Extraction Of Polysaccharides From Dendrobium Huoshanense By Ultrasonic Wave And Its Effect On The Physicochemical Property And Antioxidant Activity Of Polysaccharides

Dendrobium huoshanense which belongs to a genus of the orchidaceae family is one of the rare medicinal plants.The polysaccharides,alkaloids and polyphenols are the main bioactive constituents in Dendrobium huoshanense. Modern pharmacological studies have shown that Dendrobium huoshanense polysaccharide has a variety of medicinal functions, such as antioxidant, anti-tumor, immune modulation and anti-diabetic properties.The effect of ultrasonic on the extraction from Dendrobium huoshanenseand the physicochemical property and antioxidant activity of polysaccharides were investigated.1. Ultrasonic-assisted extraction and optimization of Dendrobium huoshanense polysaccharides.The influence of liquid-to-solid ratio, extraction temperature, extraction time, ultrasonic power and ultrasonic time on the yield of polysaccharides were investigated and Response surface methodology (RSM) was employed to optimize the extraction process.The optimal conditions for ultrasonic-assisted extraction were as follows: liquid-to-solid ratio 30:1, extraction temperature 81 ℃, extraction time 120min, ultrasonic power 423W, ultrasonic time 8min. Under these conditions, the average yield of polysaccharides was 19.96mg/g, which was well close agreement with the value predicted, and the relative error was 0.65%.Under the same conditions, the average yield of polysaccharides extracted by hot water only 11.73mg/g. The results showed that the ultrasonic-assisted method was better than hot water method for the extraction of Dendrobium huoshanense polysaccharide.2.Isolation and purification of the crude polysaccharides from Dendrobium huoshanenseThe crude polysaccharides were extracted from Dendrobium huoshanense by ultrasonic-assisted extraction method and hot water method.DHP and UDHP were obtained afterethanol deposition, deproteinization, decolorization, and dialysisand freeze-drying. DHP and UDHPwere analyzed by HPLC. The results indicated that DHP and UDHP were the mixture of several polysaccharides.The molecular weight distribution of DHP and UDHP was 8.11×103 Da～ 4.37×105Da and 5.87×103 Da～2.89×105 Da, respectively.Three polysaccharides fractions were obtained from DHP and UDHP by anion-exchange chromatography, respectively. DHP-1, DHP-2 and DHP-3 were receivedfrom DHP. DHP-1, DHP-2 and DHP-3were occupies 75.4%, 14.3% and 3.5% of original mass, respectively. UDHP-1, UDHP-2 and UDHP-3 were received from UDHP.UDHP-1, UDHP-2 and UDHP-3 were occupies 86.4%,5.4% and 0.9% of original mass, respectively.Among these fractions, DHP-1 and UDHP-1 were further fractionated by Sephadex G-100 gel chromatographic column. UDHP-1A was obtained from UDHP and accounted for 73.2% of UDHP. DHP-1 Aand DHP-1B were received from DHP-1, and accounted for40.3% and 47.7%, respectively.3. Physical and chemical properties of Dendrobium huoshanense polysaccharidesPhysical and chemical properties of DHP-1 A and UDHP-1A were studied, according to their strong activity of antioxidant. Scanning electron microscope (SEM) images showed that the configurationof DHP-1 A was moresmooth and small than that of UDHP-1A. The configuration of UDHP-1A looked more loosely and it showed a kind of accumulation from small fragments. The results of the viscosity determination indicated that the DHP-1 A and UDHP-1A possesed low viscosity, and the viscosity of DHP-1A was higher than that of UDHP-1A. Experiments of Congo red showed that there was no triple helix structure in DHP-1 A and UDHP-1A. The results of UV spectroscopy scanning showed that there was no obvious absorption peak at 260-280nm.It was shown that DHP-1A and UDHP-1A do not contain protein and nucleic acids.Compared with the infared ray spectrum of DHP-1A and UDHP-1A, DHP-1A exhibited an absorption peak at 881 nm, which demonstrated thatβ-glycosidelinkage in its structure. However, there was no such structure in UDHP-1A.HPLC was adapted to measure purityand molecular weight of polysaccharides.The results showed that DHP-1A and UDHP-1A were homogeneous polysaccharides, and the average molecular weight of DHP-1A and UDHP-1A was 1.17×104Da and 6.08×103Da,respectively.4. Antioxidant activity of Dendrobium huoshanense polysaccharideThe reducing power, DPPH radical system, ABTS radical system and hydroxyl radical systemof polysaccharides extracted by hot water method and ultrasonic-assisted extraction method were studied. The experimental results showed that polysaccharides extracted by hot water method and ultrasonic-assisted extraction methoddemonstrated antioxidant activity, furthermore, their antioxidant ability enhanced according to the increase of polysaccharide’s concentration. Generally, the antioxidant property of ultrasonic-assisted extraction polysaccharides was higher than that of hot water extraction polysaccharides.