A validated stripping voltammetric procedure for quantification of the anti-hypertensive and benign prostatic hyperplasia drug terazosin in tablets and human serum

The electrochemical behavior of terazosin at the hanging mercury drop electrode was studied in Britton-Robinson buffer (pH 2-11), acetate buffer (4.5-5.5), and in 0.1 M solution of each of sodium sulfate, sodium nitrate, sodium perchlorate and potassium chloride as supporting electrolytes. The square-wave adsorptive cathodic stripping voltammogram of terazosin exhibited a single well-defined two-electron irreversible cathodic peak which may be attributed to the reduction of CO double bond of the drug molecule. A fully validated, simple, high sensitive, precise and inexpensive square-wave adsorptive cathodic stripping voltammetric procedure was described for determination of terazosin in bulk form, tablets and human serum. A mean recovery for 1×10⁻⁸ M terazosin in bulk form, following preconcentration onto the hanging mercury drop electrode for 60 s at a −1.0 V (versus Ag/AgCl/KCl_s), of 99±0.7% (n=5) was obtained. Limits of detection (LOD) and quantitation (LOQ) of 1.5×10⁻¹¹ and 5×10⁻¹¹ M bulk terazosin were achieved, respectively. The proposed procedure was successfully applied to determination of the drug in its Itrin^® tablets and human serum samples. The achieved LOD and LOQ of the drug in human serum samples were 5.3×10⁻¹¹ and 1.8×10⁻¹⁰ M THD, respectively. The pharmacokinetic parameters of the drug in human plasma were estimated as: C_max=77.5 ng ml⁻¹, t_max=1.75 h, AUC_0-t=602.3 ng h ml⁻¹, K_e=0.088 h⁻¹ and t_1/2=11.32 h) which are favorably compared with those reported in literature.     

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.     

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).     

Quantitative determination of zinc in milkvetch by anodic stripping voltammetry with bismuth film electrodes

Bismuth film electrode (BiFE) was shown to be an attractive alternative to common mercury film electrode (MFE) for anodic stripping voltammetric measurements. In this study, bismuth film, that was in situ deposited onto glassy carbon electrode, was used to detect zinc content of milkvetch, used in traditional Chinese medicine. Variables affecting the response have been evaluated and optimized. Experimental results showed a high response, with a good linearity (between 0.5 × 10⁻⁶ mol L⁻¹ and 3 × 10⁻⁶ mol L⁻¹) a good precision (R.S.D. = 3.58%) and a low detection limit (9.6 × 10⁻⁹ mol L⁻¹ with a 120 s anodic). The anodic stripping performance makes the bismuth film electrode very desirable for measurements of trace nutritive element zinc in milkvetch and should impart possible restrictions on the use of mercury electrode.     

Square wave adsorptive stripping voltametric determination of the mixture of nalidixic acid and its main metabolite (7-hydroxymethylnalidixic acid) by multivariate methods and artificial neural network

Nalidixic acid (NA) and its main metabolite, 7-hydroximethylnalidixic acid (OHNA), are quinolones antibacterial used as agents used for the treatment of urinary tract infection. For both compounds an adsorption process on a hanging mercury electrode (HMDE). On this basis, a square wave adsorptive stripping voltammetry (SWadSV) method has been developed for the individual and simultaneous determination of NA and OHNA. The variables that affect to accumulation process, such as concentration of perchloric acid, accumulation potential and accumulation time have been optimised by using an experimental design (concretely a Box-Behnken design with three levels) together with the response surface methodology (RSM). Calibration curves were linear in the range (0-1.38) × 10⁻⁷ mol L⁻¹ for NA and (0-3.23) × 10⁻⁸ mol L⁻¹ for OHNA, in the optimized conditions, with detection limits of 9.48 × 10⁻⁹ mol L⁻¹ and 8.06 × 10⁻¹⁰ mol L⁻¹ for NA and OHNA, respectively. The method was applied to urine samples containing only one of the analytes with satisfactory recoveries. As the voltammetric signals of these compounds show a high overlapping, different chemometric methods, such as classical least squares (CLS), partial least squares (PLS), principal component regression (PCR) and artificial neural network (ANN) have been used for the resolution of the mixture. The analysis of these compounds in urine samples were carried out using the different chemometric tools and the best recoveries were obtained by using ANN. No pre-treatment of the sample was necessary.     

Voltammetric determination of Imatinib (Gleevec) and its main metabolite using square-wave and adsorptive stripping square-wave techniques in urine samples

The voltammetric behaviour of Imatinib (STI 571) and its main metabolite (N-demethylated piperazine derivative) were studied by square-wave techniques, resulting in to two methods for their determination in aqueous and urine samples at pH 2. The application of the square-wave (SW) without the adsorptive accumulation and voltammetric stripping (AdSV) exhibit a peak at a reduction potential of −0.70 V for an accumulation potential of −0.45 V. The sensitivity was higher for the stripping technique because a signal four times higher than that provided by the square-wave method without the previous accumulation was obtained. Due to the fact that Imatinib and its metabolite show the same voltammetric reduction process, some experiments were performed in order to compare the voltammetric response of Imatinib and its main metabolite in a similar ratio than that of the therapeutic concentration. The calibration curve for Imatinib in urine was linear in the range from 1.9 × 10⁻⁸ to 1.9 × 10⁻⁶ M in stripping mode with an accumulation time (t_acc) of 10 s. The relative standard deviations obtained for concentration levels of Imatinib as low as 2.0 × 10⁻⁷ M for square-wave was 2.17% (n = 9) and for stripping square-wave was 2.65% (n = 9) in the same day. The limits of detection for square-wave and stripping square-wave were 5.55 × 10⁻⁹ and 5.19 × 10⁻⁹ M, respectively. Thus, the presented method are straightforward, rapid and sensitive and has been applied to the determination of Imatinib and its main metabolite altogether in urine samples from real patients.     

Adsorptive stripping voltammetric determination of the anti-inflammatory drug celecoxib in pharmaceutical formulation and human serum

Celecoxib is a cyclooxygenase inhibitor, that has been recently and intensively prescribed as an anti-inflammatory drug in rheumatic osteoarthiritis. A robust, highly reliable and reproducible square-wave (SW) adsorptive cathodic stripping voltammetric procedure was developed for the determination of celecoxib in pharmaceutical formulation and human serum. The analytical procedure was based on the reduction of the CN of the pyrazole ring of the drug molecule at the mercury electrode surface in Britton-Robinson buffer of pH 7.0. The SW adsorptive cathodic stripping voltammogram of celecoxib showed a single well-defined peak at −1.54 V (vs. Ag/AgCl/KCl_s) using an accumulation potential of −0.70 V, frequency of 120 Hz, scan increment of 10 mV and pulse amplitude of 25 mV. Repeatability was examined for 1×10⁻⁸ M CXB drug solution after 30 s pre-concentration and a mean recovery of 99.4±0.4% (n=5) was achieved. For 90 s preconcentration, a linear concentration range of 1×10⁻⁹-2×10⁻⁸ M CXB and a detection limit of 1.86×10⁻¹⁰ M were achieved. The proposed procedure was successfully applied for the determination of the drug in capsules and human serum with mean recoveries of 101.5±0.6 and 98.8±1.1%, respectively. A detection and quantitation limits of 1.0×10⁻⁹ M (0.4 ng ml⁻¹) and 4.7×10⁻⁹ M (1.3 ng ml⁻¹) were achieved for the determination of the drug in human serum. Moreover, the procedure was useful for study of the pharmacokinetic profile of celecoxib in a healthy volunteer after administration of a single oral dose (celebrex®, 200 mg).     

Stripping voltammetric determination of silymarin in formulations and human blood utilizing bare and modified carbon paste electrodes

Silymarin is one of the most powerful natural substances that have the ability to protect and rebuild the liver cells damaged by alcohol and other toxic substances. Silymarin showed two irreversible anodic peaks in buffered solutions (pH 2.5-8.0) at either the bare carbon paste electrode or the montmorillonite-Ca modified carbon paste one. These two peaks have been attributed to oxidation of the two phenolic OH groups at positions C-20 and C-7 of silymarin molecule. A square-wave adsorptive anodic stripping voltammetry method was optimized for determination of silymarin utilizing the bare and the modified carbon paste electrodes. The method was fully validated and successfully applied for the determination of silymarin in commercial formulations and human serum without prior extraction utilizing both carbon paste electrodes. Limits of quantitation of 1 × 10⁻⁷ and 7 × 10⁻⁹ mol L⁻¹ silymarin have been achieved in bulk form or in formulations while 2 × 10⁻⁷ and 8 × 10⁻⁹ mol L⁻¹ silymarin were achieved in spiked human serum utilizing the bare carbon paste electrode and the modified one, respectively. The two electrodes exhibited excellent selectivity towards silymarin even in the presence of 10²to 10³-fold excess of its co-formulated drugs, common excipients, and common metal ions. The pharmacokinetic parameters of silymarin in plasma of healthy human volunteers were estimated following the administration of a single oral dose of 120 mg silymarin utilizing the modified carbon paste electrode. The estimated pharmacokinetic parameters were favorably compared with those reported in literature.     

An efficient approach to designing and optimizing the analysis of Ni(II) by AdCSV in seawater

A highly sensitive voltammetric method was developed for the determination of nickel in seawater at nanomolar concentrations. The measurement is based on the differential pulse cathodic adsorptive stripping of Ni(II) complexed with pyridoxal salicyloylhydrazone at a hanging mercury drop electrode. Optimal conditions were found following a two-step study strategy based on a Plackett Burman design and subsequently a modified simplex method. They were: deposition potential −0.8 V; deposition time 120 s; differential pulse scan mode; pulse amplitude −0.07 V; pulse time 0.04 s; voltage step 0.017 V; time interval for voltage step 0.05 s; supporting electrolyte ammonium chloride/ammonia (0.08 M, pH = 8.9) and concentration of PSH 5.32 × 10⁻⁶ M. The response of the system was found to be linear in a range of Ni concentrations from 0 to 306.7 × 10⁻⁹ M. The detection limit was found to be 0.04 × 10⁻⁹ M of Ni(II). The precision of the method was 1.4% for 3.4 × 10⁻⁸ M of Ni(II) and 1.48% for the blank at a significance level of 95% (n = 9). The method was free from interferences of inorganic salts and trace metals at usual concentrations in seawater. The application to seawater was demonstrated by analysis of CRM 505 and LGC 6016 certified reference estuarine water and real seawater samples from Tangier Bay (Morocco).     

Cu nanoparticles incorporated polypyrrole modified GCE for sensitive simultaneous determination of dopamine and uric acid

Cu nanoparticles have been electrochemically incorporated polypyrrole film that was used for modification of the glassy carbon electrode surface. The performance of the electrode has been characterized by cyclic voltammetry and atomic force microscopy. The electrode has shown high electrocatalytic activity towards the oxidation of dopamine (DA) and uric acid (UA) simultaneously in a phosphate buffer solution (pH 7.00). The electrocatalytic oxidation currents of UA and DA were found linearly related to concentration over the range 1 × 10⁻⁹ to 1 × 10⁻⁵ M for UA and 1 × 10⁻⁹ to 1 × 10⁻⁷ M for DA using DPVs method. The detection limits were determined as 8 × 10⁻¹⁰ M (s/n = 3) for UA and 8.5 × 10⁻¹⁰ M (s/n = 3) for DA at a signal-to-noise ratio of 3.     

Potentiometric stripping analysis of methyl and ethyl parathion employing carbon nanoparticles and halloysite nanoclay modified carbon paste electrode

Carbon nanoparticles (CNPs) and halloysite nanoclay (HNC) modified carbon paste electrode (HNC–CNP–CPE) was developed for the determination of methyl parathion (MP) and ethyl parathion (EP). The electrochemical behavior of these molecules was investigated employing cyclic voltammetry (CV), chronocoulometry (CC), electrochemical impedance spectroscopy (EIS) and potentiometric stripping analysis (PSA). After optimization of analytical conditions employing this electrode at pH 5.0 in acetate buffer (0.1 M), the peak currents were found to vary linearly with its concentration in the range of 1.55 × 10⁻⁹ to 3.67 × 10⁻⁶ M and 1.21 × 10⁻⁹ to 4.92 × 10⁻⁶ M for MP and EP, respectively. The detection limits (S/N = 3) of 4.70 × 10⁻¹⁰ M and 3.67 × 10⁻¹⁰ M were obtained for MP and EP, respectively, using PSA. The prepared modified electrode showed several advantages such as simple preparation method, high sensitivity, very low detection limits and excellent reproducibility. The proposed method was employed for the determination of MP and EP in fruits, vegetables, water and soil samples.     

The versatility of salicylaldehyde thiosemicarbazone in the determination of copper in blood using adsorptive stripping voltammetry

The adsorptive cathodic stripping voltammetry technique (AdCSV) is used to determine copper(II) using salicylaldehyde thiosemicarbazone (N, S- donor) as a complexing agent on hanging mercury drop electrode at pH 9.3. Variable factors affecting the response, i.e. the concentration of ligand, pH, adsorption potential and adsorption time are assessed and optimized. The adsorbed complex of copper(II) and salicylaldehyde thiosemicarbazone gives a well defined cathodic stripping peak current at −0.35 V, which has been used for the determination of copper in the concentration range of 7.85 × 10⁻⁹ to 8.00 × 10⁻⁶ M with accumulation time of 360 s at −0.1 V versus Ag/AgCl. This technique has been applied for the determination of copper in various digested samples of whole blood at trace levels.     

Electrooxidation of the antiviral drug valacyclovir and its square-wave and differential pulse voltammetric determination in pharmaceuticals and human biological fluids

The electrochemical properties of valacyclovir, an antiviral drug, were investigated in pH range 1.8-12.0 by cyclic, differential pulse and square-wave voltammetry. The drug was irreversibly oxidized at a glassy carbon electrode in one or two oxidation steps, which are pH-dependent. For analytical purposes, a very resolved diffusion controlled voltammetric peak was obtained in Britton-Robinson buffer at pH 10.0 using differential pulse and square-wave modes. Limits of detection were 1.04 × 10⁻⁷ and 4.60 × 10⁻⁸ M for differential pulse and square-wave voltammetry, respectively. The applicability to direct assays of tablets, spiked human serum and simulated gastric fluid, was described.     

Voltammetry and determination of metronidazole at a carbon fiber microdisk electrode

The electrochemistry of metronidazole, 1-(hydroxyethyl)-2-methyl-5-nitroimidazole, was investigated at a carbon fiber microdisk electrode in pH 9 Britton Robinson buffer. Under these conditions, the reduction of metronidazole is controlled by both mass transport to the microdisk and adsorption with an equilibrium constant of 4 × 10³ mol⁻¹ dm³ and a saturation coverage of 0.88 × 10⁻⁸ mol cm⁻². The adsorption and accumulation of metronidazole on the surface of the carbon fiber allows its determination at low concentrations by square wave adsorptive stripping voltammetry. A detection limit for metronidazole of 5 × 10⁻⁷ mol dm⁻³ and a R.S.D. of 3.7% at 1 × 10⁻⁶ mol dm⁻³ (n = 4) were obtained with a two electrode system with no stirring during the accumulation step. Based on this method, a simple procedure for the determination of metronidazole in urine is described which requires no pre-treatment of the sample before analysis.     

Square wave voltammetric determination of nitrofurantoin in pharmaceutical formulations on highly boron-doped diamond electrodes at different boron-doping contents

The influence of the boron-doping levels in boron-doped diamond film electrodes on the electrochemical response of nitrofurantoin (NFT) and the development of an electroanalytical procedure for NFT determination were investigated. The investigations were carried out using the techniques of cyclic voltammetry and square wave voltammetry on diamond film electrodes with different boron-doping levels (i.e., 5000, 10,000 and 20,000 mg L⁻¹). The level of boron-doping in the diamond film electrodes influenced the electrochemical reduction of NFT. The appropriate cyclic voltammetric response of NFT was obtained with Britton-Robinson buffer at pH 4 and for diamond films doped with 10,000 and 20,000 mg L⁻¹ of boron. These two films were selected for the development of the electroanalytical procedure. The use of square wave voltammetry with the optimized parameters demonstrated a good linear relationship between the peak current and the NFT concentration for a wide range of concentration. The lower limit of detection for the electrodes doped with 10,000 and 20,000 mg L⁻¹ of boron were 2.69 × 10⁻⁸ mol L⁻¹ (6.40 μg L⁻¹) and 8.15 × 10⁻⁹ mol L⁻¹ (1.94 μg L⁻¹), respectively, while the lower limits of quantification were 8.96 × 10⁻⁸ mol L⁻¹ (21.33 μg L⁻¹) and 2.72 × 10⁻⁸ mol L⁻¹ (6.47 μg L⁻¹), respectively. The applicability of the proposed procedure was tested using a commercial pharmaceutical formulation of NFT, and the results were compared with the procedure recommended by the British Pharmacopeia. The proposed procedure was sensitive, accurate and precise for analysis of NFT and did not require complex preparations or renovations of the electrode surface. This presents the advantage of eliminating mercury waste and minimizing the adsorptive problems related to the use of other electrodic solid surfaces.     

Indirect determination of cyanide ion and hydrogen cyanide by adsorptive stripping voltammetry at a mercury electrode

An indirect voltammetric method is described for determination of cyanide ions and hydrogen cyanide, using the effect of cyanide on cathodic adsorptive stripping peak height of Cu-adenine. The method is based on competitive Cu complex formation reaction between adenine at the electrode surface and CN⁻ ions in solution. Under the optimum experimental conditions (pH=6.42 Britton-Robinson buffer, 1×10⁻⁴ M copper and 8×10⁻⁷ M adenine), the linear decrease of the peak current of Cu-adenine was observed, when the cyanide concentration was increased from 5×10⁻⁸ to 8×10⁻⁷ M. The detection limit was obtained as 1×10⁻⁸ M for 60 s accumulation time. The relative standard deviations for six measurements were 4 and 2% for the cyanide concentrations of 5×10⁻⁸ and 2×10⁻⁷ M, respectively. The method was applied to the determination of cyanide in various industrial waste waters such as electroplating waste water and also for determination of hydrogen cyanide in air samples.     

Application of cement as new electrode material and solid-phase microextraction material demonstrated by electrochemiluminescent detection of perphenazine

Because of its unique pore network, good strength, and low cost, cement was used as a new electrode material and solid-phase microextraction (SPME) material for the first time. It was mixed with carbon to make a new electrode, cement carbon electrode (CCE). The as-prepared CCE was used to demonstrate the application of cement in SPME by Ru(bpy)₃²⁺ electrochemiluminescent detection of perphenazine (PPZ). The calibration plot for PPZ is linear from 1.0 × 10⁻⁹ to 3.0 × 10⁻⁶ M with a detection limit of 3.1 × 10⁻¹⁰ M. The method was successfully applied to the detection of PPZ in urine sample. Cement-based electrode material may also find broad applications in electrochemistry industry, such as electrochemical wastewater treatment.     

Zeolite A functionalized with copper nanoparticles and graphene oxide for simultaneous electrochemical determination of dopamine and ascorbic acid

A novel Cu-zeolite A/graphene modified glassy carbon electrode for the simultaneous electrochemical determination of dopamine (DA) and ascorbic acid (AA) has been described. The Cu-zeolite A/graphene composites were prepared using Cu²⁺ functionalized zeolite A and graphene oxide as the precursor, and subsequently reduced by chemical agents. The composites were characterized by X-ray diffraction, Fourier transform infrared spectra and scanning electron microscopy. Based on the Cu-zeolite A/graphene-modified electrode, the potential difference between the oxidation peaks of DA and AA was over 200 mV, which was adequate for the simultaneous electrochemical determination of DA and AA. Also the proposed Cu-zeolite/graphene-modified electrode showed higher electrocatalytic performance than zeolite/graphene electrode or graphene-modified electrode. The electrocatalytic oxidation currents of DA and AA were linearly related to the corresponding concentration in the range of 1.0 × 10⁻⁷–1.9 × 10⁻⁵ M for DA and 2.0 × 10⁻⁵–2.0 × 10⁻⁴ M for AA. Detection limits (<!-- no-mfc -->S/N<!-- /no-mfc --> = 3) were estimated to be 4.1 × 10⁻⁸ M for DA and 1.1 × 10⁻⁵ M for AA, respectively.     

In situ copper oxide modified molecularly imprinted polypyrrole film based voltammetric sensor for selective recognition of tyrosine

Organic-inorganic hybrids are promising functional materials as they combine the special characteristics of both organic (polymer) and inorganic phases. Among different existing approaches for the preparation of such polymer-inorganic hybrid coatings, in situ electrochemical methods are very advantageous because of their high sensitivity and simplicity. In the present study, voltammetric sensors for tyrosine are designed and developed via various modifications on glassy carbon electrode such as polypyrrole coated GCE, molecularly imprinted polypyrrole coated GCE (MIPPy) and in situ copper oxide modified MIPPy coated GCE. Of these, in situ copper oxide modified MIPPy coated GCE sensor responds to tyrosine concentrations in the range 1 × 10⁻⁸ to 1 × 10⁻⁶ and 2 × 10⁻⁶ to 8 × 10⁻⁶ M with a very low detection limit of 4.0 × 10⁻⁹ M and by far the most sensitive one. Detailed linear sweep voltammetric and chronoamperometric experiments were undertaken to investigate the electrocatalytic behavior of tyrosine. The electron transfer coefficient, diffusion coefficient and charge transfer rate constants involved in the sensing process using in situ copper oxide modified MIPPy film coated GCE are 0.47, 1.88 × 10⁻⁶ cm² s⁻¹, 4.7 × 10⁶ L mol⁻¹ s⁻¹, respectively. Furthermore, the designed sensor is highly selective and has been applied successfully for the analysis of synthetic and real samples of human urine.     

Simultaneous determination of copper and lead in seawater using optimised thin-mercury film electrodes in situ plated in thiocyanate media

In the present work the anodic stripping voltammetric (ASV) methodology using a thin mercury film electrode in situ plated in thiocyanate media was re-assessed in order to allow the simultaneous determination of copper and lead in seawater. Under previously suggested conditions [6], i.e. using a concentration of thiocyanate of 5 mM, the ASV peaks of copper and lead overlapped due to the formation of a stable copper(I)-thiocyanate species, limiting the analytical determinations. Therefore, the best value for the thiocyanate concentration was re-evaluated: for 0.05 mM a trade-off between good resolution of the copper and lead peaks and high reproducibility of the mercury film formation/removing processes was achieved. In this media, the ASV peaks for Pb and Cu occurred, separated by 140 mV. Also, the in situ thin mercury film electrode was produced and removed with good repeatability, which was confirmed by the relative standard deviation values for the ASV determinations: 0.5% for Pb and 2.0% for Cu (10 replicate determinations in a solution with metal concentrations 1.5×10⁻⁸ M for lead and 2.2×10⁻⁸ M for copper). The optimised methodology was successfully applied to the determination of copper in the presence of lead, in certified seawater (NASS-5).