Fluorimetric determination of nucleic acids with terbium and lutetium

In this paper, fluorescence-enhancement of Tb-nucleic acids [fish sperm DNA (fsDNA) and yeast RNA (yRNA)] by Lu³⁺ is studied in detail and is applied to determine nucleic acids. The experiments indicated that under the optimum conditions, a linear relationship was obtained between the fluorescence intensity (I_f) and the concentration of nucleic acids. The linear range is 1.2×10⁻⁸-1.0×10⁻⁴ g/ml for DNA and 3.0×10⁻⁸-8.0×10⁻⁴ g/ml for RNA. The detection limits (signal/noise=3) for DNA and RNA were 4.8×10⁻⁹ and 7.0×10⁻⁹ g/ml, respectively. The mechanism of the co-luminescence effect is also discussed.     

Flow injection kinetic spectrophotometric method for the determination of trace amounts of nitrite

In this work, a new, simple and sensitive flow injection catalytic kinetic spectrophotometric determination of nitrite is reported based on catalytic effect of nitrite on the redox reaction between sulfonazo III and potassium bromate in acidic media. The reaction was monitored by measuring the decrease in the absorbance of sulfunazo III at 570 nm. Various chemical (such as the effect of acidity, reagents concentrations) and instrumental parameters (flow rate, reaction coil length, injection volume and temperature) were studied and were optimized. Under the optimum conditions calibration graph was linear in the nitrite concentration ranges of 8.00 × 10⁻³-3.00 × 10⁻¹ μg/ml (with slope of 2.40) and 3.50 × 10⁻¹-1.80 μg/ml (with slope of 0.42). The detection limit was 6.00 × 10⁻³ μg/ml of nitrite, the relative standard deviation (n = 10) was 1.25% and 0.88% for 5.00 × 10⁻² and 2.00 × 10⁻¹ μg/ml of nitrite respectively. About 60 samples in 1 h can be analyzed. The interfering effects of various chemical species were studied. The method was successfully applied in the determination of nitrite in food and environmental samples.     

Determination of flurbiprofen in pharmaceutical formulations by UV spectrophotometry and liquid chromatography

In this study, a new and rapid UV spectrophotometric (UV) method and a reversed phase high performance liquid chromatographic (LC) method were developed for quantitative estimation of flurbiprofen, a non-selective, non-steroidal, anti-inflammatory drug (NSAID), in pure form and in pharmaceutical dosage form. The solvent system, wavelength of detection, chromatographic conditions were optimized in order to maximize the sensitivity of both the proposed methods. The linear regression equations obtained by least square regression method were Abs=7.5906×10⁻² concentration (μg/ml) + (−) 4.6210×10⁻² for the UV method, and peak area=1.2652×10² concentration (ng/ml) + 1.4830×10³ for the LC method. The detection limit as per the error propagation theory was found to be 0.34 μg/ml for UV method and 15 ng/ml for LC method. The developed methods were successfully employed with high degree of precision and accuracy for the estimation of total drug content in two commercial ophthalmic drops of flurbiprofen. The results of analysis were treated statistically, as per USP 2000 and International Conference on Harmonization (ICH) guidelines for validation of analytical procedures, and by recovery studies. The results obtained from UV method were comparable with those obtained by using LC. It was concluded that both the developed methods are equally accurate, sensitive, precise, reproducible, robust and rugged and could be applied directly and easily to the pharmaceutical preparations of flurbiprofen. However, LC method is useful at very low level (ng/ml), whereas UV method is suitable at μg/ml level.     

A label-free amperometric immunosenor based on multi-layer assembly of polymerized o-phenylenediamine and gold nanoparticles for determination of Japanese B encephalitis vaccine

A label-free amperometric immunosensor for fast and sensitive assay of Japanese B encephalitis vaccine is presented. Antiserum of Japanese B encephalitis were immobilized on bilayer nano-Au/o-phenylenediamine polymer film with deposited Prussian blue as an electronic mediator on the Pt electrode. The electrochemical behavior of the biosensor was studied with Fe^2+/3+ as probe on the Pt surface using cyclic voltammetry technique. The variation of amperometric response to the concentration of Japanese B encephalitis vaccine, the target antigen, was evaluated by cyclic voltammetry in PBS. The immunosensor showed a specific response to Japanese B encephalitis vaccine in the range 1.1 × 10⁻⁸ to 1.9 × 10⁻⁶ lg pfu/ml (pfu/ml is plaque forming unit and lg is common logarithm) with a detection limit of 6 × 10⁻⁹ lg pfu/ml. The correlation coefficient is 0.9955. The incubation time, incubation temperature, pH, reproducibility and stability of the immunosensor were also studied. The present work supplied a promising test method for biological products.     

Adsorptive stripping voltammetric determination of trace concentrations of molybdenum at an in situ plated lead film electrode

An in situ plated lead film electrode has been applied for adsorptive stripping voltammetric determination of trace concentrations of molybdenum in the presence of Alizarin S. The procedure is based on the preconcentration of the molybdenum-Alizarin S complex at an in situ plated lead film electrode held at −0.6 V (versus Ag/AgCl), followed by a negatively sweeping square wave voltammetric scan. The peak current is proportional to the concentration of molybdenum over the range 2 × 10⁻⁹ to 5 × 10⁻⁸ mol L⁻¹, with a 3σ detection limit of 9 × 10⁻¹⁰ mol L⁻¹ with an accumulation time of 60 s. The measurements were carried out from underaerated solutions. The proposed procedure was validated in the course of Mo(VI) determination in water certified reference materials.     

Study and application of parallel catalytic hydrogen wave of lincomycin in the presence of persulfate

The polarographic currents of lincomycin in the absence and the presence of persulfate are studied by linear potential scan polarography and cyclic voltammetry. The reduction wave of lincomycin in phosphate buffer is a catalytic hydrogen wave, which is the reduction of the proton combined with lincomycin in nature. When S₂O₈²⁻ is present, the atomic hydrogen as intermediate product from the reduction of the combined proton is oxidized by both S₂O₈²⁻ and its reduction intermediate, sulfate radical anion SO₄⁻, to regenerate the original proton, producing the parallel catalytic hydrogen wave. Based on the parallel catalytic hydrogen wave, a novel method for the determination of lincomycin is proposed. In 0.48 mol l⁻¹ KH₂PO₄-Na₂HPO₄ (pH 7.4)-8.0×10⁻³ mol l⁻¹ K₂S₂O₈ supporting electrolyte, the peak potential of the parallel catalytic hydrogen wave is −1.82 V (vs. SCE). The second-order derivative peak current is rectilinear to the lincomycin concentration in the range of 8.5×10⁻⁸-9.0×10⁻⁵ mol l⁻¹ and the detection limit is 4×10⁻⁸ mol l⁻¹. The parallel catalytic hydrogen wave is three orders in magnitude higher than that of the corresponding catalytic hydrogen wave in analytical sensitivity. The proposed method is applied to the rapid determination of lincomycin hydrochloride in eye drops without previous separation.     

Simultaneous determination of zirconium and hafnium as ternary complexes with 5-Br-PADAP and fluoride using solid-phase extraction and reversed-phase liquid chromatography

Solid-phase extraction (SPE) along with reversed-phase liquid chromatography (RP-LC) was used for the simultaneous determination of Zr(IV) and Hf(IV) by means of their ternary chelates with fluoride and 2-(5-bromo-2-pyridylazo)-5-diethylaminophenol (5-Br-PADAP). The conditions of SPE sorption were examined in detail: type of SPE column, volume of the sample, volume of the eluent, concentrations of metal ions, fluoride salt, chromogenic reagent, organic phase, and pH. It was established that the sorption of Zr(IV) and Hf(IV), as their ternary chelates, on SPE Zorbax SPE C18 (EC) cartridge was the most efficient, when the sample containing metal ion (Zr(IV), Hf(IV), both, up to 2 μg), 5-Br-PADAP 1.5×10⁻⁴, NaF 7.5×10⁻⁵ mol l⁻¹, methanol 40%, pH 4.5±1 was applied for the SPE sorption. The chelates were discarded from SPE cartridge using acetonitrile/water (99.75+0.25, v/v) eluent containing 3.8×10⁻⁴ mol l⁻¹ sodium fluoride and subsequently separated by RP-LC method. The RP-LC separation of both chelates was optimized and Zorbax SB-C18 analytical LC column along with acetonitrile/water (65+35, v/v) eluent containing the 1.5×10⁻⁴ mol l⁻¹ sodium fluoride was used. The established SPE/LC conditions allow Zr(IV) 0.08-2.0 μg and Hf(IV) 0.04-2.0 μg determination in a sample volume up to 150 ml. The detection limits, 0.03 μg Hf(IV) and 0.05 μg Zr(IV), were obtained. Recoveries, (94±2)% for Hf(IV) chelate and (106±2)% for Zr(IV) chelate were obtained, when 1 μg of Zr(IV) and Hf(IV) ions were determined by the present SPE/LC method from the sample volume of 100 ml. The established, pre-concentration SPE conditions, along with the LC separation and determination allow the assay of Zr(IV) and Hf(IV) in complicated matrix materials. The present SPE/LC method was applied to the determination of Zr(IV) and Hf(IV) in tap water and reference geological material (rock, NCS DC 73303; certified content: Zr, 27.7×10⁻³% (w/w) and Hf, 6.5×10⁻⁴% (w/w)).     

New triiodomercurate-modified carbon paste electrode for the potentiometric determination of mercury

A new tetrazolium-triiodomercurate-modified carbon paste electrode has been described for the sensitive and selective determination of mercury. The electrode shows a stable, near-Nernstian response for 1×10⁻³ to 6×10⁻⁶ M [HgI₃]⁻ at 25 °C over the pH range of 4.0-9.0, with an anionic slope of 55.5±0.4 mV. The lower detection limit is 4×10⁻⁶ M with a fast response time of 30-50 s. Selectivity coefficients of a number of interfering anions and iodo complexes of some metal ions have been estimated. The interference from many of the investigated ions is negligible. The determination of 1-200 μg/ml of mercury in aqueous solutions shows an average recovery of 98.5% and a mean relative standard deviation of 1.6% at 50.0 μg/ml. The direct determination of mercury in spiked wastewater, metal amalgams and dental alloy gave results that compare favorably with those obtained by the cold vapor atomic absorption spectrometric method. Potentiometric titration of mercury and phenylmercury acetate with standard potassium iodide has been monitored using the developed triiodomercurate-carbon paste electrode (CPE) as an end point indicator electrode.     

A novel method for determination of magnesium in urine and water samples with mercury film-plated carbon paste electrode

A square wave adsorptive stripping voltammetric (SWAdSV) method for the indirect determination of trace amounts of magnesium with thiopentone sodium (TPS) as an electroactive ligand, at carbon paste mercury film electrode (CP-MFE) is proposed. It is observed that the increase of the square wave voltammetric cathodic peak current of TPS, under alkaline conditions, is linear with the increase of Mg concentration. Under optimum experimental conditions viz.; pH 10.75, 3×10⁻⁵ M TPS and 0.05 M phosphate buffer (Na₂HPO₄-NaH₂PO₄), a linear relation in the range 6×10⁻⁹ to 9×10⁻⁸ M Mg²⁺ (0.14-2.16 ppb), at 60 s deposition time, is obtained. The detection limit of Mg²⁺ is 0.14 ppb for 60 s deposition time with the relative standard deviation is 0.5% (n=5). The proposed method was successfully applied to the determination of magnesium in urine and tap water samples with satisfactory results. The data obtained are compared with the standard flame atomic absorption spectrophotometric method (FAAS).     

Fluorimetric determination of nucleic acid using the enhancement of terbium-gadolinium-nucleic acid-cetylpyridine bromide system

It is found that the fluorescence intensity of Tb-cetylpyridine bromide (CPB)-nucleic acid system can be enhanced by La³⁺, Gd³⁺, Lu³⁺, Sc³⁺ and Y³⁺, of which Gd³⁺ has the greatest enhancement. The experiments indicate that under the optimum condition, the fluorescence intensity of the system is in proportion to the concentration of nucleic acids in the range from 9×10⁻⁸ to 1×10⁻⁵ g ml⁻¹ for yeast RNA, from 1×10⁻⁷ to 1×10⁻⁵ g ml⁻¹ for fish sperm DNA. The detection limits are 3.2 and 4.1 ng ml⁻¹, respectively. This method has been used satisfactorily for the determination of both synthetic and actual samples. In comparison with most fluorescence method for the determination of nucleic acids, this method has higher sensitivity and stability.     

Comparison of pyrimethanil-imprinted beads and bulk polymer as stationary phase by non-linear chromatography

A pyrimethanil-imprinted polymer (P1) was prepared by iniferter-mediated photografting a mixture of methacrylic acid and ethylene dimethacrylate onto homemade near-monodispersed chloromethylated polydivinylbenzene beads. The chromatographic behaviour of a column packed with these imprinted beads was compared with another column packed with irregular particles obtained by grinding a bulk pyrimethanil-imprinted polymer (P2). The comparison was made using the kinetic model of non-linear chromatography, studying the elution of the template and of two related substances, cyprodinil and mepanipyrim. Extension of the region of linearity, capacity factors for the template and the related substances, column selectivity, binding site heterogeneity, apparent affinity constant (K) and lumped kinetic association (k_a) and dissociation rate constant (k_d) were studied during a large interval of solute concentration, ranging between 1 and 2000 μg/ml. From the experimental results obtained, in the linearity region of solute concentration column selectivity and binding site heterogeneity remained essentially the same for the two columns, while column capacity (at 20 μg/ml, P1 = 23.1, P2 = 11.5), K (at 20 μg/ml, P1 = 8.3 × 10⁶ M⁻¹, P2 = 2.5 × 10⁶ M⁻¹) and k_a (at 20 μg/ml, P1 = 3.5 μM⁻¹ s⁻¹, P2 = 0.47 μM⁻¹ s⁻¹) significantly increased and k_d (at 20 μg/ml, P1 = 0.42 s⁻¹, P2 = 0.67 s⁻¹) decreased for the column packed with the imprinted beads. These results are consistent with an influence of the polymerisation method on the morphology of the resulting polymer and not on the molecular recognition properties due to the molecular imprinting process.     

Study of the electrochemical behavior of isorhamnetin on a glassy carbon electrode and its application

The electrochemical behavior of isorhamnetin (ISO) at a glassy carbon electrode was studied in a phosphate buffer solution (PBS) of pH 4.0 by cyclic voltammetry (CV) and differential pulse voltammetric method (DPV). A well-defined redox wave of ISO involving one electrons and one proton appeared. The electrode reaction is a reactant weak adsorption-controlled process with a charge transfer coefficient (α) of 0.586. Based on the understanding of the electrochemical process of ISO at the glassy carbon electrode, analysis of ISO can be realized. Under optimal conditions, the oxidation peak current showed linear dependence on the concentration of ISO in the range of 1.0 × 10⁻⁸ to 4.0 × 10⁻⁷ M and 1.0 × 10⁻⁶ to 1.0 × 10⁻⁵ M. The detection limit is 5.0 × 10⁻⁹ M. This method has been successfully applied to the detection of ISO in tablets.     

Rapid simultaneous analysis of 1-hydroxypyrene, 2-hydroxyfluorene, 9-hydroxyphenanthrene, 1- and 2-naphthol in urine by first derivative synchronous fluorescence spectrometry using Tween-20 as a sensitizer

A novel method of first derivative synchronous fluorescence was developed for the rapid simultaneous analysis of trace 1-hydroxypyrene (1-OHP), 1-naphthol (1-NAP), 2-naphthol (2-NAP), 9-hydroxyphenanthrene (9-OHPe) and 2-hydroxyfluorene (2-OHFlu) in human urine. Only one single scan was needed for quantitative determination of five compounds simultaneously when Δλ = 10 nm was chosen. In the optimal experimental conditions, there was a linear relationship between the fluorescence intensity and the concentration of 1-OHP, 1-NAP, 2-NAP, 9-OHPe and 2-OHFlu in the range of 1.75 × 10⁻⁹ to 4.50 × 10⁻⁶ mol L⁻¹, 3.64 × 10⁻⁸ to 2.20 × 10⁻⁴ mol L⁻¹, 8.18 × 10⁻⁹ to 1.20 × 10⁻⁴ mol L⁻¹, 3.26 × 10⁻⁹ to 8.50 × 10⁻⁵ mol L⁻¹ and 4.88 × 10⁻⁹ to 5.50 × 10⁻⁶ mol L⁻¹, respectively. The limits of detection (LOD) were found to be 5.25 × 10⁻¹⁰ mol L⁻¹ for 1-OHP, 1.10 × 10⁻⁸ mol L⁻¹ for 1-NAP, 2.46 × 10⁻⁹ mol L⁻¹ for 2-NAP, 9.77 × 10⁻¹⁰ mol L⁻¹ for 9-OHPe and 1.46 × 10⁻⁹ mol L⁻¹ for 2-OHFlu. The proposed method is reliable, selective and sensitive, and has been used successfully in the determination of traces of 1-OHP, 1-NAP, 2-NAP, 9-OHPe and 2-OHFlu in human urine samples, whose results were in good agreement with those gained by the HPLC method.     

Urea as new stabilizing agent for imipenem determination: electrochemical study and determination of imipenem and its primary metabolite in human urine

Imipenem shows a fast chemical conversion to a more stable imin form (identical to that of biochemical dehydropeptidase degradation) in aqueous solutions and stabilizing agents used avoid its electrochemical study and determination.The aim of this work is the proposal of urea as stabilizing agent which allows the electrochemical study of imipenem and the proposal of electrochemical methods for the determination of imipenem and its primary metabolite (M1) in human urine samples. Electrochemical studies were realized in phosphate buffer solutions over pH range 1.5-8.0 using differential-pulse polarography, DC-tast polarography, cyclic voltammetry and adsorptive stripping voltammetry. In acidic media, a non-reversible diffusion-controlled reduction involving a two steps mechanism which involves one electron and one proton in the first step and two electrons and two protons in the second step occurs and the mechanism for the reduction was suggested.A differential-pulse polarographic method for the determination of imipenem in the concentration range 3.2 × 10⁻⁶ to 2 × 10⁻⁵ M (0.95-3.4 mg/L) and its primary metabolite in the concentration range 1.4 × 10⁻⁶ to 10⁻⁴ M (0.43-26.1 mg/L) with detection limits of 9.6 × 10⁻⁷ M (0.28 μg/L imipenem) and 4.3 × 10⁻⁷ M (0.14 μg/L M1) was proposed. Also, a method based on controlled adsorptive pre-concentration of imipenem on the hanging mercury drop electrode followed by voltammetric measure, allows imipenem determination in the concentration range 1.8 × 10⁻⁸ to 1.2 × 10⁻⁶ M (5.42-347 μg/L) with a detection limit of 5.4 × 10⁻⁹ M (1.63 μg/L). The proposed methods have been used for the direct determination of the analytes in a pharmaceutical formulation and human urine.     

Development of a flow manifold for Fe(III) ion determination with a fluoride ion-selective electrode

Continuous-flow (CF) and flow-injection (FI) analysis using the fluoride ion-selective electrode (FISE) as detector have been investigated. The measurements were performed in a home-made cell under appropriate flow conditions (2.86 or 3.45 ml min⁻¹, 0.2 ml samples, 10⁻⁶ M sodium fluoride). The calibration graph was obtained by plotting the signal height versus concentration of iron in the range of Fe(III) concentration from 10⁻⁵ to 10⁻¹ M in acetate buffer (pH 2.8 or 3.4). In all described procedures, the range of linear response extends to the Fe(III) concentration from 1×10⁻³ to 1×10⁻¹ M, with detection limit 9×10⁻⁵ M. The effect of double-line, two-line flow manifold and CF was investigated and discussed.     

Study on the colour reaction of Te-KI-RhB system in the presence of PVA-OP

A new supersensitive spectrophotometric method for the determination of tellurium (IV) has been developed which is based on the formation of an ion-association complex with potassium iodide and rhodamine B (RhB) in the presence of polyvinyl alcohol (PVA) and polyethylene glycol octyl phenyl ether (OP). The molar absorptivity at 560 nm is 2.8 × 10⁶ 1 mol^–1 cm^–1. Beer's law is obeyed over the range of 20–70 ng/ml Te. The method has been applied successfully to determine tellurium in alloy, steel and some other samples.     

Highly sensitive detection of silybin based on adsorptive stripping analysis at single-sided heated screen-printed carbon electrodes modified with multi-walled carbon nanotubes with direct current heating

A new disposable multi-walled carbon nanotubes modified single-sided heated screen-printed carbon electrode (MWNT/ss-HSPCE) was fabricated. The electrochemical behavior of silybin was investigated by cyclic voltammetry and the probable electrode reaction mechanism was proposed. A simple and cheap direct current heating supplier was used to heating the electrode for adsorptive accumulation of silybin. The square wave voltammetric stripping peak current of silybin at MWNT/ss-HSPCE with an elevated electrode temperature of 50 °C only during accumulation step was dramatically improved compared with that at bare single-sided heated screen-printed carbon electrode (ss-HSPCE) without heating. This enhancement was mainly contributed to the combination of the advantages of multi-walled carbon nanotubes and electrically heated electrodes. Under optimum conditions, two detection linear ranges of silybin were from 1.0 × 10⁻⁹ to 1.0 × 10⁻⁷ M and 3.0 × 10⁻⁷ to 1.0 × 10⁻⁶ M. A detection limit of 5.0 × 10⁻¹⁰ M could be obtained (S/N = 3), which was more than two magnitudes lower than that at bare ss-HSPCE without heating. To the best of our knowledge, this was also at least two magnitudes lower than any others for electrochemical detection of silybin in the literature. Finally, the method was successfully applied to the determination of silybin in pharmaceutical tablets.     

Flow chemiluminescence sensor for determination of clenbuterol based on molecularly imprinted polymer

In this paper, a novel flow chemiluminescence (CL) clenbuterol sensor based on molecularly imprinted polymer (MIP) on line enrichment nanogram clenbuterol and chemiluminescence reaction of potassium permanganate and formaldehyde in the polyphosphate enhanced by clenbuterol. Clenbuterol in the urine was selectively adsorbed on the clenbuterol-imprinted polymer, which was packed into the flow cell. The formaldehyde and the polyphosphate with potassium permanganate flowed through the flow cell and reacted with the on line adsorbed clenbuterol and produced strong CL. The results show that the sensor was reversible. The CL intensity was linear with clenbuterol concentration from 1.0 × 10⁻⁹ g/mL to 5.0 × 10⁻⁸ g/mL. The detection limit was 3.0 × 10⁻¹⁰ g/mL. The R.S.D. for ng/mL clenbuterol was less than 5% (n = 3). The present method offered a high selectivity and sensitivity that made the quantitative analysis of trace clenbuterol (ng/mL) in the animal urine sample.     

Voltammetric behavior of methaqualone and its determination by single-sweep oscillopolarography

A well derivative reduction wave of methaqualonc (MTQ) was obtained in 0.033 mol dm⁻³ B-R (pH 3.76) buffer solution by single-sweep oscillopolarography. The peak potential is −1.36 V (vs. saturated calomel reference electrode, SCE). The peak current is proportional to the concentration of methaqualone over the range of 7.0×10⁻⁸-9.0×10⁻⁶ mol dm⁻³. The method has been successfully applied to the determination of methaqualone in tablets. The wave is believed to result from irreversible adsorption through studies of its electrochemical behavior and reaction mechanism.     

Application of 2-mercaptobenzothiazole self-assembled monolayer on polycrystalline gold electrode as a nanosensor for determination of Ag(I)

Fabrication and application of a voltammetric sensor based on gold 2-mercaptobenzothiazole self-assembled monolayer (Au-MBT SAM) for determination of silver ion is described. Preliminary experiments were performed to characterize the monolayer. The surface pK_a determined for the MBT monolayer is 7.0. This value was obtained by impedimetric titration of the monolayer in the presence of Fe(CN)₆^3−/4− as a redox probe. The extent of surface coverage was evaluated as 1.52 × 10⁻⁹ mol cm⁻² based on charged consumed for reductive desorption of the monolayer in the 0.50 M NaOH solution. Then the sensor was used for determination of Ag(I) by square wave voltammetry. The parameters affecting the sensor response, such as pH and supporting electrolyte, were optimized. A dynamic calibration curve with two linear parts was obtained in the concentration ranges of 5 × 10⁻⁸-8 × 10⁻⁷ and 1 × 10⁻⁶-1 × 10⁻⁵ M of Ag(I). The detection limit adopted from cathodic striping square wave voltammetry was as 1 × 10⁻⁸ M for n = 7. Furthermore, the effect of potential interfering ions on the determination of Ag(I) was studied, and an appropriate method was used for the elimination of this effect.