EXPERIMENT NO.
:- 4
AIM: To estimate the ascorbic acid content in given solution by 2,6 dichlorophenol indophenol
method
MATERIALS REQUIRED:
Apparatus: Burette, Pipettes, conical flask, measuring cylinders, glass markers,
Reagents: 2,6-Dichlorophenol Indophenol solution, 3% metaphosphoric acid, stock solution of
ascorbic acid, ascorbic acid standard solution (1 mg/5 ml).
THEORY:
In 1933 King and Waugh isolated an antiscurvy substance from orange having strong reducing
nature. The structure was established in 1938 by Hawarth. Vitamin C is a water-soluble natural
antioxidant also known as ascorbic acid. Ascorbic acid is a six-carbon lactone molecule
(C6H8O6). It is synthesized from glucose in many animals and plants, but not in human beings
due to the absence of the enzyme L-gluconolactone oxidase. The reducing nature of vitamin C
is due to its strong tendency to donate reducing equivalents from its enediol group. The
structure of ascorbic acid resembles that of hexose sugars. It can exist as both L- and D-
isomers. However, only L-Ascorbate has vitamin activity
PRINCIPLE:
The titrimetric method is a redox titration method which depends on the reduction of the blue
dye 2,6 dichlorophenolindophenol to a colourless leuco compound by ascorbic acid in solution
or in extracts made from foodstuff. Ascorbic acid is a strong reducing agent because of which
it reduces the dye 2,6 dichlorophenolindophenol and itself gets converted to dehydroascorbic
acid. After equivalence point is reached the next drop of dye gives a pink colour to the solution
indicating the end point. The dye in this titration is coloured in the oxidized form and colourless
in the reduced form.
Role of different reagents:
3% metaphosphoric acid: prevent the air to oxidize ascorbic acid
DCPIP: Used as an indicator in the titration of ascorbic acid. As ascorbic acid is reducing agent,
its concentration can be determined by titrating it with a solution of DCPIP. ascorbic acid is
oxidized to dehydroascorbic acid, and DCPIP is reduced from its blue form to a colourless
form.
PROCEDURE:
The step-by-step procedure involved in titrimetric estimation of vitamin C. Rinsed all apparatus
including burettes and pipettes with 3% metaphosphoric acid at the beginning of the
experiment.
Standard titration
5 ml of standard ascorbic acid solution was pipetted in a 100 ml conical flask. The burette was
filled with the dye solution. The solution was titrated against the dye solution till a light pink
colour appeared which persisted for 30 seconds. The dye solution was standardised.
Sample titration
The given unknown solution was taken in a 100 ml volumetric flask. The sample ascorbic acid
solution to the mark was diluted with 3% metaphosphoric acid. Mixed well. 10 ml of this dilute
solution was pipetted into a 100 ml conical flask and titrated against the dye solution till a light
pink colour appeared. which persisted for 30 seconds. The ascorbic acid content of the given
solution was calculated.
RESULT:
The estimated concentration of ascorbic acid in the unknown sample is found out to be
25.4mg/100ml.
