Biomedical subjects

M A Koupparis

Publications and source records attributed to M A Koupparis.

At least 19 recordsLinked to original sources

PubMed2004

Simultaneous HPLC determination of ketoprofen and its degradation products in the presence of preservatives in pharmaceuticals.

A novel and quick high-performance liquid chromatography (HPLC) method with UV spectrophotometric detection was developed and validated for the determination of five compounds in topical gel. The described method is suitable for simultaneous determination of active component ketoprofen, two preservatives methylparaben and propylparaben and two degradation products of ketoprofen--3-acetylbenzophenone and 2-(3-carboxyphenyl) propionic acid--in a topical cream after long-term stability tests using ethylparaben as an internal standard. The chromatographic separation was performed on a 5microm Supelco Discovery C18 column (125mm x 4mm i.d., Sigma-Aldrich); the optimal mobile phase for separation of ketoprofen, methylparaben, propylparaben, degradation products 3-acetylbenzophenone and 2-(3-carboxyphenyl) propionic acid and ethylparaben as internal standard consists of a mixture of acetonitril, water and phosphate buffer pH 3.5 (40:58:2, v/v/v). At a flow rate of 1.0ml min(-1) and detection at 233nm, the total time of analysis was less than 10min. The method was applied for routine analysis (batch analysis and stability tests) of these compounds in topical pharmaceutical product.

Chromatography, High Pressure Liquid↗

PubMed2001

Nonaqueous catalytic fluorometric trace determination of vanadium based on the pyronine B-hydrogen peroxide reaction and flow injection after cloud point extraction.

The catalytic effect of vanadium on the pyronine B-H2O2 system is examined. Enhancement of the catalytic reaction rate along with the efficiency and selectivity against vanadium is achieved in a formic acid environment in the presence of a nonionic surfactant (Triton X-114). Elimination of drastic interference caused by inorganic acids and aqueous matrix along with a 50-fold preconcentration of vanadium are facilitated through cloud point extraction of its neutral complex with 8-quinolinol in an acidic solution. Subsequent flow injection analysis (FIA) with fluorometric detection renders the proposed method ideal for selective and cost-effective determination of as little as 0.020 microng L(-1) vanadium in environmental, biological, and food substrates. The preconcentration step can be applied simultaneously to multiple samples, allowing for massive preparation prior to analysis, compensating, thus, for the time-consuming procedure.

Catalysis↗

PubMed2001

Improved antibody coating protocol using a second antibody antiserum. Application to total thyroxin immunoassay.

A complete antibody coating protocol for the preparation of dry antibody coated tubes is presented. This protocol is based on a recently described antibody immobilization principle. We modify this immobilization principle in order to improve and simplify the coating procedure. In addition, we propose a drying procedure that provides long-term storage stability of the antibody coated tubes. According to the modified protocol, polystyrene plastic tubes are first coated with rabbit gamma-globulins. The tubes are incubated with a sheep anti-rabbitIgG antiserum dilution. After incubation, antigen-specific antibody antiserum raised in rabbits is added directly into the tubes containing the sheep anti-rabbit IgG antiserum solution (difference from the original protocol). Finally, the tubes are washed, blocked, and dried following the drying procedure developed. The suitability of the modified protocol for the development of immunoassays requiring high loading of antibody was exemplified through the development of a RIA for total thyroxin. The estimated assay characteristics (detection limit 4 microg/L, dynamic range up to 210 microg/L, within-run CV 2.7-5.7%, between-run CV 5.1-7.3%, recovery 84.4-112%, cross-reactivity for T3 1.9%) were comparable with those provided by commercially available RIA kits for the determination of thyroxin.

Animals↗

PubMed2000

Automated flow-injection spectrophotometric determination of catecholamines (epinephrine and isoproterenol) in pharmaceutical formulations based on ferrous complex formation.

A novel automated flow-injection spectrophotometric method for the determination of catecholamines (epinephrine and isoproterenol) has been developed based on the formation of their coloured complexes with Fe(II) in aminoacetic-carbonate buffer pH 8.3 and measuring of the absorbance peaks at the lambda(max) of 530 nm. A fully automated FIA system controlled by home-made software (FIA-MOD) was used for optimising the chemical and manifold parameters and running of routine measurements. The calibration graph was linear in the range of 5-200 mg l(-1) for epinephrine with an RSD of 0.24% (n = 5; c = 150 mg l(-1)) and 10-300 mg(-1) for isoproterenol with an RSD of 0.13% (n = 5; c = 200 mg l(-1)). Measurement throughput was 120 h(-1) ensuring a sample throughput of 40 h(-1) analysed in triplicate. Common excipients for tablets and injections were found not interfering. The proposed method was applied for the assay of various commercial pharmaceutical formulations containing epinephrine and isoproterenol and for the content uniformity test for the isoproterenol tablets. The assay results with RSD 2-4% (n = 3) were comparable with those obtained with the official USP XXIII methods (mean difference 1.9%).

Artifacts↗

PubMed1999

Automated flow injection gradient technique for binding studies of micromolecules to proteins using potentiometric sensors: application to bovine serum albumin with anilinonaphthalenesulfonate probe and drugs.

An automated flow injection (FI) gradient technique is described for the binding study of the potentiometric probe 1-anilino-8-naphthalenesulfonate (ANS) to bovine serum albumin (BSA). Using a single-channel FI system with a mixing chamber and a flow ANS electrode, the binding parameters (binding constant and number of binding sites) were calculated using the Scatchard model. The concentration gradient was calibrated by injecting ANS in the stream, and the binding experiment was performed by injecting ANS-BSA solution in the carrier solution of equal albumin concentration. The equations describing the concentration gradient and the corresponding electrode potential curve are presented. A systematic study of the factors affecting the complexation equilibrium and the electrode response was performed. For the ANS binding to BSA, two binding classes were determined with binding constants of (2.1 +/- 0.3) x 10(5) and (3.3 +/- 0.8) x 10(3) M-1 and 3.8 +/- 0.6 and 10 +/- 2 binding sites per class, respectively, at 27 +/- 1 degrees C, in 0.10 M phosphate pH 7.4. Competitive binding experiments of sulfamethoxazole, salicylate, azapropazone, ketoprofen, and tolmetin to albumin were also performed by monitoring ANS binding inhibition (decrease of apparent binding constant). This technique takes advantage of FI gradients and direct potentiometry and utilizes the total information contained in FI peaks, providing fast and accurate binding information in a wide range of concentration ratios.

Anilino Naphthalenesulfonates↗

PubMed1999

Flow-injection fluorimetric determination of 1,4-benzodiazepines in pharmaceutical formulations after acid hydrolysis.

A simple, rapid and fully automated flow injection method with fluorimetric detection after hydrolysis with H2SO4 in ethanolic or methanolic medium at room temperature has been developed for the determination of 1,4-benzodiazepines (oxazepam, diazepam and nitrazepam) in pharmaceutical formulations. The calibration curves are linear in the ranges (mg ml(-1)) of oxazepam (0.025-0.150), diazepam (0.010-0.125) and nitrazepam (0.010-0.150), with detection limits of 0.01, 0.005 and 0.005 mg ml(-1), respectively, and RSD (1% (n = 10). The measurement throughput is 60 h(-1) using a 200-microl sample volume obtained by the direct dissolution of formulations in alcohol.

Anti-Anxiety Agents↗

PubMed1999

Studies on the interaction of diflunisal ion with cyclodextrins using ion-selective electrode potentiometry.

The interaction of diflunisal ion (DF) with beta-cyclodextrin (betaCD), gamma-cyclodextrin (gammaCD), and hydroxypropyl-beta-cyclodextrin (HPbetaCD) was studied in phosphate buffer, pH 7.4, at 5-37 degrees C and various CD concentrations using a home-made diflunisal ion-selective electrode. Typical direct binding plots and Scatchard plots were obtained with HPbetaCD. The Scatchard model for one class of binding sites was used for the estimation of binding parameters for the DF/HPbetaCD interaction. The estimates for n (number of binding sites per CD molecule) were in all cases very close to unity, indicating 1:1 complexation. The association constant (K) estimates decrease with increasing temperature. Sigmoidal direct binding plots and concave-downwards Scatchard plots were obtained with various betaCD or gammaCD concentrations. The Hill model was used for the estimation of the binding parameters for the DF/betaCD and DF/gammaCD interactions. Both the Hill coefficients and the binding constants were markedly dependent on the CD concentration. These findings indicate the cooperative character of DF/betaCD and DF/gammaCD interactions. The free energy change, DeltaG, and the thermodynamic parameters, DeltaH and DeltaS, were estimated for each of the interactions studied using the Van't Hoff equation.

Algorithms↗

PubMed1998

Antibody coating approach involving gamma globulins from non-immunized animal and second antibody antiserum.

An alternative protocol for immobilization of antibodies onto plastic solid supports is presented. According to the proposed protocol, tubes are first coated with gamma-globulins from non-immunized animal of the same species as that from which the antigen-specific antibody has been developed. Then, an excess of second antibody is added to the tubes and the anti-species specific antibodies present in the antiserum are immunoadsorbed on the immobilized gamma-globulins. Finally, the antigen specific antibody is immunoadsorbed on the immobilized second antibody. We found that the coating protocol developed allows the use of antigen-specific and second antibody antisera dilutions, thus avoiding the need for affinity purification of antibodies. Additionally, it provides solid-phase second antibody with increased binding capacity compared to the directly adsorbed onto the solid second antibody. The advantages of the proposed coating protocol were demonstrated through the development of a solid-phase radioimmunoassay for the determination of total triiodothyronine in human serum samples.

Animals↗

PubMed1998

An in vitro evaluation of fluoxetine adsorption by activated charcoal and desorption upon addition of polyethylene glycol-electrolyte lavage solution.

BACKGROUND: In drug overdoses, treatment with activated charcoal is frequently used because of its adsorptive properties. Recently, whole-bowel irrigation with polyethylene glycol-electrolyte lavage solution has been used as a gastrointestinal decontamination procedure for ingestions of toxins not well adsorbed to activated charcoal and for toxins with a delayed absorption phase, although well adsorbed to activated charcoal. While a combined approach using activated charcoal and whole-bowel irrigation could theoretically enhance the efficacy of both modalities, this improvement remains speculative, since data demonstrating its clinical advantage in overdose treatment are lacking. Fluoxetine, a selective serotonin uptake inhibitor, is one of the most frequently prescribed antidepressants. Fluoxetine is well adsorbed onto activated charcoal. This in vitro investigation was undertaken to study: a) the effect of polyethylene glycol, as well as polyethylene glycol-electrolyte lavage solution, on the adsorption of fluoxetine to laboratory grade-activated charcoal and a commercial activated charcoal formulation (Carbomix powder) in simulated gastric (pH= 1.2) and intestinal (pH=7.2) fluid environment; b) whether the order of polyethylene glycol-electrolyte lavage solution addition would have any effect on the adsorption of fluoxetine to activated charcoal. METHODS: Adsorption of fluoxetine to charcoal in the presence of polyethylene glycol was examined: a) by the simultaneous addition of polyethylene glycol and charcoal to fluoxetine solution and b) by the addition of charcoal to fluoxetine solution and subsequent addition of polyethylene glycol. In both cases, the slurries were incubated at 37 degrees C for 1 hour and filtered. Free fluoxetine concentration was determined in the diluted filtrate by a reversed-phase high-performance liquid chromatography method. RESULTS: The amount of fluoxetine adsorbed to activated charcoal (or Carbomix) was dramatically decreased at gastric and intestinal pH by the presence of polyethylene glycol or polyethylene glycol-electrolyte lavage solution added either concurrently or sequentially to activated charcoal. CONCLUSIONS: In cases of fluoxetine overdose, administration of activated charcoal is recommended, while a combined approach using activated charcoal and whole-bowel irrigation with polyethylene glycol-electrolyte lavage solution is not recommended since it causes a significant desorption of the drug from activated charcoal.

Adsorption↗

PubMed1997

In vitro adsorption study of fluoxetine onto activated charcoal at gastric and intestinal pH using high performance liquid chromatography with fluorescence detector.

BACKGROUND: This in vitro investigation was performed to study the adsorption characteristics of fluoxetine to activated charcoal and its commercial formulation Carbomix powder in simulated gastric (pH = 1.2) and intestinal (pH = 7.2) fluid environments. METHODS: Solutions containing fluoxetine and charcoal were incubated at 37 degrees C for one hour. Reversed phase high performance liquid chromatography was used for the determination of free fluoxetine concentrations (range 0.2-8 micrograms/mL) in the diluted filtrate. RESULTS: The maximum adsorption capacities at pH 1.2 for activated charcoal and Carbomix were 223 and 333 mg/g, respectively; at pH 7.2 they were 301 and 453 mg/g, respectively. The affinity constant values at pH 1.2 of activated charcoal and Carbomix were 441 and 122 L/g, respectively, while at pH 7.2 they were 482 and 589 L/g, respectively, indicating a strong binding of fluoxetine onto charcoals. CONCLUSIONS: Relative to the toxic and lethal doses in cases of fluoxetine intoxications, both types of charcoals tested were found effective for adsorption at gastric and intestinal pH. Adsorbed fluoxetine was significantly increased at intestinal pH, consistent with predominant adsorption of the undissociated form of the drug. We conclude that activated charcoal and Carbomix have adsorptive properties appropriate to medical treatment in cases of fluoxetine overdose.

Adsorption↗

PubMed1996

The effect of polyethylene glycol on the charcoal adsorption of chlorpromazine studied by ion selective electrode potentiometry.

BACKGROUND: This investigation was undertaken to study: a) the adsorption characteristics of chlorpromazine to activated charcoal and its formulations Carbomix powder and Ultracarbon tablets at gastric pH; b) the effect on chlorpromazine adsorption of polyethylene glycol and its combination with electrolyte lavage solution; c) the effect of the order of addition of polyethylene glycol-electrolyte lavage solution. METHOD: Ion selective electrode potentiometry, based on the selective, direct and continuous response of a chlorpromazine-ion selective electrode to the concentration of the free drug, was used. Successive additions of microvolumes of a chlorpromazine solution were made into a charcoal slurry in acidic medium of pH 1.2 with measurement of the chlorpromazine-ion selective electrode potential at equilibrium. RESULTS: The maximum adsorption capacity values of activated charcoal, Carbomix and Ultracarbon, were 297, 563, and 382 mg/g respectively, while the affinity constant values were 40.2, 70.4, and 40.5 L/g, respectively. The adsorption of chlorpromazine to each of the Ultracarbon and Carbomix components was compared to the total adsorption of the formulations. The addition of polyethylene glycol-electrolyte lavage solution causes a slight desorption of chlorpromazine from activated charcoal at gastric pH, more pronounced when polyethylene glycol-electrolyte lavage solution follows the addition of activated charcoal, suggesting the possibility of a nonspecific binding of chlorpromazine to polyethylene glycol. The amount of chlorpromazine absorbed to Carbomix and Ultracarbon was not significantly affected at gastric pH by the presence of polyethylene glycol or polyethylene glycol-electrolyte lavage solution added either concurrently or sequentially to these formulations.

Adsorption↗

PubMed1995

Construction of a diflunisal ion sensor and its use in automated flow-injection methods for assay, content uniformity, and dissolution studies of formulations.

A diflunisal ion selective electrode of the PVC membrane type with an ion-exchanger consisting of the tetraheptylammonium-diflunisal ion pair is described. The sensor exhibits a rapid, near-Nernstian, selective response to diflunisal anion in the pH range 7-10, with a (batch) detection limit of 1 x 10(-5) M. The ion sensor was used as a flow detector in an automated flow-injection analyzer to develop routine methods for assays (concentration range 1-50 x 10(-4) M, (flow) detection limit 2.6 x 10(-5) M), content uniformity, and dissolution studies of diflunisal formulations. No serious interference from common ions and tablet excipients was found, and the drug can be directly determined in colored samples without separation steps. Fourty measurements can be performed automatically per hour with a precision of 0.5-1.8% relative standard deviation. The automated method for the dissolution test provides a complete dissolution profile by the end of the experiment. Using the constructed ion sensor, the intramolecular hydrogen bonding of the diflunisal anion was studied, thereby revealing a new application of ion sensor potentiometry.

Electrodes↗

PubMed1994

General treatment of competitive binding as applied to the potentiometric ion probe technique: application to the interaction of nonsteroidal anti-inflammatory drugs with bovine serum albumin.

The binding of naproxen, ketoprofen, phenylbutazone, salicylic acid, azapropazone, and indobufen to bovine serum albumin was studied by applying the potentiometric ion probe technique. An ion-selective electrode for the ion probe 1-anilino-8-naphthalene-sulfonate was utilized for the purposes of this study. A modified site-oriented competitive binding model was used for the estimation of the drugs' binding parameters, considering different number of binding sites on the competing binding class(es) for the probe and the drug. Calculations were based exclusively on the concentration data of the free probe. The model's ability for accurate estimations of binding parameters was evaluated by simulation studies. The following values of binding parameters were found at 25 degrees C for the drugs under study; naproxen, n1 = 9.1, k1 = 9.4 x 10(5) M-1; ketoprofen, n1 = 8.8, k1 = 10.8 x 10(5) M-1; phenylbutazone, n1 = 3.2, k1 = 1.4 x 10(5) M-1; salicylic acid, n1 = 2.6, k1 = 1.8 x 10(5) M-1, n2 = 21.5, k2 = 1.0 x 10(4) M-1; azapropazone, n1 = 0.5, k1 = 7.8 x 10(5) M-1, n2 = 26.3, k2 = 1.9 x 10(4) M-1; indobufen, n1 = 5.8, k1 = 5.8 x 10(5) M-1, n2 = 19.9, k2 = 3.8 x 10(5) M-1, where ni the number of binding sites of the i class and ki the corresponding association constant.

Anilino Naphthalenesulfonates↗

PubMed1994

Automated flow-injection technique for use in dissolution studies of sustained-release formulations: application to iron(II) formulations.

The application of flow-injection analysis (FIA) to automated dissolution studies of sustained-release formulations is described. The long-term stability of the dissolution-FIA analyser was checked during unattended operation for 42 h. The construction of multiple calibration curves with the so-called electronic dilution FIA procedure was used to extend the linear range of the determination. The computer-controlled FIA system and the principles of associated software are described and applied to dissolution studies of sustained-release formulations of iron(II) using its sensitive reaction with the colour reagent, ferrozine. The extended linear range of the determination is 1-130 ppm iron(II) and the precision (RSD) better than 3% (n = 3).

Calibration↗

PubMed1994

Effect of cyclodextrins on protein binding of drugs: the diflunisal/hydroxypropyl-beta-cyclodextrin model case.

The binding of diflunisal to hydroxypropyl-beta-cyclodextrin (HP beta CD), bovine serum albumin (BSA), human serum albumin (HSA), normal human plasma, and mixed solutions of HP beta CD/protein was studied at 25 degrees C, pH 7.4, by potentiometry using an electrode selective to diflunisal. The experimental data for diflunisal/HP beta CD fit well to the 1:1 binding model. The binding of diflunisal with each of the studied proteins was compatible with a model having two independent classes of binding sites. The binding of diflunisal in mixed solutions HP beta CD/BSA, HP beta CD/HSA, and HP beta CD/plasma increased considerably when the HP beta CD concentration was increased. The binding behavior of the two biomolecules in the mixed solutions of HP beta CD/BSA or HP beta CD/HSA was described with an "additive" model formulated on the basis of the estimates of the binding parameters of diflunisal derived from the separate experiments with each one of the binders tested. The lower than theoretical binding observed in HP beta CD/plasma solutions was ascribed to the competitive displacement of diflunisal from the HP beta CD cavity by plasma cholesterol.

2-Hydroxypropyl-beta-cyclodextrin↗

PubMed1993

Micellar acid-base potentiometric titrations of weak acidic and/or insoluble drugs.

The effect of various surfactants [the cationics cetyl trimethyl ammonium bromide (CTAB) and cetyl pyridinium chloride (CPC), the anionic sodium dodecyl sulphate (SDS), and the nonionic polysorbate 80 (Tween 80)] on the solubility and ionization constant of some sparingly soluble weak acids of pharmaceutical interest was studied. Benzoic acid (and its 3-methyl-, 3-nitro-, and 4-tert-butyl-derivatives), acetylsalicylic acid, naproxen and iopanoic acid were chosen as model examples. Precise and accurate acid-base titrations in micellar systems were made feasible using a microcomputer-controlled titrator. The response curve, response time and potential drift of the glass electrode in the micellar systems were examined. The cationics CTAB and CPC were found to increase considerably the ionization constant of the weak acids (delta pKa ranged from -0.21 to -3.57), while the anionic SDS showed negligible effect and the nonionic Tween 80 generally decreased the ionization constants. The solubility of the acids in aqueous micellar and acidified micellar solutions was studied spectrophotometrically and it was found increased in all cases. Acetylsalicylic acid, naproxen, benzoic acid and iopanoic acid could be easily determined in raw material and some of them in pharmaceutical preparations by direct titration in CTAB-micellar system instead of using the traditional non-aqueous or back titrimetry. Precisions of 0.3-4.3% RSD and good correlation with the official tedious methods were obtained. The interference study of some excipients showed that a preliminary test should be carried out before the assay of formulations.

Acids↗

PubMed1992

Determination of association constants in cyclodextrin/drug complexation using the Scatchard plot: application to beta-cyclodextrin-anilinonaphthalenesulfonates.

The appropriate Scatchard equation was developed for a system involving the formation of 1:1 and 1:2 substrate:cyclodextrin complexes. Simulation of this system was performed under the most common experimental conditions encountered in this type of study. The use of the equation allows for nonlinear least-squares estimation of the association constants. The interaction of the model compounds 1-anilino-8-naphthalenesulfonate (1,8-ANS) and 2(p-toluidinyl)-6-naphthalenesulfonate (2,6-TNS) with beta-cyclodextrin (beta-CD) was used to evaluate the theoretical model. Binding experiments were performed using either potentiometric titration or fluorimetric detection. The experimental data for 1,8-ANS/beta-CD fit well to the 1:1 binding model, with an association constant of 87 +/- 1 M-1. The association constants of the 1:1 and 1:2 2,6-TNS/beta-CD complexes utilizing direct potentiometry were 3737 +/- 6 and 149 +/- 2 M-1. It is shown that fluorimetry can give biased estimates for the association constants of the complexation 2,6-TNS/beta-CD, since the assumption of an equivalent quantum yield of bound species is not valid.

Anilino Naphthalenesulfonates↗

PubMed1992

Complexation studies of cyclodextrins with tricyclic antidepressants using ion-selective electrodes.

The complexation of six tricyclic antidepressant drugs [amitriptylin (AMN), nortriptylin (NRN), imipramin (IMN), doxepin (DXN), protriptylin (PTN), and maprotilin (MPN)] with alpha- and beta-cyclodextrins (CDs) using ion-selective electrodes (ISEs) as drug ion sensors is described. Binding parameters were calculated by nonlinear fitting of the model described by the Scatchard equation, to the experimental data of a titration of a CD solution with the ion of interest. One binding site (the CD cavity) was found in all cases with both CDs. The calculated association constants at 25 degrees C using CD concentrations in the range of 0.0100-0.0010 M, varied from 4.81 x 10(3) M-1 (MPN) to 23.9 x 10(3) M-1 (AMN) in the case of beta-CD and from 50 M-1 (DXN) to 123 M-1 (MPN) in the case of alpha-CD. The precision for the estimation of the binding parameters was 0.1-5.0% (within-run RSD%) and 8-10% (between-run RSD%; n = 3). The complexation of the drugs with beta-CD was also examined as a function of temperature in the range of 5-37 degrees C; it was found to decrease by increasing temperature. Van't Hoff analysis gave good correlations (r greater than or equal to 0.989) for all drug ions studied. The estimates of the thermodynamic parameters indicate that the formation of inclusion complexes is enthalpy driven. A compensation plot based on the thermodynamic parameters delta H and delta S resulted in a linear relationship, which is indicative of a common type of force involved in the complexation of drugs to beta-CD.

Antidepressive Agents, Tricyclic↗