In most of the papers polysaccharide was quantified by converting them into their sugar constituents or some acids like sialic acid, uronic acids. Can we quantify polysaccharides without converting them into their constituents? or not. Also, how can I characterize and identify exo-polysaccharide?
You can use Phenol-Sulphuric Acid Assay to quantify EPS on their native form (crude-eps). For characterizing EPS it all depend on what you are looking for. So to determine the monomer sugars from which your EPS are constitute and their relative ratios, you need to hydrolyse the EPS with TFA then run it in HPAEC. Alternatively you can hydrolyse the eps to its monomers then convert those monomers to their alditol acetate derivatives, and you can run them using GC-MS.
You can make structure characterisation for the crude-eps using NMR analysis, e.g. Proton-NMR, carbon13-NMR, Dept135-NMR, COSY, HSQC etc.
You need to do molecular weight distribution analysis using SEC-MALLS.
Other chemical analysis are useful as well, for instance to identify if your eps have uronic acids in it or not.
If there are specific glycans you are looking for you could run a standard on HPLC and then your sample. Comparing the area under the curve for that peak with a known amount of control will allow you a very good approximation of your glycan of interest. You could simplify these complexity of exo glycans with "quick-spin" lectin columns or lectin conjugated beads. Thereby eluting a specific type of glycan while the remaining glycans are washed away. Theoretically you would only need to reduce and permethylate if you were going into a mass spec study after you had figured out these other things.
Food for thought: the media you grow the bacteria in may be very rich with contaminating glycans. Optimizing a custom growth media for such study would seem very important.
All my best,
Chris
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