Molisch Test Procedure: How to Detect Carbohydrates Step-by-Step
The Molisch Test is essential in chemistry and helps students understand the identification of carbohydrates in different chemical and biological samples. Recognizing this test is important for grasping the basics of biochemistry and practical laboratory skills.
What is Molisch Test in Chemistry?
A Molisch Test refers to a qualitative chemical test used to detect the presence of carbohydrates in a given substance. This test comes up when studying topics like qualitative analysis, biochemical tests, and carbohydrate chemistry, making it a foundational part of your chemistry syllabus.
Principle of Molisch Test
The Molisch Test is based on the dehydration of carbohydrates by concentrated sulfuric acid, which produces furfural or hydroxymethylfurfural. These compounds then react with α-naphthol in Molisch reagent to form a purple or violet-colored ring. This purple ring is proof of carbohydrate presence in the sample.
Molisch Test Procedure
The step-by-step procedure of the Molisch Test is as follows:
- Take 2 mL of the test solution in a clean test tube.
- Add 2 drops of freshly prepared Molisch reagent (a solution of α-naphthol in ethanol).
- Carefully add about 1 mL of concentrated sulfuric acid by letting it flow gently along the side of the slanted test tube to form a lower layer.
- Do not mix or shake. Let the test tube stand undisturbed.
- Observe the junction of the two layers for the appearance of a purple or violet-colored ring.
Observation & Positive Result
A positive Molisch Test is confirmed when a purple or violet ring appears at the interface of the acid and sample layers. This color change indicates the presence of carbohydrates. If the colored ring does not form, the test is negative, suggesting no carbohydrates in the sample.
This color change is often called the "purple ring test". It is easy to observe and gives quick results in the chemistry lab.
Step-by-Step Reaction Example
- Dehydration of glucose by concentrated sulfuric acid:
C6H12O6 (glucose) → HMF (hydroxymethylfurfural) + H2O - Formation of colored compound:
HMF + α-naphthol → Violet or purple colored complex (at the interface)
Uses of Molisch Test in Real Life
The Molisch Test is widely used in laboratory analysis of food, plant extracts, pharmaceuticals, and clinical samples to detect carbohydrates. It is also useful for quickly screening if an ingredient or formula labeled "sugar-free" is truly free of carbohydrates. In biology labs, it helps students differentiate between carbohydrates and non-carbohydrate biomolecules.
Some glycoproteins and nucleic acids may also respond to this test because they contain carbohydrate groups in their structure.
Limitations and Precautions of Molisch Test
- Some compounds such as citric acid, lactic acid, and other organic acids may give a faint positive result, leading to confusion.
- Tetroses and trioses (four and three carbon sugars) generally do not give a clear positive Molisch Test, as they do not form furfural easily.
- Always add sulfuric acid slowly along the test tube wall to prevent violent reaction and splashing.
- Do not shake the tube after the purple ring forms, as shaking may cause the ring to disappear.
- Concentrated sulfuric acid is highly corrosive—handle with gloves and eye protection.
Comparison with Other Carbohydrate Tests
| Test Name | Detects | Result/Indicator | Specificity |
|---|---|---|---|
| Molisch Test | All carbohydrates | Purple/violet ring at interface | Universal for carbohydrates |
| Benedict’s Test | Reducing sugars | Brick red precipitate | Specific for reducing sugars |
| Fehling’s Test | Reducing sugars | Brick red precipitate | Only for reducing sugars |
| Barfoed’s Test | Monosaccharides | Red precipitate | Monosaccharides only |
Lab or Experimental Tips
Remember the Molisch Test by the "purple ring" at the acid-layer junction. Vedantu educators often use diagrams in live classes to help students memorize the correct order and safety steps for this experiment. Practice drawing and labeling a test tube with the purple ring for exam success.
Try This Yourself
- Write the name of the colored compound formed in the Molisch Test.
- List two differences between Molisch Test and Benedict’s Test.
- Explain why proteins do not produce a positive Molisch Test.
Final Wrap-Up
We explored the Molisch Test—its principle, procedure, reaction, and real-life importance in identifying carbohydrates. For clear steps, more chemistry topic explanations, and exam-prep help, check live classes and interactive notes on Vedantu.
Explore related topics for a deeper understanding:
Benedict’s Test.
FAQs on Molisch Test: Definition, Principle, Procedure & Result
1. What is the Molisch test?
The Molisch test is a universal biochemical test for detecting the presence of carbohydrates. It involves adding Molisch reagent (α-naphthol) to a sample, followed by carefully adding concentrated sulfuric acid. A purple or violet ring at the interface confirms the presence of carbohydrates.
2. Which reagent is used in the Molisch test?
The main reagent in the Molisch test is Molisch reagent, which contains α-naphthol dissolved in alcohol.
Other chemicals used:
- Concentrated sulfuric acid (added gently to form separate layers)
- Sample solution (containing suspected carbohydrate)
3. What is the principle of the Molisch test?
The Molisch test is based on the dehydration of carbohydrates to form furfural derivatives, which react with α-naphthol to give a colored complex.
- Concentrated sulfuric acid dehydrates any carbohydrate.
- Furfural or hydroxymethylfurfural formed reacts with α-naphthol.
- A violet ring indicates a positive result.
4. What colour indicates a positive Molisch test?
A positive Molisch test is shown by the appearance of a purple or violet ring at the interface of the liquids.
Key points:
- Only at the junction of acid and sample.
- Indicates presence of one or more carbohydrates.
5. Which types of carbohydrates give a positive Molisch test?
All types of carbohydrates—monosaccharides, disaccharides, and polysaccharides—give a positive Molisch test.
- Examples: Glucose, fructose, sucrose, starch, cellulose, glycogen
- The test does not distinguish between different classes.
6. What are the steps of the Molisch test procedure?
To perform the Molisch test:
1. Take 2 mL of the sample solution in a test tube.
2. Add 2-3 drops of Molisch reagent (α-naphthol).
3. Mix gently.
4. Tilt the tube and slowly add 1-2 mL of concentrated sulfuric acid along the side to form a layer.
5. Observe for the formation of a violet ring at the interface.
7. Why does the Molisch test not react with proteins or fats?
The Molisch test is specific to carbohydrates because only carbohydrates undergo acid-induced dehydration to form furfural derivatives, which react with α-naphthol.
- Proteins and fats do not form furfural under test conditions.
- No color change indicates their absence.
8. What are the limitations and precautions of the Molisch test?
Limitations:
- Cannot distinguish between reducing and non-reducing sugars.
- May give false positives with certain glycoproteins or impurities.
Precautions:
- Add concentrated sulfuric acid slowly along the tube's side.
- Perform test in a well-ventilated lab.
- Wear gloves and safety goggles during the procedure.
9. How is Molisch test different from Benedict's and Fehling's tests?
Molisch test detects all carbohydrates, while Benedict's and Fehling's tests specifically detect reducing sugars only.
- Molisch: Universal carbohydrate test
- Benedict’s/Fehling’s: Positive for reducing sugars (not sucrose)
- Results: Molisch (violet ring), Benedict’s/Fehling’s (brick-red precipitate)
10. Can Molisch test be used for glucose identification?
Molisch test will confirm the presence of glucose if present, but it cannot uniquely identify glucose among other carbohydrates.
- All carbohydrates respond positively.
- For specific glucose detection, other tests like glucose oxidase test are used.
11. Why is α-naphthol used as the Molisch reagent?
α-naphthol is used in the Molisch test because it readily reacts with furfural derivatives from carbohydrates to form a sensitive, visible purple complex.
- Enhances color intensity
- Provides specificity for all carbohydrate types
12. What safety measures should be followed during the Molisch test?
Always practice laboratory safety:
- Add concentrated sulfuric acid slowly, along the test tube wall.
- Wear protective gloves and goggles.
- Work under supervision if you are new to handling acids.
