Ag(â…¢) Complex Chemiluminescence System:Establishment Of New Methods And Its Application In Forensic Toxicological Analysis

Chemiluminescence (CL) has become one of the focuses in the area of analytical chemistry due to its advantages such as higher sensitivity, wider linear range, simpler instrumentation, versatility of use and the easy automation. Combined with flow-injection analysis (FIA) or with capillary electrophoresis (CE) technique, the CL method can be utilized in environmental and biological samples analysis. Previously, we firstly reported a new CL system, i.e., Ag (III) complex-luminol system. In this study, we discussed its application in forensic toxicological analysis. The main contents are shown as follows:Part1Flow-injection analysis combined with luminol-Ag(III) complex chemiluminescence detection.(1) A novel flow injection analysis-direct chemiluminescence (FI-CL) method has been developed for determinations of trace amounts of dopamine (DA) based on the enhancing effect of DA on the CL reaction of luminol with an Ag(III) complex in alkaline solution. Under optimum conditions, CL intensities are proportional to the concentration of DA in the range of1.0x10-10-4.0x10-8mol·L-1. The detection limit is3.0x10-11mol·L-1for DA (3s), with a relative standard deviation (n=13) of2.3%for1.0x10-8mol·L-1DA. This method has also been applied for the determination of DA in commercial pharmaceutical injection samples. On the basis of the CL spectra and the results of the free-radical trapping experiment of this work, a reaction mechanism for this CL reaction is proposed and discussed.(2) A novel flow injection analysis-direct chemiluminescence(CL) method is proposed for the determination of ketamine, based on the enhancing effect of ketamine on the CL reaction of luminol with Ag (III) complex in alkaline solution. Under optimum conditions, CL intensities were proportional to the concentration of ketamine in the0.1-2.5mg/L range. The detection limit was0.05mg/L, with a relative standard deviation (n=11) of1.9%. This method has been applied for the determination of ketamine in injection.Part2Capillary electrophoresis analysis coupled with luminol-Ag(III) complex chemiluminescence detection.(1) A sensitive, simple method is presented for the determination of three major catecholamines in human urine by capillary electrophoresis (CE) with on-line chemiluminescence (CL) detection. This was also the first time that the luminol-Ag(III) complex CL system was used for CE detections. This method was based on the enhancing effect of epinephrine (EP), norepinephrine (NE) and dopamine (DA) on the CL reaction between luminol and the Ag(III) complex in alkaline solution. The separations and determinations were performed with an electrophoretic buffer consisting of20.0mM sodium borate and1.0mM luminol. Under optimized conditions, the three catecholamines were baseline separated and detected in less than8min. Detection limits of7.9×10-8M,1.0×10-7M and6.9×10-8M were observed for EP, NE and DA, respectively. RSD values for the peak height were4.7-5.4%(n=5). Our proposed method was applied to the determinations of the catecholamines in urine samples from12healthy individuals and26pheochromocytoma patients. Our results suggest that this method might be useful to monitor the catecholamine levels in routine screening and to diagnose pheochromocytoma.(2) A capillary electrophoresis instrument coupled with chemilumine-scence detection was designed for the determination of ketamine in urine samples. An important enhancement of the CL emission of luminol with Ag(III) complex was observed in the presence of ketamine; so this system was selected for its detection after CE separation. Under optimal conditions, the limits of detection (LODs, S/N=3) for ketamine is1.0×10-7M, with linear range of2.0x10-7-1.0×10-5M. RSD value for the peak height was4.3%(n=5). The proposed method was tested by analyzing ketamine in urines. The results demonstrate the suitability of this approach to analyze trace amount of ketamine in urine.(3) This method was based on the enhancing effect of salbutamol, terbutaline and ractopamine on the CL reaction of luminol with an Ag(III) complex in alkaline solution. Under optimum conditions, the three (32-agonists were separated on baseline and detected in less than10minutes. The detection limits for salbutamol, terbutaline and ractopamine were determined to be2.5mg/L,0.75mg/L and5.0mg/L, respectively. RSD values for the peak height were3.28-6.1%(n=5).This method was also applied successfully to the analyses of the three β2-agonists contents in some animals’forages and urine samples.(4) Under alkaline conditions, papaverine can significantly enhance the chemiluminescence of the luminol-Ag(III) complex CL system, and the chemiluminescent intensity show good linearity to the concentration of the analytes within a certain range. Based on this fact, a novel method for the analysis of papaverine was developed. The detection limits (S/N=3) for papaverine was0.03g/L, with a linear range of4.0-80.0g/L. RSD value for the peak height was4.3%for6.0g/L of papaverine (n=5). Finally, the presented method was used for the analysis of papaverine in papaverine hydrochloride injection and traditional Chinese medicine preparation Qiangli Pipa Syrup successfully.Part3Pretreatment of tissue and distribution of ketamine in rats poisoned to death.Sixteen male Wistar rats were divided into two groups at random and given intragastric administration of ketamine respectively. Group I was given with the dcse of894mg/kg (twice as much as LD50) and group II was given with the dose of1788mg/kg (four times as much as LD50). The poisoning rats were rapidly dissected after breathing and heartbeat all disappeared, in order to stablish death model of rats died of intragastric administration of ketamine arid to explore the distribution of ketamine in rats died of ketamine poisoning. Peripheral blood, heart, liver, spleen, lung, kidney and brain, muscle tissue were taken and saved in fridge at low temperature. The content of ketamine was determined by high performance liquid chromatographic. Ketamine content order of group Ⅰ was as follows:brain> liver> kidney> lung, spleen, muscle, heart, and peripheral blood. Ketamine content order of group Ⅰ was as follows:brain>kidney> liver> lung, spleen, heart, and peripheral blood muscles. The results indicated that ketamine widely distributed in poisoning rats in vivo and blood, the highest content of ketamine appeared in brain, liver and kidney. It indicates that the inspection material for toxicological analysis should be selected reasonablely in the forensic identification.