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In the practical exam you may be handed a white powder or a food extract and asked whether it contains carbohydrate, fat or protein. There are eleven named tests in NCERT's lab manual, and students often run them in a random order. This post explains what each test detects, why it works, and a sensible sequence, with a worked unknown.
Where is this in the syllabus?
The CBSE Chemistry curriculum for 2026-27 lists "Characteristic tests of carbohydrates, fats and proteins in pure samples and their detection in given foodstuffs" in the Class 12 practicals. The method is in Unit 11 of the NCERT Class 12 chemistry lab manual. The theory behind the sugars is in the NCERT Class 12 chapter on Biomolecules. Check the practical list for your exam year.
Which tests detect carbohydrates?
Molisch's test, for all carbohydrates. Add alcoholic 1-naphthol, then pour concentrated H₂SO₄ down the side. A violet ring forms at the junction. The acid breaks glycosidic bonds and dehydrates the sugar to a furfural, which condenses with 1-naphthol. The manual warns that some other organic compounds also give this test.
Fehling's and Benedict's tests, for reducing sugars. On heating, Cu²⁺ is reduced to orange-red Cu₂O. Tartrate (Fehling's) or citrate (Benedict's) keeps Cu²⁺ in solution in alkali so that copper(II) hydroxide does not precipitate.
Tollens' test, for reducing sugars. Ammoniacal silver nitrate is reduced to a silver mirror. The manual says to warm on a water bath, never a direct flame.
Barfoed's test, monosaccharide or disaccharide. Copper(II) acetate in acetic acid is weakly acidic. Monosaccharides give a brick-red precipitate within about five minutes; disaccharides react more slowly because the aldehyde is masked in the acetal link.
Seliwanoff's test, ketose or aldose. Ketohexoses such as fructose give a red colour within two minutes with resorcinol in acid; aldoses do not.
Iodine test, for starch. A blue colour from the starch-iodine complex.
Two points examiners like. Fructose reduces Fehling's reagent even though it is a ketone, because, as the manual explains, reducing sugars contain an aldehyde group or an α-hydroxy ketone group. And sucrose is not a reducing sugar; the manual hydrolyses it with dilute HCl, neutralises, and then the products give the reducing-sugar tests. Our post on biomolecules in chemistry explains why sucrose does not reduce.
How do you test for fats and oils?
The acrolein test. Heat a few crystals of potassium hydrogen sulphate with the oil or ghee. Glycerol, free or combined in the triglyceride, is dehydrated to acrolein, which has a pungent smell. The test detects the glycerol part of a fat.
Which tests detect proteins?
Biuret test. Alkaline copper sulphate gives a violet colour with compounds containing two or more peptide bonds. The manual notes it is not completely specific, since other compounds with two carbonyls linked through nitrogen or carbon also respond.
Ninhydrin test. A blue-violet product, Ruhemann's purple. Ammonia, primary amines, amino acids and peptides also react.
Xanthoproteic test. Heat with concentrated HNO₃; a yellow colour appears, turning orange when 10 M NaOH is added after cooling. Aromatic groups in the protein are nitrated.
Worked unknown: three white powders
Sample problem. You are given glucose, sucrose and starch, unlabelled as A, B and C.
Molisch's test. All three give a violet ring. All are carbohydrates; this step rules out nothing among them.
Iodine. Only C turns blue. C is starch.
Fehling's test on A and B. A gives an orange-red precipitate; B does not. A is a reducing sugar, glucose. B is non-reducing, so sucrose.
Confirm B. Hydrolyse B with HCl, neutralise and repeat Fehling's. A positive result now confirms sucrose.
The same logic applies to a peptide question. A dipeptide has one peptide bond (two amino acids, one link), so it does not meet the biuret condition of two or more. A tripeptide has two and does.
Which mistakes cost marks?
Heating Tollens' reagent over a flame.
Reporting sucrose as reducing because it is a sugar.
Over-boiling in Barfoed's test. The manual warns that prolonged boiling can hydrolyse disaccharides and give a false positive.
Treating ninhydrin as protein-specific. Free amino acids also give it.
A practice routine
Day 1: For each of the eleven tests, write the reagent, the positive result and what it detects.
Day 2: Write the order you would use for an unknown sugar, and justify each step.
Day 3: Plan how to test wheat flour, rice flour and gram flour extracts, as the manual describes.
Day 4: Compare with the functional-group sequence in our post on organic functional group tests.
If you are unsure why a reagent works, the Fermi Concept Library covers every concept in physics, chemistry, biology, mathematics and computing, each with its own lesson.
Before your next practical, write a one-page flowchart that takes any unknown from Molisch's test to a named sugar.
