March 23, 2025 | UR Gate
Seliwanoff's Test: Principle, Procedure, Reaction Mechanism, and Result
Seliwanoff test, Seliwanoff reagent, sugar detection, fructose test,
ketose sugar reaction, monosaccharide detection, resorcinol reaction,
difference between glucose and fructose, sugar detection in foods,
chemical analysis of sugars, furfural reaction, sugar testing in the
laboratory, comparison of sugar tests, common errors in Seliwanoff test,
applications of Seliwanoff test
Theoretical Principle
Seliwanoff's test is used to distinguish between ketose and aldose
monosaccharides. Ketose sugars, such as fructose, give a positive result,
while aldose sugars do not. When fructose reacts with hydrochloric acid, it
forms 5-hydroxymethylfurfural, which condenses with resorcinol to produce a
cherry-red complex. This test is commonly used to differentiate between
fructose and glucose.
Note: Heating for more than 30 seconds may yield a false positive with
aldose sugars due to their conversion into ketose sugars by hydrochloric
acid and heat. Sucrose also gives a positive result if heated for longer
than 30 seconds because it hydrolyzes into glucose and fructose.
Importance and Applications:
- In Biochemistry: Used to distinguish between ketose sugars (e.g., fructose) and aldose sugars (e.g., glucose).
- In the Food Industry: Helps detect the presence of sugars in food products such as beverages and juices, particularly identifying fructose or hydrolyzed sucrose.
- In Clinical Analysis: Applied in laboratory tests to detect sugars in biological fluids like urine, especially in cases of metabolic disorders.
- In Chemical Research: Utilized as a qualitative test to identify the presence of ketose sugars in various samples.
Chemical Reaction Mechanism:
- Acidic Medium Preparation: The sugar sample is heated with hydrochloric acid (HCl), leading to the hydrolysis of disaccharides into monosaccharides (if present).
- Dehydration of Fructose: In the acidic environment, fructose (a ketose sugar) loses water molecules and forms 5-hydroxymethylfurfural.
- Condensation Reaction: 5-hydroxymethylfurfural reacts with resorcinol (a phenolic compound in Seliwanoff's reagent), forming a cherry-red complex.
- Sugar Differentiation: Ketose sugars (e.g., fructose) produce a rapid and intense red color, while aldose sugars (e.g., glucose) yield a slower and weaker reaction.
Reaction Result:
- Positive Result: A rapid formation of a cherry-red color indicates the presence of fructose or fructose-containing sugars (e.g., sucrose after hydrolysis).
- Negative Result: Aldose sugars like glucose give a slower reaction and a faint or absent red color.
Materials Used:
- Micropipette
- Timer
- Balance
- Cylinder
- Test tube
- Water bath
- Beaker
- Heater
- Dropper
Chemicals Used:
- Resorcinol (C6H6O2) - 0.5 g
- Hydrochloric acid (HCl) - 3M
- Fructose (C6H12O6)
- Water (H2O)
Purpose of Seliwanoff's Test:
- To distinguish between aldose sugars (e.g., glucose) and ketose sugars (e.g., fructose) based on the speed of red color formation.
- It is specific for ketoses.
- Ketose sugars produce a rapid red color, while aldose sugars respond more slowly.
Preparation of Seliwanoff's Reagent:
- Dissolve 0.5 g of resorcinol in 1 liter of 3M hydrochloric acid.
Procedure:
- Add 1.5 mL of Seliwanoff's reagent (resorcinol and diluted hydrochloric acid) to a clean, dry test tube.
- Add 1.5 mL of the sugar solution to be tested to the tube.
- Place the test tube in a boiling water bath for 30 seconds.
- Observe the color change: a cherry-red or brownish-red color indicates the presence of ketose sugars (e.g., fructose or sucrose after hydrolysis).
Discussion:
Why do aldose sugars not give a positive Seliwanoff's test?
Aldoses like glucose do not form furfural derivatives under the same conditions (12% HCl) because they are less prone to dehydration compared to ketoses like fructose.
When do aldose sugars give a positive result?
Prolonged heating (beyond 30 seconds) converts aldoses into ketoses, eventually yielding a positive red color.
How does sucrose give a positive Seliwanoff's test?
When heated for more than 30 seconds, sucrose hydrolyzes into glucose and fructose in the presence of HCl. The released fructose reacts positively with Seliwanoff's reagent.
How is Seliwanoff's reagent prepared?
Dissolve 0.5 g of resorcinol in 1 liter of 3M hydrochloric acid.
What happens when Seliwanoff's reagent is added to a solution containing ketoses and aldoses?
Ketoses rapidly produce a red color, indicating a positive test, while aldoses produce a faint pinkish solution.
Why do fructose and sucrose give a positive Seliwanoff's test?
Sucrose gives a positive result because it hydrolyzes into glucose and fructose, and the fructose reacts with the reagent.
What is the principle of the Seliwanoff's reaction?
It is similar to Molisch's reaction. Fructose or glucose reacts with hydrochloric acid (3N or 12%) to form hydroxymethylfurfural, which then reacts with resorcinol to form a red complex.
Explain why the color spread throughout the test tube instead of forming a ring, as seen in the Molisch test?
◼︎ In Seliwanoff's test:
- The chemical reaction occurs between 5-hydroxymethylfurfural (formed from ketoses) and resorcinol in a homogeneous medium.
- When the sample is heated, the formed furfural disperses evenly throughout the solution, causing the red color to spread across the entire test tube.
◼︎ In contrast, during the Molisch test:
- Concentrated sulfuric acid is added carefully to prevent mixing with the sugar solution, forming distinct layers.
- The reaction occurs at the interface between the layers, producing a purple ring at the boundary where the substances meet.
◼︎ Key difference:
- Seliwanoff's test occurs in a homogeneous medium, resulting in the color spreading throughout the tube.
- Molisch test occurs in a heterogeneous medium due to layer separation, causing the formation of a purple ring at the point of contact.
Comparison with Other Sugar Detection Tests
Seliwanoff's Test, Molisch's Test, Benedict's Test, Barfoed's Test, Fehling's Test
Common Errors and How to Avoid Them:
- Use a water bath at the appropriate temperature instead of direct heating, as excessive heat can decompose other sugars and give false positives.
- Ensure accurate measurement of HCl concentration to prevent side reactions.
- Use freshly prepared Seliwanoff's reagent and store it properly to maintain accuracy.
- Allow sufficient reaction time for clear color observation, as some samples may react slowly.
Conclusion:
- Seliwanoff's test is a qualitative method for detecting ketose sugars, especially fructose, through the formation of a cherry-red complex with resorcinol.
- The test provides a quick and relatively accurate distinction between fructose and glucose but does not quantify sugar concentration.
- This method is valuable in biochemistry, food industry, and clinical diagnostics.
- Proper handling and avoiding common errors ensure reliable results.