Qualitative tests for biological molecules

What is a qualitative test

They indicate the presence or absence of a substance. They are descriptive and the results are often seen as a colour change.

How can a solid sample be tested

By grinding or crushing the solid sample and dissolving it in a suitable solvent to form a solution.

Method to test for the presence of lipids

Shake sample with ethanol which will dissolve any lipids present and then add an equal volume of cold water.

Positive a negative results for lipids

Positive result: A white emulsion will form.

Negative result: Solution remains colourless

Explanation of ethanol emulsion test

Lipids are non-polar so they dissolve in ethanol (a non-polar solvent). However the lipid comes out of solution when water (a polar solvent) is added and are dispersed in small droplets which scatter light making it appear cloudy.

Method to test for the presence of proteins

Add biuret reagent to sample, no heat is required.

Positive and negative results for proteins

Positive result: Colour of solution changes from blue to purple

Negative result: Colour of solution remains blue.

Explanation of Biuret test

Copper (II) ions form a complex with the amine groups in the peptide bonds.

Method to test for the presence of starch

Add iodine in potassium iodide solution to the sample.

Positive and negative results for starch

Positive result: Colour of solution changes from yellow/brown to blue/black.

Negative result: Colour of solution remains yellow/brown.

Explanation for iodine test

Iodine forms ions that fit inside the amylose helix.

Method to test for the presence of reducing sugars

Add benedict’s reagent to sample and heat it to 80°c in a water bath for 3 minutes.

Positive and negative results for reducing sugars

Positive result: Colour of solution changes from blue to green/yellow/orange/red (depending on concentration).

Negative result: Colour of solution remains blue.

Explanation for Benedict’s test

The sugar reduces Cu²⁺ ions to Cu⁺ ions which form a red precipitate.

Why can Benedict’s test be described as semi-quantitative

The colour of the solution indicates the concentration of reducing sugar. If equal volumes of solution and excess Benedict’s reagent are used then the concentrations can be compared (blue-none, green-trace, yellow-low, orange-moderate, red-high).

Method to test for the presence of non-reducing sugars

Carry out Benedict’s test first to confirm no reducing sugars are present. Boil a sample with acid (dilute hydrochloric acid is usually used) to hydrolyse the glycosidic bond. Allow the solution to cool and then neutralise it with an alkali (sodium hydrogen carbonate is usually used). Finally, repeat Benedict’s test.

Positive and negative results for non-reducing sugars

Positive results: Colour of solution changes from blue to green/yellow/orange/red (depending on concentration).

Negative results: Colour of solution remains blue.

Explanation of non-reducing sugar test

The non-reducing disaccharides are hydrolysed into monosaccharides which are all reducing sugars.