Amperometric Detection In Capillary Electrophoresis Research Articles

Graphene Oxide Modified Electrode for DNA Amperometric Detection Using Microfluidic Electrophoresis Chip

The amperometric technique of capillary electrophoresis (CE) is an electrochemical detection method with various advantages that is suitable for biosensors. The amperometric detection system can be easily integrated with the CE system without the need for complicated additional equipment for optical detection. For this reason, the size of the sensor can be reduced to micro-scale, also reducing the cost of the fabrication process, enabling this system to be used in point-of-care applications [1]. The optical method depends on the detection of the fluorescence intensity of the target sample. However, amperometric detection omits the pretreatment process of the sample, such as purification extraction of samples and fluorescent staining for detection [2]. These features allow the device system to be miniaturized and having a rapid response for applications like lab-on-a-chip [3]. Modified Au electrodes integrated with PDMS based microfluidic channels were proposed as an on-chip amperometric detection of CE for DNA sensors. The graphene oxide (GO)-modified Au electrodes were fabricated via photolithography process and drop-casting deposition. First, the deposited Au electrodes were patterned with positive photoresist (PR) using the photolithography method. Next, the graphene oxide was deposited 5 layers on the bare Au using drop-casting technique. Finally, the PR was dissolved by acetone and the unwanted GO was lifted-off. The surface characterization of this the GO-modified Au electrodes is shown in Fig. 1(a) by microscopy image. It was observed that the GO was deposited on Au electrodes following the desired pattern. This treatment amplifies the current signal of amperometric detection of the CE microchip. The performance of the CE amperometric detection device has been demonstrated by sensing an amplicon lambda phage DNA. Then, 1 µL of amplified lambda phage DNA (1 ng/µL) using the conventional polymerase chain reaction (PCR) with a 30-cycle reaction was used as target sample. Figure 1(b) shows the current peak of amperometric detection before (1.9 µA) and after (6.6 µA) GO-modified on the Au electrodes which amplified the sensitivity more than 3.5 times before electrode modification. The emphasized detection of lambda phage DNA using modified electrodes gives a further possibility of biomedical applications through our developed microchip.

A decoupler-free simple paper microchip capillary electrophoresis device for simultaneous detection of dopamine, epinephrine and serotonin.

This paper demonstrates a new and simplified configuration for capillary electrophoresis-amperometric detection (CE-AD) using a paper microfluidic chip incorporating inexpensive wax printing and screen printing based methods and then used for electrophoretic separation and simultaneous in-channel amperometric detection of three clinically relevant neurochemicals in a single run without using any decouplers. Detection of neurochemicals e.g., dopamine, epinephrine and serotonin is crucial for early prediction of neurological disorders including Parkinson's, Alzheimer's, dementia, as well as progressive neuro-psychiatric conditions such as depression, anxiety, as well as certain cardiovascular diseases. The plasma concentrations of such neurochemicals are as important as those present in cerebrospinal fluid (CSF) and can be useful for rapid and convenient biosensing. However, simultaneous detection of such neurochemicals in a complex mixture such as human serum requires their separation prior to detection. With the developed microchip, separation and detection of the neurochemicals were exhibited within 650 seconds without pre-treatment and the procedure was validated with spiked fetal bovine serum samples. Beside this, the developed paper microfluidic chip has potential to be integrated in point-of-care diagnosis with onsite detection ability. Moreover, the use of a straight channel capillary, a screen-printed carbon electrode without decoupler, in-channel amperometric detection and low sample volume requirements (2 μL) are shown as additional advantages.

Sensitive amperometric detection for capillary electrophoresis of phenol carbamates with in-line thermal hydrolysis strategy.

A strategy on amperometric detection for CZE of phenol carbamates as model analytes with a facile in-line thermal hydrolysis was presented, in which a thermal hydrolysis, subsequent CZE separation and final column-end amperometric detection were accomplished in an intact capillary. Key parameters of hydrolysis dynamics of carbamates and electrochemical detection of the hydrolysates were studied, as well as electrophoretic conditions. Under the optimal conditions, the capillary was utilized as chambers for insitu hydrolysis, CZE separation, and electrochemical detection. The successive separation of hydrolysates of five carbamates (propoxur, carbofuran, 3-OH-carbofuran, carbaryl and bendiocarb) were achieved within 17min. Applied to vegetable samples, the recoveries of carbamates fortified at 0.02 and 0.05 mg/kg were ranging in 88-107.2 and 86.3-107.3%, respectively. The success in the implementation of such a scheme resulted in a simple instrument as compared with those current analytical methods with post-column derivization or pre-column hydrolysis, or online enrichment in chip, respectively. This protocol might possess a potential utility for the sensitive amperometric detection of phenol carbamates.

Fabrication of a carbon nanotube-polyurethane composite electrode by in situ polyaddition for use in amperometric detection in capillary electrophoresis

The article describes the preparation of an electrode for amperometric detection in capillary electrophoresis (CE). It consists of a copper wire that was coated with a composite consisting of carbon nanotubes and polyurethane that was fabricated by in-situ polyaddition from a mixture of polyurethane prepolymer, curing agent, and carbon nanotubes (CNTs) inside a fused silica capillary. The structure of the composite was characterized by scanning electron microscopy, X-ray diffraction, thermogravimetry and FT-IR. The results indicated that CNTs were well dispersed and embedded throughout the composite to form an interconnected conducting network. The performance and advantages of the detection electrode are demonstrated by the separation and detection of standard mixtures of the hesperidin, hesperetin, naringenin and naringin by CE. The four flavanones are well separated within 12 min in a 40 cm long capillary at a separation voltage of 12 kV using a 50 mM sodium borate buffer (pH 9.2). The CNT-based electrode offers lower detection potential (0.8 V), enhanced detection limits (0.22–0.31 μM), lower costs of operation, high resistance to surface fouling, and improved stability. It shows long-term stability and repeatability, and relative standard deviations are <5 % for the peak current (for n = 15). The method was applied to the determination of flavanone in the peels of citrus fruits.

Online sample focusing technique in capillary electrophoresisamperometric detection for biogenic amines

Online sample focusing technique in capillary electrophoresisamperometric detection for biogenic amines

Graphene–epoxy composite electrode fabricated by in situ polycondensation for enhanced amperometric detection in capillary electrophoresis

This report describes the development and application of a novel graphene–epoxy composite electrode as a sensitive amperometric detector of capillary electrophoresis. The composite electrode was fabricated on the basis of the in situ polycondensation of a mixture of graphene and 1,2-ethanediamine-containing bisphenol A epoxy resin in the inner bore of a piece of fused silica capillary under heat. The structure of the material was investigated by scanning electron microscopy, thermogravimetric analysis, X-ray diffraction, and Fourier transform infrared spectroscopy. The results indicated that graphene sheets were well dispersed and interconnected throughout the composite to form an electrically conductive network. The performance of this unique electrode was demonstrated by separating and detecting two naturally occurring phenolic compounds in rosemary in combination with capillary electrophoresis. The graphene-based detector offered significantly lower operating potentials, higher sensitivity, satisfactory resistance to surface fouling, and lower expense of operation, indicating great promise for a wide range of applications.

Study on Capillary Electrophoresis–Amperometric Detection Profiles from Propolis and its Medicinal Preparations

Propolis is a resinous substance collected by honeybees from leaf buds and cracks in the bark of various plants, the pharmacologically active components in the propolis are flavonoids and phenolic acids and their esters. A simultaneous determination of flavonoids and phenolic acids from propolis and its medicinal preparations by capillary electrophoresis with electrochemical detection (CE-ED) was reported. Under the optimum conditions, the analytes could be separated in a 100 mmol/l borate buffer (pH=8.7) within 25 min. The response was linear over three orders of magnitude with detection limit (S/N=3) ranging from 1×10-7 g/ml to 5×10-7 g/ml for the analytes. This method has been used for the determination of flavonoids and phenolic acids from propolis and its medicinal preparations, and the assay result was satisfactory.

Fabrication of graphene/poly(ethyl 2-cyanoacrylate) composite electrode for amperometric detection in capillary electrophoresis

This report describes the development and application of a novel graphene/poly(ethyl 2-cyanoacrylate) composite electrode as a sensitive amperometric detector of capillary electrophoresis. The composite electrode was fabricated on the basis of the in situ polymerization of a mixture of graphene and ethyl 2-cyanoacrylate in the microchannel of a piece of fused silica capillary under heat. The structure of the material was investigated by scanning electron microscopy, X-ray diffraction, thermogravimetric analysis, and infrared spectroscopy. The results indicated that graphene sheets were well dispersed and interconnected throughout the composite to form a conductive network. The performance of this unique detector was demonstrated by separating and detecting five naturally occurring phenolic compounds in combination with capillary electrophoresis. The graphene-based detector offered significantly lower operating potentials, substantially enhanced signal-to-noise characteristics, and lower expense of operation, indicate great promise for microchip capillary electrophoresis, flowing injection analysis, and other microfluidic analysis systems.

Simultaneous determination of three β-blockers at a carbon nanofiber paste electrode by capillary electrophoresis coupled with amperometric detection

In the paper, a novel carbon nanofiber paste electrode (CFPE) was fabricated and firstly used as a sensitive amperometric detector in capillary electrophoresis (CE) for the simultaneous determination of three β-blockers: sotalol, alprenolol and atenolol. Compared with the bare carbon paste electrode, the CFPE exhibited enhanced oxidation peak current responses to the analytes due to its excellent electrocatalytic activities, high conductivity and large effective surface area. Subsequently, effects of several important factors such as detection potential, pH and concentration of running buffer, separation voltage and injection time on the analysis were investigated. Under the optimum conditions, the three analytes could be separated and detected in a phosphate buffer (pH 8.5) within 11min. The linear ranges were 0.1–100μM for sotalol, 0.2–150μM for alprenolol and 0.1–50μM for atenolol and the detection limits were as low as 10−8M magnitude (S/N=3). Moreover, the CFPE exhibited good repeatability and long-time stability. The proposed method was applied to determine the three β-blockers in spiked urine samples with satisfactory assay results. The good performance, low cost and straightforward preparation method of CFPE demonstrated that it could be used as a detector for CE–amperometric detection system for drug analysis.

An integrated PCR microfluidic chip incorporating aseptic electrochemical cell lysis and capillary electrophoresis amperometric DNA detection for rapid and quantitative genetic analysis

A fully integrated microchip for performing cell lysis, polymerase chain reaction (PCR) and quantitative analysis of DNA amplicons in a single step is described herein. The chip was built on glass substrate using an indium-tin-oxide (ITO) microheater and PDMS engraved microchannels, which integrated an electrochemical cell lysis zone, a continuous flow PCR module and capillary electrophoresis amperometric detection (CE-AD) system. The total length of the microchannel was 4625 mm for performing 25 cycles of flow-through PCR and was laid on a handheld form factor of 96 × 96 mm(2) area. The key to the fabrication of such a device lies in the use of a single medium to carry out different kinds of biochemical reactions and hence, a reagentless electrochemical cell lysis protocol was integrated on the microchip which was capable of lysing most cell types, including difficult to lyse gram positive bacteria. The lysate contained genomic DNA from a sample which was proven to be suitable for PCR reactions. Genetic analysis was successfully performed on the microchip with purified lambda phage genomic DNA and various cell types, including non-tumorigenic MCF-10A and tumorigenic MCF-7 human cell lines, gram negative bacteria Escherichia coli O157:H7, and gram positive bacteria Bacillus subtilis, at an optimized flow rate of 5 μl min(-1). For the detection of amplicon DNA, a CE-AD system was used, with semisolid alkaline agarose within the capillary microchannel to minimize interference from cell debris and for efficient resolution of DNA fragments. High signal to noise ratio during amperometric detection and the use of online FFT filtering protocol enhanced the limit of detection of DNA amplicons. Therefore, with a combination of portability, cost-effectiveness and performance, the proposed integrated PCR microchip can be used for one step genetic analysis of most of the cell types and will enable more accessible healthcare.

Determination of bioactive constituents in Flos Sophorae Immaturus and Cortex Fraxini by capillary electrophoresis in combination with far infrared-assisted solvent extraction

A method based on far infrared-assisted solvent extraction (FIASE) and capillary electrophoresis–amperometric detection (CE–AD) has developed for the rapid determination of rutin and quercetin in Flos Sophorae Immaturus and esculin and esculetin in Cortex Fraxini. The effects of the irradiation time and the voltage applied on the infrared generator were investigated to acquire the optimum extraction conditions. It was demonstrated that far IR radiation substantially enhanced the extraction efficiency and the extraction time was significantly reduced from 3h for conventional hot solvent extraction to 6min for FIASE. The obtained extracts were subsequently analysed by CE–AD. The detection electrode was a 300-μm-diameter carbon disc electrode at a detection potential of +0.90V (vs. saturated calomel electrode). The relation between peak current and analyte concentration was linear over about three orders of magnitude. The proposed method has been applied to determine the bioactive constituents in real samples.

Determination of Phenolic Disinfectants in Consumer Products by Capillary Electrophoresis with Amperometric Detection

Numerous disinfection products are widely used in daily life to kill pathogenic microorganisms. However, most disinfectants are organic compounds that might be hazardous to the environment and humans when used excessively. Phenolic disinfectants in disinfection products are investigated using a high-performance capillary electrophoresis-amperometric detection method. Under the optimum conditions, five commonly used disinfectants can be well-separated within 19 min at the separation voltage of 18 kV in a 80 mmol/L borax running buffer (pH 9.2), and adequate extraction was obtained with ethanol for the determination of the five compounds. Satisfactory recovery (93.5-106.0%), intra-day repeatability of the peak current (< 2.9%), and detection limits (1.6 x 10(-7) - 3.8 x 10(-8) g/mL) for the method are achieved. This proposed procedure is successfully used to analyze different samples of disinfection products.

Highly sensitive transient isotachophoresis sample stacking coupling with capillary electrophoresis-amperometric detection for analysis of doping substances

A simple and effective method of capillary electrophoresis-amperometric detection (CE-AD) coupled with transient isotachophoresis (tITP) was developed for the trace determination of doping substances. Compared with the conventional capillary electrophoresis method, the maximum enhancement factor in terms of peak heights was up to 5500-fold when the tITP technique was adopted. Under the optimum conditions, the detection limit (S/N=3) for methylephedrine (MDP), celiprolol (CEL), sotalol (SOT) and indapamide (IDP) were 4.2 x 10(-14), 6.3 x 10(-13), 5.8 x 10(-14) and 9.5 x 10(-13)molL(-1), respectively. The RSDs of four analytes were 1.0-2.3% for migration time and 2.6-3.8% for peak current, respectively. The proposed method was successfully applied to determine the contents of SOT and IDP in real urine sample, and the excretion curve of IDP within 48h was also investigated. The recoveries of the four doping in urine ranged from 90.0 to 102%.

A capillary electrophoresis microchip for amperometric detection of DNA

Capillary electrophoresis amperometric detection (CE–AD) microchip was fabricated for separation and detection of DNA fragments. The chip was fabricated on glass substrate and microchannels were laid in PDMS mold. The capillary was filled with 5% polyacrylamide gel and separation of DNA fragments of different molecular weight was achieved by application of 100 V DC potential across sample and waste reservoirs made at the capillary ends. The amperometric detection (AD) system involved in-channel gold microelectrodes to analyze oxidation peak for adenine residues in DNA chain. This technique was used in resolving single as well as double stranded DNA fragments.

Electrostacking online sample pre‐concentration capillary electrophoresis with amperometric detection for β2‐agonists in human urine

A simple, rapid, sensitive and specific method using capillary electrophoresis (CE) coupled with electrostacking and amperometric detection (AD) has been developed for the simultaneous separation and determination of clenbuterol (CLB), terbutaline (TER), salbutamol (SAL) and formoterol (FMT). In this paper, the CE separation and AD conditions were investigated in detail. The optimum conditions were: pH 8.6 Na(2)B(4)O(7)-H(3)BO(3) buffer solution (20.0 mmol/L), 9 kV for the separation voltage, and 1000 mV (versus Ag/AgCl) for the detection potential. When the sample which was dissolved in 70% ACN-water mixture solution was injected 60 s by 15 kV electrokinetic injection, the best stacking effects was obtained. Under the optimum conditions, the enhancement factors of these beta(2)-agonists had been greatly improved more than 5500-fold compared with the conventional electrokinetic injection. And then an excellent linear response was obtained with LODs (S/N = 3) of 0.098, 0.024, 0.063 and 0.920 pmol/L for CLB, TER, SAL and FMT in urine, respectively. The precision was determined in both intra-day (n = 5) and inter-day (n = 15) assays, and the RSDs were not more than 2.1 and 3.4% for migration time and peak current, respectively. The proposed method has been applied to analyze human urine samples successfully.

Microwave-assisted extraction followed by capillary electrophoresis-amperometric detection for the determination of antioxidant constituents in Folium Eriobotryae

A method based on capillary electrophoresis-amperometric detection has been developed for the determination of catechin, rutin, kaempferol, gentistic acid, and quercetin in Folium Eriobotryae with the aid of microwave-assisted solvent extraction. The effects of the acidity and the concentration of the running buffer, separation voltage, injection time, detection potential, and irradiation time were investigated to acquire the optimum analysis conditions. The detection electrode was a 300-μm-diameter carbon disc electrode at a detection potential of +0.90 V. The five analytes could be well separated within 12 min in a 40-cm length fused-silica capillary at a separation voltage of 12 kV in a 50-mM borate buffer (pH 9.0). The relation between peak current and analyte concentration was linear over about three orders of magnitude with detection limits (S/N = 3) ranging from 2.1 to 4.5 μM for all the analytes.

An online field-amplification sample stacking method for the determination of diuretics in urine by capillary electrophoresis-amperometric detection

A simple and sensitive online field-amplification sample stacking (FASS) pre-enrichment method following by capillary electrophoresis with amperometric detection has been developed for the determination of diuretics, such as indapamide (IDP), hydrochlorothiazide (HCT) and bumetanide (BMTN) in urine. Under the optimum conditions, it was found that the low concentration buffer solution could be used as the diluents for simultaneous field-amplification injection of three diuretics after electrokinetically injecting a short water plug (15 kV, 3 s). Three analytes could be well separated within 10 min in an uncoated fused-silica capillary with H 3BO 3–Na 2B 4O 7 (BB) buffer solution (pH 8.98). The detection limits (S/N = 3) were 9.0 ng/mL for IDP, 20 ng/mL for HCT and 1.5 ng/mL for BMTN, respectively. The detection limits of three diuretics were much lower by FASS than that by conventional sample injection, of which the detection limits were 340, 890 and 330 ng/mL for IDP, HCT and BMTN, respectively. Especially, for bumetanide the detection limit was 220-time lower by FASS. The linear ranges of three diuretics were all over three orders of magnitude. The proposed method has been successfully applied to analyze the diuretics in human urine samples without off-column sample pre-concentration.

Application of a Square‐Wave Potential Program for Time‐Dependent Amperometric Detection in Capillary Electrophoresis

AbstractUse of a square‐wave potential program for time‐dependent amperometric detection of analyte zones in capillary electrophoresis (CE) is described. Electrochemical detection for CE requires that the separation field be isolated from that of the electrochemical detection. This is generally done by physically separating the CE separation field from that of the detection. By applying a time variant potential program to the detection electrode, the detector current has a time dependence that can be used to help isolate the electrochemical detection current from that of the separation. When using a 20 μm inner‐diameter capillary, we find that a square‐wave potential program decreases the RMS baseline current from 4.5×10−10 A, found with a constant potential amperometric detection, to 1.1×10−10 A when using a square‐wave potential program. With a 75 μm inner‐diameter capillary, the improvement is even more dramatic, from 2.3×10−9 A with amperometric detection to 2.06×10−10 A when using a 1 Hz square‐wave potential program. When not using the time‐dependent detection with the 75 μm capillary, the analyte zones were beneath the S/N for the system and not detected. With the square‐wave potential program and time‐dependent detection, however, the analyte zones for an electrokinetic injection of 200 μM solution of 2,3‐dihydroxybenzoic acid were observed with the 75 μm inner‐diameter capillary. The improvement in the ability to discriminate the analytical signal from the background found experimentally is consistent with modeling studies.

Study on capillary electrophoresis–amperometric detection profiles of different parts of Morus alba L.

A high-performance capillary electrophoresis with amperometric detection (CE–AD) method has been developed for the determination of the pharmacologically active ingredients in different parts of Morus alba L. after a relatively simple extraction procedure. This method was also used in the comparison of bioactive constituent difference in the five parts, based on their electropherograms or characteristic “CE–AD profiles”. The effects of several factors such as the acidity and concentration of running buffer, separation voltage, applied potential and injection time were investigated to find the optimum conditions. Method detection limits (S/N = 3) ranged from 1.5 × 10 −7 to 1.4 × 10 −8 g/mL for all 10 analytes, and the assay results were satisfactory.

Integrated capillary electrophoresis amperometric detection microchip with replaceable microdisk working electrode: II. Influence of channel cross-sectional area on the separation and detection of dopamine and catechol

The interference of separation high voltage with the electrochemical detection is a major challenge to the microchip capillary electrophoresis–electrochemical detection systems with end-channel detection mode. Using dopamine and catechol as model analytes, the influences of channel cross-sectional area and channel-to-electrode distance on the high-voltage interference, accordingly on the separation and detection performances of the microchip capillary electrophoresis–electrochemical detection system were investigated. With the increase of the channel cross-sectional area from 312 through 450–615 μm 2, the apparent half-wave potentials of hydrodynamic voltammetry for dopamine at the field strength of 288 V/cm shifted positively from 285 through 330–400 mV. By using a chip with the smallest channel cross-section (312 μm 2 with top width of 37.3 μm and depth of 8.9 μm) the residual high-voltage field in the detection cell was small, so that detection was conducted at a channel-to-electrode distance of 20 μm to achieve better performances of separation and detection.