Electrochemistry of capillary systems with narrow pores. IV. Dialysis potentials

The dialysis potentials of different collodion membranes with graded pore sizes and electrochemical activities have been measured in dilute aqueous KCl solutions as functions of concentration. It is possible to predict the value of the diffusion potential within a few millivolts on the basis of electrical conductivity data obtained with the same membranes. In general, the measured values are lower than those calculated. It is assumed that this difference is caused by the membranes having a distribution of pore sizes.     

Multifold curdlan gel formation by dialysis into aqueous solutions of metal salts

We have found that the dialysis of curdlan dissolved in alkaline solution into aqueous solutions of metal salts yielded multifold gel structures. Aqueous sodium chloride and potassium chloride as well as pure water induced isotropic gels. Aqueous calcium salts induced liquid crystalline gel with refractive index gradient/amorphous gel alternative structure. Aqueous salts of trivalent aluminum and ferric cations induced a rigid liquid crystalline gel, which shrank above a threshold concentration of each salt. On the other hand, Liesegang ring-like pattern was observed with aqueous solutions of mixed salts of calcium chloride and magnesium chloride. The patterns have been classified to discuss the mechanism of forming the variety of structures.     

Automated determination of verapamil and norverapamil in human plasma with on-line coupling of dialysis to high-performance liquid chromatography and fluorometric detection

A fully automated method for the simultaneous determination of verapamil and its main metabolite norverapamil in human plasma is described. This method is based on on-line sample preparation using dialysis followed by clean-up and enrichment of the dialysate on a precolumn and subsequent HPLC analysis with fluorometric detection. All sample handling operations were performed automatically by a sample processor equipped with a robotic arm (ASTED system). The plasma samples were dialysed on a cellulose acetate membrane (cut-off: 15 kD) and the dialysate was purified and enriched on a short pre-column filled with cyanopropyl silica. Before starting dialysis, this trace enrichment column (TEC) was first conditioned with the HPLC mobile phase and then with pH 3.0 acetate buffer. 370 μl of plasma sample spiked with the internal standard (gallopamil) were dialysed in the static-pulsed mode. The solution at the donor side was pH 3.0 acetate buffer containing Triton X-100 while the acceptor solution was made of the same acetate buffer. When dialysis was discontinued, the analytes were desorbed from the TEC by the HPLC mobile phase and transferred to the C₁₈ analytical column by means of a switching valve. This mobile phase consisted of a mixture of acetonitrile, pH 3.0 acetate buffer and 2-aminoheptane. The influence of different parameters of the dialysis process on the recovery of verapamil and norverapamil has been studied. The effect of the volume, the aspirating and dispensing flow-rates of the dialysis solution has been investigated. The recoveries of verapamil and norverapamil in plasma were close to 75% and the limits of quantification were 5 ng/ml for both analytes. The method was found to be linear in the concentration range from 5 to 500 ng/ml (r²: 0.9996 for both analytes). The intra-day and inter-day reproducibilities at a concentration of 100 ng/ml were 2.3% and 5.6% for verapamil and 1.7% and 5.1% for norverapamil, respectively.     

Dialysis-Chelex method for determination of exchangeable copper in human plasma

A method based on dialysis in the presence of histidine, and subsequent copper adsorption and preconcentration using Chelex-100 resin, has been developed for selective extraction of exchangeable copper from blood plasma. The method was verified by comparative analysis using two different instrumental techniques, ICP–MS and AAS. The results obtained were in excellent agreement. Accurate quantification of the exchangeable copper and the firmly bound copper was achieved. The method was further validated by analysis of a lyophilised human serum certified reference material.     

在柱透析-毛细管电泳联用分析--一种带有透析采样功能的毛细管电泳的实现与初步应用

在石英毛细管的进样端端口内采用相转移法制作聚砜膜,使该毛细管同时具有采样、样品净化与分离功能.建立了一种透析-毛细管电泳在柱联用方法.膜能有效地拦截大分子和颗粒物,性能稳定,12h内连续使用,膜的透过率和对大分子的拦截性能无显著变化.该方法避免了大分子和颗粒物的干扰,可直接对复杂样品中的小分子进行分析.将其应用于咖啡牛奶中游离咖啡因的含量测定,测得其值为0.68mmol/L;用于药物与蛋白的相互作用研究,获得了盐酸异丙嗪与牛血清白蛋白的结合常数,其值为1.47×104L/mol.     

膜渗透与离子色谱联用技术测定蔬菜中亚硝酸盐和硝酸盐

建立了膜渗析与离子色谱联用技术分析蔬菜等复杂基体中亚硝酸盐和硝酸盐的方法。蔬菜样品经沸水浴后,试样液通过膜渗析除去水溶性大分子杂质和颗粒,直接进入离子色谱分析。渗析池采用0.20μm的醋酸纤维为渗析膜,以水为接收液,分析柱为Metrosep A Supp4-250阴离子交换柱,以1.8mmol/L Na2CO3 1.7mmol/L NaHCO3为流动相,采用化学抑制电导检测。以20g称样计量,方法中NO2-、NO3-的检出限分别为0.075mg/kg和0.105mg/kg;实际样品的加标回收率分别为80.6%~98.7%和90.5%~114%;相对标准偏差(RSD)≤10%。本方法前处理简单,重现性好,准确度高。     

Electrochemistry of capillary systems with narrow pores. I. Overview

In capillary systems with narrow pores the Helmholtz electrochemical double layer located at the pore wall extends over the entire cross section of the pores. It loses its character as the “charge on the wall”. It will be shown that not only the electrokinetic phenomena but also the electrical conductivity and the dialysis potential of membranes with narrow pores can be understood from the same point of view, namely: the electrolyte solution in the pores of a membrane with narrow pores is considered to be an approximately homogeneous solution in contact with immobilised charges located at the pore wall. In this case the electrochemical equations contain the fixed ion concentration as a parameter instead of the ζ potential. This makes it possible to describe quantitatively to a good approximation data on the electroosmosis, the electrical conductivity, the streaming potential and the dialysis potential taken from the literature, as well as results of our own measurements, by using a single membrane constant.     

Transferrin immunoextraction for determination of carbohydrate‐deficient transferrin in human serum by capillary zone electrophoresis

CZE‐based assays for carbohydrate‐deficient transferrin (CDT) in which serum is mixed with an Fe(III) ion‐containing solution prior to analysis are effective approaches for the determination of CDT in patient samples. Sera of patients with progressed diseases, however, are prone to interferences comigrating with transferrin (Tf) that prevent the proper determination of CDT by CZE in these samples. The need of a simple and economic approach to immunoextract Tf from human serum prompted us to investigate the use of a laboratory‐made anti‐Tf spin column containing polyclonal rabbit anti‐human Tf antibodies linked to Sepharose 4 Fast Flow beads. This article reports extraction column manufacturing and column characterization with sera having normal and elevated CDT levels. The developed procedure was applied to a number of relevant hepatology and dialysis patient samples and could thereby be shown to represent an effective method for extraction and concentration of all Tf isoforms. Furthermore, lipemic sera were delipidated using a mixture of diisopropyl ether and butanol prior to immunoextraction. CDT could unambiguously be determined in all pretreated samples.     

Dialysis-assisted two-dimensional gel electrophoresis

2-DE is an important tool in proteomics research. However, intrinsic gel-to-gel variability of 2-DE often masks the biological differences between the samples and compromises quantitative comparison of protein expression levels. Here, we describe a modification of 2-DE that results in improved matching and quantification of proteins. This was accomplished by performing IEF of two samples in two IPG strips separated by a dialysis membrane. After IEF running, the strips were separated and the SDS-PAGE dimension was accomplished on two individual gels. After gel staining with CBB, ImageMaster 2D Platinum software (Amersham) was used for spot detection and quantification. Analysis of protein extracts from C2C12 myoblasts by this method resulted in 99% spot-matching efficiency and CV in stain intensity (% volume) was less than 0.5 for 98% of spots. We conclude that this technique, called dialysis-assisted gel electrophoresis, gives superior spot matching and quantitative reproducibility compared to IEF conducted on separate strips.