A novel solid-state electrochemiluminescence detector for capillary electrophoresis based on tris(2,2'-bipyridyl)ruthenium(II) immobilized in Nafion/PTC-NH2 composite film

A new electrochemiluminescence (ECL) detector for capillary electrophoresis (CE) based on tris(2,2′-bipyridyl)ruthenium(II) (Ru(bpy)₃²⁺) immobilized in Nafion/PTC-NH₂ (an ammonolysis product of 3,4,9,10-perylenetetracarboxylic dianhydride (PTCDA)) composite film was presented for the first time. The Nafion/PTC-NH₂ composite film could effectively immobilize tris(2,2′-bipyridyl)ruthenium(II) via ion-exchange and electrostatic interaction. Cyclic voltammetric and ECL behavior of Nafion/PTC-NH₂/Ru composite film was investigated compared to Nafion/Ru composite. The Nafion/PTC-NH₂/Ru composite film exhibited good ECL stability and simple operability. Then the CE with solid-state ECL detector system was successfully used to detect sophora - a quinolizidine type - alkaloids as sophoridine (SR) and matrine (MT). The CE-ECL parameters that affected separation and detection were optimized. Under the optimized conditions, the linear range was from 2.5 × 10⁻⁸ to 2 × 10⁻⁶ mol/L for SR, 1.0 × 10⁻⁸ to 1.0 × 10⁻⁶ mol/L for MT. The detection limit (S/N = 3) was estimated to be 5 × 10⁻⁹ and 10⁻⁹ mol/L for SR and MT, respectively. It was shown that the CE coupling with solid-state ECL detector system exhibited satisfying sensitivity of analysis.     

Tris(2,2'-bipyridyl)ruthenium(II)-Zirconia-Nafion composite modified electrode applied as solid-state electrochemiluminescence detector on electrophoretic microchip for detection of pharmaceuticals of tramadol, lidocaine and ofloxacin

Tris(2,2′-bipyridyl)ruthenium(II) (Ru(bpy)₃²⁺)-Zirconia-Nafion composite modified glassy carbon disk electrode as a solid-state electrochemiluminescence (ECL) detector is successfully applied to an electrophoretic microchip system with a wall-jet configuration. Pharmaceuticals such as tramadol, lidocaine and ofloxacin were selected to characterize the performance of this microchip capillary electrophoresis (CE)-ECL detection system. Voltammetric and ECL behaviors of immobilized Ru(bpy)₃²⁺ were investigated in lidocaine system. Influences of the separation electric field to cyclic voltammograms (CVs) of the immobilized Ru(bpy)₃²⁺ were also investigated. Tramadol, lidocaine and ofloxacin can be baseline separated without any additives. The detection limits (S/N = 3) were 2.5 × 10⁻⁵ mol L⁻¹ for tramadol, 5.0 × 10⁻⁶ mol L⁻¹ for lidocaine, 1.0 × 10⁻⁵ mol L⁻¹ for ofloxacin under the sample injection of picoliters, and the linear ranges were from 5.0 × 10⁻⁵ to 2.5 × 10⁻³ mol L⁻¹ for tramadol, 1.0 × 10⁻⁵ to 1.0 × 10⁻³ mol L⁻¹ for lidocaine, and 1.0 × 10⁻⁵ to 2.5 × 10⁻³ mol L⁻¹ for ofloxacin, respectively.     

Determination of diphenhydramine by capillary electrophoresis with tris(2,2'-bipyridyl)ruthenium(II) electrochemiluminescence detection

Tris(2,2′-bipyridyl)ruthenium(II) electrochemiluminescence detection in a capillary electrophoresis separation system was used for the determination of diphenhydramine. In this study, platinum disk electrode (300 μm in diameter) was used as a working electrode and the influence of applied potential and buffer conditions were investigated. Under optimal conditions: 1.2 V applied potential, pH 8.50, 15 kV separation voltage and 10 mmol l⁻¹ running buffer, the calibration curve of diphenhydramine was linear over the range of 4×10⁻⁸ to 1×10⁻⁵ mol l⁻¹. This technique gave satisfactory precision, and relative standard deviations of migration times and chemiluminescence peak intensities were less than 1 and 6%, respectively. The technique was applied to animal studies for determination of diphenhydramine extracted from rabbit plasma and urine samples, and the extraction efficiency were between 92 and 98.5%.     

Direct tris(2,2'-bipyridyl)ruthenium (II) electrochemiluminescence detection of polyamines separated by capillary electrophoresis

Capillary electrophoresis (CE) with tris(2,2'-bipyridyl) ruthenium (II) (Ru(bpy)3(2+)) electrochemiluminescence (ECL) detection technique was developed for the analysis of four polyamines (putrescine (Put), cadaverine (Cad), spermidine (Spd), and spermine (Spm)) analysis. The four polyamines contain different amine groups, which have different ECL activity. There are several parameters which influence the resolution and ECL peak intensities, including the buffer pH and concentrations, separation voltage, sample injection, electrode materials, and Ru(bpy)3(2+) concentrations. Polyamines are separated by capillary zone electrophoresis in an uncoated fused-silica capillary (50 cmx25 micro m (ID) filled with acidic phosphate buffer (200 mmol/L phosphate, pH 2.0) - 1mol/L phosphoric acid (9:1 v/v) and a separation voltage of 5 kV (25 micro A), with end-column Ru(bpy)3(2+) ECL detection. A 5 mmol/L Ru(bpy)3(2+) solution plus 200 mmol/L phosphate buffer (pH 11.0) is added into the reagent reservoir. The calibration curve is linear over a concentration range of two or three orders of magnitude for the polyamines. The analysis time is less than 25 min. Detection limits for Put and Cad are 1.9x10(-7) mol/L and 7.6x10(-9) mol/L for Spd and Spm, respectively. Intraday and interday relative standard deviations of ECL peak intensities are less than 8%. The main advantages of this CE-ECL detection technique for polyamines analysis presented herein are the omission of chemical derivatization of the analytes and the high selectivity.     

Simultaneous determination of pethidine and methadone by capillary electrophoresis with electrochemiluminescence detection of tris(2,2′-bipyridyl)ruthenium(II)

In this paper, we described a simple and rapid method, capillary electrophoresis with electrochemiluminescence (CE–ECL) detection using tris(2,2′-bipyridyl)ruthenium(II) (Ru(bpy)₃²⁺), to simultaneously detect pethidine and methadone. Analytes were injected to separation capillary of 67.5 cm length (25 μm i.d., 360 μm o.d.) by electrokinetic injection for 10 s at 10 kV. Under the optimized conditions: ECL detection at 1.20 V, 30 mM sodium phosphate (pH 6.0) as running buffer, separation voltage at 14.0 kV, 5 mM Ru(bpy)₃²⁺ with 50 mM sodium phosphate (pH 6.5) in the detection cell, the linear range from 2.0 × 10^− 6 to 2.0 × 10^− 5 M for pethidine and 5.0 × 10^− 6 to 2.0 × 10^− 4 M for methadone and detection limits of 0.5 μM for both of them were achieved (S/N = 3). Relative standard derivations of the ECL intensity were 2.09% and 6.59% for pethidine and methadone, respectively.     

Tris(2,2'-bipyridyl)ruthenium(II) electrochemiluminescence sensor based on carbon nanotube/organically modified silicate films

In this paper, a novel electrochemiluminescence (ECL) sensor was constructed to determine herring sperm (HS) double-stranded (ds) DNA. Tetramethoxysilane and dimethyldimethoxysilane were selected as co-precursors to form an organically modified silicate (ORMOSIL) film for the immobilization of multiwall carbon nanotubes (MWNTs) wrapped by poly(p-styrenesulfonate) (PSS), and then Tris(2,2′-bipyridyl)ruthenium(II) (Ru(bpy)₃²⁺) was successfully immobilized on a glassy carbon electrode via ion-association. PSS was employed to increase the conductivity of the ORMOSIL film and disperse the cut MWNTs, which were cut and shortened in a mixture of concentrated sulfuric and nitric acids, in the film. It was found that MWNTs could adsorb Ru(bpy)₃²⁺ and acted as conducting pathways to connect Ru(bpy)₃²⁺ sites to the electrode. MWNTs also played a key role as materials for the mechanical and thermal properties. The ECL performance of this modified electrode was evaluated in a flow injection analysis (FIA) system, and the detection limit (S/N = 3) for HS ds-DNA was 2.0 × 10⁻⁷ g mL⁻¹ with a linear range from 1.34 × 10⁻⁶ to 6.67 × 10⁻⁴ g mL⁻¹ (R² = 0.9876). In addition, the ECL sensor presented excellent characteristics in terms of stability, reproducibility and application life.     

Electrochemical detector based on sol-gel-derived carbon composite material for capillary electrophoresis microchips

An on-chip disk electrode based on sol-gel-derived carbon composite material could be easily and reproducibly fabricated. Unlike other carbon-based electrodes reported previously, this detector is rigid, convenient to fabricate, and amenable to chemical modifications. Based on the stable and reproducible characters of this detector, a copper particle-modified detector was developed for the detection of carbohydrates which extends the application of the carbon-based electrode. In our experiments, the performance of the new integrated detector for rapid on-chip measurement of epinephrine and glucose was illustrated. Experimental procedures including the fabrication of this detector, the configuration of separation channel outlet and electrode verge, and the performance characteristics of this new electrochemical detector were investigated.     

毛细管电泳电致化学发光检测血浆中布比卡因

布比卡因是一种外科局部麻醉剂,使用过量会导致中枢神经系统和心脏血管系统中毒^[1],可引起心脏停博.高效液相色谱和毛细管电泳(CE)^[2]是该药常用的检测方法.     

Tris (2,2′-bipyridyl)ruthenium (III) as a chemiluminescent reagent for detection in capillary electrophresis

A post- column chemiluminescent technique for thedetection of compounds that are poor chromoshores using electorogenerated chemiluminescence following separation by capillartgy electrophoresis is described. The luminrescent signal is generated followintg the reaction of anlyres (e.g. amines) with Ru(bpy)₃³⁺, which isx electrochemically generated post-columan from Ru(bpy)₃²⁺. Tripropylamine and proline are used as two model compounds to demostrate the feasibility of the method. Detection limits for the prototype system were in the micromolar rage, suggesting that this technnique offers an alternative to indirect detection of compounds that are poor chromophores with an added selectivity advangage. The system includes the use of a conductive joint to isolate the separation field from the potential necessary to drive the elctrogenerated chemiluminescent reactiion. Addition of the chemiluminescent reagent Ru(bpy)₃²⁺ post-column did not decrease the efficiency of the separation. The design and favrication of the novel cell is discussed.     

One-step immobilization of tris(2,2'-bipyridyl)ruthenium(II) via vapor-surface sol-gel deposition towards solid-state electrochemiluminescence detection

A novel method for immobilization of tris(2,2′-bipyridyl)ruthenium(II) (Ru(bpy)₃Cl₂) on electrode surfaces based on the vapor-surface sol-gel deposition strategy is first demonstrated in this paper. Ru(bpy)₃Cl₂ immobilized sol-gel (Ru(bpy)₃Cl₂/sol-gel) films were characterized by UV-vis spectroscopy and field-emitted scanning electron microscopy (FE-SEM). These results showed that Ru(bpy)₃Cl₂ was successfully incorporated into the silica sol-gel film. It was found that many irregular Ru(bpy)₃Cl₂/sol-gel clusters were formed on surfaces through one deposition and thick sol-gel films were observed after further deposition. Electrochemical properties and electrochemiluminescence (ECL) behaviors of Ru(bpy)₃Cl₂/sol-gel films could be easily adjusted by deposition numbers and time. At last, the Ru(bpy)₃Cl₂/sol-gel film modified electrode was used for solid-state ECL detection of tripropylamine. The linear range was from 5.8 × 10⁻⁸ to 2.4 × 10⁻⁴ M with the detection limit of 5 nM, which was three orders of magnitude lower than that from pure Nafion-modified electrodes. The ECL sensor also exhibited high stability, and still remained 92% response after being stored in air for 35 days. This method for immobilization of Ru(bpy)₃Cl₂ is simple, convenient and low-cost relative to others, so it shows promising applications in solid-state ECL detection.     

Determination of atenolol and metoprolol by capillary electrophoresis with tris(2,2'-bipyridyl)ruthenium(ii) electrochemiluminescence detection.

Capillary electrophoresis (CE) coupling with a tris(2,2'-bipyridyl)ruthenium(II) (Ru(bpy)(3)(2+)) electrochemiluminescence (ECL) detection technique was developed for the analysis of two beta-blockers, atenolol (AT) and metoprolol (ME). The parameters that influence the separation and detection, including the buffer pH and concentration, the separation voltage, the detection potential and Ru(bpy)(3)(2+) concentration, were optimized in detail. The calibration curve was linear over a concentration range of two or three orders of magnitude for the two beta-blockers. The detection limits for AT and ME were 0.075 and 0.005 microM (S/N = 3). The relative standard deviations (n = 8) of the ECL intensity and the migration time were 2.65 and 0.22% for AT, 2.82 and 0.34% for ME, respectively. The proposed method was applied to determine AT and ME in spiked urine samples; satisfactory results were obtained.     

Analysis of quinolizidine alkaloids in Sophora flavescens Ait. by capillary electrophoresis with tris(2,2'-bipyridyl) ruthenium (II)-based electrochemiluminescence detection

A capillary electrophoresis method coupled with electrochemiluminescence detection for the analysis of quinolizidine alkaloids was established, especially, oxymatrine (OMT) which could not be measured by previous electrochemiluminescence methods was detected sensitively herein. Complete separation of sophoridine (SR), matrine (MT) and OMT was achieved within 13 min using a background electrolyte of 50mM phosphate buffer at pH 8.4 and a separation voltage of 15 kV. The calibration curves showed a linear range from 2.8 x 10(-8) to 4.4 x 10(-7) M for SR, 2.7 x 10(-8) to 4.4 x 10(-7) M for MT, and 2.5 x 10(-7) to 4.0 x 10(-6)M for OMT, respectively. The relative standard derivations for all analytes were below 3.1%. Good linear relationships were showed with correlation coefficients for all analytes exceeded 0.987. The detection limits were 1.0 nM for SR and MT, and 40 nM for OMT under the optimal conditions, respectively. The developed method was nearly harmless to the human and environment.     

Determination of chlorpheniramine and its binding with human serum albumin by capillary electrophoresis with tris(2,2'-bipyridyl)ruthenium(II) electrochemiluminescence detection.

Tris(2,2'-bipyridyl)ruthenium(II) electrochemiluminescence (ECL) detection in a capillary electrophoresis separation system was used for the determination of chlorpheniramine (CPM). The experimental conditions, such as the applied potential, separation voltage, injection voltage, injection time and the pH of the separation buffer were considered in detail. The ECL intensity showed two linear responses to CPM, i.e., from 15 microM to 1 mM and from 0.8 microM to 15 microM with a detection limit of 0.5 microM. The binding of CPM with human serum albumin was also monitored using this method and the binding constant was estimated to be 4.1 x 103 M(-1).     

毛细管电泳-473 nm固体激光诱导荧光检测系统的建立

以发射波长473nm的半导体激光泵浦固体激光器(LD DPSSL)为激发光源,研制了一种小型模块化激光诱导荧光检测器。以异硫氰酸荧光素(FITC)为荧光探针,毛细管电泳柱上检测(0.05mmi.d)评价了该体系,得到了5×10-12mol L的浓度检出限。利用该系统考察了氨基酸、实际样品中B族维生素的检测。     

Detection of clindamycin by capillary electrophoresis with an end-column electrochemiluminescence of tris(2,2'-bypyridine)ruthenium(II)

Coupled capillary electrophoresis (CE) with end-column electrogenerated chemiluminescence (ECL) was adopted for the quantitative detection of clindamycin. Clindamycin enhanced ECL intensity of tris(2,2'-bypyridine)ruthenium(II) (Ru(bpy)(3)(2+)) as a coreactant. Under the optimized conditions, the ECL intensity was linear with the concentration of clindamycin over the range from 5.0 x 10(-7) to 1.0 x 10(-4)M with a detection limit of 1.4 x 10(-7)M. The proposed CE-ECL was successfully applied for the detection of clindamycin in pharmaceutical and clinic samples. The interaction of clindamycin with hemoglobin was also investigated. The binding constant of clindamycin with hemoglobin was estimated to be 3.6 x 10(3)M(-1).     

Tris(2,2′-bipyridyl)ruthenium(II) electrogenerated chemiluminescence sensor based on carbon nantube dispersed in sol-gel-derived titania-Nafion composite films

A highly sensitive and stable tris(2,2′-bipyridyl)ruthenium(II) (Ru(bpy)₃²⁺) electrogenerated chemiluminescence (ECL) sensor was developed based on carbon nanotube (CNT) dispersed in mesoporous composite films of sol-gel titania and perfluorosulfonated ionomer (Nafion). Single-wall (SWCNT) and multi-wall carbon nanotubes (MWCNT) can be easily dispersed in the titania-Nafion composite solution. The hydrophobic CNT in the titania-Nafion composite films coated on a glassy carbon electrode certainly increased the amount of Ru(bpy)₃²⁺ immobilized in the ECL sensor by adsorption of Ru(bpy)₃²⁺ onto CNT surface, the electrocatalytic activity towards the oxidation of hydrophobic analytes, and the electronic conductivity of the composite films. Therefore, the present ECL sensor based on the CNT-titania-Nafion showed improved ECL sensitivity for tripropylamine (TPA) compared to the ECL sensors based on both titania-Nafion composite films without CNT and pure Nafion films. The present Ru(bpy)₃²⁺ ECL sensor based on the MWCNT-titania--Nafion composite gave a linear response (R² = 0.999) for TPA concentration from 50 nM to 1.0 mM with a remarkable detection limit (S/N = 3) of 10 nM while the ECL sensors based on titania-Nafion composite without MWCNT, pure Nafion films, and MWCNT-Nafion composite gave a detection limit of 0.1 μM, 1 μM, and 50 nM, respectively. The present ECL sensor showed outstanding long-term stability (no signal loss for 4 months).     

Effect of Ionic Strength on the Electrochemiluminescence Generation by Tris(2,2'-bipyridyl)ruthenium(II)/Tri-n-propylamine

Electrochemiluminescence(ECL) is a powerful transduction technique used in biosensing and in vitro diagnosis, while the mechanism of ECL generation is complicated and affected by various factors. Herein the effect of ionic strength on ECL generation by the classical tris(2,2'-bipyridyl)ruthenium(II)[Ru(bpy)₃²⁺]/tri-n-propylamine(TPrA) system was investigated. It is clear that the ECL intensity decreases significantly with the increase of ionic strength, most likely arising from the reduced deprotonation rate of TPrA^+·. We further combined microtube electrode(MTE) with ECL microscopy to unravel the evolution of ECL layer with the variation of ionic strength. At a low concentration of Ru(bpy)₃²⁺, the thickness of ECL layer(TEL) nearly kept unchanged with the ionic strength, indicating the surface-confined ECL generation is dominated by the oxidative-reduction route. While at a high concentration of Ru(bpy)₃²⁺, ECL generation is dominated by the catalytic route and TEL increases remarkably with the increase of ionic strength, because of the extended diffusion length of Ru(bpy)₃³⁺ at a reduced concentration of TPrA^·.     

Effective sorbents for solid-phase extraction in the analysis of quinolones in animal tissues by capillary electrophoresis

In this work, several commercial sorbents (Zorbax C18, Bond Elut C18, Isolute ENV+, Oasis HLB, Oasis MAX, SDB-RPS, and MPC-SD) were compared for the solid-phase extraction of the series of quinolones regulated by the European Community in chicken tissues in order to establish a method for the determination of this series of quinolones by capillary electrophoresis and diode array detection. Sorbents were chosen in order to achieve maximum recoveries and optimal clean-up efficiency. Better results were obtained using SDB-RPS and Oasis MAX which would provide suitable limits of detection, below of the maximum residue limits (MRLs) regulated.