Separation and determination of emetine dithiocarbamate metal complexes by capillary electrophoresis with chemiluminescence detection of the tris(2,2'-bipyridine)-ruthenium(II) complex

A new cellulose-based anion-exchanger was prepared by grafting polyallylamine onto cellulose. The material was obtained by partial oxidation of a size-exclusion grade cellulose gel by aq. NaIO4, forming dialdehyde cellulose, followed by Schiff base formation with a polyallylamine (PAA, molecular mass 5000) and subsequent reduction for stabilization. Three grades of PAA-cellulose gels, with amino group contents of 0.78, 1.01 and 1.28 mmol/g cellulose, were examined for their ionic interaction with mono- and divalent carboxylic acids at pH 2.5-5.5. While the retention factor for monovalent acids was nearly proportional to the amino group content of the gel, that for divalent acids was remarkably greater for the PAA-cellulose gel than for the conventional diethylaminoethyl (DEAE) cellulose gel bearing more amino groups (1.97 mmol/g cellulose). Such high capacity can be explained by the high local density of amino groups on grafted PAA, in contrast to the random and sparse charge distribution in conventional exchangers.     

Immobilization of protein on cellulose hydrogel

A technique of immobilizing an enzyme/antibody was developed using cellulose hydrogel prepared from an aqueous alkali-urea solvent. Partial oxidation by sodium periodate activated the cellulose gel for introducing aldehyde groups. Proteins were covalently introduced to cellulose gel by a Schiff base formation between the aldehyde and the amino groups of proteins, and stabilized by a reduction of imines. Coloring reactions confirmed the high activity of the immobilized enzymes. The activity of the immobilized enzymes increased with aldehyde content, but the effect leveled off at a low degree of oxidation, at approximately 8.1 of oxidized glucose/100 glucose unit. The amount of immobilized peroxidase calculated from the activity was 8.0 ng/g for an aldehyde content of 0.18 mmol/g: 14.6 ng/g for both 0.46 mmol/g and 1.04 mmol/g. The same method could be applied to the peroxidase antibody. Thus, various active proteins could be immobilized on cellulose gels by mild and facile processing. Owing to high mechanical and chemical stability of cellulose, this technique and resulting materials are potentially useful in biochemical processing and sensing technologies.     

Hydrophile Polymergele mit reaktiven Gruppen, 4. Mitt.: Chelatharze mit schwefelhaltigen Ankergruppen auf Basis von Saccharosemethacrylaten

Chelating resins with thioglycolate anchor groups have been synthesized by reaction of sucrosemethacrylate gels with thioglycolic acid. The capacities of the gels were found to be extremely high for Ag⁺ (5,3 mmol/g) and Hg²⁺ (4,9 mmol/g). About 1 mmol Hg²⁺/g could be recovered from the gel reversibly by treatment with hydrochloric acid. The removal of Hg²⁺ from aqueous 3M alkalichloride solutions was possible with capacities of 1 mmol/g.Sucrosemethacrylate gels with primary aromatic amino group were reacted with CS₂/NH₃ to yield gels with dithiocarbamate groups. Gels with thiorea groups were prepared by reaction of the amino groups with NH₄SCN/HCl. Diazotation of the amino groups, subsequent reaction with potassium xanthogenate and hydrolysis afforded gels with thiol groups. Thiol containing gels were synthesized also by reaction of the diazotised gels with Na₂S₂ and subsequent reduction with Na₂S. Reaction of the diazotised gels with mercaptans yielded resins with thioether anchor groups. The capacities of the sulfur containing gels were found to be max. 4,1 mmol/g for Hg²⁺ and 5,9 mmol/g for Ag⁺. About 35% of the bonded Hg²⁺ could be eluted resersibly with 3N-HCl.

     

7种胺基键合硅胶的制备及其对重金属Pb2+的吸附

制备了氨丙基键合硅胶（SiO₂-N）、乙二胺-N-丙基键合硅胶（SiO₂-2N）、二乙烯三胺基键合硅胶（SiO₂-3N）、三乙烯四胺基键合硅胶（SiO₂-4N）、四乙烯五胺基键合硅胶（SiO₂-5N）、五乙烯六胺基键合硅胶（SiO₂-6N）和聚乙烯亚胺基键合硅胶（SiO₂-nN）,一步法制备的SiO₂-N和SiO₂-2N的胺基键合密度高达2.07 mmol/g和1.71mmol/g,两步法制备的SiO₂-nN的胺基键合密度为0.02mmol/g,其余胺基键合硅胶中胺基密度约为0.50mmol/g。这7种胺基键合硅胶被用于水溶液中常见重金属离子Pb²⁺的吸附研究。结果表明,在30℃条件下,分别加入10 mL 400 mg/L的Pb²⁺溶液（pH 5）和20 mg胺基键合硅胶进行吸附,10 h后,Pb²⁺吸附量达到最大,吸附过程符合Freundlich等温方程。SiO₂-N、SiO₂-2N、SiO₂-3N、SiO₂-4N、SiO₂-5N、SiO₂-6N和SiO₂-nN对Pb²⁺的吸附量依次为131.28、138.98、85.37、75.22、61.87、79.12和114.06 mg/g,这些胺基键合硅胶在吸附Pb²⁺方面均非常具有潜力。     

Ion-exchange chromatography by dicarboxyl cellulose gel.

A new column packing material for ion-exchange chromatography was prepared from cellulose gel by periodate oxidation followed by chlorite oxidation to form spatially paired carboxyl groups (dicarboxyl cellulose, DCC). The carboxyl group was quantitatively introduced to spherical cellulose gel by controlling the extent of oxidation. The DCC gels were examined for their ion-exchange activity for various amines at pH of 2.5-5.5. In this pH range, aromatic amines with acid dissociation constant (pKa) below 2.7 showed no interaction with DCC gels as expected from their lack of protonation. The amines with pKa greater than 3.3, both aromatic and aliphatic, showed strong interaction corresponding to the amount of carboxyl introduced to the gel. However, these amines showed anomalous dependence on pH of the mobile phase, showing a maximum in retention factor at around pH 4. This is in contrast with the nearly constant retention factor of these amines on conventional carboxylated cellulose packing at pH greater than 4.0. The maximum retention factor at pH 4 of DCC gel was 4-5-times greater than that of conventional gel having a similar amount of carboxyls. Since pKa of dicarboxyl groups ranges 3-5 as determined by acid-base titration, the pH giving maximum retention corresponds to the pH at which one of paired carboxyls is dissociated. Possible cause of this anomaly is presented in terms of dissociation state of dicarboxyl groups and its interaction with amines.     

新型两性球形离子交换纤维素的制备

本文在自制的球形纤维素基础上,以Ｆｅ＾２＋Ｈ２Ｏ２－二氧化硫脲为氧化还原引发系统,通过甲基丙为稀酸与丙烯酰胺的二元混合物与珠体纤维素接枝共聚,制得了含－ＮＨ２和－ＣＯＯＨ两种官能团的两性离子交换纤维素,其中阳离子交换容量在０．１５～０．６７ｍｍｏｌ／ｇ,阴离子交换容量在０．０５～０．５０ｍｍｏｌ／ｇ。该离子交换剂可用于生物大分子如蛋白质、肽的分离。     

Investigation of the structure of cellulose in LiCl/DMAc solution and its gelation behavior by small-angle X-ray scattering measurements

Cellulose gels were prepared from cellulose in lithium chloride/N,N-dimethylacetamide (LiCl/DMAc) solution. When the cellulose concentration in the solution is above the one at which cellulose molecules overlap, cellulose gels were formed. While the gel prepared by the addition of water was turbid, the one prepared by the ion exchange was colorless, transparent, and optically anisotropic. In order to explain this gelation behavior of cellulose, small-angle X-ray scattering (SAXS) measurements of the cellulose solutions and the gels were performed. The SAXS profiles of the cellulose solutions and the gels suggested that the large-scale fluctuation of the molecular chain density in the solution can be the origin of the molecular aggregates formed in the gel. Furthermore, the differences in the structure of the gels at the macroscopic and the molecular level were discussed in terms of the phase separation and the molecular association.     

Metal ion promoted hydrogels for bovine serum albumin adsorption: Cu(II) and Co(II) chelated poly[(N-vinylimidazole)-maleic acid

Poly[(N-vinylimidazole)-maleic acid] (poly(VIm-MA)), copolymeric hydrogels were prepared by gamma-irradiating ternary mixtures of N-vinylimidazole-maleic acid-water in a (60)Co-gamma source. Cu(II) and Co(II) ions were chelated within the gels at pH=5.0. The maximum adsorption capacity of the gels were 3.71 mmol/g dry gel for Cu(II) and 1.25 mmol/g dry gel for Co(II) at pH=5.0. The swelling ratios of the gels were 1200% for poly(VIm-MA), 60 and 45% for Cu(II) and Co(II)-chelated poly(VIm-MA) gels at pH=5.0 in acetate buffer solution. These affinity gels with different swelling ratios for plain poly(VIm-MA), Cu(II)-, and Co(II)-chelated poly(VIm-MA), in acetate and phosphate buffers were used in the bovine serum albumin (BSA) adsorption/desorption studies in batch reactor. The maximum BSA adsorption capacities of the gels were 0.38 g/g dry gel for plain, 0.88 g/g dry gel for Cu(II)-chelated poly(VIm-MA) and 1.05 g/g dry gel for Co(II)-chelated poly(VIm-MA) gels. Adsorption capacity of BSA by the gels was reduced dramatically by increasing the ionic strength adjusted with NaCl. More than 95% of BSA were desorbed in 10 h in desorption medium containing 0.1M of EDTA for metal ion-chelated gels at pH=4.7.     

pH-induced reversible conformational and morphological regulation of polyleucine grafted polyallylamine assembly in solution

One of the essential parts in the molecular mechanism of biological properties is the structural changes of proteins induced by stimuli. An amphiphilic copolymer, poly(L-leucine) grafted polyallylamine as a simple model of proteins, has been prepared by NCA polymerization with free amino groups of polyallylamine as an initiator. Here, we report the pH-induced reversible conformational and morphological regulation of the amphiphilic copolymer, whose hydrophobic peptide graft chains have no pH-sensitive groups, in an aqueous solution containing 50 vol % trifluoroethanol. The conformation of the poly(L-leucine) graft chain was found to be strongly pH dependent. Under acidic conditions, where electrostatic repulsion existed between the neighboring protonated amine moieties of the polyallylamine main chain, the rapid aggregation of the poly(l-leucine) graft chains was disturbed, and the peptide graft chains formed a beta-sheet structure owing to the intramolecular hydrogen bonding among the graft chains. Under this condition, the amphiphilic polymer formed amyloid-like fibrils, and then the fibrils grew into a planer plate composed of staked beta-sheets. On the other hand, under basic conditions, the poly(L-leucine) graft chains showed conformational transitions from a beta-sheet structure to an alpha-helical conformation owing to a distortion of the regular arrangement of the peptide graft chains by the conformational change of the polyallylamine main chain, whose amino groups were deprotonated. The conformational transition resulted in a disturbance of the regular sheet assembly of the amphiphilic copolymer and induced morphological changes to the amorphous globular aggregates. The pH-induced conformational and morphological changes of the poly(L-leucine) graft polyallylamine were reversible and synchronized with the protonation of the polyallylamine main chain.     

Capillary column high-performance liquid chromatographic-electrospray ionization triple-stage quadrupole mass spectrometric analysis of proteins separated by two-dimensional polyacrylamide gel electrophoresis application to cerebellar protein mapping

A method is presented for the structural characterization of proteins separated by two-dimensional poly-acrylamide gel electrophoresis (2D-PAGE). The method includes separation of a protein mixture by 2D-PAGE, recovery of proteins from the gel spots revealed by copper staining and analysis of the proteins by triple-stage quadrupole mass spectrometry using an electrospray ionization interface (ESI-TSQMS). Prior to the mass spectrometric analysis, the extracted proteins were passed through a small reversed-phase column (10 × 4.0 mm I.D.) to remove salts and gel-derived contaminants and then introduced into the mass spectrometer through a reversed-phase capillary column with 0.25 mm I.D. Application of the method to the analysis of rat cerebellar proteins suggests that the molecular mass could be accurately determined with sub-picomole amounts of protein samples derived from one or two 2D gels. The method was also useful for peptide mapping and determination of amino acid sequences of proteins micro-prepared from the 2D gel. Because 2D-PAGE has an excellent resolving power in protein separation and because capillary LC-ESI-TSQMS provides structural information with very small amounts of samples, the combined system of 2D-PAGE and capillary LC-ESI-TSQMS described here should allow wide applications to molecular studies of genes and proteins, such as identifications of protein spots on 2D gels, confirmation of gene/protein sequences and analysis of post-translational modification of proteins present naturally in tissue/cell extracts or expressed by recombinant DNA techniques.     

聚吡咯纳米线凝胶的模板制备及储能与电化学传感性能

以配位聚合物凝胶为模板,构筑均一的聚吡咯纳米线网络,聚合后经简单处理除去模板,得到性能优异的聚吡咯凝胶.结果表明,模板法合成的聚吡咯凝胶为由均一纳米线组成的三维网络结构,具有良好的力学性能、较大的比表面积及优异的电化学特性,在0.28 A/g电流密度下,比电容可达450 F/g,在2.8 A/g电流密度下充放电1000次,比电容仍可保持88.6%.聚吡咯纳米线网络凝胶经葡萄糖氧化酶负载后得到柔性传感电极,对低浓度(0.2 mmol/L)的葡萄糖具有快速响应性能,有望用于超级电容器及生物电化学传感器等领域.     

改性生物凝胶对重金属离子的吸附性能研究

以桔子汁加工残渣为原料,制备钙型和氢型生物凝胶作为吸附剂,用于去除水溶液中的重金属离子.结果表明,上述凝胶在水溶液中稳定性较好,对重金属离子的吸附性能优良.钙型凝胶的吸附选择性顺序为:Fe3 ＞Pb2 ＞Cd2 ＞Zn2 ,饱和吸附容量分别为:Pb2 、Cd2 、Zn2 均为约1.1mmol/g、Fe3 为1.5mmol/g;氢型凝胶的吸附选择性顺序为:Pb2 ＞Zn2 ＞Cd2 .钙型凝胶对Fe3 的吸附行为明显不同于氢型凝胶,钙型凝胶以离子交换机理以及Fe3 与Ca2 之间的共沉淀作用为主;而氢型凝胶对Fe3 的吸附则以离子交换机理为主.     

Enantiomeric Separations of Acidic and Neutral Compounds by Capillary Electrochromatography with β‐Cyclodextrin‐Bonded Positively Charged Polyacrylamide Gels

The novel enantiomeric separation of acidic and neutral compounds by capillary electrochromatography with β‐cyclodextrin‐bonded positively charged polyacrylamide gels was examined. The columns used are capillaries filled with a positively charged polyacrylamide gel, a so‐called monolithic stationary phase, to which allyl carbamoylated β‐CD derivatives covalently bind. The capillary wall was activated first by bifunctional reagent to make the resulting gel bind covalently inside the fused‐silica tubing. Enantiomeric separations of sixteen acidic and two neutral compounds were achieved using the above‐mentioned columns and 200 mmol dm^–3 Tris–300 mmol dm^–3 boric acid buffer (pH 8.1) as a mobile phase. High efficiencies of up to 150 000 plates m^–1 were obtained for dansyl‐DL‐amino acids. The within‐run and between‐run reproducibilities of retention time and separation factor were examined for three dansyl‐DL‐amino acids and warfarin. The relative standard deviations of the within‐run and between‐run reproducibilities of retention time were less than 1.2 and 1.3% over the six injections, respectively. Those of the separation factor were less than 0.3 and 0.2%, respectively. The gel‐filled capillaries were stable for at least four months with intermittent use.     

绵杆纤维的改性(Ⅱ)──氮杂冠醚化纤维素的制备及其对金属离子的吸附性能

经环氧活化的棉杆纤维素醚分别与3-羟基-1，5-二氯杂环庚烷和3-羟基-1，5-二氮杂环辛烷反应，制备了二种氮杂冠醚化棉杆纤维衍生物，测定了它们对金属离子的吸附性能及溶液pH值的影响。结果表明二者对Hg2 ，Cd2 有较好的吸附性能，对Cu2 的吸附-解吸-再吸附实验表明，二种氮杂冠醚化棉杆纤维衍生物具有重复使用的性能。     

Rheology and gelation of cellulose/ammonia/ammonium thiocyanate solutions

Liquid crystalline solutions of cellulose in an ammonia/ammonium thiocyanate solvent will form thermoreversible gels at temperatures below 30°C. These gels are of interest both for processing the cellulose/ammonia/ammonium thiocyanate system and because they have an unusual structure, containing neither crystalline nor covalently bonded crosslinks. Although these gels contain neither crystalline nor covalently bonded crosslinks, the dynamic rheological behavior of the system at the gel point was found to be the same as for gels with covalent or crystalline crosslinks with a loss tangent, tan δ, independent of frequency. The kinetics of the gelation process was monitored via dynamic elastic modulus, G′. All samples revealed an exponential increase in G′ with time during gelation, very different from that observed in covalently bonded or crystalline crosslinked systems. Measurements of the loss tangent enabled precise determination of the gelation time for these systems as a function of cellulose concentration and temperature. We found the gel time to be inversely related to cellulose concentration and directly related to temperature. The strong dependence of gel time on these parameters offers a windows of spinnability that can be tailored for processing high modulus cellulose fibers. © 1996 John Wiley & Sons, Inc.     

Liquid chromatography on unmodified and modified Spheron gels III. Phenolic compounds

Summary Chromatographic behaviour of phenolic compounds is studied on the ethyleneglycol methacrylate gel Spheron and on ion-exchangers produced by the chemical modification of this gels (cation exchanger Spheron S and anion exchanger Spheron DEAE) as compared to octadecyl silica. The hydrophobic effects obviously predominate in the retention mechanism on Spherons in aqueous methanolic mobile phases, but a selectivity differring from the behaviour on octadecyl silica was found for a number of phenolic compounds This is due to interactions with the functional groups in the unmodified and modified Spheron materials and may be utilized for the separation of phenols by liquid chromatography.     

新型含能纤维素基凝胶推进剂的流变性能研究

制备了以羧甲基纤维素硝酸酯(CMCN)为胶凝剂的硝基甲烷(NM)凝胶, 系统研究了CMCN 修饰基团比例变化所引起NM/CMCN凝胶流变性能差异性的规律. 探讨了凝胶的形成机理并采用线性的流变学方法研究了NM/CMCN凝胶的屈服性、触变性、蠕变性、温敏性等动态粘弹性质, 分别利用Herschel-Bulkley 模型、Burger 模型、Carreau-Yasuda模型、Andrade-Eyring 模型对凝胶的流动曲线、蠕变曲线、频率曲线和温敏曲线进行了数据拟合. 研究发现, NM/CMCN凝胶是由CMCN 大分子链上两亲性基团与有机小分子基团间通过分子链间疏水键、氢键等非共价键相互作用形成的一种物理交联网络型凝胶. 凝胶的非牛顿系数n均小于0.60. 随着硝酸酯基团含量的减少, NM/CMCN 凝胶的结构强度变弱, 表现出屈服应力逐渐减小、触变恢复性逐渐增强、粘性与弹性柔量均减小但粘性柔量比例增加的趋势; 凝胶的弹性响应性逐渐减弱而粘性响应性渐强, 内耗也逐渐增大;凝胶的活化能总体上增大, 温度稳定性变弱.     

Gel permeation properties of cellulose. I. Preliminary comparison of unmodified and crosslinked,decrystallized cotton cellulose

Decrystallized cotton cellulose was found to be have gel permeation properties comparable to those of the highly crosslinked dextran and polyacrylamide gels used in chromatography. Partial separations of pairs of sugars, ranging in molecular weight from 120 to 738, were studied by determining the optical rotations of fractions eluted from chromatographic columns, employing a sensitive automatic polarimeter. The elution volumes of individual sugars of known molecular weights provide a quantitative measure of changes produced in the cellulose polymer structure by crosslinking. A significant difference was found between unmodified cellulose and the same material crosslinked by formaldehyde treatment; although crosslinking reduced permeability to larger molecules, the crosslinked material was found to be more permeable than the untreated cellulose to compounds having molecular weights below approximately 1000.     

Preparative electrochromatography of proteins in various types of porous media

The performance of commercial and enzymatically modified size-exclusion (SE) gels in electrochromatography was compared for preparative protein separations. Dextran and agarose-based SE gels were subjected to enzymatic digestion under mild conditions. This treatment partially hydrolyzed the gel matrix modifying its pore size distribution. Enzymatic treatment of agarose-based SE gels was found to increase the resolution of the separation. Successful separation of preparative amounts of the A and B forms of beta-lactoglobulin (difference in electrophoretic mobility of 8.5%) was achieved with a high degree of purity using agarose-based SE gels. The four major whey proteins, beta-lactoglobulin, alpha-lactalbumin, BSA and immunoglobulins, were purified from an acid whey preparation. The degree of retention of a protein in electrochromatography followed their free-solution electrophoretic mobility (mu) when the protein was able to enter the gel pores and the ratio of diffusion/mu when the protein was excluded.     

Protein Loading Capacity and Textural Properties of Column Packings in Reversed-Phase HPLC

Abstract

The relationship between the textural properties (pore size, pore volume and surface area) of reversed-phase silica gel packings for HPLC and the dynamic loading capacity of large biomolecules was studied by using silica gels manufactured by similar processes. Several silica gels whose unbonded pore diameters range from 100 to 250 A and whose pore volumes range from 1.0 to 1.4 ml/g have been prepared and characterized. The bonded phase is monomeric C18. The textural properties of the bonded silica gels are also presented and related to the properties of the unbonded silica gels.

Chromatographic evaluation with typical proteins in an underload-to-overload condition was performed in order to relate the influence of textural properties of silica gel to loading capacity and resolution. The packings with larger pore size and pore volume produced better column performance and higher loading of proteins.