Molischs Test - Definition, Test Procedure & Reaction, Uses, FAQs

Edited By Team Careers360 | Updated on Jul 02, 2025 04:50 PM IST

What is Molisch’s test?

Molisch test was named after an Austrian botanist Hans Molisch. Molisch test for carbohydrates is chemical test sensitive to carbohydrates. The carbohydrates can be bound to lipids or proteins or can be in Free State. It requires precision for the detection of carbohydrates.

Molisch meaning

Molisch in English means biochemical test for the presence of sugar.

State the principle of molisch test

Molisch test is based on the principle that concentrated acid helps catalyzing the dehydration of sugars (carbohydrates) to produce furfural (from pentose) or hydroxymethylfurfural (from hexoses) where furfural is an aldehyde with -CHO group attached to the 2-position of furan ring. This furfural molecule condenses with two molecules of phenol (although generally α-naphthol is used but occasionally other phenols like resorcinol and thymol can also be used) to form purple or violet colored complex at the interface of acid-liquid layer.

This Story also Contains

What is Molisch’s test?
State the principle of molisch test
Molisch test reaction
Furfural test vs. Molisch test
Uses of Molisch test
Limitation of Molisch test

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All carbohydrates such as monosaccharides, disaccharides, and polysaccharides (besides triose and tetrose) show positive reactions. Both polysaccharides and disaccharides are first converted into monosaccharides by strong concentrated acid. Other compounds like glycoproteins (or glycolipids) and nucleic acid can give positive reactions as they are first hydrolyzed into monosaccharide by the action of strong acids. Sometimes a green ring can be observed due to presence of impurities in the reagent which might react with the phenol or the acid present in solution. If a solution of concentrated sugar is used, then a rind ring is observed due to charring of sugar by the acid.

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Molisch reagent preparation

Molisch’s reagent is a solution mixture of phenol dissolved in ethanol. This mixture should always be prepared fresh. The exact Molisch reagent composition can be listed as 3.75g of α-naphthol dissolved in 25 ml solution of 99% ethanol.

Molisch test procedure

Molisch test is performed by combining a small amount of Molisch reagent (i.e., α-naphthol dissolved in solution of 99% ethanol) with the solution that needs to be examined. A small amount of concentrated sulfuric acid is poured along the sides of the test tube after combining Molisch reagent and ethanol. The mixture should not be stirred to form two separate layers of acid and test solution along the junction of the test tube.

A purple or violet ring observed at the interface of two liquid layers indicating the presence of carbohydrates in the given solution. When the contents of the test tube (test tube with violet ring at junction) are shaken in a stream of cold water, a deep purple solution is obtained which on dilution with cold water yields a violet precipitate. On shaking the contents of the test tube again and adding a small amount of suspension to an excess of concentrated ammonia, a dull brown color is obtained.

Molisch test procedure with accurate measurements can be performed as:

Take a test solution in a test tube along with 2ml distilled water.
Separate the solution in 4 test tubes to ensure backup if the reaction doesn’t go according to procedure.
In each of the test tubes, add 2 drops of Molisch reagent.
Add 1ml of concentrated sulfuric acid slowly along the sides of each test tube and leave the solution mixture for 2 minutes. DO NOT STIR THE SOLUTION. If the concentrated acid is not poured slowly with precautions, charring of carbohydrates takes place and a black ring may form.

(Tip: to ensure stability of test tube incline the test tube in a test tube holder to restrict movement and prevent charring)

A reddish violet ring is observed between the interface of the acid layer and the solution layer.

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Molisch test reaction

The reaction can be summarized as:

Polysaccharides H₂SO₄→ Monosaccharides (pentoses + hexoses) conc. H₂SO₄→ furfural (and hydroxyfurfural) ∝-naphthol→ Triarylmethane compounds (violet color)Polysaccharides are first converted into monosaccharide in the presence of strong acid. These monosaccharides are converted into furfural; an aldehyde which on reaction with 2 molecules of phenol produces a violet colored complex.

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Furfural test vs. Molisch test

Furfural test; also known as rapid furfural test, is based on the same principle that concentrated acid helps catalyze the dehydration of sugars (carbohydrates) to produce furfural (from pentose) or hydroxymethylfurfural (from hexoses). Although in furfural test or rapid furfural test concentrated hydrochloric acid is used in the place of concentrated sulfuric acid followed by boiling of solution.

The same procedure as the Molisch test is applied. Initially to the dilute solution of sugar, ethanolic naphthol is added followed by slow addition of concentrated hydrochloric acid. Finally, the solution is boiled. In case of fructose or sucrose, a violet color is immediately obtained when the solution starts boiling but in case of glucose the appearance of color is delayed and may not be produced before one minute boiling.

Uses of Molisch test

The Molisch test is used to determine the presence of carbohydrates in different samples.
Molisch test can also be used to distinguish between byproducts of reaction which form carbohydrates as byproducts.

Limitation of Molisch test

Sugars like trioses and tetroses do not contain five carbon atoms which are required for formation of furfural which is a five membered ring compound. And thus does not give a positive Molisch test result.
Some other organic substances which can form furfurals like oxalic acid, citric acid, lactic acid, formic acid etc., can also give a positive Molisch test result which means the Molisch test is not specific for carbohydrates.

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NCERT Chemistry Notes:

Frequently Asked Questions (FAQs)

1. What is the Molisch test?

Molisch test is a chemical test performed to detect carbohydrates in a given mixture or solution. The carbohydrates can be bound to lipids or proteins or can be in Free State. It can also be used to determine and distinguish between compounds like glycolipids which can be hydrolyzed into monosaccharides. Concentrated sulfuric acid is used as a catalyst to hydrolyze polysaccharide into monosaccharide and for the formation of furfural.

2. What is a furfural test?

Furfural test; also known as rapid furfural test, is also used to detect sugars by using concentrated hydrochloric acid which helps catalyzing the dehydration of sugars (carbohydrates) to produce furfural (from pentose) or hydroxymethylfurfural (from hexoses).

3. State principle of Molisch test.

The Molisch test is based on the principle that concentrated acid helps catalyze the dehydration of sugars (carbohydrates) to produce furfural (from pentose) or hydroxymethylfurfural (from hexoses).

4. What is Molisch reagent?

Molisch’s reagent is a solution mixture of phenol dissolved in ethanol. This mixture should always be prepared fresh.

5. Write steps for Molisch test reagent preparation.

To prepare Molisch reagent, 3.75g of alpha naphthol reagent is dissolved in a 25 ml solution of 99% ethanol. The mixture must always be prepared fresh.

6. What precautions must be taken in performing the Molisch test?

Following precautions must be taken in performing Molisch test:

Since the test involves usage of concentrated acids, special care must be taken while handling concentrated acids.
To ensure formation of a reddish violet ring at the junction of the test tube, the test tube must stand still on unmovable test tube holders.
The concentrated acid must be poured slowly to prevent charring of sugars.
Four to five test tubes must be prepared for accurate results.
Distilled water must be used to decrease the chances of impurities interacting with reagent or concentrated acid. Impurities can lead to the formation of green rings which can mislead inference.

7. Can Molisch's test detect non-sugar carbohydrates like cellulose or starch?

Yes, Molisch's test can detect non-sugar carbohydrates like cellulose and starch. These polysaccharides are composed of sugar units that can be dehydrated to form furfural derivatives, which then react with α-naphthol to produce the characteristic purple color.

8. What is the chemical name of the purple compound formed in Molisch's test?

The exact chemical name of the purple compound formed in Molisch's test is not well-established. It is generally referred to as a condensation product between furfural derivatives and α-naphthol.

9. What is the chemical equation for the formation of furfural in Molisch's test?

The general equation for furfural formation from pentoses (5-carbon sugars) in Molisch's test can be represented as:

10. What is the difference between a positive Molisch's test and a false positive result?

A positive Molisch's test is characterized by the formation of a distinct purple ring at the interface of the test solution and sulfuric acid layer, indicating the presence of carbohydrates. A false positive result may occur when non-carbohydrate compounds produce a similar color change. To distinguish between them, additional confirmatory tests or controls may be necessary.

11. Can Molisch's test be used to detect carbohydrates in mixtures containing proteins or lipids?

Yes, Molisch's test can detect carbohydrates in mixtures containing proteins or lipids. However, the presence of these other biomolecules may interfere with the test or make the results more difficult to interpret. In complex mixtures, it may be necessary to isolate or extract the carbohydrate fraction before testing.

12. Can Molisch's test detect all types of carbohydrates?

Yes, Molisch's test can detect all types of carbohydrates, including monosaccharides, disaccharides, and polysaccharides. This is because all carbohydrates contain the basic sugar structure that can undergo dehydration and form furfural derivatives.

13. Can Molisch's test distinguish between different types of carbohydrates?

No, Molisch's test cannot distinguish between different types of carbohydrates. It is a general test that detects the presence of carbohydrates but does not provide information about the specific type of carbohydrate present in the sample.

14. Is Molisch's test specific to carbohydrates only?

While Molisch's test is primarily used for carbohydrates, it can also give positive results with some other organic compounds that can form furfural derivatives under strong acidic conditions. However, it is most commonly used and reliable for carbohydrate detection.

15. What is the sensitivity of Molisch's test?

Molisch's test is quite sensitive and can detect carbohydrates in very small quantities, typically in the range of micrograms. However, the exact sensitivity can vary depending on the specific carbohydrate and test conditions.

16. Can Molisch's test be used to quantify carbohydrates?

No, Molisch's test is a qualitative test and cannot be used to quantify carbohydrates. It only indicates the presence or absence of carbohydrates in a sample, not their concentration.

17. What reagents are used in Molisch's test?

The two main reagents used in Molisch's test are:

18. Why is α-naphthol used in Molisch's test?

α-Naphthol is used in Molisch's test because it reacts with the furfural derivatives formed from carbohydrates to produce a purple-colored complex. This color change is the key indicator of a positive result in the test.

19. What is the role of concentrated sulfuric acid in Molisch's test?

Concentrated sulfuric acid serves two purposes in Molisch's test:

20. How is Molisch's test performed?

Molisch's test is performed by following these steps:

21. What does a positive result in Molisch's test look like?

A positive result in Molisch's test is indicated by the formation of a purple-colored ring at the interface between the test solution and the sulfuric acid layer. This ring appears within a few minutes of adding the acid.

22. What is the principle behind Molisch's test?

The principle behind Molisch's test is the dehydration of carbohydrates by concentrated sulfuric acid, which leads to the formation of furfural or hydroxymethylfurfural. These compounds then react with α-naphthol to produce a purple-colored complex at the interface of the two liquids.

23. What is Molisch's test?

Molisch's test is a qualitative chemical test used to detect the presence of carbohydrates in a sample. It is based on the reaction between carbohydrates and α-naphthol in the presence of concentrated sulfuric acid, resulting in the formation of a purple-colored compound at the interface of the two liquids.

24. How does Molisch's test relate to the structure of carbohydrates?

Molisch's test relates to the structure of carbohydrates by exploiting their ability to form furfural derivatives under acidic conditions. This is possible because carbohydrates contain multiple hydroxyl groups and a carbonyl group, which undergo dehydration reactions in the presence of strong acids.

25. Why is it important to add sulfuric acid carefully down the side of the test tube?

Sulfuric acid should be added carefully down the side of the test tube to form a distinct layer beneath the test solution. This layering is crucial for observing the purple ring formation at the interface, which is the key indicator of a positive result.

26. How does the concentration of carbohydrates affect the intensity of the purple ring in Molisch's test?

The concentration of carbohydrates generally correlates with the intensity of the purple ring in Molisch's test. Higher concentrations of carbohydrates typically result in a more intense and broader purple ring. However, this relationship is not strictly linear and should not be used for quantitative analysis.

27. Can Molisch's test be used to detect carbohydrates in biological samples?

Yes, Molisch's test can be used to detect carbohydrates in various biological samples, including plant extracts, food products, and some bodily fluids. However, it's important to consider potential interfering substances in complex biological matrices.

28. How does Molisch's test differ from other carbohydrate tests like Benedict's or Fehling's test?

Molisch's test differs from Benedict's and Fehling's tests in several ways:

29. Can Molisch's test be used to detect carbohydrates in solid samples?

Yes, Molisch's test can be used to detect carbohydrates in solid samples. The solid sample should first be dissolved in water or an appropriate solvent before performing the test. If the sample is insoluble, it may need to be finely ground or extracted before testing.

30. What safety precautions should be taken when performing Molisch's test?

When performing Molisch's test, the following safety precautions should be taken:

31. Can Molisch's test be used to detect artificial sweeteners?

No, Molisch's test cannot detect artificial sweeteners. It is specific to carbohydrates and compounds that can form furfural derivatives. Most artificial sweeteners do not have the chemical structure required to produce a positive result in this test.

32. Who developed Molisch's test?

Molisch's test was developed by Austrian botanist Hans Molisch in the late 19th century. He discovered this method while studying plant physiology and carbohydrate chemistry.

33. What are some limitations of Molisch's test?

Some limitations of Molisch's test include:

34. What is the chemical structure of α-naphthol used in Molisch's reagent?

α-Naphthol is an aromatic organic compound with the chemical formula C10H8O. Its structure consists of two fused benzene rings with a hydroxyl (-OH) group attached to one of the rings at the alpha position.

35. How does temperature affect Molisch's test?

Temperature can affect the rate and intensity of the color formation in Molisch's test. Higher temperatures generally accelerate the reaction, but extreme heat should be avoided as it can lead to charring of the sample. Room temperature is typically sufficient for the test.

36. What is the shelf life of Molisch's reagent?

The shelf life of Molisch's reagent (α-naphthol in ethanol) is typically several months when stored properly in a dark, cool place. However, it's best to check the reagent's effectiveness periodically and prepare fresh solutions if needed to ensure reliable results.

37. How does the structure of furfural contribute to the color formation in Molisch's test?

Furfural, formed from the dehydration of carbohydrates, has a reactive aldehyde group and an aromatic ring. This structure allows it to undergo condensation reactions with α-naphthol, forming conjugated systems that absorb light in the visible spectrum, resulting in the characteristic purple color.

38. Can Molisch's test be used to detect carbohydrates in plant cell walls?

Yes, Molisch's test can detect carbohydrates in plant cell walls. Plant cell walls are primarily composed of cellulose, hemicellulose, and pectin, all of which are carbohydrates that will give a positive result in the test. However, the cell wall may need to be broken down or extracted for effective testing.

39. How does the presence of other organic compounds affect Molisch's test results?

The presence of other organic compounds can potentially interfere with Molisch's test results. Some compounds may produce similar color changes or mask the purple ring formation. Others might react with the reagents or alter the pH, affecting the test's sensitivity or specificity. It's important to consider potential interferents when interpreting results.

40. What is the role of ethanol in Molisch's reagent?

Ethanol in Molisch's reagent serves as a solvent for α-naphthol. It helps to dissolve and stabilize α-naphthol, ensuring its even distribution in the test solution. Ethanol also helps to mix the reagent with the aqueous test sample before the addition of sulfuric acid.

41. Can Molisch's test be used to detect carbohydrates in forensic science?

Yes, Molisch's test can be used in forensic science to detect carbohydrates in various samples. It might be employed to analyze trace evidence, such as residues from foods or plant materials. However, due to its non-specific nature, it would typically be used in conjunction with other more specific tests in forensic applications.

42. How does the molecular weight of carbohydrates affect their detection by Molisch's test?

Molisch's test can detect carbohydrates of various molecular weights, from simple sugars to complex polysaccharides. However, larger molecules like starch or cellulose may react more slowly or require more time for the color to develop fully. The test's sensitivity might also vary slightly with molecular weight, but it generally remains effective across a wide range.

43. What is the environmental impact of chemicals used in Molisch's test?

The chemicals used in Molisch's test, particularly concentrated sulfuric acid, can have significant environmental impacts if not disposed of properly. Sulfuric acid is corrosive and can harm aquatic life if released into water systems. Proper neutralization and disposal methods should be followed to minimize environmental risks.

44. Can Molisch's test be adapted for use in field testing or portable kits?

While Molisch's test is traditionally performed in a laboratory setting, it could potentially be adapted for field testing or portable kits. However, the use of concentrated sulfuric acid presents safety challenges. Modified versions using less hazardous reagents or microfluidic devices might be developed for portable applications, but these would require validation against the standard method.

45. How does Molisch's test compare to instrumental methods for carbohydrate detection?

Molisch's test is a simple, rapid, and cost-effective method for detecting carbohydrates, but it lacks the specificity and quantitative capabilities of instrumental methods. Techniques like high-performance liquid chromatography (HPLC) or mass spectrometry can provide detailed information about the types and quantities of carbohydrates present. Molisch's test is often used as an initial screening tool before more advanced analyses.

46. Can Molisch's test be used to detect carbohydrates in archaeological samples?

Yes, Molisch's test can be used to detect carbohydrates in archaeological samples, such as residues on ancient pottery or preserved organic materials. However, the test's effectiveness may be limited by the degradation of carbohydrates over time and the presence of interfering substances. In archaeological applications, it would typically be used in conjunction with other analytical techniques.

47. How does the pH of the test solution affect Molisch's test results?

The pH of the test solution generally does not significantly affect Molisch's test results because the addition of concentrated sulfuric acid overwhelms any initial pH differences. However, extremely basic solutions might neutralize some of the acid, potentially affecting the reaction. It's best to start with a neutral or slightly acidic solution for optimal results.

48. Can Molisch's test be used to detect carbohydrates in dairy products?

Yes, Molisch's test can be used to detect carbohydrates in dairy products, primarily lactose. However, the presence of proteins and fats in dairy products may interfere with the test or make the results more difficult to interpret. Sample preparation techniques, such as protein precipitation or fat extraction, might be necessary for clear results.

49. What is the difference between Molisch's test and the anthrone test for carbohydrates?

Both Molisch's test and the anthrone test are used to detect carbohydrates, but they differ in several ways:

50. Can Molisch's test be used to detect carbohydrates in urine samples?

Yes, Molisch's test can be used to detect carbohydrates in urine samples. It's sometimes used as a preliminary screening test for diabetes, as elevated glucose levels in urine will give a positive result. However, more specific and quantitative tests are typically used for clinical diagnosis and monitoring of diabetes.

51. How does the structure of α-naphthol contribute to its role in Molisch's test?

The structure of α-naphthol is crucial to its role in Molisch's test: