I have a plant extract [aqueous] which has shown positive results for qualitative tests for phenolics and sugars. Though the sample is showing good HPLC profile [mobile phase-acetonitrile: acidified water] for phenolis, I am unable to identify individual components. LCMS both positive and negative modes are showing only two to three components mass [mobile phase -acetonitrile: water], can somebody suggest me the procedure.
Hi, You have LCMS data for your extract. Did you use (ESI HR) MS? I think with your MS data, you can predict the molecular ion using their adducts, after you determine the molecular ion(s), you can predict the molecular formula using isotope pattern, after this you can use many free on-line MS data bases such as MassBAnk, Metfrag, Metlin, mzCloud or others to predict your compounds, if you have known compounds I think the data bases can predict your compounds. You can search the lonk for MS data bases by Google., Best regards
I read the problem as "the mass spectrometer only shows two to three components and I expect to see more peaks". Does the UV trace show more peaks? I'm assuming you are using ESI, and not APCI or another ionization technique.
Assuming my interpretation of the question is correct, I'd try the following:
I'd try running the chromatography with some model compounds, such as catechin, to represent phenolic compounds. Also run some test chromatograms with some model sugars.
After I'm sure everything is working, I'd check for adducts and fragments as described above. For example, I found that capsaicins tend to be observed as [M+Na] in my system. Many phenolics appear in both positive and negative ESI.
consider the suggestions that others have sent you, but is quite important the comparation with reference compounds of phenolics and sugars
hi shubha, go for TLC separation (separately for sugars and phenolics) and isolate individual compounds. inject the purified compounds in LCMS.
else, after TLC separation go for a good staining reagent for sugars and phenolics
best wishes
Dear Shubha
Did you use the same amount of aliquot both in LCMS and HPLC? Another thing you used acidified water in HPLC but LCMS only used water/ Is there any reason to omit the acid? Use acidified water it will give you good result in LC as well.
Thank you kishore and Nusrat Subhan
Acidified water usage creates some problem with the machine(LCMS), hence not allowed.
I am guessing some glycosides are also present.
What sort of MS and column are you using? This would help us to provide better answers. Which sort of ionization? Assuming ESI, acidic modifiers such as trifluoroacetic, acetic, and formic acid are commonly used for LC-MS. They are volatile acids, and, with the possible exception of triflouroacetic acid, can enhance detection in ESI+. These are compatible with C18 columns too. Mineral acids, such as phosphoric, sulfuric, and so forth are not MS friendly because they are not volatile.
Hi..You can also try NMR-based metabolomics of aqueous fraction for the qualitative and quantitative analysis of sugars, amino acids and phenolics, one of the major advantage of using NMR over LC-MS is that NMR doesn't need any standard compound and its destructive nature as you can recover your sample.For this firstly you have identify each compound in the fraction or extract and then quantify by using some internal standard.or if you have biological sample you can try other method of quantification such as eretic and eretic 2 .
Hi Dr. Aslam, What did you mean compound identified.....not present in standards? Did you analyzed a mixture of certain standards using LCMS? I mean, did you find others compound(s) in addition of the standards mixture? Did you have evaluated the stability of the standard(s) in the selected solvent? If you apply different mobile phase,it could be possible the compounds could not well separated; but I think you can still evaluate the MS and MS MS to confirm the compound(s), Kind regards
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