Preparation Of Modified Pectin And The Structure Characteristics Of Its Binding To GAL-3

Pectin is a kind of macromolecular polysaccharide with complex structure, it is widely used in juice, bakery and other food technology, since it has the function of keeping stability and thickening. In addition, the pectin can also be used in health care products, medicines. A class of pectin which is mainly modified by enzymatic, chemical and physical method, called modified citrus pectin (MCP), is proved to have enhance immunity, reduce the incidence of cancer and cancer metastasis physiological functions. MCP is used as a tumor-suppressor due to its ability binding to carbohydrate recognition of Galectin-3 (GAL-3), it has rare reports about the structure-activity relationship. As raw material, commercial citrus pectin is in turns treated with acid hydrolysis, membrane separation, enzyme hydrolysis and ion exchange chromatography method to produce MCP with different molecular weight and structure characteristics, the ability of different pectin structures binding to GAL-3 were observed by Elisa.Firstly, the commercial citrus pectin with high relative molecular mass (Mp=3.30×105) was treated with acid degradation, the change of relative molecular mass distribution during acid hydrolysis process was observed, acid hydrolysis conditions were determined as follows: pectin powders with an initial concentration of 24%(w/v) were completely dissolved in 1.5 mol/L HCl at 80? within 210 min. It produced big HG fragments (Mp=2.20×105-2.50×105) and small RG fragments (Mp=3.00×104-5.00×104). The pectin acid hydrolysate was purified by ultrafiltration, the MWCO of membrane was 10 kDa, the operating temperature and pressure was 35? and 10.0 bar respectively. After 130 min, most of NaCl was dislodged, and the ultrafiltration pectin product showed unimodal distribution form(Mp=1.90×105). Using Novozyme pectinex(Ultra SP-L) for further degradation, under the condition of 50?, pH=5.0, 0.2%(v/v) enzyme was added to 1%(w/v) pectin solution within 60 min, the lower molecular weight (Mp=6.89×104) pectin was produced.Then, the galacturonic acid content, reducing sugar content, esterification degree and sugar composition were measured. The monosaccharide types mainly include galacturonic acid, galactose, rhamnose, arabinose and glucose. The galacturonic acid content of pectin acid hydrolysate, ultrafiltration and enzymolysis products were 45.7±2.37%,73.9±1.32%, 78.4±1.93%, the reducing sugar content of them were 43.66±0.17%,18.07±0.81%, 29.54±0.27%, and the DM content were 10.3±0.06%,11.3±0.16%,37.22±1.36%, respectively. Pectin structure was further observed through scanning electron microscope, pectin ultrafiltration product chain aggregation state changed, and enzymolysis products show a filament shape. Besides infrared spectra further verified the esterification degree of pectin hydrolysates reduced.Finally, the DEAE chromatographic column was used for further separation, elution with pure water,0.05 mol/L-1.0 mol/L NaCl in turns, MCP-1, MCP-2, MCP-3, MCP-4 were obtained. The peak relative molecular mass of them were 0.12×105,1.25×105,1.46×105, 1.93×105, and 1.52×105, their monosaccharide composition included rhamnose, arabinose, galactosamine, galactose, glucose, xylose, mannose and galacturonic acid. Elisa was used to observe the ability of MCP binding to GAL-3, it found that the MCP with rich RG region has higher ability binding to GAL-3, MCP with pure HG region has the weakest ability binding to GAL-3.