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How to Separate & Identify Chloroplasts in Plant Cells

Chloroplasts, or the photosynthetic components of a plant cell, perform some of the most fascinating biochemical functions in any living tissue. Their task of producing energy for the plant is made possible by several pigments, including chlorophylls, carotenes and xanthophylls. These pigments can be separated by using paper chromatography, forming a distribution that would identify the chloroplast type much like a fingerprint identifies a human. In this particular application, the varied solubility of the different pigments allows the distribution to form.

Things You'll Need

Spinach leaves
Mortar and pestle
Ethyl alcohol or C2H5OH
Chromatography paper
Pencil
Dropper
Large test tube
Acetone ((CH3)2CO)
Tape
Test tube stand

Instructions

- 1
  
  Grinding the leaves helps break apart their cellular structure.
  
  Grind two or three spinach leaves to a fine paste, using the mortar and pestle. Add 10 drops of ethyl alcohol to the mixture to extract the chloroplast pigments. Keep grinding until a few droplets of green tinted liquid appear.
- 2
  Cut the chromatography paper into a strip that will fit into your test tube. Two centimeters up from the bottom of the paper, draw a line in pencil. This serves as the pigments' baseline.
- 3
  Collect some of the ethyl alcohol solution with your dropper, and place one drop in the center of your baseline. Let the drop dry and continue the process until the spot is dyed a rich green.
- 4
  
  Do not jostle the test tube when taking it to the stand.
  
  Fill a test tube with 1.5cm acetone. Place the paper strip into the test tube so the baseline lies just slightly above the acetone, and tape the top of the paper to the outside rim of the test tube. Carefully rest the test tube in its stand.
- 5
  Allow the acetone to filter up the paper until it has almost reached the top of the test tube. Remove the paper strip from the test tube. Mark the furthest extent of the acetone and let the paper dry.
- 6
  Examine the bands of color that should appear on your paper. Each band represents a different pigment, with chlorophyll a appearing blue-green, chlorophyll b a greenish-yellow, xanthophyll a pale yellow, and carotene a bright yellow-orange. Their presence or lack thereof and their relative positions on the chromatography paper are one way of identifying chloroplasts from different species by their pigments.

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