Components Isolation & Purification, Physiochemical Properties & Three-dimensional Structure Identification And Biological Activities Of MPs From Mytilus Coruscus

Objective:To isolate and purify active polysaccharide MPs from Mytilus coruscus with various purity and determine its physiochemical properties including sugar content, purity, monosaccharide components, molecular weight, glycoside type and ID- and 2D-structure. To investigate the biological activities of different fractions of MPs.Methods:1 .Isolation & purification of MPsAfter homogenization, extraction by hot water, deproteinized by Sevage method and alcohol fractional precipitation, polysaccharide MPs was isolated from Mytilus coruscus, then purified by ion-exchange chromatographies on DEAE-cellulose 52, Sephacryl S-200HR column and HPLC.2.Physiochemical properties & structure identification of MPsPrepared MPs samples' sugar content was determined by A.max of full wave-length scan and Sulphuric acid-Phenol Colorimetry. Ultraviolet absorption spectrum, Staining-combined colorimetry, HPLC and SDS-polyacrylamide gel electrophoresis were applied to prove the homogeneity of this polysaccharide. The molecular weight were obtained from gel filtration while TLC and GC showed the composition of MPs.We performed IR spectrum to analyze the samples' characteristic IR aborption spectrum and glycoside type. Besides, NMR analysis will help to explore the ID- and 2D-structure of purified samples.3.Biological activities of MPsMPs-B₁, the best purified fraction of all components of MPs, were solved into solutions of certain concentration in pursuit of tests on its proliferation of sertorli cells of the rat testes and induction of K562 cell apoptosis.Results:With excellent hydrophilia and dissolubility, the amber powder, prepared CMPs were purified by DEAE-cellulose 52, Sephacryl S-200 HR column chromatography into semi-purified MPs: MPs-A, MPs-B, MPs-C, which were resulted into eight full-purified fractions: MPS-A₁/A₂/B₁/B₂/B₃/C₁/C₂/C₃ by HPLC. Sulphuric acid-Phenol Colorimetry, UV Absorption Spectrum, Staining-combined colorimetry,performed on the carbonhydroprotein content determination of MPs proved them wholly as neutral polysaccharides without protein nearly. Analysis of gel filtration (HPLC) showed that its fractions were homogenous except MPS-A2 and SDS-polyacrylamide gel electrophoresis illustrateded that its seven fractions were all homogenous. According to the standard curve of elution time-comparative molecular weight, the apparent average molecular weight of MPS-A1/A2/B1/B2/B3/C1/C2/C3 were 3.8X10\ 4.139X10% 3,51X10% 5.2X10% 4.168X10% 4.22X10% 6.85X10% 5.50X104Da respectively. IR spectrum exhibited that the other three fractions, of which MPs-Bi was a-D-pyranose, MPS-C2 & MPS-C3 were both B-type furanose, were structure-complicated carbohydrate complex except of the MPs-Ai,with nonsignificant characteristic carbohydrate IR aborption spectrum. GC results told us that MPs-Ai was composed of glucose only while MPs-Bi & MPS-C2 were both composed of glucose and galactose whose molar ratio was 39.07:60.93 and 57.30:42.70. Among them MPs-Bjthe best purified fraction , were identified MPs-Bi as a-D-hexapyranose by 'HNMR. The two-dimension structure of MPs-Bi is on the further study. MPs-Bj could enhance the multipliction of sertorli cells of the rat testes as well as surpress the proliferation of K562 cell and induce cell apoptosis.Conclusion:With the improvement of preparation processing, purified MPs' fractions we obtained were significantly different from the ones derived from the same materials in similar or former studies. MPs-Bl could enhance the multipliction of sertorli cells of the rat testes as well as surpress the proliferation of K562 cell and induce cell apoptosis.