Quality Analysis of Edible Bird’s Nest and the Separation and Structure Characteristics Research of Sialoglycoprotein Extracted from Indonesia BAIYANZHAN

Edible Bird’s Nest (EBN) has many nutrients such as sialoglycoprotein andminerals. It has been widely used for stimulating cell division and growth, enhancingimmunization, enhancing tissue growth and regeneration and so on. So far, there aremany researches on EBN authentication, but for the quality analysis and itsimmunocompetence study is not that comprehensive and perfect, the separation andcharacteristics research on sialoglycoprotein in EBN, which is EBN’s characteristiccomponent, is rarely reported among domestic and foreign journals.Here we report the basic composition analysis of eight kinds of EBN as rawmaterials, take the Indonesia BAIYANZHAN as major material for active functionalevaluation and establishing extraction, separation and characteristics analysis methodsfor EBN’s glycoprotein. The main research contents and the results are as follows:1. The content of water, proteins, sialic acid and amino acid of eight kinds ofEBNs from Indonesia and Malaysia are determined, monosaccharide and elementalcomposition are detected at the same time, analysis and compares the nutrients andquality of different EBNs, the significant differences in monosaccharide compositionand elemental composition among different EBNs are revealed, giving a research andtheory reference for further research on EBN.2. The immunomodulatory effect of Indonesia BAIYANZHAN on hypoimmunemice is investigated. Different dosage of EBN is given to hypoimmune model miceinduced by injecting Kunming mice with Hydrocortisone. The spleen and thymusindex, delayed-type hypersensitivity (DTH), hemolysin content and the phagocyticability of celiac macrophage are measured, respectively. The result is that EBN hasremarkable function in immune regulation. It can increase the spleen index (P<0.01)and thymus index (P<0.01and P<0.05), hemolysin content (P<0.01) and DTH level (P<0.01), promote the phagocytic index and rate, obviously. They can promoteimmune function by cell immunity, humoral immunity and non-specific immunity inimmunodepression mice. EBN can stimulate both specific and non–specific immunefunction in mice, providing the theory basis for clarifying EBN’s health effection andreasonable application.3. Using Indonesia BAIYANZHAN as main material, the extraction of rawglycoprotein is extracted, purified and property analyzed, choose60℃as extractionconditions. DE52, DEAE650M and QFF three kinds of anion exchange resins areapplied for separating glycoprotein from the raw extraction sample, sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE) combined lectin blottingmethod are used for determining the separation effect, QFF gradient elution at lowNaCl concentration is selected for preliminary separation, affinity chromatography isused for glycoprotein’s further purification. A single sialoglycoprotein band close to43kDa molecular weight is obtained, baring the structure ofNeu5Acα2-3Galβ1-4GlcNAc/Glc, which one has a Neu5Acα2-3Gal sialic acidterminal connection structure. The extraction, purification and structure characteristicof sialoglycoprotein in EBN lay a foundation for EBN’s targeted compoundspurification, identification and activity evaluation.4. EBN has high sialic acid content. SDS-PAGE analysis combined with lectinblotting analysis with the sialic acid specific lectins MAAⅡ, SNA and MAA areperformed on the glycoprotein extracted from eight kinds of EBN, most of theglycoproteins have been sialylated. The expression of sialic acid residues’ linkage indifferent EBNs is identified. MAAⅡ, SNA and MAA lectins are used to identifyterminally expressed sugars——the sialic acid residues. The lectin reaction showsthat most EBNs have the50kDa and43kDa sialoglycoprotein bands with theNeu5Acα2-3Gal terminally sialic acid linkage structure; the43kDa one has aNeu5Acα2-3Galβ1-4GlcNAc/Glc structure of oligosaccharides. IndonesiaBAIMAOYANWO and Malaria LONGYAZHAN also have aNeu5Acα2-6Galβ1-4GlcNAc oligosaccharides structure at the glycoprotein band between30kDa~40kDa molecular. Two-dimensional gel electrophoresis (2D) isperformed on protein point distribution between two representative kinds of EBN.The structure of purified sialoglycoprotein ahead is analyzed by lectin blotting forsialic acid linkage and matrix-assisted laser desorption/ionization tandemtime-of-flight mass spectrometry (MALDI-TOF MS) for peptides sequence of proteinfraction.