Safe Removal of Green Pigment for Starch Testing in Leaf Cells

Introduction

To test a leaf for the presence of starch, the primary green pigment, chlorophyll, must be removed safely. This is typically achieved through a process called decolorization, which involves several steps, including boiling in ethanol and rinsing. This article will guide you through the process, highlighting the importance of safety and proper techniques.

Decolorization Process - Boiling in Ethanol

The first step in the decolorization process is to boil the leaf in water to kill it and soften its tissues. Here are the detailed steps:

Place the leaf in boiling water for about 2-3 minutes. This step is crucial as it not only kills the leaf but also softens the tissues. After boiling, carefully transfer the leaf to a beaker containing 70% ethanol. Heat the beaker gently in a water bath to further aid in the removal of chlorophyll. Ethanol effectively removes the chlorophyll, turning the leaf a pale yellow or whitish color. This step should be performed with caution due to the flammability of ethanol. Rinse the leaf in warm water for a few seconds. This helps soften the leaf further and prepares it for the final starch test.

Testing for Starch

Once the leaf is decolorized, the next step is to test for the presence of starch with an iodine solution:

Immerse the decolorized leaf in an iodine solution (iodine in potassium iodide). If starch is present, the leaf will turn a blue-black color. This confirms that starch is indeed present in the leaf.

Alternative Method - Methylated Spirits and Higher Temperature

An alternative method to the ethanol process involves using methylated spirits and a higher temperature:

Place the leaf in a test-tube containing methylated spirits. This can be a suitable substitute if alcohol is not available. Set the test-tube in a water bath at a temperature above 78°C (the boiling point of ethanol). Keep the test-tube in the water bath until the leaf turns white. This may take longer depending on the leaf's condition. Once the leaf has turned white, remove the test-tube and allow it to cool. Then, remove the leaf and soak it in water to soften it. The alcohol may have dehydrated the leaf, making it brittle. Test the leaf with an iodine solution to confirm the presence of starch. A bluish-black coloration confirms the presence of starch.

Additional Considerations

It's important to note that if parts of the leaf have been covered, such as with aluminum foil, to prevent light exposure, these areas will not undergo photosynthesis. Consequently, the test will be negative in these regions. However, the iodine solution will remain brown, indicating that light is necessary for photosynthesis to occur.

In conclusion, the safe removal of green pigment before starch testing is crucial for accurate results. By following these detailed steps and taking appropriate safety measures, you can ensure reliable and consistent results in your experiments.