ANALYSIS OF MANNOBIOSE SOLUTION BY HPAEC

University Of Georgia, Athens GA

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Abstract

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Mannobiose composition analysis by HPAEC The sample was analyzed at two different concentrations: first by directly pipetting 100 [unreadable]L from the original container, and second, by diluting the sample 2 times. Four concentrations of 2[unreadable]-Mannosbiose standard (Sigma cat. # M1050) were prepared (0.03125 mg/mL, 0.0625 mg/mL, 0.125 mg/mL, and 0.25 mg/mL) to establish a calibration curve. The concentration of mannobiose in the sample was quantified by linear interpolation from the calibration equation. The mannobiose was analyzed by HPAEC using a Dionex ICS3000 system equipped with a gradient pump, an electrochemical detector, and an autosampler. The disaccharide was separated by a Dionex CarboPac PA20 (3 x 150 mm) analytical column with an amino trap. The gradient program used eluents A - degassed nanopure water and B - 200 mM NaOH. The autosampler was set deliver 10 [unreadable]L per injection and injection was made every 40 minutes. All methods were based on protocols described by Hardy and Townsend (Hardy, M. R., and Townsend, R. R., "High-pH anion-exchange chromatography of glycoprotein-derived carbohydrates", 1994, Methods Enzymol. 230: 208-225).