Preparation of "click" binaphthyl stationary phase and its application for separation of anthraquinones from Rheum palmatum L

An organosilane containing binaphthyl functional group was synthesized by clicking 2,2'-bis(prop-2-ynyloxy)-1,1'-binaphthyl with 3-azidopropyltriethoxysilane (AzPTES). The "click" binaphthyl stationary phase was then synthesized by covalently bonding the organosilane onto silica beads. In this synthetic method, the residue of copper iodide (CuI) catalyst can be controlled at a very low level. Polycyclic aromatic hydrocarbons (PAHs) were well retained on the "click" binaphthyl column, and showed different retention factors (k) and separation ratio (α) values from those on octadecylsilyl (ODS) column due to the π-π interaction between the analytes and the stationary phase. The anthraquinone compounds in Rheum palmatum L. were selectively separated and enriched into a fraction by the "click" binaphthyl column. By using this way, the complexity of the sample was largely reduced. Twelve of anthraquinones were recognized by UPLC-ESI-MS/MS. This stationary phase will be very useful for separating and enriching similar compounds with planar aromatic structures from complex traditional Chinese medicine (TCM) samples.     

Single-step fabrication of asymmetric dual-phase composite membranes for oxygen separation

Asymmetric dual-phase composite membranes for oxygen separation were conveniently fabricated by an acid leaching technique. A thin dense layer of Ce_0.85Sm_0.15O_1.925/Sm_0.6Sr_0.4FeO_3−δ was left by controlling the degree of acid leaching, and a porous substrate of Ce_0.85Sm_0.15O_1.925 with a fluorite structure was formed after dissolution of Sm_0.6Sr_0.4FeO_3−δ with a perovskite structure in HCl. Thus, a thin dense layer and a porous substrate can be fabricated in a single step in which traditional shrinkage mismatch and chemical reaction between thin dense layers and porous substrates can be avoided. The thickness of the dense layer can be controlled by varying the acid leaching time. Hence, dual-phase composite membranes with high oxygen flux can be obtained.     

Fabrication of microporous organic network@silica composite for high-performance liquid chromatographic separation of drugs and proteins

Microporous organic networks (MONs) that exhibit good stability and hydrophobicity are promising candidates for performing HPLC separation of small organic compounds. However, their applications in separating large analytes as well as biomolecules are still limited by the microporous nature of MONs. Herein, we demonstrated the fabrication of a MON-functionalized silica (MON@SiO₂), exhibiting micro and mesopores for the HPLC separations of small drugs as well as large analytes, such as flavones, nonsteroidal anti-inflammatory drugs (NSAIDs), endocrine disrupting chemicals (EDCs), and proteins. MON was successfully modified on SiO₂ microspheres to yield the uniform and mono-dispersed MON@SiO₂. The separation mechanisms and performance of the MON@SiO₂ packed column were evaluated for a wide range of analytes, including neutral, acidic, basic compounds, drugs, and proteins. Compared with commercial C18 and SiO₂–NH₂ packed columns, the proposed MON@SiO₂ column afforded superior performance in the separations of flavones, NSAIDs, EDCs, and proteins. Moreover, the MON@SiO₂ column also offered good repeatability with intraday RSDs (n = 7) of <0.1%, <2.0%, <2.3%, and <0.7% for the retention time, peak height, peak area, and half peak width, respectively, for separating EDCs. This work proved the potential of using MONs in the HPLC separations of drugs and proteins.     

Preparation of weak cation exchange packings for chromatographic separation of proteins using "click chemistry'

"Click chemistry" is defined as a class of robust and selective chemical reactions affording high yields and is tolerant to a variety of solvents (including water), functional groups, and air. In this study, click chemistry was used as an effective strategy for coupling three alkyne-carboxylic acids onto the azide-silica to obtain three novel stationary phases of weak cation exchange chromatography, which were characterized with FTIR and elemental analysis. Six kinds of standard proteins, such as myoglobin, RNase A, RNase B, cytochrome C, α-chymotrypsin A, and lysozyme, were separated completely with the three novel weak cation exchange chromatography stationary phases. Compared with commercial weak cation exchange chromatography columns, the three kinds of novel weak cation exchange chromatography packings prepared by click chemistry approach have better resolution and selectivity. The mass recovery of more than 97% was obtained for all the tested proteins, and the bioactivity recovery of lysozyme on the prepared column was determined to be 96%. In addition, lysozyme was purified successfully from egg white with the novel weak cation exchange chromatography column by one step. The purity was more than 97% and a high specific activity was achieved to be 81 435 U/mg. The results illustrate the potential of click chemistry for preparing stationary phase for ion-exchange chromatography.     

Preparation of Novel Macroporous Silica-Based Amide-Polymer-Bonded Packing and Its Application to the Separation of Proteins

ABSTRACT

A novel silica-based amide-polymer-bonded packing to be used for protein separation is described. A macroporous silica support was bonded with diethoxymethylvinyl silane, and then copolymerized with methylacrylamide and divinylbenzene to produce a tailored stationary phase. This packing has high resolution and stability in a large pH range; it was characterized through the separation of a standard protein mixture containing bean trypsinogen inhibitor, glucose oxidase, ribonuclease and pepsin. Such manipulation of the proteins elution profile is achievable in less than 12 minutes. The effect of pH on the protein separation is discussed, as is the effect of ammonium sulfate concentration. It is suggested that the macroporous silica-based amide-polymer-bonded packing is a better alternative for the biopolymer separation.     

Preparation of a novel composite particles and its application in the fluorescent detection of proteins

A new fluorescence method for the detection of proteins with novel composite nanoparticles (CdS/PPA) has been developed. The composite nanoparticles have been prepared through an in-situ polymerization method under ultrasonic irradiation. The surface of the composite nanoparticles was covered with functional groups (-COOH). These groups may play a major role in the improving the water solubility and biocompatibility of the nanoparticles. The composite particles is combined with proteins in NaAc-HCl buffer solution (pH=1.99), which can result in strong fluorescence, and the response is linearly proportional to the concentration of proteins. In λem/λex=650 nm/365 nm place (the stoke’ shift is 285 nm), its fluorescent strength reaches the maximum. Under the optimum conditions, the linear range is 0.10–20.0 μg·ml⁻¹ with the detection limit of 41 ng·ml⁻¹ for HSA, and 0.10–15.0 μg·ml⁻¹ with the detection limit of 35 ng·ml⁻¹ for Human γ-IgG . The method has been applied to the determination of the total protein in human serum samples collected from the hospital and the results are satisfactory.     

Inertness and stability of a maltose-silica for the separation of proteins by HPLC

Summary A stationary phase consisting of maltose on a silica support, designed for the separation of proteins, is described. When maltose is coupled to aminopropylsilica an effective shielding of the silica surface can be obtained. A size exclusion experiment is used to investigate pore structure and the behaviour of proteins. The stability of the surface layer is studied, as influenced by silica pretreatments and cross-linking reactions. Inertness is studied via the denaturation of trypsin in columns of different length at a number of temperatures. An example of a derivatization of the maltose layer is given which demonstrates its use as an interactive separation medium.     

核壳硅碳复合微球固定相的制备及其在糖类分离中的应用

采用一步法在二氧化硅(SiO₂)表面涂覆酚醛树脂聚合物(PF),并在氮气气氛下碳化,制备了核壳硅碳复合微球(Sil@MC)固定相。实验对Sil@MC固定相进行形貌观察和孔结构分析,表明制备出的Sil@MC固定相具有良好的单分散性,包覆后的Sil@MC材料比表面积为302 m²/g,平均孔径为9.5 nm,孔容为0.63 cm³/g,说明通过共聚反应成功地将碳材料固定在二氧化硅上。将制备的Sil@MC材料作为HPLC固定相,采用匀浆法装柱,以乙腈-水(含0.1%(v/v)甲酸)作为流动相,发现Sil@MC色谱柱在高效液相色谱-质谱中可以实现4种极性糖类化合物(D-(+)-氨基葡萄糖盐酸盐、葡萄糖、D-(+)-海藻糖二水合物和棉子糖)的分离,然而未涂覆酚醛树脂的SiO₂材料未能对这4种极性糖类化合物实现分离。实验进一步对Sil@MC固定相的性能进行了评价,代表性的低聚糖异构体松三糖和棉子糖、耐斯糖和水苏糖及乳寡糖异构体3′-唾液酸乳糖和6′-唾液酸乳糖、乳-N-四糖和乳-N-新四糖在Sil@MC色谱柱中...     

Measuring the extent of phase separation during polymerization of composite latex particles using modulated temperature DSC

The morphological features of composite latex particles predominantly develop during the polymerization process and depend upon a significant number of variables. In this study, we have concentrated on the relative polarities of the two polymers in the particles and the rate at which we added the monomers during semibatch reactions containing the seed polymer latex. Our particular interest was to develop data that could reveal the extent of polymer phase separation as a function of the amount of monomer fed, and to characterize the morphology resulting from it. While TEM is the most common analytical technique employed, we show in this paper that modulated temperature DSC can generate data that allows us to follow the phase separation process as the monomer feed progresses. By considering the possibilities of having “phases” within the particles of pure polymer, homogeneously mixed (but nonequilibrium) polymers, gradient and interfacial polymer, we have been able to quite successfully simulate the DSC data. This results in quantitative estimates of the relative amounts of these “phases” and their polymer compositions. Combining these results with TEM photos showing the spatial characteristics of the morphology, we can achieve a much greater understanding of the physical structure of the composite latex particles. In many cases we find that phase separation is far from complete at the end of the reaction process. © 2005 Wiley Periodicals, Inc. J Polym Sci Part B: Polym Phys 43: 2790–2806, 2005     

“Click chemistry” preparation of WCX packings for protein separation

Click chemistry was applied to immobilize three kinds of alkyne-carboxylic acids onto azide-modified silica gel to prepare three novel stationary phases for weak cation exchange chromatography(WCX).The developed protocol combines the benefits of operational simplicity,exceptionally mild conditions and high surface loadings.Six kinds of standard proteins were separated completely on the novel packings.Compared with commercial WCX columns,the three kinds of novel WCX packings prepared by click chemistry approach have better resolution and selectivity.Lysozyme was purified successfully from egg white with the novel WCX column by one step.The purity was more than 97%and a high specific activity was achieved to be 81,435 U/mg.The results illustrate the potential of click chemistry for preparation of stationary phase for IEC.     

Chiral separation of enantiomers of amino acid derivatives by high-performance liquid chromatography on a norvancomycin-bonded chiral stationary phase

A novel norvancomycin-bonded chiral stationary phase (NVC-CSP) was synthesized by using the chiral selector of norvancomycin. The chiral separation of enantiomers of several dansyl-amino acids by high-performance liquid chromatography (HPLC) in the reversed-phase mode is described. The effects of some parameters, such as organic modifier concentration, column temperature, pH and flow rate of the mobile phase, on the retention and enantioselectivity were investigated. The study showed that ionic, as well as hydrophobic interactions were engaged between the analyte and macrocycle in this chromatographic system. Increasing pH of buffers usually improved the chiral resolution for dansyl--amino-n-butyric acid (Dns-But), dansyl-methionine (Dns-Met) and dansyl-threonine (Dns-Thr), but not for dansyl-glutamic acid (Dns-Glu) which contains two carboxylic groups in its molecular structure. The natural logarithms of selectivity factors (ln ) of all the investigated compounds depended linearly on the reciprocal of temperature (1/T), most processes of enantioseparation were controlled enthalpically. Interestingly, the process of enantioseparaton for dansyl-threonine was enthalpy-controlled at pH of 3.5, while at pH of 7.0, it was entropy-controlled according to thermodynamic parameters Δ_R,SΔH° and Δ_R,SΔS° afforded by Van’t Hoff plots. In order to get baseline separation for all the solutes researched, norvancomycin was also used as a chiral mobile phase additive. In combination with the NVC-CSP, remarkable increases in enanselectivity were observed for all the compounds, as the result of a “synergistic” effect.     

Controlling morphological and electro-optical properties via the phase separation in polymer/liquid-crystal composite materials

ABSTRACT

The polymer/liquid-crystal composite materials have been extensively studied for their potential applications. Various optical devices based on this composite material have been proposed and realised. The device performance is highly dependent on the phase separation of this composite material. Here, we investigate the photopolymerisation-induced phase separation in this composite material. Depending on the mass ratios between the polymer and the liquid crystal, the phase separation can be well controlled and subsequently affect the morphological and electro-optical properties. At a fixed ratio, we can realise either phase-separated composite films or conventional polymer-dispersed liquid crystal films with completely different optical properties. By carefully controlling the exposure conditions, the morphologies and electro-optical properties have been studied and optimised in details. With in-depth studies and optimisation, the photopolymerisation-induced phase separation technique could be utilised to realise many different optical functions based on the polymer/liquid-crystal composite materials.     

芘基丁酸改性锆-镁复合氧化物HPLC固定相的制备及其在分离富勒烯中的应用

制备了用于分离C₆₀和C₇₀的芘基丁酸改性锆-镁复合氧化物高效液相色谱固定相,并对其进行了表征;考察了流动相中甲苯含量、柱温和进样量对C₆₀和C₇₀分离的影响.结果表明,芘基丁酸改性锆-镁复合氧化物固定相对富勒烯有较强的保留,并对富勒烯表现出较强的分离能力,具有制备分离富勒烯的潜力.     

联萘酚手性拆分方法及其用于实验室废水的检测

联萘酚是具有手性的药物中间体,其对映体的拆分和检测具有重要意义.建立一种反相液相色谱手性拆分方法,用于联萘酚对映体的拆分.考察了6种不同手性色谱柱的拆分效果,并进一步对流动相配比、柱温和流速等因素进行了优化.结果表明,CHIRALPAK?IA手性柱对联萘酚对映体有较好的拆分性能,在流动相为甲醇-1％ 乙酸(90:10,...     

Assembly of poly(dopamine)/poly(acrylamide) mixed coatings by a single‐step surface modification strategy and its application to the separation of proteins using capillary electrophoresis

In this work, a facile approach was developed to modify a fused‐silica capillary inner surface based on poly(dopamine) and poly(acrylamide) mixed coatings for protein separation by capillary electrophoresis. The surface morphology, thickness, and chemical components of poly(dopamine)/poly(acrylamide) mixed coatings on glass slides and silicon wafers were studied by atom force microscopy, ellipsometry, and X‐ray photoelectron spectroscopy, respectively. The hydrophilicity and stability of the mixed coatings on glass slides were investigated by static water contact angle measurements. A comparative study of electroosmotic flow showed that the poly(dopamine)/poly(acrylamide) mixed coatings could provide effective suppression of electroosmotic flow. Meanwhile, the fast and efficient separations of the mixture of four alkaline proteins, the mixture of acidic, basic, and neutral proteins and egg white proteins were obtained by capillary electrophoresis. Furthermore, the consecutive protein separation runs and low RSDs of migration time demonstrated that these poly(dopamine)/poly(acrylamide) mixed coatings were capable of minimizing protein adsorption during the protein separation by using capillary electrophoresis.     

聚(L-谷氨酸)水凝胶介导羟基磷灰石的生物矿化

以1-乙基-3-(3-二甲基氨基丙基)碳二亚胺盐酸盐(EDC·HCl)为羧基活化剂, 己二酸二酰肼(ADH)为交联剂, 制备了生物活性聚(L-谷氨酸)(PLGA)水凝胶. 通过X射线衍射和扫描电子显微镜等表征了在不同浓度模拟体液(SBF)中羟基磷灰石(HA)的形成和生长. PLGA水凝胶的表面和内部均可观察到HA的形成和生长. 同时探讨了PLGA水凝胶矿化前后的力学性能. 将矿化前后PLGA水凝胶用于脂肪干细胞(ASCs)的培养, 研究其细胞相容性.     

The preparation of a novel polydopamine-graft-poly(2-methyl-2-oxazoline) protein-resistant coating and its applications in protein separation

A novel polydopamine-graft-poly(2-methyl-2-oxazoline)(PDA-g-PMOXA) coating was prepared by immobilizing poly(2-methyl-2-oxazoline)(PMOXA) onto material surfaces through polydopamine (PDA) anchored coating for the first time.And then,the chemical composition,hydrophilicity,and protein-resistant properties of the PDA-g-PMOXA coating were studied using X-ray photoelectron spectroscopy(XPS),contact angel(CA) test,surface plasmon resonance(SPR),and quartz crystal microbalance with dissipation(QCM-D) measurement.Finally,the coating was applied to the capillary inner surface for protein separation by capillary electrophoresis(CE).     

无孔单分散亲水性中强阳离子交换树脂的制备及其在蛋白质快速分离中的应用研究

以3.0μm无孔单分散亲水性交联聚甲基丙烯酸环氧丙酯树脂为基质,将其表面经新的化学方法改性后制备了一种新型的无孔中强阳离子交换色谱填料。详细考察了该无孔填料对标准蛋白分离性能,有机溶剂、pH、流动相盐种类和流速等对蛋白质保留的影响。实验结果表明,在流速为4 mL/min时,线性梯度时间在2.0 min内可快速分离4种标准蛋白,蛋白质的保留符合阳离子交换色谱规律。将其应用于快速纯化鸡蛋清中的溶菌酶,取得较好效果。     

A comparison of the adsorption of saliva proteins and some typical proteins onto the surface of hydroxyapatite

Adsorption of protein from saliva on hydroxyapatite was compared with adsorption of several typical proteins with different electric charges, i.e. lysozyme, human serum albumin, β-lactoglobulin and ovalbumin. Adsorbed amounts of these proteins were determined and electrophoretic mobilities of protein-covered hydroxyapatite particles were measured, at different values for the adsorbed mass and, therefore, at various degrees of surface coverage. Also, adsorption kinetics were investigated by streaming potential measurements of a hydroxyapatite surface in contact with a protein solution, allowing monitoring of changes in the zeta-potential of the protein-covered hydroxyapatite surface in real time. The adsorbed amounts show that, as compared to most of the other proteins, the saliva proteins have remarkably low adsorption affinity. The measured values for the electrophoretic mobilities indicate that the positively charged proteins in the saliva mixture preferentially adsorb onto the negatively charged hydroxyapatite surface; this is most pronounced at low protein concentration in solution (i.e. at low coverage of the surface by the protein). Preferential uptake of the positively charged saliva proteins during the initial stages of the adsorption process is also concluded from the results of the kinetics experiments. Preferential adsorption of positive proteins is somewhat suppressed by the presence of Ca²⁺ ions in the medium. The results suggest that an acquired pellicle on a tooth in an oral environment contains a significant fraction of positively charged proteins. The positively charged proteins in the pellicle reduce the zeta-potential at the tooth surface to low values; consequently, electrostatic forces are expected to play only a minor role in the interaction with other components (e.g. bacterial cells).     

Capillary zone electrophoretic separation of basic proteins and drugs using guaran as a buffer modifier

Summary Guaran, a neutral polysaccharide, has been used as a buffer modifier to improve the separation of basic proteins and drugs. Migration reproductibility, peak shape and efficiency were improved when 0.1% guaran was added to the buffer. The concentration of guaran, ionic strength, and pH of buffer solution were optimized to obtain the optimum separation of proteins. Possible separation efficiencies of 700,000 plates per meter were obtained for test proteins. The relative standard deviation (% RSD) of the migration time of all test proteins was less than 0.5%. Improved separation of β-blockers was also observed when guaran was added to the buffer.