BIOLOGICALLY IMPORTANT MOLECULES

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EXERCISE 6!

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BIOLOGICALLY IMPORTANT
MOLECULES!

REPORT !

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INTRODUCTION !

In this experiment, we will learn about biological important molecules by performing tests to detect the presence of carbohydrates, proteins, lipids, and nucleic acids. Also, we will use biochemical tests to identify the unknown compound that react in the following biochemical tests: Benedict’s test, Iodine test, Sudan IV test, Biuret's Test and the Grease-Spot test, also explain the importance of a positive and negative control. !

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ALL MATERIALS!

1. Test tubes!
2. Benedict’s solution!
3. Iodine!
4. Water-bath !
5. Onion juice!
6. Potato juice!
7. Sucrose juice!
8. Glucose juice!
9. Distilled water!
10. Reducing-sugar solution!
11. Starch solution!
12. Sodium hydroxide (NaOH)!
13. Egg albumen!
14. Honey!
15. Amino acid solution!
16. Protein solution!
17. Water!
18. Acetone!
19. Vegetable Oil!
20. Salad oil!
21. Known lipid solution !
22. Sudan IV!
23. Brown wrapping paper or brown paper bag!
24. Eyedropper!
25. Pipette!
26. Test tube holder!

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METHODS !

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Benedict’s test for reducing sugars !
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Benedict's Test tests are for the presence of simple sugar!
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- First, obtain seven test tubes and label them from 1 to 7 to make it easy to

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recognize when doing the procedure ( See the picture). !

- Second, add the materials to be tested into each tube in the right order and

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quantities base on the information shown in table 6.1 at column “Tube" and
“Solution” ( in the TABLE, CHARTS, GRAPHS Section ). After that, add 2mL of Benedict’s solution into each tube. !

- Then, submerge all of the test tubes into gently boiling water-bath for three

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minutes and observe how the color changes during this time. !

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tubes from the water-bath.!

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you see in table 6.1 ( at column “Benedict’s Color Reaction" ).!

- After 3 minutes have elapsed, using the test tube holder to to remove all of the
- Let the tubes cool down to room temperature and write down the colors that
- Finally, when finished the procedure, dispose the contents as directed by the instructor.! !

The result of Benedict’s test for reducing sugars!

Iodine Test for Starch!

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The Iodine Test will test positive for starches!

- First, collect seven test tubes and label them from 1 to 7 to make it easy to

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recognize when doing the procedure ( See the picture). !

- Second, add the materials to be tested into each tube in the right order and quantities base on the information shown in table 6.1 at column “Tube" and
“Solution” ( in the TABLE, CHARTS, GRAPHS Section ). !

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- After that, add 2 drops of iodine solution into each tube. !
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- Finally, observe the color of these tubes and write down the result that you see in table 6.1 (at column “Iodine Color Reaction" ).!
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The result of Iodine Test for Starch!

Perform the Biuret test for protein!

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The Biuret Test will test positive for proteins!

- First, collect seven test tubes and label them from 1 to 4!

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- Second, add the materials to be tested into each tube in the right order and quantities base on the information shown in table 6.2 at column “Solution” ( in the TABLE, CHARTS, GRAPHS Section ). !

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- Then pour 2mL of 2.5% Sodium Hydroxide ( NaOH ) into each test tube. !
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- After that, pour 20 drops of Biuret reagent into each tube and mix gently. !
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- Finally, observe the color of these tubes and record the result that you see in table 6.2 (at column “Color" ).!

The result of the perform the Biuret test for protein!

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Solubility of lipids in polar and non-polar solvents!

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- First, collect two test tubes. !
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- Then, add 5mL of water into one of the tubes and add 5mL of acetone into the other tube!

- After that, pour a few drops of vegetable oil into each tube.!

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- Finally, observe the result that you see and write down the conclusion about the solubility of lipids in polar and non-polar solvents. !
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Performing the Sudan IV test for lipid !
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The Sudan IV test will test positive for lipids!

- First, collect seven test tubes and label them from 1 to 4!

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- Second, add the materials to be tested into each tube in the right order and

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quantities base on the information shown in table 6.3 at column “Solution” ( in the TABLE, CHARTS, GRAPHS Section ).!

- Then, pour five drops of water into tube 1 and five drops of Sudan IV into each of the remaining tubes ( tube 2,3,4)!

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- Mix the contents of each of the test tubes gently.!
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- Finally, observe the color of these tubes and record the result that you see in table 6.3 (at column “Description of Reaction" ).!

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The result of the performing the Sudan IV test for lipid!

Performing the grease-spot test lipids !

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- First, obtain a piece of brown wrapping paper or brown paper bag which is

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provided by instructor.!

- By using an eyedropper, place a drop of salad oil near the opposite corner of the piece of paper.!

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- Then, allow for the fluids to evaporate.!
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- Finally, examine the paper as you hold up to to a light and write down the

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result and description of what you see in table 6.4 ( column “Description of
Grease-Spot Reaction" )!

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The result of the Performing the grease-spot test lipids!

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TABLE, CHARTS, GRAPHS!
Table 6.1
SOLUTION AND COLOR REACTIONS FOR (1) BENEDICT’S TEST FOR REDUCING SUGARS !
AND (2) IODINE TEST FOR STARCH
Tube

Solution

Benedict’s Color Reaction

Iodine Color Reaction

1

10 drops onion juice

Green

Yellow

2

10 drops potato juice

Blue

Black

3

10 drops sucrose solution

Blue

Yellow

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10 drops glucose solution

Reddish Orange

Yellow

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10 drops distilled water

Blue

Yellow

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10 drops reducing-sugar solution

Reddish Orange

Yellow

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10 drops starch solution

Blue

Black

Table 6.2
SOLUTION AND COLOR REACTIONS FOR THE BIURET TEST FOR PROTEIN
Tube

Solution

Color

1

2mL egg albumen

Light Purple

2

2mL honey

Same Color

3

2mL amino acid solution

Clear Color

4

2mL distilled water

Clear Color

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2mL protein solution

Very Light Purple

Table 6.3
SOLUTION AND COLOR REACTIONS FOR THE SUDAN IV TEST FOR LIPIDS
Tube

Solution

Description of Reaction

1

1mL salad oil + water

2 layers - Oil on top, water on bottom

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1mL salad oil + Sudan IV

Orange

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1mL honey + Sudan IV

Light Orange

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1mL distilled water

Light Orange

Table 6.4
MATERIALS AND GREASE-SPOT REACTION AS A TEST FOR LIPID CONTENT
Food Product

Description of Grease-Spot Reaction

Salad Oil

Oil absorbed in the paper faster than water

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RESULTS!
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Benedict’s test for reducing sugars!

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- Positive - Abundance amount of reducing sugars : glucose and reducing-sugar

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solution (because of the color are reddish orange )!

- Negative - Nonreducing sugars : onion juice ( because of the color is green )and potato juice, sucrose juice, distilled water, starch solution ( because of the color are blue )!

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Iodine Test for Starch!
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- Positive : potato juice and starch solution ( because of the color are black )!
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- Negative : onion juice, sucrose solution, glucose solution, distilled water, reducing-sugar solution ( because of the color are yellow )!

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Perform the Biuret test for protein!
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- Positive : egg albumen and protein solution ( because of the color is light purple)!
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- Negative : honey ( because of the color is the same), amino acid solution and distilled water ( because of the color are clear )!

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Solubility of lipids in polar and non-polar solvents!
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- Oil totally dissolved in acetone, so acetone in non-polar.!
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- Oil does not dissolve in water, so water is polar - oil is non-polar !
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Performing the Sudan IV test for lipid!
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- Salad oil is not soluble in water!
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- Positive : salad oil ( because of the color is deep orange )!
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- Negative : honey and distilled water ( because of the color are light orange )!

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Performing the grease-spot test lipids!
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- Oil absorbed in the paper faster than water, so lipid is present in oil.!
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CONCLUSION!
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!! ! The purpose of this lab was to test various substances for the presence or absence of reducing sugars, starch, sugar, proteins, and lipids. Throughout this lab we observed that glucose and reducing-sugar solution indicates amount of sugar, beside that, starch reacted to iodine along with the potato proving starch present. We also learned that albumen turned out to contain protein by testing it with Biuret Reagent. Another thing we observed was that vegetable oil proved to have lipids by showing translucent spots. Overall, the results was made sense and after this experiment, we have known more about how to use and perform biochemical tests to identify an unknown compound, also detect the presence of carbohydrates, proteins, lipids.

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CITATION!
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!! ! Biology: A Laboratory Manual by Vodopich and Moore, 10th ed., McGraw-Hill,
2014, page 59 - 64.