Abstract: The extraction of crude polysaccharides (YP) from the tubers of Habenaria ciliolaris Kranzl was performed by hot water extraction and alcohol precipitation. DEAE cellulose 52 ion exchange column and Toyopearal HW-65 gel filtration column chromatographies were successively used to separate and purify the crude polysaccharides, yielding four fractions: YP1-1, YP2-1, YP2-2 and YP3-1. The purities and molecular masses of the purified polysaccharides were determined by high performance liquid chromatography (HPLC) with evaporative light-scattering detection and their structures were investigated by Fourier transform infrared (FT-IR) spectroscopy, ultraviolet (UV) spectroscopy, and HPLC. The monosaccharide composition was determined by HPLC with precolumn derivatization. The in vitro bile acid-binding capacity of these polysaccharides was evaluated in a simulated intestinal environment. The results showed that the yield of YP was 2.41% and the total sugar content of YP was 96.35%. YP1-1, YP2-1, YP2-2 and YP3-1 were purified to homogeneity and did not contain nuclear acid or protein. Their molecular masses were 3.52 × 106, 3.08 × 106, 1.93 × 106 and 3.35 × 106 Da, respectively. The four polysaccharides mainly contained pyranose rings connected through β-glucoside bonds. Monosaccharide composition analysis demonstrated that YP1-1 and YP3-1 were composed of rhamnose, arabinose, mannose, glucose, and galactose while YP2-1 and YP2-2 were composed of rhamnose, arabinose, mannose, glucose, galactose, and fucose. All four polysaccharides exhibited strong ability to bind bile acid.

Key words: Habenaria ciliolaris Kranzl, polysaccharides, isolation and purification, bile acid binding