High-performance liquid chromatographic assay of cefotaxime,desacetylcefotaxime and ceftriaxone in rat plasma

A simple and selective high-performance liquid chromatographic method is described for the analysis of the cephalosporins cefotexime (CXM), desacetylcefotaxime (DACXM) and ceftriaxone (CFX) in rat plasma. Plasma was deproteinized with methanol, and the supernatant was directly injected into the chromatograph and monitored at 254 nm. For determination of the unbound drugs, a centrifugal ultrafiltration method was employed. The calibration curves were linear (r=0.999) from 2.5 to 500 μg/ml; the detection limits were 100 ng/ml for DACXM and 250 ng/ml for CXM and CFX. The method was not interfered with by other plasma components, nor by barbital sodium or caffeine, and has been applied to study the pharmacokinetics of the cephalosporins in rats.     

Determination of cefotaxime and desacetylcefotaxime in plasma and urine by high-performance liquid chromatography

A high-performance liquid chromatographic method is described for the analysis of the anti-bacterial agent cefotaxime and desacetylcefotaxime in physiological fluids. Plasma or serum samples were mixed with chloroform—acetone to remove proteins and most lipid material. The aqueous phase was then freeze-dried, reconstituted in mobile phase and chromatographed on a reversed-phase column using UV detection at 262 nm. Urine was analysed directly after centrifugation to remove particulate matter. The detection limit was 0.5–1.0 μg/ml for serum and 5 μg/ml for urine. The method has been applied to the analyses of cefotaxime and desacetylcefotaxime in plasma, serum, urine, cerebrospinal fluid, saliva, and pus from infected wound secretions. Two additional metabolites, which are lactones, in which the β-lactam ring has been opened, could be separated by this method.     

Simple liquid chromatographic method for the determination of cefotaxime in human and rat plasma

A high-performance liquid chromatographic method with ultraviolet (UV) detection was developed for measuring cefotaxime in rat and human plasma. The method used direct injection of the plasma supernatant after deproteinization with 70% perchloric acid. Degradation of cefotaxime in acidic medium was retarded by adding phosphate buffer before centrifuging the sample. The mobile phase was 0.05 M aqueous ammonium acetate-acetonitrile-tetrahydrofuran (87:11:2, v/v) adjusted to pH 5.5. Analysis was run at a flow-rate of 1.0 ml/min, and a detection wavelength of 254 nm was used. The method has a quantification limit of 0.20 microgram/ml. The within- and between-day coefficients of variation and accuracy values were less than 8% and +/-3%, respectively, while the recovery values were greater than 87% over the concentration range tested (0.20-50 microgram/ml). The speed, sensitivity, specificity and reproducibility of this method make it particularly suitable for the routine determination of cefotaxime in human plasma. Moreover, only a relatively small sample plasma volume (100 microliter) is required, allowing this method to be applied to samples taken from neonates.     

Liquid chromatographic determination of irbesartan in human plasma

A simple and sensitive method was developed for determination of irbesartan by liquid chromatography with fluorescence detection. Irbesartan and losartan (I.S.) in human plasma were extracted using diethyl ether:dichloromethane (7:3, v/v) followed by back extraction with 0.05 M sodium hydroxide. Neutralized samples were analyzed using 0.01 M potassium dihydrogen phosphate buffer (containing 0.07% triethylamine as peak modifier, pH was adjusted with orthophosphoric acid to pH 3.0) and acetonitrile (66:34, v/v). Chromatographic separation was achieved on an ODS-C-18 column (100 mm x 4.6 mm i.d., particle size 5 microm) using isocratic elution (at flow rate 1.25 ml/min). The peak was detected using a fluorescence detector set at Ex 259 nm and Em 385 nm, and the total time for a chromatographic separation was approximately 13 min. The validated quantitation ranges of this method were 15-4000 ng/ml with coefficients of variation between 0.75 and 12.53%. Mean recoveries were 73.3-77.1% with coefficients of variation of 3.7-6.3%. The between- and within-batch precision were 0.4-2.2% and 0.9-6.2%, respectively. The between- and within-batch relative errors (bias) were (-5.5) to 0.9% and (-0.6) to 6.9%, respectively. Stability of irbesartan in plasma was >89%, with no evidence of degradation during sample processing and 60 days storage in a deep freezer at -70 degrees C. This validated method is sensitive and simple with between-batch precision of <3% and can be used for pharmacokinetic studies.     

Liquid chromatographic determination of amodiaquine in human plasma

A normal-phase high-performance liquid chromatographic method using dichloromethane- methanol-1M perchloric acid (100:10:0.9, v/v/v) at a flow rate of 1.0 ml min(-1) on a LiChrospher Si column with UV (254 nm) detection has been developed for the determination of amodiaquine and its metabolites desethyl amodiaquine and bisdesethyl amodiaquine in plasma. The limit of quantification was 5 ng ml(-1). Mean within-day and day-to-day coefficients of variation (CV) were 4.10 and 6.27% for amodiaquine, 3.43 and 4.80% for desethyl amodiaquine and 3.53 and 5.23% for bisdesethyl amodiaquine, respectively. Mean extraction recovery of amodiaquine, desethyl amodiaquine and bisdesethyl amodiaquine from plasma were 82.48, 74.50 and 69.65%, respectively. Chloroquine and its metabolite desethyl chloroquine, quinine, sulfadoxine and primaquine do not interfere in the detection of amodiaquine, desethyl amodiaquine and bisdesethyl amodiaquine in plasma.     

Liquid chromatographic method for the determination of ticlopidine in human plasma

A simple high-performance liquid chromatographic method for determination of ticlopidine in human plasma using ultra violet detection was developed. The separation of the investigated compound and internal standard was achieved on a C₁₈ BD column with a 0.01 M potassium dihydrogen phosphate buffer (pH 4)–acetonitrile–methanol (20:40:40, v/v) mobile phase. The detection was performed at 215 nm. The compounds were isolated from plasma by Bond Elut C₁₈ solid-phase extraction, the mean absolute recovery was 84.9%. The limit of quantitation was 10 ng ml⁻¹, the limit of detection was 5 ng ml⁻¹. The bioanalytical method was validated with respect to linearity, within- and between-day accuracy and precision, system suitability and stability. All validated parameters were found to be within the internationally required limits. The developed analytical method for ticlopidine was found to be suitable for application in pharmacokinetic studies and human drug monitoring.     

Liquid chromatographic method for the determination of rizatriptan in human plasma

A high-performance liquid chromatographic (HPLC) method with fluorescence detection has been developed for the determination of rizatriptan in human plasma. Following a single-step liquid-liquid extraction with methyl tertiarybutyl ether, the analytes were separated using a mobile phase consisting of 0.05% (v/v) triethylamine in water (adjusting to pH 2.75 with 85% phosphoric acid) and acetonitrile (92:8, v/v). Fluorescence detection was performed at an excitation wavelength of 225nm and an emission wavelength of 360nm. The linearity for rizatriptan was within the concentration range of 0.5-50ng/ml. The intra- and inter-day precisions of the method were not more than 8.0%. The lower limit of quantification (LLOQ) was 0.5ng/ml for rizatriptan. The method was sensitive, simple and repeatable enough to be used in pharmacokinetic studies.     

Liquid chromatographic method for the determination of rosiglitazone in human plasma

A robust, accurate and sensitive high-performance liquid chromatographic method for the determination of rosiglitazone (I) in human plasma has been developed. Pioglitazone (II) was used as internal standard. Both I and II are extracted from plasma using a liquid-liquid extraction procedure. Isocratic separation of I and II is carried out using a reversed-phase Zorbax SB C(18), 15-cm column with mobile phase consisting of methanol and a mixed phosphate buffer (10 mM monobasic sodium phosphate and dibasic sodium phosphate, pH adjusted to 2.6 with ortho-phosphoric acid) in the ratio 30:70 (v/v) and quantified by UV detection at 245 nm. Linearity was established over the range 5-1250 ng/ml using 1 ml human plasma. The method is specific, the endogenous components in plasma do not interfere with I and II. C.V. (%) of intra-day samples is less than 5.0% at four concentrations tested namely 5, 10, 500 and 1000 ng/ml. Similarly, over the same nominal concentrations, the precision of inter-day (5 days) samples also results in C.V. (%) less than 5.0%. The recoveries of I and II from human plasma were about 79 and 60%, respectively. This method can be used for routine clinical monitoring of I.     

Liquid chromatographic determination of oxcarbazepine and its metabolites in plasma of epileptic patients after solid-phase extraction

A method based on high-performance liquid chromatography with UV detection in combination with solid-phase extraction for sample pretreatment has been developed for the simultaneous analysis of the antiepileptic drug oxcarbazepine and its main metabolites in human plasma. The extraction of the analytes from plasma samples was carried out by means of a selective solid-phase extraction procedure using hydrophilic-lipophilic balance cartridges. The separation was obtained on a reversed-phase column (C(18), 150x4.6 mm I.D., 5 micrometer) using a phosphate buffer-acetonitrile-methanol-triethylamine mixture (final apparent pH* 3.5) as the mobile phase. Under these chromatographic conditions, oxcarbazepine and its metabolites 10,11-dihydro-10-hydroxycarbamazepine, 10,11-dihydro-10,11-dihydroxycarbamazepine and the internal standard are baseline separated in less than 9 min. The extraction yield values were >94% for all analytes and the precision, expressed by the RSD%, was in the low percentage range. For the entire method, including sample pre-treatment and HPLC determination, the linearity of the calibration lines, expressed by the linear correlation coefficient, was better than 0.995; the limit of quantitation was 15 ng ml(-1). The method was applied to plasma samples from patients undergoing chronic treatment with oxcarbazepine, both in monotherapy and in polytherapy. Based on the analytical parameters precision, accuracy, limit of quantitation and analysis time the method is suitable for routine application in therapeutic drug monitoring.     

Liquid chromatographic determination of florfenicol in the plasma of multiple species of fish

A simple method was developed for determining florfenicol concentration in a small volume (250 micro l) of plasma from five phylogenetically diverse species of freshwater fish. Florfenicol was isolated from the plasma matrix through C(18) solid-phase extraction and quantified by reversed-phase high-performance liquid chromatography with UV detection. The accuracy (84-104%), precision (%RSD相似文献   

Liquid chromatographic method for the quantitative determination of Nepsilon-carboxymethyllysine in human plasma proteins

The modification of the lysine moieties of proteins to Nepsilon-carboxymethyllysine (CML) is supposed to play a major role in the development of long-term complications in patients with diabetes mellitus. This paper presents an analytical method for the quantitative determination of CML in plasma proteins, which could be used for studying the development of diabetic complications. The method is based on isolating proteins from plasma by precipitation with trichloroacetic acid and hydrolysing these under acidic conditions (6M hydrochloric acid at 110 degrees C for 20 h) to the individual amino acids. After hydrolysis, CML is derivatised along with the other amino acids to 9-fluorenylmethoxycarbonyl (FMOC) derivatives, which are subsequently separated by reversed-phase column liquid chromatography using a 150 mm x 4.6 mm C8 column and a mobile phase of 25 mM potassium phosphate buffer (pH 2.0) and acetonitrile (80:20 (v/v)) and detected using fluorescence detection (excitation at 260 nm and emission at 310 nm). Quantification of the protein-bound CML content of a plasma sample is achieved using standard addition. The impact of several aspects of the sample preparation and chromatography on method performance is discussed. Method evaluation results are reported and show that this method is capable of determining CML with good accuracy and precision (below 10%) in the relevant concentration range (1-10 microg/ml), with a limit of detection of 0.2 microg/ml.     

Liquid chromatographic determination of total homocysteine in blood plasma with photometric detection

A rapid and sensitive method for quantification of homocysteine total forms and glutathione levels in blood plasma via HPLC was developed. Dithiotreitol as a water soluble agent has been used as a reductant for both protein and nonprotein disulphides. Dithiotreitol reacts with the mixed disulphides under 60 degrees C treatment within 10 min. Reduced aminothiols and homocystein were easily derivated with 5,5'-dithiobis-(2-nitrobenzoic acid) and the resultant ultraviolet absorbance within 330 nm was detected by the HPLC method. The concentration of total plasma homocysteine was significantly higher in groups of patients: with the end stage of renal disease: 45.5+/-40.9 micromol/l (n=79), with cerebral vascular disorders 12.3+/-7.0 micromol/l (n=65), and with coronary atherosclerosis 15.4+/-10.9 micromol/l (n=15) than that in healthy subjects (6.2+/-1.74 micromol/l, n=20). Some major advantages of the method include: simultaneous measurement of both total homocysteine and total glutathione, no loss of oxidized form during processing of blood plasma for aminothiols measurement, use of protein-bound aminothiols solution as a calibrator.     

Gas chromatographic determination of low concentrations of benzoic acid in human plasma and urine

A method for the determination of benzoic acid down to concentrations of 10 ng/ml in plasma or urine is described. After addition of an internal standard, benzoic acid is extracted at acid pH into diethyl ether. Both compounds are derivatized with pentafluorobenzyl bromide. The derivatives are determined by gas chromatography using a ⁴³Ni electron-capture detector. Hippuric acid is hydrolysed in plasma and urine and total benzoic acid is determined by the same technique.     

Liquid chromatographic determination of sodium mercaptoundecahydrododecaborate in rat urine and plasma after precolumn derivatization

A high-performance liquid chromatographic (HPLC) method was developed for the determination of disodium mercaptoundecahydrododecaborate (BSH) in biological fluids. Monobromobimane was used as a precolumn derivatizing agent. A stable derivative was obtained. The derivative was separated on a C₁₈ column using reversed-phase ion-pairing chromatography and detected by a spectrophotometric detector at 373 nm. The detection limit was 200 ng/ml (0.1 ppm boron). Calibration curves were prepared for rat urine and plasma samples. The calibration curves were linear in the range of 1 μg/ml to 100 μg/ml for urine samples and 0.2 μg/ml to 50 μg/ml for plasma samples.     

Liquid chromatographic determination of the enantiomers of ibuprofen in plasma using a chiral AGP column

A reversed-phase high-performance liquid chromatographic method has been developed for the determination of the R- and S-enantiomers of ibuprofen. The enantiomers and the internal standard 4-pentylphenylacetic acid are extracted from plasma, separated and quantified on a Chiral-AGP column using ultraviolet detection. The simplicity, sensitivity and precision of the method makes it convenient for use in pharmacokinetic studies.     

Simultaneous quantification of cefotaxime, desacetylcefotaxime, ofloxacine and ciprofloxacine in ocular aqueous humor and in plasma by high-performance liquid chromatography

Cefotaxime, given intravenously, is currently used as a broad-spectrum antibiotic for prophylaxis of intra- and postoperative infections in ocular lens surgery. A proposed therapeutic and economic alternative is the use of orally active fluoroquinolone ofloxacine as prophylactic agent. A HPLC method was developed for determination of both antibiotics in ocular aqueous humor and plasma in order to optimize dosage for safe surpassing minimal inhibitory concentration in the humor compartment. For plasma determinations a solid-phase extraction procedure was used with ciprofloxacine as internal standard. Detection limits for direct HPLC-analysis of ocular aqueous humor was 0.08 μg/ml for all compounds, whereas in plasma 0.31 μg/ml could be determined after solid-phase extraction.