Macrocyclic antibiotics as chiral selectors in the design of enantioselective, potentiometric membrane electrodes for the determination of l- and d-enantiomers of methotrexate

Three enantioselective, potentiometric membrane electrodes (EPMEs) based on macrocyclic glycopeptide antibiotics—vancomycin and teicoplanin (modified or not with acetonitrile)—were proposed for the determination of l- and d-enantiomers of methotrexate (Mtx). The linear concentration ranges for the proposed enantioselective membrane electrodes were between 10⁻⁶ and 10⁻³ mol l⁻¹ for l- and d- methotrexate. The slopes of the electrodes were 58.00 mV/pl-Mtx for vancomycin-based electrode; 57.60 mV/pd-Mtx for teicoplanin-based electrode and 55.40 mV/pd-Mtx for teicoplanin modified with acetonitrile-based electrode. The detection limits of the proposed electrodes were of 10⁻⁸ mol l⁻¹ magnitude order. The surfaces of the electrodes are stable and easily renewable by polishing on alumina paper. All proposed electrodes proved to be successful for the determination of the enantiopurity of Mtx as raw material and of its pharmaceutical formulations (tablets and injections).     

A novel potentiometric biosensor for selective L-cysteine determination using L-cysteine-desulfhydrase producing Trichosporon jirovecii yeast cells coupled with sulfide electrode

Trichosporon jirovecii yeast cells are used for the first time as a source of l-cysteine desulfhydrase enzyme (EC 4.4.1.1) and incorporated in a biosensor for determining l-cysteine. The cells are grown under cadmium stress conditions to increase the expression level of the enzyme. The intact cells are immobilized on the membrane of a solid-state Ag₂S electrode to provide a simple l-cysteine responsive biosensor. Upon immersion of the sensor in l-cysteine containing solutions, l-cysteine undergoes enzymatic hydrolysis into pyruvate, ammonia and sulfide ion. The rate of sulfide ion formation is potentiometrically measured as a function of l-cysteine concentration. Under optimized conditions (phosphate buffer pH 7, temperature 37 ± 1 °C and actual weight of immobilized yeast cells 100 mg), a linear relationship between l-cysteine concentration and the initial rate of sulfide liberation (dE/dt) is obtained. The sensor response covers the concentration range of 0.2-150 mg L⁻¹ (1.7-1250 μmol L⁻¹) l-cysteine. Validation of the assay method according to the quality control/quality assurance standards (precision, accuracy, between-day variability, within-day reproducibility, range of measurements and lower limit of detection) reveals remarkable performance characteristics of the proposed biosensor. The sensor is satisfactorily utilized for determination of l-cysteine in some pharmaceutical formulations. The lower limit of detection is ∼1 μmol L⁻¹ and the accuracy and precision of the method are 97.5% and ±1.1%, respectively. Structurally similar sulfur containing compounds such as glutathione, cystine, methionine, and d-cysteine do no interfere.     

Sequential fluorometric quantification of malic acid enantiomers by a single line flow-injection system using immobilized-enzyme reactors

A method for the sequential enantiomeric quantification of d-malate and l-malate by a single line flow-injection analysis was developed using immobilized-enzyme reactors and fluorescence detection. An immobilized d-malate dehydrogenase (d-MDH) reactor and an immobilized l-malate dehydrogenase (l-MDH) reactor were introduced into the flow line in series. Sample and coenzyme (NAD⁺ or NADP⁺) were injected into the flow line by an open sandwich method. d-Malate was selectively oxidized by d-MDH when NAD⁺ was injected with a sample. When NADP⁺ was injected with a sample, l-malate was oxidized only by l-MDH. NADH or NADPH produced by the immobilized-enzyme reactors was monitored fluorometrically at 455 nm (excitation at 340 nm). Linear relationships between the responses and concentrations of d-malate and l-malate were observed in the ranges of 1 × 10⁻⁶-1 × 10⁻⁴ M and 1 × 10⁻⁶-2 × 10⁻⁴ M, respectively. The relative standard deviations for ten successive injections were less than 2% at the 0.1 mM level. This analytical method was applied to the sequential quantification of d-malate and l-malate in fruit juices and soft drinks, and the results showed good agreement with those obtained using conventional method (F-kit method).     

Solid phase extraction of Co ions using l-tyrosine immobilized on multiwall carbon nanotubes

A study was performed to assess the performance of aminoacids immobilized on carbon nanotubes (CNTs) for their employment as a sorbent for solid phase extraction systems. An immobilization method is introduced and the aminoacid l-tyrosine was chosen as a case study. A spectrophotometric study revealed the amount of aminoacid immobilizated on CNTs surface, and it turned to be of 3174 μmol of l-tyr g⁻¹. The material was tested for Co retention using a minicolumn inserted in a flow system. At pH 7.0, the amount of Co retained by the column was of 37.58 ± 3.06 μmol Co g⁻¹ of CNTs. A 10% (v/v) HNO₃ solution was chosen as eluent. The pH study revealed that Co binding increased at elevated pH values. The calculation of the mol ratio (moles of Co bound at pH 9 to moles of l-tyr) turned to be 3:1. The retention capacity was compared to other bivalent cations and showed the following tendency: Cu²⁺ > Ni²⁺ > Zn²⁺ ? Co²⁺. The analytical performance was evaluated and an enrichment factor of 180 was obtained when 10 mL of 11.37 μg L⁻¹ Co solution was loaded onto the column at pH 9.0; reaching a limit of detection (LoD) of 50 ng L⁻¹. The proposed system was successfully applied to Co determination in QC-LL2 standard reference material (metals in natural water).     

Simultaneous determination of tiopronin and d-penicillamine in human urine by liquid chromatography with ultraviolet detection

d-Penicillamine and tiopronin are drugs widely used for the treatment of many diseases. Because of the relatively high frequency of side effects to these compounds, some of which are dose-related, drug monitoring in urine samples during treatment is advisable. In this paper, we describe a simple method for the determination of tiopronin and d-penicillamine in human urine. The method was based on derivatization with 2-chloro-1-methylquinolinium tetrafluoroborate followed by ion-pairing reversed-phase liquid chromatography separation and ultraviolet-absorbance detection. 2-S-quinolinium derivatives of thiols were detected at 355 nm. The derivatization was optimized in terms of pH and time of the reaction. Baseline separation was achieved on an analytical Zorbax SB C-18 (5 μm, 150 mm × 4.6 mm) column with a mobile phase consisting of pH 2.0 0.09 mol L⁻¹ trichloroacetic acid buffer (component A) and acetonitrile (component B) pumped at 1.0 mL min⁻¹. Gradient elution was used: 0-4 min, 12% B; 4-8 min, 12-40% B; 8-12 min, 40-12% B. The d-penicillamine and tiopronin standards added to the urine show that the response of the detector is linear within the range studied, from 1 to 200 μmol L⁻¹ urine. The imprecision ranges for tiopronin and d-penicillamine were within 1.61-8.24% and 2.92-10.60%, respectively. The analytical accuracy for determined compounds was from 97.24 to 109.39%. The lower limits of detection and quantitation were 0.5 μmol L⁻¹ and 1.0 μmol L⁻¹ urine, respectively. This method can be used for routine clinical monitoring of the title thiol-drugs. Cysteine can be measured concurrently, if needed.     

Size-dependent electrochemiluminescence behavior of water-soluble CdTe quantum dots and selective sensing of l-cysteine

Water-soluble CdTe quantum dots (QDs) with five sizes (2.25, 2.50, 2.77, 3.12, and 3.26 nm) were synthesized with the hydrothermal method. The electrochemiluminescence (ECL) of CdTe QDs was investigated in detail in air-saturated solution without adding foreign oxidant. It was found that the ECL of CdTe QDs displayed a size-dependent property. With the increasing in the particle size of the CdTe QDs, the ECL intensity was gradually increased, in addition, both ECL peak potentials and ECL onset potentials of CdTe QDs were shifted positively. Influences of some factors on the ECL intensity were investigated. Under the optimal conditions, the ECL intensity had a linear relationship with the concentration of l-cysteine (l-Cys) in the range from 1.3 × 10⁻⁶ to 3.5 × 10⁻⁵ mol L⁻¹ (R² 0.996) with a detection limit of 8.7 × 10⁻⁷ mol L⁻¹ (S/N = 3). The proposed method was applied to the determination of l-Cys in real samples with satisfactory results. Compared with previous reports, it has better selectivity for the determination of l-Cys.     

Determination of l-ascorbic acid in wines by direct injection liquid chromatography using a polymeric column

A rapid method for the identification and quantification of l-ascorbic acid in wines by direct injection liquid chromatography equipped with a UV detection was developed. The levels of ascorbic acid were determined using a polymeric PLRP-S 100 A (5 μm) column (150 mm × 4.6 mm) with a mobile water/trifluoroacetic acid (99/1, v/v) phase. The method is rapid (less than 5 min) and sensitive (LOQ of 5 mg L⁻¹). The calibration curve of ascorbic acid was linear (r = 0.999) over a concentration range between 1 and 200 mg L⁻¹. Repeatability was less than 2.5% and the recovery over 95%.     

Development of a voltammetric sensor for diospyrin determination in nanomolar concentrations

A sensor based on glassy carbon (GC) electrode modified with cobalt tetrasulfonated phthalocyanine (CoTSPc) and a poly-l-lysine (PLL) film is proposed for diospyrin determination in nanomolar concentrations with differential pulse voltammetry (DPV) technique. The modified electrode showed excellent catalytic activity presenting much higher peak currents than those measured on a bare GC electrode. Linear response range, sensitivity and limit of detection (LOD) were of 1-120 nmol l⁻¹, 220.46 nA l nmol⁻¹ cm⁻² and 0.3 nmol l⁻¹, respectively. The repeatability of the proposed sensor, evaluated in term of relative standard deviation (R.S.D.), was measured as 4.4% for 10 experiments in 50 μmol l⁻¹ diospyrin samples. The developed sensor was applied for the determination of diospyrin in the crude extracts of the stem-bark of Diospyros montana Roxb. and the average recovery for these samples was 101.9 (±3.1)%.     

Electrochemical determination of L-phenylalanine at polyaniline modified carbon electrode based on β-cyclodextrin incorporated carbon nanotube composite material and imprinted sol-gel film

A sensitive and selective electrochemical sensor based on a polyaniline modified carbon electrode for the determination of l-phenylalanine has been proposed by utilizing β-cyclodextrin (β-CD) incorporated multi-walled carbon nanotube (MWNT) and imprinted sol-gel film. The electrochemical behavior of the sensor towards l-phenylalanine was investigated by cyclic voltammetry (CV), differential pulse voltammetry (DPV), and amperometric i-t curve. The surface morphologies of layer-by-layer assembly electrodes were displayed by scanning electron microscope (SEM). The response mechanism of the imprinted sensor for l-phenylalanine was based on the inclusion interaction of β-CD and molecular recognition capacity of the imprinted film for l-phenylalanine. A linear calibration plot was obtained covering the concentration range from 5.0 × 10⁻⁷ to 1.0 × 10⁻⁴ mol L⁻¹ with a detection limit of 1.0 × 10⁻⁹ mol L⁻¹. With excellent sensitivity, selectivity, stability, reproducibility and recovery, the electrochemical imprinted sensor was used to detect l-phenylalanine in blood plasma samples successfully.     

Synthesis of cross-linked chitosan possessing N-methyl-d-glucamine moiety (CCTS-NMDG) for adsorption/concentration of boron in water samples and its accurate measurement by ICP-MS and ICP-AES

A chitosan resin derivatized with N-methyl-d-glucamine (CCTS-NMDG) was synthesized by using a cross-linked chitosan (CCTS) as base material. The N-methyl-d-glucamine (NMDG) moiety was attached to the amino group of CCTS through the arm of chloromethyloxirane. The adsorption behavior of 59 elements on the synthesized resin was systematically examined by using the resin packed in a mini-column, passing water samples through it and measuring the adsorbed elements in eluates by ICP-MS. The CCTS-NMDG resin shows high ability in boron sorption with the capacity of 0.61 mmol ml⁻¹ (= 2.1 mmol g⁻¹). The sorption kinetics of this resin was faster than that of the commercially available resins. Other advantages of the synthesized resin are: (1) quantitative collection of boron at neutral pH regions; (2) complete removal of large amounts of matrices; (3) no loss of efficiency over prolonged usage; (4) effective collection of boron in wide range concentration using a mini column containing 1 ml resin; (5) complete elution of boron with 1 mol l⁻¹ nitric acid. The resin was applied to the collection/concentration of boron in water samples. Boron in tap water and river water was found to be in the range of 6-8 μg l⁻¹. The limit of detection (LOD) of boron after pretreatment with CCTS-NMDG resin and measurement by ICP-MS was 0.07 μg l⁻¹ and the limit of quantification (LOQ) was 0.14 μg l⁻¹ when the volume of each sample and eluent was 10 ml.     

A fully automated system for inorganic antimony preconcentration and speciation in urine

A study was undertaken to ascertain the analytical capabilities of l-methionine immobilized on controlled pore glass for Sb preconcentration and speciation. A fully automated on-line system, implemented with hydride generation (HG) and inductively coupled plasma optical emission spectrometry (ICP OES), was used. Sb(III), at pH 10 was selectively retained in the column containing the immobilized aminoacid, while Sb(V) was not retained at all. A 30% HCl solution was used as eluent agent. Prior to total Sb determination, a pre-reduction step with thiourea was necessary. An on-line pH adjusting and pre-reduction of Sb(V) was achieved in a fully automated system. The detection limit for the preconcentration of 10 mL of an aqueous solution was 70 ng L⁻¹ with a relative standard deviation of 2%. An enrichment factor of 20 was achieved when 10 mL of sample was passed through the system, reaching a throughput of 23 samples per hour. The method was successfully applied to the determination of Sb(III) and total Sb in urine.     

Continuous-flow/stopped-flow system for determination of ascorbic acid using an enzymatic rotating bioreactor

The high sensitivity that can be attained using an enzymatic system and mediated by hydroquinone, has been verified by on-line interfacing of a rotating bioreactor and continuous flow/stopped-flow/continuous-flow processing. Horseradish peroxidase, HRP, [EC 1.11.1.7], immobilized on a rotating disk, in presence of hydrogen peroxide catalyses the oxidation of hydroquinone to p-benzoquinone, whose electrochemical reduction back to hydroquinone is detected on glassy carbon electrode (GCE) surface at −0.15 V. Thus, when l-ascorbic acid is added to the solution, this acid is reduced chemically (p-benzoquinone to hydroquinone) and acts as mediator of HRP, decreasing the peak current obtained proportionally to the increase of its concentration. The recovery of l-ascorbic acid from four samples ranged from 99.09 to 101.10%. This method could be used to determine l-ascorbic acid concentration in the range 12 nM-3.5 μM (r = 0.998). The determination of l-ascorbic acid was possible with a limit of detection of 6 nM in the processing of as many as 25 samples h⁻¹. The method was successfully applied for the analysis of l-ascorbic acid in pharmaceutical formulations.     

Interactions of aluminum(III) with the biologically relevant ligand d-ribose

Aluminum(III) can be absorbed when it is appropriately complexed. There are several plasma components which can bind weakly Al(III). Many proteins bind Al(III) in solution quite strongly. Carbohydrates bearing an abundance of electronegative functional groups can interact with metal cations. In solution, d-ribose exists as a mixture at equilibrium of many isomers and only a few of them bear a ‘complexing’ sequence of the hydroxyl groups. The presence of d-ribose in an Al(III) solution experiences a decrease of its Brönsted-acid sites. The lowering of the Brönsted acidity of an Al(III)-d-ribose mixture suggests the existence of attractive interactions (‘association’) between Al(III) ion and the complexing sequence of the hydroxyls of d-ribose. There is enhancement in the stability of the interaction complexes between Al(III) and d-ribose through strong intramolecular hydrogen bonding, which offers the possibility to investigate the kinetics of the subsequent proton release reactions. On the basis of the kinetic results, it may be concluded that proton release reactions, which are associated with the complexation reactions, are associatively activated. The complexes (Al(H₂O)_6−n(d-ribose_−nH)⁽³⁻ⁿ⁾⁺) resulting from the various ‘complexing’ forms of d-ribose are formed at mainly acidic pH. As the pH increases, the values of the activation enthalpy, ΔH^≠, are changing, because of the formation of mixed hydroxo-complexes (Al(H₂O)_6−n−m(OH)_m(d-ribose_−nH)^(3−n−m)+); finally, OH⁻ displaces d-ribose from the coordination sphere of Al(III) in a rather slow process, i.e. with high values of ΔH^≠; the activation enthalpy values, ΔH^≠, decrease with the progression of the displacement, becoming finally very small due to the formation of a precipitate. Chelate coordination of d-ribose with some divalent and trivalent metal ions has been also reported.     

An amperometric method for the determination of trace mercury(II) by formation of complexes with l-tyrosine

A selective and sensitive amperometric method of analysis has been developed for determination of the trace amounts of mercury in waters at a platinum electrode based on the effect of the presence of mercury ions on the current due to oxidation of l-tyrosine. A decrease of signal was observed due to the formation of a complex of tyrosine with the Hg(II) ion adsorbed on the electrode surface. Several parameters were varied, such as applied potential, pH and concentration of tyrosine. The calibration plot was linear in the range from 0.02 to 3 μmol l⁻¹ Hg(II) with r=0.997 and the detection limit (3σ) was 0.014 μmol l⁻¹; the relative standard deviation was 2.2%. The study of interferences from other metal ions revealed a good selectivity of this method towards mercury(II). The stoichiometry of the mercury-tyrosine complex was determined to be 1:2 and the formation constant 627±19. Formation of complexes with mercury ions was also demonstrated with several catechol compounds and other amino acids. The method was applied to the analysis of contaminated waters.     

Time measurement-visual analysis of l-cysteine using the autocatalytic sodium sulfite/hydrogen peroxide reaction system and its application to length detection-flow analysis

Trace amounts of l-cysteine can function as a trigger, i.e., reaction initiator, in the autocatalytic sodium sulfite/hydrogen peroxide reaction system. Rapidly changing of pH after induction time is visually confirmed by color changing of bromothymol blue in this autocatalytic reaction. Based on this finding, μg L⁻¹ levels of l-cysteine were measured over time using the autocatalytic reaction system. The determination range using the above method was 5.0 × 10⁻⁸-2.5 × 10⁻⁶ M, the detection limit (3σ) was 1.8 × 10⁻⁸ M (1.94 μg L⁻¹), and the relative standard deviation was 2.41% at an l-cysteine concentration of 5 × 10⁻⁷ M (n = 5). This method was also applied to length detection-flow injection analysis. The determination range for the flow injection analysis was 2.0 × 10⁻⁷-1.0 × 10⁻⁵ M. The detection limit (3σ) was 1.4 × 10⁻⁷ M (17.0 μg L⁻¹), and the relative standard deviation was 0.91% at an initial l-cysteine concentration of 10⁻⁶ M (n = 5).     

Electrochemical behaviour of epinephrine and uric acid at a Sonogel-Carbon l-Cysteine modified electrode

The Sonogel-Carbon electrode is a special class of sol-gel electrode that exhibits favourable mechanic and electric properties to be used as electrochemical sensor. In this study, Sonogel-Carbon modified with l-Cysteine was used to prepare a novel electrochemical sensor. The objective of this novel electrode modification was to seek new electrochemical performances for detection of epinephrine in the presence of uric acid. The response of catalytic current with epinephrine concentration shows a linear relation in the range from 1 × 10⁻⁷ to 5 × 10⁻⁴ M with a correlation coefficient of 0.998, and a detection limit of 8.7 × 10⁻⁸ M. The modified electrode had also been applied to the determination of epinephrine and uric acid in biological samples with satisfactory results. A surface characterisation of this modified electrode was carried out helped by scanning electron microscopy (SEM) and X-Ray energy dispersive spectroscopy (EDS).     

Determination of l-histidine by modified carbon paste electrode using tetra-3,4-pyridinoporphirazinatocopper(II)

This paper describes a potentiometric method for determination of l-histidine (l-his) in aqueous media, using a carbon paste electrode modified with tetra-3,4-pyridinoporphirazinatocopper(II) (Cu (3,4tppa)). The electrode exhibits linear response to the logarithm of the concentration of l-histidine from 2.4 × 10⁻⁵ to 1.0 × 10⁻² M, with a response slope of −49.5 ± 1 mV and response time of about 1.5 min. The detection limit according to IUPAC recommendation was 2.0 × 10⁻⁵ M. The proposed electrode shows a good selectivity for l-his over a wide variety of anions. This chemically modified carbon paste electrode was successfully used for the determination of l-his in a synthetic serum and RANDOX control serum solutions.     

Separation and determination of l-tyrosine and its metabolites by capillary zone electrophoresis with a wall-jet amperometric detection

A method of capillary electrophoresis with wall-jet amperometric detection (AD) has been developed for separation and determination of l-tyrosine (Tyr) and its metabolites, such as Tyramine (TA), p-hydroxyphenylpyruvic (pHPP), homogentisic acid (HGA) and some dipeptides containing Tyr, such as Tyr-Gly-Gly (YGG), Tyr-Arg (YR) and Tyr-d-Arg (Y-d-R). A carbon disk electrode was used as the working electrode and the optimal detection potential was 1.00 V (versus Ag/AgCl). At 18 kV of applied voltage, the seven compounds were completely separated within 20 min in 110 × 10⁻³ mol/L Na₂HPO₄-NaH₂PO₄ buffer (pH 7.10) containing 3 × 10⁻³ mol/L β-cyclodextrin (β-CD). Good linear relationship was obtained for all analytes and the detection limits of seven analytes were in the range of 0.95-4.25 ng/mL. The proposed method has been applied to examine the metabolic process of l-tyrosine in rabbit's urine.     

Electrochemical polymerization of 3,4-ethylenedioxythiophene in aqueous solution containing N-dodecyl-β-d-maltoside

Poly(3,4-ethylenedioxythiophene) (PEDOT) films were synthesized electrochemically by direct anodic oxidation of 3,4-ethylenedioxythiophene (EDOT) at relatively low monomer concentration (0.005-0.01 mol L⁻¹) in aqueous solution containing green nonionic sugar-based surfactant N-dodecyl-β-d-maltoside (DM), which has good biocompatibility and biodegradability. The moderate interaction between DM through the hydroxy groups and EDOT monomer led to the decrease of monomer oxidation onset. Different surfactants, such as anionic sodium dodecylbenzenesulfate (SDBS), nonionic triton X-100 (TX100), were also tested for comparison. As-formed PEDOT films were characterized electrochemically and spectroscopically using FTIR and UV-visible techniques. PEDOT nano-materials with good thermal stability and conductivity of 26.2 S cm⁻¹ can be synthesized in water-DM micellar solution, which can be proved by the results of scanning electron microscopy (SEM). PEDOT prepared from water-DM media with good biocompatibility can be a good candidate for application in biosensors.     

On-line solid phase extraction of Ni and Pb using carbon nanotubes and modified carbon nanotubes coupled to ETAAS

A study about the capabilities of three kinds of nanomaterials namely, carbon nanotubes (CNT), oxidized carbon nanotubes (ox-CNT) and l-alanine immobilized on carbon nanotubes (ala-CNT) to serve as sorbents for preconcentrating Ni and Pb using an on-line system coupled to electrothermal atomic absorption spectroscopy (ETAAS) technique, was accomplished. The solid phase extraction was performed in a conical minicolumn used as sorbent holder. After loading a fixed volume of the analytes, they were eluted with a discrete volume (50 μL) of nitric acid, placed directly into the platform of a L’Vov tube. After that, each analyte was determined individually. Ni and Pb retention was strongly influenced by pH but exhibiting different behaviors. The study demonstrated that the sorbent based on ox-CNTs was the one that exhibited the highest capacity and linearity for both analytes when compared with CNT or ala-CNT. The analytical performance was evaluated for the three sorbents to establish the best conditions regarding sensitivity, reproducibility and accuracy. The precision expressed as relative standard deviations (n = 6) were 3.9 and 0.5% for Ni²⁺ and Pb²⁺, respectively The limit of detection (LOD), calculated as the concentration required to yield a net peak equal to three times the standard deviation of the background signal (3σ) was 30 and 10 ng L⁻¹ for Ni²⁺ and Pb²⁺ respectively. Alternatively, the limit of quantification (10σ) was calculated and resulted to be 0.79 and 0.07 μg L⁻¹ for Ni²⁺ and Pb²⁺ respectively. After optimization, the method that involved the use of ox-CNT associated to an on-line preconcentration was tested in samples of relevant environmental importance. Accuracy was evaluated analyzing a certified reference material namely, Municipal Sludge (QC MUNICIPAL SLUDGE A) and a reference sample of Lake Sediment (TRAP-LRM from IJS).