BIOLOGICALLY IMPORTANT MOLECULES 1

Biologically Important Molecules

Madison Rice

Lourdes University, Sylvania Ohio

Bio Lab-201

Ms. Carnicorn
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INTRODUCTION:

Macromolecules are large biological molecules consisting of thousands of atoms that are

responsible for all of life’s functions. Macromolecules can be divided into four dominant types

including carbohydrates, lipids, proteins, and nucleic acids. Such macromolecules exist in two

structural forms comprised of monomers, which are small molecular subunits, and polymers,

which are large units composed of multiple monomers. Carbohydrates possess carbon,

hydrogen, and oxygen in a 1:2:1 ratio and consist of monosaccharides, or simple sugars,

disaccharides, or paired monosaccharides, and polysaccharides, or linked monosaccharides.

Many monosaccharides can be defined as reducing sugars seeing as they are not bound to

aldehyde or ketone groups. Such characteristics allow for the use of the Benedict’s test to trial

for reducing sugars and the iodine test to distinguish starch within food items containing glucose.

Lipids are composed of carbon, hydrogen, and some oxygen elements that assemble to form

three fatty acid chains on glycerol known as triglycerides. Fats, oils, and waxes consist of lipids

that are popularly characterized by their nonpolar status, making lipids insoluble in water. By

observing lipids physical characteristics, the grease-spot test can be applied to identify lipids in a

substance. Furthermore, proteins are made up of carbon, hydrogen, oxygen, nitrogen, and

sometimes sulfur that form amino acid monomers and peptide polymers. Peptide bonds form

between the amino groups which are targeted by the Biuret test to distinguish proteins.

The purpose of this lab is to use assorted procedures and tests to find the presence of

different macromolecules in food items including unknowns. Such unknowns can be reliably

interpreted with the use of positive and negative controls. When testing for lipids using the

grease-spot reaction, if a substance produces a grease ring then it will contain lipids. While

conducting the Benedict’s test for reducing sugars, if a substance changes to a light blue color,
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then the substance will test positive for reducing sugars. Similarly, while conducting the iodine

test for starch, if a substance turns black then it will test positive for starch molecules and while

testing for proteins if a substance turns purple it will test positive for protein molecules.

PROCEDURE:

The first step was to obtain a brown paper bag and cut out a rectangular square 1 layer

thick that had a length of about 5 inches. The given reactants were then transferred by

eyedropper to the paper as well as the two unknown substances and the distilled water. Each

spot was labeled and given 45 minutes to evaporate. After the correct amount of time elapsed,

the grease rings were observed and compared to the positive and negative controls.

In the second step, 9 test tubes were collected and labeled 1-9 in preparation for

Benedict’s reagent test. Each test tube received 10 drops of Benedicts reagent and 10 drops of a

solution, including the unknown solutions. A hot plate was then obtained along with a 1000mL

beaker with 300mL of water in it and then brought to a boil. Paper towels were placed inside the

beaker to prevent the water from boiling over. All test tubes were then submerged in the boiling

water bath for 3 minutes. Colors were then observed and compared after test tubes were cooled.

In the third step, a second set of 9 test tubes were collected and labeled 1-9 in preparation

for the iodine test. Each test tube received 3 drops of iodide along with 10 drops of a particular

solution. The reactants were the mixed thoroughly and observed.

The final step required 7 test tubes labeled 1-7 with each test tube containing 10 drops of

2.5% sodium hydroxide (NaOH) and 3 drops of Bluret reagent. Solutions were then mixed

thoroughly and observed.

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RESULTS:

Table 1. Grease-Spot Reaction. Olive oil produced the largest ring, establishing olive oil as the

positive control. Additional substances that contained lipids included vegetable oil, unknown

(B), onion juice, and peanut butter. Distilled water produced no ring and remains negative

control.

Food Product Description of Grease-Spot

1. Olive oil Large translucent ring
2. Vegetable oil Large translucent ring
3. Unknown (A) No movement
4. Unknown (B) Small ring
5. Honey No movement
6. Onion Juice Small translucent ring
7. Peanut Butter Small translucent ring
8. Distilled water No movement

Table 2. Benedict’s Test for Reducing Sugars. The positive control can be identified as the

glucose solution, producing a dark orange color. The negative control remains distilled water,

displaying a bright light blue color. Light blue can be identified as having no concentration of

sugars, green having a weak amount, and orange and red having a large amount. Other high

concentrated sugar food substances include sucrose, and reducing-sugar solution.

Tube Solution Benedict’s Color Reaction

1. 10 drops onion juice Bright green
10 drops Benedict’s Reagent
2. 10 drops potato juice Muddy dark green
10 drops Benedict’s Reagent
3. 10 drops sucrose solution Reddish orange
10 drops Benedict’s Reagent
4. 10 drops glucose solution Dark orange
10 drops Benedict’s Reagent
5. 10 drops distilled water Bright light blue
10 drops Benedict’s Reagent
6. 10 drops reducing-sugar solution Light orange
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10 drops Benedict’s Reagent

7. 10 drops starch solution Bright light blue
10 drops Benedict’s Reagent
8. 10 drops unknown (A) Green
10 drops Benedict’s Reagent
9. 10 drops unknown (B) Blue and white separation
10 drops Benedict’s Reagent

Table 3. Iodine Test for Starch. The positive control can be identified as the starch solution,

displaying a dark brown color. The negative control remains distilled water, depicting an orange

brown color. Thus, the orange brown color can be labeled as having little to no starch molecules,

and the dark browns and blacks can be identified as having a large concentration of starch

molecules. Other high concentrated starch substances include, onion juice, potato juice, and

unknown (A).

Tube Solution Iodine Color Reaction

1. 10 drops onion juice Brown
3 drops Iodine
2. 10 drops potato juice Black
3 drops Iodine
3. 10 drops sucrose solution Dark orange/brown
3 drops Iodine
4. 10 drops glucose solution Orange/brown
3 drops Iodine
5. 10 drops distilled water Orange/brown
3 drops Iodine
6. 10 drops reducing-sugar solution Orange/brown
3 drops Iodine
7. 10 drops starch solution Dark brown
3 drops Iodine
8. 10 drops unknown (A) Brown
3 drops Iodine
9. 10 drops unknown (B) Yellow/orange
3 drops Iodine
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Table 4. Buiret Test for Protein. The positive control can be identified as the protein solution,

producing a clear purple color. The negative control remains distilled water, displaying a bright

light blue color. Light blue can be identified as having no concentration of proteins, yellow

having a weak amount, and purple having a large amount. Other high concentrated protein

substances include unknown (A), unknown (B), and egg albumen.

Tube Solution Buiret Color Test

1. 10 drops egg albumen Light purple
3 drops Bluret reagent
10 drops 2.5% (NaOH)
2. 10 drops honey Yellow
3 drops Bluret reagent
10 drops 2.5% (NaOH)
3. 10 drops amino acid solution Light blue
3 drops Bluret reagent
10 drops 2.5% (NaOH)
4. 10 drops distilled water Light blue
3 drops Bluret reagent
10 drops 2.5% (NaOH)
5. 10 drops protein solution Purple
3 drops Bluret reagent
10 drops 2.5% (NaOH)
6. 10 drops Unknown (A) Purple
3 drops Bluret reagent
10 drops 2.5% (NaOH)
7. 10 drops unknown (B) Purple
3 drops Bluret reagent
10 drops 2.5% (NaOH)

Table 5. Chemical Testing to Identify an Unknown.

Biochemical test Color Unknown Results
Sample + Control (+/-)
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Grease-spot test Olive oil Distilled water (A) -

(lipids) Translucent ring No diffusion (B) +
Benedict’s test Glucose solution Distilled water (A) +
(sugars) Red/green variations Light blue (B) -
Iodine test Starch solution Distilled water (A) +
(starches) Back Brown (B) -
Biuret test Protein solution Distilled water (A) +
(proteins) Purple Blue (B) +

DISCUSSION:

To identify the various macromolecules within each unknown solution, a known positive

control can be compared to a negative control. The variation in colors discloses the saturation of

macromolecules within each substance. For the lipid test, the concentrations were interpreted by

distinguishing the largest translucent ring. With these characteristics in mind, the hypothesis

stating substances containing sugars will transition to a light blue color during the Benedict’s test

is incorrect. The light blue color becomes the negative control seeing as the solution is dyed blue

and no reaction can take place when adding the Benedict’s solution to water.

CONCLUSION:

Possible errors could have included cross contamination between the test tubes that would

have weakened the visual displays of the colors and given imprecise results. During the grease-

spot test the diffusion of the highly concentrated lipids such as olive oil and vegetable oil could

have diffused through nearby substances, giving the illusion that they did not diffuse or diffused

significantly, impacting further results.