Capillary Electrophoresis Chemiluminescence Research Articles

Simultaneous determination of four phenolic acids in traditional Chinese medicine by capillary electrophoresis-chemiluminescence.

Chlorogenic, ferulic, vanillic, and caffeic acids are phenolic acids found in natural drugs. They possess the biological activities of scavenging free radicals and inhibiting thrombus formation. Phenolic acids can inhibit the oxidation of low-density lipoprotein, as well as have anti-inflammatory effects. This paper reports for the first time a capillary electrophoresis–chemiluminescence (CE–CL) method for the simultaneous determination of the four phenolic acids found in traditional and proprietary Chinese medicine, including Lycium chinense Miller, Shuanghuanglian oral liquid, and Taraxacum mongolicum granules. Capillary electrophoretic separation was performed on a self-assembled CE–CL device with an uncoated fused-silica capillary (66 cm effective length, 50 μm i.d.), and the background electrolyte was composed of 3.0 × 10−5 M Ag(iii) (pH = 12.01), 3.0 mM luminol (pH = 9.20), and 10 mM sodium tetraborate solution. The injection time was 12 s (under gravity) and the separation voltage was 22 kV. The combination of solid-phase extraction (SPE) and CE–CL improves the sensitivity. Under optimal conditions, calibration graphs displayed a linear range between 0.625 and 20.0, 1.000 and 30.0, 0.150 and 1.50, and 0.045 and 1.00 μg mL−1 for chlorogenic, ferulic, vanillic, and caffeic acid, respectively. The detection limit ranged from 0.014 to 0.300 μg mL−1. The practicality of using the proposed method to determine the four target analytes in traditional Chinese medicine was also validated, in which recoveries ranged from 90.9% to 119.8%. Taken together, these results indicate that the developed method is sensitive and reliable. Furthermore, the method was successfully applied to real traditional Chinese medicine samples.

A novel chemiluminescence signal amplification strategy based on a capillary electrophoresis platform for highly sensitive competitive immunoassay of biomolecules

A novel chemiluminescence (CL) signal amplification strategy based on luminophore and antigen co-loaded gold nanoparticles was developed to improve the sensitivity of capillary electrophoresis CL assay.

Capillary electrophoresis chemiluminescence assay of naphthol isomers in human urine and river water using Ni(IV) complex-luminol system.

A capillary electrophoresis method involving online indirect chemiluminescence (CL) detection was used to determine naphthol (NAP) isomers. The method was based on the quenching effect of 1- and 2-NAP on a new CL reaction of luminol with Ni(IV) complex in an alkaline medium. Separation was conducted with a 25.0 mM sodium borate buffer containing 0.8 mmol/L luminol. Under optimized conditions, 1- and 2-NAP were baseline separated and detected in less than 8 min. The limits of detection of 1- and 2-NAP were 3.1 and 2.7 μg/L, respectively (S/N = 3), with a linear range of 4.0-80.0 μg/L (r > 0.995). Analysis of real samples demonstrated that the spiked recoveries were in the range of 89.2-107.5% (n = 3). The proposed method was successfully used to determine 1- and 2-NAP contents in three environmental water samples and 14 human urine samples. No derivatization or tedious pretreatment was required in the analysis. The proposed method is a potential approach for routine tests of naphthol isomers in a facile CE-CL system.

Quantum dot-modified aptamer probe for chemiluminescence detection of carcino-embryonic antigen using capillary electrophoresis

A new method is proposed for determination of carcino-embryonic antigen (CEA) based on quantum dot (QD)-modified aptamer probe by capillary electrophoresis-chemiluminescence (CE-CL) detection system. An oligonucleotide sequence A contains two parts, one is CEA aptamer and the other is bridging segment. Horseradish peroxidase (HRP) conjugates with sequence A (HRP-DNAA) and then mixes with QD labeling complementary bridging segment (QD-DNAB), forming HRP-DNAA-B-QD probe. When CEA exists, the specific combination of the aptamer in sequence A and CEA can form CEA/HRP-DNAA-B-QD complex. After separation of CEA/HRP-DNAA-B-QD complex and HRP-DNAA-B-QD probe by capillary electrophoresis (CE), the chemiluminescence (CL) catalyzed by CEA/HRP-DNAA-B-QD complex can be detected without any interference, and the content of CEA can be estimated by the CL intensity. It has been proved that the separation efficiency of HRP-DNAA-B-QD and CEA/HRP-DNAA-B-QD complex is improved greatly after QD modification, then the interference issue resulted from free HRP-DNAA-B-QD probe is solved well; and the specific combination of HRP-DNAA-B-QD and CEA leads to a closer distance of HRP and QD, then chemiluminescent resonance energy transfer (CRET) occurs, which confirms the validity of the strategy. Results show that the CL intensity has a linear relationship with the concentration of CEA in the range from 0.0654 to 6.54ng/mL, and the limit of detection is approximately 8pg/mL (S/N=3). This proposed method with high specificity has been applied for the accurate analysis of content of CEA in patient's serum.

Capillary electrophoresis-chemiluminescence detection for carcino-embryonic antigen based on aptamer/graphene oxide structure

A new strategy is proposed for determination of carcino-embryonic antigen (CEA) based on aptamer/graphene oxide (Apt/GO) by capillary electrophoresis-chemiluminescence (CE-CL) detection system. CEA aptamer conjugated with horseradish peroxidase (HRP) firstly mixes with GO, and the CL will be quenched because the stack of HRP–Apt on GO leads to chemiluminescence resonance energy transfer (CRET). When CEA exists, the specific combination of HRP–Apt and CEA can form HRP–Apt–CEA complex, which dissociates from GO. Then, the CL catalyzed by HRP–Apt–CEA complex can be detected without any CRET, and the content of CEA can be estimated by the CL intensity. It has been proved that the interference issue resulted from free HRP–Apt is solved well by mixing GO firstly with HRP–Apt, which blocks the free HRP–Apt's CL signal due to CL quenching effect of GO; and the interference resulted from GO to CL is also solved by CE, then the sensitivity and accuracy can be greatly improved. Results also showed that the CL intensity had a linear relationship with the concentration of CEA in the range from 0.0654 to 6.54ng/mL, and the limit of detection was approximately 4.8pg/mL (S/N=3). This proposed method with high specificity offers a new way for separation and determination of biomolecule, and has good potential in application of biochemistry and bioanalysis.

Electrophoresis–chemiluminescence detection of phenols catalyzed by hemin

Based on the catalytic activity of hemin, an efficient biocatalyst, an indirect capillary electrophoresis-chemiluminescence (CE-CL) detection method for phenols using a hemin-luminol-hydrogen peroxide system was developed. Through a series of static injection experiments, hemin was found to perform best in a neutral solution rather than an acidic or alkaline medium. Although halide ions such as Br(-) and F(-) could further enhance the CL signal catalyzed by hemin, it is difficult to apply these conditions to this CE-CL detection system because of the self-polymerization of hemin, as it hinders the CE process. The addition of concentrated ammonium hydroxide to an aqueous/dimethyl sulfoxide solution of hemin-luminol afforded a stable CE-CL baseline. The indirect CE-CL detection of five phenols using this method gave the following limits of detections: 4.8 × 10(-8) mol/L (o-sec-butylphenol), 4.9 × 10(-8) mol/L (o-cresol), 5.4 × 10(-8) mol/L (m-cresol), 5.3 × 10(-8) mol/L (2,4-dichlorophenol) and 7.1 × 10(-8) mol/L (phenol).

Capillary Electrophoresis Chemiluminescence for the Analysis of Flavonoids in Pharmaceuticals and Human Plasma

A capillary electrophoresis (CE)-chemiluminescence (CL) method has been developed for the determination of the pharmacologically active flavonoids, including rutin and quercetin in pharmaceuticals and human plasma containing Ginkgo biloba leaves extract (EGb). The separation was conducted in borate buffer and luminol. The post-column CL reagent was K3Fe(CN)6 in NaOH medium. Rutin and quercetin were baseline separated within 10 min with detection limits of 1.0 and 5.0 nM, respectively. The maximum intra- and inter-day relative standard deviations (RSD) of migration time of analytes were <3.5 %. Moreover, the high selectivity of the CL detection and the high-separation efficiency of CE render the method the potential of quick analyzing the pharmacologically active substances in complex matrix with satisfactory results.

Determination of alkaloids inSinomenium acutumby field-amplified sample stacking in capillary electrophoresis with chemiluminescene detection

A simple and rapid capillary electrophoresis (CE) with an acidic potassium permanganate chemiluminescence (CL) detection method was developed to determine three alkaloids (curine, sinomenine and magnoflorine) simultaneously. A laboratory-built CE-CL detection interface was used. The field-amplified sample stacking technique was applied to the online concentration of alkaloids. Experimental conditions for CE separation and CL detection were investigated in detail to acquire optimum conditions. Under optimal conditions, the three alkaloids were baseline separated within 6 min, and the detection limits (S/N = 3) ranged from 0.03 µg/mL to 0.49 µg/mL. This method was successfully applied to determine the above three alkaloids in Sinomenium acutum, and the result of the determination of sinomenine was in good agreement with those given by high-performance liquid chromatography and CE methods. In addition, a possible CL reaction mechanism of sinomenine-KMnO4-H2SO4 was proposed.

Ultrasensitive determination of epicatechin, rutin, and quercetin by capillary electrophoresis chemiluminescence

Summary An ultrasensitive and rapid method for the determination of epicatechin, rutin, and quercetin was developed using capillary zone electrophoresis with on-line chemiluminescence detection. Under the optimal conditions, the analytes were baseline separated within 12 min. The limits of detection in turn were 0.60 pg mL−1 for epicatechin, 0.50 pg mL−1 for rutin, and 1.0 pg mL−1 for quercetin. The developed method was an easy and reliable method of determining these analytes concentrations in tea, extract Ginkgo biloba, and rutin tablet, demonstrating the feasibility and reliability of the proposed method.

Development of a compact capillary electrophoresis–chemiluminescence system with ultra‐fast peroxyoxalate reaction to monitor the hydrolysis of rhodamine 6G

A simple and effective capillary electrophoresis-chemiluminescence (CE-CL) detection system was developed based on an ultra-fast bis(2,4,6-trichlorophenyl)oxalate (TCPO) chemiluminescence (CL) reaction (0.6 s duration) that avoided overlapping peaks and peak tailing. Through a series of static injection experiments, this unusually rapid CL reaction was ascribed to the catalytic effect of imidazole in the tetrahydrofuran solvent, which has been rarely utilized in such investigations. A possible mechanism is given to explain the results. Under the optimized conditions, rhodamine 6 G (R6G) and its hydrolysis product (R6G-COOH) could be efficiently separated through electrophoresis in 7 min, with sensitive CL detection in the proposed CE-CL system. In this way, the alkaline hydrolysis of R6G was monitored, followed by estimation of relative rate constants and activation energy. This finding and application should be helpful in further study for the TCPO CL reaction, and revealed an attractive opportunity for simplifying the CE-CL system, such as in a microchip device.

Quantum dot-enhanced chemiluminescence detection for simultaneous determination of dopamine and epinephrine by capillary electrophoresis

A sensitive method based on quantum dot (QD)-enhanced capillary electrophoresis–chemiluminescence (CE–CL) detection was developed for simultaneous determination of dopamine (DA) and epinephrine (E). In this work, CdTe QD was added into the running buffer of CE to catalyze the post-column CL reaction between luminol and hydrogen peroxide, achieving higher CL emission. Negative peaks were produced due to the inhibitory effects on CL emission from DA and E eluted from the electrophoretic capillary. The decrease in CL intensity was proportional to the concentration of DA and E in the range of 8.0×10−8–5.0×10−6M and 4.0×10−8–5.0×10−6M, respectively. Detection limits for DA and E were 2.3×10−8M and 9.3×10−9M, respectively. Using this method, the levels of DA and E in human urine from healthy donors were determined.

On-line chemiluminescence determination of mitoxantrone by capillary electrophoresis

A novel capillary electrophoresis (CE) with chemiluminescence (CL) detection method for the determination of mitoxantrone (MTX) has been developed, which based on the CL reaction of potassium ferricyanide with luminol in sodium hydroxide medium sensitized by MTX. Under optimum analytical conditions, MTX is determined over the range of 7.0 × 10 −8–1.0 × 10 −6 M with a detection limit of 1.0 × 10 −8 M. The relative standard deviation (RSD) was 3.7%, 2.6% and 3.0% for 7.0 × 10 −8, 5.0 × 10 −7 and 1.0 × 10 −6 M MTX ( n = 11), respectively. In laboratory-built CE–CL apparatus, the proposed method has been applied to determination of MTX in commercial drug and spiked in human urine and plasma with satisfactory results.

An on-column fracture/end-column reaction interface for chemiluminescence detection in capillary electrophoresis

Chemiluminescence (CL) offers a sensitive detection method for capillary electrophoresis (CE), but the implementation of CE–CL is usually under compromised operating conditions for CE, such as the prerequisite of extreme pH buffer for optimal CL reaction at the capillary outlet. This has sometimes significantly deteriorated the separation of CE. In this study, the development of a new interface makes it possible to optimize the operating conditions for CE separation and CL detection independently. The interface consists of an on-column fracture being installed in a reservoir near the capillary end to create an electrical connection and also serve as reagent addition entrance. The capillary terminal is inserted into an end-column reservoir for CL reaction and detection. In this arrangement, the applied electric field has been decoupled from the CL detection, which is proved to effectively improve CE's performance by allowing the use of optimal CE buffers. At the same time, it enables the optimization of CL detection independently. The applicability of this interface was evaluated by using acridinium ester (AE) and luminol systems. For AE system, the interfering products of CL reagent ( −OH, HO 2 −) have been prevented, and the pH range of CE buffer can be independent to the optimal pH value of AE CL reaction, which is usually below 3. The AE was detected using running buffer at pH 8.7, giving a detection limit of 0.1 nM (S/N = 3), and the theoretical plate numbers is as high as 56 000. The on-column fracture based configuration is simple, sensitive and easy to implement.

Development and Evaluation of a Rotary Cell for Capillary Electrophoresis−Chemiluminescence Detection

Many efforts have been made toward the advancement of capillary electrophoresis chemiluminescence (CE-CL) detection and its applications through continuous development and improvement of interfaces. In this study, a novel rotary cell for CE-CL detection was fabricated and evaluated. A ring-shaped narrow channel with a quartz bottom is made in a cell body to hold CL reactants and act as the reaction chamber. The CE capillary is placed closely to the bottom of the reaction chamber where analyte is deposited into the CL reactants for reactions to occur. Detection is achieved with a photomultiplier tube below the reaction channel. An advantage of the rotary reaction cell is that it renews the reactants at the capillary end as it revolves at a preset speed during experiments. The rotary detection cell presents a new concept to the conventional coaxial-flow configuration by solving the problems of bubble formation and flow blockage that often interrupt the electrophoresis. Two standard proteins, horseradish peroxidase (HRP) and hemoglobin (Hb), were used to evaluate the interface's performance with luminol/H(2)O(2) CL system. Satisfactory sensitivities (LOD of 0.91 x 10(-9) M for HRP, and 4.37 x 10(-8) M for Hb at S/N = 3) were obtained in this proof-of-concept experiment. This novel design provides a straightforward and robust interface for CE-CL hyphenation.

Electrogeneration of Ferrate(VI) in Low-concentration NaOH Solution for Capillary Electrophoresis–Chemiluminescence Detection

Abstract A novel, eco-friendly, and stable homogeneous luminescent system for capillary zone electrophoresis (CE) detection was established, based on the chemiluminescence (CL) flash caused by mixing ferrate(VI) with luminol solution. It was found that the stable ferrate(VI) could be freshly electrogenerated in low-concentration NaOH solution for direct analytical application. To test this CL system, 8-hydroxyquinoline (HQ) was chosen as a model sample by CE detection.

Ultrasensitive determination of cobalt in single hair by capillary electrophoresis using chemiluminescence detector

By use of the high separation capability of capillary electrophoresis (CE) and ultrasensitive chemiluminescence (CL) detection, a CE–CL method was proposed for the ultrasensitive determination of trace cobalt in a single hair, which is potentially useful in construction of the fingerprint of trace elements in the hair collected from crime scenes. In this work, the CE experimental conditions, CL experimental conditions and the digestion methods for the analysis of a single hair sample for cobalt were investigated in detail. The relative limit of detection (LOD, 3 σ) was 0.01 ng/mL, and the absolute LOD was 2.4 × 10 − 16 g considering the sampling volume of 24 nL. Using a dry digestion method, the analytical results for certified reference hair samples by the proposed method were in good agreement with the certified values. Finally, this method was successfully used to detect trace cobalt in a single hair from three adults. It has potential applications in forensic analysis.

Quantification of Carcinoembryonic Antigen by Capillary Electrophoresis–Chemiluminescence Detection using Internal Standard Method

Sensitive determination of tumor marker carcinoembryonic antigen (CEA) by capillary electrophoresis–chemiluminescence detection (CE-CL) has been developed using an internal standard method. Parameters affecting the CE separation and CL detection were systematically optimized. Under the optimal conditions, the free horseradish peroxidase (HRP)–labeled antibody (Ab*), the bound Ab*-antigen complex (Ab*-Ag), and HRP used as internal standard were well separated within 5 min. The proposed method was successfully applied for the quantification of CEA in human sera obtained from both healthy persons and patients.

Development of a novel capillary electrophoresis chemiluminescence system for amino acid analysis

In the present study, a novel peroxyoxalate CE-CL system was developed to achieve high signal stability and sensitivity based on a design of a new interface including a new mixing mode and a new grounding electrode mode. Amino acids fluorescently tagged with dansyl chloride and naphthalene-2,3-dicarboxaldehyde(NDA) were used for the study. Experiment results show this new system is quite effective to separate and detect amine acid with high stability and resolute. The detection limits were 1.1 nmol/L for dansyl-leucine (Leu) and 2.0 nmol/L for dansyl-aspartic acid (Asp). The relative standard deviations of peak height and migration time were in the ranges of 2.3-3.8% and 1.2-1.5%, respectively.

Determination of uric acid in human urine and serum by capillary electrophoresis with chemiluminescence detection

A simple and sensitive method based on capillary electrophoresis (CE) with chemiluminescence (CL) detection has been developed for the determination of uric acid (UA). The sensitive detection was based on the enhancement effect of UA on the CL reaction between luminol and potassium ferricyanide (K 3[Fe(CN) 6]) in alkaline solution. A laboratory-built reaction flow cell and a photon counter were deployed for the CL detection. Experimental conditions for CL detection were studied in detail to achieve a maximum assay sensitivity. Optimal conditions were found to be 1.0 × 10 −4 M luminol added to the CE running buffer and 1.0 × 10 −4 M K 3[Fe(CN) 6] in 0.2 M NaOH solution introduced postcolumn. The proposed CE–CL assay showed good repeatability (relative standard deviation [RSD] = 3.5%, n = 11) and a detection limit of 3.5 × 10 −7 M UA (signal/noise ratio [S/N] = 3). A linear calibration curve ranging from 6.0 × 10 −7 to 3.0 × 10 −5 M UA was obtained. The method was evaluated by quantifying UA in human urine and serum samples with satisfactory assay results.

A sensitive immunoassay for determination of hepatitis B surface antigen and antibody in human serum using capillary electrophoresis with chemiluminescence detection

A sensitive and homogeneous immunoassay (IA) based on capillary electrophoresis (CE) with enhanced chemiluminescence (CL) detection has been developed for the determination of hepatitis B surface antigen (HBsAg) and antibody (HBsAb) in human serum. The conditions for the CL reaction and electrophoresis were investigated in detail using horseradish peroxidase (HRP) labeled HBsAg (HBsAg*) as a marker because of its catalytic effects on the luminol–hydrogen peroxide reaction. The CL reaction was enhanced by para-iodophenol and the CL detector was designed uniquely without any dead volume or diluents effect. The present method has been used for assaying HBsAg and HBsAb in human serum using a competitive format and a non-competitive format, respectively. Under the optimal conditions, the linear ranges were from 1 to 400 pmol/L ( R = 0.9988) for HBsAg and 2 to 200 mIU/mL ( R = 0.9981) for HBsAb. The detection limits were 0.4 pmol/L and 1 mIU/mL for HBsAg and HBsAb, respectively. The relative standard deviations of peak area were 4.2% and the errors of it were from −0.03% to +0.05% for 80 pmol/L HBsAg* ( n = 7). In this study, the free HBsAg* and the bound HBsAg* (HBsAg*–HBsAb) were separated in the separation capillary within 6 min using a borate run buffer. To verify the experimental reliability, the result was comparable with that of enzyme linked immunosorbent assay (ELISA) and demonstrated the feasibility of the CE–CL immunoassay method for clinical diagnosis.