Biology Concept. Vitamin C Content in Various Fruit Juice

Categories: Biology Food Fruit Lemon Titration Vitamin C

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This experiment examined the effect of vitamin C content in fruit juice on DCPIP’s colour changes using acid-based titration method. A titration is a method of analysis that will allow us to determine the precise endpoint of a reaction and therefore the precise quantity of reactant in the titration flask.

The objective of the experiment was to determine vitamin C content in various fruit juice which it is include freshly squeezed juice( orange and lemon), commercial juice(orange and lemon) and vitamin C tablet.

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As vitamin C is important for human and it cannot be stored in the body, vitamin C should be extracted and human can get the vitamin C by eating fruits and vegetables that have vitamin C. Other than that, the objective is to identify which fruit juices has higher vitamin C content and the lowest vitamin C content. To identify which fruit juices has higher vitamin C content and the lowest vitamin C content. To determine the standard curve of vitamin C based on concentrate of vitamin C.

To differentiate between fresh squeezed fruit juices and commercial carton fruit juices. The best way to extract vitamin C, and for human to benefit from fruit juice is when fruit juice is freshly blended or squeezed. The concentration of vitamin C is more in freshly blended fruit juice that can be useful for human. In short, drinking freshly prepared fruit juice is better than drinking commercial fruit juice such as TWISTER and PEEL FRESH.

Introduction
Vitamin C, also known as L-ascorbic acid, is a water-soluble vitamin that is naturally present in some foods, added to others, and available as a dietary supplement.

Humans, unlike most animals, are unable to synthesize vitamin in their body, it is have to constantly supplied in the daily diet. Vitamin C is required for the biosynthesis of collagen , and certain neurotransmitters . Collagen is an essential component of connective tissue, which plays a vital role in wound healing. If it is applied topically may protect skin from free radical damage after exposure to UV rays. Vitamin C is also involved in protein metabolism. Vitamin C is also an important physiological antioxidant and has been shown to regenerate other antioxidants within the body, including vitamin E. Ongoing research is examining whether vitamin C, by limiting the damaging effects of free radicals through its antioxidant activity, might help prevent or delay the development of certain cancers(protect cell and DNA from damage and mutation),and cardiovascular disease. Diabetes also can benefit from extra vitamin C. It can help regulate blood sugar level. People with diabetes may not have enough vitamin C inside many of their cells. Supplements vitamin C can force it into body cells, where it can protect against many complications of diabetes.Vitamin C helps to prevent heart disease by preventing free radicals from damaging artery walls, which could lead to plaque formation.

This nutrient also keeps cholesterol in the bloodstream from oxidizing, another early step in the progression towards heart disease and stroke. Vitamin C may help people who have marginal vitamin C status to obtain favorable blood cholesterol levels. High blood pressure may also
improve in the presence of this wonder vitamin. All these factors combined make vitamin C an inexpensive and easy way to lower one's risk of heart disease and strokes. Vitamin C during pregnancy is required to convert folic acid into its active form. It increases the absorption of iron from non-animal foods such as spinach and also plays a role in the storage of iron in major organs e.g. the liver. Vitamin C therefore aids the body in the efficient use of folic acid and iron. Some studies suggest that taking vitamin C along with vitamin E may help prevent preeclampsia in women who are at high risk. Preeclampsia, characterized by high blood pressure and too much protein in the urine, is a common cause of premature births. Not all studies agree, however.

DCPIP is a chemical compound used as a redox dye. When oxidized, DCPIP is blue with a maximal absorption at 600 nm; when reduced, DCPIP is colorless. DCPIP used as an indicator for Vitamin C. If vitamin C, which is a good reducing agent, is present, the blue dye, which turns pink in acid conditions, is reduced to a colorless compound by ascorbic acid.

DCPIP (blue) + H+ ——→ DCPIPH (pink)
DCPIPH (pink) + VitC ——→ DCPIPH2 (colorless)
C6H8O6 + C12H7NCl2O2 ——→ C6H6O6 + C12H9NCl2O2

In this titration, when all the ascorbic acid in the fruit juice has been used up, there will not be any electrons available to reduce the DCPIP and the solution will remain pink due to the DCPIP. The end point is a pink color that persists for 10 seconds or more. The objective for the experiment was to determine vitamin C content in the fruit juices( freshly prepared & commercial ) and also in vitamin C tablet. When the first drop of DCPIP is added to the fruit juice, the initial blue colour should quickly disappear - as a consequence of the reaction between any ascorbic acid present and the DCPIP. If this does not happen then the ascorbic acid concentration is very low (or even zero) and a larger volume of fruit juice may be needed in the test tube, because the measurement of the volume fruit juice needed to decolourise DCPIP need to record in a table.

However, because of only vitamin C tablet known its vitamin C content in each mg instead of fruit juice, then when the data is obtained, only the percentage and concentration of vitamin C in each mg is calculated using the formula below to check whether the vitamin C level in the tablet is true or not as claimed by the manufacturer.

Percentage of vitamin C in tablet = weight of vitamin C tablet(mg) 0.1 %
volume of water

Concentration of Vitamin C in tablet = weight of vitamin C tablet(mg) 1.0mgcm¯3 volume of water
PROBLEM STATEMENT

In this experiment we used different types of fruit juice that was freshly squeezed fruit juice and commercial . For freshly squeezed fruit juice we used orange and lemon, same with commercial . We also used different weight of vitamin C tablet (1/4, 1/2, 1, 3/2, 7/4, 2) and mix it with 100 ml of water in order to get real value of vitamin C content since we know the content of vitamin C in each mg from the manufacturer. So then, we used the same procedure with different sample tested by using DCPIP solution.

So that the problem statement is : Do different types of fruit juice sample and weight of vitamin C tablet have similar content of vitamin C?

HYPOTHESIS
Based on the knowledge that I have, my hypothesis of this experiment is : * Freshly squeezed fruit juices and high weight of vitamin C tablet have higher vitamin C content compared to commercial fruit juices and low weight of vitamin C tablet.

VARIABLES
MANIPULATED VARIABLE : Type of fruit juices / weight of vitamin C tablet solution

RESPONDING VARIABLE : Volume of fruit juice needed to decolourise DCPIP solution / volume of vitamin C tablet solution needed to decolourise DCPIP solution

CONSTANT VARIABLE : Volume of DCPIP solution,Volume of water mix with
vitamin C tablet solution

APPARATUS
test tubes, test tube rack, pipette , beaker , syringe , pipette filler , pestle and mortar

MATERIALS
DCPIP solutions, freshly squeezed orange juice, freshly squeezed lemon juice , commercial orange juice , commercial lemon juice , vitamin C tablet , distilled water.

TECHNIQUE
1. Measure and record the volume of fruit juice needed to decolourise 1 ml of DCPIP solution with a graduated syringe. 2. Calculate and record the concentration of vitamin C contain in vitamin C tablet using the formula .

PROCEDURE
Procedure of this experiment was splitted into two part :
1 ) Preparation of standard solution of tablet

(1)The vitamin C tablet is cut and divided into six (2) Each portion is crushed using pestle different portion which is ¼ , ½ , 1 , 1 ½ , 1 ¾ and mortar until it become a powder . and 2.

(3)Each vitamin C powder is poured into a (4) Each of solution filled up in beaker and mixed with 100ml of distilled different syringe and tested with water. DCPIP solution.

2 )Preparation in laboratory

(1)The freshly squeeze(orange,lemon) (2)The 6ml of syringe is filled with first 5 juice and commercial(orange,lemon) ml of commercial orange juice . juice is prepared in a beaker.
(3) 1 ml of 1.0% DCPIsolution is sucked up (4)The DCPIP solution is poured into a into a pipette using a pipette filler . test tube.
(5)The needle of the syringe is placed into (6) The commercial orange juice is added the DCPIP solution drop by drop to the DCPIP solution.The mixture is shaked gently after the addition of each drop of the commercial orange juice. The commercial orange juice is continuosly added until Dcpip solution is decolourised .
More than 5ml needed to decolourised the (7) Steps 2-6 is repeated using commercial Dcpip, then the syringe is filled up again with lemon juice,freshly squeezed orange juice , another 5ml of orange juice.The volume of freshly squeezed lemon juice and ½,1/4,1, commercial orange juice used until DCPIP is 1 ½,1 ¾ ,2 vitamin C tablet that has mixed
decolourised is recorded. with 100 ml of distilled water.The volume of fruit juice and vitamin C tablet solution required to decolourised DCPIP solution in each case is recorded in the following table.

PERCENTAGE OF VITAMIN C =
Weight of vitamin C (mg) × 0.1%
Volume of water (cm3)
PERCENTAGE OF VITAMIN C =
Weight of vitamin C (mg) × 0.1%
Volume of water (cm3)

CONCENTRATION OF VITAMIN C =
Weight of vitamin C (mg) × 1.0mgcm¯3
Volume of water (cm3)
CONCENTRATION OF VITAMIN C =
Weight of vitamin C (mg) × 1.0mgcm¯3
Volume of water (cm3)
(8)The results are tabulated.The percentage and the concentration of vitamin C in vitamin C tablet solution are calculated using the formulae.

½ tablet (500 )| 4.0| 5.0|
1 tablet (1000)| 2.0| 10.0|
1 ½ tablet(1500)| 2.0| 15.0|
1 ¾ tablet(1750)| 2.0| 17.5|
2 tablet (2000)| 3.0| 20.0|
TABLE 1.2
TABLE 1.2

TABLE 1.3

GRAPH 1.1

DISCUSSION
The experiment was about vitamin C contained in the freshly squeezed (orange ,lemon) juice , commercial (orange, lemon) juice and vitamin C tablet solution was titrated with DCPIP solution and the vitamin C concentration in each fruit juice and vitamin C tablet was calculated using the formulae. The results of the experiment is tabulated in a table below Type of freshly squeezed fruit juice| Volume of fruit juice needed to decolourise DCPIP solution(ml)| Orange| 6.0 |

In this study, my hypothesis stated ‘Freshly prepared fruit juices
and high weight of vitamin C tablet have higher vitamin C content compared to artificial fruit juices and low weight of vitamin C tablet’. This hypothesis is not proven. My prediction is supposed to be correct with our group experiment’s result. From the observation, the colour of DCPIP solution changed to colourless when it is added drop by drop with freshly squeezed fruit juice but when added by commercial fruit juice it needed high volume to decolourise it.

According to Table 1.1, volume of orange juice needed to decolourise DCPIP solution is 6.0 ml. It is same with lemon juice. It is not supposed to be like that because generally lemon juice juice contain less vitamin C compared than orange juice. So the volume is supposed to be more than 6.0 ml. The reason for this case is actually our error when taking the reading. Moreover, we did not repeat the experiment three times and that is why we did not get the average volume of fruit juice needed to decolourise DCPIP solution.

½ tablet (500 )| 4.0| 5.0|
1 tablet (1000)| 2.0| 10.0|
1 ½ tablet(1500)| 2.0| 15.0|
1 ¾ tablet(1750)| 2.0| 17.5|
2 tablet (2000)| 3.0| 20.0|
Referring to Table 1.2 , volume of commercial lemon juice needed to decolourise DCPIP solution is 5.0 ml and volume of lemon juice is also 5.0 ml. In this case, the problem is same with Table 1.1. The process of taking the reading is also have an error when the fruit juic e is added drop by drop into DCPIP solution instead of repeat the experiment three times to get the average reading .

TABLE 1.3 GRAPH 1.1
One vitamin C tablet contain 1000 mg of vitamin C as claimed by the manufacturer. Then the experiment conducted to determine the vitamin C content . Based on Table 1.3 and graph 1.1 , there is something wrong with the data. The graph supposed to get a perfect straight line. The higher the volume needed to decolourise DCPIP solution , the lower the concentration of vitamin C in the tablet . Actually when the weight of vitamin C tablet is 2000 mg, the volume of fruit juice needed to decolourise must be less than 2.0 instead of in the Table 1.3. The tablet is going many process before it mixed with distilled water. It may have probability to lose it is chemical properties such as vitamin C content. There a some suggesting way on how to make improvement in this experiment . Oxidation of vitamin C in the juices may happen. Work quickly with the freshly prepared juices. Other than that DCPIP is also easy oxidized .Drop fruit juice quickly in DCPIP solution after being transferred into a test tube.

FURTHER STUDIES
Instead of using DCPIP solution , there is another method to determine vitamin C content in fruit juice. The way to determine the amount of vitamin C in food is to use a redox titration. The redox reaction is better than an acid-base titration since there are additional acids in a juice, but few of them interfere with the oxidation of ascorbic acid by iodine.

Iodine is relatively insoluble, but this can be improved by complexing the iodine with iodide to form triiodide:
I2 + I- <--> I3-
Triiodide oxidizes vitamin C to form dehydroascorbic acid:
C6H8O6 + I3- + H2O --> C6H6O6 + 3I- + 2H+
As long as vitamin C is present in the solution, the triiodide is converted to the iodide ion very quickly. Howevever, when the all the vitamin C is oxidized, iodine and triiodide will be present, which react with starch to form a blue-black complex. The blue-black color is the endpoint of the titration.

This titration procedure is appropriate for testing the amount of vitamin C in vitamin C tablets, juices, and fresh, frozen, or packaged fruits and vegetables. The titration can be performed using just iodine solution and not iodate, but the iodate solution is more stable and gives a more accurate result.

The essential vitamin known as vitamin C is one of the most important elements in an individual’s daily diet. We need Vitamin C to help contribute to good cellular growth, promote function of the circulatory system, and generally help our bodies to develop and maintain themselves. A vitamin C deficiency is a serious issue, and should be caught early and treated with natural foods or dietary supplements that contain plenty of this critical ingredient.

Lots of medical experts agree that a vitamin C deficiency is fairly uncommon in most modern societies. Infant formulas and other food sources are fortified with vitamin C in order to prevent some forms of deficiency of this essential nutrient.

Symptoms of a Vitamin C Deficiency:
A variety of symptoms will show that an individual is suffering from a vitamin C deficiency. Here are some of the top signs of this kind of shortage in the system: * Fatigue – Early on, someone with a vitamin C deficiency will tend to get tired easily and experience reduced energy. Because chronic fatigue is a symptom of so many illnesses, it can be hard to catch a specific condition based on this symptom. * Mood Changes – Individuals with a vitamin C deficiency may become irritable or short tempered. * Weight Loss – As with other “wasting conditions” someone with a vitamin C deficiency may experience sudden weight loss. * Joint and Muscle Aches – Chronic pain in the limbs or joints can be a symptom of a vitamin deficiency. * Bruising – Individuals with a vitamin C deficiency will tend to bruise easily.

Excessive bruising is a sign that the body’s chemistry needs to be improved. * Dental Conditions – Just as a healthy daily dose of vitamin C contributes to healthy teeth and gums, a deficiency can cause deterioration of the gums. Periodontal problems are a symptom of a vitamin C deficiency that has been allowed to develop to a hazardous level. This kind of deficiency was classically called "scurvy" when it happened to mariners who had no access to nutrients (or sometimes even fresh water) on long trips at sea. In today’s world, it is a rare but frightening condition. * Dry Hair and Skin – A change in hair and skin conditions can also signal that the body is not getting enough of vitamin C and other essential vitamins and minerals. * Infections – When an individual does not have enough vitamin C over time, this can have a negative impact on general healing of wounds and the fighting of infections. Generally, the body’s immune system will be compromised.

Vitamin Deficiency Treatments
A shortage of vitamin C in the system can be treated with dietary supplements. However, it’s important to seek professional medical help, rather than self-medicating with natural medicine. Vitamin C supplements are generally not known to have side effects, although at extremely high levels, they can cause nausea or indigestion. Those who experience any of the above negative symptoms should see qualified family practice doctors to talk about what may be the issue and whether a dietary deficiency may be the root cause.

Recommended Intakes
Intake recommendations for vitamin C and other nutrients are provided in the Dietary Reference Intakes (DRIs) developed by the Food and Nutrition Board (FNB) at the Institute of Medicine (IOM) of the National Academies (formerly National Academy of Sciences) [8].

DRI is the general term for a set of reference values used for planning and assessing nutrient intakes of healthy people. These values, which vary by age and gender [8], include: * Recommended Dietary Allowance (RDA): average daily level of intake sufficient to meet the nutrient requirements of nearly all (97%–98%) healthy individuals. * Adequate Intake (AI): established when evidence is insufficient to develop an RDA and is set at a level assumed to ensure nutritional adequacy. * Tolerable Upper Intake Level (UL): maximum daily intake unlikely to cause adverse health effects [8].

Table 1 lists the current RDAs for vitamin C [8]. The RDAs for vitamin C are based on its known physiological and antioxidant functions in white blood cells and are much higher than the amount required for protection from deficiency [4,8,11]. For infants from birth to 12 months, the FNB established an AI for vitamin C that is equivalent to the mean intake of vitamin C in healthy, breastfed infants.

Table 1: Recommended Dietary Allowances (RDAs) for Vitamin C [8] Age Male Female Pregnancy Lactation
0–6 months40 mg*40 mg*
7–12 months50 mg*50 mg*
1–3 years15 mg15 mg
4–8 years25 mg25 mg
9–13 years45 mg45 mg
14–18 years75 mg65 mg80 mg115 mg
19+ years90 mg75 mg85 mg120 mg
SmokersIndividuals who smoke require 35 mg/day
more vitamin C than nonsmokers.
* Adequate Intake (AI)
Sources of Vitamin C
Food
Fruits and vegetables are the best sources of vitamin C (see Table 2) [12]. Citrus fruits, tomatoes and tomato juice, and potatoes are major contributors of vitamin C to the American diet [8]. Other good food sources include red and green peppers, kiwifruit, broccoli, strawberries, Brussels sprouts, and cantaloupe (see Table 2) [8,12]. Although vitamin C is not naturally present in grains, it is added to some fortified breakfast cereals. The vitamin C content of food may be reduced by prolonged storage and by cooking because ascorbic acid is water soluble and is destroyed by heat [6,8]. Steaming or microwaving may lessen cooking losses. Fortunately, many of the best food sources of vitamin C, such as fruits and vegetables, are usually consumed raw. Consuming five varied servings of fruits and vegetables a day can provide more than 200 mg of vitamin C.

Table 2: Selected Food Sources of Vitamin C [12] Food
Milligrams (mg) per serving Percent (%) DV*
Red pepper, sweet, raw, ½ cup 95 158
Orange juice, ¾ cup 93 155
Orange, 1 medium 70 177
Grapefruit juice, ¾ cup 70 117
Kiwifruit, 1 medium 64 107
Green pepper, sweet, raw, ½ cup 60 100

Broccoli, cooked, ½ cup 51 85
Strawberries, fresh, sliced, ½ cup 49 82
Brussels sprouts, cooked, ½ cup 4880
Grapefruit, ½ medium 3965
Broccoli, raw, ½ cup 39 65
Tomato juice, ¾ cup 3355
Cantaloupe, ½ cup 2948
Cabbage, cooked, ½ cup 2847
Cauliflower, raw, ½ cup 2643
Potato, baked, 1 medium 1728
Tomato, raw, 1 medium 1728
Spinach, cooked, ½ cup915
Green peas, frozen, cooked, ½ cup813

*DV = Daily Value. DVs were developed by the U.S. Food and Drug Administration (FDA) to help consumers compare the nutrient contents of products within the context of a total diet. The DV for vitamin C is 60 mg for adults and children aged 4 and older. The FDA requires all food labels to list the percent DV for vitamin C. Foods providing 20% or more of the DV
are considered to be high sources of a nutrient.

SAFETY PRECAUTION
Safety precaution in laboratory during experiment is most important for student to follow. 1) For individual :
* As always be sure to follow any additional instructions given by your lecturer. * Always ensure that you wear proper fitting clothes and nothing that is too loose. You should also wear your lab coat to avoid spilling anything on your clothes. * Wear proper footwear that is closed. Sandals and open toed slippers are a strict no no and should be avoided. * You must also know how to use the lab equipments properly and should ask the right way of usage, in case you do not know. * Make sure you read the bottle labels carefully and only use what you are supposed to use during an experiment. You should not try out anything on your own unless you are very sure of what reagents or chemicals you need to use and their results. * Be prepared for your work in the laboratory. Read all procedures thoroughly before entering the laboratory. Never fool around in the laboratory. Horseplay, practical jokes, and pranks are dangerous and prohibited. * Do not consume food in a laboratory , or any food used for experiment , because it may be contaminated. * Do not touch anything that you are not authorized to touch especially if it is a chemical or an acid that could cause harm to you

2) For experiment :
* Although Vitamin C and 2,6-dichloroindophenol are not considered hazardous, students should wash their hands thoroughly after handling.
* Make sure there is not a bubble inside pipette before sucked up DCPIP solution. * Read the meniscus at eye-level.
* Place white paper behind the syringe so that the meniscus level can properly be seen. * Use a clamp stand if draining a solution from a syringe into a beaker (e.g. in an experiment to measure the rate of flow of the solution) to keep the syringe at a vertical angle (the same as the beaker) to help achieve the most reliable measurement of the time it takes for a solution to drain, etc. * Observe the color changes of solutions against a white background. * Wash and dry equipment before each reuse.

* When timing (using a stop watch) the rate of flow of a certain volume of solution from a syringe into a test tube, choose the same end point to stop the stop watch for each repeat of the experiment. * Do repeat experiments.

* Do a control experiment.
* Keep variables, that are not being tested, controlled.

CONCLUSION
The hypothesis of my study is that I believe that freshly squeezed juice and higher weight of fruit juice contain higher vitamin C compared than commercial fruit juice and lower weight of vitamin C tablet . I made this choice because in my research I learned that vitamin C found more in orange fruit and the manufacturer of the tablet claimed 1 tablet contain 1000 mg of vitamin C. Based on my experiment I cannot conclude that my hypothesis is correct, because I found concentration in vitamin C same in both freshly squeezed juice and commercial fruit juice. When I compared my results to facts in my research, the vitamin C contain was not supposed to be same in these two items.

I did not have the experience to make an adequate conclusion, so I surfed in internet to get answer for my questions. After surfed, I have concluded that the test to determine which food groups contain vitamin C is an extremely sensitive test to the presence of vitamin C; however, I cannot rule out that the test could have been sensitive to another acidic vitamin found in the both materials. It is possible that the test detected same volume of vitamin C in both items because the error in process of taking reading I believe that my experiment is important because it is important to eat foods that contain vitamin C. Our body uses Vitamin C immediately, but our body does not make any of its own vitmain C. So you can see, it is very good if we consume vitamin C from fresh fruit, eaten just like that or make fresh orange of yourself. It is because actually commercial fruit juice doed not have hight content of vitamin C as well as they have preservative and others sweetener that reducing vitamin C properties. In assessing my study design, I have found a strength of this test to be that it is extremely sensitive to the presence of Vitamin C, and a weakness that it may indicate the presence of substances other than just vitamin C. I am planning to continue this experiment to determine what other substances my test solution detects besides vitamin C, in order to understand why the same volume of vitamin C was found in both freshly squeezed fruit juice and commercial fruit juice.

REFERENCE
BOOK :

INTERNET :

* Benefits of vitamin C, Office of Dietary Supplements, National Institute of Health, June 05 2013, http://ods.od.nih.gov/factsheets/vitaminc-healthprofessional/ * Benefits of vitamin C, Jennifer Brett, N.D,

http://health,howstuffworks.com/wellness/food.nutrition/vitamin-supplements/cell.htm * http://nowcomment.com/documents/4543#.Ug72W8rJZMJ
* www.netdoctor.co.uk/focus/nutrition/facts/lifestylemanagement/smokingnutrition.htm * www.En.wikipedia.org/wiki/dichlorophenolindophenol
* http://malaysia.answers.yahoo.com/question/index?qid=20110307233132AADciuQ