Sensitive Determination Of Drugs By Capillary Electrophoresis With Electrochemiluminescence Detection And Applications To Investigate Interaction Of Drug With Protein

CE, with advantageous properties including high separation efficiency, short analysis time, low consumption of chemicals, and ease of installation, operation, and maintenance, is a particularly interesting candidate for drug analysis. ECL is a powerful analytical technique with the advantages of high sensitivity, wide linear range, and simple instrumentation. ECL has been widely used in the areas of, for example, immunoassay, food and water testing, and biowarfare agent detection. CE combining with ECL （CE-ECL） has good performances for the determination of drugs and applications to investigate interaction of drug with protein. The main contents of this thesis are as follows:1. A new method for determination of metoprolol tartrate （ME） and bisoprolol fumarate （BF） by CE coupled with tris（2,2’-bipyridyl）-ruthenium（II） ECL detection was developed for the simultaneous separation and determination of ME and BF. The parameters including pH and concentration of the running buffer, separation voltage and detection potential that affected CE separation and ECL detection were optimized. Under the optimized conditions, ME and BF were well separated and detected within 10 min, the linear range of ME and BF are 0.05-60 μmol L^-1 and 0.5-40μmol L^-1, the limits of detection （LODs S/N= 3） are 1.9×10^-8 mol L^-1 for ME and 9.4 ×10^-8 mol L^-1 for BF, separately. RSDs of the peak area of ME and BF （n=5） are 2.1% and 3.8% within a day （intraday）,3.1% and 5.5% in 3 consecutive days （interday）. RSDs of the migration time of ME and BF are 4.5 and 6.7% for intraday,7.4 and 7.9% for interday. The limits of quantitation （LOQs S/N=10） in human urine sample are 3.3×10^-7 mol L^-1 for ME and 1.4 × 10-6 mol L^-1 for BF. The recoveries （n=5） of ME and BF in human urine are from 89.0 to 126.0% with less than 7.4% in RSD. This method was also successfully applied to the determination of ME and BF in human urine and study on interaction between ME or BF and human serum albumin （HSA）. The number of binding sites of ME and BF with HSA are 1.2 and 1.1, and the binding constants are 2.8×103 and 2.7×103 L mol^-1, respectively.2. A new method of CE coupled with tris（2,2’-bipyridyl） ruthenium（II） ECL detection has been developed to detect four local anesthetics procainamide （PAH）, tetracaine （TCH）, proparacaine （PCH） and cinchocaine （CIN） simultaneously. A europium （Ⅲ）-doped prussian blue analogue film （Eu-PB） modified platinum electrode was prepared and applied to improve the detection sensitivity. The parameters including additives, concentration and pH of the running buffer, separation voltage and detection potential that affect CE separation and ECL detection were optimized in detail. The four local anesthetics was baseline separated and detected within 10 min under the optimized conditions. The linear range of PAH, TCH, PCH and CIN are 0.2^-75 μmol L^-1,0.5-50 μmol L^-1,0.5^-100μmol L^-1 and 0.5^-100μmol L^-1. The LOD of PAH, TCH, PCH and CIN are 5.5×10^-8,9.6×10^-8,2.5×10^-8 and 3.5 ×10^-8 mol L^-1 （S/N=3）, respectively. RSDs of the migration time for four analytes range from 1.2 to 2.5% within intraday and from 2.4 to 4.9% in interday, RSDs of the peak area for four analytes are from 1.7 to 3.3% within intraday and from 2.2 to 5.6% in interday, respectively. The LOQ （S/N=10） for PAH, TCH, PCH and CIN in human urine sample are 5.9×10^-7,9.2 ×10^-7,8.3 × 10^-7 and 5.0 × 10^-7 mol L^-1, separately. The recoveries （n=5） of four analytes in human urine are from 87.6 to 107.7% with less than 5.9% in RSDs. The developed method was used to determine four local anesthetics in human urine samples and investigate the interaction between PAH and HSA. The number of binding sites and the binding constant of PAH with HSA were calculated to be 1.03 and 2.4 × 104 L mol^-1, respectively.3. A sensitive CE coupled with ECL system of tris（2,2’-bipyridyl） ruthenium （Ⅱ） was described for detection of proprnolol （Pro） and acebutolol （Ace）. AuNPs were added in the end column luminescent reagents solution to enhance the sensitivity of the CE-ECL system. Experimental conditions of separation and detection as well as the amount of AuNPs were optimized. Under the optimum conditions, the linear range of Pro is 0.01^-100μmol L^-1 with the LOD of 3.6×10^-9 mol L^-1 （S/N= 3） and with the LOQ of 1.1 ×10^-7 mol L^-1 in human urine samples （S/N= 10）. For Ace, the linear range is 0.02^-100 μmol L^-1 with the LOD of 5.0×10^-9 mol L^-1 （S/N= 3） and with the LOQ of 9.5 ×10^-8 mol L^-1 in human urine samples （S/N=10）. RSDs （n=5） of the migration time for Pro and Ace are from 1.9 to 2.4% intraday and from 2.9 to 3.8% interday. RSDs of the peak area for Pro and Ace are from 3.2 to 3.7% intraday and from 4.3 to 4.6% interday, respectively. The developed method was successfully used to determine two analytes in human urine samples. In addition, the interaction between Pro as a model analyte and HSA was investigated; the number of binding site and the binding constant of Pro with HSA are 1.0 and 2.3 x 104 L mol^-1, respectively.