Ilar, Quenie Mariel M.

Date Performed:

BS Chemistry-3 Date Submitted: January 5, 2016

ASCHEM3

Experiment 4

Vitamins: Their Solubility and Antioxidizing Activity

Abstract: Vitamins are essential organic substances needed in small amounts in the diet for normal
function, growth, and maintenance of body tissues. Vitamins A and E dissolve in organic solvents
whereas the B vitamins and vitamin C dissolve in water. Vitamins C and E are essential antioxidants since
they can defend against free radicals through reduction. Among the four tested vitamin samples
however, vitamin C is the most vulnerable to air, light, and heat exposure. Thus, degradation of the
quality of the vitamins, especially for vitamin C, is probable when exposed to open, humid, and warm
environments.

Introduction

Vitamins are needed in small quantities viruses more effectively. It is essential to

to perform invaluable functions. They are
required for normal function, growth, and
maintenance of body tissues. Without a
sufficient quantity of any vitamin, a deficiency
will occur with a subsequent decline in health.
Fortunately, a balanced diet is often sufficient
enough to meet your needs.
Vitamins fall into two classes: fat-
soluble and water-soluble. Their solubility will
determine how the vitamin is absorbed and
transported by the bloodstream, whether or
not it can be stored in the body, and how easily
it can be lost from the body. Fat-soluble
vitamins are not excreted readily and because
they are stored in body cells, accumulation may
cause toxicity.
Vitamin A is the blanket term for
retinoids, biologically active compounds that
occur naturally in both plant and animal tissues.
It plays a vital role in bone growth, reproduction
and immune system health. It also helps the
skin and mucous membranes repel bacteria and
healthy vision, and may slow declining retinal
function in people with retinitis pigmentosa.
The vitamin A that comes from animal sources
is fat-soluble, and in the form of retinoic acid,
retinal and retinol. Because these retinoids are
very bioavailable and stored in our tissues, too
much animal-derived vitamin A can build up in
the body and become toxic (Weil & Becker,
2012).
The vitamin B-complex refers to all of
the known essential water-soluble vitamins
except for vitamin C. These include thiamine
(vitamin B1), riboflavin (vitamin B2), niacin
(vitamin B3), pantothenic acid (vitamin B5),
pyridoxine (vitamin B6), biotin, folic acid and
the cobalamins (vitamin B12). Each member of
the B-complex has a unique structure and
performs unique functions in the human body.
Vitamins B1, B2, B3, and biotin participate in
different aspects of energy production, vitamin
B6 is essential for amino acid metabolism, and
vitamin B12 and folic acid facilitate steps
required for cell division.
Vitamin E is a vitamin that dissolves in
fat. Vitamin E is an important vitamin required
for the proper function of many organs in the
body. It is also an antioxidant. Antioxidants
work by preventing oxidation reactions that
produce free-radicals which can cause harm to
the body. This means it protects body tissue
from damage caused by substances called free
radicals, which can harm cells, tissues, and
organs.
The body needs vitamin E to help keep
the immune system strong against viruses and
bacteria. Vitamin E is also important in the
formation of red blood cells and the widening of
blood vessels to keep blood from clotting inside
them.
Vitamin C, also known as ascorbic acid,
is a water soluble vitamin found in fruits and
vegetables. It is an antioxidant that is very
essential for human nutrition and proper
functioning of the body (Davey et al., 2000).
Vitamin C is instrumental in neutralizing free
radicals, which are harmful to the body,
assimilation of iron, healing of wounds, helps to
build collagen, which aids the skin, defense
against bacterial and viral infection. Deficiency
of vitamin C in human can lead to a disease
known as scurvy, whose symptoms include
hemorrhaging especially in gums and skin,
loosening of the teeth, joint pains and
exhaustion (Brody, 1994). Many fruits and
vegetables contain vitamin C, but excess
amount of heat can destroy the vitamin
completely. At high temperature, in the
presence of sunlight and oxygen in air, vitamin
C reacts and it is oxidized.
In this experiment, the four vitamins
introduced above will be tested according to
their solubility and antioxidizing activity. The
objective of this is to categorize the vitamins
into water-soluble or fat-soluble. Moreover, the
experiment aims to identify which among the
vitamins are capable of reacting with free
radicals and to also observe the effect of heat
to a certain vitamin like ascorbic acid.
Methodology
The apparatuses used in conducting the
test for the solubility and antioxidizing activity
of vitamins were: 10 mL test tubes to contain
the vitamin samples and a test tube rack to hold
these containers; mortar and pestle to grind the
vitamins and maximize solubility; droppers; and
a spot plate to set the vitamins to be tested for
its antioxidizing activity. For the standardization
of vitamin C in a certain brand of tablet, a
titration set-up must be assembled with the
following apparatuses: iron stand, buret clamp,
50-mL buret, and a 100 mL Erlenmeyer flask to
contain the solution of vitamin C. The same set-
up must be used for the determination of the
amount of vitamin C destructed after applying
heat for 3 and 8 minutes. A heat source, an
alcohol lamp or a hot plate, is necessary to heat
the ascorbic acid solution before cooling it at
room temperature and subjecting it to titration.
The chemicals needed to test for
Vitamins A, B-complex, C and E were:
Methylene chloride as the organic solvent; 0.1
N iodine solution as the reacting substance for
the antioxidants; 1% starch solution as the
titration indicator; and 0.1N acetic acid.
The experiment was divided into four
parts. The first two is on the solubility of
vitamins in water and organic solvent, and the
antioxidizing activity of these vitamins. While
the last two parts are focused on the amount of
vitamin C present before and after subjecting it
to heat.

A. Solubility of Vitamins

To test for the water solubility of the

vitamins, 2 mL of water was added to adequate
amounts of each pulverized sample placed in
separate test tubes. The phase of the resulting
solution was then observed.

Another sample of those vitamins that

formed two layers with water was obtained and
placed in another test tube. Each of the water
insoluble vitamins was allowed to react with 20
drops of methylene chloride, an organic
solvent, to see if the sample is fat-soluble.

B. Antioxidizing Activity of Vitamins

Whole tablets of vitamins C and B-

complex were set separately in the spot plate.
While vitamins A and E was obtained from their
encapsulating films and were also placed in in
the plate. To test whether the vitamin is an
antioxidant or not, a drop of iodine solution was
allowed to react with it. The transformation of
iodine to colorless indicates a positive
antioxidant.
Figure 1.Titration set-up for Vitamin C-iodine
solution (Source: Google Images)
The initial and final volumes of the
iodine solution were noted.
D. Heat destruction of Vitamin C

C. Standardization of Vitamin C The same titration in the previous step

Using an analytical balance, 25 mg of
powdered vitamin C was weighed. The weighed
ascorbic acid was transferred into a 100-mL
Erlenmeyer flask and was added with 125 mL
H2O, 0.5 mL of 0.1N CH3COOH, and 3 drops of
1% starch solution.
The titration set-up shown in Figure 1
was assembled with the iodine solution as the
titrant. The ascorbic acid solution was added
with the titrant until the deep-blue endpoint.
was performed but this time the samples must
be boiled first. Two aqueous vitamin C solutions
were prepared by adding 25 mg of the sample
and 12.5 mL of H2O. The first solution was
boiled for 3 minutes while the other one was
boiled for 8 minutes. Each was allowed to cool
before adding 0.5 mL of 0.1 N acetic acid and 3
drops of starch indicator. Both the boiled
ascorbic acid solutions were titrated to a deep-
blue endpoint and the dispensed volume of
titrant for each sample was determined.
Results and Discussion
Table 1.Solubility and Antioxidizing Activity of
Vitamins
Reduction
Soluble Soluble of red violet
Vitamin in H2O in CH2Cl2 I2 to
(+/-) (+/-) colorless
(+/-)
A - + -
B-
+ - - free-radical molecule. But the antioxidant
complex
C + - +
E - + +
Among the four tested vitamins,
vitamins B-complex and C are soluble in water.
Water soluble vitamins act as coenzymes or
carriers that help the body obtain energy from
food. They are not stored in the body and are
secreted in their urine thus they must be
replaced each day. These vitamins are easily
destroyed or washed out during food storage
and preparation. For instance, when vegetables
are boiled, the water soluble vitamins tend to
seep into the water.
For fat-soluble vitamins, which include
vitamins A and E, are not readily excreted and
stored in fat cells. Adequate absorption of these
vitamins is dependent on efficient fat intake and
absorption since excessive amounts can cause
harm. However, these vitamins are essential in
performing a variety of functions in our body.
According to WebMD, vitamin A is important
for healthy vision, immune function, and proper
cell growth. On the other hand, vitamin E has
been shown to have a wide array of health
benefits, including prevention of stroke,
diabetes, cancer, heart disease, arthritis,
cataracts and improved immune function.
Those vitamins that promoted color
change for the I2 solution from red violet to
colorless are considered as antioxidants. The
color change indicates the reduction of
molecular iodine to iodide ions by the reducing
activity of the antioxidizing vitamin.
Antioxidants are molecules that can neutralize
free radicals by accepting or donating an
electron to eliminate the unpaired condition.
Typically this means that the antioxidant
molecule becomes a free radical in the process
of neutralizing a free radical molecule to a non-
molecule will usually be a much less reactive
free radical than the free radical neutralized.
The antioxidant molecule may be very large
allowing it to "dilute" the unpaired electron, it
may be readily neutralized by another
antioxidant and/or it may have another
mechanism for terminating its free radical
condition. Vitamin E, as an antioxidant, traps
HOO and ROO radicals and forms a less
reactive tocopherol radical. For Vitamin C, it can
donate a hydrogen atom to a free radical
molecule, R, thereby neutralizing the free
radical while becoming a very stable ascorbate
radical.
Table 2.Standardization of Vitamin C
Unboiled Vit. C sample
solution
Mass of pure Vit. C,
27.10 mg
mg
Initial buret reading of
0.00 mL
I2 solution, mL
Final buret reading of
1.10 mL
I2 solution, mL
Volume of I2 solution
1.10 mL
used, mL
Vitamin C-iodine
24.6 mg/mL
solution titer, mg/mL
This method determines the vitamin C
concentration in a solution by a redox titration
using iodine. As the iodine is added during the Table 3.Heat destruction of Vitamin C
titration, the ascorbic acid is oxidized to
Vit. C sample Vit. C sample
dehydroascorbic acid, while the iodine is
solution boiled solution
reduced to iodide ions. for 3 minutes boiled for 8
minutes
+ 2 2 + Initial mass of 25.10 mg 26.00 mg
pure Vit. C, mg
Due to this reaction, the iodine formed Initial buret 0.00 mL 2.00 mL
is immediately reduced to iodide as long as reading of I2
there is any ascorbic acid present. Once all the solution, mL
ascorbic acid has been oxidized, the excess Final buret 2.00 mL 3.30 mL
reading of I2
iodine is free to react with the starch indicator, solution, mL
forming the deep-blue starch-iodine complex. In Volume of I2 2.00 mL 1.30 mL
Figure 2, the endpoint was reached upon the solution used, mL
addition of 1.10 mL of the titrant to the ascorbic Concentration of 0.1 N 0.1 N
I2, N
acid solution. Therefore 24.6 mg of the unboiled
Moles of Vit. C, 0.200 mmol 0.130 mmol
vitamin C reacts with 1 mL of the iodine mmol
solution. Final mass of Vit. 35.22 mg 22.90 mg
C, mg
Mass of Vit. C -10.12 mg 3.10 mg
destroyed, mg (erroneous)

Sample calculation for Table 3

.
1 .
= 2 2
1 2
1 .
= 0.1 1.30
1 2
= 0.130

176.12
. = 0.130

= 22.90

. =
= 26.00 22.90
= 3.10

When 25.10 mg ascorbic acid solution

was boiled for 3 minutes, cooled and titrated
with the iodine solution, the final mass
becomes 35.22 mg. This makes the result
Figure 2.Deep-blue endpoint of Vitamin C
erroneous since there is an increase in the value
-iodine titration
of vitamin C instead of a decrease of its amount
after heat destruction. Significant instrumental
and human errors might have occurred which
resulted to such unreliable data.
However, for the vitamin C sample
boiled for 8 minutes, the data goes to
agreement that when ascorbic acid is exposed
to high temperature, the amount of vitamin
decreases. From an initial mass of 26.00 mg,
only 22.90 mg of the vitamin sample reacted
with iodine. The decrease in the amount
signifies that 3.10 mg of vitamin C was
destroyed by heat.
Conclusion and Recommendation
Vitamins are categorized into two
according to their nature of solubility: the
water-soluble and fat-soluble vitamins.
Vitamins C and B-complex are water soluble
and can be excreted daily by our body. On the
other hand, vitamins A and E belong to the fat-
soluble type and are not excreted readily by our
system. And because they are stored in body
cells, accumulation may cause toxicity.
Antioxidants work by preventing
oxidation reactions that produce free-radicals
which can cause harm to the body. Vitamins C
and E are considered to be antioxidants and can
provide strong protection against major disease
risk including cancer, atherosclerosis,
inflammation, neurodegenerative, and
cardiovascular diseases.
One of the tested vitamins was vitamin
C. Vitamin C, or ascorbic acid, is a water soluble
antioxidant that plays a vital role in protecting
the body from infection and disease. However,
ascorbic acid is susceptible to oxidation by
atmospheric oxygen over time and the quality
of the vitamin is degraded when exposed to
warm and humid environment. This is why
vitamin C tablets should be stored in a cool and
dry place. In addition, cooking at high
temperature destroys vitamin C since it easily
leaches into the cooking water being a water-
soluble vitamin. Hence, overcooking the
vegetable should be avoided in order to retain
the nutrients and health promoting compounds.
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