| In this paper, we used 3-amino-9-ethylcarbazole (AEC) as a derivatizing reagent for derivatization analysis chitooligosaccharides and N-glycan chain of glycoproteins based on analysis AEC-labeled monosaccharides. The derivatives were separated by reverse-phase HPLC and identified by LC-ESI-MS. Finally, we have obtained these results as follows:1. In this paper, we use AEC for derivatization analysis chitooligosaccharides. The derivatives were separated by reverse-phase HPLC and identified by ESI-MS and online LC-ESI-MS. We optimized the HPLC conditions of LC-ESI-MS analysis, The column used was filled with RP-C18 packing (5μm,250 mm x 4.6 mm). The mobile phase was comprised of 10 mmol/L ammonium acetate, pH 4.5 (solvent A), and acetonitrile (solvent B), the AEC-labeled chitooligosaccharides were eluted using a gradient program as following: from 90%A to 75%A for 80 min at a flow rate of 0.2 mL/min. The AEC-labeled chitooligosaccharides was detected by UV at 254 nm To achieve a simultaneous separation and structure identified the different degree of polymerization of oligosaccharides.2. We further expand the application of AEC, use it as a derivatizing reagent for derivatization analysis N-glycan chain of ribonuclease B and hen ovalbumin. Established the method for derivatization analysis N-glycan chains separated by reverse-phase HPLC and identified by ESI-MS and online LC-ESI-MS. At first, we use PNGase F enzyme to released N-glycan chain of glycoprotein and isolation and purification it. Complete this step, we use AEC as a derivatizing reagent for derivatization analysis N-glycan chain of ribonuclease B. and hen ovalbumin. The third step, to analysis AEC-labeled N-glycan chain by reverse-phase HPLC and ESI-MS, we also use LC-ESI-MS to analysis the derivative, this method has good repeatability and save sample. The suitability of the developed method for analysis N-glycan chain of biological specimen is satisfactory.
|
PDF Full Text Request