Vitamin C Amount Estimation


     Vitamin C (ascorbic acid) is a very important vitamin to the body. Vitamin C
promotes healthy teeth and gums, helps absorption of iron, aids in maintenance
of normal connective tissue, promotes wound healing, and helps boost the immune
system. With vitamin C being such a useful substance to our bodies, finding good
sources of vitamin C is important. Many people today rely on vitamin supplement
tablets. But fruit juices, vitamin-supplemented drinks, or vitamin supplemented
foods may contain just as much vitamin C as a supplement tablet. Which one is
better though, commercially sold drinks or fresh fruit juices? This was the
research question: Are commercially sold and popularly consumed juices (in

Japan) a good substitute fro fresh fruits in terms of dietary vitamin C? What
this experiment sought to find out was exactly what kind of drink was better in
terms of dietary vitamin C. The juices were titrated into a set amount of DCPIP
and measuring how many millilitres it took for the DCPIP to turn from blue to
clear. The hypothesis was that fresh fruit juices should contain more vitamin C
since they had not been heat treated and probably had spent less time on a shelf
or being transported than commercially sod drinks. This is important since
vitamin C is heat labile. This means that vitamin C is susceptible to change and
unstable or that the vitamin C can break down easily if exposed to high
temperatures or is kept for a long time on a shelf. The experiment and results
showed that vitamin C is more abundant in fresh fruit juices. This was true for
all the juices tested except for lemon. Therefore, it is safe to say that fresh
fruit juices tend to contain more vitamin C than commercially bought juices.

Introduction The body needs a good balance of foods, which must contain
carbohydrates, proteins, and fats along with mineral salts, water, fibre, and
vitamins. All of these are required in different amounts according to different
people. However, there are recommended daily allowances. For example, the
recommended daily allowance for vitamin C is 60mg. Vitamins are easily absorbed
into the bloodstream from the gut. A diet lacking in any particular vitamin will
lead to a deficiency disease. Such diseases are rickets that is caused by lack
of vitamin D, and night blindness that is caused by lack of vitamin A. However,
these can be remedied by using vitamin supplements if the dietary intake is
inadequate. The aim of the experiment was to see the difference of vitamin C
content between fresh fruit juices and commercially sold and popularly consumed
juices (in Japan) a good substitute for fresh fruits in terms of dietary vitamin

C? This research question was established because in the modern day and age
people are too busy, especially in winter, to stock up on fresh fruit and many
people rely on commercially sold drinks as a source of vitamins. However,
vitamin C, in particular, is known to be labile and therefore likely to be
absent from a cooked food diet. In temperate climates, such as Japan or Europe,
people ear fresh fruits in summer, but eat tinned, preserved, or cooked foods in
the winter. The latter being more susceptible to heat, possibly breaking down
the amount of vitamin C in them. This experiment tested for the vitamin C
content in fresh fruit juices and commercially sold drinks. This experiment was
conducted mostly on citrus fruits because vitamin C is said to be abundant in
citrus fruits. The experiment was also performed on non-citrus fruits. The
experiment was performed on these two types of fruit drinks because vitamin C
contributes to maintaining a healthy body, especially during the winter, when
citrus fruits are not in season. AS a result, the amount of vitamin C found in
each type of juice would be essential in knowing what drinks to choose during
the winter to provide the most or the optimum amount of vitamin C. Using this
information, the following hypothesis was formed. Since vitamin C is labile
(meaning susceptible to change and unstable), the commercially sold juices,
which have most likely been heat treated and stored in various conditions for
various periods of time, should have lower vitamin C content than fresh fruit
juices. The commercially sold juices would have most likely been exposed to the
conditions leading to the deterioration in the content of vitamin C. In this
experiment the independent variables were the juices that were being tested for
their vitamin C content. The volume of each required to make a standard volume
of DCPIP (dichlorophenolindophenol) change from blue to clear was measured.1

This was the dependent variable. These juices were first filtered and then
titrated using a burette. The fixed variable of the experiment was the amount of

DCPIP in each beaker and the room temperature. Both of these remained constant
throughout the experiment. The importance of the room temperature being constant
and not too high is because otherwise the vitamin C content of any and all the
juices may have been altered, since vitamin C is heat labile. Moreover, if the
temperature varied, the measured results might have varied also. The DCPIP was
carefully made to the concentration of 0.1%. In each case 2 millilitres of DCPIP
was taken. The amount, 2 millilitres of DCPIP, was chosen because it was not too
much or too little an amount for the reactions to be seen clearly, without
taking too much time. Vitamin C was first discovered because of its absence,
during the age of exploration. Sailors on long sea voyages suffered very often
from bleeding gums, loosened teeth, and aching joints. These were the symptoms
of the disease now called scurvy. It is called scurvy because of "the presence
of scurfís (or scales) on the skin". It was James Lind that showed that
scurvy could be cured and prevented by eating "greens, fresh vegetables, and
ripe fruits". However, it was the Polish biochemist, Casimir Funk, which named
the missing group vitamines. He named this because he believed that they
contained an amine group. Vitamines means "life amines". It is from the word
that we get the word, vitamin. When vitamin C was finally isolated in 1925, it
was given the name ascorbic acid because ascorbic means "no scurvy".2

Vitamin C has many functions in the body. One of the most important functions is
as an antioxidant. This means that it helps prevent oxidation of water-soluble
molecules that could otherwise create radicals, which may generate cellular
injury, disease, and damage skin cells. It can also be said that it helps
neuralise or counteract damage to cells caused by free radicals, which can cause
the aging of skin and damage to different cells around the body. Indeed,
ascorbic acid (vitamin C) is commonly added to processed food as an antioxidant.

3 In some roles, vitamin C may act as a coenzyme, helping a particular enzyme to
do its work, especially where metallic ions play a role. Where there are two
oxidation states of metals such as Fe2+ (Iron II) and Fe3+ (Iron III), in the
presence of vitamin C the reduced form (Fe2+) prevails. For iron-deficiency
anemia, vitamin C helps the absorption of iron (especially the nonheme or
vegetable-source iron) from the gastrointestinal tract.4 Specifically, ascorbic
acid works as a coenzyme to convert proline and lysine to hydoxyproline and
hydroxylysine, both important to the collagen structure. 5 Vitamin C also helps
in the stimulation of production of collagen. Collagen is the basis of
connective tissue. It is found in ligaments, skin, cartilage, vertebral discs,
capillary walls, bones, and teeth. As a result, vitamin C helps heal wound in
the ligaments, blood vessels, skin, and cartilage. It also helps prevent hernias
as it protects the inside part of the disc in the vertebral discs where hernias
may occur. Vitamin C is also used in skin treatments because it softens the skin
and prevents or delays the aging of skin.6 It also helps form serotonin which is
an important brain chemical, it stimulates adrenal function, it aids in
cholesterol metabolism, helps wounds heal, and helps maintain healthy blood
vessels.7 In diabetes, vitamin C is commonly used to improve the utilisation of
blood sugar and thereby reduce it, but there is no clear evidence that regular
vitamin C usage alone can prevent diabetes.8 There are some preliminary reports
that ascorbic acid may help prevent cataract formation (probably through its
antioxidant effect) and may be helpful in the prevention and treatment of
glaucoma, as well as certain cases of male infertility caused from the clumping
together of sperm, which decreases sperm function. Ascorbic acid is also said to
act as a detoxifier and may reduce the side effects of drugs such as cortisone,
aspirin, and insulin; it may also reduce the toxicity of the heavy metals lead,
mercury, and arsenic, either by controlling Oxidation State or by facilitating
excretion. There are other proposed functions for vitamin C, but they remain
controversial. For example, it is said that it aids in the production of
interferon, which stimulates the immune system, that it is an antihistamine and
therefore prevents or lessens the affects of allergies, and it may help prevent
certain forms of cancer. In short, vitamin C helps prevent scurvy, promotes
healthy teeth and gums, helps absorption of iron, aids in maintenance of normal
connective tissue, promotes wound healing, and helps boost the immune system.

However, vitamin C is also a natural laxative and may help with constipation
problems. In fact, the main side effect of too much vitamin C intake is
diarrhea. However, this will not happen if you go over the recommended daily
allowance (RDA). For this side effect to occur there would have to be a very
high consumption of vitamin C, very fast because it is a water soluble
molecule.9 Vitamin C is an important substance in the body. This is why it is
vital that we take in the right amount of vitamin C by eating or drinking the
substances that supply us with it. Materials ∑ Fresh grapefruit ∑ Fresh lemon
∑ Fresh orange ∑ Fresh pomegranate ∑ Fresh apple ∑ Fresh mikan (tangerine)
∑ Bottle of C100 Vitamin Lemon drink ∑ Can of Nichirei Acerola drink ∑ Bottle
of Sawayaka apple juice ∑ Carton of Zakuro (pomegranate) Water drink ∑ Carton
of grapefruit juice ∑ Carton of Dole orange juice ∑ Tin of Sanyo mikan ∑

Carton of Ringo No Oishii Mizu (Delicious Apple Water) ∑ Knife ∑ Cutting board
∑ Blender ∑ Lemon squeezer Filter ∑ Distilled water ∑ Burette ∑ DCPIP
(0.1%) solution ∑ Funnel ∑ White marble tile ∑ Paper napkins ∑ 14 beakers ∑

6M hydrochloric acid ∑ Electronic scales ∑ Spatula ∑ Pipette Procedures An
amount of 0.5g of DCPIP (dichlorophenolindophenol) powder was measured on an
electronic scale. Next, 500ml of distilled water were then mixed together to
form 500ml of 0.1% DCPIP solution, which was stored in a dark bottle. The
solution was a dark blue colour. DCPIP is used for the testing of vitamin C.

When tested for vitamin C, a colour change will take place either from blue to
clear, or from blue to pink to clear. Once the full colour change is observed,
the amount of solution taken to change it can be recorded. The burette was
cleaned thoroughly using hydrochloric acid. The acid was poured in gently so as
not to spill while the burette was slowly rotated over a sink. Once this was
finished, water was run down the sink so the acid would be diluted and be less
harmful to the pipe system. Distilled water was then poured down the burette to
make sure the acid was fully rinsed out. The grapefruit had the juice extracted
from it using a regular lemon squeezer. The squeezer was then cleaned using
distilled water. The juice was hen filtered using a tea strainer and then a
filter paper. The juice was collected in a beaker. This procedure was repeated
with an orange, lemon, and a mikan (tangerine). The pomegranate was cut in half
and the juice was extracted by squeezing each half using the hand. This juice
was also filtered in the same fashion as all the juices. The apple was diced and
put into a blender where it was blended until it looked like a puree. This was
then filtered and placed into a beaker like the previous juices. All the juices,
including the commercially sold ju8ices, were filtered to prevent blockage of
the burette. Next, 16 beakers were each filled with 2ml of 0.1% DCPIP solution.

The burette was filled with the fresh grapefruit juice just past the zero line
using a funnel to pour it into the burette so the juice would go directly into
the burette. The grapefruit juice was then drained until it came exactly to the
zero line. Looking at eye level to see if the bottom part of the curvature was
exactly at zero checked this. The grapefruit juice was slowly dripped into the
beaker of DCPIP until a clear observation in the colour change was observed. The
beaker was swirled gently to ensure mixing. It was properly observed, as a white
tile had been placed under the beaker of DCPIP. This made the colour change more
clearly visible. The amount of juice taken for a full colour change to take
place in the beaker containing DCPIP was recorded. The burette was then cleaned
by pouring distilled water through it twice. The lemon juice was then poured
into the burette and the mount of lemon juice taken to observe a clear colour
change in the beaker containing DCPIP was recorded. The burette was cleaned once
again and the process was repeated with each fresh fruit juice and commercially
sold drink. Data Chart showing The Amount of Fruit Juice Needed in mlís to

Turn DCPIP From Blue To Clear Types of Juices Amount of commercially sold juices
in ml needed to turn 2mlís of DCPIP clear Amount of fresh juices in mlís
needed to turn 2 mlís of DCPIP clear Grapefruit 50+ ml 2.2 ml Lemon 0.13 ml

2.0 ml Orange 2.9 ml 2.3 ml Pomegranate 50+ ml 11.3 ml Apple 1.4 ml 23.1 ml

Mikan (tangerine) 6.5 ml 2.9 ml The above chart shows how many mlís of each
juice, both commercially sold and fresh, it took to turn 2mlís of DCPIP clear.

All fruits used to make the fresh fruit juices in the experiment were purchased
fresh so heat and length of time wouldnít have affected the vitamin C content
too much. All the commercially sold juices were bought on the basis of
popularity among teenagerís in Japan. They were bought to represent the likely
amount of vitamin C intake that Japanese teenagerís would have during the
winter when cooked vegetables would lose a lot of their vitamin C content. The
fewer amounts of millilitres of juice it took to turn DCPIP from blue to clear,
the larger the amount of vitamin C there was in the drink. In procuring
commercial fruit juices, it soon became apparent that all was not what it
seemed. Some were heavily supplemented with vitamin C (e.g. the commercially
sold lemon drink "C1000 Lemon" and other, while labeled as fruit juices,
contained only 10% juice!) When the commercially sold lemon drink was first
measured, the colour changed with a mere 0.1ml of lemon drink, or just four
drops. So the experiment was repeated with a one in ten dilution of the lemon
drink, this time giving a reading of 1.3ml. Tinned mikan (tangerine) juice was
used and juice prepared for other fruits, i.e. the tin juice was discarded and
the mikan sections squeezed and filtered. The carton of grapefruit juice was
labeled as containing 20% real fruit juice. This may be why the amount of ml of
juice it took to turn the DCPIP from blue to clear was not established. Over

50ml's of this grapefruit juice was titrated into the beaker containing DCPIP
with little colour change observed. This was shown as more than 50ml on the bar
chart, but in each case 75ml was run in without colour change observed. After
the DCPIP was too dilute to read the colour. The more than 50ml readings should
be interpreted as effectively zero vitamin C content. The juice prepared with a
fresh grapefruit showed that there was nearly as much vitamin C content as in
the fresh lemon juice. Therefore, real grapefruit is high in vitamin C content.

The lemon drink tested was supposed to provide the daily intake of vitamin C. It
only contained 10% real fruit juice. This means that vitamin C that was not
naturally produced was inserted into the drink. This is obvious because when
compared to the commercially sol lemon juice, the amount of fresh lemon juice
needed to turn DCPIP from blue to clear was almost five times the mount of the
commercially sold lemon juice. The commercially sold orange juice contained 100%
real fruit juice. It is easy to tell that this juice was either heat treated or
old, as it is 100% as real as the fresh orange juice made, but it took 0.6 more
ml to turn the DCPIP from blue to clear. The vitamin C content in the
commercially sold orange juice was probably broken down a bit by being heat
treated and being in storage and on a shelf for too long. It is because vitamin

C is heat labile that the vitamin C broke down under these conditions. The
carton of pomegranate 'water' contained 10% real juice. More than 50ml of the
pomegranate 'water' were used to measure the change of DCPIP from blue to clear.

On the bar chart it is shown as greater than 50ml though. However, this change
was not observed even with that amount of the drink being used, therefore the
effective vitamin C content was zero. In the case of fresh pomegranate juice the
change in colour was hard to observe/measure as the colour of the pomegranate
juice and the pink stage of DCPIP was similar in colour. The difference in
colour between the pomegranate juice with the DCPIP pink stage and the
pomegranate juice by itself was observed better using a white tile beneath the
beaker containing the DCPIP and a beaker of fresh pomegranate juice was place on
an other white tile right beside it to make a clearer comparison. The apple
drink (acerola) contained 10% real fruit juice. The acerola apple is different
from the common eating apple used in the fresh fruit juice comparison. It is
more closely related to the crab apple. It was found, after some more research,
that acerola apples have a very high vitamin C content, more so than the common
eating apple. Although extra vitamin C may have been introduced into the drink
[the package label was not helpful], the vitamin C content was still very high.

It contained twenty times the mount of the fresh apple juice tested. Another
popular Japanese apple drink called Ringo No Oishii Mizu (delicious apple water)
was tested for vitamin C content. It contained 20% real fruit juice made of
apples similar to those used for the fresh fruit juice tested. With this apple
drink it took 26.9ml before DCPIP turned clear. The mikan juice extracted from
tinned mikan contained 100% real fruit juice. Tinned goods are heat treated, and
are normally cooked as part of the canning process. Thins tend to have longer
shelf and storage lives too, that would probably account for the decomposition
of vitamin C that gave the relatively low reading in comparison to the fresh
mikan juice. However, given the famed lability of vitamin C, the readings for
canned mikan were surprisingly high and confound the accepted wisdom that
"canned fruit contains no vitamin C". The graph shows the different
amount of each juice in millilitres needed to turn DCPIP from blue to clear.
i.e. a lower reading means more vitamin C. Showing it in graph from makes it
easier to see the differences between the commercially sold drink in comparison
the fresh fruit juice. The Amount of Each Drink in Millilitres Needed to Fulfill
the Required Daily Allowance (RDA) of Vitamin C Commercially Sold Drinks Drink

Amount of Juice in ml's Needed to Turn 2ml's of DCPIP from blue to clear Amount
of Juice in ml's Needed to Fulfill the RDA of Vitamin C Lemon (C1000) 0.13ml

8.4ml Orange 2.9ml 187.34ml Apple (acerola) 1.4ml 90.44ml Mikan 6.5ml 419.9ml

Apple ( Oishii Mizu) 26.9ml 1737.74ml The above chart shows the amount of each
commercially sold drink needed, in millilitres, to fulfill the required daily
allowance (RDA) of vitamin C, which is 60mg of vitamin C. This was figured out
because the lemon drink contained 1000mg of vitamin C and was a bottle of 140ml.

The following equation was then used to figure out how many millilitres of the
lemon drink would provide a person with the RDA for vitamin C. The amount of
mg's of vitamin C in drink = The required daily allowance The amount of ml's of
drink x 1000 = 60 140 x In this equation x was 8.4. Then 8.4 was divided by 0.13
(the amount of lemon C1000 in ml's needed to turn DCPIP from blue to clear). The
number gotten by doing this was 64.6. This number was then multiplied by the
amount of juice, in ml's, needed to turn DCPIP from blue to clear to get the
amount of juice in ml's needed to fulfill the RDA of vitamin C. This also
applies to the next chart. For the lemon, orange, and apple (acerola) drink, the
amounts needed to fulfill the RDA are relatively small. These are amount that
could be easily consumed without much effort and disgust. For the mikan and
apple drinks, the amounts needed to fulfill the RDA are sizeable in comparison
to the lemon, orange, and acerola drinks. Pomegranate and grapefruit juice were
not included in this chart since the amount of juice needed to turn DCPIP clear
was more than the 75ml's measured. The amount of apple juice needed to fulfill
the RDA is impractical for somebody. Drinking almost two litres of the apple
juice in one day would be highly unlikely. Fresh Fruit Juices Drink Amount of

Juice in ml's Needed to Turn 2ml's of DCPIP from blue to clear Amount of Juice
in ml's Needed to Fulfill the RDA of Vitamin C Lemon 2.0ml 129.2ml Orange 2.3ml

148.58ml Apple 23.1ml 1492.26ml Grapefruit 2.2ml 142.12ml Pomegranate 11.3ml

729.98ml For the lemon, orange, mikan, and grapefruit, the amounts needed to
fulfill the RDA are relatively small. For the apple and pomegranate and apple
juices, the amounts needed to fulfill the RDA are quite large. It would be
impractical to drink that much apple juice just to get the RDA of vitamin C. The
juice also tasted bad. It would be fairly hard or expensive to get enough
pomegranate juice to fulfill the RDA of vitamin C. However, it is also
impractical to have about 130 millilitres of fresh lemon juice as it is very
sour and not that tasty. Evaluation From the two types of drinks (commercially
sold drinks and fresh fruit juices), fresh fruit juices tended to contain more
vitamin C than the commercially sold juices of the same fruit. The commercially
sold juices that had a larger vitamin c content than its equivalent fresh fruit
juice were the lemon juice and the first apple juice tested. The lemon juice
contained a lot more vitamin C because it was a vitamin C supplement drink for
those in the winter with colds that don't want to drink the hot cough drinks.

However, no other commercially sold lemon drink, that wasn't a vitamin C
supplement drink, was found. The first apple drink tested for vitamin C had
extra vitamin C added and the type of apple used in the drink had a higher
amount of vitamin C than the normal apple, which was used for the fresh fruit
juice. Not all commercially sold drinks had a lower vitamin c content than their
equivalent fresh fruit juice. This was especially not expected for the first
apple juice tested. Therefore, the hypothesis: since vitamin C is labile
(susceptible to change and unstable), the commercially sold juices, which have
most likely been heat treated and stored in various conditions for various
periods of time, should have lower vitamin c content than fresh fruit juices,
was not fully supported. This was due to the fact that assumptions were made on
the vitamin C content of apples. It was thought that all apples would have
toughly the same vitamin C content, as a result the expected measurements were
not as expected. However, this was somewhat remedied by testing a different
popular apple drink. The results from this test proved to be like those expected
that were stated in the hypothesis. If the experiment were to be repeated, the
most likely change would be to get a wider variation of commercially sold drinks
of the same fruit and, if possible, fresher fruit. Testing for the vitamin C
content in, for example, three different commercially sold apple drinks may have
given a more accurate picture of the vitamin C content in commercially sold
apple drinks. The amount of vitamin C broken down in the canning or packaging
process, along with the shelf life, may have also become more apparent. To get
more accurate results, the experiment should have been done several times. With
all t he results collected an average should have been calculated to give a more
concise amount of vitamin C in the drinks tested, but time was limited. Another
thing that would be good to do if the experiment were repeated would be to test
how much pure vitamin C (ascorbic acid) it takes to turn DCPIP from blue to
clear. This was not achieved because there was no ascorbic acid powder
available. Had it been available, it would have been used as a control. As a
result, the amount of each juice in millilitres to meet the recommended daily
allowance of vitamin C was figured out which served a purpose almost as good as
the control method. In some cases it is conceivable that the volume of fruit or
commercially sold drinks needed to meet the RDA would not be practicable. In
short, it took from 50 - 500 millilitres less of fresh fruit juice than
commercially sold drink to fulfill the RDA of vitamin C. This is for all fruits
except the lemon and acerola commercially sold drinks as they had vitamin C
added. The results that were accumulated through this experiment were nearly all
backed up by the hypothesis, with the exception of the lemon drink comparison
for reasons stated earlier on in the paper. The conclusion was made, in answer
to the research question: are commercially sold and popularly consumed juices
(in Japan) a good substitute for fresh fruits in terms of dietary vitamin C?

That commercially sold and popularly consumed juices (in Japan) are not a good
substitute for fresh fruits (in the form of juices for the purpose of this
experiment). This is because the vitamin C content for all, except the lemon
juice and the first apple (acerola) juice tested, was higher in the fresh fruits
than it was in the commercially sold drinks. So, it would benefit the majority
of teenagers who buy the commercially sold drinks (thinking they contain more
vitamin C among other vitamins and minerals) to drink fresh fruit juices if they
want the proper amount of vitamin C.

Bibliography

Endnotes 1) "Vitamin C Content of a Lemon " The Chemicals of Life p.47

2) Bates, Chris "Vitamin C, The Chameleon of the Vitamins" Biological

Science Review November, 1991, p.11 3) Pitt, George "The Dark Side of

Vitamins" Biological Science Review May, 1994, p.38 4) Bates, Chris
"Vitamin C, The Chameleon of the Vitamins" Biological Science Review

November, 1991, p.12 5) http://www.cforyourself.com 6) http://www.cforyourself.com
& Bates, Chris "Vitamin C, The Chameleon of the Vitamins"

Biological Science Review November, 1991, p.12 7) http://www.cforyourself.com 8)
"Vitamin C Content of a Lemon" The Chemicals of Life p.47 9) http://www.cforyourself.com

Endnotes 1) "Vitamin C Content of a Lemon " The Chemicals of Life p.47

2) Bates, Chris "Vitamin C, The Chameleon of the Vitamins" Biological

Science Review November, 1991, p.11 3) Pitt, George "The Dark Side of

Vitamins" Biological Science Review May, 1994, p.38 4) Bates, Chris
"Vitamin C, The Chameleon of the Vitamins" Biological Science Review

November, 1991, p.12 5) http://www.cforyourself.com 6) http://www.cforyourself.com
& Bates, Chris "Vitamin C, The Chameleon of the Vitamins"

Biological Science Review November, 1991, p.12 7) http://www.cforyourself.com 8)
"Vitamin C Content of a Lemon" The Chemicals of Life p.47 9) http://www.cforyourself.com