摘要
黑木耳(Auricularia auricula (L.ex Hook.)Underwood)隶属于真菌门、担子菌纲、木耳目、木耳科、木耳属,是著名药食兼用真菌,其主要活性成分—黑木耳多糖具有抗肿瘤、抗血栓、抗衰老、降血糖、降血脂、调节免疫等功能。
本文以黑木耳多糖为对象,系统地研究了其分级方法以及分子修饰(硫酸酯化、超声波辐射降解与过渡金属离子的相互作用)方法,通过对黑木耳多糖及其分子修饰多糖功能(抗氧化、抗肿瘤、抗凝血)研究,并探索其在膜材料上的应用,为黑木耳多糖、修饰多糖在医药、保健品、食品及膜材料等方面的应用奠定基础。得到以下主要结果:
1、用阳离子表面活性剂十六烷基三甲基溴化铵(CTAB)对黑木耳多糖进行分级,得到黑木耳中性多糖和酸性多糖(AAP)两部分,其中酸性多糖含量占80.33%,中性多糖占18.67%。用乙醇将黑木耳多糖分为AAP1、AAP2、AAP3三部分。
2、确定了黑木耳多糖的最佳硫酸酯化工艺:氯磺酸与吡啶体积比5:1;多糖溶液与酯化试剂的体积2:1:酯化温度为50℃,酯化反应时间为1.5h。FTIR光谱显示硫酸基与AAP形成硫酸酯化合物。并制备了取代度分别为0.32、0.64、1.0的三个黑木耳多糖硫酸酯AAP-S_1、AAP-S_2、AAP-S_3。
3、黑木耳多糖(AAP)对ZnSO_4·7H_2O中Zn(Ⅱ)的螫合能力最强,其最佳螯合工艺为:Zn(Ⅱ)的初始浓度为3mg/mL、pH值为4.0、黑木耳多糖与Zn(Ⅱ)的比值为4:1、螯合6h。制备了黑木耳多糖(AAP)与5种过渡金属离子(Fe~(2+)、Co~(2+)、Ni~(2+)、Cu~(2+)、Zn~(2+))的螯合物。
4、黑木耳多糖经超声波辐射降解后,随时间的增加,黏度逐步降低、溶解性增加,葡萄糖醛酸含量基本保持不变。黑木耳多糖经超声波辐射20min、40min、60min后,制备了三种多糖AAP-US_1、AAP-US_2、AAP-US_3
5、硫酸酯化黑木耳多糖对金黄色葡萄球菌、大肠杆菌有不同程度的抑制,抑制作用可达16~20h。对黑曲霉的孢子萌发也有一定的抑制现象。
AAP-S有直接清除氧自由基及提高机体抗氧化的功能。在生物体内可以通过显著地提高SOD、GSHPX、T-AOC的活力,降低MDA的含量,提高机体非特异性免疫器官—胸腺、脾脏的重量等作用,达到清除自由基的功能。
硫酸酯化黑木耳多糖对肿瘤细胞表现出显著的抑制功能,并表现出量效关系,对癌细胞的最大抑制率出现在72h,可达49.6%。抗凝血活性存在量效关系,并随硫酸化取代度的增加,抗凝血活性逐渐增强。
6、超声波改性黑木耳多糖对除羟自由基、超氧离子自由基均有一定的清除作用;对小鼠的生长无明显不良影响,并可以显著地促进胸腺和脾脏的生长;可以不同程度地提高小鼠血清T-AOC的能力、升高SOD、GSH-PX的活力、降低MDA含量。其抗氧化能力呈量效关系,即随剂量的增加,抗氧化能力逐渐提高。
Auricularia auricular (Lex Hook.) Underw.) is known to be a highly nutritious and officinal edible fungus. Polysaccharide, as one of main active components extracting from A. auricular, has various functions such as antitumor, antithrombus, anticoagulation, anticaducity, decreasing blood sugar and blood fat, and modulating immunity, and so on.
This paper systematically studied that fractionation methods and molecule modification (Sulfated esterification and the interaction of ultrasonic radiation degradation and transition metal ions) of polysaccharide of A. auricula. Through studies on the functions of polysaccharide and molecule modified polysaccharide in A. auricula (antioxidation, antitumor and anticoagulation), their applications on membrane materials were studied, which established a basis for the applications of polysaccharide and modified polysaccharide of A. auricula in
it
medicine, health supplies, foodstuffs, and membrane materials, etc. The main results are as follows:
1 The polysaccharides of A. auricula were fractionated by the cation surface active agent-CTAB, and the neutral (18.67%) and acid polysaccharides (80.33%) of A. auricula (AAP) were obtained. The polysaccharide of A. auricula was divided into three parts of AAP1, AAP2, and AAP3.
2 The optimum sulfated esterifing technics was determined: the volume ratio of chlorosulfonic acid and pyridine is 5:1, the volume ratio of polysaccharide solution and esterifing reagents is 2:1, esterifing temperature is 50°C, and esterifing reaction time is 1.5h. FTIR spectra show that sulfate and AAP may form sulfated compounds. And the three AAP sulfates of AAP-S1, AAP-S2 and AAP-S3, whose substitutions are 0.32, 0.64 and 1.0, respectively, were prepared.
3 The polysaccharide of A. auricular (AAP) has best chelation on the Zn (II) of ZnSO_4·7H_2O, and the optimum chelated technics: initial concentration of Zn (II) is 3mg/mL, pH value is 4.0, the volume ratio of polysaccharide and Zn (II) is 4:1, and chelating reaction time is 6h. The chelates of five kinds of transition metal ions (Fe~(2+), Co~(2+), Ni~(2+), Cu~(2+) and Zn~(2+)) with AAP were prepared.
4 After the AAP is degraded by ultrasonic radiation, and along with the increase of time, its viscosity reduces gradually, dissolving quality increases, and the content of glucose uronic acid keeps constant. AAP was radiated by ultrasonic for 20min, 40min, and 60min separately, and three polysaccharides of AAP-US1, AAP-US2 and AAP-US3 were gained.
5 The antimicrobial tests indicate that sulfated AAP has a certain of inhibition for the yellow staphylococcus and E. coli, with an inhibiting time of 16~20h. Moreover, AAP also has a
引文
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