Studies On The Active Polysaccharides Constituents From Sea Cucumber Apostichopus Japonicus

The cold water sea cucumber A. japonicus Selenka belongs to Genus Apostichopus, Family Stichopodidae, Order Aspidochiroia, Class Holothuroidea, and Phylum Echinodermata. With the rapid development of studies of the ocean polysaccharide, the bioactive functions of polysaccharide in sea cucumber are being discovered.In this paper, I take a study for A. japonicus Selenka from Dalian, Liaoning province, taking the method of ethanol precipitation, removing protein by Sevage method, bleaching by hydrogen peroxide, to propose polysaccharides of A. japonicus Selenka. The yield average of the process from A. japonicus Selenka frozen powder to polysaccharides of A. japonicus Selenka is 8.55%. The polysaccharides of A. japonicus Selenka separated by multiple separate technologies such as using ultrafiltration device, DEAE - cellulose column chromatography, Sephadex G - 100 column chromatography, HPGFC and so on to get the following uniform sample polysaccharides: AJP-1 (the rate of production is 5.840%),AJP-2 (the rate of production is 6.480%),AJP-3 (the rate of production is 6.816%),AJP-4 (the rate of production is 6.344%),AJP-5 (the rate of production is 8.768%). The uniformity of these polysaccharides extraction is of good quality through HPGFC testing, and they do not contain protein and amino acids through ultraviolet scanning.These polysaccharides samples are ivory floe powder, soluble in water and dimethyl sulfoxide, not soluble in ethanol, acetone, etc. Sulfuric acid - phenol reaction and Molish reaction are both positive. AJP-1: relative molecular weight is 101,291 Da. The content of uronic acid in AJP-1 is the highest, quality percentage reached 1.1170%. AJP-2: relative molecular weight is 73,010 Da. It contains uronic acid of which quality percentage is 0.4202%. AJP-3: relative molecular weight is 39,638 Da. It contains uronic acid of which quality percentage is 0.5046%. AJP-4: relative molecular weight is 9,716 Da. Uronic acid could not be detected specific quality percentage. AJP-5: relative molecular weight is 2,213 Da. Sample could not be detected uronic acid.Every polysaccharides sample is different from each other, but they mainly contain fucose (Fuc), glucose (Glc), amino sugar and uronic acid, except for AJP-5 excluding fucose (Fuc) and uronic acid. Relative molecular weight of these polysaccharides samples has very big difference. Polysaccharides which have same compositions, the content ratio of their monosaccharide is different, and indican key type has some difference. Polysaccharides samples almost contain hydroxyl, carboxyl, acetamido, sulfuric acid base and the methyl of fucose. Among these different polysaccharides, sulfuric acid based replace site vary slightly. Those polysaccharides samples areα- configuration. In addition to AJP-5, the rest are mainly containα-L-Fucose-2, 4-di OSO₃^-,and most of them also containα-L-Fucose-3, 4-di OSO3-,α-L-Fucose-4-OSO₃^- and Fucose–unsulfated.Anticoagulant experiment showed that polysaccharides samples significantly prolong thrombin time (TT) in certain doses. AJP-1 which relative molecular weight above 100,000 has the best anticoagulant effect; AJP-2 and AJP-3 which relative molecular weight between 30,000 and 80,000, have anticoagulant effect; AJP-4 which relative molecular weight about 10,000, does not have obviously anticoagulant effect. The anticoagulant effect of high relative molecular weight polysaccharide is better than low relative molecular weight polysaccharide. The anticoagulant effect of polysaccharide relative molecular weight below 10,000 is not well. This concerned with the structure of polysaccharide.The PTPσsuppression test of polysaccharide sample shows that these polysaccharides have nerve protection. The inhibition ratio is 99.8% while the concentration is 20μg/mL. Through in-depth study, it might be applied to the research and development of improving neural function drugs or health care function food, or prevention of Alzheimer's drug.