Determination of water-soluble vitamins by liquid chromatography with a parallel dual-electrode electrochemical detector

A method for the determination of water-soluble vitamins (ascorbic acid, pyridoxine hydrochloride, pyridoxal hydrochloride, pyridoxamine dihydrochloride, p-aminobenzoic acid, folic acid) by liquid chromatography, with a parallel dual-electrode electrochemical detector, is described. One electrode was controlled at +0.80 V (vs. SCE), the other at +1.20 V (vs. SCE). The possibility of interference by eight other water-soluble vitamins (riboflavin, nicotinamide, cyanocobalamin, menadione, dextro calcium pantothenate, thiamine, nicotinic acid, dextro biotin) was studied. These vitamins did not interfere when a parallel dual-electrode detector system was used. The estimation of five of the vitamins was studied in detail. The linear ranges found were 10 ng-1.2 mug for pyridoxine hydrochloride, 2 ng-2mug for pyridoxal hydrochloride, 10 ng-3 mug for pyridoxamine dihydrochloride, 5-200 ng for folic acid and 0.6-200 ng for p-aminobenzoic acid, the limits of detection being 3, 0.6, 1, 2 and 0.06 ng respectively. Application of the technique to the estimation of vitamin B(6) in tablets is illustrated. The results indicate that the vitamin B(6) in these tablets existed in the pyridoxine hydrochloride form and the B(6) content agreed well with liquid chromatograph by spectrophotometric analysis.     

Liquid chromatography of vitamin B6 with electrochemical detection using a carbon fibre electrode

A flow-through electrochemical detector with a carbon fibre electrode was fabricated. A chromatographic method for the determination of vitamin B₆ using this detector is described, and the results are compared with those obtained by detection with a thin-layer glassy carbon electrode or spectrophotometry. The detection limit of vitamin B₆ is 2.5 ng for pyridoxine, 1 ng for pyridoxal and 1 ng for pyridoxamine. The technique was applied to the determination of vitamin B₆ in tablets.     

Cathodic adsorption stripping analysis of vitamin B12

Amalgam formation preconcentration is usually performed in electrochemical stripping analysis, but adsorption also can be employed for preconcentration. Vitamin B₁₂ is adsorbed on a mercury electrode in the potential region between ca. ?0.3 and ?1.5 V vs. SCE. Vitamin B₁₂ also yields a polarographic catalytic wave at ca. ?1.6 V. It is considered that the adsorbed protonated reduced form of vitamin B₁₂ plays an important role in producing the catalytic current. Stripping analysis for vitamin B₁₂ employing adsorption preconcentration was found to be possible. Ammonium acetate was used as the supporting electrolyte. It was found that the peak for vitamin B₁₂ was maximal in 0.3 M ammonium acetate. The suitable preconcentration potential was ?1.45 V. The preconcentration time was 5 min while stirring the solution. The scan rate used was 50 mV/s, considering the pen speed of the X-Y recorder. The optimum temperature was 35°C. Under the optimum conditions described above, the calibration curve was linear in the concentration range up to 0.1 μ M. The detection limit was 2 n M.     

Electrochemical and Spectroelectrochemical Studies on Pyridoxine Hydrochloride Using a Poly(methylene blue) Modified Electrode

The electrochemical redox processes of pyridoxine hydrochloride (VB₆) at a poly(methylene blue) film modified glass carbon electrode (PMBE) in a phosphate buffer solution (PBS, pH 8.0) were studied by cyclic voltammetry. The VB₆ electrode reaction with quasi‐reversible characteristics was diffusion‐controlled at low scan rates and adsorption‐controlled at high scan rates. The anodic peak current positive to 0.6 V (vs. SCE) was found to be proportional to the concentration of VB₆ in the range of 0.010 to 1.03 mg?mL^?1 with a detection limit of 1.34 μg mL^?1. Fluorescence and UV‐vis absorption spectroelectrochemical measurements suggest that the pyridine ring was not destroyed over the potential range from ?0.8 to 1 V (vs. SCE), and the electrocatalytic generation of pyridoxal was anodically started at 0.57 V.     

Electrode Modified with Langmuir–Blodgett (LB) Film of Calixarenes for Preconcentration and Stripping Analysis of Hg(II)

A new type of current sensor, Langmuir–Blodgett (LB) film of calixarene on the surface of glassy carbon electrode (GCE) was prepared for determination of mercury by anodic stripping voltammetry (ASV). An anodic stripping peak was obtained at 0.15 V (vs. SCE) by scanning the potential from ?0.6 to +0.6 V. Compared with a bare GCE, the LB film coated electrode greatly improves the sensitivity of measuring mercury ion. The fabricated electrode in a 0.1 M H₂SO₄+0.01 M HCl solution shows a linear voltammetric response in the range of 0.07–40 μg L^?1 and detection limit of 0.04 μg L^?1 (ca. 2×10^?10 M). The high sensitivity, selectivity, and stability of this LB film modified electrode demonstrates its practical application for a simple, rapid and economical determination of Hg²⁺ in a water sample.     

Determination of vitamin B6 using high-performance liquid chromatography with electrochemical detection

A chromatographic method for the determination of vitamin B₆ utilizing a thin-layer amperometric detector with a glassy carbon electrode has been described. The redox behavior of vitamin B₆ has been studied by means of cyclic voltammetry. The limit of detection is 4 ng for pyridoxine and 1 ng for pyridoxamine. Application of the technique for detection of vitamin B₆ in drugs was illustrated.     

Boron‐Doped Diamond Dual‐Plate Deep‐Microtrench Device for Generator‐Collector Sulfide Sensing

A BDD‐BDD dual‐plate microtrench electrode with 6 μm inter‐electrode spacing is investigated using generator‐collector electrochemistry and shown to give microtrench depth‐dependent sulfide detection down to the μM levels. The effect of the microtrench depth is compared for a “shallow” 44 μm and a “deep” 180 μm microtrench and linked to the reduction of oxygen to hydrogen peroxide which interferes with sulfide redox cycling. With a deeper microtrench and a fixed collector potential at ?1.4 V vs. SCE, two distinct redox cycling potential domains are observed at 0.0 V vs. SCE (2‐electron) and at 1.1 V vs. SCE (6‐electron).     

Automated determination of molybdenum(VI) in seawater by means of constant-current reduction of the adsorbed 8-quinolinol complex in a computerized flow potentiometric stripping analyzer

Molybdenum(VI) in seawater is determined by means of potentiostatic adsorption of the 8-quinolinol complex onto a mercury film electrode at ?0.2 V vs. SCE and subsequent reduction of the complex by means of constant-current stripping in 5 M calcium chloride medium with a fully automated stripping analyzer. A single stripping peak at –0.42 V vs. SCE was obtained. The molybdenum(VI) concentration in reference seawater NASS-1, with a certified value of 11.5 ± 1.9 μg 1^?1, was found to be 8.9 ± 1.3 μg 1^?1 (n = 10).     

Light‐emitting‐diode‐induced chemiluminescence detection for capillary electrophoresis

In this work, an LED‐induced‐chemiluminescence (LED‐CL) system was developed to extend the application of CL detection in CE. In the LED‐CL, the analyte photooxidizes luminol under the irradiation of LEDs and generates CL. Taking the advantage of the small size nature of LEDs, the constructed photoreactor is greatly miniaturized, and especially suitable as a CE detector. The feasibility of the proposed detector was evaluated by detection of riboflavin (RF), flavin mononucleotide (FMN) and flavin adenine dinucleotide (FAD) after CE separation. Under the optimized conditions, the LODs for RF, FMN and FAD were 0.007, 0.02 and 0.1 μg/mL, respectively, better than those by UV detection. The RSDs were 3.4, 3.6 and 4.1% for 0.5 μg/mL RF, 2 μg/mL FMN and 5 μg/mL FAD, respectively. The LED‐CL detector features low cost, miniaturization, fast response, high sensitivity and good reproducibility.     

Determination of Riboflavin Using the Polarographic Reduction Wave of Its Photochemical Reaction Products

Abstract

A new, simple, and sensitive method for the determination of riboflavin based on the single-sweep polarographic reduction wave of the photolysis products of riboflavin has been developed. Under the irradiation of an iodine-tungsten lamp, the photolysis reaction of riboflavin in pH 9.3 NH₃-NH₄Cl buffer solution can be complete in about 20 min. The reaction products yield a single adsorptive reduction wave at ?0.63 V (vs. SCE) negative to the reduction wave of riboflavin by 0.18 V. The derivative wave height is linearly proportional to the concentration of riboflavin in the range of 0.01–17.5 µg/mL (correlation coefficient 0.996). The detection limit is 0.00 5µg/mL. The proposed method has been applied to the determination of multivitamin tablets with satisfactory results. In comparison with the methods directly using the reduction wave of riboflavin for the determination, this method eliminates the effect of the degradation of riboflavin on the determination accuracy and is much more sensitive. The new method could be useful in many fields.     

Electrochemical reduction and determination of cimetidine at nanomolar to micromolar levels of concentration

The electrochemical reduction of cimetidine, an H₂-antagonist of histamine used in the treatment of duodenal and gastric ulcers, has been examined by using a variety of electrochemical techniques. Reduction in 0.1. M HCl occurs at ?0.8 to ?1.1 V vs. SCE depending on the experimental conditions. Square-wave and cyclic voltammetry were found to be the best techniques for the determination of cimetidine whereas sampled-d.c., normal pulse, and differential pulse polarography were not useful. The detection limit for the determination depends on the condition employed but can be as low as 1 ng ml^?1 (4 nM). A linear calibration plot is obtained up to 2 μg ml^?1 and calibration curves can be used at higher concentrations. The high sensitivity is due to reactant adsorption. The electrode process appears to involve a 4-electron reduction of the cyano group in cimetidine. The major metabolites of cimetidine can also be determined by this method.     

Preconcentration and voltammetric determination of trace myoglobin at a 6-mercaptopurine modified silver electrode

6-Mercaptopurine(6-MP) is covalently modified onto a silver electrode to construct a chemically modified electrode (CME). It exhibits the capability of selectively complexing myoglobin and can be applied as a selective biosensor for this compound. Myoglobin is accumulated onto the CME at 0.32 V (vs. SCE); after exchanging the medium it is determined by differential pulse stripping voltammetry. A cathodic stripping peak is obtained at 0.15 V (vs. SCE) by scanning the potential from 0.35 to –0.1 V. The peak currents are linearly proportional to the concentration of myoglobin in the range of 0.2 ～ 4 μg/mL with a relative standard deviation of 7.8%. The detection limit is about 0.1 μg/mL. The mechanism of the complexation is also discussed.     

Manganese Oxide Nanoparticles/Reduced Graphene Oxide as Novel Electrochemical Platform for Immobilization of FAD and its Application as Highly Sensitive Persulfate Sensor

In this study, manganese oxide nanoparticles/reduced graphene oxide(MnOxNPs/rGO) was used as support for strong immobilization of flavin adenine dinucleotide(FAD). A thin film of rGO cast on the electrode surface, followed by performing electrodeposition of MnOxNPs at applied constant potential of +1.4 V vs. Ag/AgCl for 200 s. Finally, FAD was electrodeposited onto the rGO/MnOxNPs film by potential cycling between 1.0 to ?1.0 V in solution containing 1 mg ml^?1 FAD. Electrochemical properties and catalytic activity of GCE/rGO‐MnOxNPs/FAD toward persulfate (S₂O₈^2?) reduction was investigated. Under optimized condition, the concentration calibration range, detection limit, and sensitivity were 0.1 μM–2 mM, 90 nM and 125.8 nA/μM, respectively, using hydrodynamic amperometry technique.     

Adsorptive Stripping Voltammetric Detection of Single‐Stranded DNA at Electrochemically Modified Glassy Carbon Electrode

Electrochemically modified glassy carbon electrode (GCE) was used to study the electrochemical oxidation and detection of denatured single‐stranded (ss) DNA by means of adsorptive stripping voltammetry. The modification of GCE, by electrochemical oxidation at +1.75 V (vs.SCE) for 10 min and cyclic sweep between +0.3 V and ?1.3 V for 20 cycles in pH 5.0 phosphate buffer, results in 100‐fold improvement in sensitivity for ssDNA detection. We speculated that the modified GCE has a high affinity to single‐stranded DNA through hydrogen bond (specific static adsorption). Single‐stranded DNA can accumulate at the GCE surface at open circuit and produce a well‐defined oxidation peak corresponding to the guanine residues at about +0.80 V in pH 5.0 phosphate buffer, while the native DNA gives no signal under the same condition. The peak currents are proportional to the ssDNA concentration in the range of 0–18.0 μg mL^?1. The detection limit of denatured ssDNA is ca. 0.2 μg mL^?1 when the accumulation time is 8 min at open circuit. The accumulation mechanism of ssDNA on the modified GCE was discussed.     

The Convenient Determination of Palladium at a Solid Electrode via Adsorptive Stripping Voltammetry at a Glassy Carbon Electrode Modified with a Random Array of Mercury Nanodroplets

The detection of palladium using adsorptive stripping voltammetry reported by Wang et al. (J. Wang, K. Varughese Anal. Chim. Acta 1987, 199, 185 [3]) at a hanging mercury drop electrode is extended to a more convenient solid electrode. To this end a random array of 3.5×10⁸ mercury nanodroplets per cm² (65 nm average diameter) was electrodeposited on a glassy carbon substrate. Adsorptive stripping voltammetry was performed using 2×10^?4 M dimethylglyoxime as a chelating agent for the Pd(II) ion, with accumulation at ?0.20 V vs. SCE for 120 s and a linear detection range of 5–150 μM was determined with a limit of detection of 1.6 μM.     

A glassy-carbon electrode modified by dispersed alpha-alumina as an enhanced electrochemical detector in liquid chromatography for cysteine and glutathione

The dispersion of alumina particles on a glassy-carbon surface serving as a modified electrode significantly enhances the amperometric detection of cysteine and glutathione following liquid chromatography. With an applied potential of 0.8 V vs. SCE, the detection limits were 1.2 ng for cysteine and 8 ng for glutathione and the electrode response was linear up to 600 ng for cysteine and 1.8 mug for glutathione. The modified electrode displayed high sensitivity and stability and was easy and inexpensive to prepare.     

Tuning the redox potential of riboflavin by zirconium phosphate in carbon paste electrodes

Modified carbon paste electrodes were prepared by inclusion of riboflavin together with zirconium phosphate (ZrP) into carbon paste. The midpoint potential for riboflavin in this electrode was found to be -0.259 V vs. SCE and shifted by 0.207 V to the positive direction, as compared to carbon paste electrode not containing ZrP. The electrode prepared was shown to electrocatalyse the anodic oxidation of the coenzyme NADH in the potential range of 0.0 to 0.25 V.     

Ion‐pair cloud‐point extraction: A new method for the determination of water‐soluble vitamins in plasma and urine

A novel, simple, and effective ion‐pair cloud‐point extraction coupled with a gradient high‐performance liquid chromatography method was developed for determination of thiamine (vitamin B₁), niacinamide (vitamin B₃), pyridoxine (vitamin B₆), and riboflavin (vitamin B₂) in plasma and urine samples. The extraction and separation of vitamins were achieved based on an ion‐pair formation approach between these ionizable analytes and 1‐heptanesulfonic acid sodium salt as an ion‐pairing agent. Influential variables on the ion‐pair cloud‐point extraction efficiency, such as the ion‐pairing agent concentration, ionic strength, pH, volume of Triton X‐100, extraction temperature, and incubation time have been fully evaluated and optimized. Water‐soluble vitamins were successfully extracted by 1‐heptanesulfonic acid sodium salt (0.2% w/v) as ion‐pairing agent with Triton X‐100 (4% w/v) as surfactant phase at 50°C for 10 min. The calibration curves showed good linearity (r² > 0.9916) and precision in the concentration ranges of 1‐50 μg/mL for thiamine and niacinamide, 5–100 μg/mL for pyridoxine, and 0.5–20 μg/mL for riboflavin. The recoveries were in the range of 78.0–88.0% with relative standard deviations ranging from 6.2 to 8.2%.     

Ultrasensitive Determination of Vitamin B12 Using Disposable Graphite Screen‐Printed Electrodes and Anodic Adsorptive Voltammetry

A facile, rapid and ultra‐sensitive method for the determination of vitamin B₁₂ (cyanocobalamin) at the sub‐nanomolar concentration range by using low‐cost, disposable graphite screen‐printed electrodes is described. The method is based on the cathodic preconcentration of square planar vitamin B_12s, as occurred due to the electro reduction of Co(III) center in vitamin B_12a to Co(I), at ?1.3 V versus Ag/AgCl/3 M KCl for 40 s. Then, an anodic square wave scan was applied and the height of the peak appeared at ca. ?0.73 V versus Ag/AgCl/3 M KCl, due to the oxidation of Co(I) to Co(II) in the adsorbed molecule, was related to the concentration of the vitamin B₁₂ in the sample. EDTA was found to serve as a key‐component of the electrolyte by eliminating the background signal caused by metal cations impurities contained in the electrolyte (0.1 M phosphate buffer in 0.1 M KCl, pH 3). It also blocks trace metals contained in real samples, thus eliminating their interference effect. The method was optimized to various working parameters and under the selected conditions the calibration curve was linear over the range 1×10^?10–8×10^?9 mol L^?1 vitamin B₁₂ (R²=0.994), while the limit of detection for a signal‐to‐noise ratio of 3 (7×10^?11 mol L^?1 vitamin B₁₂) is the lowest value of any reported in the literature for the electrochemical determination of vitamin B₁₂. The sensors were successfully applied to the determination of vitamin B₁₂ in pharmaceutical products.     

Bienzyme amperometric lactate-specific electrode

The electrode, based on a lactate dehydrogenase and a diaphorase, permits the assay of l-lactate in the concentration range 0.2–8 mM with a response time of about 40 s. Both the enzymes are commercially available. The amperometric detection of hexacyanoferrate(II) at a platinum electrode is done at 0.3 V (vs. SCE) instead of 0.8 V as in the detection of NADH, improving the selectivity of the sensor.