L9. PROTEIN IDENTIFICATION

1-Introduction:

Biuret's test is a chemical test used for detecting the presence of peptide bonds. ( covalent bond formed between two peptides when the carboxyl group of the first one reacts with amino group of the second peptide). Polypeptides as proteins, are chains of amino acids link together by peptides bonds. 

A peptide bond can be broken by hydrolysis (the adding of water); and in organims, the protein molecules called enzymes makee the process easier.

The biuret reaction can be used to assess the concentration of proteins. The intensity of the colour is directly proportional to the protein concentration. 

The solution to be tested is treated with a strong base followed by a few drops of copper (II) sulphate. 

-If the solution turns purple, it will mean that the protein is present. But only peptides with a chain of at least two peptide bonds give a significant measurable colour shift with these reagents.

Proteins in the alkaline environment reduce Cu2+ to Cu+, which forms a coordination complex with proteins, leading to a blue to pink-purple colour change.

2-Objectives

- Identify peptides bonds.

- Compare protein concentration in different foods.

3-Materials:

- 7x250mL

- 6 test tubes

- test tube rack

- 6x10mL pipet

- mortar

- glass marking pen

- gloves

- goggles

- milk

- rice milk

- egg

- yogurt

- potato

- distilled water

- NaOH 20%

- 10 drops of CuSO4

4-Procedure: 

We are going to determine protein concentration in: milk, rice, yogurt, potato and egg. You've to be careful, cause we are going to test separately the egg white and the yolk. 

First of all we are going to dilute the protein.

1. Add 100mL of distilled water to each 250mL beaker. Label them with (M, RM, EW, EY, Y, P).

2. Separate the egg white and the yolk in another beaker.

3. Smash the potato in a mortar and add some amount of the smashed potato to the P beaker. 

Prepare the samples: 

1. Add 10mL of a dispersion of each food (M, RM, EW, EY, Y, P) to the indicate beaker. Calculate the final concentration. All the groups will use the same dispersion from the beakers.

2. Prepare 6 test tubes and label them. Add 2mL of each test tube of the every food dilution of each beaker.

3. Add 2mL of 20% NaOH dissolution.

4. Shake gently and add 5 drops of CuSO4 in each tube. Allow the mixture to stand for 5 minutes.

5. Note any colour change. Remember that proteins will turn solution pink or purple.

6. Compare the test tubes. 

5-Results and conclusions:

-Milk: positive
-Rice milk: negative
-Egg white: positive
-Egg yolk: negative
-Yogurt: positive
-Potato: positive ------- (it's not correct because potato had starch so it had to be negative.) 
The positive result, are the foods of animals, because all they have proteins. Rice milk have starch like potato so they are negative. 

We ordered from more concentration to less concentration according to the colour. 
1. Egg white.
2. Potato and yogurt.
3. Milk.

6-Questions:

1.- Which food has protein?
Egg white, milk, yogurt, potato, (the animal food)

2.- Which food has more protein? why?
1 Egg white

2 Yogurt 

3 Cow milk 

3.- Do you find any difference between rice and cow milk?
 Cow milk has animal proteins and rice milk has also protein but vegetal ones. 

4.- Is there any difference among milk and yogurt? why?
Yes, yogurt has more protein than milk. Because yougurt has ben fermentade.