I am trying to establish the effect of priming (using a fungus) to a plant.I want to determine the level of and changes in the amount of salicylic acid as time progresses.
I was not allowed to use liquid nitrogen for extraction due to some technical reasons, so I am hoping for a method which will allow me to extract salicylic acid without liquid nitrogen and quantify it using spectrophotometry.
Ok my dear this is going to be a bit more tricky...
1. Use a mortar with some sand in it to destroy the leave cells. You dont need it to be cool or anything we dont care about the proteis, we just want the small molecules. If you have access to it use a Soxhlet-Extraction tool instead of a mortar or better after crushing it in the mortar. This should allow you to pull almost all the salicylic acid out of the leaves
2. Direct spectrophotometry? I have to pull that tooth right away my dear, you are going to have a variety of small molecules and dye like chlorophyll in your extract. You are not going to directly quantify it. Best possiblity would be to use an HPLC or other wise a HPTLC to seperate the compounds and quantify the salicylic acid....Also keep in mind that they might be glycosylated in the plant so before putting them in an HPLC cook them for an hour under reflux in KOH.
3. Use an internal or extarnal standard of salicylic acid to quantify it via UV-spectroscopy
Mr. Silva,
You can use the following lmax:
- 263 nm (C2H5OH)
- 296 nm (H2O)
- 340 nm (H2O/NaCl)
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