一种新的粒毛盘菌多糖结构表征、化学修饰及生物活性研究

发布时间：2018-06-05 16:14

  本文选题：粒毛盘菌胞外多糖结构 + 抗肿瘤活性　； 参考：《合肥工业大学》2017年硕士论文

【摘要】：本研究经发酵提取、分离纯化获得一种新的粒毛盘菌YM130胞外多糖(LEP-2a),采用单糖组成分析、甲基化分析、傅里叶变换红外光谱分析和核磁共振光谱分析对LEP-2a进行结构表征,并评价其体外抗肿瘤活性;分别对LEP-2a进行乙酰化和苯甲酰化修饰,并研究LEP-2a及其衍生物对STZ诱导的糖尿病小鼠的心血管保护作用。通过高效凝胶渗透色谱(HPGPC)检测LEP-2a为均一组分,且平均分子量约为1.61×106 Da。单糖组成分析结果表明,LEP-2a由甘露糖和半乳糖组成,且摩尔比为3.8:1.0。甲基化分析、FT-IR分析和NMR分析结果表明,LEP-2a的主链由1,2-linked-α-D-Manp、1,2,6-linked-α-D-Manp、1,3,4-linked-α-D-Manp和1,3-linked-β-D-Galp组成。体外抗肿瘤活性研究表明,在浓度为100、200、300和400μg/m L时,LEP-2a能够显著增强5-氟尿嘧啶对Hela细胞的抑制作用。分别对LEP-2a进行乙酰化和苯甲酰化修饰,获得取代度为0.220的乙酰化LEP-2a(ALEP-2a)和取代度为0.574的苯甲酰化LEP-2a(BLEP-2a)。FT-IR分析和13C NMR分析结果进一步证实了乙酰化和苯甲酰化修饰成功。基于13C NMR分析的结果,我们推测出→2)-α-D-Manp-(1→的C-3、C-4和C-6位,α-D-Manp-(1→的C-2、C-3、C-4和C-6位,→2,6)-α-D-Manp-(1→的C-3和C-4位,→3,4)-α-D-Manp-(1→的C-2、C-4和C-6位,→3)-β-D-Galp-(1→的C-1、C-2、C-4和C-6位以及β-D-Galp-(1→的C-2、C-4和C-6位可能被乙酰基部分取代;→3)-β-D-Galp-(1→的C-1位被完全苯甲酰化,→2)-α-D-Manp-(1→、α-D-Manp-(1→和→2,6)-α-D-Manp-(1→的C-4位以及→2)-α-D-Manp-(1→、α-D-Manp-(1→、→3,4)-α-D-Manp-(1→、→3)-β-D-Galp-(1→和β-D-Galp-(1→的C-6位可能被苯甲酰基部分取代。通过腹腔注射链脲佐菌素(STZ)构建糖尿病小鼠模型,并研究不同剂量LEP-2a、ALEP-2a和BLEP-2a对糖尿病小鼠的心血管保护作用。结果表明,LEP-2a及其衍生物具有心血管保护作用且存在剂量-效应关系;与模型组相比,治疗组小鼠心脏、肾脏和肝脏指数显著降低,心脏组织中超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-PX)和过氧化氢酶(CAT)活性显著增加,丙二醛(MDA)水平显著降低,血清中晚期糖基化终产物(AGEs)、高敏C反应蛋白(hs-CRP)、可溶性细胞间粘附分子1(sICAM-1)和内皮素1(ET-1)水平显著降低,一氧化氮(NO)水平显著增加;与同剂量的LEP-2a相比,ALEP-2a和BLEP-2a表现出更显著的心血管保护作用。此外,组织病理学观察显示,治疗组小鼠受损心肌结构得到改善。综上所述,LEP-2a及其衍生物的作用机制可能与改善抗氧化防御系统、抑制氧化应激以及促进受损心肌再生有关。
[Abstract]:In this study, a new extracellular polysaccharide (Lep-2aa) was isolated and purified by fermentation and purification. The structure of LEP-2a was characterized by monosaccharide composition analysis, methylation analysis, Fourier transform infrared spectroscopy (FTIR) and nuclear magnetic resonance spectroscopy (NMR). The antitumor activity of LEP-2a was evaluated in vitro, the acetylation and benzoylation of LEP-2a were modified, and the cardiovascular protective effects of LEP-2a and its derivatives on STZ-induced diabetic mice were studied. LEP-2a was homogenized by high performance gel permeation chromatography (HPGPC), and the average molecular weight was about 1.61 脳 106 Da. The results of monosaccharide composition analysis showed that the LEP-2a was composed of mannose and galactose, and the molar ratio was 3.8: 1.0. The results of FT-IR and NMR analysis of methylation analysis show that the main chain of LEP-2a is composed of 1k2-linked- 伪 -D-Manpan- 1, 6-linked- 伪 -D-Manpan- (34-linked- 伪 -D-Manp) and 1-3-linked- 尾 -D-Galp (1-3-linked- 尾 -D-Galp). In vitro antitumor activity studies showed that the inhibitory effect of 5-fluorouracil on Hela cells was significantly enhanced at concentrations of 100200300 渭 g / mL and 400 渭 g / mL. The results of acetylation and benzoylation of LEP-2a with a degree of substitution of 0.220 and benzoylation of LEP-2a with a degree of substitution of 0.574 were obtained respectively. The results of FT-IR and 13C NMR further confirmed that the modification of acetylation and benzoylation was successful. 鍩轰簬13C NMR鍒嗘瀽鐨勭粨鏋，

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