Experiment No.

4 Dateː 5 October 2023

Objectiveː Extraction of vitamin A from margarine

Theoryː
Vitamins are the foundation of life. These are organic compounds with a low molecular
weight that belong to xenobiotic compounds essential for both animals and humans. They
should be supplied in small amounts with food or as supplements because they are necessary,
for the proper course of a number of metabolic processes and for controlling important
reactions in the body.
Vitamin A is a term referring to several chemical compounds belonging to the group of
carotenoids, including retinol and its derivatives and beta-carotene. All forms of vitamin A
include substances belonging to polyenes, so they have a similar structure and have a β-
ionone ring configuration. There are three main forms of vitamin A, apart from retinol
(vitamin A1), derived from animal products, i.e., retinoic acid, retinal, and 3-dehydroretinol
(vitamin A2), as well as carotenoids (provitamin A), of plant origin.
As this vitamin belongs to fat-soluble vitamins, its absorption in the body is facilitated by
fats. There are two main sources of vitamin A. One is foods of animal origin, such as liver,
fatty fish, egg yolk, milk, ripened cheeses, and butter. A good source of retinol is meat
products (55%), followed by animal fats (about 23%), milk and dairy products (about 11%),
and food of plant origin: carrots, pumpkin, peach, apricot, chard, spinach, papaya, mango,
parsley leaves, broccoli, kale, and lettuce. 80% of provitamin A is supplied by vegetables and
only 5% by fruit. Vitamin A has numerous physiological functions, playing important roles in
living organisms.
In selecting of method to determine vitamin A content, from the nature sources, include
processes such as esterification, saponification, liquid-liquid extraction, crystallization, and
distillation. Combinations of these techniques are usually used to obtain fractions of the
highest purity. The choice of the method can be determined by the lipid composition of the
food. In the case of low lipid concentration, direct extraction is possible, but, when their
concentration is high, the saponification process should be performed to destroy structures to
which vitamins bind or to eliminate interferences caused by proteins and carbohydrates. The
original physicochemical properties of vitamins remain unchanged through the formation of
the so-called fraction of unsaponifiable compounds, so their further extraction with organic
solvents is possible. The potassium hydroxide (KOH) solution is commonly used for
saponification. It is usually followed by extraction, which is an important step leading to the
separation of the desired compounds from the remaining components of the sample. In the
analysis of fat-soluble vitamins, the most frequently used methods of extraction include:
liquid–liquid extraction, solid phase extraction, supercritical fluid extraction, pressurized
liquid extraction, or microwave-assisted extraction, as techniques alternative to conventional
methods.
Vitamin A (retinol) is an unsaturated monohydric alcohol with the
Molecular formula= C20H30O
Molecular weight=286.48

Molecular structure of Vitamin A

Chemical name of Vitamin A is 3,7-dimethyl-9-(2,6,6-trimethylcyclohex-1-yl) nona-2,4,6,8-
tetraen-1-ol

Physiological functions
The physiological functions of vitamin A is divided into five major classes;
(i) overall growth, (ii) vision, (iii) bone growth, (iv) epithelial growth and (v) reproduction

Properties of Vitamin A:
 Retinol is a viscid, colourless oil.
 Retinol is a steroid hormone.
 It has also been isolated as pale yellowish needles.
 It gives a characteristic absorption band in Ultraviolet spectrum at 328 nm .
 It is fat soluble but insoluble in water.
 Loss of vitamin A occurs while cooking, canning and freezing is less, however,
oxidizing agents destroy it.
 It is destroyed on exposure to UV light.

Material requiredː Margarine, weighing balance

Glass wareː Conical flask, measuring cylinders, china dish, separating funnel,
Chemicalsː Ethanol 95%, KOH 50%, Petroleum ether
Methodː
 Take 10g of margarine in a conical flask.
 Add 25ml of 95% ethanol.
 Add 25ml of 50% KOH.
 Heat and boil it for 5 minutes.
 Transfer the solution in separating funnel, add 100ml petroleum ether and shake for
10 minutes.
 Separate lower layer that contain vitamin A.
 Separate it in china dish and measure the volume.

Calculation A
Weight of Vitamin A extract (g) = 41.9 gm

Result:
We used 250gm margarine box of well known brand “Blue Band”. The Label claims that
250gm margarine box contains 24% Vitamin A. We seperated a layer of Vitamin A from
Blue Band Margarine in Lab which contains 41.9gm Vitamin A content along with little
amount of impurities.

Observationː