfation
3) Glutathione conjugation
What is the toxic intermediate of paracetamol
NAPQI
Which CYP enzyme converts paracetamol to it’s intermediate?
CYP2E1 and CYP1A2
How is overdose on paracetamol treated? (antidote)
N-acetylcysteine (NAC) / (Methionin also)
- Increases GSH synthesis leading to conjugation of NAPQI –> protect protein thiols
- Increases sulfation by relieving saturation
Overall result is reduced levels of NAPQI and oxidized protein thiols.
Where are the enzymes responsible for hydrolysis located?
1) Cytosol (cytosolic enzymes)
2) Microsomal (gut enzymes)
3) Blood
What are the substrates for hydrolysis?
Esters, thioesters, hydrazines, carbamates, peptides
Which groups are often reduced in vivo?
Aldehydes, ketones, disulfide, sulfoxide, quinone, N-oxide, alkenes, azo and nitro groups
Where is MAO located and which group of enzymes does it belong to?
MAO is located in the mitochondria, and it belongs to the group of enzymes called flavoproteins (FAD)
Which is the most abundant CYP in the liver?
CYP3A4
What is the main location of CYP and which co-factors are involved?
Location: ER
Co-factor: NADPH
Where are FMO located and which co-factors are involved?
Location: ER
Co-factor: FAD, NAD(P)H
Where are MAO located and which co-factors are involved?
Location: Mitochondira
Co-factor: FAD, NAD(P)H
Where are the esterases / amidases located and which co-factors are involved?
Location: Cytosol, Plasma (ER)
Co-factor: (H2O)
Where are alcohol dehydrogenase (ADH) / aldehyd dehydrogenase (ALDH) located and which co-factors are involved?
Location: Cytosol, ER, mitochondria
Co-factor: FAD, NAD(P)H
Where are epoxide hydrolase located and which co-factors are involved?
Location: Cytosol, ER
Co-factor: (H2O)
Which two enzymes metabolize a majority of large compounds?
CYP3A4 / CYP3A5
(CYP 3A5 are the garbage bins – metabolizes compounds that the others didn’t)
Which CYP enzyme metabolizes small, planar aromatics (amines)?
(e.g. caffeine)
CYP1A1 and CYP1A2
Which CYP enzyme metabolizes acidic and lipophilic medium size substrates?
(e.g. warfarin)
CYP2C9
Which enzymes likes nitrogen – amines and amides medium size?
(e.g. omeprazol)
CYP2C19
Which CYP enzyme is highly active but cannot be induced and metabolize basic medium size substrates?
(e.g. codeine)
CYP2D6
Which enzyme is considered the garbage bin – very open pocket – takes all of the compounds that none of the other takes (broad range) but is not specific?
(e.g. paracetamol)
CYP3A4 and CYP3A5 (mostly CYP3A5)
Which mechanism of inductions are there?
Mechanism of induction are:
1. Increased synthesis of mRNA or precursor rRNA,
2. Increased stability of mRNA or rRNA
3. Decreased heme degradation
4. Decreased apoprotein degradation
5. Increased transport of RNA
6. Effects on DNA-dependent RNA polymerase
Which substrates induces the polycyclic hydrogen carbon and phenobarbital type CYPs respectively?
Planar substrates induces the polycyclic hydrogen carbon type (CYP1A1)
Non-planar substrates induces the phenobarbital type (CYP2B1, CYP2B2 and CYP2B6A)
What is TCDD and how does it work?
TCDD is the most toxic compound (known of): 2,3,7,8-tetrachlorodibenzodioxin.
The mechanism is receptor mediated induction of CYP1A1
Which CYP enzyme is induced by alcohol?
CYP2E1
What is the mechanism of induction of the CYP3A and CYP4A family?
It is receptor mediated induction by a chemical.
Which similarities and differences are there between CYPs and FMO (flavin-containing monooxygenase)?
- Both require NADPH, both are found in sER
- FMO differes from CYP as it is thermolabile and metal free.
Where does glucuronide conjugation take place?
ER
Where does sulfation / sulfate conjugation take place?
Cytosol
Where does gluathione conjugation take place?
Cytosol, mitochondria and nucleus
How does glutathione detoxify?
It has a nucleophilic thiol group, and it can detoxify substances in one of three ways:
1. Conjugation catalyzed by a GSH transferase
2. Chemical reaction with a reactive metabolite to form a conjugate
3. Donation of a proton or hydrogen atom to reactive metabolites or free radicals, respectively
Where are the enzymes involved in xenobiotic metabolism produced?
Microsomes: CYP + UGT (UDP-glucuronosyl transferase)
Cytosol: NAT (N-acetyltransferase) + GST (Glutathione-S-transferase)
Which reactions are catalyzed by CYPs?
Hydroxylation, epoxidation, (N, O, S)-dealkylations
What are required in CYP reactions?
O2, NAD(P)H and NADPH reductase
