A specific enzyme electrode for urea

A truly specific, simple enzyme electrode is described for the assay of urea in blood serum. The sensor used is the newly developed air-gap electrode of R??i?ka and Hansen, and has advantages of speed of response and specificity over earlier enzyme electrodes for urea. Potassium, sodium and ammonium ions and other organic and inorganic species present in blood do not interfere. Linear curves are obtained from 2 · 10^-2M to 1 · 10^-4M urea with slopes close to Nernstian (about 0.90 pH/decade). Urea in blood was assayed with an accuracy of 2.2% and a precision of 2.0% with immobilized urease; only 3–5 min is required per assay. The electrode was used for a month and almost 500 assays with excellent results. Since the sensor never touches the sample solution, problems caused by blood components which block membrane pores are avoided.     

Flow injection analysis for glucose and urea with enzyme reactors and on-line dialysis

A flow injection system for glucose and urea determination is described. The glucose determination uses immobilized glucose oxidase in a reactor designed to give 100% substrate conversion. The hydrogen peroxide formed is converted to a coloured complex with 4-aminophenazone and N,N-dimethylaniline. The coupling is catalysed by a reactor containing immobilized peroxidase. The coloured complex is measured in a flow-through spectrophotometric cell. Urea is converted to ammonia in a reactor with immobilized urease and detected with an ammonia gas membrane electrode. Proteins and other interfering species from serum samples are removed in an on-line dialyzer. Calibration curves are linear for glucose in the range 1.6 × 10^-4–1.6 × 10^-2 M and for urea in the range 10^-4–10^-1 M. The samples are 25 μl for glucose determination and 100 μl for urea determination. Linear ranges can be changed by varying the sample sizes. The effects of the dialyser, enzyme reactors and detectors on dispersion are evaluated.     

An improved electrode for the assay of urea in blood

An improved urea enzyme electrode is applied for the determination of urea in blood samples. The electrode is based on the enzymatic hydrolysis of urea, and potentiometric detection of the ammonium ion produced. A silicone rubber-based nonactin ammonium ion-selective electrode serves as the sensor. The selectivity coefficients of this electrode were 6.5 for NH₄⁺/K⁺; 750 for NH₄⁺/Na⁺, and much higher for other cations. The reaction layer of the electrode was made of urease enzyme chemically immobilized on polyacrylic gel. The prepared gel was stable at 4° for over four months. The electrodes retained their activity for over one month. A three-electrode system, which allowed dilution to a constant interference level, was applied to avoid interfering effects in blood samples. Analyses of blood sera showed good agreement with a standard spectrophotometric method. Routine clinical assays of blood urea are feasible.     

Part I. A specific electrode for urea.

A specific simple enzyme stirrer electrode is described for the assay of urea in blood serum. The enzyme is placed directly on a magnetic stirrer and held in place with a nylon net. The enzyme stirrer both stirs the solution and effects an enzymatic transformation, permitting the direct assay of a substrate such as urea. Potassium, Na⁺ , NH₄⁺ and other organic and inorganic species present in blood do not interfere. Linear curves are obtained from 5· 10^-2M to 1· 10^-4M urea with slopes close to Nernstian, 0.95 pH/decade. Urea in blood was assayed with an accuracy of 1.8% and a precision of 2.0% with immobilized urease in the stirrer. The stirrers were used for 15 weeks and over 500 assays with excellent results.     

An enzyme reactor electrode for urea determinations.

An enzyme reactor electrode system for the determination of urea is described. A buffer is pumped through an enzyme reactor (0.4 ml) containing urease immobilized with glutaraldehyde to glass. The effluent is mixed with sodium hydroxide pumped through a second channel and fed through an ammonia gas electrode. Samples are introduced via a third flow channel and mixed with the buffer. The conversion of urea to ammonia is quantitative for sample concentrations of less than 0.03 M for a flow rate of 40 ml h^-1. The reactor electrode shows a Nernstian slope of 57 mV/decade for 5·10^-5–3·10^-2 M urea. The response is independent of variations in the flow rate, enzyme activity or temperature of the reactor.     

Rapid and accurate amperometric determination of acetaminophen in pharmaceutical preparations and spiked human blood serum samples at cadmium pentacyanonitrosylferrate modified glassy carbon electrode

The present work describes a rapid and accurate amperometric technique for the determination of acetaminophen (ACT) in pharmaceutical preparations and human blood serum, based on electrocatalytic oxidation of ACT at a glassy carbon electrode modified by cadmium pentacyanonitrosylferrate (CdPCNF) film. The electrocatalytic response of the modified GC electrode was linear over the concentration of 1.64-52.90 μM. The limit of detection was found to be 2.04 μM by amperometric technique. The method was successfully utilized for the determination of ACT in various pharmaceutical preparations and the results have been statistically compared with those obtained by the official method. The interference of some pharmaceutical and biological compounds was investigated. The results of interference study showed that the Nafion-coated CdPCNF|GC electrode can be utilized as a selective amperometric sensor for acetaminophen determination in human blood serum. The mean value of rate constant k for catalytic reaction, and the diffusion coefficient of ACT (D) in the phosphate buffer solution of pH 7.2 were found to be 4.27 × 10² M?¹ s?¹, and (4.25 ± 0.33) × 10^?6 cm² s^?1, respectively.     

Highly sensitive determination of hydroxylamine using fused gold nanoparticles immobilized on sol-gel film modified gold electrode

We are reporting the highly sensitive determination of hydroxylamine (HA) using 2-mercapto-4-methyl-5-thiazoleacetic acid (TAA) capped fused spherical gold nanoparticles (AuNPs) modified Au electrode. The fused TAA-AuNPs were immobilized on (3-mercaptopropyl)-trimethoxysilane (MPTS) sol-gel film, which was pre-assembled on Au electrode. The immobilization of fused TAA-AuNPs on MPTS sol-gel film was confirmed by UV-vis absorption spectroscopy and atomic force microscopy (AFM). The AFM image showed that the AuNPs retained the fused spherical morphology after immobilized on sol-gel film. The fused TAA-AuNPs on MPTS modified Au electrode were used for the determination of HA in phosphate buffer (PB) solution (pH = 7.2). When compared to bare Au electrode, the fused AuNPs modified electrode not only shifted the oxidation potential of HA towards less positive potential but also enhanced its oxidation peak current. Further, the oxidation of HA was highly stable at fused AuNPs modified electrode. Using amperometric method, determination of 17.5 nM HA was achieved for the first time. Further, the current response of HA increases linearly while increasing its concentration from 17.5 nM to 22 mM and a detection limit was found to be 0.39 nM (S/N = 3). The present modified electrode was also successfully used for the determination of 17.5 nM HA in the presence of 200-fold excess of common interferents such as urea, NO₂⁻, NH₄⁺, oxalate, Mn²⁺, Na⁺, K⁺, Mg²⁺, Ca²⁺, Ba²⁺ and Cu²⁺. The practical application of the present modified electrode was demonstrated by measuring the concentration of HA in ground water samples.     

A novel PVC membrane sensor for determination of loratadine in pharmaceutical compounds and blood serum

A novel PVC membrane-selective electrode based on loratadine-tetraphenyl borate ion-pair was prepared for the determination of loratadine potentiometrically. This electrode exhibits a Nernstian slope of 59.1 ± 0.3 mV decade^-1 for loratadine in a concentration range of 5.0 × 10^-6?1.0 × 10^-2 M at pH 2.2 with a detection limit of 2.9 × 10^-6 M. The potential of the electrode is very stable and exhibits good reproducibility with very fast response time (??3 s). The selectivity of the proposed electrode towards some cations and organic compounds was tested and the selectivity coefficients were calculated. The electrode was successfully applied to the determination of loratadine in tablets and blood samples.     

Enzyme coated glass pH-electrode: Its fabrication and applications in the determination of urea in blood samples

A new enzyme coated electrode for the determination of urea in blood samples has been developed. It is based on the encapsulation of urease enzyme in the porous silicate matrix by the sol-gel technique on a glass electrode for the purpose of sensing urea in blood samples. Various parameters like the effect of pH, selection of a suitable buffer of appropriate concentration and interference of common substances in blood samples have been evaluated to optimize the conditions for the determination of urea. The electrode can be used for the determination of urea in the concentration range 0.03-30.0 mM in a solution. The detection limit of the present enzyme-coated electrode is found to be 52 μg/ml of urea. The relative standard deviation for the electrode-to-electrode reproducibility is found to be 2.4% for the determination of 0.1 mM of urea (six replicate electrodes). Sol-gel matrix containing immobilized enzyme was stable for about 25 days at ∼4 °C with 80% urease activity. Urea content in various clinical blood samples has been estimated using this electrode and the results are found to be in good agreement with the standard clinical methods as reported in the literature.     

Electrochemical determination of acetaminophen in the presence of propranolol using an electrode modified with a Schiff base from 2-hydroxy-1-naphthaldehyde and ethylenediamine and multi-walled carbon nanotubes

A carbon paste electrode, modified with N,N′-bis-(2-hydroxy-1-naphthalidene)ethylenediamine and multi-walled carbon nanotubes (HNED-MWCNPE), was used for the determination of acetaminophen (ACOP) and propranolol (PP). Cyclic voltammetry (CV), chronocoulometry, chronoamperometry and differential pulse voltammetry (DPV) techniques were employed to study electro-oxidation of ACOP. The results revealed that the modified electrode showed an electrocatalytic activity toward the anodic oxidation of acetaminophen by a marked enhancement in the current response in buffered solution at pH 8.0. Some kinetic parameters such as the electron transfer coefficient (α) were also determined for the ACOP oxidation. The linear concentration range of 1 × 10^?3?1 × 10^?6 M with a detection limit of 4.6 × 10^?8 M (n = 16) for ACOP was obtained using DPV (pH 8.0). The modified electrode shows good sensitivity, selectivity and stability. The prepared electrode was also applied for the determination of ACOP in human blood serum.     

Preparation and comparison of Proteus vulgaris and Proteus mirabilis bacterial electrodes for the determination of DL-phenylalanine

Bacterial electrodes for DL-phenylalanine were prepared by immobilizing the bacteria Proteus vulgaris and Proteus mirabilis on an ammonia gas-sensor. The response of the Proteus vulgaris bacterial electrode, when 10 mg of the bacteria was used, had a linear range between 3.0 × 10⁻⁴ and 1.0 × 10⁻² M DL-phenylalanine with a response slope of 43 mV/decade in pH 7.0, 0.1 M phosphate buffer solution at 30°C, while the response of the Proteus mirabilis bacterial electrode, when 3 mg of the bacteria was used, had a linear range between 3.0 × 10⁻⁴ and 3.0 × 10⁻² M DL-phenylalanine with a response slope of 49 mV/decade in pH 7.2, 0.1 M phosphate buffer solution at 30°C. The most important interferents were urea and l-asparagine, and inorganic salts reacted as an inhibitor. The Proteus vulgaris bacterial electrode could be used directly for the determination of DL-phenylalanine in nearly the same linear range during 3 days. On the other hand, the Proteus mirabilis bacterial electrode could be used continuously during 7 days in the above linear range.     

Voltammetric determination of mercury(II) using a modified pencil graphite electrode with 4-(4-methylphenyl aminoisonitrosoacetyl)biphenyl

A sensitive and selective method for determination of mercury(II) with “4-(4-methylphenyl aminoisonitrosoacetyl)biphenyl (TKO)-modified pencil graphite electrode” was developed. All factors affecting determination process were optimized. Differential pulse voltammetry with 4-(4-methylphenyl aminoisonitrosoacetyl)biphenyl-modified electrode showed a linear response between 1.0 × 10^?5 and 1.0 × 10^?3 M (R ² = 0.9994). The detection limit of this electrode was found as 5.85 × 10^?7 M (S/N = 3). The effects of different cations on the determination of mercury(II) were investigated and found that modified electrode is highly selective. The developed method was applied for mercury determination in different water samples.     

Characterization of biosensors based on membranes containing a conducting polymer

A new type of biosensor based on the coupling of an enzyme to an ion-selective membrane containing a conducting polymer is evaluated. The results obtained with the enzyme field- effect transistor (ENFET) and the ion-selective electrode (ISE) for the determination of creatinine and urea are compared. The presence of the conducting polymer significantly lowers the detection limit for creatinine by one decade to 10^?7 and 10^?4 M for the ENFET and ISE, respectively. The determination of urea in urine and serum with the ENFET was carried out, and the results correlated well with those obtained by spectrophotometry.     

Kinetic study of the tartaric acid-periodate reaction: Determination of tartaric acid using an improved periodate-selective electrode

The preparation of an improved liquid-membrane periodate-selective electrode is described. The electrode exhibits rapid and near-Nernstian response in the range 2 × 10⁻⁶-10⁻² M. The electrode was used to monitor the course of the tartaric acid-periodate reaction. A potentiometric reaction rate method for the rapid and accurate determination of tartaric acid has been developed. A total of 0.4–120 μmol of tartaric acid has been determined with relative error of about 2.0%. The method has been applied to the determination of tartaric acid in pharmaceutical preparations and to the determination of small amounts of tartaric acid in impure citric acid.     

Voltammetric,potentiometric and amperometric studies with a rotated aluminum wire electrode : A comparison of the baker and morrison cell with the raie.

In the amperometric determination of fluoride at the RAIE a half cell composed of 5% cadmium amalgam in equilibrum with a solution 1 M in cadmium sulfate and saturated with potassium chloride can be used as a reference electrode in a short-circuited cell instead of applying a potential of -0 75 V versus the saturated calomel electrode The standard addition method can be used in the presence of air, although removal of oxygen is recommended Using the Baker and Morrison electrode versus the above half cell and following their directions (10 ml solution, magnetic stirring) proportionality between current and fluoride concentration in a range between 1 · 10^-5 and 1 · 10^-4M was found in oxygen-free solutions Halides and perchlorates do not interfere. The standard addition technique can be used in the determination of fluoride in an unknown.     

A coated-metal enzyme electrode for urea determinations

A potentiometric enzyme electrode is reported in which an enzyme immobilized in polyvinyl chloride is used to coat an antimony metal electrode to detect changes in pH when the electrode is immersed in a solution of the enzyme substrat. As an example, urea is determined in solution by using immobilized urease on an antimony electrode, giving a linear concentration range of 5.0 × 10^-4–1.0 × 10^-2 M urea with a slope of 44 mV per decade change in urea concentration. The response slope is stable for about 1 week, with response times in the range 1–2 min, but with absolute potential changes occurring from day to day.     

Improved Sensing Performance of Amperometric Urea Biosensor by Using Platinum Nanoparticles

A novel Platinum nanoparticle (PtNPs) modified Poly(pyrrole-co-1-(2-Aminophenyl)pyrrole)/Urease film coated Au electrode was designed for amperometric detection of urea. PtNPs quantity, film density and pH were optimized and interference effect of some substances readily found in municipal wastewater and blood was investigated. The biosensor responded to urea with a measurement concentration range of 0.1 to 30 mM, a sensitivity of 31.8 μA mM⁻¹ cm⁻², a LOD of 7.58 μM, an accuracy of 104 % and a RSD% of only 0.82. It sensed the concentration of urea in the municipal sewage water with recovery of 97.6 % (n=3) and remained 78 % of its initial response at 28^th day. Results confirmed that PtNPs with strong conductivity improved the electron transfer ability of the working electrode.     

Sputtering deposition of Pt nanoparticles on vertically aligned multiwalled carbon nanotubes for sensing L-cysteine

A highly sensitive electrochemical sensor for determination of L-cysteine (CySH) is presented. It is based on vertically aligned multiwalled carbon nanotubes modified with Pt nanoparticles by magnetron sputtering deposition. The morphology of the nanocomposite was characterized by scanning electron microscopy, transmission electron microscopy and energy-dispersive. The electrochemistry of CySH was investigated by cyclic voltammetry, differential pulse voltammetry and chronoamperometry. The mechanism for the electrochemical reaction of CySH at the modified electrode at different pH values is discussed. The electrode exhibits a higher electrocatalytic activity towards the oxidation of CySH than comparable other electrodes. It displays a linear dependence (R ²?=?0.9980) on the concentration of CySH in the range between 1 and 500 μM and at an applied potential of +0.45 V, a remarkably low detection limit of 0.5 μM (S/N?=?3), and an outstandingly high sensitivity of 1.42?×?10³ μA?mM^?1?cm^?2, which is the highest value ever reported. The electrode also is highly inert towards other amino acids, creatinine and urea. The sensor was applied to the determination of CySH in urine with satisfactory recovery, thus demonstrating its potential for practical applications.

Development of non-enzymatic strip for simple and selective determination of urea in water and biological samples

A simple and selective method for the determination of urea based on the paptode technique is described. The sensor was constructed by immobilizing an ionophore on a TLC strip. The procedure is based on the nucleophilic displacement of urea with tetrachloro-p-benzoquinone (chloranil) as an ionophore, and the formed violet-color product was detected using a flatbed scanner. The color of each spot was analyzed to red (R), green (G) and blue (B) values from 0 to 255 using a program written in visual basic (VB) programming language. The calibration graph obtained with the proposed sensor was linear over the range of 0.05?C10.00?mg?L^?1 with a detection limit of 0.01?mg?L^?1 for urea. Parameters such as pH and concentration of chloranil were optimized. The proposed sensor was successfully applied for the determination of urea in bovine serum, urine and tap water samples.     

Immobilized enzyme electrode for the determination of salicylate in blood serum

This determination of salicylate in blood serum is based on application of an immobilized enzyme electrode. Salicylate hydroxylase (E.C.1.14.13.1) is chemically immobilized onto a pig intestine mounted on an oxygen electrode. The signals are monitored amperometrically and the resulting output voltage is read using a simple adapter. The experimental parameters and possible interferences are discussed. Samples containing 1.0 × 10^?5?1.87 × 10^?3 M (1.6–300 μg ml^?1) salicylate were assayed with relative standard deviations between 1.3% and 6% and recoveries between 98.7 and 103%. Results obtained by the proposed method and by the established clinical method for randomly spiked pooled serum samples correlated well (r = 0.99).