I'm extracting phytochemicals from different plant parts by using different solvents (both polar and non-polar). To some extent through literature I know a particular plant has some compounds. All compounds can't be extracted with one solvent so I used different solvents and I want to know the compounds in every solvents as it is showing antibacterial activity. Is there any general procedure to know them by any techniques like HPLC, GC, NMR etc. I'm not satisfied with the qualitative phytochemical analysis.
I've done similar studies in the past. To find the antibacterial or antifungal compounds, we first used TLC to separate the components, and use a number of sprays and UV wavelengths to help with a partial identification. The TLC plate can also be overlayed with hot agar and spread fungi or bacteria on the agar. Zones of inhibition (no growth) could be seen around spots on the TLC plate (previously visualized by UV or other means). But to your question of identifying the compounds, I prefer to use LC-MS with a ToF mass spec to be able to determine the molecular formula from the mass. From the formula, using searches on ChemSpider, google, etc, reasonable structures can be obtained many times. If you already used a silica TLC, then silica HPLC will work fine, but C18 columns usually work better with MS. And if the compound is volatile enough for GC, GC-MS, with its Wiley/NIST database, is great for identifying compounds. Depending on the amount of pure compound you can isolate, NMR is the ultimate method of identification. I hope this helps.
If I want to process my sample to LC-MS, I need to give the technicians conditions of running the samples. Is there any condition which can be used for any kind of plant extracts.
Uma- for such a situation, I use a technique I term "Wide Polarity Range Chromatography", which is essentially chained gradients. Although the examples I list below are for flash chromatography, they should work well for HPLC for silica bonded phase media. Since the functional group is chemically bound, it won't be washed off by changing solvents. This lets one evaluate compounds that elute in a wide variety of solvents.
FLASH with MS detection (see the section headed with Purification of Curcuma longa extract) :
http://www.isco.com/WebProductFiles/Applications/101/Poster_and_Paper_Reprints/Purification%20of%20Natural%20Products%20Using%20Mass%20Directed%20Fractionation.pdf
Diol column (see example 1):
http://www.isco.com/WebProductFiles/Applications/101/Poster_and_Paper_Reprints/All_Wavelength_Detection_MPLC_for_Natural_Products.pdf
C18 (Don't try with polymeric reverse phase, since this material swells in organic solvent):
http://www.isco.com/WebProductFiles/Applications/101/Application_Notes/AN77_Wide_Polarity_Range_Chromatography.pdf
Technique overview:
http://www.isco.com/WebProductFiles/Applications/101/Poster_and_Paper_Reprints/Wide%20Polarity%20Range%20Flash%20Purifications.pdf
Thank you Mr. Jack Silver. I will refer your papers. Hoping to find a clue.Thanks once again.
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