酵母β葡聚醣结构特性监定及活化巨噬细胞之探讨.doc

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1、酵母卩葡聚醵結構特性鑑定及活化巨噬細胞之探討StructuralcharacterizationandmacrophageactivationbyP-glucanisolatedfromSaccharomycesserevisiae研究生：黃柏銘Huang,Bo・Ming指導教授：黃卓治Huang,Tzou-Chi【摘要】卩-glucan為真菌類細胞壁上的主要成份之一，具有調節免疫活性的能力。卩・ghcari的免疫活性會受其結構所影響。本研究中利用酸鹼萃取的方式從酵母細胞壁中分離製備出水不溶性的particulateglucan，再進一步進行磷酸化及硫酸化製備成

2、水溶性的衍生物(glucanphosphate及glucansulfate)-利用NMR、FTIR、GC・MS、HPSEC進行結構特徵分析後指出酵母glucans具有卩・1—3鍵結所形成的骨架及P-l->6分支的結構，其分子量超過16萬，分支度為0.25〜0.43。進一步探討水不溶性的particulateglucan及二個水溶性衍生物對RAW264.7巨噬細胞株活性之影響。結果顯示glucansulfate具有較顯著的提升吞噬活性能力以及增加IFN*誘發RAW264.7巨噬細胞分泌NO和iNOS的表現。經由西方墨點法分析的結果指出glucansulfate能刺

3、激IkB-«的磷酸化和降解並活化NFkB°關鍵字：酵母、卩■葡聚陋、NO合成酶、NFkB[Abstract]P-glucanswereconservedcomponentsoffungalcellwallswhichpossessimmunomodulatoryactivities.Theactivityof卩・glucansappearstodependontheirstructuralproperties.Inthisstudy,weusedanalkaline-acidhydrolysismethodtoisolatedwater-insolublepar

4、ticulateglucanfromSaccharomycescerevisiaecellwell,andthenphosphorylationandsulfationtopreparewater-solublederivatives(glucanphosphateandglucansulfate).TheirstructurewerecharacterizedbyusingNMR,FTIR,GC-MS,IIPSEC,andtheresultsindicatedthatthisglucanconsistedofabackbonecomposedof(If3)■卩

5、・D-glucosylresiduesandcarriedasingle(1—>6)-P-linkedD-glucosylresidue.Themolecularweightoftheglucanswerelargerthan1.6幻0‘daltons.Thedegreeofbranchingwas0.25-0.33.Theimmunomodulatoryeffectofwater-insolubleparticulateglucanandthetwowater-solublederviativesontheRAW264.7macrophagewasstudie

6、d.Theresultsshowedthatglucansulfatesignificantlyincreasedphagocyticactivityandinducednitricoxide(NO)productioninTFN-y-primedRAW264.7macrophagesthroughtheincreasedexpressionofinduciblenitricoxidesynthase(iNOS).Westernimmunoblottingassaysrevealedthatglucansulfatestimulatedphosphorylati

7、onanddegradationofIkB-«andsubsequentNFkBactivation.Keywords:Saccharomycescerevisiae.卜glucan,iNOS、NFkB