Investigation And Application Of High Performance Liquid Chromatography Combined With Ru(bipy)32+-ce(s04)2 Post-column Chemiluminescence Detection In Pharmaceutical Analysis

With high sensitivity and fast analytical time of chemiluminescence (CL) and high selectivity of high performance liquid chromatography (HPLC) technology, HPLC-CL has become a good technique in different fields of analytical chemistry. This thesis describes the application of high performance liquid chromatography (HPLC) combined with Ru(bipy)32+-Ce(SO4)2 post-column chemiluminescence (CL) detection in pharmaceutical analysis, which mainly contains two parts.The first part was a review that introduced chemiluminescence detection technique and HPLC-CL technique which contains principles, equipment, assortment, the main chemiluminescence reaction systems. Among them, Ru(bipy)32+-Ce(SO4)2 CL system was mainly reviwed, the application and development of HPLC-CL was summarized.The second part is research reports, which are composed of five parts as follows.1 Determination of indometacin in compound indomethacin tincture and in human urine samplesThe reaction of Indomethacin and tris(2,2’-bipyridyl) ruthenium(II) with intermediate reagent cerous sulfate oxygenant in the presence of acid. And the CL radiation was emitted in the reaction of their procreant oxidation productions. The optimal conditions for the CL detection were studied and a new method for determining indommethacin is described by HPLC-CL. The linear range is 22.95-61.20μg/mL with the detection limit of 6.12μg/mL, and the relative standard detection is 2.64%for 61.2μg/mL. This method has obtained satisfactory results in the analysis of indomethacin in compound Indometacin Tincture and in human urine.2 Determination of carbamazepine in human urine and serum samplesIt was found that carbamazepine (CBZ) could significantly enhance the chemiluminescence (CL) of the reaction of cerium sulfate and tris(2,2’-bipyridyl) ruthenium(II) CL system in the presence of acid. According to the observation, influence factors of the CL signal and the chromatograph separation were investigated, and a novel and rapid HPLC-CL method for the determination of carbamazepine was developed in the work. Under the optimal conditions, the linear range is 2.0×10-8-4.0×10-5 g/mL with the detection limit of 6.0 x 10-9 g/mL, and the relative standard detection is 2.5%for 2.0×10-6 g/mL(n=11). This method has obtained satisfactory results in the analysis of carbamazepine in pharmaceutical tablets and human urine and serum biological fluids samples.3 Determination of carbamazepine in human serum and urine samplesA sensitive, novel and rapid chemiluminescence (CL) method combined with high-performance liquid chromatography (HPLC) separation for the determination of salicylic acid (SA) is described in this work. The method was based on the fact that SA could significantly enhance the CL of the reaction of cerium sulfate and tris(2,2’-bipyridyl) ruthenium(II) CL system in the presence of acid. Under the optimal conditions, the CL intensity was linear over concentrations of SA in the range 0.02-10μg/mL, with a detection limit of 8 ng/mL (S/N= 3). And the relative standard detection was 2.2%for 0.1μg/mL (n= 11). The proposed method has been successfully applied to the analysis of SA in human serum samples and urine samples with satisfactory results.4 Determination of captopril in human urine samplesA new approach to improving for determination of captopril in human urine samples using high performance liquid chromatography (HPLC) with chemiluminescence (CL) detection is developed. The method is based on the CL reaction of captoril with Cerium sulfate (Ce(SO4)2) and tris(2,2’-bipyridyl) ruthenium(Ⅱ) (Ru(bipy)32+), which was added in the mobile phase. The chromatographic separation was performed on a Kromasil TM RP-C18 column (i.d.:150 mm×4.6 mm, particle size:5μm, pore size:100 A) with a mobile phase consisting of methanol-0.01 mol/L KH2PO4-1 mg/mL Ru(bipy)32+(80:20:2, v/v/v) at a flow rate of 0.9 mL/min, the total analysis time was within 10 min. Under the optimal conditions of 0.1 mol/L H2SO4,8.0×10-4 mol/L Ce(SO4)2, the linear range was 2.0 X 10-7-1.0 X 10-4 mol/L (r2= 0.9988) with the detection limit of 6.0 X 10-8 mol/L, and the relative standard detection (RSD) was 1.8%for 1.0 X 105 mol/L (n= 11). This method had obtained satisfactory results in the analysis of captopril in human urine samples. The possible mechanism of the CL reaction was also discussed briefly.5 Determination of sulfite in beer samplesThe CL radiation was emitted in the reaction of cerium sulfate oxidation reagent with tris(2,2’-bipyridyl) ruthenium(II) in the presence of acid. The existence of sodium sulfite makes the reaction more sensitive and strong. The optimal conditions for the CL detection were studied and a new method for determining sodium sulfite is described by HPLC-CL. The linear range is 2.0×10-6-5.0×10-4g/mL with the detection limit of 6.0×10-7 g/mL, and the relative standard detection is 3.7%for 5.0×10-3 g/mL. This method has obtained satisfactory results in the analysis of sodium sulfite in beer samples.