Determination of prednisolone and prednisone in plasma by liquid chromatography with fluorescence detection

After a single-step extraction from plasma (250 μl) with dichloromethane, the drugs and dexamethasone (internal standard) are oxidized by copper(II) acetate to the corresponding glyoxal and converted into the fluorescent quinoxalines by reaction with 1,2-diamino-4,5-methylenedioxybenzene. The quinoxalines are separated within 55 min by reversed-phase liquid chromatography with isocratic elution. The detection limits for prednisolone and prednisone added to plasma are 3 ng ml^?1 in plasma (signal-to-noise ratio=3).     

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.     

Simultaneous voltammetric determination of levodopa, carbidopa and benserazide in pharmaceuticals using multivariate calibration

An analytical methodology based on differential pulse voltammetry (DPV) on a glassy carbon electrode and the partial least-squares (PLS-1) algorithm for the simultaneous determination of levodopa, carbidopa and benserazide in pharmaceutical formulations was developed and validated. Some sources of bi-linearity deviation for electrochemical data are discussed and analyzed. The multivariate model was developed as a ternary calibration model and it was built and validated with an independent set of drug mixtures in presence of excipients, according with manufacturer specifications. The proposed method was applied to both the assay and the uniformity content of two commercial formulations containing mixtures of levodopa-carbidopa (10:1) and levodopa-benserazide (4:1). The results were satisfactory and statistically comparable to those obtained by applying the reference Pharmacopoeia method based on high performance liquid chromatography. In conclusion, the methodology proposed based on DPV data processed with the PLS-1 algorithm was able to quantify simultaneously levodopa, carbidopa and benserazide in its pharmaceuticals formulations using a ternary calibration model for these drugs in presence of excipients. Furthermore, the model appears to be successful even in the presence of slight potential shifts in the processed data, which have been taken into account by the flexible chemometric PLS-1 approach.     

Anodic voltammetric behavior and determination of cefixime in pharmaceutical dosage forms and biological fluids

The voltammetric behavior of cefixime was studied using cyclic, linear sweep, differential pulse and square wave voltammetric techniques. The oxidation of cefixime was irreversible and exhibited diffusion controlled process depending on pH. The oxidation mechanism was proposed and discussed. Different parameters were tested to optimize the conditions for the determination of cefixime. The dependence of current intensities and potentials on pH, concentration, scan rate, nature of the buffer was investigated. According to the linear relationship between the peak current and the concentration, differential pulse (DPV) and square wave (SWV) voltammetric methods for cefixime assay in pharmaceutical dosage forms and biological fluids were developed. For the determination of cefixime were proposed in acetate buffer at pH 4.5, which allows quantitation over the 6 × 10⁻⁶-2 × 10⁻⁴ M range in supporting electrolyte and spiked serum sample; 8 × 10⁻⁶-2 × 10⁻⁴ M range in urine sample; 6 × 10⁻⁶-1 × 10⁻⁴ M range in breast milk samples for both techniques. The repeatability, reproducibility, precision and accuracy of the methods in all media were investigated. No electroactive interferences from the excipients and endogenous substances were found in the pharmaceutical dosage forms and in the biological samples, respectively.     

Adsorptive voltammetric procedure for the determination of platinum baseline levels in human body fluids

Summary An extremly sensitive procedure for the determination of platinum in human body fluids is presented. A high pressure decomposition of the samples is followed by adsorptive voltammetric measurement. A detection limit down to 0.2 ng Pt/l sample allowed baseline levels of platinum in body fluids (urine: 0.5–15 ng/l, blood and blood plasma: 0.8–6.9 ng/l) to be evaluated. The concentration ranges in body fluids of occupationally exposed people were determined to 21–2900 ng/l (urine), 32–180 ng/l (blood) and 95–280 ng/l (blood plasma).     

Spectrofluorometric determination of nicardipine,nifedipine and isradipine in pharmaceutical preparations and biological fluids

A simple and highly sensitive spectrofluorometric method was developed for the determination of some 1,4-dihydropyridine compounds namely, nicardipine, nifedipine and isradipine in pharmaceutical preparations and biological fluids. The method is based on the reduction of nicardipine, nifedipine and isradipine with Zn/HCl and measuring the fluorescence intensity obtained (λ_em/λ_ex) at 460/364, 450/393 and 446/360 nm, respectively. The factors affecting the development of the fluorophore and its stability were studied and optimized. The effect of some surfactants such as β-cyclodextrin (βCD), carboxymethylcelullose (CMC), sodium dodecyl sulphate (SDS) and triton X-100, on the fluorescence intensity was studied. The fluorescence intensity-concentration plots of nicardipine, nifedipine and isradipine were rectilinear over the ranges 0.4–6.0, 0.2–4.0 and 0.1–9.0 μg ml⁻¹ with detection limits of 0.0028, 0.017 and 0.016 μg ml⁻¹, respectively. The proposed method was successfully applied to commercial tablets containing the compounds; the percentage recovery agreed well with those obtained using the official methods. The method was further extended to the in vitro determination of the compounds in spiked human plasma and urine samples. A proposal of the reduction reaction pathway was postulated.      

Sequential voltammetric determination of trace metals in meals

An analytical procedure for the sequential determination of Zn(II), Cr(VI), Cu(II), Sb(III), Sn(II), Pb(II) by square wave anodic stripping voltammetry (SWASV) and Fe(III), Mn(II), Mo(VI) by square wave voltammetry (SWV) in matrices involved in foods and food chain as wholemeal, wheat and maize meal is described.The digestion of each matrix was carried out using a concentrated HCl–HNO₃–H₂SO₄ attack mixture, employing dibasic ammonium citrate buffer solution (pH 6.9 and 8.7) as supporting electrolytes. The analytical procedure was verified by the analysis of the standard reference materials Wholemeal BCR-CRM 189, Wheat Flour NIST-SRM 1567a and Rice Flour NIST-SRM 1568a.For all the elements in the certified matrix, the precision as repeatability, expressed as relative standard deviation (s_r) was of the order of 3–5%; the accuracy, expressed as relative error (e) was generally of the order of 3–6%.In presence of reciprocal interference, the standard addition method considerably improved the resolution of the voltammetric technique.Finally, the analytical procedure was transferred and applied to commercial meals sampled on market.A critical comparison with atomic absorption spectroscopic measurements is also discussed.     

Simultaneous determination of cefotaxime and desacetylcefotaxime in real urine sample using voltammetric and high-performance liquid chromatographic methods

Two rapid, accurate and sensitive methods are developed and validated for the quantitative simultaneous determination of cefotaxime (CFX) and its active metabolite desacetylcefotaxime (DCFX) in urine.Based on the previous results which showed the four electron reduction of CFX at ≈ −0.5 V, and the new findings that DCFX reduction occurred at more positive potential (−0.23 V), the new adsorptive stripping differential pulse voltammetric (AdSDPV) method was developed for determination of CFX in the presence of DCFX. Linear responses were observed over a wide concentration range (0.07-0.52 μg/ml for CFX and 0.22-1.3 μg/ml for DCFX) in urine.The second assay involves subsequent separation on a reversed-phase HPLC column, with ultraviolet detection at 262 nm. Retention times were 4.057 and 1.960 min for CFX and DCFX, respectively. Linear responses were observed over a wide range, 0.55-6.60 μg/ml for CFX and 1.10-11.00 μg/ml for DCFX, in urine.The statistical evaluation for both methods was examined by means of within-day repeatability (n = 5) and day-to-day precision (n = 3) and was found to be satisfactory with high accuracy and precision.     

Electrochemical characterisation of nefazodone hydrochloride and voltammetric determination of the drug in pharmaceuticals and human serum

Nefazodone, an antidepressant was electrochemically studied in various buffer systems and at different pH using glassy carbon electrode. Nefazodone was electrochemically oxidized at all pH values. According to the linear relation between the peak current and the nefazodone concentration differential pulse (DPV) and square wave (SWV) voltammetric methods for its quantitative determination in pharmaceuticals and human serum were developed. For analytical purposes, a very well resolved diffusion controlled voltammetric peak was obtained in 0.1 M H₂SO₄ at 0.99 and 1.03 V for DPV and SWV techniques, respectively. The linear response was obtained in the ranges of 8×10⁻⁷ to 6×10⁻⁴ M with a detection limit of 2.1×10⁻⁷ M for DPV and 1.17×10⁻⁷ M for SWV techniques. The repeatability and reproducibility of the methods were within 1.03, 0.81% relative standard deviations (R.S.D.) for peak currents and 0.40, 0.20% R.S.D. for peak potentials, for DPV and SWV, respectively. Precision and accuracy of the developed method was checked by recovery studies. The proposed methods were successfully applied to the individual tablet dosage form and human serum.     

Flow-injection enhanced chemiluminescence method for determination of ciprofloxacin in pharmaceutical preparations and biological fluids

A novel rapid and sensitive analytical method, enhanced chemiluminescence with flow-injection sampling, is described for determination of ciprofloxacin. The method is based on the chemiluminescence reaction of the potassium permanganate–sodium thiosulfate–ciprofloxacin system. An enhanced chemiluminescence reaction was developed, and optimum conditions for CL emission were investigated. The chemiluminescence intensity was linearly dependent on ciprofloxacin concentration in the range 1.0×10⁻⁸–1.0×10⁻⁵ g mL⁻¹. The detection limit was 4×10⁻⁹ g mL⁻¹. The relative standard deviation was 1.8% for eleven measurements of 2.0×10⁻⁷ g mL⁻¹ ciprofloxacin standard solution. The new method enables simple, sensitive, and rapid determination of ciprofloxacin and has been successfully used for determination of ciprofloxacin in biological fluids and in ciprofloxacin hydrochloride tablet and injection.     

Simultaneous determination of prednisolone acetate, prednisolone, prednisone, cortisone and hydrocortisone in swine plasma using solid-phase and liquid-liquid extraction techniques

A high-performance liquid chromatographic (HPLC) method for the simultaneous determination of prednisolone acetate (PA), prednisolone (PO), prednisone (PN), cortisone and hydrocortisone in swine plasma is described. Extraction of the steroid mixture from swine plasma with dexamethasone as internal standard was accomplished by solid-phase extraction (SPE) or the more traditional liquid-liquid extraction (LLE) techniques. These compounds were analyzed by normal-phase HPLC with ultraviolet detection. Although a detectable sensitivity of 5 ng/ml is achieved by the SPE technique, the practical sensitivity is established as 10 ng/ml. Conversely, the practical sensitivity is 5 ng/ml for all compounds by the LLE technique. Calibration curves were found to be linear between 10 and 500 ng/ml by the SPE technique and between 5 and 100 ng/ml by the LLE technique. The average recovery of the steroids PA, PO and PN at 20 ng/ml is between 70 and 90%. PA is stable for up to 3 h in swine plasma at room temperature (22 degrees C) but is completely converted to PO within 24 h. PA is stable in swine plasma in an ice bath for over 24 h. The usefulness of this analytical technique is demonstrated by the intraperitoneal administration of 125 mg of PA to swine and the quantitative determination of PA, PO and PN in the plasma as a function of time.     

Potentiometric determination of antihistaminic diphenhydramine hydrochloride in pharmaceutical preparations and biological fluids using screen-printed electrode

The performance characteristic of sensitive screen-printed (SPE) and carbon paste (CPE) electrodes was investigated for the determination of diphenhydramine hydrochloride (DPH) drug in pure, pharmaceutical preparations and biological fluids. Different experimental conditions namely types of materials used to prepare the working electrode (plasticizer), titrant, pH, temperature and life time were studied. Under these conditions, the SPE shows the best performance than CPE with respect to total potential change and potential break at the end point. The SPE and CPE exhibit suitable response to DPH in a concentration range of 1.0.10^{− 2} to 1.0.10^{− 6} mol/L with a limit of detection 9.70.10^{− 7} and 9.80.10^{− 7} mol/L, respectively. The slope of the system was 55.2 ± 1.0 and 54.7 ± 1.0 mV/decade over pH range 3.0–8.0 and 3–7 for SPE and CPE, respectively. Selectivity coefficients for DPH relative to a numbers of potential interfering substances were investigated. The SPE and CPE show a fast response time of 10 and 16 s and were used over a period of 2 months with a good reproducibility. The sensors were applied successfully to determine DPH in pharmaceutical preparations and biological fluids. The results are compared with the official method.     

An overview of the analytical methods for the determination of organic ultraviolet filters in biological fluids and tissues

Organic UV filters are chemical compounds added to cosmetic sunscreen products in order to protect users from UV solar radiation. The need of broad-spectrum protection to avoid the deleterious effects of solar radiation has triggered a trend in the cosmetic market of including these compounds not only in those exclusively designed for sun protection but also in all types of cosmetic products.     

Development of a validated HPLC method for the determination of four 1,4-benzodiazepines in human biological fluids

A simple and sensitive HPLC method was developed and validated for the determination of four frequently prescribed 1,4-benzodiazepines: alprazolam (ALP), bromazepam (BRZ), diazepam (DZP), and flunitrazepam (FNZ). Separation was achieved on an Inertsil C8 analytical (250 mm x 4 mm, 5 microm) column, after selective extraction of benzodiazepine drugs from biological matrices by means of SPE. Isocratic elution was performed with a mobile phase consisting of CH3COONH4, 0.05 M CH3OH, and CH3CN (33:57:10 by volume). Quantification was performed at 240 nm with mefenamic acid (6 ng/microL) as the internal standard. DSC-18 Supelco cartridges provided high absolute recoveries (81-115%). The developed method was fully validated in terms of selectivity, linearity, accuracy, precision, stability, and sensitivity. Repeatability (n = 8) and between-day precision (n = 8) revealed RSD <12%. Recoveries from biological samples ranged from 81.2 to 115%. The detection limit of the method was calculated as 3.3-10.2 ng in blood plasma and 2.6-12.6 ng in urine for 20 microL injection volume. The method was applied to spiked biological matrices. Moreover, the method was applied to real samples of urine after an oral administration.     

Analytical procedures for the simultaneous voltammetric determination of heavy metals in meals

An analytical procedure regarding the determination of copper(II), lead(II), cadmium(II), zinc(II) and antimony(III) in matrices involved in foods and food chain as wholemeal, wheat and maize meal is proposed. The digestion of each matrix was carried out using concentrated HCl suprapure at 130 °C for 3 h. Differential pulse anodic stripping voltammetry (DPASV) was employed for simultaneously determining all the elements, using a conventional three-electrode cell and 0.5 M HCl as supporting electrolyte. The analytical procedure has been verified on the reference standard materials Wholemeal BCR-CRM 189, Wheat Flour NIST-SRM 1567a and Rice Flour NIST-SRM 1568a. For all the elements in the certified matrix, the precision as repeatability, expressed as relative standard deviation (s_r), and the accuracy, expressed as relative error (e), were of the order of 3 to 6%. The limits of detection were in the range 0.009–0.096 μg/g.     

Development and validation of an HPLC method for the determination of benzodiazepines and tricyclic antidepressants in biological fluids after sequential SPE

A novel method for the simultaneous determination of six benzodiazepines (BZDs) and four tricyclic antidepressants (TCAs) in biological fluids by HPLC with UV detection at 240 nm has been developed. After a deproteinization step biological fluids were analyzed by direct injection. SPE on Nexus cartridges was also applied. Since two compounds, namely imipramine and diazepam, were coeluting, a sequential SPE protocol has been developed. BZDs were eluted by a mixture of methanol/ACN(1:1), followed by the elution of TCAs with methanol. Separation was performed on a Kromasil C8 column (250 x 64 mm(2) id, 5 microm) using a mobile phase of 0.05 MCH3COONH4/ACN/methanol (initial composition 55:15:30 v/v/v) at a flow rate of 1.0 mL/min delivered by a gradient program within 15 min. Colchicine was used as the internal standard (4 ng/microL). The method was linear for all analytes up to 20 ng/lL, with coefficients of regression between 0.996 and 0.99996. LODs and LOQs were 0.08-1.17 and 0.28-3.91 ng/lL, respectively. Recovery was in the range of 92.8-108.7% for within-day and 91.9-109.9% for between-day assays, with RSD values lower than 10.0% for all matrices.     

Adsorptive stripping voltammetric determination of antimony

A new method is described for the determination of antimony based on the cathodic adsorptive stripping of Sb(III) complexed with 2′,3,4′,5,7-pentahydroxyflavone(morin) at a static mercury drop electrode (SMDE). The reduction current of the adsorbed antimony complex was measured by 1.5th-order derivative linear-sweep adsorption voltammetry. The peak potential is at −0.51 V (vs. SCE). The effects of various parameters on the response are discussed. The optimized analytical conditions were found to be: supporting electrolyte, chloroacetic acid (0.04 mol/l, pH 2.3); concentration of morin, 5×10⁻⁶ mol/l; accumulation potential, −0.25 V (vs. SCE); scan rate, 100 mV/s. The limit of detection and the linear range were 7×10⁻¹⁰ mol/l and 1.0×10⁻⁹3.0×10⁻⁷ mol/l Sb(III) for a 2-min accumulation time, respectively. This method has been applied to the determination of Sb(III) in steel and brass samples and satisfactory results were obtained. The adsorptive voltammetric characteristics and composition of the Sb(III)–morin complex were studied.     

Determination of chlorproguanil and metabolites in biological fluids

Summary The retention behaviour of chlorproguanil and its putative metabolites chlorcycloguanil and 3,4-dichlorophenylbiguanide has been studied on a reversed phase chromatographic system incorporating sodium lauryl sulphate as hydrophobic pairing ion. On the basis of data obtained and comparison of standard compounds with components present in urine following the administration of chlorproguanil, the identity of the metabolites have been confirmed chromatographically. A third unidentified metabolite is also observed. The retention study allows selection of a solvent system which, when used with a small volume, 100 × 2 mm, 3μm ODS column, enables chlorproguanil and its two major metabolites to be determined in plasma, whole blood and urine. The analytical characteristics of the method are reported and the usefullness of the method in obtaining pharmacokinetic data on the drug and its metabolites is discussed.     

Voltammetric determination of anabolic steroid nandrolone at gold nanoparticles modified ITO electrode in biological fluids

The electrochemical behavior of nandrolone decanoate (ND) at gold nanoparticles modified indium tin oxide (ITO) electrode was investigated. Oxidation of ND has been carried out in phosphate containing supporting electrolyte in the pH range 2.1-9.2 and a well-defined oxidation peak was noticed. The peak potential (E_p) of the oxidation peak decreases linearly with increasing pH. Linear calibration curve is obtained over the nandrolone decanoate concentration range of 50 nM to 1.5 μM at pH 7.2 with a detection limit of 1.36 × 10⁻⁷ M. The proposed method is effectively applied to detect the concentration of ND in human blood serum and urine samples after 24 and 72 h of intramuscular injection. The method is rapid and does not require any pre-treatment.     

Studies on voltammetric determination of cadmium in samples containing native and digested proteins

This work focuses on determination of cadmium ions using anodic stripping voltammetry (ASV) on thin film mercury electrode in conditions corresponding to those obtained after digestion of cadmium-based quantum dots and their conjugates. It presents the impact of selected proteins, including potential receptors and surface blocking agents on the voltammetric determination of cadmium. Experiments regarding elimination of interferences related to proteins presence using sodium dodecyl sulfate (SDS) are also shown. Effect of SDS on selected analytical parameters and simplicity of analyses carried out was investigated in the framework of current studies. The significant differences of influence among tested proteins on ASV cadmium determination, as well as the variability in SDS effectiveness as the antifouling agent were observed and explained. This work is especially important for those, who design new bioassays and biosensors with a use of quantum dots as electrochemical labels, as it shows what problems may arise from presence of native and digested proteins in tested samples.