魔芋葡甘聚糖DEAE阴离子交换介质的制备及表征

以交联的魔芋葡甘聚糖微球(KGM)为基质,制备了DEAE型的阴离子交换层析介质,考察了反应物浓度、温度、时间等反应因素对魔芋葡甘聚糖层析介质离子交换容量的影响,确定最优反应参数.结果表明,层析介质的离子交换容量达到3.42mmol/g,蛋白吸附容量高达126.84mg/mL,蛋白洗脱率为90.42%.     

交联羧甲基魔芋葡甘聚糖空心微球制备及应用

以魔芋葡甘聚糖(KGM)为壁材,通过羧甲基化并经乳状液化学交联制备球体较为完好的羧甲基魔芋葡甘聚糖(CMKGM)空心微球.通过扫描电镜(SEM)和透射电镜(TEM)对CMKGM空心微球的形貌和粒径进行了表征.结果表明,KGM溶胶浓度为0.5%-1%、乳化剪切速率为8000r/min、环氧氯丙烷(ECH)为5ml时,可以得到粒径均匀、形貌规整、溶胀性能良好的CMKGM空心微球.增大溶液的pH和羧甲基试剂的用量均有利于CMKGM空心微球对水中Cu2+的吸附.     

交联羧甲基魔芋葡甘聚糖吸附重金属离子的研究

以异丙醇为分散剂,环氧氯丙烷为交联剂,在碱性介质中由一氯乙酸和魔芋葡甘聚糖(KGM)反应,制备了取代度为0.265和0.550的两种交联羧甲基魔芋葡甘聚糖(CMKGM),并将其用于吸附溶液中Cu2 、Pb2 和Cd2 。结果表明,CMKGM对3种重金属离子的吸附约在20min内达到平衡,与金属离子类型无关,吸附遵从二级动力学方程;pH对吸附量影响较大,适宜范围为5~6;吸附能较好地服从Langmuir等温吸附方程,CMKGM(DS=0.550)吸附Pb2 的最大吸附容量(Qm)为41.7mg/g,Langmuir常数(b)为0.305mg/L,均大于Cu2 和Cd2 相应值;再生后的CMKGM吸附性能好,脱吸附百分率高。     

酶催化魔芋葡甘聚糖的可控降解

魔芋葡甘聚糖 (KGM)是一种来自植物的天然高分子 .它具有优异的可生物降解性和生物相容性 ,并具有许多独特的生理和药理功能 .本实验首先测定了 β 甘露糖酶在不同条件 (温度、pH值、介质 )下的活性 ,发现 β 甘露糖酶在 5 0℃左右 ,pH 9 4附近 ,乙醇含量低于 5 %的水介质中具有较高的活力 ;而在pH 7 0以下 ,或温度低于 3 0℃ ,或加入 2 0 %乙醇的条件下均基本上失活 .在此研究基础上 ,探讨了 β 甘露糖酶催化KGM降解反应的规律 ,通过调节反应条件制备了一系列分子量不同的降解样品 ,并确定了KGM的分子量与特性粘数之间的关系为 :[η]=5 0 6× 1 0 - 4M0 754w ,使得酶催化KGM的可控降解成为可能 ,从而为深入研究KGM及其衍生物的结构与性能 ,扩展其应用领域奠定了良好的理论和实验基础     

基于脱乙酰基魔芋葡甘聚糖固定酶的H2O2生物传感器

对魔芋葡甘聚糖进行了脱乙酰基改性处理,并用红外光谱表征了其脱乙酰基前后的结构变化.探讨了脱乙酰基魔芋葡甘聚糖溶胶-凝胶的制备条件对其成膜性能及酶固定化的影响.将脱乙酰基魔芋葡甘聚糖用于辣根过氧化物酶的固定,制备出了H2O2生物传感器.传感器在优化的工作条件下线性范围为1.0×10-6 ～6.0×10-4 mol/L,检出限为8×10-7 mol/L,稳定性良好.实验表明脱乙酰基魔芋葡甘聚糖是一种适用于生物传感器领域酶固定化的优良材料.     

分子量及酰基供体对酶促魔芋葡甘聚糖酰化反应的影响

研究了在有机介质叔丁醇中魔芋葡甘聚糖(KGM)的分子量及酰基供体对固定化脂肪酶Novozym 435催化KGM乙酰化反应的影响.KGM的分子量对酶促其酰化反应的活性及产物取代度有显著影响.随着KGM分子量的增大,酶催化反应的速率逐渐下降,产物的取代度逐渐减小.KGM分子量对该反应的影响与不同分子量KGM的溶解度、体系粘度、空间位阻及颗粒形态等因素有关.以不同链长的脂肪酸乙烯酯为酰基供体时,随着酰基供体中脂肪酸碳链的增长,酶促KGM酰化反应速率逐渐下降,产物的取代度逐渐减小,且该酰化反应具有高度的区域选择性,反应均发生在C6-OH上.     

羟丙基甲基纤维素与魔芋葡甘聚糖复配体系相互作用的研究

采用黏度测试、振荡测试以及动态力学分析方法,以结构典型的羟烷基烷基纤维素醚———羟丙基甲基纤维素(HPMC)与魔芋葡甘聚糖(KGM)混合体系为研究对象,通过对两种天然多糖混合水溶液表观黏度、流变性及溶液动态力学参数的变化,分析了结构差异性天然多糖大分子间相互作用的机理并建立分子模型.实验表明,HPMC-KGM复配后,KGM分子链上未被取代的甘露糖单元与HPMC分子链上憎水性基团通过疏水作用形成弱交联的疏水缔合区域,而形成以HPMC分子为骨架并与KGM分子相联结的结构.HPMC与KGM按质量比3∶1到1∶1比例范围内复配效应最为明显.通过疏水缔合作用,缓解KGM分子在高温条件下的降解,还对HPMC的热凝胶化作用起到延缓效果.     

纤维素酶降解-离子色谱法测定魔芋中的亚硝酸盐

建立了以纤维素酶降解魔芋葡甘聚糖释放阴离子,然后采用离子色谱测定亚硝酸盐含量的方法。考察了纤维素酶降解葡甘聚糖的最佳条件,结果表明:在50 ℃、pH 5.5条件下酶解24 h,可以释放出被包埋的亚硝酸盐。NO-2的检出限为0.039 mg/L,回收率为96.7%～97.8%。运用本法对施用稀土微肥后魔芋中亚硝酸盐的含量进行测定,发现施用稀土微肥能够在一定程度上降低魔芋中亚硝酸盐的含量。     

固定化辣根过氧化物酶在有机/水混合溶液中的电化学

用魔芋多糖(KGM)将辣根过氧化物酶(HRP)固定在玻碳电极(GCE)表面, 制备了HRP-KGM膜修饰电极. 在乙醇等亲水性有机溶剂与水的混合溶液中, 包埋在KGM中的HRP 可以与电极发生直接电子传递, 且能催化还原过氧化氢、氢过氧化异丙基苯、氢过氧化叔丁基、过氧化丁酮等过氧化物. HRP-KGM膜修饰电极具有较好的稳定性和重现性, 可用于这些物质的定量检测.     

魔芋葡甘聚糖功能材料研究与应用进展

为了更好地开发利用魔芋葡甘聚糖(KGM),本文阐明了KGM独特的结构、优良的理化性质,分析了其结构与性能的关系, 综述了在食品、化工、医药、石油钻探等应用领域中形成的以KGM为基本组成的功能材料研究进展。并对近几年国内外KGM在该领域的研究动向及其应用前景进行了探讨。     

Electroactive hemoglobin-surfactant-polymer biomembrane-like films

Polyions complex 2C12N+ PSS- was prepared by reacting poly(sodium styrenesulfonate) (Na+ PSS-) with didodecyldimethylammonium bromide (2Cl2N+ Br-). Stable thin films made from 2C12N+ PSS- with incorporated redox protein hemoglobin (Hb) on pyrolytic graphite (PG) electrodes were then characterized by electrochemistry and other techniques. Cyclic voltammetry (CV) of Hb-2C12N+ PSS- films showed a pair of well-defined and nearly reversible peaks for HbFe(III)/Fe(II) couple at about -0.17 V vs. saturated calomel electrode (SCE) in pH 5.5 buffers. The electron transfer rate between Hb and PG electrode was greatly facilitated in microenvironment of 2C12N+ PSS- films. Positions of Soret absorption band suggest that Hb keeps its secondary structure similar to its native state in 2C12N+ PSS- films at the medium pH. The results of X-ray diffraction and differential scanning calorimetry (DSC) suggest synthesized lipid 2C12N+ PSS- films have an ordered bilayer structure intercalated between PSS- polyion layers, and the incorporated Hb expands the layer spacing of the films. HbFe(I), a highly reduced form of Hb, might also be produced in these films at about -1.09 V, and could be used to catalytically reduce organohalide pollutants.     

碳糊电极上无机膜固载血红蛋白的直接电化学

报道了用硅溶胶-凝胶(Sol-gel)膜将血红蛋白(Hb)固载于碳糊电极上的直接电化学行为.研究结果表明,Hb-Sol-gel修饰的碳糊电极在pH=7.0的缓冲溶液中于-0.275V(vs.Ag/AgCl)处有一对可逆的循环伏安氧化-还原峰,为Hb血红素辅基Fe(Ⅲ)/Fe(Ⅱ)电对的特征峰.HbFe(Ⅲ)/Fe(Ⅱ)电对的式量电位在pH5.0～11.0范围内与溶液pH值呈线性关系,表明Hb的电化学还原很可能是一个质子伴随着一个电子的电极过程.FTIR光谱证实,Sol-gel膜对Hb的固载没有破坏其天然结构.Hb-Sol-gel修饰的碳糊电极能够催化还原H₂O₂,可望将其用于制作第三代生物传感器.     

血红蛋白在碳纳米管修饰碳糊电极上的直接电化学行为

利用吸附法将血红蛋白(Hb)固定在碳纳米管修饰碳糊电极表面,制成稳定的固载Hb碳纳米管修饰电极,研究了Hb在碳纳米管修饰电极上的直接电化学行为.固载Hb的碳纳米管修饰电极在pH=7.0的PBS（磷酸盐缓冲溶液）中有一对相当可逆的循环伏安氧化还原峰,为Hb血红素辅基Fe(Ⅲ)/Fe(Ⅱ)电对的特征峰.式电位为－0.160 V(vs SCE),随扫描速度变化很小.电子转移数为1.021,近似为一个辅基发生电子转移.Hb在碳纳米管修饰电极表面的电子转移常数为0.0816 s－1,远大于亚甲蓝作媒介体时Hb的电子转移反应速率常数.应用于过氧化氢、三氯乙酸和硝基苯等的电催化还原,固定在碳纳米管修饰碳糊电极的血红蛋白表现出稳定且较高的催化活性.     

NO* release from MbFe(II)NO and HbFe(II)NO after oxidation by peroxynitrite

In this work, we showed that the reaction of peroxynitrite with MbFe(II)NO, in analogy to the corresponding reaction with HbFe(II)NO (Herold, S. Inorg. Chem. 2004, 43, 3783-3785), proceeds in two steps via the formation of MbFe(III)NO, from which NO* dissociates to produce iron(III)myoglobin (Mb = myoglobin; Hb = hemoglobin). The second-order rate constants for the first steps are on the order of 10(4) and 10(3) M(-1) s(-1), for the reaction of peroxynitrite with MbFe(II)NO and HbFe(II)NO, respectively. For both proteins, we found that the values of the second-order rate constants increase with decreasing pH, an observation that suggests that HOONO is the species responsible for oxidation of the iron center. Nevertheless, it cannot be excluded that the pH-dependence arises from different conformations taken up by the proteins at different pH values. In the presence of 1.2 mM CO2, the values of the second-order rate constants are larger, on the order of 10(5) and 10(4) M(-1) s(-1), for the reaction of peroxynitrite with MbFe(II)NO and HbFe(II)NO, respectively. The pH-dependence of the values for the reaction with MbFe(II)NO suggests that ONOOCO2- or the radicals produced from its decay (CO3*-/NO2*) are responsible for the oxidation of MbFe(II)NO to MbFe(III)NO. In the presence of large amounts of nitrite (in the tens and hundreds of millimoles range), we observed a slight acceleration of the rate of oxidation of HbFe(II)NO by peroxynitrite. A catalytic rate constant of 40 +/- 2 M(-1) s(-1) was determined at pH 7.0. Preliminary studies of the reaction between nitrite and HbFe(II)NO showed that this compound also can oxidize the iron center, albeit at a significantly slower rate. At pH 7.0, we obtained an approximate second-order rate constant of 3 x 10(-3) M(-1) s(-1).     

Direct electrochemistry and electrocatalysis of hemoglobin immobilized in bimodal mesoporous silica and chitosan inorganic–organic hybrid film

Hemoglobin modified electrode was successfully fabricated to realize direct electrochemistry by immobilizing of Hemoglobin (Hb) in bimodal mesoporous silica (BMS) and chitosan (CS) inorganic–organic hybrid film. Here, BMS acted as a support to immobilize Hb due to its large pores and CS acted as a binder to increase film adherence and stabilizer to prevent the leakage of Hb. The resulting electrode (Hb/BMS/CS) gave a well-defined, reversible redox couple for HbFe(III)/Fe(II) with a formal potential of about −0.32 V (vs. Ag/AgCl) in pH 7.0 phosphate buffer solution. Hb/BMS/CS electrode showed a better electrocatalytial performance to H₂O₂ with wider linear detection range, lower detection limit, and higher sensitivity than that at electrode without BMS. The improved electrocatalytic performance for Hb/BMS/CS electrode was possibly contributed to BMS bimodal structure, whose large pores with 10–40 nm provide favorable conditions for protein immobilization and small pores with 2–3 nm avoid the mass-transfer limitations. In addition, UV–Vis and FTIR spectra indicated that Hb well maintained its native structure in the hybrid film.     

Direct electrochemistry and electrocatalysis of hemoglobin entrapped in composite matrix based on chitosan and CaCO3 nanoparticles

The direct electron transfer between hemoglobin (Hb) and the underlying glassy carbon electrode (GCE) can be readily achieved via a high biocompatible composite system based on biopolymer chitosan (CHT) and inorganic CaCO₃ nanoparticles (nano-CaCO₃). Cyclic voltammetry of Hb-CHT/nano-CaCO₃/GCE showed a pair of stable and quasi-reversible peaks for HbFe(III)/Fe(II) redox couple in pH 7.0 buffer. The electrochemical reaction of Hb immobilized in CHT/nano-CaCO₃ composite matrix exhibited a surface-controlled process accompanied by electron and proton transfer. The electron transfer rate constant was estimated to be 1.8 s⁻¹. This modified electrode showed a high thermal stability up to 60 °C. The apparent Michaelis–Menten constant was calculated to be 7.5 × 10⁻⁴ M, indicating a high catalytic activity of the immobilized Hb toward H₂O₂. The interaction between Hb and this nano-hybrid material was also investigated using FT-IR and UV–vis spectroscopy, indicating that Hb retained its native structure in this hybrid matrix.     

Pulse radiolysis studies of the reactions of CO3*- and NO2* with nitrosyl(II)myoglobin and nitrosyl(II)hemoglobin

The reactions of carbonate radical anion [CO3*-, systematic name: trioxidocarbonate*1-] with nitrosyl(II)hemoglobin (HbFe(II)NO) and nitrosyl(II)myoglobin (MbFe(II)NO) were studied by pulse radiolysis in N2O-saturated 0.25 M sodium bicarbonate solutions at pH 10.0 and room temperature. The reactions proceed in two steps: outer-sphere oxidation of the nitrosyliron(II) proteins to their corresponding nitrosyliron(III) forms and subsequent dissociation of NO*. The second-order rate constants measured for the first reaction steps were (4.3 +/- 0.2) x 10(8) and (1.5 +/- 0.3) x 10(8) M(-1) s(-1), for MbFe(II)NO and HbFe(II)NO, respectively. The reactions between nitrogen dioxide and MbFe(II)NO or HbFe(II)NO were studied by pulse radiolysis in N2O-saturated 0.1 M phosphate buffer pH 7.4 containing 5 mM nitrite. Also for the reactions of this oxidant with the nitrosyliron(II) forms of Mb and Hb a two-step reaction was observed: oxidation of the iron was followed by dissociation of NO*. The second-order rate constants measured for the first reaction steps were (2.9 +/- 0.3) x 10(7) and (1.8 +/- 0.3) x 10(7) M(-1) s(-1), for MbFe(II)NO and HbFe(II)NO, respectively. Both radicals appear to be able to oxidize the iron(II) centers of the proteins directly. Only for the reactions with HbFe(II)NO it cannot be excluded that, in a parallel reaction, CO3*- and NO2* first react with amino acid(s) of the globin, which then oxidize the nitrosyliron(II) center.     

基于聚亚甲基蓝-碳纳米管复合膜固载血红蛋白的H2O2生物传感器

采用吸附和电化学聚合修饰方法,制得了聚亚甲基蓝-碳纳米管聚合膜玻碳电极(PMB-MWNTs/GCE),再将血红蛋白(Hb)固定在PMB-MwNTs/GCE表面,制备了稳定的Hb/PMB-MwNTs//GCE的H2O2生物传感器,并用循环伏安法对修饰电极的生物电催化行为进行了表征.研究结果表明,固定在PMB-MWNTs/...     

生物分子电催化的研究(Ⅲ)——血红蛋白在甲酚固紫修饰碳糊电极上的电催化

血红蛋白是脊椎动物红细胞内的呼吸蛋白,但由于其电活性中心不容易暴露以及在电极表面强烈吸附造成电极的钝化,使得它在一般固体电极上的电子传递速率很慢,得不到有效的电流响应。有关血红蛋白在固体电极上的电化学行为的研究很少。众多的分析化学家都在积极寻求加速血红蛋白电子传递速度的途径,使蛋白质直接电化学分析成为可能。最近的研究表明,某些染料修饰电极可以促进血红蛋白的电极过程,从而获得有效的电流响     

Direct electron-transfer of native hemoglobin in blood: kinetics and catalysis

A novel approach that uses nature biological tissues, fish blood, for the study of the direct electron-transfer of hemoglobin and its catalytic activity for H(2)O(2) and NO(2)(-) is observed. The direct electron-transfer of hemoglobin in red blood cells in fish blood on glassy carbon electrode was observed for the first time. By simply casting fish blood on GC electrode surface and being air-dried, a pair of well-defined redox peaks for HbFe (III)/HbFe (II) appeared at about -0.36 V (vs SCE) at the fish blood film modified GCE in a pH 7.0 phosphate buffer solution. Ultraviolet visible (UV/VIS) characterization and the enhancement of the redox response of Hb by adding pure Hb in fish blood suggested that Hb preserved the native second structures in the fish blood film. Optical micrographs showed that the RBCs retained its integrity in blood. Hb in blood/GCE maintained its activity and could be used to electrocatalyze the reduction H(2)O(2) and NO(2)(-).