Extraction Of Polysaccharides From Malus Hupehensis By Aqueous Two-phase System And Its Physiological Activity

Malus hupehensis(Pamp.)Rehder(M.hupehensis)is the most important ornamental plant of Malus.Its leaves show edible and medicinal value.polysaccharides,polyphenols and flavonoids show antioxidant,anti hyperglycemic,anti-tumor and other pharmacological activities.Aqueous two-phase system(ATPS)is composed of two immiscible polymers or one polymer,salt and water.It is widely used in one-step extraction and purification of polysaccharides,proteins and other biomolecules.Compared with traditional extraction methods,aqueous two-phase extraction is a green and efficient solvent system,which provides a mild and biocompatible environment for the target compounds.It has the advantages of simple operation,fast phase separation and good solubility.The purpose of this study is:(1)to optimize the extraction conditions of Polysaccharides from Malus hupehensis(MHP)by response surface methodology,and to establish a rapid and effective extraction method based on ultrasonic assisted aqueous twophase;(2)to preliminarily characterize MHP by chemical analysis and modern chromatographic analysis;(3)to study the potential antioxidant and hypoglycemic activities of MHP in the field of food and medicine.The results are as follows(1)The extraction rate of MHP in different ethanol salt ion aqueous two-phase systems was compared.The ethanol ammonium sulfate system showed a relatively high extraction rate.Therefore,the ethanol ammonium sulfate system was selected for single factor and response surface experiments to optimize the process.The results of single factor experiment showed that ultrasonic power,ultrasonic time and solid-liquid ratio show significant influence on the extraction rate of MHP,but temperature was not significant influence to it.The results showed that the optimum extraction conditions were as follows: 30.31%(w / w)ethanol and 19.86%(w /w)ammonium sulfate aqueous two-phase,extraction temperature 50 °C,ultrasonic power 65 W,ultrasonic time 33 min,solid-liquid ratio of 1:23(g / m L).The yield and purity of MHP were47.63% and 83.63%,respectively.(2)The structure of MHP was characterized by UV full wavelength scanning,Fourier transform infrared spectroscopy(FTIR),high performance liquid chromatography(HPLC),Congo red test,Circular dichroism(CD)and scanning electron microscopy(SEM).The results showed that the MHP extracted by UAATPS did not contain or contained a small amount of nucleic acid and protein,indicating that the impurities such as nucleic acid and protein in MHP could be effectively removed by UAATPS extraction,and the purity of MHP could be improved.MHP was composed of Mannose,Glucuronic acid,Rhamnose,Galacturonic acid,Glucose,Galactose,Xylose,Arabinose and Fucose in the molar ratio of3.22:1.85:4.25:5.08:6.69:38.46:4.68:30.90:4.87.Congo red test and CD analysis showed that MHP had triple helix and asymmetric structure.(3)The results of chemical method showed that MHP showed good antioxidant activity and inhibition of ages activity.The results of cytological test showed that MHP could significantly increase the glucose consumption rate of IR cells,reduce the levels of TG and LDL-C in IR cells,and increase the levels of SOD,CAT,LDL-C and T-AOC.MHP had good antioxidant and hypoglycemic activities.In conclusion,response surface methodology(RSM)was used to optimize the UAATPS extraction process of polysaccharides from Malus hupehensis.The preliminary structure of MHP was characterized.The antioxidant and hypoglycemic activities of MHP in vitro were studied by chemical and cell biological methods.However,further animal experiments and the mechanism of action still need to be explored.It not only provides a green,simple and efficient technical reference for the extraction of polysaccharides from plants,but also provides a theoretical basis for improving the added value of Malus hupehensis leaves and promoting its comprehensive utilization.