去除内毒素的柱亲和介质与膜亲和介质的制备和特征

以琼脂糖和尼龙６６微孔滤膜为原料，发展了两种用于去 内毒素的不同类型的亲和介质。在对原料进行化学改性后，键合了含六个碳原子的间隔臂和组氨酸亲和配基。它们对内毒素的亲和容量分别为２．０ｍｇ／ｇ和０．２ｍｇ／ｇ，内毒素和介质之间的表观解离常数分别为８．１＊１０＾－１０ｍｏｌ／Ｌ和３．０＊１０＾－８ｍｏｌ／Ｌ。     

固定化铜离子亲和膜色谱柱吸附血红蛋白的研究

将纤维素滤纸进行碱处理及环氧活化、偶联亚氨基二乙酸、固定化铜离子等处理,并将其装入自制的色谱柱管,制得固定化铜离子亲和膜色谱柱。该柱可用于吸附血红蛋白(hemoglobin,Hb),吸附率可达到90%以上。考察了上样量、pH值、温度、上样速度等因素对固定化铜离子亲和膜吸附Hb的影响。实验结果表明,固定化铜离子亲和膜色谱柱吸附血红蛋白的最佳条件为：室温下实验,缓冲体系的pH值控制在6~8,上样速度0.5~1.0 mL/min,上样量为3.16~7.90 mg/g。     

固定化金属螯合亲和膜色谱柱制备及纯化铜锌超氧化物歧化酶的研究

 以大孔纤维素滤纸为基质 ,通过碱处理、环氧活化、偶联亚氨基二乙酸二钠、固定化Cu2 +,装柱后制得固定化金属螯合亲和膜色谱柱。对Cu/Zn SOD粗品进行了纯化研究 ,将其比活从 645U/mL提高到 6882U/mL ,纯化倍数为 1 0 7,蛋白回收率为 92 3% ,活性回收率达 985%。设计了一种解决金属离子泄露问题的方案 ,将Cu2 +泄露量降至 86μg/L。     

金属螯合亲和膜吸附分离与纯化溶菌酶的研究（Ⅱ）——不同金 …

采用低温氧或氨等离子体法改性聚丙烯微孔膜，基于等离子体改性膜的活化，偶联及螯合过程的机理，制备了Ｆｅ＾３＋、Ｎｉ＾２＋、Ｃｕ＾２＋等金属离子螯合亲和膜，并用于溶菌酶的吸－脱附实验。两种膜的重复吸－脱附性能相近，而Ｆｅ＾３＋螯合亲和膜基本上不能用于重复吸－脱附实验，采用补充金属螯合离子，能部分恢复亲和膜对溶菌酶的吸附量，是实现亲和膜重复使用的有效方法。     

聚偏氟乙烯中空纤维亲和膜分离γ—球蛋白的研究（I）——聚偏氟乙烯中空纤维亲和膜的制备及其吸附性能

对聚偏氟乙烯(PVDF)中空纤维膜进行表面亲水化处理，再经活化、偶联制得了带有苯丙氨酸(Phe)的PVDF中空纤维亲和膜。制备过程中以光电子能谱(XPS)及红外光谱分析膜表面基团的变化，经亲水化处理及活化偶联后，膜表面的O和N含量分别从5．6％和0.6％增加到16．5％和3．0％，而F含量由38．3％降低为23．0％。将该亲和膜用于室温下血浆中γ—球蛋白的分离实验，在pH＝7．4，离子强度I＝0.18的缓冲液体系及进料质量浓度为1．0mg／mL条件下，该亲和膜对γ—球蛋白的吸附量可达到5．85mg／g膜；与市售γ-球蛋白相比，血浆中提取纯度可达83.9%。     

纤维素亲和膜用于内毒素的去除

报道了用于内毒素去除的３种新型纤维素亲和膜的制备及应用,并对它们的性能进行了比较。结果表明,这３种亲和膜均能有效去除盐溶液中的内毒素,去除率在９０％以上。对人血清白蛋白溶液,季铵盐和壳聚糖亲和膜对内毒素的去除率较高,仍然在９０％以上,金属螯合物亲和膜的去除效果不如前两种,去除率仅为８３．３％。实验选择不同的亲和膜,考察了它们对内毒素的吸附容量、样品处理量以及再生效果,结果表明吸附量在 ２．４×１０６ＥＵ／ｇ以上,样品处理量较大,适合用于小批量医药制剂的生产过程;再生效果较好,可重复使用。     

鸟嘌呤—铜配合物在汞电极表面上的电吸附性的研究

在０．０２ｍｏｌ／Ｌ ＮａＨＣＯ３－０．０５ｍｏｌ／Ｌａ２ＳＯ４介质中、用极谱法和伏安法研究表明,铜－乌嘌呤配合物在汞电极表面上的吸附服从Ｌａｎｇｍｕｉｒ吸附,测得其饱和吸附量Γｍ＝１．１×１０＾－１０ｍｏｌ／ｃｍ＾２,吸附系数β＝４．７×１０＾４,吸咐自由能△Ｇ°＝－３６．６ｋＪ／ｍｏｌ。在８×１０＾－６ｍｏｌ／Ｌ Ｃｕ＾２＋离子存在下,可用示差脉冲阴极吸附溶出法测定８×１０＾－１０～１×１０＾     

聚芳醚酮膜的溶剂吸附与诱导结晶行为

研究了聚芳醚酮系列在１，２－二氯乙烷中的吸附，及溶剂与温度诱导结晶行为。得到６０℃时的最大吸附量Ｃ∞及假扩散系数Ｄ为：ＰＥＥＫ，Ｃ∞＝４６％，Ｄ＝３．１０×１０＾－１２ｍ＾２·ｓ＾－１；ＰＥＥＫＫ，Ｃ∞＝５０％，Ｄ＝６．５７×１０＾－１２ｍ＾２·ｓ＾－１；ＰＥＫＫ，Ｃ∞＝５５％，Ｄ＝９．４１×１０＾－１２ｍ＾２·ｓ＾－１。实验表明，吸附及脱附行为均与分子链刚性或羰基含量有关，膜的分子链受溶剂作用而     

α—胰凝乳蛋白酶的固定化及其对DL—苯丙氨酸的光学拆分

以氨丙基多孔球为载体，戊二醛为交联剂制得固定化α－胰凝乳蛋白酶。其含酶量为３．６ｍｇ／ｇ，米氏常数Ｋｍ＝１．２×１０＾－３ｍｏｌ／Ｌ，最适宜温度为３５℃，ｐＨ值为７．０，对热、酸碱、甲醇以及尿素的稳定性有较大提高，Ｃａ＾２＋激活效应明显。用该固定化酶连续拆分ＤＬ－苯丙氨酸制备Ｌ－苯丙氨酸时，拆分产率高于９０％，产品纯度超过９６％；连续使用２个月后仍能保持较高酶活力。     

二溴邻羧基偶氮氯膦光度测定铝合金中微量锶

本介绍在一定量钙、钡共存下测定锶的新方法。在０．４８ｍｏｌ／Ｌ Ｈ２ＳＯ４介质中，二溴邻羧基偶氮氯膦与锶形成２：１的蓝色络合物，最大吸收波长为６３０ｎｍ。摩尔吸光系数为３．２×１０＾４Ｌ．ｍｏｌ＾－１．ｃｍ＾－１，锶量在０－１ｍｇ／Ｌ范围内符合比耳定律，经过二氧化锰共沉淀分离富集后，可测定各种复杂铝合金样品中痕量锶。     

Immobilized metal affinity composite membrane based on cellulose for separation of biopolymers

Ｕｐｔｏｄａｔｅ,ｒａｐｉｄｐｕｒｉｆｉｃａｔｉｏｎｏｆｍｉｘｅｄｐｒｏｔｅｉｎｉｎｌａｒｇｅｓｃａｌｅｈａｓｂｅｅｎａｎｉｍｐｏｒｔａｎｔｒｅｓｅａｒｃｈｐｒｏｊｅｃｔｉｎｂｉｏｅｎｇｉｎｅｅｒｉｎｇｐｒｏｄｕｃｔｐｒｏｃｅｓｓｉｎｇ.Ｉｍｍｏｂｉｌｉｚｅｄｍｅｔａｌｉｏｎａｆｆｉｎｉｔｙｃｈｒｏｍａｔｏｇｒａｐｈｙ(ＩＭＡＣ)ｉｓａｎｅｆｆｉｃｉｅｎｔｍｅｔｈｏｄｅｘｔｅｎｓｉｖｅｌｙｕｓｅｄｆｏｒａｆｆｉｎｉｔｙｐｕｒｉｆｉｃａｔｉｏｎｏｆｂｉｏｌｏｇｉｃａｌｌｙａｃｔｉｖｅｓｕｂｓｔａｎｃｅ…     

新型亲和膜色谱用于去除胆红素的研究

 以纤维素膜为亲和基质 ,接枝后分别以聚赖氨酸和季铵盐为配基 ,制备了两种亲和膜介质 ,用于去除磷酸缓冲液 (pH 7 4)及人血白蛋白 (HSA)溶液中的胆红素。实验结果表明 ,该方法对磷酸缓冲液中胆红素的去除率能达到 70 %以上 ;对HSA溶液中的胆红素 ,其去除率稍低 ,但在较低浓度的HSA溶液中 ,也能获得高于 5 0 %的去除率。并对亲和膜与胆红素间的亲和作用进行了研究 ,考察了温度、流速、HSA浓度及胆红素初始浓度等相关条件对去除效果的影响。结果表明 ,在较高的温度条件下 ,去除效果较佳 ;在较高的初始浓度情况下 ,胆红素的吸附速率较大。     

硅胶-环糊精固定金属离子亲合吸附剂的制备及性能

以硅胶键合β-环糊精(-βCD)为载体,亚氨基二乙酸(IDA)为螯合基,制备了新型固定金属离子亲合吸附剂.通过13C固体核磁、元素分析对其进行了表征.每克吸附剂分别含IDA分子158μmol,-βCD分子136μmol.研究了吸附剂对α-淀粉酶的吸附特性.     

One-pot preparation of silica-supported hybrid immobilized metal affinity adsorbent with macroporous surface based on surface imprinting coating technique combined with polysaccharide incorporated sol--gel process

A simple and reliable one-pot approach using surface imprinting coating technique combined with polysaccharide incorporated sol-gel process was established to synthesize a new organic-inorganic hybrid matrix possessing macroporous surface and functional ligand. Using mesoporous silica gel being a support, immobilized metal affinity adsorbent with a macroporous shell/mesoporous core structure was obtained after metal ion loading. In the prepared matrix, covalently bonded coating and morphology manipulation on silica gel was achieved by using one-pot sol-gel process starting from an inorganic precursor, -glycidoxypropyltrimethoxysiloxane (GPTMS), and a functional biopolymer, chitosan (CS) at the atmosphere of imprinting polyethylene glycol (PEG). Self-hydrolysis of GPTMS, self-condensation, and co-condensation of silanol groups (Si-OH) from siloxane and silica gel surface, and in situ covalent cross-linking of CS created an orderly coating on silica gel surface. PEG extraction using hot ammonium hydroxide solution gave a chemically and mechanically stabilized pore structure and deactivated residual epoxy groups. The prepared matrix was characterized by using X-ray energy dispersion spectroscopy (EDX), scanning electron microscopy (SEM) and mercury intrusion porosimetry. The matrix possessed a high capacity for copper ion loading. Protein adsorption performance of the new immobilized metal affinity adsorbent was evaluated by batch adsorption and column chromatographic experiment using bovine serum albumin (BSA) as a simple model protein. Under the optimized coating conditions, the obtained macroporous surface resulted in a fast kinetics and high capability for protein adsorption, while the matrix non-charged with metal ions offered a low non-specific adsorption.     

基于磁性微球的固定化金属亲和载体及其在蛋白质/多肽纯化中的应用

采用模板法制备的单分散磁性硅胶微球,经过表面修饰偶联上亚氨基二乙酸(IDA),与过渡金属离子Cu2 螯合,制成一种新型的磁性固定化金属亲和纯化载体。用牛血清白蛋白(BSA)作为模型进行磁性固定化金属亲和吸附蛋白的研究,结果表明,BSA在磁性亲和载体上的吸附可用Langmuir吸附方程描述,对BSA的饱和吸附量为90mg/g。将磁性亲和载体用于带有组氨酸标签的镇痛抗肿瘤多肽(analgesic-antitumorpeptide,AGAP)纯化,在未经过滤的细胞裂解液中可以将AGAP一步纯化,非特异性吸附低,操作简便,完全适用于含有组氨酸标记的重组多肽或蛋白的分离纯化。     

Elaboration,characterization and study of a new hybrid chitosan/ceramic membrane for affinity membrane chromatography

A new kind of metal affinity membrane based on a ceramic support was prepared. It was elaborated in four steps: (i) deposition of a chitosan layer in order to functionalize the ceramic support, (ii) cross-linking with epichlorohydrin to stabilise the polymer layer and to enable the grafting, (iii) iminodiacetic acid grafting, (iv) Cu²⁺ adsorption. Due to the ceramic support, this membrane is highly resistant and the chitosan layer brings its biocompatibility properties. Each step of the membrane elaboration was studied and the membrane structure was characterized. Both thin coating of the polymer on the alumina grains of the support and the chemical modification of the membrane were proved. Then, bovine serum albumin (BSA) was used as a model protein to test protein retention of the affinity membrane. The protein/membrane interactions were investigated showing that some non-specific ones are involved. Finally, the effect of buffer concentration was checked and it appears that, in the studied range, an increase of the buffer concentration entailed a limitation of the non-specific interactions inducing a better BSA recovering and a higher selectivity.     

Solution‐blown nylon 6–chitosan core–shell nanofiber for highly efficient affinity adsorption

A facile spinning‐based strategy was developed to fabricate chitosan (CS) surface nanofiber‐based affinity membranes for protein adsorption. The core–shell nanofiber mat of nylon 6–CS was prepared via coaxial solution blowing process. The nanofibers have a diameter range of 60–300 nm. The core–shell structure was confirmed by transmission electron microscopy, and CS was observed as a thin layer that uniformly adhered to the core. The dye ligand of cibacron blue F3GA (CB F3GA) was further covalently immobilized on the nanofibers with a content of 425 µmol/g. The pristine and CB F3GA‐attached mats were studied in protein adsorption. High bovine serum albumin adsorption capacities of 91.9 and 219.6 mg/g were obtained for pristine and CB F3GA‐attached mats, respectively. Given its properties of high flux rate and low pressure drop, CB F3GA‐attached nylon 6–CS nanofiber mat meets the requirements of highly effective affinity membrane chromatography. Copyright © 2016 John Wiley & Sons, Ltd.     

Analysis of protein adsorption on regenerated cellulose-based immobilized copper ion affinity membranes

Immobilized metal affinity membranes (IMAMs) were prepared by immobilizing copper ions on microporous regenerated cellulose membranes through different types of chelating agents (dentate and triazine dye). The resulting chelator utilization percentages were 95% for iminodiacetic acid, 56% for N,N,N-tris(carboxymethyl)ethylenediamine, 52% for Cibacron blue 3GA, and 140% for Cibacron red 3BA. On the other hand, triazine dyes were slightly superior to dentate chelators on metal ion utilization for protein adsorption. In batch single-protein adsorptions, the protein adsorption capacity decreased with increasing molecular size and number of accessible surface histidine residues [lysozyme>bovine serum albumin(BSA)>gamma-globulin], while the binding strength order was the opposite (gamma-globulin>BSA>lysozyme). Moreover, the proportions of specific and nonspecific bindings were evaluated by varying pH and salt concentration conditions. A large fraction of the adsorption capacity was found to come from the nonspecific interactions for the prepared IMAMs. Lastly, batch three-protein adsorptions were performed and weak adsorption competition was observed.