PROTEINS

Introduction:
Proteins are nitrogenous - substances of high molecular weights and are
fundamental constituents of living things. Elements present in nearly all proteins are
C, N, H, O, and S. Other elements may also be present, i.e., Fe in hemoglobin, I in
thyroglobulin and P in casein. Certain chemical reactions are used to characterize
protein and their composite amino acids. The following include composite tests, a
number of color tests devised to detect presence of proteins and some amino acids
as well as denaturation and precipitation of proteins.
Procedure:
1.Composition Test:
a. In a dry test tube, place about 1.0 g of egg albumin.
b. At the mouth of the tube, suspend a strip of filter paper moistened with lead
acetate solution and a piece of litmus paper moistened with water.
c. Heat the tube diagonally using a test tube holder and carefully without moving it.
QUESTIONS:
1. What happened to the litmus paper? To filter paper? To egg albumin?

2. What is deposited on the walls of the test tube?

3. Based on your results, what can you deduce about the constituent elements
of proteins?

II. Color Reaction Test:

The egg albumin solution to be used in color reaction tests and precipitation
reaction is prepared by heating the egg white in 10 volume of water and filtering it
after through a cheese cloth or coarse filter paper:

1. Milion's Test
a. To about 2 mL of egg albumin solution in a test tube, add 1 to 2 drops of Millon's
reagent. Mix.
b. Heat the mixture gradually to boiling.
c. Note observations and explain the reaction and principle involved.

2. Xanthoproteic Test
a. To about 2 mL of egg albumin solution in a test tube, add 1 ml of conc. HNO3.
Observe the color.
b. Add carefully NH4OH in excess.
c. Record your observations and explain the reaction and principle involved.

3. Biuret Test
a. To about 2 mL of egg albumin solution in a test tube, add an equal amount of 10%
NaOH. Mix well. :
b. Add a drop of 0.5% CuSO4 solution, mixing between drops.
C. Record your observations and explain your results.
 QUESTION: Will amino acid give positive result in Biuret reaction? Why?

III. Precipitation Reactions:

1. Concentrated Mineral Acids and Alkali
a. In 5 separate test tubes, place about 1 mL of egg albumin solution. b. Add drop by
drop conc. HCl, conc. H2SO4, conc. HNO3, 10% NAOH and glacial acetic acid,
separately, until there is an excess. Label the test tube carefully.
c. Record your observations in tabulated form and explain your results.
Heiler's Ring Test: To 1 mL of egg albumin solution, add carefully, down the side of
the tube, 1 ml conc. HNO3. What is formed at the junction of the 2 liquids? Explain
the significance of Heller's Test.

2. Salt of Heavy Metals

a. Place about 1 mL of egg albumin solution in 6 separate test tubes,
b. Add drop by drop solution of HgCl 2, Pb (CH3C00)2, AgNO3, CuSO4, FeCl2, and
BaCl2 (Label the test tubes) separately in the 6 test tries until there is an excess.
c. Record your observations and explain your results.

QUESTION: Why is egg white a good antidote for mercury poisoning?

3. Alkaloidal Reagents
a. Place about 1 mL of egg albumin solution in 3 separate test tubes.
b. Add about 2 drops of the following reagents:
1st tube -5% tannic acid
2nd tube. -- unsaturated picric acid
3rd tube -- 5% potassium ferrocyanide
c. Record your observations and explain your results.

QUESTION: Why is picric and tannic acids used in treating burns and diarrhea?

4. Alcohol
a. Place about 1 mL of ethanol in 3 test tubes containing 1 mL of egg albumin.
b. To the first tube, add 1 drop of dilute HCI solution.
c. To the second, add 2 drops of dilute NaOH solution. Leave the third neutral.
d. Record your observations and explain your results. Note the tube that will have a
maximum precipitation to occur
IV. Coagulation Test:
a. Boil 1 ml of egg albumin solution for 1 minute.
b. Record your observations and explain your results.
V. Test on Plant Proteins (Gluten):
a. Moisten with just enough water to form dough about 5 g of wheat flour. Set aside
for about 30 minutes.
b. Knead and wash off with water the starch particles until you form a rubbery
material.
c. Melt a portion of the material (Gluten) in about 10 mL of dilute HCl in a test tube,
place in a boiling water bath.
d. Test for color reactions using: Millon's, Xanthoproteic and Biuret Tests. e.
Tabulate your results and explain.
 PROTEINS

Name : ___
Group No. : ________ Score
Year & Sec : ___
Date Performed : ___
Date Submitted : ___

Observation and Results

I. Composition Test

Reagent/Substrate Observation

Litmus paper

Filter paper

Egg albumin

From your observation, what can you deduce about the constituent elements of
protein?
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II. Color Reaction Test

Reagent Observation Explanation

Millon’s Test

Xanthoproteic Test

Biuret Test

Will amino acid give positive result in Biuret reaction? Why?

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III. Precipitation Reactions

A. Concentrated Acids and Alkali

Substance Observation

HCl

H2SO4

HNO3

Acetic Acid

NaOH

C. Alkaloid Reagents

Reagent Observation

5% Tannic Acid

Unst’d Picric acid

5% Potassium
ferrocyanide

D. Alcohol

Reagent Observation

HCl

NaOH

Neutral
Questions:
1. Why is egg white a good antidote for mercury poisoning?
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2. Why is picric and tannic acids used intreating burns and diarrhea?
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IV. Coagulation Test

Substance Observation

Egg albumin

V. Test on Plant Protein

Test Results Remarks

1. Millon’s

2. Xanthoproteic

3. Biuret

Conclusion:
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