hello Sir/Madam
I want to know that what is the suitable technique used to isolation of desire compound from plant extract. I have a plant extract that show my desire biological activity, further my aim is to purify that compound that show biological activity in extract. I have used column for purification. I am successfully some extant purified my desire compound(that fraction have biological activity) but their is other compound impurity. then how to overcome these impurity from my desire compound on a large scale. Please help me.
Try doing TLC, Single spot in TLC indicates purity of the compound. If your compound is impure you will get many bands on TLC. After getting the many bands point out which is your desired compound among the bands. Carefully mark with pencil on TLC plate and after that, scratch that band put in eppendorf tube carefully pour some solvent in which it gets dissolved centrifuge take the supernantant after this again run TLC see whether you get single desired band. You can try this method.
Please read Plant drug analysis by Wagner and Bladt.
It depends which kind of compound you are interest in. As you are working with a plant extract you coul use Extraction with Supercritical Fluid. If the molecule target has apolar characteristic, you could use CO2 Supercritical ou CO2 Supercritical with co-solvent, as ethanol, for example. It is even possible to "clean" the plant matrix, concentrating the compounds by proceeding sequencial extraction at different pressures (increasing at each extraction). Pay attention to the temperature ,of the extraction. CO2 supercritical is available above 32-33°C but it could happen decomposition of the biological actives molecules if the temperature is higher than, for example, 50°C.
Hi, in addition of the nice comments above, I will share what we have done. For isolating the possible active compounds from extracts, we did generally LC hr ms ms of our extracts (maybe many fractions), than we did also bio activities studies of the (fractions) extracts. By combining those data using MVA (PCA, PLS-DA) we can predict the active compound (peak) using S plot or loading plot of the PCA. After these , we apply the system of LC MS, to analytical/preparative HPLC to isolate the active compound(s) only. If you do not have LC MS, you can do it with TLC (Densitomety) as well, kind regards
Generally it depends on the desired compounds. The following link may be helpful to you in making a decision.
Article General Extraction, Isolation and Characterization Technique...
Kind regards
You will need to prepare your samples through extraction then purify using chromatographic techniques. HPLC is one of the best there are. You can use TLC to check the purity of your isolated fractions from time to time
Thank you to all for your informative suggestions. But my problem is continue, because i have tried TLC for separation single band show. In GC-MS their is also a higher single peak with some minor peaks. But when i go for LC-MS there is about 13 ms peaks. I am very disappoint for this. what am i doing for isolating my desire compound from these 13 peaks of MS and how to justify which one is the compound that have biological activity.
Really depend upon the plant source and compounds you want to extract. My suggestion to you is to use several solvents with different polarity to obtain most if not all compounds in the sample. Later, you can check the extracts using thin layer chromatography. Make sure you select the best eluent so you can get good separation of the compounds...hope it helps...
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