Voltammetric/amperometric detection for flow-injection systems

A dual-electrode voltammetric/amperometric detector for flow-injection systems is described. The detector provides qualitative voltammetric information without the charging currents associated with scanning the potential. Selectivity is enhanced relative to direct voltammetric detection at a single electrode because only chemically reversible redox couples are detected. A preliminary evaluation with hexacyanoferrate(III) and ascorbic acid is presented.     

Determination of Organic Peroxides by High Performance Liquid Chroma-Tography with Electrochemical Detection

Abstract

The application of electrochemical detection to the high performance liquid chromatographic determination of organic peroxides has been studied. The use of a buffered mobile phase was found to be critical to the successful analysis of samples containing hydroperoxides. Using amperometric detection, mixtures of peroxide containing compounds were readily determined. The sensitivity of the amperometric detector was in the one nanogram range for both benzoyl peroxide and cumene hydroperoxide. Polar-ographic detection was found to be a highly reproducible method for the analysis of samples containing peroxides as components of mixtures in the range of 5-2000 ng. The peroxide containing compounds determined in this manner were t-butyl hydroperoxide, cumene hydroperoxide, and 13-hydroperoxy-9(Z)-11(E)-octadecadien-oic acid. The polarographic detection system was used to obtain observed half-wave potentials for the peroxides under different chromatographic conditions. These observations correlated closely with literature results on the polarography of these compounds.     

毛细管电泳扫描伏安电化学法分离检测肾上腺素、氯丙嗪和异丙嗪

采用自制的高效毛细管电泳扫描伏安电化学检测装置,在磷酸盐缓冲溶液中进行了肾上腺素、异丙嗪和氯丙嗪的分离检测,在优化的条件下获得了较好的检测重现性和低的检出限。结果表明,采用扫描伏安电化学方法不但能够减少电极的污染,而且可以在电泳过程中获得被测物质的伏安特性及检测体系的动态伏安图,更有利于被测物质的识别。     

Investigation and comparison of the electrochemical behavior of some organic and biological molecules at various conducting polymer electrodes

Electrodes modified by the electrodepozition of conducting organic polymers such as poly(3-methylthiophene)(PMT), polypyrrole (PPY) and polyaniline (PAN) were used as chemical sensors for voltammetric analysis and flow injection detection of some organic and biological molecules. The electrochemical behaviors of catechol, ascorbic acid, hydroquinone, dopamine, epinephrine, acetaminophen and p-aminophenol were examined by differential pulse voltammetry. The electrochemical behavior of these molecules at different electrodes was compared and the effects on behavior of electrolyte type and its pH and the film thickness were systematically examined. The results showed that the proposed modified surface catalyzes the oxidation of these compounds. Electrocatalytic 'efficiency' decreases in order of poly-3-methylthiophene, polypyrrole and polyaniline. Voltammetric peak positions were affected by the nature of the electrolyte and its pH. Also, the effect of increasing film thickness was to observe increased peak heights. Polymer coated electrodes were also used in an amperometric detector for flow injection analysis of most of the these compounds. The responses of the polymer electrode were 5-15 times larger as compared with those of bare platinum. PMT showed improved performance as an amperometric detector for flow injection analysis systems over other types of polymer electrodes. Detection limits as low as 10(-8)-10(-9) M were achieved using the PMT, compared with 10(-6)-10(-8) M using platinum electrodes. In the flow injection analysis, with increasing molecular weight of analyte molecules was to observe decreased peak heights.     

Comparative study of coulometric and amperometric detection for the determination of macrolides in human urine using high-performance liquid chromatography

A high-performance liquid chromatography (HPLC) method using chromatographic conditions optimised in a previous work was applied for the separation of three macrolide antibiotics roxithromycin (Rox), oleandomycin (Ole) and rosamicin (Ros) and further determination of two of them, roxithromycin (Rox) and oleandomycin (Ole), in human urine samples. A comparative study of the behaviour of these macrolides under the two types of electrochemical detection (EC) widely coupled with HPLC, that is coulometric (EC-C) and amperometric (EC-A), was carried out by applying the same multiresidue method. From the assays performed using both detectors the comparison was made taking relevant criteria such as detection limits, linearity, recovery and precision values into account. As a result of this comparison, the coulometric detector appears slightly more suitable than the amperometric one for macrolide analysis.     

Non-invasive electrochemical hand-held biosensor as diagnostic indicator of dental diseases

A new hand-held amperometric biosensor for assay of salivary peroxidase in human secretions is described. The biosensor comprises a flow through amperometric detector coupled with a micropipette, and electronics (an amplifier, peak detector, microprocessor, and display). A micropipette, amplifier, peak detector, and microprocessor have been built in-house. All operations, such as sample injection, electrochemical detection and data processing are accomplished using only one action, by pressing the plunger of the micropipette. Because the use of a peristaltic pump is completely eliminated, the consumption of both samples and reagents is significantly reduced. The biosensor has been tested as a diagnostic indicator of dental diseases. It is shown that there is a functional relationship between the total level of salivary peroxidase and the clinical status of the periodontal disease. By using commercial available kits with peroxidase-labeled conjugates, the biosensor has an excellent potential for detection of different hormones, drugs, viruses, antibodies, and bacteria.     

Determination of hydralazine and metabolites in urine by liquid chromatography with electrochemical detection

The cyclic voltammetric behavior of hydralazine and its primary metabolites, the pyruvate and acetone hydrazones, was examined in the positive potential range at both conventional and electrochemically pretreated glassy carbon electrodes. The enhanced oxidations observed at the treated surface were used as the basis of amperometric electrochemical detection of the compounds following reversed-phase liquid chromatography. The detection limits so obtained at +0.75 V vs. Ag/AgCl (1, 3, and 5 ng injected, respectively) were comparable to those previously reported for absorption and fluorescence detection approaches employing derivatization/preconcentration procedures. For liquid chromatography with electrochemical detection, however, direct quantitation of all three species in urine samples was readily accomplished without any chemical derivatization or sample treatment operations other than particulate filtration.     

Electroanalytical voltammetry in flowing solutions

A review is given of the principles, key developments and representative applications of small electrodes in flow-through electrochemical (ec) detectors having very low effective dead volume (<10 μl) for voltammetric and amperometric detection in flowing solutions. Emphasis is placed on factors contributing to high sensitivity, reliability and selectivity of ec detection as an integral part of larger analytical systems utilizing continuously flowing, unsegmented streams, e.g., flow-injection and liquid chromatographic analyses. Solid and mercury electrodes are considered under potentiostatic and potentiodynamic control. A review is given also of auxiliary chemical and photolytic derivatizations coupled to ec detection. The majority of the literature on the subject relates to liquid chromatography with electrochemical detection (l.c./ec); however, applications to these concepts to specific examples in flow-injection systems, as well as for on-line process control, should be obvious. Details of chromatographic separations and design of total analytical systems are not reviewed.     

Determination of the catecholamines and serotonin, their precursors tyrosine and tryptophan, and their main metabolites in rat brain using reversed-phase high-performance liquid chromatography with fluorimetric and oxidative amperometric detection in series.

A high-performance liquid chromatographic method for the determination of catecholamines and serotonin, their precursors and their main metabolites was developed applied to rat cerebellum, hypothalamus, striatum and cortex. A fluorimetric and an oxidative amperometric detector were used in series. For both detectors, detection limits (25-520 pg) were useful for this application, linearity of standards was excellent (average r greater than 0.9997), between-run precision for sample analytes was generally acceptable (coefficient of variation less than 10% with appreciable concentrations present) and average recoveries of standard additions to sample analytes were better than 90%. Particular attention was paid to peak identification, including both a thorough treatment of retention time agreement of peaks in standards and sample analytes, and a comparison of results for the seven compounds amenable to quantitation by both detectors. Considerable attention was also given to determining the stability of standards and sample analytes under a wide variety of conditions, and practical recommendations were made.     

Characterization of etched electrochemical detection for electrophoresis in micron inner diameter capillaries

An enhanced etched electrochemical (EC) detection technique has been developed for CE in micron inner diameter capillaries. The design improvements allow for better alignment between the capillary bore and the electrode. This new method involves utilizing a carbon fiber microelectrode and etching both the carbon fiber and the detection end of a micrometer-sized inner diameter capillary to limit dead volume and analyte diffusion at the amperometric EC detector. To understand the factors affecting enhanced detector efficiency, a detailed examination of the relationship between detector design and performance has been completed by exploring the effects of varying electrode diameter, tip shape, and size, in addition to the etch length of the capillary outlet. The enhanced detection provides peak efficiencies as high as 75000 theoretical plates and estimated detection limits as low as 40 nM for dopamine. This etched detection method should further facilitate volume-limited sample analysis by CE.     

Determination of fat-soluble vitamins in yogurt by HPLC with electrochemical detection

A method employing HPLC with electrochemical detection for the rapid and simultaneous determination of vitamins A, D(3) and E is described. The method uses a C-18 reverse phase column and 2.5 mM HAcO-NaAcO in methanol-water (99:1, v/v) solution as the mobile phase. The compounds are quantified using amperometric detection with a glassy carbon electrode at a potential of + 1300 mV (vs. Ag/AgCl) and the results are compared with those obtained using UV detection at a wavelength of 280 nm. The method was successfully applied to the analysis of vitamins A, D(3) and E in yogurt samples. After saponification, fat-soluble vitamins were extracted and the methanolic solution of the extracts was injected directly into the chromatographic system, avoiding the clean-up step which is necessary when no electrochemical detection is used. Good recovery percentages were obtained.     

Comparison of pulsed amperometric detection and integrated voltammetric detection for inorganic sulfur compounds in liquid chromatography

A Variety of potential–time waveforms are useful in pulsed electrochemical detection (PED) when applied for the amperometric detection of numerous polar organic compounds following their separation by liquid chromatography (LC). Here, we compare the waveforms for pulsed amperometric detection (PAD) and integrated voltammetric detection (IVD) applied for detection of organosulfur compounds at Au electrodes in acidic media. In PAD waveforms, electrodes response is measured at a constant detection potentials. In IVD waveforms, electrodes current is integrated throughout a fast cyclic scan of the detection potential. As a consequence of this difference in detection strategy, the background signal for IVD is significantly smaller for PAD in the detection of organosulfur compounds whose response mechanisms require the concomitant formation of surface oxides on Au electrodes. Furthermore, in comparison to Pad, IVD has a larger sensitivity and a diminished system peak from 0₂ dissolved in the sample. Use of a preadsorption step increases detection sensitivity in both PAD and IVD. The limit of detection (S/N=3)for cysteine in LC-IVD is ca. 6 nM for a 50-μl injection (i.e., 300 fmol) using a detection waveform that includes a 1000-ms preadsorption period.     

Simultaneous determination of allopurinol and oxypurinol by liquid chromatography using immobilized xanthine oxidase with electrochemical detection

A novel liquid chromatographic method using an immobilized xanthine oxidase reactor and an electrochemical detector was developed for the simultaneous determination of allopurinol and oxypurinol in rat plasma, intestinal wash and bile. Xanthine oxidase was immobilized on 5-microns aldehyde silica (prepacked into a 2 mm x 10 mm cartridge) in a simple procedure. Allopurinol eluted from an analytical column was converted to oxypurinol in the enzyme reactor with the eluent as the reaction medium and detected with high selectivity using an amperometric detector with a glassy carbon electrode at the applied potential of +0.85 V. High specificity of the enzymatic reaction combined with selectivity of the electrochemical detection eliminated the need for an extensive sample preparation. The assay was linear in the range 15-500 ng/ml of rat plasma, intestinal wash and bile with a low limit of detection of 10 pg on-column (signal-to-noise ratio = 4) for both allopurinol and oxypurinol.     

Temperature dependence of electrochemical detection for liquid chromatography

The temperature dependence of amperometric response to compounds commonly assayed by liquid chromatography with electrochemical detection is reported. Temperature dependence ranges from 1.5 to 9% change in response per degree. Both diffusion and heterogeneous electron transfer contribute to temperature-induced variability. Controlling the temperature of the chromatographic column markedly reduces the effect of ambient temperature on the electrochemical detector. The efficiency of using different internal standards to compensate for temperature effects is shown to depend on the electrochemical similarity of the standard and the analyte. Several additional approaches to minimization of temperature dependence are discussed. Alteration of detector temperature for the enhancement of signal-to-noise ratio has little advantage.     

Rapid-scanning electrochemical detector with micro working electrode for micro high-performance liquid chromatography

Summary A voltammetric system controlled by a personal computer was developed using a PC-8801 mk II computer (NEC, Tokyo, Japan) for rapid-scanning voltammetry. The system was combined with an ultra-small-volume electrolytic cell with a single-carbon-fiber (7-μm diameter and 3-mm length) working electrode and used as a new detector for micro high-performance liquid chromatography. The system's capability for the detection of catecholamines and their related compounds was investigated. Three-dimensional voltamo-chromatograms were obtained by adding the potential axis of the electrode to the conventional current and time axes, leading to a dramatic improvement in the identification of each chroamtographic peak.     

Capillary electrophoresis of some tetracycline antibiotics coupled with reductive fast cyclic voltammetric detection.

The separation and quantitative performance parameters for tetracycline, chlortetracycline and oxytetracycline antibiotics were investigated by capillary zone electrophoresis coupled with fast cyclic voltammetric detection. Optimization of pH and complexation with a boric acid-sodium tetraborate buffer provided good resolution of all compounds. Detection by electrochemical reduction using fast on-line cyclic voltammetric detection with a Hg-film-microm electrode gave detection limits (2 x peak-to-peak baseline noise) of 7 x 10(-7) mol/l for tetracycline and chlortetracycline, and 1.5 x 10(-6) mol/l for oxytetracycline. The influence of electrode material, potential range and scan rate was examined and discussed. Optimal electrochemical detection was obtained at a Hg-film electrode with a waveform that consisted of an initial constant potential of -0.6 V for 200 ms followed by a cyclic voltammetry (CV) scan at 300 V/s from - 0.6 V to a vertex potential of 1.7 V. The analytical signal was obtained by plotting the integrated values of the CV current from each applied waveform as a function of time. The calibration plot (peak areas) for each separated peak was found to be linear over three-orders of magnitude.     

Hand-held amperometric sensor for saliva and other oral fluid-based diagnostics

A new hand-held amperometric sensor for monitoring of saliva and other oral fluid-based components is described. The amperometric sensor includes a flow through amperometric detector coupled with a micropipette, and an electronic block which consists of an amplifier, peak detector, microprocessor and display. All operations, such as sample injection, electrochemical detection and data processing are accomplished using only one action, by pressing the plunger of the micropipette. The instrument has been applied for assay of salivary peroxidase, human luteinizing hormone and human chorionic gonadotropin. The low detection limit of peroxidase is 0.5 ng/ml (defined as the concentration which gives a signal with S/N>2). The response time of the sensor is 1-3 s. The linear dynamic range of the hormones is from 1.0 to 80 mUI/ml. The life time of the hand-held amperometric device without regeneration of the electrode surface is 12 months. The sensor has the potential as a tool for noninvasive physiological studies and diagnostic strategies.     

Pulsed discharge emission detector — Application to analytical spectroscopy of permanent gases

Summary This publication contains our initial development of a pulsed discharge emission detector (PDED) (patent pending). It uses a pulsed high voltage discharge in helium, which provides a stable source for atomic and polyatomic emission spectroscopy. We have evaluated this detector for both quantitative and qualitative analysis of a range of chemical compounds. Emission spectra observed from the pulsed high voltage discharge are valuable for spectral analysis. The results obtained for selected permanent gases in the ultraviolet, visible and infrared regions of the optical spectrum indicate that these spectra can be used for compound identification after a chromatographic separation. The data are unique in our opinion and serve as a basis for the future development and investigation of the analytical significance of this detection method.Dedicated to Professor Leslie S. Ettre on the occasion of his 70th birthday.     

Swept-potential oxidative detection in flow streams

A swept-potential electrochemical detector, operated in the oxidative staircase voltammetric metric mode, is demonstrated for the high-performance liquid chromatography of a mixture of catecholamines. Voltammetric limits of detection are approximately 30 pg or 1 nM and chromatographic limits of detection are approximately 250 pg. The use of a platinum working electrode in a wall-jet cell configuration, with potential pulses for cleaning and activation between each sweep, results in a cell that has maintained a constant response for over a year without mechanical refinishing of the electrode surface.     

Determination of aminoglycoside antibiotics by reversed-phase ion-pair high-performance liquid chromatography coupled with pulsed amperometry and ion spray mass spectrometry.

This work constitutes a preliminary investigation of a high-performance liquid chromatographic (HPLC)-mass spectrometric (MS) method for confirming aminoglycoside residues in bovine tissues. A reversed-phase ion-pair HPLC method for the separation of four aminoglycosides was developed using volatile ion-pairing agents and optimized for detection with an ion spray HPLC-MS interface. The method is also compatible with a commercial pulsed amperometric detector that was used for HPLC method development and that may be useful for the screening and quantification phases of a regulatory method. Several column phases, eluent compositions, and pairing ions were evaluated for optimum HPLC-MS sensitivity. Detection limits are in the low nanogram range with the pulsed amperometric detector and with HPLC-MS in the selected ion monitoring mode. Results with bovine kidney, fortified to 20 ppm and extracted by matrix solid-phase dispersion, obtained using both detectors are presented.