Simple determination of betaine,l‐carnitine and choline in human urine using self‐packed column and column‐switching ion chromatography with nonsuppressed conductivity detection

A sequential online extraction, clean‐up and separation system for the determination of betaine, l ‐carnitine and choline in human urine using column‐switching ion chromatography with nonsuppressed conductivity detection was developed in this work. A self‐packed pretreatment column (50 × 4.6 mm, i.d.) was used for the extraction and clean‐up of betaine, l ‐carnitine and choline. The separation was achieved using self‐packed cationic exchange column (150 × 4.6 mm, i.d.), followed by nonsuppressed conductivity detection. Under optimized experimental conditions, the developed method presented good analytical performance, with excellent linearity in the range of 0.60–100 μg mL⁻¹ for betaine, 0.75–100 μg mL⁻¹ for l ‐carnitine and 0.50–100 μg mL⁻¹ for choline, with all correlation coefficients (R²) >0.99 in urine. The limits of detection were 0.15 μg mL⁻¹ for betaine, 0.20 μg mL⁻¹ for l ‐carnitine and 0.09 μg mL⁻¹ for choline. The intra‐ and inter‐day accuracy and precision for all quality controls were within ±10.32 and ±9.05%, respectively. Satisfactory recovery was observed between 92.8 and 102.0%. The validated method was successfully applied to the detection of urinary samples from 10 healthy people. The values detected in human urine using the proposed method showed good agreement with the measurement reported previously.     

Simultaneous Electrochemical Detection of Co(II) and Cu(II) by 1‐Diazo‐2‐Naphthol‐4‐Sulfonic Acid/MWCNTs Modified Electrode

This work describes a simple preparation of 1‐diazo‐2‐naphthol‐4‐sulfonic acid (1,2,4‐acid) and multiwalled carbon nanotubes (MWCNTs) modified glassy carbon electrode (GCE) for the simultaneous detection of Co(II) and Cu(II). MWCNTs, with their good conductivity and large surface area, were drop‐casted onto the surface of the GCE prior to the electrodeposition of 1,2,4‐acid, a metal chelating agent. Co(II) and Cu(II) were simultaneously measured by differential pulse anodic stripping voltammetry (DPASV) in a batch system. Under optimum conditions, the linear range of Co(II) was between 0.10 and 2.5 μg mL⁻¹ with an LOD of 80 ng mL⁻¹. Two linear ranges were obtained for Cu(II), 0.0050 to 0.030 μg mL⁻¹ and 0.040 to 0.25 μg mL⁻¹,with an LOD of 2.4 ng mL⁻¹. The method offered a high operational stability for up to 52 measurements (RSD=3.4 % for Co(II) and 2.6 % for Cu(II)) and good reproducibility (RSD=1.2 % for Co(II) and 1.7 % for Cu(II)). In the simultaneous detection of Co(II) and Cu(II), there was no effect from common interferences found in wastewater. The method was successfully applied in real water samples with good recoveries (88.2±0.8 to 102.0±0.8 % for Co(II) and 96.5±0.4 to 103.8±0.9 % for Cu(II)) and the results were in good agreement with those obtained from inductively coupled plasma optical emission spectrometry (ICP‐OES) (P >0.05).     

Simultaneous Kinetic‐Potentiometric Determination of Hydrazine and Thiosemicarbazide by Partial Least Squares and Principle Component Regression Methods

Simultaneous determination of hydrazine (HZ) and thiosemicarbazide (TSC) by partial least squares (PLS) and principle component regression (PCR) was carried out based on kinetic data of novel potentiometry. The rate of chloride ion production in reaction of HZ and TSC with N‐chlorosuccinimide (NCS) was monitored by a chloride ion‐selective electrode. The experimental dada shows not only the good ability of ion‐selective electrodes (ISEs) as detectors for the direct determination of chloride ions but also for simultaneous kinetic‐potentiometric analysis using chemometrics methods. The methods are based on the difference observed in the production rate of chloride ions. The results show that simultaneous determination of HZ and TSC can be performed in their concentration ranges of 0.7‐20.0 and 0.5‐20.0 μg mL^?1, respectively. The total relative standard error for applying PLS and PCR methods to 9 synthetic samples in the concentration ranges of 0.8‐10 μg mL^?1 of TSC and 1.0‐12.0 μg mL^?1 of HZ was 4.62 and 4.98, respectively. The effects of certain foreign ions upon the reaction rate were determined for the assessment of the selectivity of the method. Both methods (PLS and PCR) were validated using a set of synthetic sample mixtures and then applied for simultaneous determination of HZ and TSC in water samples.     

Voltammetric Method for the Simultaneous Determination of Melatonin and Pyridoxine in Dietary Supplements Using a Cathodically Pretreated Boron‐doped Diamond Electrode

The simultaneous voltammetric determination of melatonin (MT) and pyridoxine (PY) has been carried out at a cathodically pretreated boron‐doped diamond electrode. By using cyclic voltammetry, a separation of the oxidation peak potentials of both compounds present in mixture was about 0.47 V in Britton‐Robinson buffer, pH 2. The results obtained by square‐wave voltammetry allowed a method to be developed for determination of MT and PY simultaneously in the ranges 1–100 μg mL⁻¹ (4.3×10⁻⁶–4.3×10⁻⁴ mol L⁻¹) and 10–175 μg mL⁻¹ (4.9×10⁻⁵–8.5×10⁻⁴ mol L⁻¹), with detection limits of 0.14 μg mL⁻¹ (6.0×10⁻⁷ mol L⁻¹) and 1.35 μg mL⁻¹ (6.6×10⁻⁶ mol L⁻¹), respectively. The proposed method was successfully to the dietary supplements samples containing these compounds for health‐caring purposes.     

Voltammetric Analysis of Cadmium and Lead (Undesirable Elements) in Brine,Supported by Solid Phase Extraction – the Right Solution of the Analytical Challenge

The stripping voltammetry at HMDE is proposed for Cd and Pb (undesirable ingredients) determination in the natural brine (C_Cl >43 g L⁻¹). Samples with so high salinity have to be significantly diluted. For ICP MS, a 10^5–6 times dilution is required, which disqualifies this method. The proposed procedure allows to determine Cd (0.001 μg L⁻¹) and Pb (0.005 μg L⁻¹) after only 100 times dilution. The thermal chloride stripping or isolation by Chelex 100 increase the quality of obtained data. The recovery study was performed. The LOQs are below recommendations related to the use of brines in balneology.     

Mesoporous Bimetallic PtPd Nanoflowers as a Platform to Enhance Electrocatalytic Activity of Acetylcholinesterase for Organophosphate Pesticide Detection

Mesoporous bimetallic PtPd nanoflowers (MPtPdN) were synthesized by a surfactant‐directing method. The MPtPdN was characterized by scanning electron microscopy (SEM), transmission electron microscopy (TEM), and wide‐angle X‐ray diffraction (XRD). The use of MPtPdN as a platform for the indirect electrochemical detection of organophosphate pesticides (OPs) via enzymatic inhibition pathway was demonstrated. Due to the nanostructure of MPtPdN and the synergy effect of the noble metal nanoparticles, a novel and sensitive acetylcholinesterase (AChE) biosensor for OPs detections based on MPtPdN and enzyme inhibition was prepared. The inhibition of omethoate exhibited a linear relationship from 4.7×10⁻¹¹ to 4.7×10⁻⁸ M with a detection limit of 1.7×10⁻¹² M (S/N=3). The proposed AChE biosensor was reliable and can be used to measure the concentration of omethoate in different spiked samples with high accuracy and satisfactory recovery. The preparation of biosensors based on the MPtPdN can be further extended to construct many other important enzyme biosensors.     

Efficient kefiran production by Lactobacillus kefiranofaciens ATCC 43761 in submerged cultivation: Influence of osmotic stress and nonionic surfactants,and potential bioactivities

Kefiran is a water soluble polysaccharide produced by Lactobacillus kefiranofaciens ATCC 43761. It has wide potential applications in food, pharmaceutical and nutraceutical industries. To the best of our knowledge, there have been no previous reports on the effect of osmotic stress and ionic surfactants on kefiran production by L. kefiranofaciens ATCC 43761. Accordingly, the current work aimed at optimizing kefiran production as affected by osmotic stress and nonionic surfactants in submerged cultivation system. Afterwards, the work was extended to investigate cytotoxic as well as antioxidant potentials of kefiran. Firstly, different osmolarities, different ionic surfactants (Triton X-100, Tween 20, Tween 80) as well as their concentrations and addition time were evaluated. The kinetics of cell growth and kefiran production were evaluated before and after the addition of surfactants. Results clearly demonstrated that osmotic stress and surfactant addition had a stimulatory effect on kefiran production. Using the optimal medium osmolality, 550 mOsmol.kg⁻¹, kefiran production was enhanced from 1.29 to about 1.38 g.L⁻¹. Furthermore, Triton X-100 was found to be the best surfactant stimulating kefiran production when added at a concentration of 1.0 g.L⁻¹ at the onset of cultivation process (0 h). This increased kefiran production from 1.38 g.L⁻¹ to 1.62 g.L⁻¹. To summarize, the maximal kefiran production can be enhanced using 550 mOsmol.kg⁻¹ and by adding 1.0 g.L⁻¹ of Triton X-100 at 0 h. The new optimized medium showed an increase of about 25.6% in kefiran production (1.29 up to 1.62 g.L⁻¹). After this step, the process was further optimized in 16-L stirred tank bioreactor. Maximal kefiran production reached 2.32 g.L⁻¹ and 1.87 g.L⁻¹ in bioreactor under control and un-controlled pH conditions, respectively, corresponding to 72.9 and 45.0% increase from the initial production titer, respectively. The produced kefiran exhibited promising anticancer activity against breast cancer (MCF-7) cells, with an IC₅₀ value of 193.89 μg.mL⁻¹. Also, kefiran showed 96.58% radical scavenging activity at 100 μg/mL, with an ED₅₀ recorded of 12.29 ± 0.98 μg.mL⁻¹.     

Simultaneous Quantitation of Captopril and NSAID's in API,Dosage Formulations and Human Serum by RP‐HPLC

An accurate, sensitive and least time consuming reverse phase high performance liquid chromatographic (RP‐HPLC) method for the estimation of captopril in the presence of non steroidal anti‐inflammatory drugs in formulation and human serum has been developed and validated. Chromatographic separation was conducted on prepacked Purospher star C18 (5 μm, 25 × 0.46 cm) column at room temperature using methanol:water (80:20 v/v) as a mobile phase, pH adjusted at 2.8 with o‐phosphoric acid and at a flow rate of 1.0 mL min⁻¹, while UV detection was performed at 227 nm. The limit of detection and quantification for captopril were 1 and 0.35 ng mL⁻¹, while that for (NSAID's) i.e. flurbiprofen, ibuprofen, diclofenac sodium and mefenamic acid LOD were 0.2, 1, 2 and 0.4 ng mL⁻¹ respectively and LOQ were 0.9, 2.9, 8 and 1 ng mL⁻¹ Analytical recovery was > 98.1%. The method used for the quantitative analysis of commonly administered non steroidal anti‐inflammatory drugs (NSAID's) i.e. ibuprofen, flurbiprofen, diclofenac sodium and mefenamic acid alone or in combination with captopril from API (active pharmaceutical ingredients), dosage formulations and in human serum. The established method is rapid (RT < 12 min), accurate (recovery > 98.1%), selective (no interference of excepients and other commonly used drugs and food) and sensitive (LOQ 3.5 ng mL^;‐1) and reproducible (SD ± 0.003).     

Development and validation of a generic RP‐HPLC PDA method for the simultaneous separation and quantification of active ingredients in cold and cough medicines

A novel generic reverse phase high performance liquid chromatography (RP‐HPLC) method is developed and validated for simultaneous determination of seven pharmaceutically active ingredients, namely, acetaminophen, dextromethorphan, doxylamine, phenylephrine, guaifenesin, caffeine and aspirin. All seven ingredients were quantified in soft gel, syrup and tablet formulations of the over‐the‐counter US‐marketed products, as per the guidelines of the International Conference on Harmonization. The separation was achieved in a 16 min run time on an Agilent Zorbax Phenyl column using a gradient method with two mobile phases. Mobile phase A was 0.15% trifluoro acetic acid in purified water and while mobile phase B was a mixture of acetonitrile and methanol (750:250 v/v) with 0.02% trifluoro acetic acid. The flow rate was 1.0 mL min^?1 and injection volume was 10 μL. Detection was performed at 280 nm using a photodiode array detector. As part of the method validation, specificity, linearity, precision and recovery parameters were verified. The concentration and area relationships were linear (R² > 0.999), over the concentration ranges 20–120 μg mL^?1 for acetaminophen, 75–450 μg mL^?1 for dextromethorphan, 31.25–187.5 μg mL^?1 for doxylamine, 25–150 μg mL^?1 for phenylephrine, 25–150 μg mL^?1 for aspirin, 6.5–39 μg mL^?1 for caffeine and 12–72 μg mL^?1 for guaifenesin. The relative standard deviations for precision and intermediate precision were <1.5%. The proposed RP‐HPLC generic method is applicable for routine analysis of cold and cough over‐the‐counter products.     

Relative tissue distribution and excretion studies of gastrodin and parishin from powder and extract of Gastrodiae Rhizoma in rat by UHPLC‐ESI‐MS/MS

New research has indicated that Gastrodiae Rhizome (GR) has potential anti‐diabetic and anti‐asthmatic effects in mouse models. On the basis of our previous study of the relative bioavailability of gastrodin (GAS) and parishin (PA) from extract and powder of GR, we performed further research on the tissue distribution and excretion of the two analytes. A reliable bioanalytical method for the quantification of GAS and PA in rat tissues and excretion is required. Chromatographic separation was carried out on a gradient mobile phase of acetonitrile–water with 0.1% formic acid. Calibration curves (1/x ² weighted) offered satisfactory linearity (r ² > 0.9835) within 100–3000 ng mL⁻¹ for GAS and (r ² > 0.9862) within 10–1000 ng mL⁻¹ for PA. The relative standard deviations of the intra‐day and inter‐day precision were all <14.98%, whilst the relative errors of the intra‐day and inter‐day accuracy were all within ±14.71%. The matrix effect and recovery values were satisfactory in all of the biological matrices examination. The data of relative differences in tissue distribution and excretion of GAS and PA from powder and extract of GR indicated that higher bioavailabilities for GAS and PA were obtained when a dosage of 4 g kg⁻¹ GR powder was used.     

Potentiometric Immunosensor Based on Immobilization of Hepatitis B Surface Antibody on Platinum Electrode Modified Silver Colloids and Polyvinyl Butyral as Matrixes

A highly sensitive immunosensor based on immobilization of hepatitis B surface antibody (HBsAb) on platinum electrode (Pt) modified silver colloids and polyvinyl butyral (PVB) as matrixes has been developed for potentiometric immunoanalysis to detect hepatitis B surface antigen (HBsAg) in this study. HBsAb molecules were immobilized successfully on nanometer‐sized silver colloid particles associated with polyvinyl butyral on a platinum electrode surface. The modification procedure was electrochemically monitored by electrochemical impedance spectroscopy (EIS) and cyclic voltammetry (CV). The HBsAb‐silver‐PVB‐modified electrode exhibited direct electrochemical behavior toward HBsAg. The factors influencing the performance of the resulting immunosensor were studied in detail. More than 94.7% of the results of human serum samples obtained by this method were in agreement with those obtained by enzyme‐linked immunosorbent assays (ELISAs). The resulting immunosensor exhibited a sigmoid curve with log HBsAg concentration, high sensitivity (39.8 mV/decade), wide linear range from 16.0 to 800 ng mL^?1 with a detection limit of 3.6 ng mL^?1, fast potentiometric response (<3 min) and long‐term stability (>4 months). The response mechanism of the immunosensors was also studied with AC impedance techniques.     

New Insights on Potentiometric Immunosensor at Carbon Fiber Microelectrode for Alpha-Fetoprotein in Hepatocellular Carcinoma

Herein, we investigated the analytical features of potentiometric immunosensors for detection of alpha-fetoprotein (AFP) in hepatocellular carcinoma at different electrodes, such as carbon fiber microelectrode (CFME) and carbon-disk electrode (CDE), respectively. To construct such an immunosensor, anti-AFP capture antibodies were first conjugated covalently onto the activated electrodes through typical carbodiimide coupling. Thereafter, one-step immunoreaction protocol was successfully introduced to develop a new potentiometric immunoassay upon addition of AFP. Accompanying the antigen-antibody reaction, the surface charges of the modified electrodes were changed for the readout of electric potential. Results indicated that the linear range of CDE-based immunosensor was 0.1–100 ng mL⁻¹ AFP, whereas the assay sensitivity by using CFME could be further increased to 3.2 pg mL⁻¹ with the linear range from 0.01 to 500 ng mL⁻¹ AFP. Meanwhile, CFME-based immunosensor showed high sensitivity, good reproducibility and specificity, and could be utilized for the analysis of human serum specimens with consistent results relative to commercialized ELISA kit.     

Direct and Rapid Detection of Diphtherotoxin via Potentiometric Immunosensor Based on Nanoparticles Mixture and Polyvinyl Butyral as Matrixes

In this paper a novel potentiometric immunosensor for direct and rapid detection of diphtherotoxin (D‐Ag) has been developed by means of self‐assembly of monoclonal diphtheria antibody (D‐Ab) onto a platinum electrode based on nanoparticles mixture (containing gold nanoparticles and silica nanoparticles) and polyvinyl butyral (PVB) as matrixes. At first, D‐Ab was absorbed onto the surface of nanoparticles mixture, and then they were entrapped into polyvinyl butyral sol‐gel network on a platinum electrode. The detection is based on the change in the potentiometric response before and after the antigen‐antibody reaction in a phosphate buffer solution (pH 7.0). The immobilized D‐Ab exhibited direct potentiometric response toward D‐Ag. In comparison to the conventional applied methods, this strategy could allow antibodies immobilized with higher loading amount and better retained immunoactivity, as demonstrated by potentiometric response, cyclic voltammetry and electrochemical impedance spectroscopy of the immunosensor. The immunosensor with nanoparticles mixture exhibited much higher sensitivity, better reproducibility, and long‐term stability than that with gold nanoparticles or silica nanoparticles alone. The linear range was from 5.0×10^?3 to 1.2 μg?mL^?1 with a detection limit of 1.1×10^?3 μg?mL^?1. Up to 16 successive assay cycles with retentive sensitivity were achieved for the probes regenerated with in 0.2 mol?L^?1 glycine‐hydrochloric acid (Gly‐HCl) buffer solution and 0.25 mol?L^?1 NaCl. Moreover, the immunosensor with nanoparticles mixture was applied to evaluate a number of practical specimens with potentiometric results in acceptable agreement with those given by the ELISA method, implying a promising alternative approach for detecting diphtherotoxin in the clinical diagnosis.     

Implementation of a Simple Nanostructured Bio‐electrode with Immobilized Rhus Vernicifera Laccase for Oxygen Sensing Applications

In this work a simple nanostructured direct‐electron transfer bio‐electrode based on tree laccase from Rhus vernicifera is described. The electrode was implemented on a 2 mm diameter graphite mine casted with a reduced graphene surface presenting the specific capacitance of 195.8 F g⁻¹. About 10 μl of mixture between 25 mg mL⁻¹ laccase suspension and 5 mg mL⁻¹ single‐walled carbon nanotubes in 2 % SDS is dropped over the surface followed by 5 μl of the biological friendly tetrakis(2,3‐dihydroxypropyl)‐silane monomer sol to provide physical entrapment in a silica matrix after gelation. The rigidity of enzyme encapsulation allowed to obtain a constant enzyme turnover of about 16 min⁻¹ in the extended pH range of 6.0‐7.5, being the activity almost proportional to the temperature used in the interval between 25 and 40 °C. The graphite‐graphene/SWCNT‐laccase/sol‐gel electrode enabled a proportional response to molecular oxygen up to the concentration of 0.45 mmol L⁻¹ and is capable to generate the maximum power of 4.5 μW cm⁻² at 0.250 V vs the AgCl/Ag reference electrode in quiescent oxygen saturated solution.     

In-Syringe Electrokinetic Protein Removal from Biological Samples prior to Electrospray Ionization Mass Spectrometry

Here, an electrokinetic extraction (EkE) syringe is presented allowing for on-line electrokinetic removal of serum proteins before ESI-MS. The proposed concept is demonstrated by the determination of pharmaceuticals from human serum within minutes, with sample preparation limited to a 5× dilution of the sample in the background electrolyte (BGE) and application of voltage, both of which can be performed in-syringe. Signal enhancements of 3.6–32 fold relative to direct infusion of diluted serum and up to 10.8 fold enhancement, were obtained for basic and acidic pharmaceuticals, respectively. Linear correlations for the basic drugs by EkE-ESI-MS/MS were achieved, covering the necessary clinical range with LOQs of 5.3, 7.8, 6.1, and 17.8 ng mL⁻¹ for clomipramine, chlorphenamine, pindolol, and atenolol, respectively. For the acidic drugs, the EkE-ESI-MS LOQs were 3.1 μg mL⁻¹ and 2.9 μg mL⁻¹ for naproxen and paracetamol, respectively. The EkE-ESI-MS and EkE-ESI-MS/MS methods showed good accuracy (%found of 81 % to 120 %), precision (≤20 %), and linearity (r>0.997) for all the studied drugs in spiked serum samples.     

A New Validated Potentiometric Method for Batch and Continuous Quality Control Monitoring of Oseltamivir Phosphate (Taminil) in Drug Formulations and Biological Fluids

A new validated potentiometric method is described for batch and continuous quality control monitoring of the drug oseltamivir phosphate (Taminil) (OST). The method involves the development of a potentiometric sensor responsive to the drug based on the use of the ion‐association complex of (OST⁺) cation with phosphomolybdate anion (PMA^?) as an electroactive material in a poly(vinyl chloride) matrix membrane plasticized with o‐nitrophenyloctyl ether (o‐NPOE). Optimization of the performance characteristics of the sensor is described. A membrane incorporating the OST‐PMA‐NPOE complex in a tubular flow through detector is used in a two channel flow injection set up for continuous monitoring of the drug at a frequency of ～30 samples h^?1. The sensor shows fast near‐Nernstian response for OST over the concentration range 5.2×10^?5–0.8×10^?2 M (21.34 µg mL^?1–3.23 mg mL^?1) with a detection limit of 9.1×10^?6 M (3.73 µg mL^?1) over the pH range 4.6–6.1. The sensor displays good selectivity for OST drug over some basic drugs, inorganic cations, excipients and diluents commonly used in the drug formulations. Validation of the assay method is tested by measuring the lower detection limit, range, linearity, bias, trueness, accuracy, precision, and between‐day‐variability, within day reproducibility, selectivity and ruggedness (robustness). The results reveal good potentiometric performance of the proposed sensor for determination of OST in pharmaceutical capsules and in biological fluid matrices as well as for testing the dissolution profile of the drug and drug homogeneity.     

Solid phase extraction with electrospun nanofibers for determination of retinol and α-tocopherol in plasma

A novel packed-fiber solid phase extraction procedure based on electrospun nanofibers for simultaneous determination of vitamins A (retinol) and E (α-tocopherol) in human plasma has been developed. Parameters affecting extraction efficiency were investigated in detail. The limit of detection is 0.01 μg mL^?1 for retinol, and 0.3 μg mL^?1 for α-tocopherol. The linear range is from 0.05 to 2.0 μg mL^?1 for retinol, and from 0.5 to 30 μg mL^?1 for α-tocopherol. The precision (RSD) is <6%, and the relative recovery >90%. The method was applied to analysis of retinol and α-tocopherol in human plasma with satisfactory results.     

A 4-Methylumbelliferone-based Fluorescent Probe for Sodium New Houttuyfonate

A new fluorogenic probe for sodium new houttuyfonate (SNH) was proposed. 4‐Methylumbelliferyl‐2,4‐dinitrobenzenesulfonate (4‐MUDNBS) was a nonfluorescent compound and was synthesized via the one‐step reaction of 4‐methylumbelliferone (4‐MU) with 2,4‐dinitrobenzenesulfonyl chloride. In basic media, SNH was decomposed to produce sodium sulfite, which then reacted with 4‐MUDNBS to yield highly fluorescent 4‐MU, hence leading to the fluorescence increase of the reaction solution. A linear correlation existed between the emission intensity and the concentration of SNH within the range from 0.5 to 15 μg·mL⁻¹ with a detection limit of 0.15 μg· mL⁻¹ (3δ). The effect of substituents on the benzenesulfonyl moiety of the probe is discussed, and the presence of electronegative groups is favorable for the proposed cleavage reaction.     

Carbon‐nanotube Modified Screen‐printed Electrode for the Simultaneous Determination of Nitrite and Uric Acid in Biological Fluids Using Batch‐injection Amperometric Detection

A portable electroanalytical system applied for rapid and simultaneous determination of uric acid (UA) and nitrite (NIT) in human biological fluids (urine, saliva and blood) is reported. The system is based on batch‐injection analysis with multiple‐pulse amperometric (BIA‐MPA) detection using screen‐printed electrodes (SPEs) modified with multi‐walled carbon nanotubes. Sample dilution in optimized electrolyte (0.1 mol L⁻¹ Britton‐Robinson buffer pH 2) followed by injection of 100 μL on the electrode surface using an electronic micropipette is performed. UA is detected at +0.45 V and both UA+NIT at +0.70 V. Linear calibration plots for UA and NIT were obtained over the range of 1–500 μmol L⁻¹ with detection limits of 0.05 and 0.06 μmol L⁻¹, respectively. For comparison, a differential‐pulse voltammetric (DPV) method was optimized, and linear calibration plots for UA and NIT were obtained over range of 1–30 μmol L⁻¹ and 1–40 μmol L⁻¹ with detection limits of 0.1 and 0.3 μmol L⁻¹, respectively. BIA‐MPA is highly precise (RSD<1.3 %), fast (160 h⁻¹) and free from sample‐matrix interferences as recovery values ranged from 77 to 121 % for spiked samples (short contact time of sample aliquot with SPE). Contrarily, recovery tests conducted using DPV did not provide adequate recovery values (>150 %), probably due to the longer contact time of the SPE with the biological samples during analysis leading to a severe interference of sample matrices.     

Acetylcholine Biosensor Based on Dendrimer Layers for Pesticides Detection

We tested a new design of an enzyme biosensor based on acetylcholinesterase (AChE) and choline oxidase (ChO) immobilized on the supported monomolecular layer composed of poly(amidoamine) (PAMAM) dendrimers of the fourth generation (G4) mixed with 1‐hexadecanethiol (HDT). The resulting enzymatic activity, measured amperometrically, was substantially depressed in the presence of the organophosphate pesticide dimethyl‐2,2‐dichlorovinylphosphate (DDVP, Dichlorvos), carbamate pesticides carbofuran and carbamate drug eserine. The detection limits (1.3×10^?3 ppb for DDVP, 0.01 ppb for carbofuran and 0.03 for eserine) were considerably lower than so far reported for AChE based amperometric and potentiometric sensors. The relative simple protocol of biosensor preparation, high sensitivity and stability is very promising for determination of environmental pollutants in field conditions.