Study On The Degradation Of Porphyra Haitanensis Polysaccharides And The Activity And Structure Characterization Of Its Degraded Products

Porphyra haitanensis,as a common macroalgae,accounts for more than 80%of the total amount of Porphyra in China.It has been recorded in the "Compendium of Materia Medica" and other traditional Chinese medicine books.It is rich in nutrients such as protein,vitamins,polysaccharides,etc.As one of the main components of Porphyra haitanensis,polysaccharides have many biological activities,such as antioxidant and immunomodulation,which are often associated with their physical and chemical properties and molecular weight and structure,etc.Crude Porphyra haitanensis polysaccharides（PHP）were prepared by hot water extraction,alcohol precipitation,deproteinization,dialysis and freeze-drying,with a yield of 7.1%.PHP were degraded using a hydrogen peroxide-ascorbic acid system,when the degradation conditions were as follows,H₂O₂（5 mmol/L）+Vc（0.5 mmol/L）,H₂O₂（4mmol/L）+Vc（1 mmol/L）,H₂O₂（4 mmol/L）+Vc（1.5 mmol/L）,H₂O₂（20 mmol/L）+Vc（20mmol/L）,the solutions of PHP were reacted at 30℃ for 2 h under the above conditions respectively,yielding four degradation products:PHPD-I（329 kDa）,PHPD-II（203 kDa）,PHPD-III（128 kDa）,and PHPD-IV（10 kDa）.We proved that this degradation method had no major influence on the important structural units of polysaccharide using infrared spectrum analysis.The results of physical and chemical properties suggest that crude polysaccharide and the four degradation components were mainly composed of galactose.The sugar contents of PHP,PHPD-I,PHPD-II,PHPD-III and PHPD-IV were 59.11%,54.63%,60.65%,56.07%,58.76%,respectively;protein contents of them were 9.1%,7.08%,8.18%,5.62%,5.07%,respectively;sulfated group contents were 4.9%,6.2%,4.5%,6.7%,5.7%,respectively;the contents of3,6-anhydro-galactose were 11.13%,7.17%,14.34%,10.26%,9.32%,respectively;and the contents of galactose was 46.89%,46.2%,45.01%,44.51%,50.7%,respectively.The results show that PHP and its degraded products have antioxidant and immunostimulatory activities in a dose-dependent manner and the degradation products were higher than those of PHP,and the product with the lowest molecular weight（PHPD-IV）had the strongest activity.Five components,PHPD-IV-1,PHPD-IV-2,PHPD-IV-3,PHPD-IV-4 and PHPD-IV-5,were obtained after purification by DEAE-52 cellulose column.The yields of the five components were 3.3%,36.6%,20.2%,6.6%and 6.9%,respectively.The results show that PHPD-IV-4had the best immunomodulation activity.In the determination of its physical and chemical properties,it was found that the sulfated group content was the highest among the five components,which may be closely related to its activity.In addition,in the study of the MAPKs signaling pathway of PHPD-IV-4,it was found that it was involved in the immunomodulation process by up-regulating the ERK,JNK and p38 expression.Furthermore,PHPD-IV-4 was purified by Sephadex-G50,and its monosaccharide composition was analyzed by GC and GC-MS.The results show that PHPD-IV-4 was composed of galactose and 6-O-Me-galactose.The structure of PHPD-IV-4 was analyzed by IR and NMR.It was found that it contains two kinds of glycoside bonds,αandβ,and its repetitive structural units contains→3)β-D-galactose（1→4）3,6-anhydro-α-L-galactose（1→and→3）β-D-galactose（1→4）α-L-galactose(1→,the repetitive structural units were replaced by methoxy group at the C-6 position of galactose and the C-2 position of3,6-anhydro-α-L-galactose.