I want to know how to measure the activity of antioxidants of herbal plants when given to rats for anti-diabetic treatment.
As above-mentionned by Mr Hamza, antioxidant capacity is a multifactorial parameter. Therefore, to have a global picture of your plant extract, you have to test several methodologies. Among the myriads of in vitro chemical methods that have been developped, I would delineate some global method's characteristics that are the most important.
(i) First, some methods are performed in aqueous solution (ORAC for instance), while others are performed in organic solvent (DPPH for instance). If the final objective is to give your extract to rats, it seems to me better to use methods carried out in water.
(ii) Some methods measure the ability of a molecule to protect an oxidizable substrate (lipid, protein...) from being oxidized by a free radical (ORAC for instance), whereas simpler methods just measure the ability of a molecule to reduce a free radical (DPPH for instance). The first kind of methods is called "competitives" because it implies a competition between antioxidant and oxidizable substrate for giving an electron (or a Hydrogen atom) to free radical, while in the second kind of methods, called "non-competitives", there is no such a competition. Competitive methods are often more representative to what happens in natural media compared to non-competitive ones.
(iii) Some methods are performed in homogenous solution (aqueous for instance), while others are performed in multiphasic and compartmentalized media such as micelles, oil-in-water emulsions, liposomes, etc. The second type of methods are the best for me since natural systems are almost always multiphasics (homogenous solutions only exist as artificial media prepared in laboratories).
Hi,
Priliminary phytochemical studies in higher plants, Ethanobotany (3), 84-86- Brinda et. al., 1977. This article may be useful for ur preliminary studies such as presence of tannins, alkaloids,flavonoids,gums& mucilages, lignin, saponins , proteins &free amino acids etc.
Next u can quantitative the total polyphenols by Method (singleton etal.,1965) .
There are number of procedures to determine antioxidant activity as Mickael & Hamza said ORAC, TEAC etc. Some of them i have listed below
DPPH- screening antioxidant ability,
Reducing power ability- to measure the reducing ability of the sample.this exert the antioxidant activity by breaking the free radical chain by donating a hydrogen atom. The reduction of Fe3+/ ferric cyanide complex to the ferrous form.
Hydrogen peroxide scavenging assay -
Hydroxyl radical scavenging assay - shortest half-life compared with other free radicals. Ferrous reducing,
beta- carotene bleaching assay - to analyse the antioxidant activity,
Ferrous chelating ability,
Phosphomoybdate method- quantitative method total antioxidant activity capacity
Bleomycin-dependent DNA damage- sensitive & specific to examine the potential pro-oxidant of the sample.
Hi,
For invivo analysis Lipid peroxidation
Catalase activity
Glutathione peroxidase should also be analysed for antioxidant activity,
look at
Biswapriya B. Misra1, Satyahari Dey. Phytochemical Analyses and Evaluation of Antioxidant Efficacy of in vitro Callus Extract of East Indian Sandalwood Tree (Santalum album L.). Journal of Pharmacognosy and Phytochemistry. Vol. 1 No. 3 2012
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