毛细管电泳测定主客分子相互作用的结合常数

在２５℃下,用毛细管区带电泳测定了主管体分子（配体和溶质分子）β－环糊精（β－ＣＤ）和心得舒（ａｌｐｒｅｎｏｌｏ）在ｐＨ值为２．５,浓度为１００ｍｍｏｌ／Ｌ的磷酸盐缓冲溶液下的结合常数,并对４种求解方法既非线性拟合法,双倒数法,ｙ－倒数法和ｘ－倒数法的结果进行了比较,所得值分别是３０７．２、４０８．４、３３１．４和３４３．１Ｌ／ｍｏｌ,同时获得了结合物的迁移率。该方法可用于测定结合比为１：１的各种     

毛细管电泳法研究牛血清白蛋白与盐酸异丙肾上腺素的相互作用

本研究分别采用区段-区段动力学毛细管电泳法(Plug-Plug Kinetic Capillary Electrophoresis,ppKCE),及以药物为添加剂的亲和毛细管电泳法(Affinity Capillary Electrophoresis,ACE)对盐酸异丙肾上腺素(Hydrochloric Acid Isoproterenol,HAI)与牛血清白蛋白(Bovine Serum Albumin,BSA)的结合作用进行研究。实验结果显示,ppKCE法可同时测得HAI与BSA相互作用体系的结合速率常数kon和离解速率常数koff分别为163.60L·mol-1·s-1、3.50×10-2±0.95×10-2s-1(n=3)。ppKCE法和ACE法测得HAI与BSA的结合常数Kb分别为4.67×103 L·mol-1、6.02×103 L·mol-1,两种方法所得结果能够较好吻合。证明毛细管电泳法在药物-蛋白相互作用研究领域的可靠性,可用于测定药物与蛋白的相互作用,从而用于新药筛选研究。     

毛细管电泳法研究环糊精与芳香羧酸的相互作用

基于毛细管电泳同时测定了β-环糊精与苯甲酸、苯乙酸和邻羟基苯甲酸的包结常数及包结反应的热力学参数，从分子结构和热力学角度讨论了超分子化合物形成的规律。结果表明：氢键在超分子化合物中具有重要作用，环糊精与芳香羧酸的包结反应在热力学上呈现熵焓补偿效应。该方法快速、准确，并有较高的重现性。     

二元络合体系的毛细管电泳相互作用分析

建立了一种利用毛细管电泳相互作用测定络合比为1：2的二元络合体系各级络合常数。依据所设计的毛细管电泳二级峰漂移模型,利用间接紫外检测方法,在运行电压为30kV、缓冲溶液组成为咪唑和醋酸（pH5．50）的条件下,测定了Mg^2＋与草酸体系的相互作用参数,求出了Mg^2＋与草酸的一级条件稳定常数的对数值为lgK1=3．40,二级条件稳定常数对数值lgK2=0．90,与文献值具有较好的一致性。     

毛细管电泳前沿分析法研究钾离子与18-冠醚-6体系相互作用

采用毛细管电泳前沿分析(CE／FA)法，利用间接紫外检测方法，在pH为5．10，运行电压为30kV，缓冲溶液组成为咪唑和醋酸的条件下，测定了钾离子与18-冠醚-6体系的相互作用参数，结合常数对数值lgK=3．50；在同样条件下用峰漂移法作对照实验，求得体系的结合常数对数值lgK=3．40，结果与前沿分析法基本一致。验证了两种方法测定结果的正确性。     

药物与细胞膜相互作用的ppKCE动力学参数

本文在区段-区段动力学毛细管电泳（ppKCE）的理论基础上,以兔红细胞膜、人红细胞膜为受体,首次同时测得了表征药物与细胞膜间结合快慢的正向结合速率常数kon、反向解离速率常数koff、以及配体与受体间相互作用的结合常数Kb。实验中考察了迁移时间及峰高的稳定性,pH值对药物与细胞膜相互作用动力学参数的影响。该方法为评价药物疗效、毒性及药物动力学研究提供一种新的方法。     

毛细管电泳法表征离子液体与蛋白质的相互作用

基于毛细管区带电泳考察了不同离子液体的阳离子母核、烷基侧链碳原子数目和阴离子组成对牛血清白蛋白的影响,以及离子液体[C4mim]BF4与肌红蛋白、牛血清白蛋白、血红蛋白、牛凝血酶和转铁蛋白之间的相互作用。利用亲和毛细管电泳法比较了[C4mim]BF4与上述蛋白之间的相互作用,并计算得到结合常数Kb分别为1.24×107 L/mol,1.23×106 L/mol,5.75×104 L/mol和5.60×104 L/mol。结果表明,转铁蛋白、血红蛋白、肌红蛋白、牛血清白蛋白与离子液体的相互作用依次减弱,存在1～2个数量级的差异。结合毛细管区带电泳和亲和毛细管电泳模式可实现离子液体与多种蛋白质相互作用的定性与定量表征。     

毛细管电泳法测定结合常数的研究进展

综述了近年来毛细管电泳法测定结合常数的研究进展。毛细管电泳测定结合常数的方法有多种,如淌度移动法、配体分离法、前沿分析法、部分填充法、空峰法和空亲和毛细管电色谱法及Hummel-Dreyer 法等。每种方法都有各自的优缺点,应根据被测体系来选择最合适的测定方法。另外,对数据处理方法以及测定结合常数的影响因素如温度、配体及缓冲溶液的浓度等也进行了详细评述。引用文献57篇。     

毛细管电泳测定物质三元相互作用体系的结合系数

毛细管电泳一般只能对二元结合体系的结合常数进行测定。研究了三元相互竞争体系的毛细管电泳测定方法,测定了金属锌离子与酒石酸和半胱氨酸以及锌离子与乳酸和半胱氨酸两个三元结合体系的结合常数,并以此比较了两种锌形态与半胱氨酸结合能力的大小。本法可用于测定两配体相互竞争结合体系的结合常数。     

配位作用毛细管电泳方法研究

研究了一种以过渡金属离子为添加剂,基于配位相互作用的毛细管电泳技术。选择7种芳香胺类化合物作为分析对象,考察了金属离子种类和浓度、运行介质酸度、电压对配位毛细管电泳的影响。通过对邻苯二胺变质样品的鉴别以及实际合成药磺胺类药物的分离,考察了这种毛细管电泳分离技术的分离能力,效果令人满意。应用于颠茄磺苄啶片中磺胺甲噁唑(SMZ)的定量测定时,SMZ的检出限为0.12 mmol/L;线性范围为0.25~10 mmol/L(r2=0.9917);平均加样回收率为97.8%,表明所建立的配位作用毛细管电泳(C ICE)方法,具有稳定、可靠的定性和定量分析能力。     

Application of liquid pre-column capillary electrophoresis technique to the study of interaction between drug enantiomers and human serum albumin

Based on the chiral separation of several basie drugs, dimetindene, tetryzoline, theodrenaline and verapamil, the liquid pre-colunm capillary electrophoresis (LPC-CE) technique was established. It was used to determine free concentrations of drug enantiomers in mixed solutions with human serum albumin (HSA). To prevent HSA entering the CE chiral separation zone, the mobility differences between HSA and drugs under a specific pH condition were employed in the LPC. Thus, the detection confusion caused by protein was totally avoided. Further study of binding constants determination and protein binding competitions was carried out. The study proves that the LPC technique could be used for complex media, particularly the matrix of protein coexisting with a variety of drugs.     

Investigation of interaction between the drug and cell membrane by capillary electrophoresis

By introducing cell membrane into electrophoretic buffer as pseudo-stationary phase, a novel capillary electrophoresis method was established to explore the interaction between drugs and cell membrane, where the interaction between citalopram and rabbit red blood cell membrane was used as an example. A series of concentrations of cell membrane were suspended into the running buffer by peak-shift method. The binding constant of citalopram to rabbit red blood cell membrane of 0.977 g^?1·L was obtained after treatment of Scatchard plot. This method could provide not only a new way for the investigation on the interactions between drugs and cell membrane, but also a new approach for high throughput screening of the drug membrane permeability, biological activity, and evaluating drugs in vivo.     

毛细管电泳法考察全氟辛酸与人血清白蛋白的相互作用

利用毛细管电泳（CE）技术建立了全氟辛酸（PFOA,C₈HF₁₅O₂）与人血清白蛋白（HSA）相互作用的分析方法。在生理条件下构建配体（PFOA）-受体（HSA）相互作用模型,通过淌度移动法、区段-区段动力学（plug-plug kinetic,PPK）法、简化的Hummel-Dreyer（HD）法研究其与HSA的相互作用。简化的HD法运用非线性方程、Scatchard方程和Klotz方程获得PFOA-HSA体系的相互作用参数,进而分析了模型适用度。结果表明,淌度移动法、PPK法、简化的HD法均适用于PFOA-HSA体系相互作用的分析,其中简化的HD法最优。模型适用度分析得出非线性回归方程为最适理论模型。相互作用参数测试表明PFOA-HSA相互作用体系之间发生的结合反应只有单一类型的结合位点且结合稳定。相关工作阐明了人血清白蛋白与PFOA的相互作用机制,可为PFOA毒理机制的深入研究提供有益参考。     

Electrostatic interaction mechanism on the separation of phenols by non-aqueous capillary electrophoresis

The electrostatic interaction between additive and analyte is of great importance to non-aqueous cap- illary electrophoresis(NACE)separation.Three tetraalkylammonium bromides and acetonitrile were applied as additives and running solvent respectively.The effect of alkyl chain length and concentra- tion of additive on electrostatic interactions was investigated by the separation of phenols.The sepa- ration ability was found to increase with decreasing alkyl chain length of the additive,and the resolu- tion values were increased with increasing additive concentration.The separation was seriously dete- riorated after a little amount of water was added in the running solution.Furthermore,the electrostatic interaction is strong under the conditions of low electron cloud density,weak steric hindrance and multi-interaction sites.Thus,the separation result can be predicted by theoretical analysis,which is helpful for the separation of other substances in NACE based on electrostatic interaction.     

毛细管电泳应用于测定牛血清白蛋白与脂质体的相互作用

建立了一种用毛细管电泳法检测牛血清白蛋白(BSA)与脂质体相互作用的分析方法。氧化指数的测定实验结果表明经过冷冻干燥的脂质体稳定性更好;毛细管电泳表征脂质体的电荷性质实验结果表明脂质体在pH 5.0～8.0的条件下呈电中性。在pH 7.0的条件下,以各种浓度的脂质体混悬液为电泳缓冲液,以0.8%二甲亚砜(DMSO)为内标,随着缓冲液中脂质体质量浓度从0增加到2.4 mg/mL,BSA的有效淌度从-2.232×10-4 cm2·V－1·s－1变化到-3.046×10-4 cm2·V－1·s－1;结合Scatchard分析,测得BSA与脂质体的结合常数为2.522×103(g/mL)－1。该方法简单、快速,为研究蛋白质与脂质体的相互作用提供了新的技术手段。     

A study of the interaction between enantiomers of zolmitriptan and hydroxypropyl-beta-cyclodextrin by capillary electrophoresis

The enantioresolution of zolmitriptan was performed using cyclodextrin (CD)-modified capillary zone electrophoresis (CZE) with hydroxypropyl-β-CD (HP-β-CD) as the chiral selector. The influence of experimental conditions on the enantioseparation of zolmitriptan, such as pH, temperature, applied voltage, and concentrations of running electrolyte and CD, was systematically investigated, obtaining a baseline separation of two enantiomers by the use of a 25 mM sodium dihydrogen phosphate (SDPH) running electrolyte (pH 2.4) containing 30 mM HP-β-CD at 15 °C. Binding constants for each enantiomer–HP-β-CD pair at different temperatures, as well as thermodynamic parameters for binding, were calculated. A nonlinear van’t Hoff plot was obtained, indicating that the thermodynamic parameters of complexation were temperature-dependent for zolmitriptan enantiomers. The significant contribution of the enthalpy difference to the Gibbs free energy change suggested a stereomeric barrier mechanism for chiral recognition. Figure Resolution of zolmitriptan enantiomers was achieved by using CD-modified CZE Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Evaluation of interactions between RAW264.7 macrophages and small molecules by capillary electrophoresis

In this study, the affinity interactions between RAW 264.7 macrophages and three small molecules including naringin, oleuropein and paeoniflorin were evaluated by affinity capillary electrophoresis (ACE), partial filling affinity capillary electrophoresis (PFACE) and frontal analysis capillary electrophoresis (FACE), respectively. The result indicated that ACE (varying concentrations of cell suspension were filled in the capillary as receptor) may not be suitable for the evaluation of interactions between cell and small molecules due to the high viscosity of cell suspension; PFACE can qualitatively evaluate the interaction, but the difference in viscosity between RAW264.7 suspension and buffer effects on the liner relationship between filling length and injection time, which makes the calculation of binding constant difficult. Furthermore, based on the PFACE results, naringin showed stronger interaction with macrophages than the other two molecules; taking advantage of the aggregation phenomenon of cell induced by electric field, FACE was successfully used to determine the stoichiometry (n = 5×10⁹) and binding constant (K_b = 1×10⁴ L/mol) of the interaction between RAW264.7 and naringin.     

Comparison of mobility shift affinity capillary electrophoresis and capillary electrophoresis frontal analysis for binding constant determination between human serum albumin and small drugs

In this study, two capillary electrophoresis–based ligand binding assays, namely, mobility shift affinity capillary electrophoresis (ms-ACE) and capillary electrophoresis-frontal analysis (CE-FA), were applied to determine binding parameters of human serum albumin toward small drugs under similar experimental conditions. The substances S-amlodipine (S-AML), lidocaine (LDC), l -tryptophan (l -TRP), carbamazepine (CBZ), ibuprofen (IBU), and R-verapamil (R-VPM) were used as the main binding partners. The scope of this comparative study was to estimate and compare both the assays in terms of their primary measure's precision and the reproducibility of the derived binding parameters. The effective mobility could be measured with pooled CV values between 0.55% and 7.6%. The precision of the r values was found in the range between 1.5% and 10%. Both assays were not universally applicable. The CE-FA assay could successfully be applied to measure the drugs IBU, CBZ, and LDC, and the interaction toward CBZ, S-AML, l -TRP, and R-VPM could be determined using ms-ACE. The average variabilities of the estimated binding constants were 64% and 67% for CE-FA and ms-ACE, respectively.     

毛细管电泳药物筛选方法与技术

毛细管电泳（CE）在新药研发领域显示着重要的应用前景。CE使用水溶液介质作为实验体系,保证了药物筛选在类似于生命介质的环境中进行,优于其他传统体外仪器筛选方法。除了维持被筛选分子和作用对象的生物活性外,CE筛选过程着重突出配体与受体之间的相互作用。毛细管电泳药物筛选瞄准与药理学理论相关的重要参数,如结合常数K_b 、结合速率常数K_on 和解离速率常数K_off ,有利于模拟并预测机体内靶标与药物之间的相互作用过程。该文回顾了毛细管电泳进行药物筛选的历史,评述了毛细管电泳药物筛选方法所依据的理论和相对成熟的各种常用方法,并抽取了部分典型实例以及相关技术进行说明,对以亲和毛细管电泳、动力学毛细管电泳为手段的药物筛选方法进行了介绍,包括分子和细胞层次的药物筛选,以及针对不同类型的候选药物的研究工作都有提及。毛细管电泳与多种技术的联用,包括与质谱以及化学发光等联用发挥了更大的效能。联用方法还应用于中药有效成分的筛选。毛细管电泳在DNA编码化合物库筛选中将有良好应用前景。馏分收集的发展为筛选药物提供了广阔前景,它配合指数富集配体系统进化技术为毛细管电泳药物筛选提供了更多可能。总之,毛细管电泳多样可选的药物筛选方法和技术将为新概念的药物筛选与药物评价提供有力支撑。     

涂层毛细管电色谱法评价药物与血小板的相互作用

建立了一种新的开管毛细管电色谱法以用于开展药物与血小板相互作用研究.该方法利用物理吸附将血小板固定于毛细管内壁,并通过改变血小板进样时间控制涂层长度.根据涂层长度与药物淌度的关系,建立了药物与血小板结合常数的计算方程,得出阿司匹林与血小板的结合常数为0.579g-1L,大于阴性对照品帕珠沙星与血小板的结合常数0.249g-1L.最后,用亲和萃取法所得结果与本方法对比,说明了本方法能够用于评价抗凝血药物与血小板的相互作用.