Coulometric Cell Research Articles

Analysis of Food by High Performance Liquid Chromatography Coupled with Coulometric Detection and Related Techniques: A Review.

The development of single coulometric cells in combination with high performance liquid chromatography to dual cells and to the coulometric electrode array detector is described. An overview is given about the application of these methods in food chemistry. Easily oxidizable compounds, such as phenolic substances, pesticides, or vitamins, can be determined, as well as substances with high oxidation potentials or electroinactive compounds. Substances exhibiting poor electrochemical activity can be transformed to electroactive compounds by precolumn derivatization, postcolumn photochemical reactions, postcolumn enzyme reactors, or by using the oxidative/reductive mode for coulometric electrode array detection. Furthermore, it is shown that the interesting combination of high performance liquid chromatography with electrochemistry and mass spectrometry has opened further possibilities with respect to interpretation of redox reactions, drug metabolism studies, metabolomics, and electrochemical derivatization.

HPLC electrochemical detection and quantification of monoamines and their metabolites in rat brain tissue samples.

The monitoring of monoamines and their metabolites in CNS samples can be very valuable in pharmaceutical and biomedical research. A specific high performance liquid chromatography, coupled to a coulometric electrochemical detection method, for the assay of monoamines (dopamine, norepinephrine, epinephrine and serotonin) and their metabolites in rat brain tissue samples was developed. The chromatographic separation was achieved on a C8 reversed phase column with a mobile phase consisting of 0.1 M sodium formate buffer, 5 mM sodium 1-heptanesulfonate, 0.17 mM ethylenediaminetetraacetic acid disodium salt and 5% v/v acetonitrile (pH ±4.0). The detection was achieved through electrochemical detection, with a coulometric cell potential setting of +650 mV. The flow-rate was at 1 ml/min and the total run time was 50 min. The method was validated according to validation guidelines. The method was found to be linear (R2 > 0.99) over the analytical range (5 to 200 ng/ml) for all monoamines and their metabolites. All the other validation parameters were acceptable and within range. The method was applied to three rat brain areas (pre-frontal cortex, hippocampus and striatum), where the monoamines (except for epinephrine) and their metabolites were easily detected.

Indirect determination of zinc by thiol complexation and iodine coulometric titration with photocell detection

We have extended the capabilities of iodine coulometry for the determination of ionic zinc through its complexation with cysteine and subsequent titration of the thiol. The titration endpoint times of cysteine with zinc are proportionally longer as compared to cysteine itself. A homebuilt constant-current coulometer is employed for this method development. The instrument is composed of a constant-current source connected to nonisolated platinum electrodes contained in a 150-mL beaker (the coulometric cell) that sits on a photocell mounted in a styrofoam lid on top of a magnetic stirrer. Photometric endpoint detection is facilitated by connecting the photocell detector to an operational amplifier circuit; its output, recorded on a multifunctional chemical analysis system, resembled a sigmoidal curve. The titration conditions are optimized in terms of pH, temperature, stirring rate, and cysteine-to-zinc concentration ratios. Calibration curves generated as time versus increasing zinc concentrations in the presence of a constant cysteine concentration are used to determine zinc content in dietary supplements. The limit of detection is 5.5×10−5M Zn and the limit of quantification is 6.2×10−5M Zn. Percent relative standard deviations are at 4%. The recoveries of zinc from liquid zinc, enhanced zinc lozenges, and zinc tablets generally range from 80 to 120%. Titration of ascorbic acid and then zinc in the same solution is successfully performed with recoveries greater than 95% for both analytes. A comparison of the coulometry Zn data to atomic absorption results is also made for most samples.

Identification and Quantitation of Some Characteristic Phenolic Compounds in Elderberry Juice by HPLC with Coulometric Electrode Array Detection

An HPLC-coulometric electrode array detection (CEAD) based method for the determination of protocatechuic acid, chlorogenic acid, rutin (quercetin-3-O-rutinoside), nicotiflorin (kaempferol-3-Orutinoside) and quercetin in elderberry juice was developed. Sample work up for the determination of phenolic acids included solid phase extraction (SPE). For the analysis of flavonoids, simple dilution and centrifugation steps were sufficient. The compounds were separated isocratically on the same reversed phase column using two different mobile phases to prevent long retention times and tailing of the aglycone peaks. The analytes were then detected at twelve coulometric cells. The current/voltage curves contributed to compound identification and to verification of the peak purity of the separated compounds. Additionally, LC-ESI-MS in the negative ion scan mode was applied for qualitative analysis. Quantification of the compounds was carried out by standard addition method. The content in elderberry juice varied between 785 mg/l for rutin and 2.8 mg/l for quercetin. The presented rapid and reliable method can be used to differentiate between elderberry juices of different origins. Coulometric electrode array detection was applied for the first time for the determination of these compounds in elderberry juice. Keywords: Phenolic compounds, identification, quantitation, HPLC-CEAD, HPLC-ESI-MS, elderberry juice.

Electrolyte related parameters of coulometric solid state devices

Coulometric solid electrolyte sensors and measuring systems are gaining in importance for a variety of applications. One of their main advantages is their relatively precise function based on Faraday's law enabling calibration free performance over long time spans. By using devices with yttria-stabilized zirconia (YSZ) as solid electrolyte for the measurement of very small amounts of analytes like oxygen or gases reacting with oxygen ions, deviations of the measured cell current from the Faraday's law become visible. In this work, these deviations were measured at tubular YSZ cells with 6mm outer diameter, 1mm electrolyte thickness and 60mm length with the Hebb–Wagner method, using ultra clean nitrogen at different temperatures and oxygen partial pressures. At constant potentials of the measuring electrode between −850 and −350mV vs. Pt/air reference the deviation of the cell current from the Faraday's law was found to be caused completely by the hole conductivity of YSZ within the possible precision of measurement. At constant potentials below −1400mV the deviation is significantly lower than predicted by the values for electron conductivity already published for YSZ by Park and Blumenthal in 1989. Beside these static deviations at constant electrode potentials the coulometric cells show a dynamic non-Faraday charge transfer after potential steps of −50mV. This charge transfer, depending on temperature, start potential and prehistory, ranges between 0.45 and 310mAs. It is caused, inter alia, by oxygen release from the electrolyte and the porous environment of the electrode. Since the values of this charge transfer outnumber the targeted precision of coulometric measurements of traces gases by several orders of magnitude these measurements have to be carried out at constant electrode potentials.

Electrochemistry-coupled to mass spectrometry in simulation of metabolic oxidation of methimazole: Identification and characterization of metabolites

Abstract Methimazole (MMI), an antithyroid drug, is associated with idiosyncratic toxicity. Reactive metabolites resulting from bioactivation of the drug have been implicated in these adverse drug reactions. Mimicry of enzymatic oxidation of MMI was carried out by electrochemically oxidizing MMI using a coulometric flow-through cell equipped with a porous graphite working electrode. The cell was coupled on-line to electrospray ionization mass spectrometry (EC/ESI-MS). ESI spectra were acquired in both negative and positive modes. In acidic medium, ESI spectral analysis showed that the dimer was the main product, while in neutral and basic media, methimazole sulfenic acid, methimazole sulfinic acid and methimazole sulfonic acid were observed as the major electrochemical oxidation products. Oxidation of MMI and subsequent trapping with nucleophiles resulted in formation of adducts with N-acetylcysteine. Some of the electrochemically generated species observed in these experiments were similar to metabolites that have been observed from in vitro and in vivo studies. Trapping studies also showed that bioactivation of MMI proceeds predominantly through the S-oxide and not through formation of thiyl radicals. These results show that electrochemistry coupled to mass spectrometry can be used in mimicry of oxidative metabolism and subsequent high throughput screening of metabolites.

Unexpected benzimidazole ring formation from a quinoneimide species in the presence of ammonium acetate as supporting electrolyte used in the coupling of electrochemistry with mass spectrometry

Electrochemistry (EC) coupled to mass spectrometry (MS) has been used to study different phase-I reactions. Despite of the versatility of EC/MS, the effect of the nature of the supporting electrolyte on the formation of oxidation products has seldom been discussed during EC/MS experiments. Here, we present a comparison of two different supporting electrolytes and their effect on the identification of unstable intermediate oxidation species is discussed. The oxidation of acebutolol was performed with a coulometric cell in the presence of two supporting electrolytes namely ammonium acetate and lithium acetate. Ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UPLC/QTOFMS) using a binary gradient (water/acetonitrile) with positive electrospray ionization was used to identify the oxidation products in the presence and absence of glutathione. Chemical structure elucidations of the oxidation products were performed by high-resolution mass spectrometry (HRMS) and were also supported by nuclear magnetic resonance (NMR) measurements. From the electrochemical study and HRMS measurements, we demonstrate that the quinoneimide species resulting from the oxidative hydrolyses of acebutolol gives a benzimidazole ring product in the presence of ammonium acetate. Through the example of the oxidation of acebutolol, a correlation between the supporting electrolyte nature and oxidation product formation was established. The obtained results were supported by quantum mechanical calculations. We present here evidence of the side reactions induced by the presence of ammonia as supporting electrolyte during EC/MS measurements. Acebutolol was used as a model to postulate an uncommon and unexpected side reaction leading to benzimidazole ring formation. The findings may help to understand the identification of the intermediate species in the oxidative degradation process.

Oxygen nonstoichiometry and defect chemistry of the mixed conductor La0.9Ca0.1FeO3−δ at low oxygen partial pressure

Abstract The equilibrium oxygen nonstoichiometry, δ, for the mixed ionic and electronic conducting perovskite La 0.9 Ca 0.1 FeO 3 − δ (LCF91) was measured for oxygen partial pressures, p O2 , between 10 − 3 and 10 − 21 atm and temperatures of 750 to 900 °C using a tubular, two-probe Pt|YSZ|LCF coulometric titration cell. XRD experiments confirmed that LCF91 remains a single phase perovskite under the reducing conditions of a fuel electrode, and reveal no evidence of reaction with the zirconia electrolyte. The p O2 -nonstoichiometry relationship can be interpreted using a dilute-solution point-defect model involving oxygen vacancies, electrons, and electron holes.

Coulometric determination of dissolved hydrogen with a multielectrolytic modified carbon felt electrode-based sensor

A multielectrolytic modified carbon electrode (MEMCE) was fabricated by the electrolytic-oxidation/reduction processes. First, the functional groups containing nitrogen atoms such as amino group were introduced by the electrode oxidation of carbon felt electrode in an ammonium carbamate aqueous solution, and next, this electrode was electroreduced in sulfuric acid. The redox waves between hydrogen ion and hydrogen molecule at highly positive potential range appeared in the cyclic voltammogram obtained by MEMCE. A coulometric cell using MEMCE with a catalytic activity of electrooxidation of hydrogen molecule was constructed and was used for the measurement of dissolved hydrogen. The typical current vs. time curve was obtained by the repetitive measurement of the dissolved hydrogen. These curves indicated that the measurement of dissolved hydrogen was finished completely in a very short time (ca. 10 sec). A linear relationship was obtained between the electrical charge needed for the electrooxidation process of hydrogen molecule and dissolved hydrogen concentration. This indicates that the developed coulometric method can be used for the determination of the dissolved hydrogen concentration.

A solid electrolyte sensor for trace gas analysis

Coulometric solid electrolyte sensors based on yttria-stabilized zirconia were characterized with respect to the electronic conductivity of the solid electrolyte and to their electrochemical activity of oxygen, hydrogen and hydrocarbon conversion. Sensor parameters like the temperature dependency of the electronic conductivity as well as the elevated noise of the coulometric cell at the working temperature determine the limit of detection of coulometric titration. Here we describe investigations on these parameters and approaches to shift the lower detection limit into the ppb-range. Furthermore, it is shown that a coulometric solid electrolyte sensor, positioned behind a gas chromatographic separation unit, allows simultaneous, calibration-free and long-term stable detection of different oxidizable components and oxygen.

Detection Limits of Thin Layer Coulometry with Ionophore Based Ion-Selective Membranes

We report here on a significant improvement in lowering the low detection limit of thin layer coulometric sensors based on liquid ion-selective membranes, using a potassium-selective system as a model example. Various possible processes that may result in an elevated residual current reading after electrolysis were eliminated. Self-dissolution of AgCl on the Ag/AgCl inner element may result in a residual ion flux that could adversely affect the lower detection limit. It was here replaced with an Ag/AgI inner pseudoreference electrode where the self-dissolution equilibrium is largely suppressed. Possible residual currents originating from a direct contact between inner element and ion-selective membranes were eliminated by introducing an inert PVDF separator of 50 μm diameter that was coiled around the inner element by a custom-made instrument. Finally, the influence of electrolyte fluxes from the outer solution across the membrane into the sample was evaluated by altering its lipophilic nature and reducing its concentration. It was found that this last effect is most likely responsible for the observed residual current for the potassium-selective membranes studied here. For the optimized conditions, the calibration curves demonstrated a near zero intercept, thereby paving the way to the coulometric calibration-free sensing of ionic species. A linear calibration curve for the coulometric cell with valinomycin potassium-selective membrane was obtained in the range of 100 nM to 10 μM potassium in the presence of a 10 μM sodium background. In the presence of a higher (100 μM) concentration of sodium, a reliable detection of 1-100 μM of potassium was achieved.

Oxygen Nonstoichiometry and Defect Chemistry of Mixed Conductor La0.9Ca0.1FeO3- δ at Low PO2

The equilibrium oxygen nonstoichiometry, δ, for the perovskite La0.9Ca0.1FeO3-δ (LCF) was determined for oxygen partial pressures, PO2, between 10-3 and 10-20 atm and temperatures of 800, 850 and 900C using a tubular, two-probe Pt|YSZ|LCF coulometric titration cell. The PO2-nonstoichiometry relationship was interpreted using a dilute point-defect model, analogous to that of La1-xSrxFeO3-δ (LSF), and the resulting equilibrium constants were reported.

Application of coulometric electrolytic cells in absolute humidity hygrometers

We present data from studies of coulometric electrolytic cells with platinum, rhodium, and platinum–iridium electrodes. We describe the experimental apparatus, the procedure, and the results of a study of the dependence of the measurement error in analysis of inert gases, nitrogen, hydrogen, and oxygen at temperatures of 20 and 50°C. We give recommendations for use of coulometric electrolytic cells in hygrometers with measurement range 0–1000 ppm.

Quantitation of Parent Drug and Its Unstable Metabolites by in Situ Coulometric Oxidation and Liquid Chromatography−Tandem Mass Spectrometry

Recent FDA and ICH guidances on safety testing of drug metabolites have challenged the way we traditionally think about quantitative bioanalytical methods. Such assays, in general, require a reference standard for each analyte to construct calibration curves and prepare quality control samples. However, early in the drug development process, metabolite standards may not be readily available, and if they are inherently unstable, they are difficult to synthesize or purify. In this paper, we describe a novel in-line method for producing and then quantifying a very unstable metabolite which is based upon the in situ postcolumn coulometric oxidation of the parent drug. Lacking any metabolite standards, the feasibility of simultaneously quantifying a development drug (compound A) and its unstable hydroxylated metabolites (metabolite B) was investigated. Reference standards for these ostensibly major human metabolites could not be reliably obtained due to rapid degradation upon purification and/or subsequent storage. Following high-performance liquid chromatography (HPLC) separation, parent drug and its [(13)C(3)-(15)N] isotopically labeled internal standard were quantitatively converted to equal amounts of a diastereomeric pair of hydroxylated metabolites using a postcolumn coulometric electrochemical cell before reaching the mass spectrometer. The concentration of the injected parent (which is equal to the total concentration of the in-line generated metabolites since the conversion to metabolite is quantitative) and the tandem mass spectrometry (MS/MS) signals of the electrochemically generated metabolites were used to construct a calibration curve for quantifying both the parent drug and its hydroxylated metabolites. Plasma extracts from humans dosed with compound A contained chromatographically distinct liquid chromatography-mass spectrometry (LC-MS) signals (m/z 538) for in vivo formed hydroxylated metabolites and the electrochemically oxidized parent drug which had been converted in-line into its chemically identical twin. Both peaks in this study sample could be quantified using a single calibration curve obtained under the same coulometric conditions using known amounts of the parent drug. Although no attempt was made to fully validate a bioanalytical method, the practicality of this in situ quantification approach was further confirmed by the preliminary bioanalytical analysis of a selection of plasma samples collected following oral administration (50 mg) of compound A in a clinical study.

Background current elimination in thin layer ion-selective membrane coulometry

A promising method for the elimination of undesired capacitive currents in view of realizing a potentially calibration free coulometric ion detection system is presented. The coulometric cell is composed of a porous polypropylene tube doped with a liquid calcium-selective membrane and a silver/silver chloride wire as an inner electrode, forming a thin layer sample between wire and tubing. The total charge passed through the system during potential controlled electrolysis of the thin layer sample is indeed found to be proportional to the amount of calcium present, but non-Faradaic processes do contribute to the obtained signal. We introduce here a multi-pulse procedure that allows one to perform a second excitation pulse at the same excitation potential after exhaustive ion transfer voltammetry of calcium has taken place. The intercept of the calibration curve after background subtraction is found as 20.6 ± 0.6 μC, significantly lower than the value of 54.1 ± 0.8 μC for the uncorrected curve. Changes in sample temperature (from 23 °C to 38 °C) did equally not affect the background corrected coulometric readings, supporting that the procedure renders the readout principle more robust.

Online electrochemistry/mass spectrometry in drug metabolism studies: principles and applications

Importance of the field: Conventional metabolism studies in early stages of drug development include in vitro tests, often based on hepatic cells and cell extracts, as well as in vivo tests on the basis of animal models. Complementary to these existing techniques, the electrochemical study of oxidative metabolism reactions is gaining increasing attention. Electrochemistry (EC) allows the fast detection of sites labile towards oxidation in a drug molecule and can mimic the formation of potential oxidative metabolites.Areas covered in this review: The present review summarizes the developments of EC-based methods to study the oxidative metabolism of drugs in the past 10 years. The implementation of different coulometric and amperometric cells is described. Furthermore, online set-ups utilizing the hyphenation of EC, liquid chromatography and electrospray ionization mass spectrometry are discussed regarding their applicability in metabolism studies. Besides mass spectrometric detection, the isolation and advanced characterization of oxidation products are reviewed. This includes structure elucidation based on NMR spectroscopy as well as the evaluation of the reactivity of metabolites towards trapping agents or proteins.What the reader will gain: A major focus of the article is directed to the comparability between electrochemically predicted metabolites and those occurring in vitro and in vivo. This comprises the discussion of typical oxidative metabolism reactions and provides a guideline about how far EC is capable of generating a specific oxidative metabolite.Take home message: The study of drug metabolism reactions in an electrochemical cell enables the prediction as well as the synthesis of oxidative metabolites. In particular, reactive metabolites are directly detected using online EC/electrospray ionization mass spectrometry. Thus, the electrochemical technique is a promising tool that complements existing in vivo and in vitro techniques in drug metabolism studies.

Analysis of risperidone and its metabolite in plasma and saliva by LC with coulometric detection and a novel MEPS procedure

A new analytical method, based on the use of liquid chromatography with coulometric detection, has been developed and applied to quantify risperidone and its main active metabolite 9-hydroxyrisperidone in human plasma and saliva. The analytes were separated on a reversed phase C18 column, using a mobile phase composed of acetonitrile (26%) and a pH 6.5 phosphate buffer (74%). Pipamperone was used as the internal standard. A high sensitivity coulometric detection analytical cell containing two flow-through working electrodes was used: electrode 1 was set at +0.500V and electrode 2 at +0.700V. The detector response was linear over a plasma and saliva concentration range of 0.5-50.0ngmL(-1) for risperidone and 0.5-100.0ngmL(-1) for 9-hydroxyrisperidone. The limit of quantitation and the limit of detection for risperidone and 9-hydroxyrisperidone were 0.5ngmL(-1) and 0.17ngmL(-1), respectively. A novel clean-up procedure of biological samples was developed using the microextraction by packed sorbent technique, which gave good extraction yield for both the analytes, with absolute recovery values higher than 90.1%. The intra-day and the inter-day precision results, expressed by relative standard deviation values, were lower than 5.8%. Accuracy and selectivity assays were also satisfactory. The validated method has been successfully applied to the analysis of risperidone and 9-hydroxyrisperidone in plasma and saliva of psychiatric patients undergoing therapy with risperidone.

Estimate of incompleteness error of moisture absorption in coulometric hygrometers

One of the most important components of the error of flow-through coulomometric hygrometers is that due to “moisture transit,” or loss. Exact relationships between this error and the fraction of electrolytic current from the control part of a coulometric cell for both fresh (“new”) cells (i.e., those that been regenerated) and cells that are in the aging process are obtained. It is shown that it is possible to determine and take into account this error in the measurement process.

Influence of AgCl Precipitates on the Precipitation Titration of Sodium Chloride by Constant-Current Coulometry

The precipitation titration of sodium chloride with electrogenerated silver ion was studied. The production of a precipitate of silver chloride had a significant effect on the titration results because the precipitate involved unreacted chloride or unreacted silver ion. The accuracy of the method was investigated by changing the introduction time of a sodium chloride solution to the coulometric cell during the process of electrolysis, and examining the dependency on the sample size. The accuracy of the measurement of the precipitation titration is discussed.

Determination of Olanzapine in rat brain using liquid chromatography with coulometric detection and a rapid solid-phase extraction procedure

A sensitive and selective method was developed for the determination of the antipsychotic drug Olanzapine levels in rat brain tissue, based on HPLC with electrochemical detection. The analyses were carried out on a C8 reversed phase column (150 mm × 4.6 mm, 5 μm), using a mobile phase composed of methanol and a phosphate buffer (44.0 mM, pH 3.5), containing triethylamine (21:79, v/v), flowing at 1.2 mL min −1. A high sensitivity coulometric detection analytical cell containing two flow-through low volume working electrodes was used: electrode 1 was set at +0.350 V and electrode 2 at −0.200 V. Olanzapine, administered to rats in different doses or in different times, was extracted from tissue homogenate of either the whole brain or specific areas (cortex, hyppocampus, nucleus striatum) with a rapid solid phase extraction procedure (SPE) on Oasis HLB cartridges. The method provided a high extraction yield of Olanzapine and internal standard (2-methylolanzapine) from brain tissue homogenate with absolute recovery values higher than 90.0%. The detector response was linear over a concentration range of 0.2–100.0 ng mL −1 of Olanzapine. The limit of quantification (LOQ) was 0.2 ng mL −1. Precision results, expressed by the intra-day and the inter-day relative standard deviation values, were satisfactory, better than 4.6%. Accuracy was satisfactory as well. This method proved to be suitable for the analysis of Olanzapine in rat brain tissues and for the study of distribution and pharmacokinetics of Olanzapine in rat brain after a single treatment with the antipsychotic drug.