Biomedical subjects

E Roets

Publications and source records attributed to E Roets.

At least 37 records · Page 2Linked to original sources

PubMed2000

Capillary electrophoresis analysis of gentamicin sulphate with UV detection after pre-capillary derivatization with 1,2-phthalic dicarboxaldehyde and mercaptoacetic acid.

A selective, sensitive, and rapid pre-capillary derivatization method for determination of the multicomponent aminoglycoside antibiotic gentamicin is described. The derivatization reagents 1,2-phthalic dicarboxaldehyde and mercaptoacetic acid were used and the thioisoindole derivative was UV detected at 330 nm. A central composite experimental design was performed to optimize selectivity and derivatization conditions. Baseline separation of gentamicin C1, C1a, C2, C2a, C2b, sisomicin and several minor components was achieved with a background electrolyte containing 30 mM sodium tetraborate, 7.5 mM beta-cyclodextrin and 12.5% (v/v) methanol at pH 10. Quantitative analysis was performed and illustrated the potential use of capillary electrophoresis for the identification and quantitation of gentamicin as an alternative to methods prescribed in the United States Pharmacopeia and European Pharmacopoeia.

Electrophoresis, Capillary↗

PubMed2000

Analysis of doxycycline by capillary electrophoresis. Method development and validation.

An optimized capillary electrophoresis method for the analysis of doxycycline is described. The influence of methanol as organic modifier, buffer pH, buffer concentration, capillary length, column temperature, Triton X-100 and methyl-beta-cyclodextrin was systematically investigated. A central composite design was performed in order to optimize the method. The optimal separation conditions were: capillary, uncoated fused-silica [40 cm (32 cm effective length) x 50 microm I.D.]; background electrolyte, a solution of 145 mM sodium carbonate and 1 mM EDTA brought to pH 10.3-methanol (89:11, v/v); temperature, 15 degrees C; voltage, 12 kV. The method showed good selectivity, repeatability, linearity and sensitivity. Six commercial samples were quantitatively analyzed. The results were compared with those established by the liquid chromatography method from the European Pharmacopoeia.

Doxycycline↗

PubMed2000

Analysis of polymyxin B sulfate by capillary zone electrophoresis with cyclodextrin as additive. Method development and validation.

A capillary zone electrophoresis method for analysis of polymyxin B sulfate is described. In this method, triethanolamine (TEA)-phosphate buffer at pH 2.5 was employed to reduce the adsorption of analyte onto the capillary wall. Methyl-beta-cyclodextrin (M-beta-CD) and 2-propanol (IPA) were found to be necessary for selectivity enhancement. In order to optimize the method and to control its robustness, a central composite design was performed with four parameters, i.e. concentration of M-beta-CD, TEA, IPA and buffer pH. The optimal separation conditions were as follows: capillary, 55 cm (50 microm I.D., 47 cm effective length); 130 mM TEA-phosphate buffer (pH 2.5) containing 5 mM M-beta-CD and 5% IPA; 24 kV (51 microA) applied voltage; column temperature, 20 degrees C. Further, linearity and limits of detection quantification were examined. Three commercial samples were analyzed quantitatively.

Cyclodextrins↗

PubMed2000

Separation of erythromycin and related substances on base-deactivated reversed-phase silica gel columns.

An official liquid chromatographic method for the analysis of erythromycin and related substances, which is based on a polymer reversed-phase, is described in the European Pharmacopoeia and in the United States Pharmacopeia. The pH of the mobile phase used in this system is 9.0. Recent advanced technology has led to the introduction of a new generation of silica-based reversed-phase column packings, which are claimed to be much more stable towards bases. They are useful for the analysis of basic compounds. Studies to verify the separation of erythromycin and related substances on Hypersil BDS C18, Luna C18(2), Inertsil ODS-2 and Supelcosil ABZ+ have been performed and the results are presented. It is shown that these base-deactivated phases give a better sensitivity and selectivity towards erythromycins than the polymer phase, provided that an adapted mobile phase is used. This is the first liquid chromatographic method described for the separation of erythromycin D from erythromycin A.

Chromatography, High Pressure Liquid↗

PubMed2000

Liquid chromatography of polymyxin B sulphate.

A reversed-phase liquid chromatography method for analysis of polymyxin B sulphate is described. The method uses a YMC-Pack Pro, C18, 5 microm, 250x4.6 mm I.D. column maintained at 30 degrees C. The mobile phase comprises acetonitrile-sodium sulphate (0.7%, m/v)-phosphoric acid (6.8%, v/v dilution of 85%, m/m phosphoric acid)-water (22.25:50:5:22.75) at a flow-rate of 1.0 ml/min. Detection was by UV at 215 nm. The method is able to resolve polymyxin B1, the major component, from more than thirty other components present in the complex. Robustness was evaluated by performing a full-factorial design experiment. The method showed good selectivity, repeatability, linearity and sensitivity.

Anti-Bacterial Agents↗

PubMed2000

Investigation of unknown related substances in commercial erythromycin samples with liquid chromatography/mass spectrometry.

A selective reversed phase liquid chromatography/mass spectrometry (LC/MS(n)) method is described for the identification of erythromycin impurities and related substances in commercial erythromycin samples. Mass spectral data are acquired on a LCQ ion trap mass spectrometer equipped with an electrospray interface operated in positive ion mode. The LCQ is ideally suited for identification of impurities and related substances because it provides on-line LC/MS(n) capability. Compared with UV detection, this hyphenated LC/MS(n) technique provides as a main advantage efficient identification of novel substances without time-consuming isolation and purification procedures. Using this method four novel related substances were identified in commercial samples.

Animals↗

PubMed2000

Analysis of colistin sulfate by capillary zone electrophoresis with cyclodextrins as additive.

A method for the quantitative analysis of colistin sulfate by capillary zone electrophoresis is described. Since colistin components have five free amino groups, they tend to adsorb onto the capillary wall and cause peak tailing. It was found that triethanolamine (TEA)-phosphate buffer at pH 2.5 was useful to reduce such adsorption. Methyl-beta-cyclodextrin (M-beta-CD) and 2-propanol (IPA) were found necessary for selectivity enhancement. In order to optimize the separation parameters and predict the method robustness, a central composite design was performed including three variables, namely concentration of M-beta-CD, TEA, and IPA. The effects of capillary length and applied voltage on separation were also investigated. The optimal conditions established were: 140 mM TEA-phosphate buffer containing 5 mM M-beta-CD and 6% v/v IPA, a capillary with 55 cm total length (50 microm inner diameter, 47 cm from inlet to detection window) and 24 kV applied voltage. The method was found to be robust when the variables were changed in the following range: 4-6 mM M-beta-CD, 5-7% v/v IPA, and 130-150 mM TEA. Further, the linearity, limit of detection (LOD), and limit of quantitation (LOQ), as well as repeatability for both colistin A and B were examined and three commercial samples were quantitatively analyzed.

1-Propanol↗

PubMed2000

Analysis of tobramycin by liquid chromatography with pulsed electrochemical detection.

The determination of tobramycin by liquid chromatography using a column packed with poly(styrene-divinylbenzene) and pulsed electrochemical detection on a gold electrode is described. The mobile phase consisted of an aqueous solution containing 52 g/l of sodium sulfate, 1.5 g/l of sodium octanesulfonate and 50 ml/l of a 0.2 M phosphate buffer pH 3.0. The total time of analysis was not more than 30 min. The effects of the different chromatographic parameters on the separation were also investigated. When a number of commercial samples of tobramycin was analyzed using this method, nine different components were separated, five of which are of unknown identity.

Anti-Bacterial Agents↗

PubMed2000

A stability study of ticlopidine products from 18 countries.

The results of the stability study of ticlopidine formulations (250 mg and 100 mg) show that products available in many countries worldwide exhibit different stability characteristics. Stability testing under the International Conference on Harmonization (ICH) accelerated test conditions (40 degrees C/75% relative humidity [RH], 3 and 6 months) was performed on a total of 43 products obtained from 18 countries. The samples were visually examined for physical change and analyzed for their content of degradation products, remaining ticlopidine, and in vitro dissolution characteristics (in the case of tablets). Only 6 (16%) of all the samples submitted to this study had a good stability profile. Their appearance remained unchanged during the study; assay results were between 95% and 100%; their impurity content did not exceed 0.25%; and in the dissolution test, at least 75% of ticlopidine was dissolved after 30 min. Three samples had excellent dissolution properties and showed a very high purity level (viz. 21, 40, and 43) over the course of the study.

Chromatography, Liquid↗

PubMed1999

Micellar electrokinetic capillary chromatography of macrolide antibiotics. Separation of tylosin, erythromycin and their related substances.

The separation of tylosin by micellar electrokinetic capillary chromatography with a mixed micelle system is described. Good selectivity was obtained with sodium phosphate buffer (80 mM, pH 7.5) containing 20 mM sodium cholate and 7 mM cetyltrimethylammonium bromide (CTAB). This method permits tylosin to be separated from its closely related substances within 15 min. The influences of type of buffer, buffer pH, the concentrations of sodium cholate and CTAB were investigated. The robustness of the method was examined for tylosin by means of a full-fraction factorial design. Quantitative results are presented. Using a similar buffer system (80 mM sodium phosphate, pH 6.0, 20 mM sodium cholate and 5 mM CTAB), separation of erythromycin and its main related substances was also obtained. However, detection sensitivity and resolution are not sufficient for analysis of related substances in erythromycin commercial samples.

Anti-Bacterial Agents↗

PubMed1999

Micellar electrokinetic capillary chromatography for the separation of cefalexin and its related substances.

Micellar electrokinetic capillary chromatography (MEKC) was examined for analysis of cefalexin and its related substances. Good selectivity was obtained with two different buffer solutions: a sodium acetate buffer (50 mM, pH 5.25) containing sodium dodecyl sulfate (50 mM SDS) or sodium phosphate buffer (40 mM, pH 7.0) containing 100 mM SDS. Both methods permit cefalexin to be completely separated from its ten related substances within 20 min. The robustness of the method, using pH 5.25 acetate buffer, was examined by means of a full-fraction factorial design to test the influence of buffer pH, concentration of SDS and buffer concentration. The parameters for validation such as linearity, precision, limit of detection and limit of quantitation are also reported. The results show that method 1 is suitable for the analysis of cefalexin.

Cephalexin↗

PubMed1999

Analysis of spiramycin by capillary electrophoresis.

The development and validation of an analytical method for the determination of spiramycin I in the presence of its related substances by capillary electrophoresis is shown. The separation, performed in a phosphate buffer (80 mM, pH 7.5) containing 12 mM cetyltrimethylammonium bromide (CTAB) and 20 mM sodium cholate, with a 50 microm ID and 44 cm long fused-silica capillary (36 cm effective length), applying a voltage of 12 kV (l approximately 80 microA), at 25 degrees C, is achieved in 15 min. Good selectivity among spiramycin I and its related substances was obtained. The influence of the buffer pH, and of the CTAB and sodium cholate concentrations was investigated. The method robustness, examined by means of a full-fraction factorial design, shows that it can be used within the limits set for the three parameters that were investigated. The method is linear (r = 0.9992) and precise (day-to-day corrected peak area repeatability, n = 18, relative standard deviation = 1.3%). The limits of detection and quantitation are 7 pg (0.025%) and 22 pg (0.08%), respectively, relative to a 2 mg/mL solution.

Aminoglycosides↗

PubMed1999

Interlaboratory study comparing the microbiological potency of spiramycins I, II and III.

An interlaboratory study has been performed to determine the relative potencies of spiramycins (SPMs) I, II and III by diffusion or/and turbidimetric assays with Bacillus subtilis or Staphylococcus aureus as the test organisms. Six laboratories from three countries participated. Experimental procedures were according to the European Pharmacopoeia, 3rd ed. The activity of SPM I is markedly higher than that of SPM II and III. By diffusion, the activities of SPM II and III relative to SPM I were found to be 57 and 72%, respectively. The interlaboratory relative standard deviations (RSD) varied from 3.6 to 16.3%. By turbidimetry, the activities of SPM II and III relative to SPM I were found to be 45 and 52%, respectively. The interlaboratory RSD values varied from 2.6 to 7.7%. The results of the study were analyzed according to the ISO 5725-2 guidelines to determine the repeatability, the between-laboratory and the reproducibility variances of both methods.

Aminoglycosides↗

PubMed1999

Liquid chromatography method for separation of clindamycin from related substances.

A reversed-phase liquid chromatography method has been developed for the separation of clindamycin from 7-epiclindamycin, clindamycin B, lincomycin, lincomycin B, 7-epilincomycin and other impurities of unknown identity. The method uses a Hypersil ODS, 5 microm, 250 x 4.6 mm i.d. column maintained at 45 degrees C. The mobile phase comprises acetonitrile phosphate buffer (1.35% v/v phosphoric acid, adjusted to pH 6.0 with ammonium hydroxide)-water (35:40:25, v/v) at a flow rate of 1.0 ml/min. UV detection is performed at 210 nm. The method was tested on several C-18 columns and showed good robustness. Robustness was further evaluated by performing a full-fraction factorial design experiment. The method showed good selectivity, linearity, and repeatability. It is also suitable for analysis of clindamycin formulations.

Anti-Bacterial Agents↗

PubMed1999

In-house packing and testing of capillaries for capillary electrochromatography using simple equipment.

The feasibility of producing packed capillaries for capillary electrochromatography (CEC) is evaluated. Emphasis is put on the fact that only material was used that is already available in any LC/CE orientated laboratory. An experimental set-up is developed for filling the capillaries by use of an ordinary LC pump, and frits are sintered with a glowing resistance wire, fed by a d.c. power supply. Electrochromatography was carried out in an in-house built capillary electrophoresis apparatus, without pressurizing the vials. Under these conditions, the capillaries performed well, producing up to 190,000 plates per meter. Bubble formation did not appear, on condition that the mobile phase was thoroughly helium degassed. Even at ambient temperature, electrophoresis obeyed Ohm's law up to a voltage of 30 kV, proving that Joule heating was not a major concern.

Anti-Inflammatory Agents↗

PubMed1999

Nested designs in ruggedness testing.

Nested designs were performed in order to execute a ruggedness test according to the United States Pharmacopeia definition for ruggedness, in which mainly non-procedure related factors are examined. Several nested designs have been executed on a high performance liquid chromatography assay to determine tetracycline and related substances in bulk samples of tetracycline. Factors such as different laboratories, analysts, instruments, columns, days and batches were examined. The interpretation methods described in the literature were found to cause problems. In these methods the variances of the examined factors are estimated from the calculated mean square values and from the equation for the expected mean squares. Very frequently, negative variance estimates were obtained. Their absolute values were found to be dependent on the influence of the factor examined below it in the design, on the examined response. Therefore an alternative interpretation method for nested designs, based on pooled variances, was proposed and found to be appropriate to use for ruggedness testing purposes. Both approaches, the one from the literature and the one proposed here, were tested on simulated data coming from a nested design with four factors and on the experimentally measured data.

Analysis of Variance↗

PubMed1999

Liquid chromatographic analysis of dihydrostreptomycin sulfate.

The analysis of dihydrostreptomycin sulfate using a column packed with base deactivated reversed phase silica gel and ultraviolet detection at 205 nm is described. The mobile phase consists of an aqueous solution containing 4 g/l of sodium sulfate, 1.5 g/l of sodium octanesulfonate, 100 ml/l of acetonitrile and 50 ml/l of a 0.2-M phosphate buffer at pH 3.0. The method allows separation of streptidine, dihydrostreptomycin B, streptomycin, dihydrostreptomycin and deoxydihydrostreptomycin, as well as some other components which were not identified. The total time of analysis is 55 min. The effects of the different chromatographic parameters on the separation were also investigated. A number of commercial samples were analyzed using this method.

Analysis of Variance↗

PubMed1999

Evaluation of the role of endotoxin and cortisol on modulation of CD18 adhesion receptors in cows with mastitis caused by Escherichia coli.

OBJECTIVE: To determine the effect of mastitis caused by Escherichia coli on expression of CD18 cell surface receptors and to evaluate the involvement and regulation of receptors by lipopolysaccharide (LPS) and cortisol. ANIMALS: 11 clinically normal lactating Holstein-Friesian cows. PROCEDURE: Binding of CD18 monoclonal antibodies to neutrophils was studied, using flow cytometry, before and after intramammary inoculation of E. coli organisms. Effect of LPS and cortisol on expression of adhesion receptors was investigated, using a whole-blood model. RESULTS: Expression of CD18 adhesion receptors on bovine neutrophils increased 35% by 12 hours after intramammary inoculation of E. coli. By 24 hours after inoculation, the number of receptors had returned to control values. High cortisol concentrations (100 nmol/L) were seen 12 to 18 hours after inoculation. Addition of LPS to blood induced a 30% increase in the number of CD18 receptors, and maximal number of receptors was expressed at an LPS concentration of 0.1 ng/ml. A decrease in the number of CD18 receptors was induced by incubation with cortisol or dexamethasone before challenge-exposure with LPS. CONCLUSIONS: An increase in the number of CD18 receptors on neutrophils is mediated by local production of LPS. Subsequent endogenous release of cortisol may prevent additional increases in the number of receptors. CLINICAL RELEVANCE: During acute mastitis caused by E. coli, there is an increase in the number of CD18 receptors on circulating neutrophils. Cortisol induces a decrease in the number of CD18 receptors, probably modulating the acute inflammatory response in mammary glands of lactating cows.

Animals↗