Qualitative tests for biological molecules
Qualitative tests for biological molecules indicate the presence or absence of a biological molecule but not concentration. The results can be described e.g. a colour change. If testing a solid sample a solution can be obtained by grinding or crushing the solid sample and dissolving it in a suitable solvent.
Testing for lipids (ethanol emulsion test)
Method: Shake sample with ethanol which will dissolve any lipids present and then add an equal volume of cold water.
Positive result: A white emulsion will form.
Negative result: Solution remains colourless
Explanation: 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.
Testing for proteins (biuret test)
Method: Add biuret reagent to sample, no heat is required.
Positive result: Colour of solution changes from blue to purple
Negative result: Colour of solution remains blue.
Explanation: Copper (II) ions form a complex with the amine groups in the peptide bonds.
Testing for starch (Iodine test)
Method: Add iodine in potassium iodide solution to the sample.
Positive result: Colour of solution changes from yellow/brown to blue/black.
Negative result: Colour of solution remains yellow/brown.
Explanation: Iodine forms ions that fit inside the amylose helix.
Testing for reducing sugars (benedict’s test)
Method: Add benedict’s reagent to sample and heat it to 80°c in a water bath for 3 minutes.
Positive result: Colour of solution changes from blue to green/yellow/orange/red (depending on concentration).
Negative result: Colour of solution remains blue.
Explanation: The sugar reduces Cu²⁺ ions to Cu⁺ ions which form a red precipitate.
Benedict’s test can be described as semi-quantitative as 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).
Testing for non-reducing sugars
Method: 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 results: Colour of solution changes from blue to green/yellow/orange/red (depending on concentration).
Negative results: Colour of solution remains blue.
Explanation: The non-reducing disaccharides are hydrolysed into monosaccharides which are all reducing sugars.
Qualitative tests for biological molecules indicate the presence or absence of a biological molecule but not concentration. The results can be described e.g. a colour change. If testing a solid sample a solution can be obtained by grinding or crushing the solid sample and dissolving it in a suitable solvent.
Testing for lipids (ethanol emulsion test)
Method: Shake sample with ethanol which will dissolve any lipids present and then add an equal volume of cold water.
Positive result: A white emulsion will form.
Negative result: Solution remains colourless
Explanation: 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.
Testing for proteins (biuret test)
Method: Add biuret reagent to sample, no heat is required.
Positive result: Colour of solution changes from blue to purple
Negative result: Colour of solution remains blue.
Explanation: Copper (II) ions form a complex with the amine groups in the peptide bonds.
Testing for starch (Iodine test)
Method: Add iodine in potassium iodide solution to the sample.
Positive result: Colour of solution changes from yellow/brown to blue/black.
Negative result: Colour of solution remains yellow/brown.
Explanation: Iodine forms ions that fit inside the amylose helix.
Testing for reducing sugars (benedict’s test)
Method: Add benedict’s reagent to sample and heat it to 80°c in a water bath for 3 minutes.
Positive result: Colour of solution changes from blue to green/yellow/orange/red (depending on concentration).
Negative result: Colour of solution remains blue.
Explanation: The sugar reduces Cu²⁺ ions to Cu⁺ ions which form a red precipitate.
Benedict’s test can be described as semi-quantitative as 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).
Testing for non-reducing sugars
Method: 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 results: Colour of solution changes from blue to green/yellow/orange/red (depending on concentration).
Negative results: Colour of solution remains blue.
Explanation: The non-reducing disaccharides are hydrolysed into monosaccharides which are all reducing sugars.
