Sensitive High-Performance Liquid Chromatographic Determination of Mebeverine in Plasma Using Fluorescence Detection

Abstract

A sensitive high-performance liquid chromatographic (HPLC) method for mebeverine (MB) determination in plasma is described. Sample preparation involves extraction of MB and Ibuprofen (internal standard) from 0.5 ml plasma. The analysis is carried out on reversed-phase chromatographic system using U-Bondapack C₁₈ column with a mobile phase consisting of water: acetonitrile:acetic acid (59:40:1) mixture. The effluent was monitored using a fluoremetric detection at excitation and emission wave lengths 270 and 362 nm, respectively. The method gave accurate, precise and reproducible results with high sensitivity. The within-day coefficients of variation ranged from 2.5 to 6.1% and between-days from 7.5 to 13.5% at four different concentrations. Injection-volumes containing as small amount of MB as 0.5 ng in plasma was detected. This method was applied to a bioavailability study with a single 10 mg/kg oral dose in two rabbits.     

A Simple High-performance Liquid Chromatographic Assay for Loracarbef in Human Plasma

Abstract

A simple, rapid and reproducible high-performance liquid chromatographic (HPLC) method for the determination of loracarbef in human plasma has been developed and evaluated. Plasma protein was precipitated with ammonium sulfate. The drug and the internal standard (Cefetamet) were eluted from a μ-bondapak C-18 column with a mobile phase consisting of acetonitrile:methanol:water:glacial acetic acid (2.5:17.5:79.2:0.8%, v/v). The column eluent was monitored at 265 nm. Quantification was achieved by the measurement of the peak-height ratio of the analyte to the internal standard and the limit of quantification for loracarbef in plasma is 0.5 ug/ml. The within-day coefficient of variation (CV) ranged from 2.28% to 3.67%, and between-day CV from 2.38% to 5.59% at three different concentrations. The absolute recoveries ranged from 91.1% to 93.88%, and the relative recoveries from 93.4% to 108% at three different concentrations. Preliminary stability tests showed that loracarbef is stable for at least 5-weeks in human plasma after freezing. The method is applied for the determination of the pharmacokinetic parameters of loracarbef after oral administration to 2 beagle dogs.     

High-Performance Liquid Chromatographic Determination of Roxithromycin in Tablets

Abstract

For the quantitative determination of Roxithromycin in tablets a rapid, and simple HPLC assay was developed. Reversed-phase chromatography was conducted using a RPC 18 (3.9 × 150 mm, 5 μm) and guard column RP C 18 (3.0 × 3.9 mm, 5 μm), mobile phase of 0.067 M phosphate buffer, pH 4.0 and methanol (65:35), and UV detection at 210 nm. Mean recovery of 100.90% and percentual coefficient of variation (CV%) of 1.51% was obtained from five commercial samples of Roxithromycin. The calibration graph was a straight line (r = 0.9995). Linearity is observed in the concentration range from 50.0 to 250.0 μg/ml. The excipients did not interfere in the determination.     

Rapid High-Performance Liquid Chromatographic Determination of Physostigmine in Plasma

Abstract

A new method is described for the quantitative determination of physostigmine in human plasma. The drug is isolated from human plasma utilizing a C₁₈ SEP PAK Cartridge, and quantified by liquid chromatography with ultraviolet detection. The average recovery is 54.3 ± 4.3% (S.D.) with a day to day coefficient of variation of 4%.     

High-Performance Liquid Chromatographic Determination of Isofraxidin in Rat Plasma

For the first time a high-performance liquid chromatographic (HPLC) method, with liquid-liquid extraction and ultraviolet (UV) absorbance detection, has been developed for quantification of isofraxidin in rat plasma. The analysis was performed on a Diamonsil C₁₈ column (200 mm × 4.6 mm i.d., 5 μm particle size) with acetonitrile–0.05% phosphoric acid, 26:74 (v/v), as isocratic mobile phase. The linear range was 0.05–8.0 μg mL⁻¹ and the lower limit of quantification was 0.05 μg mL⁻¹. The intra and inter-day relative standard deviation (RSD) for measurement of 0.25, 2.0, and 6.0 μg mL⁻¹ quality-control (QC) samples ranged from 5.7 to 6.4% and from 6.3 to 7.9%, respectively. Accuracy, as relative error (RE), was from ±5.8% to ±7.3%. The method was validated for specificity, accuracy, and precision and was successfully used in a pharmacokinetic study of isofraxidin in rat plasma after administration of Ciwujia extract.     

High-Performance Liquid Chromatographic Determination of Acetazolamide in Human Plasma

Abstract

A simple, rapid and specific HPLC method has been developed to determine acetazolamide concentrations in human plasma. The assay procedure requires only 250 μl of sample with direct injection of the organic supernatant after protein precipitation with acetonitrile. Chlorothiazide was used as an internal standard. A reversed-phase C₁₈ μBondapak column was employed for the chromatographic separation. The eluent was monitored at 265 nm using a UV variable wavelength detector. The retention times for acetazolamide (ACZ) and chlorothiazide (CTZ) were 6 and 8 min respectively. A linear relationship (r).995) was obtained over the 1-20 μg/ml concentration range. The limit of sensitivity for ACZ was 0.5 μg/ml, with greater than 85% recovery of ACZ and internal standard. The method was applied to human plasma samples obtained after administration of a 250 mg acetazolamide tablet.     

High-Performance Liquid Chromatographic Determination of Atenolol in Tablets

Abstract

A reversed phase high-performance liquid chromatographic method was developed for the determination of atenolol in four oral 100 mg atenolol preparations.

An aliquot of the sample is dissolved in a mobile phase consisting of 0.0612 M potassium hydrogen phosphate - isopropanol-tetrahydrofuran (84:10:6) v/v). The pH was adjusted to 6.7 with phosphate buffer. Nicotinamide was used as internal standard and chromatographed on a Pinkerton column ISRP (GFF-S5–80) 5 μm, 150 × 4.6 mm i.d. The applied column is convenient for the assay at least 90 samples of atenolol without degrading column performance. The detection was performed at 272 nm. The retention time for atenolol was 5.07 min.

The proposed HPLC method was found to be suitable for the rapid and precise routine analysis of atenolol in tablets.     

Rapid Determination of Apomorphine in Brain and Plasma Using High-Performance Liquid Chromatography

Abstract

A new method for quantitative determination of apomorphine in mouse brain and rat plasma is described. The drug was extracted utilizing SEP-PAK C₁₈ cartridge, and quantified by high performance liquid chromatography with electrochemical detector. The average recovery was 92 ± 2.8% with a day-to-day coefficient of variation of 10.2%. Apomorphine concentration in mouse brain and in rat plasma, as a function of dose and time, after injection with apomorphine-HCl were determined. The results indicate that the method is adequate for pharmacokinetic studies.     

Simple and Sensitive Assay of Torasemide in Human Plasma by High-Performance Liquid Chromatography Using a Monolithic Silica Column

A rapid and sensitive method to assay torasemide in plasma was developed using a simple liquid-liquid extraction technique followed by high-performance liquid chromatography. Torasemide and the internal standard furosemide were extracted from 0.5 mL of plasma using ethyl acetate in the presence of 0.1M HCl. The analysis of the extracts was performed on a monolithic silica column with ultraviolet spectrophotometric detection. The calibration curve was linear over the concentration range of 0.05-5 μg mL^?1 in plasma. Recoveries were reasonable for routine analyses (>80%); the limit of quantification was 0.05 μg mL^?1 with a signal-to-noise ratio of 5. The coefficient of variation of the assay precision was less than 6.1%, and the accuracy exceeded 98%. This method was used to measure the torasemide concentration in plasma from healthy subjects after a single 20-mg oral dose of torasemide. This method provides a very simple, sensitive, and accurate way to determine torasemide concentrations in plasma.     

High-Performance Liquid Chromatographic Determination of SAZ-VII-23, A Novel Antiarrhythmic Agent,in Dog Plasma and Urine

Abstract

A HPLC method has been developed to determine the concentrations of SAZ-VII-23 (3-benzoyl-7-isopropyl-3,7-diazabicyclo[3.3.1]nonane HClO₄), a novel antiarrhythmic agent, in dog plasma and urine. Plasma treated with acetonitrile and alkalinized urine were extracted with chloroform- propanol (9:1). An aliquot was injected on to HPLC system using a C₆ reversed-phase column and acetonitrile-methanol-37.5 mM phosphate buffer, pH 6.8 (28.5:28.5:43 v/v) containing 4.0 mM triethylamine as mobile phase. Detection wavelength was 255 nm. The linear range were 0.04–8 μg/ml, and the lower limit of quantitation was 0.04 μg/ml in plasma and urine, respectively. The method was applied to determine plasma and urine concentrations and preliminary pharmacokinetic profiles of SAZ-VII-23 in a dog.     

High-Performance Liquid Chromatographic Assay of D-Glucose in Erythrocytes

Abstract

A procedure is presented after several attempts with different modes of chromatography for measuring high concentrations of d-glucose in erythrocytes. The procedure utilizes rapid deproteinization of hemolysate by mixing with acetonitrile. The supernatant is analyzed by strong cation exchange chromatography, using an Organic Analysis Column. Separation conditions are: eluent = 0.01 N H₂SO₄, flow rate = 0.6 ml/min, detection = 195nm at 0.05 AUFS, sample size = 20 μl and temperature = ambient. The coefficients of variation for 5 mg/ml samples were (within-run) 6.7%, and (day-to-day) 7.1%. This study shows the presence of a high concentration (1900 mg/dl) of d-glucose within the erythrocytes as a result of a high external d-glucose concentration (2000 mg/dl) in plasma, and suggests that d-glucose is rapidly transported into the cell.     

Simple Liquid Chromatographic Determination of Minocycline in Brain and Plasma

A new high-performance liquid chromatography assay was developed for the determination of minocycline in plasma and brain. A solid–liquid extraction procedure was coupled with a reversed-phase HPLC system. The system requires a mobile phase consisting of acetonitrile:water:perchloric acid (26:74:0.25, v/v/v) adjusted to pH 2.5 with 5 M sodium hydroxide for elution through a RP8 column (250 × 3.0 mm, i.d.) with UV detection set at 350 nm. The method proved to be accurate, precise (RSD < 20%) and linear between 0.15–20 μg mL⁻¹ in plasma and 1–20 μg mg⁻¹ in brain. The method was successfully applied to a blood-brain barrier minocycline transport study.     

A Rapid High Pressure Liquid Chromatographic Assay of Benoxaprofen in Plasma

Abstract

A rapid high-performance liquid chromatographic assay for the determination of the anti-inflammatory drug benoxaprofen in human plasma, is described. Plasma samples of 1.0 ml, to which benoxaprofen, and warfarin as an internal standard, had been added, were extracted with ether under acidic conditions. The samples were analyzed on a MicroPak CN-10 column using 25% acetonitrile in water (pH 2.5 with phosphoric acid). Detection was made on a variable wavelength UV absorbance detector at 309 nm.

Samples containing 0.5–10 μg benoxaprofen gave a mean extraction recovery from control plasma of 90.6 ± 6.8% (n=18). Stability tests have shown that benoxaprofen in plasma is stable for at least two weeks after freezing.     

High-Performance Liquid Chromatographic Determination in Human Plasma of a Anticonvulsant Benzodiazepine: Clonazepam

Abstract

A rapid and specific high-pressure liquid chromatography method for determination of clonazepam in human plasma is described.

The analysis is linear for concentrations ranging from 5 to 100 ng. ml^-1 plasma for clonazepam.

The method is applicable to quantitation of clonazepam in human plasma of subjects receiving 0.05 at 0.20 mg.kg^-1 orally, with satisfactory accuracy and precision.     

High Performance Liquid Chromatographic Assay of Methoclopramide in Plasma Using a Silica Gel Column and An Aqueous Meobile Phase

Abstract

A high performance liquid chromatographic method for quantitating matoclopramide in plasma is presented. Proteins ara precipitated from the plasma sample with acetonitri la containing the internal standard, procainamida. The treated samples ara than analyzad using an Ultrasphere Si column, an aqueous solution at pH 7 of 65% CH³CN and 5.0 mM (NH₄)₂HPO₄ as a mobile phase, and a fluorescence detector. The retention times for drug and intarna1 standard ara 11.2 and 13.2 min, respectively. The caibration curve is Linear from 0.89 to 44.5 ng/ml. The detection limit is 0.89 ng/ml [signaL/hoisa = 31] for 0.2 ml plasma samples Pracision is measured by intraday and intarday coefficients o f variation, which are less than 10%. This method is currently being used for pharmacokinetic studies of methoclopramide.     

Simple High-Performance Liquid Chromatographic Assay, with Post-Column Derivatization, for Simultaneous Determination of Mycophenolic Acid and its Glucuronide Metabolite in Human Plasma and Urine

Two simple high-performance liquid chromatographic (HPLC) methods have been established for simultaneous determination of mycophenolic acid (MPA) and its glucuronide metabolite (MPAG) in human urine, and of their total and unbound forms in human plasma. For total MPA and MPAG analysis sample preparation entailed precipitation of protein with acetonitrile and isolation of the free analytes from the plasma by ultrafiltration. For urine samples, fivefold dilution with water was used. MPAG was determined by UV detection whereas MPA was quantified by fluorescence detection after post-column derivatization with 0.2 M sodium hydroxide solution. For plasma, response was found to be linearly dependent on concentration over the ranges 0.1–40 μg mL^-1 and 0.01–1 μg mL^-1 for total and free MPA, respectively, and 10–200 μg mL^-1 and 2.5–100 μg mL^-1 for total and free MPAG, respectively. For urine, linearity was observed from 0.1 to 50 μg mL^-1 for MPA and 10 to 500 μg mL^-1 MPAG in the urine before dilution. The methods reported were found to be accurate and reproducible for quantifying the level of MPA and MPAG and can thus be used for clinical pharmacokinetic studies and for therapeutic drug monitoring. Contributed equally to this work An erratum to this article is available at .     

High-Performance Liquid Chromatographic Determination of Cefepime and Cefazolin in Human Plasma and Dialysate

A simple high-performance liquid chromatographic (HPLC) method was developed for the simultaneous determination of cefepime and cefazolin in human plasma and dialysate. For component separation, the method utilized a C₁₈ column with an aqueous mobile phase of dibasic potassium hydrogen phosphate (pH 7.0) and methanol gradient at a flow rate of 1 mL min⁻¹. The method demonstrated linearity from 2.0 to 100.0 μg mL⁻¹ (r > 0.999) with detection limit of 1 μg mL⁻¹ for both cefepime and cefazolin. The method was utilized for evaluation of plasma and dialysate samples in a clinical study evaluating the dialyzer clearance of cefepime and cefazolin using high-flux hemodialysis with varying blood flow rates in chronic kidney failure patients undergoing hemodialysis and peritoneal dialysis treatment.     

A High-Performance Liquid Chromatographic Method for the Determination of Doxefazepam in Human Plasma Using a Solid-Phase Extraction Column

Abstract

A procedure for the rapid, quantitative isolation of doxefazepam from plasma with Supelclean LC-18 cartridges is described together with a sensitive HPLC assay for the quantitative determination of the drug. The recovery of doxefazepam was greater than 80 % over an investigated range of 0.1–2.0 μg/ml of plasma. The column extraction of doxefazepam coupled with the versatility of HPLC make this procedure well suited for detailed pharmacokinetic studies and as well as routine plasma analysis of doxefazepam.     

A Simple Isocratic High-Performance Liquid Chromatographic (HPLC) Determination of Naproxen in Human Plasma using a Microbore Column Technique

Abstract

A simple and sensitive HPLC method was developed for the determination of naproxen in human plasma. The assay employs a microbore column packed with a C18 reversed-phase material (5 μm ODS Hypersil) with an isocratic mixture of acetonitrile and 10 mM phosphate buffer, pH 2.5 (40:60, v/v) as the mobile phase. The mobile phase was pumped at a flow rate of 0.5 ml/min. For sample analysis 200 μl of acetonitrile containing internal standard (flurbiprofen) was added to 100 μl of plasma. After centrifugation 10 mM phosphate buffer, pH 7.4 (200 μl) was added to the tube, then vortexed and centrifuged. The supernatant (20 μl) was injected onto the HPLC column. The chromatographic separation was monitored by a fluorescence detector at an emission wavelength of 350 nm with an excitation wavelength of 225 nm. The direct precipitation of plasma protein using acetonitrile gave a good recovery for both naproxen and the internal standard. The detection limit was 0.1 μg/ml for naproxen. The intra- and inter-assay coefficients of variation at different concentrations evaluated were less than 10%.     

High Performance Liquid Chromatographic Determination of Amiodarone in Plasma and Tissues

Abstract

A simple and rapid high-performance liquid chromatographic (HPLC) method for the determination of amiodarone (AD) in plasma and tissues was developed. The method involved deproteinization of plasma or homogenized tissue with acetonitrile containing an internal standard (N-Cetylpyridinium chloride) followed by reversed phase chromatography using μ bondapack C₁₈ column (10μm) with a mobile phase consisting of acetonitrile - methanol - sodium dihydrogen phosphate buffer (70:10:20%, v/v), the pH adjusted to 4.0 and pumped at flow rate of 1.0 ml/min. The column effluent was monitored at 242 nm. A linear relationship was obtained between peak height ratios (drug to internal standard) versus drug levels over the concentration range of 50–750 ng/ml. The detection limit of AD in plasma and tissues by this method was 20 ng/ml.