I am trying to separate glucose ,fructose ,mannose ,galactose ,arabinose and xylose in Agilent 1260 infinity HPLC system which has DAD and RID detectors by using Zorbax NH2 column.
conditions maintained are
Mobile phase is 75% ACN and 25% water.
flow rate:0.4ml/min
Temperature of column and detector are 35 0C.
sample injection: 5 micro liter.
sample concentration: 1 mg/ml
But I am unable separate these sugars. Could you please suggest me what parameters I should change here?
We also have some columns like poroshell 120 HILIC-Z ,C-18.Which column you will suggest for separation of sugars among these three columns?
Also check https://www.agilent.com/cs/library/applications/lc04.pdf
Hi Talari,
Consider using anion-exchange chromatography for carbohydrate analysis. In that case you need to couple it with electrochemical detection. With his technique you are able to establish very demanding separations of sugars coupled with extremely sensitive detection. Here you can find an application note based on HPAEC-PAD (Anion Exchange Chromatography in combination with pulsed ampermetric detection as an example. Although it is focussed on the analysis of trace amounts of Lactose and isomers in dairy products, it shows the capabilities of the technique. The original work was performed on an Agilent 1260 Infinity II LC system in combination with an Antec Scientific DECADE Elite electrochemical detector (there are control drivers available for the detector to operate it under OpenLab CDS or OpenLAb Chemstation edition). See application note:
https://antecscientific.com/download/19/food-applications/8049/220_009_06-lactose-free-products
Further information about the type of column used can be found here:
https://assets.thermofisher.com/TFS-Assets/CMD/manuals/Man-065686-IC-CarboPac-PA210-Fast-4um-Man065686-EN.pdf
Regards, Hendrik
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