Biomolecules – Lab #3
The four major types of large biological molecules are carbohydrates, lipids,
proteins, and nucleic acids. The basic units of three of the four large molecules
(carbohydrates, proteins, and nucleic acids) are composed of chains of smaller
molecules (monomers) connected to form larger molecules (polymers).
The objective of this laboratory exercise is to demonstrate methods of detecting
biomolecules, specifically carbohydrates, proteins, and lipids.
Procedures
CARBOHYDRATES
Carbohydrates include simple sugars and more complex molecules like starch
and cellulose. Monosaccharides such as glucose and fructose are simple
sugars, and are classified by location of the carbonyl group and by number of
carbons in the carbon skeleton (aldose or ketose sugar). Disaccharides are
double sugars and consist of two monosaccharides that are joined through
dehydration synthesis. Polysaccharides such as starch and cellulose are
macromolecules composed of many sugar building blocks.
A. Benedict’s Test
We will use Benedict’s reagent to test for the presence of carbohydrates called
reducing sugars. A reducing sugar is a monosaccharide or disaccharide with a
free aldehyde or ketone group.
In the Benedict’s Test, alkaline solutions of copper are reduced by reducing
sugars. Copper ions are reduced to cuprous oxide, a reddish brown precipitate.
1. Set up a hot water bath by setting a beaker ½ full of water on a hot plate.
When steam begins to rise, turn the temperature down.
2. Number test tubes 1-8 and place in test tube rack. Each test tube should
contain 2.5 mL of Benedict’s reagent.
3. Add eight drops of each test solution (water, glucose, starch, skim milk,
cream, egg albumin, vegetable oil, and your group’s unknown) to the
Benedict’s reagent, mix well, and heat in water bath for 2-3 minutes. Allow
test tubes to cool and observe for color changes and precipitates. A green,
yellow, or red/orange precipitate is indicative of a sugar bearing a free
aldehyde or ketone group.
4. Record observations on data sheet.
B. Potassium iodide (I2KI) Test for Starch
Potassium iodide (I2KI) or Lugol’s solution will produce a color change from light
amber to blue-black in the presence of starch.
1. Number test tubes 1-8 and place in test tube rack.
2. Add 5 mL of test solution (water, glucose, starch, skim milk, cream, egg
albumin, vegetable oil, and your group’s unknown) to the correctly labeled test
tube.
3. Add 1-2 drops of Lugol’s solution to each test tube.
3. Record observations on data sheet.
PROTEINS
Proteins are polymers made up of amino acids linked together by peptide bonds.
There are 20 different naturally occurring amino acids, common to all living
systems. Proteins can be detected by a reaction with certain reagents to form
various colored complexes. We will use the Biuret Test to detect the presence of
protein in test solutions. In the Biuret Test, the change in color from blue to
violet indicates the presence of protein.
Biuret Test
1. Number test tubes 1-8 and place in test tube rack.
2. Add 2 mL of test solution (water, glucose, starch, skim milk, cream, egg
albumin, vegetable oil, and your group’s unknown) to the correctly labeled test
tube.
3. Add 4-6 drops of Biuret solution to all test tubes and mix well.
4. Record the results on data sheet.
LIPIDS
Lipids are very important to living organisms because they serve as a
concentrated source of stored energy. The largest class of lipids, the
triglycerides, are composed of fatty acids and a molecule of glycerol.
Oils are triglycerides that are liquid at room temperature, whereas fats
are solid at room temperature.
We will use the Sudan IV stain to test for the presence of lipids. Sudan IV stain
is a lipid specific stain that only mixes with oils. In a positive test, the Sudan IV
stains the lipid a deep red color.
Sudan IV Stain Test for Lipids
1. Number test tubes 1-8 and place in test tube rack.
2. Add 4 mL of water to each tube.
3. Add 2 mL of test solution (water, glucose, starch, skim milk, cream, egg
albumin, vegetable oil, and your group’s unknown) to the correctly labeled test
tube.
4. Add 8 drops of Sudan IV stain (liquid form) and shake
the tubes vigorously.
5. Observe for 5 minutes and record results on data sheet.
Data Sheet for Biomolecules Lab
CARBOHYDRATES
A. Benedict’s Test for reducing sugars
Tube
1
2
3
4
5
6
7
8
Test Solution
water
glucose
starch
skim milk
cream
egg albumin
vegetable oil
unknown # __
Observations (use + or – and describe color)
_______________________________________
_______________________________________
_______________________________________
_______________________________________
_______________________________________
_______________________________________
_______________________________________
_______________________________________
B. Potassium iodide (I2KI) Test for Starch
Tube Test Solution Observations (use + or – and describe color)
1
water
_______________________________________
2
glucose
_______________________________________
3
starch
_______________________________________
4
skim milk
_______________________________________
5
cream
_______________________________________
6
egg albumin
_______________________________________
7
vegetable oil
_______________________________________
8
unknown # __ _______________________________________
PROTEINS
Biuret Test
Tube Test Solution
1
water
2
glucose
3
starch
4
skim milk
5
cream
6
egg albumin
7
vegetable oil
8
unknown # __
Observations (use + or – and describe color)
_______________________________________
_______________________________________
_______________________________________
_______________________________________
_______________________________________
_______________________________________
_______________________________________
_______________________________________
LIPIDS
Sudan IV Stain Test for Lipids
Tube Test Solution Observations (use + or – and describe color)
1
water
_______________________________________
2
glucose
_______________________________________
3
starch
_______________________________________
4
skim milk
_______________________________________
5
cream
_______________________________________
6
egg albumin
_______________________________________
7
vegetable oil
_______________________________________
8
unknown # __ _______________________________________
My group’s unknown is _____________________.