OBJECTIVE
Determine the contents of vitamin C (ascorbic acid)  in various food products using small scale redox titration.

INTRODUCTION
It was as long  ago as  in  mid eighteenth century that the beneficial effects of  vitamin C  were discovered.  This is when citrus fruits rich in vitamin C were used to prevent scurvy in  the sea sailors. Since then, vitamin C  has been claimed a miracle agent in curing and  preventing common colds and known as one of the  anti-cancer antioxidants.
Pure ascorbic acid is a  white,  crystalline solid with a molecular formula C₆H₈O_6.  It is well soluble in water. Its main chemical property is that it oxidizes  extremely easily in solution, thus reducing anything that can  accept electrons.  The oxidation process is impeded  in acidic environment and at low temperatures. The process is accelerated  by addition of  light, heat, in basic solutions, and in the presence of metal ions such as Cu²⁺ or Fe³⁺ .
Chemically,  ascorbic acid  is  similar to sugar molecules. In the body , it is oxidized to  dehydroascorbic acid, the fully functional molecule affecting biochemical reactions.
To read more about  vitamin C and its physiological  role as well  biochemical processes  involving  this nutrient  click HERE.
Vitamin C is not produced or stored in the human body, therefore there is a need for a  constant and regular  supply of this vitamin in our everyday diet.  Vitamin C is present in many fruits and vegetables, the best sources being  broccoli, sweet  peppers, parsley, Brussels sprouts,  strawberries, papayas, kiwi, oranges, lemons, and  grapefruits. Amazingly, potatoes are  a reasonable source of vitamin C, particularly when cooked with skin on.  Because  vitamin C  is so easily destroyed by the food processing techniques (due to its water solubility and reducing properties), you may find or ascorbic acid  listed  as an  ingredient on a nutritional label of grocery items, as foods   are frequently  artificially  enriched in  this nutrient.

Analysis of vitamin C
Redox titration is used here to determine the vitamin C content in a sample.  The oxidizing agent,  iodine,  reacts in an acidic environment  with the reducing agent, ascorbic acid,  to produce  iodide ion  and  dehydroascorbic acid.

            ascorbic acid                                      dehydroascorbic acid
Rather than using tedious large -scale burette titrations, you will perform the redox  titration using a small scale burette.
This technique is fast and easy and  allows for  comparison of the vitamin C content  in  many different food items.
The procedure consists of two steps:
1)  the standardization of I₂ solution
2)  the analysis  of a  sample
After the analysis is completed some simple calculations are required.

In this experiment, you will determine the vitamin C content  in a serving  ( typically 250mL ) of various juices  and /or concentrates. You may also  choose to bring your own items, including some solids (breakfast cereals, vegetables, fruits etc.).
If you  need  to develop a technique for analyzing solids (for extra credit), remember that they need to be processed (crushed, chopped, mashed, etc.) and  sometimes vitamin C must  be water-extracted from them. You will  also need the to know the exact mass of the food  sample and/or  the volume of extract prepared as well as  the volume of an aliquot  tested, in order to calculate the vitamin C content in a serving (e.g. 30 g is a serving for Total cereal, but will you test 30 g  or less ?).
The amount of food sampled  for testing will depend on the expected vitamin C  content. The richer the food in vitamin C, the smaller sample can be taken to analyze using our method. The less vitamin C, the more sample must be used in order to obtain good results  in titration.   It may also be possible to keep  smaller sample size of a food item lower in vitamin C, but use a less concentrated I₂ solution.
PROCEDURE
A. Standardization
To standardize the iodine solution here  means  to determine how many drops od iodine solution are required to react with 1 mg of vitamin C.  In every  1.0 ml  of standard ascorbic acid solution used here  there is  1.0 mg of vitamin C.
Therefore, the number of drops of iodine solution used to completely react  with 1.0 ml of the standard solution is equivalent to 1.0 mg of vitamin C.

1. Prepare (and clean and dry if necessary)  a plastic 24-well tray and a short stem beral pipette (=small scale burette).
2. From a provided burette, dispense 1.0 ml  portions of the standard ascorbic acid solution into three wells  of your well plate
3. Add 1 drop of 1 M sulfuric acid and two  drops of starch solution to each well with  standard solution.
4. Fill the short stem beral pipette with  provided iodine solution:  Step 1   Step 2
5. To titrate the first  solution in the well plate:
gently squeeze the small scale burette (tip down, above the solution in the well)  to dispense one  drop of iodine solution at a time, stirring   with a toothpick or small plastic stirrer after every addition, until the solution turns jet-black and the  color persists for at least 10 seconds ( this means that the iodine is no longer reacting with vitamin C and is forming a colored complex with starch = iodine indicator). Make sure to count  all drops and record the number of drops  of I₂ on your data sheet.
6. Repeat  titration for the remaining two samples of  solution.

B. Vitamin C in food
1. Use at least two different  types of food or juices for testing (e.g. orange and lemon  juice, or fresh orange juice vs reconstituted from concentrate orange juice,  or juice vs green pepper, etc.). See Table 1  for sample size and prep.
2. Perform   titrations  on two separate  portions  of the same sample ( juice, etc.) and determine the average # of drops of iodine solution used.
3. Record the manufacturer claim on vitamin C content per serving (check product  label).
IMPORTANT:  Since various foods contain different amounts of vitamin C, the amount of juice or  solid taken for titration or the required concentration of the I₂ solution depends on how rich the food  item is in vitamin C.  If  only 1 or 2 drops of the standard  iodine solution  are  used for titrating a portion of food, you need to:
a. ask the instructor to show you how to properly dilute  the I₂ solution in a 3:1 ratio. However,  I₂ solution will have to be standardized again, to find the proper number of drops per 1 mg of vitamin C,   OR
b. use  at least triple the sample size and repeat the titration with  the original  I₂ solution.
Table 1.

Clean up
Dispose of the solutions from the tray in an appropriate waste container. Wash all equipment used, using soapy water, rinse well  and replace on the instructor's cart.

Calculations and report
Part A: Standardization
1)  Calculate the average number of drops of iodine solution  per 1 mg vit.. C:
      Avg #  drops  per 1 mg vit C  = ( # drops well1 + # drops well2 + # drops well3) / 3
part B
2) Calculate the average # of drops of iodine solution used for titration of your samples:
     Avg # drops B = ( # drops for sample 1 + # drops for sample 2) / 2

3) Calculate the number of mg of vitamin C in a sample of food used:
     mg vit. C  in a sample tested =  Avg # drops B /  Avg # drops per 1 mg vit.C

4) Calculate the content of vitamin C in a serving  (for  juice  =  250 ml):
       vit. C in 250 ml juice = mg vit. C  in a sample tested x (250 ml / volume of sample in mL)
                 Example:  0.5 mg vit C x ( 250 mL /2. ml) = 0.5  mg x 125 = 62.5 mg vit C/serving
4a) For the concentrate:
     find the serving size on the product label  and convert  it  to grams = grams of concentrate /serving.
Hint:  if a serving is in  fl. oz, convert the fl. oz to mL, then use  density of 1.0 g/mL  to calculate the mass,  m= dxV
Then calculate the content of vitamin C in a serving:
     vit. C in one  serving of concentrate =  mg vit. C  in a sample tested  x  (grams of concentrate /serving)
                 Example:   (0.6 mg vit.C /1.0 g sample) x (180 g concentrate/serving) =  108 mg vit C/serving

In your REPORT show all calculations and results,  and  include answers to the  questions on your data sheet.
Extra credit :
If you chose additional  solid food items  brought from home, for which the procedure had to be  developed, please give an outline of the steps followed (include the sample preparation method ( chopped, raw, cooked...), amount taken for testing, etc.

DATA SHEET

Prelab
1. Using the links provided in this document, find the current  RDA (Recommended Daily Allowance) for  vitamin C for adults  of either sex.
 
 
 
 

2. Using the links: http://www.msdsonline.com or http://msds.pdc.cornell.edu/msdssrch.asp find the health hazards associated with ascorbic acid and iodine.
 
 
 
 
 
 

3. What precautions must be taken when working with aqueous  iodine solution?
 
 
 
 
 
 

Additional Prelab for  Extra credit (4 pts):
If you choose to bring  your own solid  food items, you need to develop a  preliminary procedure for testing (how to prepare the sample, amount of the food  used fo titration, etc.- use a step-wise, organized approach here)
If you want to test cooked food,  please cook it at home.  If you need to chop the food, please bring  your own kitchen knife or chop it at home and refrigerate until ready for lab (but no for too long, why?).
Some ideas
you may want to test:
- fresh food vs stored in the refrigerator for a long time, or
- food item cooked in a microwave vs  the same item cooked using a traditional method (in a pot) -  - cooked using various methods (e.g. traditional, pressure cooker, etc.) vs raw