Determination of Monosaccharides in Polysaccharides of Rhaponticum Uniforum by High-performance Anion Exchange Chromatography with Amperometric Detection

High performance anion-exchange chromatography with pulse-amperometric detection (HPAEC-PAD) was applied to the determination of monosaccharides in polysaccharide of Rhaponticum uniforum.Polysaccharide was separated from the sample by refluxing-extraction with C2H5OH-H2O (80+20) for 3 h to defat.After volatilization of ethanol,the drug residue was extracted twice with boiling water,maintaining 1 h for each extraction.Ethanol (95%) was added to the water extract in a volume of 200 mL to make content of ethanol in the solution achieved to 4/5 of its total volume.Polysaccharides in the ethanol solution were precipitated overnight at 4 ℃.Protein in the crude polysaccharide was removed by the Sevage method.One mg of the polysaccharide was hydrolyzed for 4 h at 100 ℃ in 2 mol·L-1 H2SO4,and then separated on the CarboPac PA10 chromatographic column.100 mmol·L-1 NaOH solution was used as eluant passing through the column at a flow-rate of 1.0 mL·min-1.Monosaccharides in the eluate were determined by PAD.Six min were taken for the whole analysis.It was found that arabinnose,glucose and fructose in the mole ratio of 1.55 to 1.00 to 2.10 were the 3 monosaccharides in polysaccharide of Rhaponticum uniforum.Detection limits found were in the range from 2.8 μg·L-1 to 12.3 μg·L-1,and recoveries found by standard addition were in the range of 93.0%-107.8%