For hydroxyl radical scavenging assay I will follow this protocol:
Hydroxyl radical scavenging assay Hydroxyl radical scavenging activity of fruit extracts was assayed by the method of Smirnoff and Cumbes17. The reaction mixture 3.0 ml contained 1.0 ml of 1.5 mM FeSO4, 0.7 ml of 6 mM hydrogen peroxide, 0.3 ml of 20 mM sodium salicylate and varied concentrations of the extracts. After incubation for an hour at 37°C, the absorbance of the hydroxylated salicylate complex was measured at 562 nm. The scavenging activity of hydroxyl radical effect was calculated as follows : [1-(A 1-A2) / A0] x 100, where A0 is absorbance of the control (without extract) and A1 is the absorbance in the presence of the extract, A2 is the absorbance without sodium salicylate.
But I've 30% H2O2 solution. Now how can I prepare 6mM H2O2 solution from 30% H2O2 solution? Please, help in this respect as soon as possible. Thanks in advance.
This is my friendly advise. If you unable to dilute H2O2 to a desired concentration, don't proceed further.
Additionally.
How are you going to determine "varied concentrations of the extracts?" 30% H2O2 solution does not have 1 M H2O2.
i think you put the Q wrong. 30% H2O2 has 9.8 M H2O2 in pure form (according to density of 1.11 g/ml) so you can just dilute that 9.8M to your desired value by formula M1V1=M2V2 where M1 is 9.8M, V1 is the required V to take while M2 is 0.006M and V2 is your required V.
Sorry for my mistake as well as late reply and Thanks for answer.
I would like to use five concentration of plant extract (1mg/ml, 0.5mg/ml, 0.250mg/ml….) to scavenge hydroxyl free radical through the above procedure.@Yurii V Geletii
The concentrations are measured in mol/L, not in mg/mL. The scavenging activity of HO radical by some molecule is characterized by the reaction rate constants of HO with these molecules. It's well establish that the hydroxyl radical react with almost all organic molecule with a diffusion control reaction rate constant. Therefore I would predict that activity of all your substrate would almost the same
With due respect, how I can measure the mol/L of a crude plant extract? Here, I share an article for your kind consideration.
Evaluation of antioxidant activity using peroxynitrite as a source of radicals
- February 2002
- Methods in Enzymology 359:366-79
- DOI:
- 10.1016/S0076-6879(02)59199-0
The Determination of Total Concentration and Activity of Antioxidants in Foodstuffs
- January 2002
- Russian Journal of Bioorganic Chemistry 28(6):501-514
- DOI:
- 10.1023/A:1021249717025
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