Study On Separation, Purification, Physicochemical And Structural Characteristics Of Glycoprotein From Jellyfish

Jellyfish (Rhopilema esculehtum Kishinouye) is a large edible and warm waterjellyfish who growth in Marine planktonic life camp. We have a wide range ofresources distribution in the South China Sea, the East China Sea, Yellow Sea andBohai Sea. There are wide ranges of resources which the annual output was300,000tons. Jellyfish play as medicine and food, which not only provides people with richnutrients, has been recorded in the ancient medical books. However, the developmentand utilization of the jellyfish only stay in curing or simple cooking stage at presentwithout deeper processing and research of the jellyfish, as a result, we are waste theresource. This paper research extracted and purified pure the glycoprotein fromjellyfish, also we study its physicochemical properties and structure characteristics. Inthis paper, the research contents and results of jellyfish are as follows:1. Jellyfish glycoprotein extraction process optimization: Basis on the singlefactor experiments, design Box-Behnken response surface experiments using theDesign V8.0.6, optimum technological conditions of extraction jellyfish glycoproteinis: NaCl concentration is0.57mol/L, extraction time is12.50h, and solid-liquid ratio is1:5.50, ultrasonic broken time for20.00min, ultrasonic power is275.00W. Basis on thecondition of the extraction yield will be: total protein content is19.89%, total sugarcontent is2.86%, the glycoprotein extraction yield is9.14%.2. The purification and electrophoresis of jellyfish glycoprotein: JGP-III wasobtained from crude glycoprotein isolated using SP Sepharose FF anionic gelatin, andJGP-III-II was obtained from JGP-III using Sephacryl S-300HR gel columnchromatography. Identify by using Sephacryl S-300HR we found a single symmetricalpeak, it is said JGP-III-II component is single, purification yield is1.8%. After SDS-PAGE electrophoresis experiment using coomassie brilliant blue staining and PASstaining for identify sugar. Results show that JGP-III-II is a kind of glycoprotein andits molecular weight is41.53KD. 3. Physicochemical properties of JGP-III-II: A single symmetrical peak wasobtained from HPLC analysis, and proved that JGP-III-II is a kind of glycoproteinsagain by connection the differential detector, measured JGP-III-II relative molecularmass is44.196KD, and it is not significant compare with SDS-PAGE electrophoresisexperiment results, and its purity is98.25%. JGP-III-II thermal denaturationtemperature is53.6oC and thermal shrinkage temperature is92oC, the existence of thesugar chain play a collaborative role on its thermal stability. Amino acid analysisshowed that JGP-III-II contains16amino acids, which contains7kinds of essentialamino acids. Glycine, valine and glutamic acid are main composition amino acids ofJGP-III-II amino acids. PMP of HPLC method showed that the monosaccharidecomponents of JGP-III-II:2-amino-D-mannose, glucose, amino galactose, lactose,glucose, galactose, xylose and fucose, their content proportions are:6.44:4.80:4.86:66.24:1.38:1.96:6.17:1.63.4. Structure characteristics of JGP-III-II: Basis on β-eliminate reaction andglycosidase (PNGase F) enzymolysis reaction, the result show that O-glycosidic andN-glycosidic consist in JGP-III-II. By infrared scanning for JGP-III-II, we found itscharacteristic absorption peak of glycoprotein or sugar wave number of2966.55cm-1and3405.47cm-1, and the secondary structure is α-helical, glycosidic bond belongs topyran type. MADLI-TOF-MS/MS mass spectrometry analysis showed that JGP-III-IIhas7N-glycosylation sites, and there may be has several O-glycosidic sites. Also theresult showed N-glycosidic and O-glycosidic are all found in the JGP-III-II, and O-glycosidic is more than N-glycosidic.