I would like to know the changes that happen in chlorophyll as a response to stress.
What level of accuracy you need? You may try a extremely easy procedure by crushing 50 mg of leaves in 10 mL 80% acetone (aqueous). The homogenate is then filtered through a paper filter (standard nitrocellulose filter), and measure the absorption of the filtrate at 645 and 663 nm. You need the Arnon equation, easily found in internet. See for example http://employees.csbsju.edu/SSAUPE/biol327/Lab/photosyn/chlorophyll_quant.htm. GOOD LUCK!!
There are many different procedures... the chloroplast responds in different ways according to the kind of stress (biotic o abiotic). If you want to see chlorophyll content or chlorophyll a/b ratio, you could start with DMF extractions and spectrophotometric analysis. But if you want a very detailed analysis, I recommend you a HPLC procedure.
I agree with Gomez. also see this attachment http://www.ncbi.nlm.nih.gov/pmc/articles/PMC440358/
What level of accuracy you need? You may try a extremely easy procedure by crushing 50 mg of leaves in 10 mL 80% acetone (aqueous). The homogenate is then filtered through a paper filter (standard nitrocellulose filter), and measure the absorption of the filtrate at 645 and 663 nm. You need the Arnon equation, easily found in internet. See for example http://employees.csbsju.edu/SSAUPE/biol327/Lab/photosyn/chlorophyll_quant.htm. GOOD LUCK!!
Hello. I agree with all of you that everything depends on the accuracy level you want to achieve :). For comparative studies the spectrophotometric determination is enough. I often use the protocol similar to this presented by Luis E. Hernandez, but I use the 1:10 ratio (w/v), namely e.g. 500 mg of leaves (FW) per 5 mL of solvent, diluting the sample while measured (if necessary). It diminishes the use of the solvent (80% acetone or 98% ethanol). In the latter case the equations have to be different that these used for acetone-based ones. Please let me know if you want more details.
Another method of estimation the Chl content is to measure so-called greening index using Hansatech or Konica Minolta chlorophyllmeters. This method is non-destructive, so this is the main advantage - you can study an individual leaf as long as you want to... until it is dead :).
Both methods are used in our lab. They may be combined.
Chlorophyll conttent as a reference for studying other leaf or chloroplast or thyalakoid parameters has been my routine once. May see my res publications available here at reserchgate.com
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