Introduction

There are four major classifications of biological compounds:

1. Carbohydrates

2. Proteins

3. Lipids

4. Nucleic Acids

Each major classification can be divided into subgroups that correspond to monomers and polymers.

• Monomers are smaller molecules that bond together in dehydration reactions to form larger molecules called polymers.

• Polymers are broken down into monomers in hydrolysis reactions.

Carbohydrates

Carbohydrates are molecules composed primarily of carbon, hydrogen, and oxygen in the form of sugar molecules.
They can be classified as simple sugars (SS) or complex carbohydrates (CC).

• Monomers: Monosaccharides (e.g., glucose, fructose, galactose)

• Disaccharides: Two monosaccharides joined together (e.g., sucrose = glucose + fructose)

• Polysaccharides: Many monosaccharides bonded together (e.g., starches)

Proteins

Proteins are essential biological molecules and fundamental components of all living cells. They include enzymes, hormones, and antibodies that are necessary for proper organism function.

• Monomers: Amino acids (20 total types)

• Polymers: Dipeptides and polypeptides (formed by peptide bonds)

Lipids

Lipids (fats) consist of carbon (C), hydrogen (H), and oxygen (O), but with a higher hydrogen-to-oxygen ratio than carbohydrates. They include triglycerides, phospholipids, and steroids.

• Monomers of triglycerides: 1 glycerol + 3 fatty acids

• Fatty acids:

  • Saturated (no double bonds)

  • Unsaturated (one or more double bonds)

Overview of Chemical Tests

In this lab, you will use various chemical reagents to test for the presence of biological molecules.
Each test includes positive and negative controls for comparison.

Experiment 1 & 2: Benedict’s Test for Sugars and Iodine Test for Starch

Purpose

• Benedict’s Test: Detects simple sugars (monosaccharides).
  Benedict’s solution reacts with reducing sugars to cause a color change.

• Iodine Test: Detects starch molecules.
  If starch is present, the solution changes to black.

Pre-Lab Questions

In this experiment, you will use:

• Negative control: No sugar present (remains blue after heating)

• Positive control: High sugar content (turns orange after heating)

Identify the following before beginning:

Benedict’s Test

1. Negative control: ______________________

2. Positive control: ______________________

Iodine Test
3. Negative control: ______________________
4. Positive control: ______________________

Hypotheses

In Table 1, indicate your hypothesis for each solution. Predict whether each will test positive or negative for sugar and starch, and explain why.

Table 1. Hypotheses for Sugar and Starch Tests

Procedure

Safety: Wear gloves and goggles. Use caution when heating substances. Always direct test tubes away from your face.

Benedict’s Test for Sugar

1. Set up 10 test tubes in a rack.

2. Label each test tube according to Table 1.

3. Add 4 mL of each solution into its corresponding test tube.

4. Add 1 mL of Benedict’s solution to each test tube and mix gently.

5. Place tubes in a boiling water bath for 3 minutes.

6. Observe any color changes and record your results in Table 2.

   • No reaction → “NR”

   • Weak, moderate, or strong reaction → record color change.

Iodine Test for Starch

1. Wash and rinse all test tubes used in the Benedict’s test (keep labels).

2. Set up 8 clean test tubes in the rack.

3. Add 4 mL of each solution.

4. Add 5 drops of iodine solution and mix gently.

5. Observe color changes and record results in Table 2.

Observations

Table 2. Results for Benedict’s and Iodine Tests

Experiment 3: Biuret Test for Proteins

Purpose

The Biuret Test detects the presence of proteins.
Sodium hydroxide (NaOH) and copper sulfate (CuSO₄·5H₂O) react with peptide bonds to form a purple color. The intensity of this color reflects the protein concentration.

Identify Controls

1. Negative control: ______________________

2. Positive control: ______________________

Hypotheses

In the table below, record your predictions for each sample — whether it will test positive or negative for proteins, and why.

Table 3. Biuret Test for Protein

Procedure

Caution: Wear gloves and goggles when handling Biuret solution.

1. Place 10 labeled test tubes in your rack.

2. Add 1 mL of each solution listed above.

3. Add 1 mL of Biuret solution and mix by tapping the tube.

4. Record color changes and indicate intensity (weak, moderate, strong).

5. Dispose of contents properly.

6. Wash, rinse, and dry tubes for the next experiment.

Experiment 4: Sudan III Test for Lipids

Purpose

The Sudan III Test detects lipids.
Sudan III dye combines with lipids to produce a vivid orange color.

Identify Controls

1. Negative control: ______________________

2. Positive control: ______________________

Hypotheses

In Table 4, record your hypothesis for each sample — positive or negative for lipids, and your reasoning.

Procedure

Caution: Do not discard Sudan III down the sink. A disposal container will be provided.

1. On a piece of filter paper, draw several 1-cm circles with a pencil. Label each circle with a sample abbreviation (e.g., W = water, B = butter).

2. Place one drop of each sample on the paper and allow to dry.

3. Immerse the paper in Sudan III stain for 5 minutes.

4. Using forceps, remove the paper and rinse gently in water for 1 minute.

5. Observe coloration and record results in Table 4 (intensity: weak, moderate, or strong).

Paper Bag Test (Alternative for Lipids)

1. Repeat steps 1–2 using a brown paper bag instead of filter paper.

2. Allow to dry. Substances containing lipids will remain translucent.

3. Record results in Table 4.

Observations

Table 4. Sudan III and Paper Bag Tests for Lipids

Conclusions

Write your conclusions based on your experimental results. You should include at least one paragraph for each of the following sections:

• Sugars

• Starches

• Proteins

• Lipids

Each paragraph should:

• Use complete sentences

• Clearly state whether your hypotheses were supported or rejected

• Reference observed results

Guiding Questions

• What did you expect, and why do you think certain substances contained sugar but not starch?

• Did any results from the sugar or starch tests surprise you? Why?

• Were the protein test results consistent with your expectations or with nutrition labels?

• Why do you think you got the results that you did?

Licenses and Attribution

CC Licensed Content, Original:

Other Licensed Content Included:

• Adapted with permission from BioScience I Laboratory Manual by Jamie Colson-Moon and Denise Bristol. Per request from these original authors, this content is now licensed (CC BY-NC-SA).