describe and carry out the Benedict’s test for reducing sugars, the iodine test for starch, the emulsion test for lipids and the biuret test for proteins

Testing for Biological Molecules

In this unit students learn how to identify the four major classes of biomolecules – reducing sugars, starch, lipids and proteins – using simple qualitative chemical tests. Each test is based on a characteristic chemical reaction that produces a visible colour change or physical change.

1. Benedict’s Test for Reducing Sugars

Principle: Reducing sugars possess a free aldehyde or ketone group that can reduce copper(II) ions (Cu²⁺) in Benedict’s reagent to copper(I) oxide (Cu₂O), a red‑orange precipitate.

Reaction (simplified):

\$\text{R–CHO} + 2\,\text{Cu}^{2+} + 5\,\text{OH}^- \rightarrow \text{R–COO}^- + \text{Cu}2\text{O}\downarrow + 3\,\text{H}2\text{O}\$

1. Place 2 mL of Benedict’s reagent in a clean test tube.
2. Add 1 mL of the sample solution (e.g., fruit juice, plant extract).
3. Mix gently and heat the tube in a boiling water bath for 2–3 minutes.
4. Observe the colour of the solution and any precipitate formed.

Observations:

• Blue – no reducing sugar present.
• Green – low concentration of reducing sugar.
• Yellow‑orange – moderate concentration.
• Brick‑red precipitate – high concentration.

Safety notes: Benedict’s reagent contains copper sulphate; avoid skin contact and wear gloves. Use a heat‑proof holder for the boiling water bath.

2. Iodine Test for Starch

Principle: Iodine molecules (I₂) fit inside the helical structure of amylose, forming a charge‑transfer complex that appears blue‑black.

1. Place a small amount of the sample (solid or solution) on a clean white tile or in a test tube.
2. Add 1–2 drops of freshly prepared iodine solution (iodine dissolved in potassium iodide).
3. Mix gently and observe the colour change within 30 seconds.

Observations:

• Blue‑black colour – starch present.
• Yellow‑brown colour – no starch (or only non‑starch polysaccharides).

Safety notes: Iodine solution can stain skin and clothing; handle with gloves and avoid inhalation of vapour.

3. Emulsion Test for Lipids

Principle: Lipids are insoluble in water but soluble in organic solvents such as ethanol. When a lipid‑containing solution is mixed with water, a cloudy emulsion forms.

1. Place 2 mL of ethanol in a test tube.
2. Add 1 mL of the sample (e.g., oil, butter extract).
3. Shake the tube gently to dissolve any lipids in the ethanol.
4. Add 2 mL of distilled water and shake vigorously.
5. Observe the formation of a milky white emulsion.

Observations:

• Milky white cloudy layer – lipids present.
• Clear solution – no detectable lipids.

Safety notes: Ethanol is flammable; keep away from open flames and use in a well‑ventilated area.

4. Biuret Test for Proteins

Principle: Peptide bonds in proteins complex with copper(II) ions in alkaline solution, producing a violet‑purple complex.

Reaction (simplified):

\$\text{Protein} + \text{Cu}^{2+} + \text{OH}^- \rightarrow \text{Protein–Cu}^{2+}\text{ complex (purple)}\$

1. Place 2 mL of Biuret reagent (copper(II) sulfate solution in alkaline medium) in a test tube.
2. Add 1 mL of the sample solution.
3. Mix gently and allow the mixture to stand for 1–2 minutes.
4. Observe any colour change.

Observations:

• Purple or violet colour – protein present.
• Blue‑green colour – no protein (or very low concentration).

Safety notes: The Biuret reagent is strongly alkaline; wear gloves and eye protection.

Summary Table

These qualitative tests provide a rapid way to screen biological samples for the four major classes of macromolecules. Mastery of the procedures, observations and underlying chemistry is essential for A‑Level examinations and practical laboratory work.