Determination of meloxicam in human plasma by ultra-high-performance liquid chromatography–tandem mass spectrometry and its application in a pharmacokinetic study

A rapid, selective and sensitive ultra-high-performance liquid chromatography–tandem mass spectrometry (UHPLC–MS/MS) method was developed to detect meloxicam in human plasma. A triple quadrupole tandem mass spectrometer equipped with an electrospray ionization source was used in positive ion mode. Protein precipitation with acetonitrile was used for sample preparation. Meloxicam and ¹³C₆-meloxicam internal standard were analyzed on an Acquity CSH C₁₈ column with a mobile phase of acetonitrile and water in 0.1% formic acid using a gradient program for separation. The retention time of meloxicam was 1.1 min and the total run time was only 2.0 min. Detection was performed in multiple reaction monitoring mode using an electrospray ionization source with optimized mass spectrometry parameters. The calibration curves were linear in the range 10.0–3.00 × 10³ ng/ml (r ≥ 0.99). The within-run and between-run RSDs were ≤14.8%. The within-run and between-run REs ranged from −4.6 to 10.7%. There was no significant matrix effect, and the recovery rate was high. This method was fully validated, including reinjection reproducibility in human plasma. The method was applied to the pharmacokinetic study. All of the incurred sample reanalysis methods met the criteria.     

An HPLC method for the determination and pharmacokinetic study of lehmannine in rat plasma

A high-performance liquid chromatographic (HPLC) method for determining lehmannine (LMN) in rat plasma was developed for application in the pharmacokinetics study. The plasma was deproteinized with acetonitrile that contained an internal standard and was separated from the aqueous layer by adding sodium chloride. The HPLC assay was carried out using a VP-ODS column at 40 degrees C. The mobile phase was acetonitrile-0.02 mol/L ammonium acetate buffer-triethylamine (35:65:0.04, v/v/v). The flow rate was 1.0 mL/min. The detection wavelength was set at 220 nm. The method was used to determine the concentration-time profiles of LMN in the plasma following oral administration or bolus injection of LMN aqueous solution. The pharmacokinetic parameters of LMN were calculated for the first time by Drug and Statistics 1.0 program.     

A simple and sensitive HPLC method for quantitation of low phenoprolamine hydrochloride concentrations in human plasma and its pharmacokinetic application

Phenoprolamine hydrochloride is a novel compound that works against a variety of types of hypertension. The purpose of this study was to develop a simple and sensitive high-performance liquid chromatographic method for quantitation of low phenoprolamine hydrochloride concentrations in human plasma and to apply it to pharmacokinetic study. The procedure involved extraction of the drug and clonidine (internal standard) from the plasma using diethyl ether. Chromatographic separations were carried out on a 4.6 x 200 mm Hypersil silica column with UV detection at 230 nm. The isocratic mobile phase, 1% ammonium acetate (pH 5.4) and methanol (0.3:99.7, v/v), was run at 1 mL/min. Extraction recovery was 84% for phenoprolamine hydrochloride at a concentration level of 200 ng/mL, and 76% for clonidine at 200 ng/mL. The method was linear in the concentration range 5-4000 ng/mL with a lower limit of quantitation of 5 ng/mL for phenoprolamine hydrochloride. Inter- and intra-day coefficients of variation were less than 10%. The validated method was successfully applied to a pharmacokinetic study in human after an oral administration of the drug, and the pharmacokinetic parameters are presented.     

Quantitative determination of meloxicam in dog plasma by high performance liquid chromatography–tandem mass spectrometry and its application in a pharmacokinetic study

A rapid, selective and sensitive high‐performance liquid chromatography–tandem mass spectrometry (HPLC‐MS/MS) method was developed to determine meloxicam in beagle dog plasma. Sample pretreatment involved a one‐step protein precipitation with methanol of 0.1 mL plasma. Analysis was performed on a Venusil ASB‐C₁₈ column with mobile phase consisting of methanol–water (containing 0.1% formic acid) (75:25, v/v). The detection was performed on a triple quadrupole tandem mass spectrometer by multiple reaction monitoring mode via electrospray ionization source. Each plasma sample was chromatographed within 4.1 min. The linear calibration curves for meloxicam was obtained in the concentration range of 10.3–4.12 × 10³ ng/mL (r ≥ 0.99). The intra‐ and inter‐day precisions (relative standard deviation) were ≤ 15%, and accuracy (relative error) was within ±7.3%. The method herein described was fully validated and successfully applied to the pharmacokinetic study of meloxicam tablets in beagle dog.     

Rapid and sensitive determination of levofloxacin in microsamples of human plasma by high‐performance liquid chromatography and its application in a pharmacokinetic study

A rapid, sensitive and simple high‐performance liquid chromatographic assay with ultraviolet detection was developed for the quantification of levofloxacin in microsamples (100 μL) of human plasma. The extraction procedure included a protein precipitation technique and a short chromatographic running time (4.5 min). Analyses were carried out on a Symmetry C₁₈ column using a mixture of acetonitrile and 0.01 m potassium dihydrogen aqueous solution (pH 3.4; 14:86 v/v) as mobile phase. The method provided specificity and was linear (r ≥ 0.9992) over the concentration range 0.1–12 µg/mL. The average absolute recovery was 93.59%. The intra‐ and inter‐day coefficients of variation were <6%. Additionally, levofloxacin was stable in all evaluations. The usefulness of this method was demonstrated in a pharmacokinetic study of levofloxacin in healthy adult volunteers. The present method offers two main advantages: (a) the use of microsamples reduces the total volume of blood to be collected from patients; and (b) it provides a good cost–effectiveness ratio. It is concluded that the method is rapid, simple, sensitive, economical and suitable for the determination of levofloxacin in human plasma using a small volume of sample. Copyright © 2014 John Wiley & Sons, Ltd.     

Development and validation of an RP‐HPLC method for the quantitation of odanacatib in rat and human plasma and its application to a pharmacokinetic study

A simple, specific, sensitive and reproducible high‐performance liquid chromatography (HPLC) assay method has been developed and validated for the estimation of odanacatib in rat and human plasma. The bioanalytical procedure involves extraction of odanacatib and itraconazole (internal standard, IS) from a 200 μL plasma aliquot with simple liquid–liquid extraction process. Chromatographic separation was achieved on a Symmetry Shield RP₁₈ using an isocratic mobile phase at a flow rate of 0.7 mL/min. The UV detection wave length was 268 nm. Odanacatib and IS eluted at 5.5 and 8.6 min, respectively with a total run time of 10 min. Method validation was performed as per US Food and Drug Administration guidelines and the results met the acceptance criteria. The calibration curve was linear over a concentration range of 50.9–2037 ng/mL (r² = 0.994). The intra‐ and inter‐day precisions were in the range of 2.06–5.11 and 5.84–13.1%, respectively, in rat plasma and 2.38–7.90 and 6.39–10.2%, respectively, in human plasma. The validated HPLC method was successfully applied to a pharmacokinetic study in rats. Copyright © 2015 John Wiley & Sons, Ltd.     

A high-performance liquid chromatographic method for determination of scopolin in rat plasma: application to pharmacokinetic studies

An analytical method based on high-performance liquid chromatographic (HPLC) with ultraviolet (UV) detection was developed for determination of scopolin in rat plasma using aesculin as internal standard (IS). After protein precipitation of plasma sample with methanol, the supernatant was directly injected and analyzed. Chromatographic separation was achieved on a C18 column using methanol and distilled water (22:78, v/v) containing 0.2% (v/v) glacial acetic acid as mobile phase with a column temperature of 30 degrees C. The UV detector was set at 338 nm. The calibration curve was linear over the range of 0.105-13.125 microg/mL with a correlation coefficient of 0.9998. The retention times of aesculin and scopolin were 10.4 and 12.8 min, respectively. The recoveries for plasma samples of 0.105, 4.725 and 13.125 microg/mL were 91.08, 95.30 and 96.10%, respectively. The RSD of intra- and inter-day assay variations was less than 7.35%. The lower limit of detection was 0.03 microg/mL .This HPLC assay is a simple, sensitive and accurate and was successfully applied to the pharmacokinetic study of scopolin in rats.     

HPLC method for determination of moxifloxacin in plasma and its application in pharmacokinetic analysis

Moxifloxacin is a representative of the fourth generation of fluoroquinolones. It possesses bacteriostatic activity against Gram-positive and Gram-negative bacteria. A simple and fast high performance liquid chromatography-ultraviolet detection (HPLC-UV) method was developed for moxifloxacin analysis. The separation was performed on C18 column. The mobile phase was a mixture of acetonitrile and 0.4% triethylamine solution. The regression curve was linear over the range 0.2–10.0?µg/mL. The validation parameters obey the European Medicine Agency limits. The in vivo study confirmed the practical application of the method.     

Development and validation of a reliable high-performance liquid chromatographic method for determination of nodakenin in rat plasma and its application to pharmacokinetic study

A simple and reliable high-performance liquid chromatographic (HPLC) method has been developed for the determination of nodakenin in rat plasma. The concentration of nodakenin was determined in plasma samples after deproteinization with methanol using hesperidin as internal standard. HPLC analysis was performed on a Diamonsil C(18) analytical column using acetonitrile-water (25:75, v/v) as the mobile phase and a UV detection at 330 nm. This method was validated in terms of recovery, linearity, accuracy and precision (intra- and inter-day variation). The extraction recoveries were 91.3 ± 10, 87.8 ± 4.8 and 92.6 ± 5.1 at concentrations of 0.500, 5.00 and 40.0 μg/mL, respectively. The standard curve for nodakenin was linear (r(2) ≥ 0.99) over the concentration range 0.250-50.0 μg/mL with a lower limit of quantification of 0.250 μg/mL. The intra- and inter-day precision (relative standard deviation, RSD) values were not higher than 12% and the accuracy (relative error, RE) was within ± 5.8% at three quality control levels. The validated method was successfully applied for the evaluation of the pharmacokinetics of nodakenin in rats after oral administration of Rhizoma et Radix Notopterygii decoction and nodakenin solution.     

Validation of a HPLC‐ESI MS/MS method for the determination of clonidine in human plasma and its application in a bioequivalence study in Chinese healthy volunteers

A rapid and sensitive liquid chromatography–tandem mass spectrometry (HPLC‐MS/MS) method to determine clonidine in human plasma was developed and fully validated. Sample preparation was involved an one‐step extraction with diethyl ether. Donepezil was employed as the internal standard (IS). Chromatographic separation was performed on a Hypersil BDS C₁₈ column (i.d. 2.1 × 50 mm, particle size 3μm) with a mobile phase of methanol–water (containing 0.1% formic acid; 60:40, v/v) at a flow rate of 200 μL/min. The peaks were detected by mass spectrometry using the electrospray ion source in selected reaction monitoring mode. The extraction recovery was 72.53–85.25%. The method was found to be linear in a concentration range of 0.02–6.00 ng/mL and the lower limit of quantification was 0.02 ng/mL. The within‐ and between‐batch precisions at three concentrations were 4.33–16.47 and 7.24–17.24% with accuracies of ?2.47–10.91 and 1.86–10.19%, respectively. This validated method was successfully used for a bioequivalence study of two clonidine transdermal patches on healthy volunteers. The results suggested that the test formulation of clonidine patch met the regulatory criterion for bioequivalence to the reference formulation, but a larger sample size should be needed for the estimation of bioequivalence. Copyright © 2015 John Wiley & Sons, Ltd.     

High‐performance liquid chromatography method for determination of carnosic acid in rat plasma and its application to pharmacokinetic study

A sensitive and reproducible high‐performance liquid chromatography method with ultraviolet detection (UV) was developed for the determination of carnosic acid (CA) in rat plasma. After simple acidification and liquid–liquid extraction of plasma samples using gemfibrozil as an internal standard, the supernatant was evaporated to dryness under a gentle stream of nitrogen. The residue was reconstituted in 200 µL before being injected into the chromatographic system. The analysis was performed on a C₁₈ column protected by an ODS guard column using acetonitrile–0.1% phosphoric acid (55:45, v/v) as mobile phase, and the wavelength of the UV detector was set at 210 nm. The calibration curve was linear over the range of 0.265–265.0 µg/mL with a correlation coefficient of 0.9997. The recovery for plasma samples of 0.530, 13.25, 132.5 and 265.0 µg/mL was 72.2, 87.9, 90.4 and 94.7%, respectively. The intra‐day and inter‐day relative standard deviations for the measurements of quality control samples were less than 3.1%. The stability of the plasma samples was also validated. This method was successfully used to study the pharmacokinetics and bioavailability of CA in rats. Copyright © 2009 John Wiley & Sons, Ltd.     

An ultraperformance liquid chromatography–tandem mass spectrometry method for determination of anastrozole in human plasma and its application to a pharmacokinetic study

A fast, selective and sensitive ultraperformance liquid chromatography–tandem mass spectrometry method was developed for determination and pharmacokinetic study of anastrozole in human plasma. Plasma sample pretreatment involved a one‐step extraction with diethyl ether of 500 µL plasma. The chromatographic separation was carried out on an Acquity UPLC^TM BEH C₁₈ column with a mobile phase consisting of methanol–10 mmol/L ammonium acetate (75:25, v/v) at a flow rate of 0.30 mL/min. The detection was performed on a triple quadrupole tandem mass spectrometer by multiple reaction monitoring via electrospray ionization source with positive mode. A high throughput was achieved with a run time of 1.5 min per sample. The standard curve for anastrozole was linear (r² ≥ 0.99) over the concentration range of 0.0550–27.5 ng/mL with a lower limit of quantification of 0.0550 ng/mL. The intra‐ and inter‐day precision (relative standard deviation) values were not higher than 14% and the accuracy (relative error) was within ±3.2% at three quality control levels. This simple, fast and highly sensitive method was fully validated and successfully applied to a clinical pharmacokinetic study of anastrozole in healthy volunteers after oral administration. Copyright © 2010 John Wiley & Sons, Ltd.     

A rapid and simple high-performance liquid chromatography method for the determination of human plasma levofloxacin concentration and its application to bioequivalence studies

A high-performance liquid chromatography method with fluorescence detection (HPLC-FLD) for the determination of levofloxacin in human plasma is described. Neutralized with phosphate buffer (pH 7.0), the sample (0.1 mL) was extracted with dichlormethane (1 mL). After voltex-mixing and centrifuged at 3000g for 6 min at 4 degrees C, the upper aqueous layer was aspirated using a micro vacuum pump and the organic layer was directly transferred to a clean test tube without pipetting. The organic solvent was evaporated and the residues were reconstituted with the mobile phase. Levofloxacin and terazosin (internal standard, IS) were chromatographically separated on a C(18) column with a mobile phase containing phosphate buffer (pH 3.0, 10 mm), acetonitrile and triethylamine (76:24:0.076, v/v/v) at a flow rate of 1 mL/min. The analytes were detected using fluorescence detection at an excitation and emission wavelength of 295 and 440 nm, respectively. The linear range of the calibration curves was 0.0521-5.213 microg/mL for levofloxacin with a lower limit of quantitation (0.0521 microg/mL). The retention times of levofloxacin and terazosin were 2.5 and 3.1 min, respectively. Within- and between-run precision was less than 12 and 11%, respectively. Accuracy ranged from -6.3 to 4.5%. The recovery ranged from 86 to 89% at the concentrations of 0.0521, 0.5213 and 5.213 microg/mL. The present HPLC-FLD method is sensitive, efficient and reliable. The method described herein has been successfully used for the pharmacokinetic and bioequivalence studies of a levofloxacin formulation product after oral administration to healthy Chinese volunteers.     

Novel LC‐ ESI‐MS/MS method for desvenlafaxine estimation human plasma: application to pharmacokinetic study

A simple, sensitive and specific liquid chromatography tandem mass spectrometry (LC‐ESI‐MS/MS) method was developed for the quantification of desvenlafaxine in human plasma using desvenlafaxine d6 as an internal standard (IS). Chromatographic separation was performed using a Thermo‐BDS hypersil C₈ column (50 × 4.6 mm, 3 µm) with an isocratic mobile phase composed of 5 mM ammonium acetate buffer: methanol (20:80, v/v), at a flow rate of 0.80 mL/min. Desvenlafaxine and desvenlafaxine d6 were detected with proton adducts at m/z 264.2/58.1 and 270.2/ 64.1 in multiple reaction monitoring positive mode, respectively. Liquid–liquid extraction was used to extract the drug and the IS. The method was linear over the concentration range 1.001–400.352 ng/mL with a correlation coefficient of ≥0.9994. This method demonstrated intra and inter‐day precision within 0.7–5.5 and 1.9–6.8%, and accuracy within 95.3–107.4 and 93.4–99.5%. Desvenlafaxine was found to be stable throughout the freeze–thaw cycles, bench‐top and long‐term matrix stability studies. The developed and validated method can be successfully applied for the bioequivalence/pharmacokinetic studies of desvenlafaxine in pharmaceutical dosage forms. Copyright © 2015 John Wiley & Sons, Ltd.     

Rapid and sensitive LC‐MS/MS method for determination of megestrol acetate in human plasma: application to a human pharmacokinetic study

A rapid, simple and fully validated LC‐MS/MS method was developed and validated for the determination of megestrol acetate in human plasma using tolbutamide as an internal standard (IS) after one‐step liquid–liquid extraction with methyl‐tert‐butyl‐ether. Detection was performed using electrospray ionization in positive ion multiple reaction monitoring mode by monitoring the transitions m/z 385.5 → 267.1 for megestrol acetate and m/z 271.4 → 155.1 for IS. Chromatographic separation was performed on a YMC Hydrosphere C₁₈ column with an isocratic mobile phase, which consisted of 10 mm ammonium formate buffer (adjusted to pH 5.0 with formic acid)–methanol (60:40, v/v) at a flow rate of 0.4 mL/min. The achieved lower limit of quantitation (LLOQ) was 1 ng/mL (signal‐to‐noise ratio > 10) and the standard calibration curve for megestrol acetate was linear (r > 0.99) over the studied concentration range (1–2000 ng/mL). The proposed method was fully validated by determining its specificity, linearity, LLOQ, intra‐ and inter‐day precision and accuracy, recovery, matrix effect and stability. The validated LC‐MS/MS method was successfully applied for the evaluation of pharmacokinetic parameters of megestrol acetate after oral administration of a single dose 800 mg of megestrol acetate (Megace?) to five healthy Korean male volunteers under fed conditions. Copyright © 2012 John Wiley & Sons, Ltd.     

Development and validation of UPLC‐MS/MS method for determination of domperidone in human plasma and its pharmacokinetic application

A sensitive, rapid and selective ultra‐performance liquid chromatography–tandem mass spectrometric (UPLC‐MS/MS) method was developed for the determination and pharmacokinetic study of domperidone in human plasma. Diphenhydramine was used as the internal standard. Plasma sample pretreatment involved a one‐step liquid–liquid extraction with a mixture of diethyl ether–dichloromethane (3:2, v/v). The analysis was carried out on an Acquity UPLC^TM BEH C₁₈ column. The mobile phase consisted of methanol–water containing 10 mmol/L ammonium acetate and 0.5% (v/v) formic acid (60:40, v/v). The detection was performed on a triple quadrupole tandem mass spectrometer in multiple reaction monitoring mode via electrospray ionizationsource with positive mode. Each plasma sample was chromatographed within 2.1 min. The standard curves for domperidone were linear (r² ≥ 0.99) over the concentration range of 0.030–31.5 ng/mL with a lower limit of quantification of 0.030 ng/mL. The intra‐ and inter‐day precision (relative standard deviation) values were not higher than 13% and accuracy (relative error) was from ?7.6 to 1.2% at three quality control levels. The method herein described was superior to previous methods and was successfully applied to the pharmacokinetic study of domperidone in healthy Chinese volunteers after oral administration. Copyright © 2012 John Wiley & Sons, Ltd.     

High-performance liquid chromatographic determination of doxazosin in human plasma for bioequivalence study of controlled release doxazosin tablets

A highly sensitive high-performance liquid chromatographic quantification method with fluorescence detection was developed and validated for the determination of doxazosin in human plasma. The developed method employed one-step extraction of doxazosin from plasma matrix with ethyl acetate using propranolol as an internal standard. Chromatographic separation was obtained within 8.0 min using a reverse-phase Capcell-Pak C(18) column (150 x 4.6 mm i.d., 5 microm) and the mobile phase consisted of methanol-water containing 10 mM perchloric acid and 1.8 mM sodium heptane sulfonic acid (50:50, v/v) and was set at a flow rate of 1.5 mL/min. The calibration curve constructed was linear in the range of 0.3-50.0 ng/mL. The proposed method achieved a lower limit of quantification of 0.3 ng/mL, better than the reported HPLC methods. Average recoveries of doxazosin and the internal standard from human plasma matrix were 87.0 and 85.9%, respectively. The present method was validated by evaluating the precision and accuracy for inter- and intraday variation in the concentration range 0.3-50 ng/mL. The precision values expressed as relative standard deviations in the inter- and intraday validation were 1.17-6.29 and 0.84-5.94%, respectively. This method was successfully applied to the bioequivalence study of two doxazosin controlled release tablets in healthy, male human subjects.     

Determination of ipriflavone in human plasma by LC-MS and its application in a pharmacokinetic study

A sensitive liquid chromatography-mass spectrometric method was developed for the quantification of ipriflavone in human plasma. The method utilized liquid-liquid extraction of plasma with ethyl acetate. A gradient elution was performed on a Hedera ODS-2 column (150×2.1 mm i.d., 5 μm), using a mobile phase consisting of 0.1% formic acid solution and methanol at a flow rate of 0.5 mL/min. The single quadrupole mass spectrometer was operated in selected-ion monitoring mode via positive electrospray ionization interface detecting m/z 239.1 and 285.1 for ipriflavone and diazepam (the internal standard), respectively. To improve the selectivity and sensitivity, the fragment ion m/z 239.1, which was produced by in-source collision-induced dissociation, was chosen as the quantitative ion for ipriflavone. The method was fully validated and applied to a pharmacokinetic study of ipriflavone. After oral administration of a single 200 mg ipriflavone tablet, the C(max,) AUC(0-72 h) , t(1/2) and T(max) were 6.3±6.3 ng/mL, 80.0±69.1 μg h/L, 23.0±8.6 h and 3.4±2.1 h, respectively.     

Development and validation of a highly sensitive method for the determination of abiraterone in rat and human plasma by LC-MS/MS-ESI: application to a pharmacokinetic study

A highly sensitive, rapid assay method has been developed and validated for the estimation of abiraterone (ART) in rat and human plasma with liquid chromatography coupled to tandem mass spectrometry and electrospray ionization in the positive-ion mode. The assay procedure involves extraction of ART and phenacetin (internal standard, IS) from rat and human plasma with a simple protein precipitation extraction process. Chromatographic separation was achieved using an isocratic mobile (10 mm ammonium acetate:acetonitrile, 10:90, v/v) at a flow-rate of 0.70 mL/min on an Atlantis dC(18) column maintained at 40 °C with a total run time of 3.5 min. The MS/MS ion transitions monitored were 350.3 → 156.0 for ART and 180.2 → 110.1 for IS. Method validation was performed as per FDA guidelines and the results met the acceptance criteria. The lower limit of quantitation achieved was 0.20 ng/mL and the linearity range extended from 0.20 to 201 ng/mL. The intra- and inter-day precisions were in the ranges 2.39-10.4 and 4.84-9.53% in rat plasma and 3.82-10.8 and 6.97-8.94% in human plasma.     

Highly sensitive method for the determination of omeprazole in human plasma by liquid chromatography-electrospray ionization tandem mass spectrometry: application to a clinical pharmacokinetic study

A highly sensitive, rapid assay method has been developed and validated for the estimation of omeprazole (OPZ) in human plasma with liquid chromatography coupled to tandem mass spectrometry with electrospray ionization in the positive-ion mode. The assay procedure involves alkalinization of plasma followed by simple liquid-liquid extraction of OPZ and lansoprazole (internal standard, IS) from human plasma with acetonitrile. Chromatographic separation was achieved with 0.01 M ammonium acetate:acetonitrile (40:60, v/v) at a flow rate of 0.25 mL/min on an Inertsil ODS 3 column with a total run time 2.5 min. The MS/MS ion transitions monitored were 346.1 --> 198.1 for OPZ and 370.1 --> 252.1 for IS. Method validation and clinical sample analysis were performed as per FDA guidelines and the results met the acceptance criteria. The lower limit of quantitation achieved was 0.05 ng/mL and the linearity was observed from 0.05 to 10.0 ng/mL. The intra-day and inter-day precisions were in the ranges 2.09-8.56 and 5.29-8.19%, respectively. This novel method has been applied to a pharmacokinetic study of OPZ in humans.