Simultaneous determination of ketoprofen enantiomers and probenecid in plasma and urine by high-performance liquid chromatography.

A rapid, sensitive, stereospecific reversed-phase high-performance liquid chromatographic method was developed for simultaneous quantitation of ketoprofen enantiomers, probenecid and their conjugates in biological fluids. Following addition of the internal standard, indoprofen, the constituents were extracted into isooctane-isopropanol (95:5), water-washed, extracted with chloroform, then evaporated and the residue sequentially derivatized with ethyl chloroformate and L-leucinamide hydrochloride. The formed diastereomers were chromatographed on a reversed-phase column with a mobile phase of 0.06 M KH2PO4-acetonitrile-triethylamine (65:35:0.1) at a flow-rate of 1 ml/min and a detection wavelength of 275 nm. The minimum quantifiable concentration was 0.5 micrograms/ml in 100 microliters of rat plasma and urine samples. The intra- and inter-day coefficients of variation for this method are less than 10%. The assay is successfully applied to a pharmacokinetic study. The simultaneous analysis of probenecid with several other non-steroidal anti-inflammatory drugs was also successful.     

Determination of Celecoxib, Rofecoxib, Sodium Diclofenac and Niflumic Acid in Human Serum Samples by HPLC with DAD Detection

A reverse-phase high-performance liquid chromatographic method has been developed for the separation and simultaneous determination of two COX-2 inhibitors, celecoxib and rofecoxib, in addition to two well-known non-steroidal anti-inflammatory drugs (NSAIDs), sodium diclofenac and niflumic acid in human serum samples. Good chromatographic separation was achieved using a C18 bonded silica column applying a gradient with acetronitrile and water, from 15 to 60% acetonitrile. The mobile phase contained 0.1% trifluoroacetic acid as an organic modifier. Detection was made using a diode array detector (DAD) and the analytical parameters were established at the wavelength maximum in the UV spectrum of each drug. Linearity was studied up to 100.0 mg L⁻¹. Calibration functions, quantification and detection limits, intra- and inter-day reproducibility and accuracy were estimated for each drug. Solid phase extraction was needed to separate and concentrate the drugs from human serum samples. The method was successfully applied to determine the drugs in human serum samples at levels of 1.0 mg L⁻¹.     

High-performance liquid chromatographic determination of 18 alpha-glycyrrhetinic acid and 18 beta-glycyrrhetinic acid in rat plasma: application to pharmacokinetic study.

A high-performance liquid chromatographic method for the separation and determination of 18 alpha-glycyrrhetinic acid and 18 beta-glycyrrhetinic acid has been developed. Indomethacin was used as an internal standard. The drugs were separated on a reversed-phase column and detected by UV detection at a wavelength of 254 nm. Methanol-water-perchloric acid-ammonia (80:20:0.4:0.4, v/v) was used as the mobile phase at pH of 7.0-7.5. The detection limit of both compounds was 0.1 microgram/ml in rat plasma. The method was applied to pharmacokinetic studies of glycyrrhetinic acids in rats. The results suggest that the pharmacokinetics appeared to be non-linear in nature.     

Development and validation of RP-HPLC method for simultaneous determination of glipizide, rosiglitazone, pioglitazone, glibenclamide and glimepiride in pharmaceutical dosage forms and human plasma

A simple, high performance liquid chromatographic method has been developed for the simultaneous determination of glipizide, rosiglitazone, pioglitazone, glibenclamide and glimepiride in pharmaceutical dosage forms and human plasma. The elution was performed using a mobile phase mixture of 0.05% Triethylamine (pH-3.5, adjusted with ortho phosphoric acid), acetonitrile and methanol in the ratio of 55:15:30 at a flow rate of 1 ml min^?1 on a phenomenex C₁₈ column (150 × 4.6 mm, i.d., 5 ??m) at ambient temperature. The drugs were monitored at a wavelength of 248 nm and were separated within 20 min. Mixtures of formulations were prepared in suitable dilutions and plasma samples were prepared by extraction with acetonitrile. The method was successful in detecting the drugs at a concentration of less than 0.1 ??g ml^?1 for each drug and %RSD for intra- and inter-day studies was found to be less than 4.34 for all the selected concentrations. Moreover, the method was validated as per ICH guidelines and the results were found to be within the acceptable range. Hence, the proposed method can be used for the routine quality control of the drugs and can also be applied to pharmacokinetic studies.     

Direct determination of non-steroidal anti-inflammatory drugs by column-switching LC-MS

A method for determination of 16 non-steroidal anti-inflammatory drugs (NSAIDs) in human plasma samples without time-consuming sample pre-treatments was developed. The system consisted of two pumps for mobile phase delivery, a six-port switching valve, a pre-column (Oasis HLB Cartridge Column), and a reversed phase analytical column (COSMOSIL 3C18-MS-II). The analytes were trapped on the precolumn and subsequently separated on the analytical column. The present method allowed on-line sample clean-up and enrichment, leading to improved sensitivity without any tedious sample preparation. The recoveries of NSAIDs from human plasma by column-switching were greater than 72.6%. The total analysis time for a single analytical run was approximately 11 min. The detection limits of NSAIDs were 0.0025 to 0.2 microg/mL using the selected ion monitoring mode.     

Determination of flurbiprofen enantiomers in plasma using a single-isomer amino cyclodextrin derivative in nonaqueous capillary electrophoresis

A nonaqueous capillary electrophoresis (NACE) assay was developed for the separation and determination of flurbiprofen enantiomers in plasma samples using 6-monodeoxy-6-mono(3-hydroxy)propylamino-beta-cyclodextrin as chiral selector. The nonaqueous background electrolyte was made up of 40 mM ammonium acetate in methanol (MeOH), and flufenamic acid was employed as internal standard. Solid-phase extraction was used for sample cleanup prior to the NACE separation. The NACE method reproducibility was optimized by evaluating different capillary washing sequences between runs. After having tested various conditions, trifluoroacetic acid (1 M) in MeOH was finally selected. Concerning the solid-phase extraction procedure, good and reproducible analyte recoveries were obtained using MeOH for protein denaturation and a polymeric phase combining hydrophobic interactions with anion exchange properties (Oasis) MAX) was selected as extraction sorbent. The method selectivity was not only demonstrated toward a blank plasma sample but also toward other non-steroidal anti-inflammatory drugs. The method was then successfully validated with respect to response function, trueness, precision, accuracy, linearity and limit of quantification.     

A Validated and Stability-Indicating LC Assay Method for Valdecoxib

A gradient reversed-phase liquid chromatographic assay was developed for the quantitative determination of the non-steroidal anti-inflammatory drug valdecoxib. The developed method was also applicable to the determination of related substances in the bulk drug. Forced degradation studies were performed on bulk valdecoxib using acid (2.0 N hydrochloric acid), base (2.0 N sodium hydroxide), oxidation (6.0% v/v hydrogen peroxide), water hydrolysis, heat (60 °C) and photolysis. Mild degradation was observed using alkaline conditions and considerable degradation observed during oxidative stress. Chromatographic separation of process-related impurities and degradation products was achieved using a 5 micron Zorbax SB-CN LC column. The mobile phase consisted of aqueous potassium dihydrogen phosphate (pH 3.0) and acetonitrile. Stressed samples were assayed using the developed LC method and determination of the mass balance accounted for 99.5%, thus indicating the suitability of this stability-indicating method. Linearity, accuracy, precision and robustness have also been evaluated.     

Determination of atorvastatin in human plasma by magnetic solid-phase extraction combined with HPLC and application to a pharmacokinetic study

A simple and sensitive analytical procedure by solid-phase extraction method combined with high-performance liquid chromatography and using of graphene–magnetite nanomaterials as sorbent has been developed for the determination of atorvastatin in human plasma. A magnetic solid-phase extraction method as a simple, fast, and efficient extraction technique has been used for sample preparation. A solid nanocomposite material, graphene nanosheets decorated with magnetite nanoparticles, was used as a magnetic adsorbent and the adsorption process was optimized in this study. RP C18 column was used with mobile phase composed of acetonitrile–10?mM orthophosphoric acid by isocratic elution with the flow rate of 1?mL?min^?1. Fluorimetric detection was used by the excitation wavelength at 282?nm and the emission wavelength at 400?nm. It was found that the calibration curve was linear in the 30–150?ng?mL^?1. Limit of detection and limit of quantitation values were found to be 10 and 30?ng?mL^?1, respectively. The intra-day and inter-day relative standard deviation values were less than 5.27%. It has been concluded that the new developed method provides fast, simple, cost reduced, and sensitive assay for atorvastatin determination in human plasma. This method is also applied to a pharmacokinetic study.     

An HPLC Method for Determination of Atropine in Human Plasma

Abstract

A development of a high performance liquid chromatographic method for the determination of atropine in human plasma is presented. Atropine is extracted from plasma basified with 0. 1N sodium hydroxide using chloroform, subsequently subjected to base hydrolysis, followed by derivatization of the generated tropic acid with 4-bromomethyl-7-methoxycoumarin (Br-Mmc). The derivative produced has a strong blue fluorescence at excitation wavelength of 328 nm and emission cutoff filter of 389 nm. d1-Mandelic acid as internal standard (I. S.) was added after hydrolysis. The chromatographic separation was achieved on a reversed phase ODS column with a mobile phase of 33% acetonitrile in 0. 01M ammonium phosphate buffer (pH 5). The minimum quantitative limit was 125 ng/ml of plasma.     

高效液相色谱法直接测定几种非甾体类解热镇痛药物中的对映体含量

用WHELK-O1手性色谱柱，在正相条件下测定了几种非甾体类解热镇痛药物萘普生、布洛芬、酮基布洛芬和苯氧布洛芬等中对映体的含量。结果表明：这种手性固定相色谱柱能够以正己烷和异丙醇为流动相，简便、快速、准确地测定非甾类药物中对映体的含量。     

Validation and pharmacokinetic application of a method for determination of doxazosin in human plasma by high-performance liquid chromatography

A simple high-performance liquid chromatographic method for the determination of doxazosin in human plasma was developed and validated. Prazosin was used as internal standard. After extraction twice with ethyl acetate, chromatographic separation of doxazosin in human plasma was carried out using a reversed-phase Apollo C18 column (250 x 4.6 mm, 5 microm) with mobile phase of methanol-acetonitrile-0.04 m disodium hydrogen orthophosphate (22:22:56, v/v/v) adjusted to pH 4.9 with 0.9 m phosphoric acid and quantified by fluorescence detection operated with an excitation wavelength of 246 nm and an emission wavelength of 389 nm. The lower limit of quantification (LLOQ) of this assay was 1 ng/mL using 500 microL human plasma. Linearity was established over the range 1-25 ng/mL (r2 > 0.9994). The intra- and inter-day accuracy ranged from 90.5 to 104.4% and the coefficient of variation were not more than 8.6% for both intra- and inter-day precision, over the range of the calibration curve. The absolute recoveries of doxazosin and prazosin from human plasma were more than 91%. Doxazosin demonstrated acceptable short-term, long-term and freeze-thaw stability in human plasma. The assay has been successfully applied to plasma sample ana-lysis for pharmacokinetic study.     

Simultaneous estimation of six anti-diabetic drugs--glibenclamide, gliclazide, glipizide, pioglitazone, repaglinide and rosiglitazone: development of a novel HPLC method for use in the analysis of pharmaceutical formulations and its application to human plasma assay

This paper describes a convenient method for the separation and simultaneous determination of six anti-diabetic drugs viz., glibenclamide (GLB), gliclazide (GLC), glipizide (GLZ), pioglitazone (PGL), repaglinide (RPG) and rosiglitazone (RGL) in pharmaceutical formulations. Also, the assay has been shown applied to support quantification of the six anti-diabetic drugs in human plasma. The analytes were either injected directly onto the column after suitable dilution (pharmaceutical formulation analysis) or a simple extraction procedure, using acetonitrile, from human plasma spiked with anti-diabetic drugs and internal standard (IS). Ternary gradient elution at a flow rate of 1 mL/min was employed on an Intertisl ODS 3V column (4.6 x 250 mm, 5 microm) at ambient temperature. The mobile phase consisted of 0.01 m formic acid (pH 3.0), acetonitrile, Milli Q water and methanol. Celecoxib was used as an IS. The six anti-diabetic drugs were monitored at a wavelength of 260 nm. The nominal retention times of RGL, PGL, GLZ, GLC, GLB, IS and RGL were 11.4, 13.3, 14.8, 17.6, 20.78, 22.1 and 25.4 min, respectively. The assay developed for formulation analysis was found to be accurate and precise. The calibration curves ranged from 0.1 to 100 microg/mL for all analytes with the exception of GLB, where the range was 0.3-100 microg/mL. The plasma assay was validated for parameters such as specificity, accuracy and extraction recovery. The proposed method is simple, selective and can be extended for routine analysis of anti-diabetics in pharmaceutical preparations and in biological matrices.     

Determination of remoxipride in human plasma and urine by reversed-phase ion-pair high-performance liquid chromatography.

A sensitive method is described for the measurement of remoxipride in human plasma and urine. Remoxipride and its internal standard are extracted from plasma or urine at pH 12 with a mixture of hexane and methyl tert.-butyl ether. After washing the organic phase with base, the compounds are extracted into acid and analyzed on a C18 column with ultraviolet detection at 214 nm. The mobile phase is composed of acetonitrile and aqueous buffer (sodium perchlorate and phosphoric acid, pH 1.7). The limits of reliable quantitation for remoxipride are 12.5 and 50 ng/ml for plasma and urine, respectively. The run times are 6 min for plasma and 3 min for urine. The method has been successfully used to assay remoxipride clinical study samples. This mobile phase has also been successfully applied to the analysis of other basic drugs such as cimetidine, codeine, diltiazem and quinidine with minor modifications.     

Selective determination of famotidine in human plasma by high performance liquid chromatography in alkaline media with solid phase extraction

A new method is described for the determination of famotidine by solid phase extraction from alkalinized human plasma followed by reversed phase (RP) HPLC in acetonitrile/alkaline buffer with molsidomine as an internal standard. Different acetonitrile/aqueous buffer mobile phases as well as various RP columns were used. Alkaline medium allowed the limit of quantitation to be lowered to 5 ng/mL of plasma as the famotidine gives more intense absorption at about 286 nm (at pH values higher than 7). Moreover, work in alkaline media and at this wavelength is highly selective as peaks corresponding to impurities present in most samples are well separated. A method using a mildly alkaline mobile phase (acetonitrile/10 mM phosphate with 10 mM 1‐heptanesulphonic acid, pH 7.5) was successfully used for determination of famotidine in human plasma in a pharmacokinetic study.     

Development and evaluation of a simple and easy high-performance liquid chromatography–ultraviolet system simultaneously suitable for determination of 24 anti-epileptic drugs in plasma

We aim to establish a simple and easy high-performance liquid chromatography system coupled with an ultraviolet detector suitable for simultaneous determination of 24 antiepileptic drugs in human plasma. Optimized chromatographic separation was performed on a ZORBAX Eclipse Plus-C18 (4.6 × 150 mm², 3.5 μm) column with acetonitrile and 5 mM potassium dihydrogen phosphate water solution as mobile phase. Note that, 24 antiepileptic drugs were divided into three groups and eluted with different gradient procedures, respectively. The column temperature was maintained at 35°C and the detection wavelength was set at 210 nm. Plasma was processed with ethyl acetate or acetonitrile. The calibration curves of 24 antiepileptic drugs demonstrated good linearity within the test range (r > 0.996). The intra- and inter-batch precision and accuracy were all less than 15%, while extraction recoveries were in the range of 74.57–90.89% with the relative standard deviation values less than 15%. The validated methods have been successfully applied to determination of some antiepileptic drugs in rat or patient plasma. Those results indicated that the developed methods were simple and easy, and could be suitable for the determination of 24 antiepileptic drugs in plasma just by changing the gradient elution procedures of mobile phase.     

Simple and extractionless high-performance liquid chromatographic determination of rosiglitazone in human plasma and application to pharmacokinetics in humans

A simple and extractionless HPLC method using fluorescence detection was developed for the determination of rosiglitazone in human plasma. After deproteinization using perchloric acid the plasma samples were directly injected onto the HPLC system. The mobile phase was composed of acetonitrile (52%) and 20 mm ammonium acetate (48%, pH 7.5), and analysis was run at a flow rate of 0.2 mL/min with the detector operating at 247 nm for excitation wavelength and at 367 nm for emission wavelength, respectively. The method has a mean recovery of 97%, while the intra-day and inter-day precisions were all less than 7%. This method is simple, specific, sensitive and requires only a small plasma volume with short analytical time, and is suitable for the determination of plasma rosiglitazone in routine measurements for pharmacokinetic studies.     

Automated Solid Phase Extraction and Hplc Analysis of Ibuprofen in Plasma

Abstract

Ibuprofen is a non-steroidal anti-inflammatory drug, widely used in arthritis and other disorders. We describe a high pressure liquid chromatographic (HPLC) method for the analysis of ibuprofen in plasma, using an automated solid phase extraction technique (the Varian AASP^R). In this method ibuprofen was extracted from 0.5 ml of plasma by application to a C₂ extraction cartridge followed by “on line” elution with the HPLC mobile phase (55% acetonitrile / 45% 0.02 H phosphate buffer; pH 3.0), at a flow rate of 1.5 ml/min. The analytical column was a Nucleosil C₁₈ column and the fluorescence detector was set at 253 nm (excitation wavelength) and 300 nm (emission wavelength). Chromatography was complete in less than 10 mins and the limit of detection was 1.3 /μg/ml. The method is linear through the range of 1.0 to 100.0 /μg/ml with a mean correlation coefficient of 0.9964. Absolute recovery of ibuprofen from the spiked plasma samples ranged from 77.8% to 86.5%. The method was shown to be precise within 11% C.V. and accurate to within 8% over the concentration range studied.     

High-performance liquid chromatographic evaluation of Med 15 and its metabolites Med 5 and tolmetin in rat plasma.

A simple and reliable high-performance liquid chromatographic method is described for the quantitative analysis of the new non-steroidal anti-inflammatory agent Med 15 and its metabolites Med 5 and tolmetin in rat plasma. After selective extraction the three analytes and an internal standard (p-phenyl-phenol) were separated on a reversed-phase Ultrasphere 5 micron column using potassium dihydrogenphosphate (0.05 M)-acetonitrile (52:48) (pH 4.7) as the mobile phase. The analytes were detected at 313 nm; the sensitivity of the method proved to be 0.05 microgram/ml for all three compounds. The method has been applied to investigate Med 15 pharmacokinetics in rats.     

配体交换色谱法测定草甘膦

建立了配体交换色谱测定草甘膦的方法.在自制的配体交换色谱柱上,以醋酸铜-醋酸水溶液为流动相,紫外(254 nm)检测,实现了草甘膦与双甘膦中间体的基线分离.考察了流动相中醋酸铜和醋酸的浓度,以及流动相流速对目标物分离的影响,确定了以0.2 mmol/L醋酸铜-醋酸水溶液为流动相,流速0.8 mL/min的优化色谱条件,并进行了系统适应性实验和方法学评价.结果表明:草甘膦与双甘膦之间的分离度Rs＞3,拖尾因子在0.8～1.0之间;在0.05～1.00 g/L范围内,草甘膦和双甘膦的线性相关系数分别为0 9998和0 9999; 回收率分别为102.2%和99.0%; 峰面积的RSD分别为0.14%和0.59%; 检出限分别为0 05和0.50 mg/L(S/N=3).     

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.