Separation Of Anaesthetics And Ephedra Alkaloids By Capillary Electrophoresis With Electrochemiluminescence Detection

Capillary electrophoresis (CE) is based on the mobilities of charged species through a filled capillary on application of a voltage. During the past few years, CE has become a robust separation technique for characterizing the molecules in free solutions. It brings a profound revolution following the gas chromatography (GC) and high-performance liquid chromatography (HPLC), develops the analysis technology from a level of microlitre to nanolitre, and makes it possible to analyze a single cell and single molecule. CE has attracted considerable attention because of its virtues of high resolution, short analysis time, small analyte consumption, rapid separation and easy automation.Electrochemiluminescence (ECL) detection is another sensitive detection technique in CE, which has advantages of high sensitivity, wide dynamic linear range, low cost, simplicity and inexpensive instrumentation. ECL detection based on tris(2,2’-bipyridyl)ruthenium(II)(Ru(bpy)32+) has been proved to be a powerful analytical technique and successfully demonstrated in analysis of proteins, ami no acids and drugs.Recently, CE coupled with ECL using Ru(bpy)32+has been proved to be a sensitive and efficient analytical technique, is suitable for the analysis of trace components in complex biological and pharmaceutical samples.In this work, the studies on the analysis of drugs by CE coupled with Ru(bpy)32+of ECL detection have been described. The main investigations are as follows.1. A new method for the determination of procaine (Pro), lidocaine (Lid), ropivacaine (Rop) and bupivacaine (Bup) was developed by CE-ECL. The use of SDS with Tween20mixed micelles in CE could improve significantly the resolution of analytes with the similar structure. The sensitivity can be increased by utilization of Eu-PB modifying Pt working electrode. The parameters affecting CE separation and ECL detection were investigated in detail. Under the optimized conditions, Pro, Lid, Rop and Bup were baseline separated within9min. The limits of detection (LOD, S/N=3) of Pro, Lid, Rop and Bup are2.5×10-8,1.3×10-8,3.0×10-8and4.1×10-8mol/L, respectively. The limits of quantitation (LOQ, S/N=10) of Pro. Lid. Rop and Bup are2.3×10-7,1.210-7mol/L,3.7×10-7and5.6×10-7mol/L in human urine and8.5×10-7，6.9×10-7,2.8 x10-6and1.1x10-6mol/L in human serum. The recoveries of the four analytes at different concentration levels in human urine and serum samples were between85.7%and107.6%. The RSDs of peak area were less than3.6%. The proposed method was successfully applied to the determination of the local anaesthetics in biofluids. In order to analyze quantitatively the four analytes in real sample, the linear ranges and the limits of quantitation in urine and serum samples were determined, respectively.2. A novel approach for the simultaneous determination of ephedrine (E), methylephedrine (ME) and pseudoephedrine (PE) was developed by capillary electrophoresis (CE) coupled with electrochemiluminescence (ECL) detection using internal standard method. The use of ionic liquid (IL) can improve the separation efficiency and sensitivity. In order to improve precisions and obtain accurate results in CE especially for biochemical analysis, the internal standard method was adopted. Parameters affecting separation, detection and IL were investigated in detail. Under the optimum conditions, three ephedra alkaloids were well separated and detected. The LOD (S/N=3) of E, ME and PE are4.0x10-8,6.5×10-8and4.6x10-8mol/L, respectively. The LOQ (S/N=10) in human urine are5.3×10-7mol/L for E,9.1×10-7mol/L for ME and6.9x10-7mol/L for PE, respectively. The precisions (RSD%) of the peak area and the migration time are less than3.5%and0.8%within a day and less than5.4%and1.4%in three days with external standard method, respectively. Using internal standard method, the RSD%of the peak area and the migration time are less than2.5%and0.2%within a day and less than3.9%and0.8%in three days using internal standard method, respectively. The recoveries of three analytes at different levels in traditional Chinese medicine and human urine samples were between85.6and108.4%. The applicability of the proposed method was illustrated in the qualitative and quantitative analysis of three ephedra alkaloids in traditional Chinese medicine. The method was also applied to the determination of E, ME and PE in human urine samples and the monitoring of pharmacokinetics for PE in human body. In order to analyze quantitatively the three analytes in real sample, the linear ranges and the limits of quantitation traditional Chinese medicine and in urine samples were determined, respectively.