Biochemical Tests

• Reducing sugars • Non-Reducing sugars • Starch • Lipid • Proteins

Reducing Sugars

Test add 2cm of a solution of the reducing sugar to a test-tube. Add an equal volume of Benedict’s solution. Shake and bring gently to the boil., shaking continuously to minimise spitting.

Observations The initial blue colouration of the mixture turns green, then yellowish and may finally form a brick-red precipitate Basis of test Benedict’s solution contains copper sulphate. Reducing sugars reduce soluble blue copper sulphate, containing copper(ll) ions to insoluble red brown copper oxide containing copper(ll). The latter is seen as a precipitate.

Non-Reducing Sugars

Test Add 2cm of sucrose solution to a test tube. Add 1cm dilute hydrochloric acid. Boil for one minute. Carefully neutralise with sodium hydrogen carbonate (check with ph paper) – care is required because effervescence occurs. Carry out benedict’s test.

Observations As Benedict’s test Basis of test A disaccharide can be hydrolysed to its monosaccharide constitutents by boiling with dilute hydrochloric acid. Sucrose is hydrolysed to glucose and fructose, both of which are reducing sugars and give the reducing sugar results with the Benedict’s test

Test Iodine/potassium iodide test Add 2cm 1% starch solution to a test tube. Add a few drops of l 2 /kl solution. Alternatively add the latter to the solid form of starch

Starch

Observations A blue-black colouration.

Basis of test A polyiodide complex is formed with starch.

Emulsion test Add 2cm fat or oil to a test-tube containing 2cm of absolute ethanol. Dissolve the lipid by shaking vigorously. Add equal volume of cold water.

Lipid

Observations A cloudy white suspension Basis of test Lipids are immiscible with water. Adding water to a solution of the lipid in alcohol results in an emulsion of tiny lipid droplets in the water which reflect light and give a white, opalescent appearance

Protein

Biuret Test Add 2cm protein solution to a test-tube. Add an equal volume of 5% potassium hydroxide solution and mix. Add 2 drops of 1% copper sulphate solution and mix. No heating is required.

Observations A mauve or purple colour develops slowly Basis of test A test for peptide bonds. In the presence of dilute copper sulphate in alkaline solution, nitrogen atoms in the peptide chain form a purple complex with copper(ll) ions. Biuret is a compound derived from urea which also contains the –CONH- group and gives a positive result.