Science Buddies: "Ask an Expert"

My 7th grade daughter decided to do her science fair project this year on gel electrophoresis of leaf pigments. She gathered leaves from trees in which some were green, some had changed to fall colors, and some were a mixture of green spots and fall color spots. She put these in a blender with isopropyl acohol and left them in containers overnight. The next day, she filtered out the leaf bits and used the remaining solution to run her gel electrophoresis.

She had many problems along the way and ran trials through her gel with food coloring to ensure her setup was working properly. After some trial and error, she was able to get the chamber working with food coloring. The samples of food coloring moved and separated into differently colored bands.

When she ran her pigments, they only moved 0.5-1.5 centimeters through the gel and did not separate into any bands.
All of the samples turned brown when they were run through the chamber.

Any ideas on what she should do differently?

She made her gel from a sodium bicarbonate buffer solution and agar agar powder. She used copper wire for her electrodes. She used 5 9-volt batteries linked in series for her power supply. She allowed this to run for 90 minutes.

Any help is greatly appreciated.

Hello!
I'm sorry that this happened and I understand that it can be frustrating after all the troubleshooting and trial runs with the food coloring.

The principle of separation by gel electrophoresis is based on charge and mass. One possible reason why the pigments did not migrate through the gel could be because most plant pigments (e.g. chlorophyll and anthocyanin) are non-polar (i.e. not charged). It's also possible that the extract contained other materials than just pigments so these complex mixtures could have behaved differently from the food coloring. The color change is very likely due to a chemical reaction with some component of the buffer used to run the gel.

If the main goal of the experiment is to separate plant pigments then I would suggest that she try to use chromatography instead of gel electrophoresis. This is a simpler process and will better separate non-polar compounds. Let me know if this is of interest and I can help you to figure out the protocol and set up.

Best wishes,
MS

Hello scarpenter21!

It should be much easier (and more accurate for you) to test leaf pigments using chromatography, and MS15 gave some great advice.

Here is a link to a project and procedure that will guide you through the leaf pigment experiment using chromatography:
https://www.sciencebuddies.org/science- ... #procedure

Just some background information about how chromatography works:
Leaves contain one or more of the 4 types of pigments that are found in leaves. Each of these pigments will travel up the chromatography paper at a different rate because the pigments do not have the same amount of solubility in water: therefore, the molecules in the pigment must dissolve to be small enough to travel up the paper, which occurs at varying rates for each pigment. You will stick the chromatography paper in the water containing the crushed leaves, then wait the directed amount of time for each pigment to travel up the paper. The colors on the chromatography strip will show you which pigment is most abundant in the leaf you are testing, as well as the least abundant, and the different pigments that are also contained in that leaf.

Hope this helps and feel free to ask any additional questions!
-cnoonan180