Is there any objective test which is able to discriminate the presence of chlorophyll from its precursors (chlorophyllides)?
Let's say you have a liquid alcoholic solution from young just-germinated embryos from seeds. The solution has a greenish colour whose origin could be both chlorophyll or any chlorophyllide, i.e. chlorophyll precursor.
How can you be sure about chlorophyll? I have tried photo-emission tests but the spectral difference is so small that indeed it is hard to discriminate between the two. Any suggestion?
Hi Eugenio
I think HPLC would be the best, because in addition to the discrimination of the spectra, there will also be variations in the separation time
Hi Eugenio
I sent your question to a PhD student in our lab working on such questions (Mathieu Lauffer). I gave him your email adress in order to answer to you.
All the best
Delphine
Dear Eugenio,,
Please send a mail to Prof. Govindjee. He will address it the best. His mail address is
You may download a book written by him along with Prof. Robinowitch from Prof. Govindjee's website.
I believe that chromatographic techniques including HPLC can help in seperating them.
I agree that HPLC or any chromatographyc method is the most suitable method to separate and distinguish between chlorophyllides and their esters.
If you don't have the possibility to run chromatography, you can try phase separation method. You should extract the pigments with 80% acetone, this will contain both types of the pigments. Then add petroleum ether (with boiling point 40-70 degree is the best) of known volume and water. Shake it and add small amount of solid sodium chloride what helps the phase separation. Add water until the volume of the petroleum ether will be the same as was the initial. This phase will sit on the top and contains all chlorophylls and carotenoids. The bottom water-acetone mixture will contain the chlorophyllides. You can collect the top phase and repeat the phase separation until the petrolic ether phase will be empty (will not show any fluorescence signal). You can test it with the spectrofluorometer you mentioned you have. You can collect the chlorophyllides from the watery phase with diethyl ether via similar phase separation as described above.
- Badges
- Science method