Iodine Test- Definition, Principle, Procedure, Result, Uses

The iodine test is a simple and widely used chemical test to detect the presence of certain types of carbohydrates in a sample. The test is based on the reaction of iodine with starch, a polysaccharide composed of long chains of glucose units. Starch forms a blue-black or purple complex with iodine, which can be easily observed by the naked eye. Other polysaccharides, such as dextrin and glycogen, also react with iodine but produce different colors. Monosaccharides and disaccharides, such as glucose and sucrose, do not react with iodine and remain colorless.

The iodine test is a simple and quick method to detect the presence of certain polysaccharides in a given sample. The main objective of this test is to identify starch, which is a common storage carbohydrate in plants and animals. Starch consists of two types of glucose polymers: amylose and amylopectin. Amylose forms a linear chain of glucose units, while amylopectin forms a branched chain of glucose units. Both amylose and amylopectin can form helical structures that can bind to iodine molecules and produce characteristic colors.

Another objective of the iodine test is to demonstrate the process of photosynthesis in plants. Photosynthesis is the process by which plants convert light energy into chemical energy and store it as glucose. Glucose is then polymerized into starch and stored in various parts of the plant, such as leaves, stems, roots, seeds, and fruits. By performing the iodine test on different parts of a plant, one can observe where starch is accumulated and how it varies with different environmental factors, such as light intensity, duration, and wavelength.

The iodine test can also be used to detect other polysaccharides that have similar helical structures as starch, such as dextrin and glycogen. Dextrin is a degradation product of starch that is formed by partial hydrolysis or enzymatic breakdown. Glycogen is the main storage carbohydrate in animals and fungi, which has a more branched structure than amylopectin. Both dextrin and glycogen can react with iodine and produce different colors than starch.

The iodine test is not specific for starch or other polysaccharides, as it depends on the shape and length of the glucose chains rather than their chemical composition. Therefore, this test cannot be used to quantify the amount of starch or other polysaccharides in a sample, nor can it distinguish between different types of starch or other polysaccharides. For these purposes, more sophisticated methods are required, such as spectrophotometry, chromatography, or enzymatic assays.

The iodine test is based on the fact that polyiodide ions form colored adsorption complex with helical chains of glucose residue of certain polysaccharides. The color of the complex depends on the type and length of the polysaccharide.

The reagent used in the iodine test is Lugol’s iodine, which is an aqueous solution of elemental iodine and potassium iodide. Iodine on its own is insoluble in water. Addition of potassium iodide results in a reversible reaction of the iodine ion with iodine to form a triiodide ion, which further reacts with an iodine molecule to form a pentaiodide ion.

Bench iodine solution appears brown, whereas, the iodide, triiodide, and pentaiodide ion are colorless. It is observed that the helix (coil or spring) structure of the glucose chain is the key to this test. Further, the resulting color depends on the length of the glucose chains.

The triiodide and pentaiodide ions formed are linear and slip inside the helix structure. It is believed that the transfer of charge between the helix and the polyiodide ions results in changes in the spacing of the energy levels, which can absorb visible light, giving the complex its color.

The intensity of the color decreases with the increase in temperature and the presence of water-miscible organic compounds like ethanol. On heating, the blue color amylase-iodine complex dissociates but is formed again on cooling because the helical structure is disrupted; thereby amylose loses its iodine binding capacity and the blue color. The blue color reappears on cooling due to the recovery of iodine binding capacity due to regaining of the helical structure.

The following table summarizes the colors produced by different polysaccharides with Lugol’s iodine:

Monosaccharides, disaccharides, and branched polysaccharides like cellulose remain colorless with Lugol’s iodine as they do not have a helical structure that can accommodate polyiodide ions.

To perform the iodine test, you will need the following:

• Reagent: Lugol`s iodine, which is an aqueous solution of elemental iodine and potassium iodide. You can prepare it by mixing 5% elemental iodine with 10% potassium iodide in distilled water. Alternatively, you can buy it from a chemical supplier or a pharmacy.
• Test sample: The sample that you want to test for the presence of starch. It can be a solid or a liquid substance, such as food, plant material, saliva, etc. If the sample is solid, you will need to grind it or dissolve it in water before testing.
• Materials required: Test tubes, test tube stand, pipette or dropper, spatula or spoon.
• Equipment: Water bath, thermometer.

That`s all you need to perform the iodine test. Make sure you wear gloves and goggles when handling the reagent and the sample, as they may be irritant or harmful to your skin and eyes. Also, work in a well-ventilated area and dispose of the waste properly after the test.😊

The iodine test can be performed in a few simple steps:

1. Take 1 ml of the sample to be tested in a clean, dry test tube. Label it as "test".
2. Take another test tube and add 1 ml of distilled water to it. Label it as "control".
3. Add 2-3 drops of Lugol`s iodine solution to both the test tubes and mix them well by shaking or vortexing.
4. Observe the color change in the test tubes. A blue-black or purple color indicates a positive result for starch, while no color change indicates a negative result.
5. To confirm the result, heat the test tubes in a water bath until the color disappears. This means that the starch-iodine complex has broken down due to high temperature.
6. Remove the test tubes from the water bath and let them cool down. Observe the color change again. The reappearance of the blue-black or purple color indicates that the starch-iodine complex has reformed due to low temperature.

The procedure of iodine test is illustrated in the following table:

The result of the iodine test depends on the appearance of color in the test tubes after adding Lugol`s solution and heating.

• The appearance of blue-black or purple color represents a positive test, indicating the presence of starch.
• If there is no change in color, the result is negative and indicates the absence of starch.
• The appearance of black color indicates the presence of dextrin.
• The appearance of reddish-brown color indicates the presence of glycogen.
• The appearance of orange-yellow color indicates the presence of amylopectin.

The intensity of the color may vary depending on the concentration and length of the glucose chains in the sample. The color may also fade or disappear with increasing temperature or the presence of organic solvents. However, the color may reappear on cooling as the helical structure of the glucose chains is restored.

The iodine test can also be used to measure the iodine levels in serum or urine samples. In this case, the result is expressed in units of micromoles per liter (µmol/L) and interpreted as follows:

• Severe deficiency: 0-0.19 µmol/L
• Moderate deficiency: 0.20-0.38 µmol/L
• Mild deficiency: 0.39-0.78 µmol/L
• Normal: 0.79-1.56 µmol/L
• Slight excess: 1.57-2.36 µmol/L
• Severe excess: Over 2.37 µmol/L

The iodine test is a simple and useful method to detect the presence of starch in various samples. Some of the uses of this test are:

• Food analysis: The iodine test can be used to check the quality and purity of food products by detecting the presence or absence of starch. For example, this test can be used to determine if honey is adulterated with starch syrup, or if flour is contaminated with other substances.
• Medical diagnosis: The iodine test can be used to diagnose certain medical conditions that affect the metabolism of starch. For example, this test can be used to detect glycogen storage diseases, which are inherited disorders that cause abnormal accumulation of glycogen in the body. The iodine test can also be used to monitor the treatment of these diseases by measuring the reduction of glycogen levels in the blood or urine.
• Plant physiology: The iodine test can be used to study the process of photosynthesis in plants. Photosynthesis is the process by which plants convert light energy into chemical energy and store it as starch. The iodine test can be used to demonstrate that starch is produced in the leaves of plants when they are exposed to light, and that starch is consumed when they are kept in the dark. This test can also be used to compare the rate of photosynthesis in different plants or under different environmental conditions.

• The iodine test is a qualitative test and does not indicate the concentration or quantity of starch in the sample. It only shows the presence or absence of starch.
• The iodine test is not specific for starch and can also give positive results with other polysaccharides like dextrin and glycogen. Therefore, it is important to use other tests to confirm the identity of the carbohydrate.
• The iodine test is not reliable for detecting branched polysaccharides like amylopectin and cellulose, which do not form a stable complex with iodine and produce a weak or no color change.
• The iodine test is affected by temperature and solvents. High temperature can disrupt the helical structure of starch and cause the color to fade. Similarly, water-miscible organic solvents like ethanol can interfere with the formation of the complex and reduce the intensity of the color.
• The iodine test is incompatible with acidic conditions, as acid hydrolyzes starch into glucose and maltose, which do not react with iodine. Therefore, the pH of the sample should be neutral or slightly alkaline for the test to work.

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