FOL-02

METHOD FOR THE ESTIMATION OF TOTAL PHENOLIC CONTENT USING THE FOLIN- C ASSAY

1.0 Title: Estimation of Total Phenolic content in Salacia Species using the Folin-C (Folin Ciocalteu) Assay.

2.0 Scope: This method is used for Estimation of Total Phenolic content in Salacia Species using the Folin-Ciocalteu Assay.

3.0 Principle: Phenolics include simple phenols, phenolic acids (benzoic and cinnamic acid derivatives), coumarins, flavonoids, stilbenes, hydrolyzable and condensed tannins, lignans, and lignins. These compounds are among the most widely occurring secondary metabolites in the plant kingdom, acting mainly as phytoalexins, attractants for pollinators, and contributors to plant pigmentation, antioxidants, and protective agents against UV light, among others. Although quantitative determination of polyphenols is hampered by their structural complexity and diversity, several methods have been used to determine polyphenols in plant extracts. Assuming that quantification of individual polyphenols does not adequately reveal the proportion of polymeric procyanidins, and then spectrophotometry in the ultraviolet region may be a useful tool to help resolve this problem. Colorimetric reactions are widely used in the UV/VIS spectrophotometric method, which is easy to perform, rapid, reproducible, reliable and applicable in routine laboratory use, and low-cost. However, it is important that colorimetric assay need to use a reference substance, and then this method measures the total concentration of phenolic hydroxyl groups in the plant extract against reference standard. Polyphenols in plant extracts react with specific redox reagents (Folin-Ciocalteu’s phenol reagent) to form a blue complex that can be quantified by visible-light spectrophotometry. The reaction forms a blue chromophore constituted by a phosphotungstic- phosphor- molybdenum complex, where the maximum absorption of the chromophores depends on the alkaline solution and the concentration of phenolic compounds.The reaction generally provides accurate and specific data for several groups of phenolic compounds.

4.0 Reagents:

Gallic acid >99% (Make: Sigma)

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• Folin-Ciocalteu’s phenol reagent (Make: Merck)

Sodium Carbonate (Make: Merck)

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• Methanol HPLC Grade (Make: Rankem)

HPLC grade Water (Make: Rankem)

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5.0 Apparatus:

• UV-Visible Spectrophotometer (Make: Shimadzu UV-1800)

Ultrasonicator (Make: PCI analytics)

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