ABSTRACT
A simple method using HPLC-DAD was developed for the determination of fluoroquinolones in human urine including ciprofloxacin (CIPRO), enrofloxacino (ENRO), marbofloxacino (MARBO) and norfloxacin (NOR). In addition, it was studied the extraction of fluoroquinolones in human urine samples using pipette tip-based molecularly imprinted polymers solid phase extraction (PT-MIPs-SPE). With the goal of finding the best procedure for extraction of four fluoroquinolones in human urine, several parameters that are likely to affect the efficiency of extraction during sample preparation, including the washing solvent, type and volume of eluent, amount of material, the volume of the sample, pH and the ionic strength were systematically optimized. Chromatographic separations of fluoroquinolones were hit within 10min using a Synergi(®) C18 (250×4.6mm, 4µm) column and mobile phase consisting of water (10mM of phosphoric acid, the pH adjusted at 3.29 with triethylamine) : acetonitrile (85.7: 14.3, v/v) at a flow rate of 1.5mLmin(-1). Detection was performed at 290nm. The average extraction recoveries/standard deviation relative to ENRO, CIPRO, NOR and MARBO were 96.40±5.51%, 42.47±4.81%, 41.82±7.99% and 87.49±4.70, respectively. The method was liner from 39 to 1260ngmL(-1) for each fluoroquinolone with correlation coefficient of 0.9904, 0.9910, 0.9914 and 0.9919, to ENRO, CIPRO, NOR and MARBO, respectively. The assays of within-day and between-day precision and accuracy for all analytes were studied at three concentration levels and were lower than 15%. The method was successfully employed in a preliminary cumulative urinary excretion study after administration of CIPRO to a healthy volunteer.
Subject(s)
Fluoroquinolones/urine , Molecular Imprinting/methods , Solid Phase Extraction/methods , Chromatography, High Pressure Liquid , Drug Stability , Fluoroquinolones/chemistry , Fluoroquinolones/isolation & purification , Humans , Limit of Detection , Linear Models , Reproducibility of ResultsABSTRACT
A phosphorimetric method was developed to enable the determination of enrofloxacin using photochemical derivatization which was used to both improve detection limits and to minimize the uncertainty of measurements. Phosphorescence was induced on cellulose containing TlNO(3). Absolute limit of detection at the ng range and linear analytical response over three orders of magnitude were achieved. A metrological study was made to obtain the combined uncertainty value and to identify that the precision was mainly affected by the changing of substrates when measuring the signal from each replicate. Pharmaceutical formulations containing enrofloxacin were successfully analyzed by the method and the results were similar to the ones achieved using a HPLC method. A solid phase extraction on an acrylic polymer was optimized to separate enrofloxacin from interferents such as diclofenac and other components from biological matrices, which allowed the successful use of the method in urine analysis.
Subject(s)
Acrylates/chemistry , Fluoroquinolones/analysis , Luminescent Measurements/methods , Polymers/chemistry , Solid Phase Extraction/methods , Temperature , Absorption , Cellulose/chemistry , Chromatography, High Pressure Liquid , Diclofenac/chemistry , Enrofloxacin , Fluoroquinolones/chemistry , Fluoroquinolones/urine , Humans , Molecular Imprinting , Pharmaceutical Preparations/chemistry , Solutions , Spectrometry, FluorescenceABSTRACT
Synchronous fluorescence spectra measured in a flow-injection system with double pH gradient modulation constitute a new second-order signal which is herein studied for the quantitative determination of three fluoroquinolone antibiotics in spiked human urine samples. Because calibration is done using aqueous solutions of each of the three analytes ciprofloxacin, norfloxacin and ofloxacin, the fluorescent urine background makes it necessary to achieve the second-order advantage. Several second-order multivariate calibration algorithms were evaluated for this purpose: parallel factor analysis, unfolded and multiway partial least-squares with residual bilinearization, and multivariate curve resolution-alternating least-squares. The best analytical figures of merit, a root mean square error of 4-6 mg L(-1) (corresponding to a relative error of 4-6% for a calibration range from 0 to 200 mg L(-1) for each analyte), and a limit of detection of 4 mg L(-1) were obtained using partial least-squares (in the specific unfolded version) combined with residual bilinearization. Reasons for the improved success of this latter technique are provided on the basis of the analysis of simulated second-order data.
Subject(s)
Factor Analysis, Statistical , Flow Injection Analysis/methods , Fluorescence , Fluoroquinolones/chemistry , Nucleic Acid Denaturation/drug effects , Spectrometry, Fluorescence/methods , Algorithms , Anti-Bacterial Agents/chemistry , Anti-Bacterial Agents/urine , Ciprofloxacin/chemistry , Ciprofloxacin/urine , Clinical Laboratory Techniques , Fluoroquinolones/urine , Humans , Hydrogen-Ion Concentration , Least-Squares Analysis , Norfloxacin/chemistryABSTRACT
Las fluoroquinolonas constituyen un grupo de antibióticos, ampliamente utilizado en infeccionesdel tracto urinario (ITU), por su excelente actividad frente a las enterobacterias y por su vía deeliminación. El objetivo de este estudio es determinar la resistencia a las fluoroquinolonas:ciprofloxacina, levofloxacina y gatifloxacina en bacilos gramnegativos aislados de ITU. Se incluyeronen el estudio todos los bacilos gramnegativos, aislados de pacientes adultos con ITU, queconcurrieron en forma consecutiva al laboratorio San Roque, desde junio de 2005 a marzo de 2006.Para determinar la resistencia, se utilizó el método de difusión en agar siguiendo normasestandarizadas del NCCLS. De las 380 cepas aisladas el 81,7% correspondió a Escherichia coli,11,6% Klebsiella pneumoniae, 3,4% Proteus mirabilis, 1% Enterobacter aerógenes, 0,8%Enterobacter cloacae, 0,5% Citrobacter koseri, 0,5% Citrobacter freundii y 0,5% Klebsiella oxytoca.El 17,6% de todos los aislamientos, fue resistente a todas las fluoroquinolonas ensayadas (58cepas de E. coli, 7 cepas de K. pneumoniae y 2 de E. cloacae). La resistencia de E. coli a las tresfluoroquinolonas fue del 18,7%, y de K. pneumoniae 15,9%. Una cepa de E. coli, sensible agatifloxacina, presentó sensibilidad intermedia a ciprofloxacina y levofloxacina. Las tresfluoroquinolonas testadas presentaron igual actividad frente a bacilos gramnegativos, aislados deITU. Esta resistencia es relativamente alta, debido a que la resistencia a un antibiótico a serutilizado en forma empírica, no debe superar el 20%