Alcohol Metabolism & Oxidative Stress Flashcards
What is the highest to the lowest in energy content between
- Protein
- Fats
- Alcohol
- Carbohydrates
-Fats 37 Kj/g
-Alcohol 29 Kj/g
-Protein 17 Kj/g
-Carbohydrates 17 Kj/g
Where is most alcohol Metabolised and what happens to the rest
Most (>90%) alcohol is metabolised by liver
Remainder excreted passively in urine and on breath.
What is alcohol oxidised to and what by
- Alcohol oxidised by alcohol dehydrogenase to acetaldehyde
and then to - Acetate by aldehyde dehydrogenase.
What happens to acetate in our body
Acetate converted to acetyl~CoA and used in TCA cycle or for fatty acid synthesis
Where can small amounts of alcohol be oxidised
Smaller amounts of alcohol can also be oxidized by the cytochrome P450 2E1 enzyme (CYP2E1), (in the liver)
or by catalase (in brain).
What is the recommended limit for alcohol
14 units/week
spread over at least 3 days for BOTH men & women
What is the Rate of alcohol metabolism
Eliminated at rate of ~7g per hour
- One unit of alcohol = 8 g
- Half pint of normal strength beer, small glass of wine
What happens when you have too much Acetaldehyde
Acetaldehyde is a toxic metabolite. Accumulation causes “Hangover”
Can also lead to liver damage
How is Acetaldehyde levels kept down
Acetaldehyde toxicity normally kept to a minimum by aldehyde dehydrogenase
(low Km for acetaldehyde) - thus higher affinity
What does Excess NADH and Acetyl-Coa Lead to
Excess NADH and Acetyl-CoA lead to changes in liver metabolism
This causes
* “Fatty liver”
* Alcoholic hepatitis
* Alcoholic cirrhosis
In Alcohol Oxidation what happens when there is a decrease in NAD+/ NADH ratio
1- inadequate NAD+ for conversion of lactate to pyruvate
Thus lactate accumulates in the blood
Causing Lactic Acidosis
2- inadequate NAD+ for conversion of lactate to pyruvate
Thus lactate accumulates in the blood
Kidney’s ability to excrete uric acid reduced
This is as lactic and Uric acid share the same transporters so one limits to other.
Urate crystals accumulate in tissues producing gout
3- Inadequate NAD+ for glycerol metabolism
Deficit in gluconeogenesis
Hypoglycaemia
In alcohol Oxidation what happens when there is a Increase in Acetyl-CoA
Increased synthesis of fatty acids and ketone bodies
Increased synthesis of Triacylglycerol
Lower lipoprotein synthesis
Fatty Liver
What is used form the treatment of alcohol dependance
The Drug Disulfiram
What does Disulfiram do
- Disulfiram can be used as an adjunct in the treatment of chronic alcohol dependence.
- It is an inhibitor of aldehyde dehydrogenase
- If patient drinks alcohol acetaldehyde will accumulate causing symptoms of a ‘hangover’ thus they will feel sick every time they drink alcohol
What diseases are caused by oxidative stress
Cardiovascular disease
Alzheimer’s disease
Rheumatoid arthritis
Crohn’s disease
COPD
Ischaemia / reperfusion injury
Cancer
Pancreatitis
Parkinson’s disease
Multiple sclerosis
What is a major reasons for diseases caused by Oxidative stress
Cellular damage caused by ROS & RNS is a significant component in a wide range of disease states
What are free radials
- Electrons of atoms, molecules & ions usually associate in pairs. Each pair moves within a defined region of space (an orbital).
- A free radical is an atom or molecule that contains one or more unpaired electrons and is capable of independent (“free”) existence
- A superscript dot used to denote free radical (e.g. OH*)
- Free radicals (usually) very reactive and tend to acquire
electrons from other atoms, molecules or ions - Reaction of a radical with a molecule typically generates a second radical thereby propagating damage.
Reactive Oxidative Species (ROS) formation .
(Molecular oxygen is a biradical. It has 2 unpaired electrons in different orbitals)
(O2) + (e-) = (O2-)
This produces a superoxide
(Produced by adding electron to molecular oxygen. Also Important source of other ROS)
(O2-) + (2H+ , e-) = (H2O2)
This produces Hydrogen Peroxide
(Not a free radical but can react e.g. with Fe2+ to produce free radicals. Readily diffusible.)
(H2O2) + (e-, H+) = ( H2O + OH)
This produces water and Hydroxyl Radical
(OH = Most reactive and damaging free radical. Reacts with anything!)
(OH*) + (e-, H+) = (H2O)
This produces water
Reactive nitrogen species
-Nitric oxide
(NO*)
turns to
-Peroxynitrite
(ONOO-)
O2- + NO –> ONOO-
- Superoxide can react with nitric oxide to produce peroxynitrite
- Peroxynitrite is not itself a free radical, but is a powerful oxidant that can damage cells
What are the two types of damage ROS can do to DNA
Two main types of damage
- ROS reacts with base
Modified base can lead to mispairing and mutation - ROS reacts with sugar (ribose or deoxyribose)
Can cause strand break and mutation on repair
What is the issue with ROS reacting with DNA
- ROS reacts with DNA
- DNA damage
- Failure in repair can lead to mutation
- Can lead to cancer
How can you measure the amount of damage causes by ROS’s
The amount of 8-oxo-dG present in cells
can be used as measurement of oxidative damage
What happens when ROS reacts with protein Sidechain
Modified amino acid (e.g.)
* Carbonyls
* Hydroxylated adducts
* Ring opened species
* Dimers (e.g. di-tyrosine)
* Disulphide bond (Cys)
This changes protein structure
This can either lead to loss or gain in function
This leads to protein Degradation
What happens when the ROS reacts with the protein backbone
Fragmentation occurs
This leads to protein degradation
-What are disulphide bonds
-How are they formed
-What does an issue in the disulphide bond cause , and how does this happen
- Play important role in folding and stability of some proteins (usually secreted proteins or in extracellular domains of membrane proteins)
- Formed between thiol groups of cysteine residues
- Inappropriate disulphide bond formation can occur if ROS takes electrons from cysteines causing misfolding, crosslinking and disruption of function (e.g. enzyme)
How does ROS damage lipids
Initiation
* Free radical (e.g. OH) extracts hydrogen atom from a polyunsaturated fatty acid in membrane lipid
Propagation
* Lipid radical formed which can react with oxygen to form a lipid peroxyl radical
- Chain reaction formed as lipid peroxyl radical extracts hydrogen form nearby fatty acid
Due to this
* Hydrophobic environment of bilayer disrupted and membrane integrity fails
What are the Endogenous (inside cell) Sources of biological oxidants
(7)
1* Electron transport chain
2* Peroxidases
3* Nitric oxide synthases
4* Lipooxygenases
5* NADPH oxidases
6* Xanthine oxidase
7* Monoamine oxidase
What are the Exogenous (outside cell) Sources of Biological oxidants
- Radiation Cosmic rays
UV light X-rays - Pollutants
- Drugs
Primaquine (anti-malarial) - Toxins
Paraquat (herbicide)
How does the election transport chain lead to formation of ROS
Electrons leaking from the ETC can prematurely react with oxygen, resulting in the generation of reactive oxygen species (ROS)
How does Nitric Oxide Synthase lead to cell damage
Nitric oxide already has a free radical
it reacts with an oxygen which has a free radical to form Peroxynitrite
Peroxynitrite is not a free radical but is a powerful oxidant that can damage cells
O2* - + NO* –> ONOO-
How does NADPH oxidases lead to formation of ROS
- NADH and FADH2 supply electrons (e−) from metabolic substrates
- e − pass through ETC and reduce oxygen to form H2O at Complex IV
- Occasionally electrons can accidently escape chain and react with dissolved O2 to form superoxide
How does NADH and FADH2 lead to formation of ROS’s
- NADH and FADH2 supply electrons (e−) from metabolic substrates
- e − pass through ETC and reduce oxygen to form H2O at Complex IV
- Occasionally electrons can accidently escape chain and react with dissolved O2 to form superoxide
What are the three enzymes involved in the formation of NOS
And how do the different enzymes differ the NOS
- iNOS: Inducible nitric oxide synthase. Produces high NO concentrations in phagocytes for direct toxic effect.
- eNOS: Endothelial nitric oxide synthase (Signalling)
- nNOS: Neuronal nitric oxide sytnthase (Signalling)
How is nitric Oxide formed
-Argenine through nitric oxide synthase using one of the enzymes (iNOS, eNOS, nNOS)
- Turning (NADPH) + (O2) into
(NADP+ )(+ H2O)
- Forms Citrulline and Nitric Oxide
What do the different levels of nitric oxide form
HIGH LEVELS
- Toxic effect
NORMAL LEVELS
- It becomes a signalling molecule for
* Vasodilation
* Neurotransmission
* S-Nitrosylation
How does Respiratory bursts help in Antimicrobial defence system
- Phagolysosome engulfs invading bacteria
- Phagolysosome contains NADPH oxidase enzyme
(a membrane bound complex) - This converts O2 in to O2*-
( Turning NADPH to NADP+ ) - O2*- turns in H2O2
- H2O2 reacts with Cl- to form HOCL* (hypochlorite)
- hypochlorite is essentially bleach
- This works by using the enzyme Myeloperoxidase
- iNOS also in Phagolysosome
- This produces Nitric Oxide (NO*)
- (NO) + (O2- ) react to form ONOO- (peroxynitrate)
hypochlorite and peroxynitrate kill bacteria
What is the cellular defence for Superoxide
- What does Superoxide do
- What does Catalase do
Superoxide dismutase (SOD)
* Converts superoxide to H2O2 and oxygen
* Primary defence because superoxide is strong initiator of chain reactions
Catalase
* Converts H2O2 to water and oxygen as (H2O2 can still be dangerous)
* Widespread enzyme. Important in immune cells to protect against oxidative burst
What are the the 3 isoenzymes of Superoxide dismutase (SOD)
3 isoenzymes:
* Cu+-Zn2+ Cytosolic
* Cu+-Zn2+ Extracellular
* Mn2+ Mitochondria
What is the cellular defence of glutathione
-Reduced form of GSH has a central amino acid of Cysteine
-Thiol group of Cys donates e− to ROS.
-The loss of an election causes GSH to react with another GSH forming a Disulphide bridge between to two cysteine groups, forming GSSG
- This required the enzyme Glutathione peroxidase, which requires Selenium.
- GSSG reduced back to GSH by glutathione reductase which catalyses the transfer of electrons from NADPH to disulphide bond
-NADPH from pentose phosphate pathway is therefore essential for protection against free radical damage
What is the rate limiting enzyme of the pentose pathway
Glucose 6-phosphate dehydrogenase
The Pentose phosphate pathway is an important source of NADPH why is NADPH important
- Reducing power for biosynthesis
- Maintainance of GSH levels
- Detoxification reactions
What vitamins a free radical scavengers and what is their importance
Vitamin E
(α-tocopherol)
* Lipid soluble antioxidant
* Important for protection against lipid peroxidatio
Vitamin C
(Ascorbic acid)
* Water soluble antioxidant
* Important role in regenerating reduced form of Vitamin E
Others
* Carotenoids
* Uric Acid
* Flavonoids
* Melatonin
What is a free radical scavenger
Free radical scavengers reduce free radical damage by donating hydrogen atom (and its electron) to free radicals in a nonenzymatic reaction
What are 5 oxidants
O2*-
H2O2
OH
NO
ONOO-
What are 5 defences agains oxidants
SOD
Catalase
GSH
Vit E
Vit C
NADPH
What enzymes lead to Galactosemia
A deficiency in the enzymes
- Galactokinase
- Uridyl Transferase
- UDP Galactose Epimerase
What does a lack of Galactokinase do
-This leads to a build up of Galactose
-This is then turned into Galactitol by Aldose reductase
by oxidising NADPH
-This leads to build up in Osmostic pressure
- Also all NADPH is being used up thus less protection against ROS damage
- This leads to Proteins (crystallin) in lens being denatured
- Causing Cataracts
What does a lack of Uridyl transferase or UDP- Galactose enzyme
- This causes a build up of Galactose - 1P
- UDP-galactose epimerase enzymes turns it to UDP- Galactose
- this turns to UDP-Glucose
this causes Glycogenisis
What happens when there is a G6PDH Deficiency
- Decreased G6PDH activity limits amount of NADPH
- Lower GSH means less protection against damage from oxidative stress
- Oxidative stress causes Infection, Drugs (e.g. anti malarial) , Broad beans
- Lipid peroxidation
- Cell membrane damage
- Lack of deformability leads to mechanical stress
Protein damage - Aggregates of cross-linked haemoglobin (Heinz bodies)
What are Heinz Bodies
- Dark staining within red blood cells resulting from precipitated haemoglobin
- Bind to cell membrane altering rigidity
- Increased mechanical stress when cells squeeze through small capillaries
- Spleen removes bound Heinz bodies resulting in “blister cells”
- Clinical sign of G6PDH deficiency
What does a normal amount of paracetamol turn in to
Glucuronide and Sulphate
What happens when you have too much paractamol
paractamol turns to (NAPQI)
this is an oxidant and causes Oxidative damage to liver cell
Lipid peroxidation,
Damage to proteins
Damage to DNA
How do you treat too much paracetamol
Acetylcysteine treatment
Antidote acetylcysteine works by replenishing glutathione levels
detoxifies the toxic metabolite of paracetamol, NAPQI,