脂质体电动色谱法评价阿魏酸与生物膜的相互作用

利用脂质体与生物膜结构的相似性,将脂质体加入毛细管电泳缓冲溶液中作为假固定相,在数分钟内测定了阿魏酸的脂水分配系数Klw,建立了脂质体电动色谱评价阿魏酸与生物膜相互作用的方法。研究了脂质体中胆固醇的含量、缓冲溶液pH值和缓冲体系对Klw的影响。结果表明,在实验条件范围内(胆固醇含量0~30%,pH值4.0~12.0),胆固醇含量升高,缓冲溶液pH值增大,Klw降低;在不同的缓冲体系中,离子强度越大,Klw越大。脂水分配系数的变化反映了阿魏酸与生物膜相互作用。     

Fluorescence Quenching Study on the Interaction of Some Schiff Base Complexes with Bovine Serum Albumin

The interaction of Schiff base ligand A and its three metal complexes [A‐Fe(II), A‐Cu(II), and A‐Zn(II)] with bovine serum albumin (BSA) was investigated using a tryptophan fluorescence quenching method. The Schiff base ligand A and its three metal complexes all showed quenching of BSA fluorescence in a Tris‐HCl buffer. Quenching constants were determined for quenching BSA by the Schiff base ligand A and its metal complexes in a Tris‐HCl buffer (pH=7.4) at different temperatures. The experimental results show that the dynamic quenching constant (K_SV) was increased with increasing temperature, whereas the association constant (K) was decreased with the increase of temperature. The thermodynamic parameters ΔH, ΔG and ΔS at different temperatures were calculated. The ionic strength of the Tris‐HCl buffer had a great influence on the wavelength of maximum emission of BSA. Under low ionic strength, the emission spectra of BSA influenced by A‐Zn(II) had a small blue shift. Compared to A‐Zn(II), the emission spectra of BSA in the presence of the Schiff base ligand A and A‐Cu(II) had no significant λ_em shift. At high ionic strength, the emission spectra of BSA upon addition of the Schiff base A, A‐Fe(II), and A‐Zn(II) all had a red shift, but the emission spectra of BSA had λ_em shift neither at low ionic strength, nor at high ionic strength in the presence of A‐Cu(II). Furthermore, the temperature did not affect the λ_em shift of BSA emission spectra.     

Determination of Hydrogen Single Ion Activity Coefficients in Aqueous HCl Solutions at 25°C

The single ion activity coefficients of hydrogen and chloride ions in aqueous HCl solutions have been estimated at 25°C at concentrations up to 1 mol-kg^–1, using potentiometric measurements with ion-selective electrodes and appropriate calibration procedures. Two methods are described for an internal calibration of the electrodes in the extended Debye–Hückel concentration range. The results are compared to the conventional pH calibration with external buffer solutions. Since the latter calibration method does not account for the liquid junction potential E _J which arises at the reference electrode, the resulting activity coefficients are quite different in HCl solutions of higher concentration. These differences between internal and external calibration decrease significantly, when a correction for E _J is introduced into the conventional pH calibration. Hence, in solutions of higher ionic strength the accuracy of the conventional pH electrode calibration using buffer solutions is very limited, when exact H⁺ activities are required. The consistency of the results indicates that the liquid junction potentials in the examined systems calculated by the Henderson/Bates approximation are of reasonable precision.     

Factors Responsible for the Electrophoretic Behavior of Carboxylic Acid and Triazine Derivatives under Conditions of Capillary Zone Electrophoresis and Micellar Electrokinetic Chromatography

The effects of the pH, nature, and ionic strength of the running buffer; temperature; and surfactants, organic solvents, and macrocyclic compounds added to the buffer solution on the electrophoretic separation parameters of anionic, cationic, and neutral components of complex mixtures of organic compounds under the conditions of capillary zone electrophoresis and micellar electrokinetic chromatography. Pesticides from the series of chlorophenoxycarboxylic acids and triazines, as well as amino acids and substituted benzoic acid derivatives, were used as model systems. The predominant role of the addition of macrocyclic reagents, in particular, -cyclodextrin, as buffer electrolyte components was noted.     

pH effect on dynamic coating for capillary electrophoresis of DNA

A buffer consisting of tris(hydroxymethyl)aminomethane, 2-(N-moropholino)ethanesulfonic acid (Mes) and EDTA with constant ion strength was used to investigate the effect of buffer pH on the dynamic coating behavior of poly(N-isopropylacrylamide) (PNIPAM) for DNA separation. The atomic force microscopy (AFM) image illustrated that PNIPAM in lower-pH buffer was much more efficient in covering a silica wafer than that in higher-pH buffer. The coating performance of PNIPAM was also quantitatively analyzed by Fourier transform IR attenuated total reflectance spectroscopy and by measuring the electroosmotic flow (EOF). These results indicated that the stability of the dynamic coating was dependent on the pH of the sieving matrix and was improved by reducing the pH to the weak-acid range. The lower pH of the sieving buffer may induce the polymer more efficiently to adsorb on the capillary wall to suppress EOF and DNA–capillary wall interaction for DNA separation. The enhanced dynamic coating capacity of PNIPAM in lower-pH buffer may be attributed to the hydrogen bonds between the hydroxyl groups of the silica surface and the oxygen atom of the carbonyl groups of PNIPAM.     

Polymeric Sorbents for Bile Acids: II. Oligopeptide-Containing Resins with Quaternary Amine Groups

Abstract

Polymeric sorbents for bile acids have been prepared by attaching lysine-containing peptide sequences onto a water-swellable polyamide resin, by solid phase peptide synthesis, and then attaching a terminal N,N-dimethyl glycine residue that was subsequently quaternized. The resins with relatively longer peptide sequences demonstrated a higher binding capacity, on a per active site basis, for bile acids in pH 7.4 aqueous buffer solutions at 20°C than cholestyramine and colestipol when tested under the same in vitro conditions. In solutions of low ionic strength, they also have a degree of specificity for bile acid anions. The resins have a higher binding affinity for cholic acid than for glycocholic acid, which indicates the importance of the hydrophobic interactions in the binding.     

Empirical predictor of conditions that support ideal-filter capillary electrophoresis

Ideal-filter CE (IFCE) is a method for the selection of affinity binders for protein targets from oligonucleotide libraries, for example, random-sequence oligonucleotide libraries and DNA-encoded libraries, in a single step of partitioning. In IFCE, protein–oligonucleotide complexes and unbound oligonucleotides move in the opposite directions, facilitating very high efficiency of their partitioning. For any given protein target and oligonucleotide library, protein–oligonucleotide complexes and unbound oligonucleotides move in the opposite directions only for a limited range of EOF mobilities, which, in turn, corresponds to a limited range of pH and ionic strength values of the running buffer. Rational design of IFCE-based partitioning requires a priori knowledge of this range of pH and ionic strength values, and here we introduce an approach to predict this range for a given type of the running buffer. The approach involves measuring EOF mobilities for a relatively wide range of pH and ionic strength (I) values and finding an empirical predictor function that related the EOF mobility with pH and ionic strength. In this work, we developed a predictor function for a running buffer (Tris-HCl) that is commonly used in CE-based partitioning of affinity binders for protein targets. This predictor function can be immediately used for the rational design of IFCE-based partitioning in this running buffer, while the described approach will be used to develop predictor functions for other types of running buffer if needed.     

Influence of buffer composition on the capillary electrophoretic separation of carbon nanoparticles

Carbon nanoparticles obtained from the flame of an oil lamp were examined by means of capillary electrophoresis. The influence of buffer composition on the separation of the mixture of negatively charged carbon nanoparticles was studied by varying buffer selection, pH, and concentration. The electrophoretic pattern was affected by both the co- and counter-ion in the buffer solution, influencing selectivity and peak shape. The capillary electrophoretic separations at different pH revealed species with large electrophoretic mobilities under a wide range of pH. The mobility of selected species in the mixture of nanoparticles showed a strong dependence upon the solution ionic strength. The mobility of these nanoparticles as a function of ionic strength was compared to classical electrokinetic theory, suggesting that under the experimental conditions utilized, the species are small, highly charged particles with appreciable zeta potentials, even at low pH.     

Measurement of pH by flow injection over a wide dynamic range with PVC/neutral carrier electrodes

Two pH-sensitive neutral carrier/PVC electrodes are used simultaneously for the measurement of pH by flow injection. One of these is based on the neutral carrier tridodecylamine and the other on octadecyl isonicotinate, and together they allow the pH range 1–13 to be covered. These electrodes have been used in a very low dispersion miniature potentiometric flow cell designed specifically for use in flow injection in conjunction with a multi-channel data acquisition system. The effects of the solution ionic strength and buffer capacity on the pH measurement are discussed. A flow-injection manifold is proposed which can be used for high-accuracy pH measurements without ionic strength adjustment of the sample and for simultaneous pH and ion concentration measurements with ionic strength adjustment. This has been tested on some synthetic samples for the simultaneous determination of potassium and calcium and the measurement of pH.     

Separation of Basic Drugs Using Pressurized Capillary Electrochromatography

A novel pressurized capillary electrochromatography(PCEC) was developed to separate baxic drugs on strong cation exchange(SCX) column.The separation result by using PCEC was better than that by using micro-HPLC.The effects of electrical field and pressure on plate height and resolution were investigated.Influences of organic modifier,ionic strength and pH value of buffer on retention behavior were evaluated,and the separation mechanism was also discussed.     

Continuous microfluidic DNA and protein trapping and concentration by balancing transverse electrokinetic forces

Sample pre-concentration can be a critical element to improve sensitivity of integrated microchip assays. In this work a converging Y-inlet microfluidic channel with integrated coplanar electrodes was used to investigate transverse DNA and protein migration under uniform direct current (DC) electric fields to assess the ability to concentrate a sample prior to other enzymatic modifications or capillary electrophoretic separations. Employing a pressure-driven flow to perfuse the microchannel, negatively charged samples diluted in low and high ionic strength buffers were co-infused with a receiving buffer of the same ionic strength into a main daughter channel. Experimental results demonstrated that, depending of the buffer selection, different DNA migration and accumulation dynamics were seen. Charged analytes could traverse the channel width and accumulate at the positive bias electrode in a low electroosmotic mobility, high electrophoretic mobility, high ionic strength buffer or migrated towards an equilibrium position within the channel in a high electroosmotic mobility, high electrophoretic mobility, low ionic strength buffer. The various migration behaviours are the result of a balance between the electrophoretic force and a drag force induced by a recirculating electroosmotic flow generated across the channel width due to the bounding walls. Under continuous flow conditions, DNA samples were concentrated several-fold by balancing these transverse electrokinetic forces. The electrokinetic trapping technique presented here is a simple technique which could be expanded to concentrate or separate other analytes as a preconditioning step for downstream processes.     

Microchip micellar electrokinetic chromatography based on one functionalized ionic liquid and its excellent performance on proteins separation

Herein, one water-soluble functionalized ionic liquid (IL), 1-butyl-3-methylimidazolium dodecanesulfonate (BAS), was designed, investigated and successfully applied to microchip micellar electrokinetic chromatography (MEKC) construction. It possessed the properties of both IL and surfactant. A fairly stable pH value approximately 7.4, which was fit to pH values of general biological buffers, was nicely placed at the optimum concentration of 20mM BAS solution. While applying BAS solution as running buffer in poly(dimethylsiloxane) (PDMS) microfluidic systems, significantly enhanced electroosmotic flow (8-fold) and resolutions between analytes were obtained than that using other supporting electrolytes or surfactants. Pure BAS solution could conveniently complete the task of MEKC establishment: not only the ionic strength of the running buffer was guaranteed, but also the analytes adsorption to PDMS surface was conquered, as was demonstrated in the sensitive determination of fluorescent dyes and well-separated protein mixtures.     

鼠李糖脂应用于微乳毛细管电动色谱快速测定化妆品中激素类物质

采用生物表面活性剂鼠李糖脂建立了无需助表面活性剂的微乳体系,并应用于微乳毛细管电动色谱快速分析化妆品中皮质类激素泼尼松、泼尼松龙和氢化可的松。考察了pH值、鼠李糖脂浓度、离子强度、油相种类和浓度、分离温度、分离电压及进样电压和时间的影响,得出微乳体系最佳组成为0.1%(w/w)鼠李糖脂+0.8%(w/w)正庚烷+99.1%(w/w)硼砂缓冲液(80 mmol/L,pH 9.2)。分离温度20℃,分离电压20kV,电动进样10 kV×3 s,泼尼松、氢化可的松和泼尼松龙在9.4 min内可基线分离。重复进样7次,迁移时间和峰面积的RSD分别小于0.2%和5.0%。3种分析物线性范围均为5～100 mg/L;检出限分别为1.0,1.1和1.3 mg/L(S/N=3)。仅需简单萃取即可用于化妆品样品测定,回收率为81.6%～108%;RSD均小于4.8%。     

Characterization of ion exchange stationary phases via pH transition profiles

New non-destructive method for characterization of ion exchange chromatographic columns based on transient pH formed by a step change in ionic strength of buffer solutions was examined. The method was used to distinguish between cation and anion or weak and strong ion exchange chromatographic supports and to determine the capacity of the chromatographic resins. The general scheme to distinguish between most commonly used types of ion exchange chromatographic columns was proposed. The duration of pH transient was shown to be linearly proportional to the total ionic capacity and was used to estimate protein dynamic binding capacity of the resin. The effects of pH, concentration and temperature on transient pH duration were examined.     

甲基丙烯酸酯整体柱毛细管电色谱法快速分析白芷中的主要活性成分

以甲基丙烯酸酯整体柱为分离柱,建立了一种快速分离、分析白芷药材提取物中的主要活性成分欧前胡素(imperatorin)、异欧前胡素(isoimperatorin)、珊瑚菜内酯(phelloptorin)和发卡二醇(falcarindiol)的毛细管电色谱(CEC)方法。在整体柱制备实验中对三元致孔剂组成成分的比例进行了系统的考察。在分离实验中对流动相的组成(包括有机相组成、缓冲液浓度和缓冲液的pH值)进行了优化。最终的优化条件为: 流动相为乙腈-20 mmol/L NaH2PO4(pH 4.95)(50:50, v/v),分离电压为－25 kV。结果表明,所制备的甲基丙烯酸酯毛细管整体柱具有良好的渗透性和重现性;4种分析物的标准曲线线性关系良好(r2＞0.997),检出限均小于0.34 mg/L,加样回收率为95.18%～98.44%。该方法快速、简便、可靠。应用该方法对18个不同产地的白芷样品进行了测定,并对其药材质量进行了评价。     

Ionic strength and buffer capacity of wide-range buffers for polarography

A procedure for the calculation of the ionic strength and buffer capacity of a series of buffers prepared by the partial neutralization of a mixture of mono- and polybasic acids is outlined and data tabulated for a mixture 0.02 M in each of CH₃COOH, H₃PO₄, and H₃BO₃. The useful polarographic range of each buffer solution is also given. Illustrative calculations have also been made to show that, at any given pH value, the minimum buffer capacity required for polarographic reduction of up to 1 × 10⁻³ M solutions of an organic compound, and which involves hydrogen ions, is 0.006.     

Buffer Choice for Tuning the Selectivity in Reverse Phase Chromatography

Abstract

Retention behaviour of ionogenic species in high-performance liquid chromatography on reversed phase materials was studied, specifically dependence of buffer quality applied to mobile phases. The buffers' effect on retention of organic acids, amino acids and dipeptides is quantified by modelling capacity factors as a function of pH-values. At constant ionic strength, increasing capacity factors were observed going from phosphate to less polar citrate buffer, modification of accessible silanol groups of the stationary phase being responsible for this effect. Application of citrate buffer for separation of a seven-component mixture is demonstrated on the basis of a computerized search for optimum chromatographic performance. The evaluated factor levels (pH, methanol content and ionic strength) differ from those found using phosphate buffer-containing mobile phases.     

Equations for calculation of the pH of buffer solutions containing sodium or potassium dihydrogen phosphate, sodium hydrogen phosphate, and sodium chloride at 25°C

Published thermodynamic data measured in aqueous mixtures of sodium or potassium dihydrogen phosphate with hydrogen phosphate and chloride at 25°C were used to test recently developed methods for calculation of the pH of phosphate buffer solutions. Equations for ionic activity coefficients are used in these methods. It is shown that all data used in the tests up to an ionic strength of about 0.5 mol-kg^-1 can be accurately predicted by the two methods recommended. In one of these methods, equations of the Hückel type are used for ionic activity coefficients and in the other equations of the Pitzer type. Several sets of phosphate buffer solutions are recommended,e.g., for calibrations of glass electrode cells. In the recommended sets, the pH of the buffer solutions can be calculated either by the Hückel or Pitzer method, and the pH predictions of these methods agree in most cases within 0.005 at least up to ionic strengths of about 0.2 mol-kg^-1. The pH values of the two primary pH standards endorsed by IUPAC based on aqueous mixtures of KH₂PO₄ and Na₂HPO₄,i.e., pH values of 6.865 and 7.413, can also be accurately predicted by the equations recommended in this study.     

Developments in the use of soluble ionic polymers as pseudo-stationary phases for electrokinetic chromatography and stationary phases for electrochromatography

This article reviews the development, characterization and application of soluble ionic polymeric materials as pseudo-stationary phases for electrokinetic chromatography and as stationary phases for electrochromatography since 1997. Polymeric pseudo-stationary phases for electrokinetic chromatography, including cationic polymers, anionic siloxane and acrylamide polymers, polymerized surfactants (micelle polymers), and chiral polymers are reviewed. Also reviewed are suspended molecularly imprinted polymer micro-particles. Application of polymeric pseudo-stationary phases with electrospray ionization mass spectrometric detection is presented. Recent progress in the development and characterization of physically adsorbed stationary phases for electrochromatography using polymers of the same or similar chemistry is also reviewed.     

Influence of moderate Joule heating on electroosmotic flow velocity, retention, and efficiency in capillary electrochromatography

The influence of Joule heating on electroosmotic flow velocity, the retention factor of neutral analytes, and separation efficiency in capillary electrochromatography was investigated theoretically and experimentally. A plot of electrical current against the applied electrical field strength was used to evaluate the Joule heating effect. When the mobile phase concentration of Tris buffer exceeded 5.0 mM in the studied capillary electrochromatography systems using particulate and monolithic columns (with an accompanying power level of heat dissipation higher than 0.35 W/m), the Joule heating effect became clearly noticeable. Theoretical models for describing the variation of electroosmotic flow velocity with increasing applied field strength and the change of retention factors for neutral analytes with electrical field strength at higher Tris buffer concentrations were analyzed to explain consequences of Joule heating in capillary electrochromatography. Qualitative agreement between experimental data and implications of the theoretical model analysis was observed. The decrease of separation efficiency in capillary electrochromatography with macroporous octadecylsilica particles at high buffer concentration can be also attributed to Joule heating mainly via the increased axial diffusion of the analyte molecules and dispersion of solute bands by a nonuniform electroosmotic flow profile over the column cross-section. However, within a moderate temperature range, the contribution of the macroscopic velocity profile in the column arising from radial temperature gradients is insignificant.