ANTIOXIDANT

ASSAYS
TOTAL PHENOLICS ASSAY

BY NOOR RABIHAH AID

The Major Classes of Plant Phenolics

*Phenolics and polyphenolic compounds are present in all plant

tissues and frequently form the most abundant secondary
metabolites in fruits, where they sometimes reach high
concentrations.

* It is estimated that plants produce 8,000 phenolic compounds.

Only plants and
* The fundamental structural element that characterizes this group microorganisms are capable
of biologically synthesizing
is the presence of at least one aromatic ring, substituted by at least the aromatic nucleus.
one hydroxyl group. This group is either free, or engaged in another
Animal organisms are
function: ether, ester, or glycoside. almost always dependent
on either their nutrition, or
a symbiosis, to obtain the
indispensable metabolites
that comprise this structural
element.
*Secondary metabolites have been divided into three groups, based
on their biosynthesis:
1. Terpenes and terpenoids
2. Alkaloids
3. Phenylpropanoids INFORMATION

*Most phenolic compounds are derived from phenylpropanoids and

include lignins, lignans and flavonoids.

The several thousand polyphenols that have been described in

plants can subsequently be grouped into several classes. What is secondary metabolites
in plant?

*Distinction between these classes is drawn, on the basis of the Primary metabolites are
number of constitutive carbon atoms, and then on the structure of compounds that are directly
involved in the growth and
the basic skeleton. development of a plant
whereas ,
secondary metabolites are
*Besides simple soluble forms, there are also polymerized forms of compounds produced in other
varyingsolubility (tannins), or completely insoluble forms (lignins) metabolic pathways that,
although important, are not
essential to the functioning of
the plant.
Lack of standardization of FCR methods can result in a difference of
several orders of magnitude in detected phenols, therefore the
following conditions have been recommended for reliable and
predictable data:
* The proper volume ratio of alkali to Folin-Ciocalteu reagent should
be maintained
* Optimal reaction time and temperature should be adhered to for
colour development
* Absorbance should be monitored at 765 nm
* Use of gallic acid as the reference standard phenol
 Method described by Singleton and Rossi
*The total phenolic content (TPC) is determined by
spectrophotometry, using gallic acid as a standard, according the
method described by Singleton and Rossi (1965).
*Briefly, 0.2 mL of the diluted sample extract is transferred in tubes
containing 1.0 mL of a 1/10 dilution of Folin-Ciocalteu’s reagent in
water.
*After waiting for 5-10 minutes, 1.5 mL of a sodium carbonate solution
(7.5% w/v or 0.566 M) is added to the mixture. The tubes are then
allowed to stand at room temperature for 30 min before absorbance at
743 nm is measured.
*The TPC is expressed as gallic acid equivalents (GAE) in mg/100 mL of
food. The concentration of polyphenols in samples is derived from a
standard curve of gallic acid ranging from 0.2 to 4 mg/L
THANK YOU

NOOR RABIHAH AID