When you have to purify the crude sample using TLC and Column chromatographic methods.
This entirely depends on the chemical nature of the plant compounds you are interested in. Polar, typically non-volatile compounds such as sugars or hydroxy-carboxylic acids are easier to analyse by LC/MS (HPLC coupled to MS). Apolar, volatile compounds such as terpenes are ideally analysed by GC/MS. Of course, one can derivatise polar compounds to make them volatile enough and thus amenable to GC/MS but from own experience this is not necessarily something you want to do after you have spent a lot of time and effort on obtaining a crude yet purified plant extract that (one hopes) contains the compounds one is interested in. Furthermore, you can use analytical TLC to pilot and develop your HPLC method (column and mobile phase) trialling silica as well as reversed-phase TLC plates.
GC - means Gas Cromatograph*; HPLC - means High Pressure Liquid Chromatograph*; MS - means Mass Spectromet*. Gas or Liquid Chormatographs are useful to help one to separate a mix of substances. Identification depends on standards or published retention times. Coupled Mass Spectrometers are useful in identifications of substances based on a fragmentation technique and comparison with published standards that you buy with MS equipment. There are diferent Mass Spectrometers that can be specilized to detected diferent molecular weight ranges. I prefer use GC with volatile substances while (HP)LC can be used with less volatile substances. I have seen people who work with plant samples using both techniques (GC/MS and HPLC/MS). If the substances are not described in your bases you have to do two more analysis usually FTIR and RMN-H to start the job.
This entirely depends on the chemical nature of the plant compounds you are interested in. Polar, typically non-volatile compounds such as sugars or hydroxy-carboxylic acids are easier to analyse by LC/MS (HPLC coupled to MS). Apolar, volatile compounds such as terpenes are ideally analysed by GC/MS. Of course, one can derivatise polar compounds to make them volatile enough and thus amenable to GC/MS but from own experience this is not necessarily something you want to do after you have spent a lot of time and effort on obtaining a crude yet purified plant extract that (one hopes) contains the compounds one is interested in. Furthermore, you can use analytical TLC to pilot and develop your HPLC method (column and mobile phase) trialling silica as well as reversed-phase TLC plates.
In my opinion the usefulness of GC-MS and HPLC-MS is a function of many parameters characterizing the experiments to be monitored by these techniques. One of the most important of these parameters is the number of chemical compounds to be monitored. Of course, as already noted earlier by my predecessor, polarity and volatility of these compounds are very important, too. I believe that for the analysis of complex, multi-component environmental samples much better suited HPLC-MS.
Yes, its simple. For volatile compounds go for GC, GC-MS and for non-volatiles go for HPLC, LC-MS etc.
if you want to identify a specific compound you must begin with preparative HPLC to separate each fraction from your extract individually then GC/MS
Indeed, in the case of complex, multi-component plant samples, as an introduction to the analysis it is necessary separation of the components. In my opinion its does not necessarily have to be done by preparative liquid chromatography. Very good results often gives thin layer chromatography, instead preparative liquid chromatography.
Thanks very much for all your contributions, but I have conducted column chromatoraphy and analytical TLC where I obtained many fractions (28)and the Rf were similar such as 0.93, 0.94, 0.95. My question is should I use GC-MS or HPLC-MS for all the fractions or to pull the fractions with similar Rf together before the Spectrometric analysis.
You can go through HPTLC with desitrometric detection (Camag) with facility for purity of peak detection with spectral homoginity detection. U can further optimized u r mobile phase to separate them or bi-directional HPTLC now HPTLC-MS/MS also available wich much cost-effective and background effect (due to mobile phase) get minimized.
Yes it totally depends on the nature of chemical compounds.
Generally plant metabolites are determined on basis of polarity from non-polar or volatiles to polar non-volatiles.
Mostly for non-polar volatiles GC is preferred while for polar non-volatiles HPLC in preffered. HPLC can be used for polar as well as non polar compouds separation and detection
as refereed by Rana, it depends on the nature of compounds, but generally HPLC has a broad spectrum of stationary phases (gel permeation, ion exhange, adsorption, partition , reverse phases etc) and this can broaden its use in comparison to the GC-MS (only used partition and adsorption stationary phases). in addition, the high temperature in GC makes its use confined stable volatile compounds which form a small number of Natural products.
This condition depends on the investigated chemical compounds. If you are searching volatile components or fatty acids, you should choose GC and GCMS. However, If you want to determine the content of plant extracts, such as vitamins and phenolic compounds, you can prefer HPLC or LCMS.
First, What type of compounds are you looking for? tannins, phenolic acid, flavonols, flavan-3-ols, steroids, terpenes??? Go ahead and do some literature review to see what people have used in your specific plant for the specific group of compounds you are looking for. Then set up that method and tweek it to perfection.
If it is a general screening that you are targeting for, i.e. you want to have an overview on a very broad spectrum of substances, I would not chose either GC-MS OR LC-MS but rather use them both as complementary techniques, because both of them have their strengths and weaknesses. Some substances you will not see in LC-MS but do so in GC-MS and vice versa.
Hello Babatunde. Concur with other people's comments - it depends on what you're interested in. Neither platform is best for separation/detection of the vast array of plant compounds. Both have been used quite effectively, especially with derivatization to either improve detection, stability, or both. It appears you've already done some sample cleanup via columns and TLC. Without knowing the specifics of your extraction/cleanup, I'm guessing your prep is already better suited to HPLC/LC-MS. Another factor for consideration is which platform is in better shape or more expertise at your institution.
HPLC will be more preferable separation technique for identification of chemical components in plant samples. And most importantly it depends upon the nature of the chemical component present in your plant samples. As you have already done TLC and column. So now its better to analyze by HPLC.
you have separated by open column chromatography.But only telling telling Rf wont help. If you have eluted with low polar solvent most probably it might be low polar compound for which you can go for GC. But you take IR of the separated samples .From IR you can make out from the functional groups acidic,carbonyl then decide . If you get polar GO FOR LC-MS.Good luck .Try to take out crystals.
If you have a precise idea of compounds which are likely to be found in your extracts and if you have standards for these compounds - HPLC is a nice technique, you can find the correct column and mobile phase with the standards and then inject your crude extract.
On the contrary, if you do not know what compounds are likely to be found in your extract, it is simplier to start with the GC-MS, its associated library (NIST) will help to identify at least a part of compounds which are composing your extract and then it would be easier to separate and investigate the still unidentified compounds.
There are many different types of plants and many different classes of compounds in plant materials. Are you trying to do a complete characterization of the plant material or are there specific compounds or classes of compounds that you are interested in? In general, LC/MS recommended for non-volatiles and GC/MS for volatiles. Many non-volatile compounds however can be derivaized to volatile analogs and then analyzed by GC/MS. Do not limit yourself however if both instruments are available. LC/MS and GC/MS can be complimentary and both may be needed as well as NMR, SEC and other techniques for more complete identification and characterization of the plant extracts. For those compounds that can be analyzed by GC, this technique offers very high chromatographic resolution, much higher than can be achieved by HPLC or UHPLC.
It depends on the type and nature of plant extract that you are interested in. If you are looking for all sort of components (a kind of general screening of all compounds) including terpenes,tannins,steroids, and flavanoids, its better to go with LC/MS. Just use longer columns preferably with a flow rate of 0.2mL/min and percentage B from 5-100 in 25 minutes. We have used Agilent Poroshell 120 EC-C18 coloumn (2.1x 150mm, 2.7um) with the above mentioned flow rate and %B for analyzing the whole unknown components from plant extracts . The resolution was found to be more better rather than having a higher %B (40-100 in 30 min) to start with .To have a better chromatographic resolution you can try using 2D-HPLC also, which is preferably applied in analyzing tannins that are difficult to resolve using normal HPLC/UHPLC. If you want to analyze compounds that are less volatile than the mobile phase its better to go with using a Evaporative light scattering detector (ELSD) that enables detection of semi-volatile and thermally-sensitive compounds at low levels. And may be you can try analyzing with GC/MS if the samples are found to be highly volatile.
Both the instruments ( GC/MS and HPLC ) are excellent in their operational behavior and result.The choice in between GC/MS and HPLC depend upon the chemistry of plant as well as type of expected chemical compound, What type of solvent used during the chemical process.
I think it all depends on what you wanted to achieve.I prefer using GC-MS due to its flexibility,cost and the not-to-cumbersome procedure.
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