Preparation of Antihypertensive Drugs in Biological Matrix with Solvent Front Position Extraction for LC–MS/MS Analysis

Solvent front position extraction procedure was used to prepare biological samples containing selected antihypertensive drugs (ramipril, lercanidipine, indapamide, valsartan, hydrochlorothiazide, perindopril, and nebivolol). Substances separated from the biological matrix components (bovine serum albumin) were quantified by means of liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). Sample preparation process was performed with the use of a prototype horizontal chamber with a moving pipette driven by a 3D printer mechanism enabling a controlled eluent flow velocity. Application of this device was advantageous for simultaneous preparation of several samples for further quantitative analysis, with a synchronized reduction of manual operations and solvent consumption. Quantitative results obtained for the majority of the investigated antihypertensive drugs in a complex biological matrix were satisfactory. The values of the %RSD were around 5% for six of the seven substances (with the exception of indapamide). The method exhibits a suitable accuracy (the relative error percentage was below 10% for most drugs). The values of LOD and LOQ were in the range of 1.19 µg/L–8.53 µg/L and 3.61 µg/L–25.8 µg/L, respectively.     

前沿色谱法对盐酸小檗碱和固定化牛血清白蛋白结合作用的研究

盐酸小檗碱(BC)是黄连的主要有效成分,它具有抗菌消炎等多种作用。采用前沿色谱法测定了不同温度下BC与固定化牛血清白蛋白(BSA)的结合常数K和结合率PPB、BC的保留因子k、BC在色谱柱上活性位点的物质的量mL,以及BC与BSA结合过程中的热力学参数。在温度为30 ℃时,BC与BSA的结合常数为4.79×104 L/mol。K、k和mL均随温度的升高而降低,其中以mL的变化程度最为显著,表明k的降低是由于K与mL共同作用的结果。而BSA分子构象变化可能是mL降低的主要原因。热力学分析结果表明:BC和BSA之间的作用力以静电作用力为主。     

亲和毛细管电泳法测定牛血清白蛋白和加替沙星的结合常数

利用亲和毛细管电泳法对牛血清白蛋白(BSA)与加替沙星(GT)间的结合反应及其相互作用做了初步探索,并应用淌度比(M)作为指标测定了两者的结合常数。以20 mmol/L pH 7.4的磷酸盐缓冲液作为运行缓冲液,分别以GT和BSA作为添加剂,另一组分为进样样品,内标为二甲基甲酰胺,于214 nm波长下检测。两种测定条件下得到的结合常数分别为4.4×104 L/mol和4.2×104 L/mol,与传统的荧光淬灭法测得的结果基本一致。该方法具有简单、高效的优点。     

三丁基锡(TBT)化合物与牛血清白蛋白(BSA)的相互作用

通过紫外、荧光和圆二色(CD)光谱,研究了船体防污漆的防污成分三丁基锡(TBT)化合物与牛血清白蛋白(BSA)的相互作用,考察了浓度、酸度和有机溶剂等因素的影响。结果表明,TBT与BSA的相互作用是双重的,既有TBT中丁基基团的疏水作用,又有锡离子与BSA的配位作用,使BSA内部的色氨酸和酪氨酸等芳香氨基酸残基裸露,导致BSA二级结构破坏,α-螺旋含量减少和构象改变。     

水杨酸键联卟啉的合成及与牛血清白蛋白的相互作用

以溴代烷氧基苯基卟啉(1)为原料与水杨酸作用,分别得到了化合物2和3,再与醋酸钴反应,得到了钴卟啉配合物4和5.新化合物分别由¹HNMR,IR,UV-Vis,MS和元素分析确证.同时,利用荧光光谱考察了金属卟啉与牛血清白蛋白的相互作用.     

反胶团相转移法提取牛血清白蛋白

介绍了在利用CTAB／正辛醇：三氯甲烷(4：1V／V)反胶团体系对牛血清白蛋白(BSA)进行相转移中,通过对萃取体系水相的pH值、离子强度、两液相的体积比、小分子糖(葡萄糖、蔗糖)及助表面活性剂(直链醇分子)等因素的改变,探讨了BSA在阳离子表面活性剂体系的萃取机理;研究结果表明选择合适的条件提取BSA时,萃取率可达到97％,反萃率达到了85％;找到实现牛血清白蛋白分离提纯的有效方法。     

荧光光谱法研究山梨醇与牛血清白蛋白的相互作用

采用荧光和紫外吸收光谱法,研究了利尿脱水药山梨醇（Sorbitol）与牛血清白蛋白（BSA）的相互作用。在正常生理条件下,山梨醇对牛血清白蛋白有较强的猝灭作用,根据不同的药物浓度、温度及紫外吸收光谱的变化,判断其猝灭方式可能为静态猝灭,考察了不同温度、药物浓度等多种条件下Sorbitol对BSA荧光猝灭的影响。通过Stern-Volmer方程和Lineweaver-Burk方程的简化形式,求出在不同温度下反应的结合常数K_D分别为7.4×10-5（25 ℃）和1.7×10-4（37 ℃）、结合位点数n为1。根据反应热力学参数确定了它们之间相互作用的主要形式为电荷作用力。采用同步荧光考察了山梨醇对BSA构象的影响,发现随药物浓度的增大,色氨酸残基的最大发射波长不变, 而酪氨酸残基所处环境的疏水性改变, 从而导致BSA的构象发生了变化。     

左氧氟沙星与牛血清白蛋白相互作用的液滴荧光法研究

采用液滴荧光技术与紫外-可见光度法研究了生理pH值条件下左氧氟沙星和牛血清白蛋白的相互作用机制。左氧氟沙星对牛血清白蛋白产生荧光猝灭,且猝灭过程是由于复合物形成而引起的静态猝灭。根据Forster偶极-偶极非辐射能量转移理论算出供体-受体的结合距离为2.68 nm。由Linewear-Burk方程求出不同温度下反应时复合物的形成常数K_LB和结合位点数n及对应温度下结合反应的热力学参数,证明二者主要靠疏水作用力结合。同时采用同步荧光分析技术,对蛋白质与药物结合时构象的变化进行了探讨。     

巴比妥钠与牛血清白蛋白结合反应的热力学研究

采用荧光光谱研究了药物巴比妥钠(BBTS)和牛血清白蛋白(BSA)的相互作用机理,运用热力学方法确定了巴比妥钠和牛血清白蛋白主要是以静电力相互作用,该过程是一个熵增加、Gibbs自由能降低的自发超分子作用过程,根据Stern-Volmer方程和Lineweaver-Burk双倒数曲线方程求出了其结合常数在高温时比在低温时小,得到巴比妥钠和牛血清白蛋白的结合比为1∶1,证实了巴比妥钠和牛血清白蛋白之间的作用是生成复合物的静态猝灭过程,并且采用同步荧光技术考察了巴比妥钠对牛血清白蛋白氨基酸残基微环境的影响。     

In-situ and one-step preparation of protein film in capillary column for open tubular capillary electrochromatography enantioseparation

In this wo rk,the phase-transitioned BSA(PTB) film using the mild and fast fabrication process adhered to the capillary inner wall uniformly,and the fabricated PTB film-coated capillary column was applied to realize open tubular capillary electrochromatography(OT-CEC) enantioseparation.The enantioseparation ability of PTB film-coated capillary was evaluated with eight pairs of chiral analytes including drugs and neurotransmitters,all achieving good resolution and symmetrical peak shape.For three consecutive runs,the relative standard deviations(RSD) of migration time for intra-day,inter-day,and column-tocolumn repeatability were in the range of 0.3%-3.5%,0.2%-4.9% and 2.1%-7.7%,respectively.Moreover,the PTB film-coated capillary column ran continuously over 300 times with high separation efficiency.Therefore,the coating method based on BSA self-assembly supramolecular film can be extended to the preparation of other proteinaceous capillary columns.