I want to assess the level of oxidative stress in blood. Which are the biochemical marker/markers to be targeted?
Hi Brijesh Dhabi!, you can measure:
Malondialdehyde/TBARS
(in plasma/serum/erythrocytes)
Protein carbonylation
(in plasma/serum)
Oxidized LDL
(in plasma/serum)
Uric acid
(in plasma/serum)
Vitamin E (α-tocopherol)
(in plasma/serum)
Vitamin C (ascorbic acid)
(in plasma/serum)
Vitamin A (retinol)
(in plasma/serum)
α-Carotene
(in plasma/serum)
β-Carotene
(in plasma/serum)
Total antioxidant capacity
(in plasma/serum)
Superoxide dismutase
(in plasma/serum/erythrocytes)
Glutathione peroxidase
(in plasma/serum/erythrocytes)
Copper
(in plasma/serum)
Ceruloplasmin (Cp)
(in plasma/serum)
Cp oxidase activity
(in plasma/serum)
Transferrin
(in plasma/serum)
Glutathione reductase
(in erythrocytes)
Catalase
(in erythrocytes)
8-hydroxyguanosine
(in lymphocytes)
You could measure: lipid peroxidation, reduced glutathione, vit A, C, E in plasma and activities of superoxide dismutase (SOD), catalase, glutathione peroxidase (GPx), glutathione reductase (GRx) and glutathione S transferase (GST) in red blood cells - That is what has been done in this paper: http://www.ncbi.nlm.nih.gov/pubmed/17668211
Good luck!
Hi Brijesh Dhabi!, you can measure:
Malondialdehyde/TBARS
(in plasma/serum/erythrocytes)
Protein carbonylation
(in plasma/serum)
Oxidized LDL
(in plasma/serum)
Uric acid
(in plasma/serum)
Vitamin E (α-tocopherol)
(in plasma/serum)
Vitamin C (ascorbic acid)
(in plasma/serum)
Vitamin A (retinol)
(in plasma/serum)
α-Carotene
(in plasma/serum)
β-Carotene
(in plasma/serum)
Total antioxidant capacity
(in plasma/serum)
Superoxide dismutase
(in plasma/serum/erythrocytes)
Glutathione peroxidase
(in plasma/serum/erythrocytes)
Copper
(in plasma/serum)
Ceruloplasmin (Cp)
(in plasma/serum)
Cp oxidase activity
(in plasma/serum)
Transferrin
(in plasma/serum)
Glutathione reductase
(in erythrocytes)
Catalase
(in erythrocytes)
8-hydroxyguanosine
(in lymphocytes)
Hi Brijesh,
The answers provided by Jose and Barbara are helpful and give you an array of options. It depends on what exactly you are looking at.
For example, we measured postprandial oxidative stress biomarkers after a high-fat meal. We used several markers as indicators of oxidative stress.
In plasma you can measure malondialdehyde (MDA) as a marker of lipid peroxidation, H2O2 (using amplex red reagent methods), advanced oxidation end products (AOPP).
You can also measure the antioxidant capacity of the blood through fairly simple techniques such as trolox equivalent antioxidant capactiy (TEAC).
This is a link to the manuscript https://www.researchgate.net/publication/221876702_Impact_of_short-term_dietary_modification_on_postprandial_oxidative_stress?ev=prf_pub
Best of luck
Article Impact of short-term dietary modification on postprandial ox...
Thanks for answer.I got the point but i want to know that if i am not able to measure all the markers as you have mentioned,Which markers should be focused for oxidative stress assessment?Are 2 0R 3 MARKERS like sod,catalase and MDA sufficient enough to assess oxidative stress?
Oxidative stress is characterized by a decrease of endogenous antioxidant enzymes and by an increase of oxidation products of lipids, proteins, DNA and carbohydrates. I think at least you should measure the enzymes SOD, GPx, GR and oxidation products: MDA (lipid oxidation product), 8-hydroxyguanosine (DNA oxidation product), carbonyl groups (protein oxidation products). And finally, total antioxidant capacity.
Does anyone have protocol for measurement of total antioxidant capacity in blood?
This is a difficult question. RBC for example greatly contribute to antioxidant activity e.g. being packed with Prx2.
So I would try to spin down the cellular fraction and measure anti-oxidant activity in plasma. Just as an really easy starter you could e.g. compare 5 blood plasma by adding DCF and recording "background" fluorescence in a plate reader and upon addition of increasing concentrations of H2O2. Alternatively use Amplex Ultra red to measure residual H2O2 - its extremely sensitive.
Be aware that freezing the samples might affect oxidative state - we have seen that with regard of cell culture medium.
You wanted a method for total antioxidant capacity of blood. The original reference is attached. However, you need to explain whether you are trying to detect blood oxidation after it has been collected or stored outside the body or total body oxidative stress occurring in vivo but measurable in a blood sample. There are many later modifications of the total antioxidant capacity assay, however, this assay will not be sensitive enough for detecting in vivo oxidative stress in most cases.
Sorry that the PDF of the paper did not get attached. Either is was not allowed, or the software did not work. Please contact me and I'll send it privately. [email protected]
Brijesh, I wanted to upload another publication that shows that the Total Antioxidant Capacity assay is not a sensitive method for measuring oxidative stress via blood plasma in a rat. If you search more recent articles in this series, you will see that the CCl4-induced oxidative stress is very high relative to other types induced, so I doubt that lack of TAC was caused by insufficient oxidative stress.
With regard to biomarkers measuring, you should consider the type of tissue and target organism. You must also monitor at least two of the components of the oxidative processes: antioxidant response , oxidative effect and / or total antioxidant capacity.
I think you can get away with measuring MDA, GSH, and protein oxidation. If you are measuring an acute oxidative stress you should see a decline in GSH and an increase in oxidized GSH (GSSG). If that's the case I think it's clear that there was an oxidative stress. I don't think there is one right answer.
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