Practical Assessment Flashcards
Test for reducing sugars
Benedict’s Test
Procedure for Benedict’s Test
1) Place 2cm3 of test solution into a test tube.
2) Add an equal volume of Benedict’s reagent.
3) Stir to mix the mixture.
4) Heat the mixture by placing it in a boiling water bath for 2 minutes.
5) Observe the contents of the test tube.
Basis of Benedict’s Test
- based on ability of reducing sugars to reduce Cu2+ to Cu+
- due to presence of carbonyl groups that are rich in electrons
- blue alkaline solution of CuSO4 receives an electron and is reduced to red, insoluble Cu2O
Observations for Benedict’s Test
Red precipitate - large Orange precipitate - moderately large Yellow precipitate - moderate Green precipitate - small Remains blue - no
Test (procedure) for non-reducing sugars
If a -ve result is obtained in the Benedict’s test:
1) Boil fresh test sample with an equal amount of dilute hydrochloric acid for about 1 minute to hydrolyse disaccharides into monosaccharides.
2) Cool contents of the test tube.
3) Neutralise contents with sodium hydrogen carbonate as Benedict’s solution only works in alkaline conditions.
4) Carry out Benedict’s test on solution.
5) Record observations.
Basis of non-reducing sugars test
- no precipitate observed when heated directly with Benedict’s reagents as there are no electron-rich carbonyl groups available to reduce other compounds
- non-reducing sugar has to hydrolysed into its reducing sugar components
Observations for non-reducing sugars test
Red precipitate - large Orange precipitate - moderately large Yellow precipitate - moderate Green precipitate - small Remains blue - no
Test for starch
Iodine test
Procedure for Iodine Test
1) Add a few drops of potassium iodide solution to 1cm3 of test sample or piece of test specimen
2) Observe any colour change
Basis of Iodine Test
- as ioidine is not very soluble in water, iodine reagent is made by dissolving iodine in water in the presence of potassium iodide; forms a soluble, linear triiodide ion complex
- blue-black colouration observed when starch is present as triiodide ion complex fits into centre of each turn of the amylose helix, forming a starch-ion complex
- boiling causes temporary unwinding of the amylose helix, releasing the iodine molecules; colouration reforms on cooling
Observations for Iodine Test
Mixture turns from yellow-brown to blue-black - starch is present
Mixture remains yellow-brown - starch is absent
Test for proteins
Biuret test
Procedure for Biuret Test
1) Place 2cm3 of test solution in a test tube.
2) Add an equal amount of 5% NaOH solution.
3) Stir to mix mixture well.
4) Add 1% copper sulphate solution, drop by drop, shaking well after each drop.
OR
- add an equal volume of Biuret reagent and shake to mix well
5) Observe any colour change.
Basis of Biuret Test
- In presence of dilute copper sulphate in alkaline solution, nitrogen atoms in peptide chain form a purple complex with copper (II) ions, Cu2+
- Biuret reagent is derived from urea which also contains the -CONH- group, also giving a postive result
Observations for Biuret Test
Mixture turns from blue to purple - proteins are present
Mixture remains blue - proteins are absent
