The Iodine Test: What Is the Test for Starch and Glycogen?

December 11, 2025 •

4 min read

The iodine test, a classic biochemical technique, is the standard method for identifying certain polysaccharides, with a blue-black color change confirming the presence of starch. But what is the test for starch and glycogen, and how can a simple solution differentiate between them? This guide will detail the procedure and the fascinating chemical principles behind this colorimetric reaction, essential for any biology or chemistry student.

Quick Summary

The iodine test distinguishes between starch and glycogen by reacting with their unique helical structures. A deep blue-black color indicates starch, while a reddish-brown color signals the presence of glycogen, providing a clear visual identification.

Key Points

Iodine Test is Definitive: The iodine test is the primary chemical method to identify starch and glycogen, distinguishing them by the distinct colors produced upon reaction.
Starch Turns Blue-Black: When tested with Lugol's iodine, starch produces an intense deep blue-black color, a result of the amylose component trapping polyiodide ions in its helical structure.
Glycogen Turns Reddish-Brown: In contrast, glycogen reacts with iodine to produce a reddish-brown or amber color because its highly branched structure does not form the same long helical complexes as starch.
Principle Depends on Helical Structure: The test's mechanism relies on the ability of polyiodide ions to be entrapped within the coiled glucose chains of polysaccharides, with the resulting color directly tied to the polymer's architecture.
Heating Causes Decolorization: The color from the iodine-polysaccharide complex fades when heated, as the helices uncoil, but the color reappears upon cooling.
Highly Specific to Polysaccharides: Simple sugars like glucose and sucrose do not react with iodine to produce a color change, confirming the test's specificity for certain complex carbohydrates.

The Principle of the Iodine Test

The iodine test relies on the distinct structural differences between polysaccharides. The test uses a solution of iodine (I₂) and potassium iodide (KI) in water, often called Lugol's solution. In this solution, iodine reacts with iodide ions to form polyiodide ions ($I_3^-$ and $I_5^-$), which are linear in shape. When these polyiodide ions are added to a sample containing a suitable polysaccharide, they can be adsorbed into the helical chains of the molecule, forming a colored absorption complex. The resulting color depends directly on the polysaccharide's structure, particularly the length of its unbranched glucose chains.

The Reaction with Starch

Starch, a storage carbohydrate in plants, is composed of two main components: amylose and amylopectin.

Amylose: The linear, unbranched component of starch, consisting of alpha-1,4-glycosidic bonds. Its unbranched chains coil into a helical structure. This spiral shape is crucial for the test.
Amylopectin: The branched component of starch, featuring both alpha-1,4 and alpha-1,6 glycosidic bonds.

When Lugol's solution is added to starch, the linear polyiodide ions from the reagent get trapped inside the amylose helices. This forms a stable starch-iodine complex that causes a shift in the absorption of light, resulting in a characteristic, intense deep blue-black color. The coiled nature of amylose is what allows for the strong color change, effectively trapping the iodine molecules.

The Reaction with Glycogen

Glycogen serves as the energy storage carbohydrate in animals and fungi. It is structurally similar to amylopectin but is far more highly branched. This dense, extensively branched structure prevents the polyiodide ions from forming the long, deep helical complexes that are possible with starch's amylose component. Instead, the shorter, more frequent branches in glycogen can only trap the iodine to a lesser extent. This results in a much weaker interaction, producing a reddish-brown or amber color, which is distinctly different from the blue-black color seen with starch.

The Laboratory Procedure

Performing the iodine test is a straightforward process in a laboratory setting. A positive test is indicated by a color change, while a negative result shows no change from the original amber color of the Lugol's solution.

Materials:

Test tubes or spot plate
Lugol's iodine solution
Test samples (e.g., starch solution, glycogen solution, distilled water as a negative control)
Dropper

Steps:

Label several clean test tubes for each sample and a control.
Add a small amount of each sample into its corresponding test tube.
Add 1 ml of distilled water to the control tube.
Using a dropper, add 2-3 drops of Lugol's iodine solution to each tube.
Swirl or mix gently to combine.
Observe the color change and compare it to the control.
Heat can temporarily break the iodine-polysaccharide complex, causing the color to disappear. Upon cooling, the color will reappear, confirming the presence of the polysaccharide.

Comparison of Starch and Glycogen Iodine Test Results


Feature	Starch	Glycogen
Biological Source	Plants (e.g., potatoes, rice)	Animals and fungi (e.g., liver, muscle)
Structural Characteristics	Less branched; contains linear amylose and branched amylopectin	Highly branched with shorter chains between branches
Interaction with Iodine	Polyiodide ions become trapped within the linear helical coils of amylose.	The high degree of branching prevents the formation of stable, long helical complexes with polyiodide ions.
Color with Iodine	Deep blue-black color	Reddish-brown or amber color
Test Conclusion	Positive for starch	Positive for glycogen

Conclusion

The iodine test remains a reliable, simple, and visually clear method for distinguishing between starch and glycogen. Its principle, based on the unique interactions between polyiodide ions and the distinct helical structures of these two polysaccharides, provides a powerful tool for biochemistry. While starch's linear amylose component forms a deep blue-black complex, the highly branched nature of glycogen results in a reddish-brown color, allowing for easy differentiation in the lab. For further detailed information, scientific journals or resources like the National Institutes of Health (NIH) provide comprehensive overviews of polysaccharide analysis.

A Closer Look at the Chemistry

The chemical foundation of the iodine test lies in the geometry of the glucose polymers. The linear component of starch, amylose, can coil tightly into a spiral shape, creating a cavity just the right size to accommodate the linear triiodide ($I_3^-$) and pentaiodide ($I_5^-$) ions. This entrapment is what leads to the intense color. The charge-transfer interaction between the polyiodide ion and the glucan chain absorbs light in the visible spectrum, producing the characteristic deep blue-black. Glycogen's excessive branching and shorter chain lengths prevent the formation of such long, stable helices, resulting in a less pronounced color change. This difference highlights how a simple reagent can reveal fundamental distinctions in molecular architecture.

Beyond Starch and Glycogen

It is important to remember that the iodine test is highly specific for certain polysaccharides. Monosaccharides (like glucose) and disaccharides (like sucrose) do not exhibit any color change because they lack the helical structure necessary to accommodate the polyiodide ions. Other polysaccharides like cellulose, which has a linear, uncoiled structure due to different glycosidic linkages, also yield a negative result. This specificity makes the iodine test a valuable tool for quickly differentiating between classes of carbohydrates based on their structural complexity.

Limitations of the Iodine Test

While highly effective for its intended purpose, the iodine test has some limitations. It is a qualitative test, meaning it can confirm the presence or absence of a polysaccharide but cannot determine its exact concentration. Factors like temperature can also affect the results; heating the solution will cause the color to fade as the polysaccharide helix uncoils and releases the trapped iodine. The test also becomes invalid under acidic conditions, which can hydrolyze starch. These considerations are important for accurate interpretation in a lab setting.

Frequently Asked Questions

The primary reagent is Lugol's iodine solution, which is an aqueous solution of elemental iodine and potassium iodide.

Starch contains amylose, a linear glucose polymer that coils into a helix. Polyiodide ions from the iodine solution get trapped inside this helix, forming a complex that strongly absorbs light, creating the deep blue-black color.

Glycogen is more highly branched and has shorter glucose chains than starch. This extensive branching prevents the formation of the long, stable helices needed to trap the polyiodide ions, leading to a weaker reaction and a reddish-brown color.

Heating the sample will cause the color to fade or disappear. This is because the heat disrupts the helical structure of the polysaccharide, releasing the trapped iodine. The color will return upon cooling.

No, the iodine test is not effective for simple sugars like glucose or sucrose. These molecules do not have the coiled structure required to interact with the polyiodide ions and will not produce a color change.

Distilled water is typically used as a negative control. When Lugol's iodine is added to distilled water, the solution remains its original amber color, indicating no polysaccharide is present.

The iodine test is a qualitative test, meaning it can only determine the presence or absence of starch or glycogen. It cannot be used to measure the precise amount or concentration of the polysaccharide.