1.0 INTRODUCTION:
Oxidative stress is the major driving factor
responsible for the initiation and progression of cancer, diabetes mellitus,
cardiovascular diseases, neurodegenerative diseases, and inflammatory diseases
among other syndromes]. The condition is brought by excessive generation of
free oxygen and nitrogen species or their inefficient quenching in the cell.
Free oxygen and nitrogen species are unstable molecules that are present in the
environment (exogenous) and are also generated in the body (endogenous) during
the normal aerobic metabolic processes in the body. The body possesses a
complex antioxidant defense system, comprising enzymatic and non-enzymatic
pathways, which in the normal physiologic state, maintain a steady equilibrium
between prooxidants and antioxidants, thereby ensuring well-being.
2.0 METHODOLOGY:
·
Five different concentrations
(From 0−1.2mg/mL) of the test sample will be prepared in methanol (analytical
grade).
· The same concentrations will also
be prepared for vitamin C, which will use as a standard antioxidant.
·
The positive control and test
sample will be mixed with 0.2 mL nitroblue tetrazolium (NBT) (0.08 mM), 0.4 mL
NADH (0.25 mM), and 0.2 mL PMS (0.06 mM), respectively.
· The reaction will be initiated
by adding 0.5 mL phenazine methosulfate (PMS) solution (0.12 mM) to the
mixture, incubated at 25 °C for 5 min in dark.
·
The absorbance will be
measured at 560 nm against a blank sample.
· The percentage superoxide
radical scavenging activity of the test sample and vitamin C will be calculated
using the following formula:
% Radical scavenging activity =
Ac – As/Ac
× 100
Where
As is the absorbance of the sample and Ac is the
absorbance of the control.
3.0 ENDPOINT PARAMETER(S):
·
% Radical scavenging activity
4.0 REFERENCE(S):
4.1 Mayank Gangwar, Manish Kumar Gautam, Amit Kumar
Sharma, Yamini B. Tripathi, R. K. Goel, and Gopal Nath. Antioxidant Capacity
and Radical Scavenging Effect of Polyphenol Rich Mallotus philippenensis Fruit
Extract on Human Erythrocytes: An In Vitro Study. e Scientific World Journal
Volume 2014, Article ID 279451, 12 pages http://dx.doi.org/10.1155/2014/279451.
4.2 Habu JB, Ibeh BO. In vitro antioxidant capacity
and free radical scavenging evaluation of active metabolite constituents of
Newbouldia laevis ethanolic leaf extract. Biol Res. 2015;48(1):16.
Published 2015 Mar 14. doi:10.1186/s40659-015-0007-x
4.3 Hardik Joshi, Manoj Pagare, Leena Patil,
Vilasrao Kadam. In–Vitro Antioxidant Activity of Ethanolic Extract of Leaves of
Buchanania Lanzan Spreng. Research J. Pharm. and Tech. 4(6): June 2011; Page
920-924.
4.4 Owen Kenny, Nigel P. Brunton, and Thomas J.
Smyth. In Vitro Protocols for Measuring the Antioxidant Capacity of Algal
Extracts. Methods in Molecular Biology, vol. 1308, DOI
10.1007/978-1-4939-2684-8_24.
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