A sensitive and selective liquid chromatographic/electrospray ionization tandem mass spectrometric assay for the simultaneous quantification of alpha-,beta-arteether and its metabolite dihydroartemisinin in plasma,useful for pharmacokinetic studies

A sensitive, selective, specific and rapid liquid chromatographic/electrospray ionization tandem mass spectrometric assay method was developed and validated for the simultaneous quantitation of alpha-,beta-arteether (alpha-,beta-AE) and its metabolite alpha-dihydroartemisinin (DHA) in monkey plasma using the propyl ether analogue of beta-arteether (PE) as an internal standard. The method involves a simple two-step liquid-liquid extraction with hexane. The analytes were chromatographed on a C(18) reversed-phase chromatographic column by isocratic elution with methanol-ammonium acetate buffer (pH 4) (92 : 8, v/v) and analysed by mass spectrometry in the multiple reaction monitoring mode. The chromatographic run time was 7 min and the weighted (1/x(2)) calibration curves were linear over the range 0.78-200 ng ml(-1). The method was validated in terms of accuracy, precision, absolute recovery, freeze-thaw stability, bench-top stability and re-injection reproducibility. The limit of detection and lower limit of quantification in monkey plasma were 0.39 and 0.78 ng ml(-1) respectively for all the analytes. The intra- and inter-batch precision and accuracy were found to be well within acceptable limits (<15%). All three analytes were stable even after three freeze-thaw cycles (deviation < 15%). The average absolute recoveries of alpha-,beta-AE, DHA and PE, used as an internal standard, from spiked plasma samples were 85.85 +/- 6.56, 70.10 +/- 7.06, 54.37 +/- 3.39 and 93.90 +/- 6.9%, respectively. The assay method described here could be applied to study the pharmacokinetics of alpha-,beta-AE and DHA in rhesus monkeys.     

Determination of sodium cromoglycate in human plasma by liquid chromatography with tandem mass

A sensitive and selective liquid chromatography-tandem mass spectrometric (LC-MS/MS) method was developed and validated for the determination of sodium cromoglycate (SCG) in human plasma after a nasal dose of 10.4 mg sodium cromoglycate nasal spray, using pravastatin sodium as the internal standard. The method was validated over a linear range of 0.300-20.0 ng/mL. SCG and I.S. were extracted from 1.0 mL of heparinized plasma by C(18) solid-phase extraction cartridges using methanol as eluting solvent. The dried residue was reconstituted with 100 microL of mobile phase, and 10 microL was injected onto the LC-MS/MS system. Chromatographic separation was achieved on a C(18) column (250 x 4.6 mm i.d., 5 microm particle size) with a mobile phase of methanol-acetonitrile-water (containing 2 mmol/L ammonium acetate; 42.5:42.5:15, v/v/v) at a flow rate of 0.4 mL/min. The analytes were detected with a triple quad LC-MS/MS using ESI with positive ionization. Ions monitored in the multiple reaction monitoring mode were m/z 469.0 (precursor ion) to m/z 245.0 (product ion) for SCG and m/z 447.2 (precursor ion) to m/z327.1 (product ion) for pravastatin sodium (internal standard) The average recovery of SCG from human plasma was 94.88% and the lower limit of quantitation was 0.3 ng/mL. Results from a 3-day validation study demonstrated excellent precision and accuracy across the calibration range of 0.3-20 ng/mL. The method was successfully applied to the pharmacokinetic study of SCG in healthy Chinese volunteers. Copyright (c) 2008 John Wiley & Sons, Ltd.     

Quantification of zolpidem in human plasma by liquid chromatography-electrospray ionization tandem mass spectrometry

A simple and robust method for quantification of zolpidem in human plasma has been established using liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI MS/MS). Es-citalopram was used as an internal standard. Zolpidem and internal standard in plasma sample were extracted using solid-phase extraction cartridges (Oasis HLB, 1 cm3/30 mg). The samples were injected into a C8 reversed-phase column and the mobile phase used was acetonitrile-ammonium acetate (pH 4.6; 10 mm) (80:20, v/v) at a flow rate of 0.7 mL/min. Using MS/MS in the selected reaction-monitoring (SRM) mode, zolpidem and Es-citalopram were detected without any interference from human plasma matrix. Zolpidem produced a protonated precursor ion ([M+H]+) at m/z 308.1 and a corresponding product ion at m/z 235.1. The internal standard produced a protonated precursor ion ([M+H]+) at m/z 325.1 and a corresponding product ion at m/z 262.1. Detection of zolpidem in human plasma by the LC-ESI MS/MS method was accurate and precise with a quantification limit of 2.5 ng/mL. The proposed method was validated in the linear range 2.5-300 ng/mL. Reproducibility, recovery and stability of the method were evaluated. The method has been successfully applied to bioequivalence studies of zolpidem.     

Determination of caudatin-2,6-dideoxy-3-O-methy-beta-d-cymaropyranoside in rat plasma using liquid chromatography-tandem mass spectrometry

A selective, sensitive and rapid liquid chromatography-tandem mass spectrometry method has been developed for the determination of caudatin-2,6-dideoxy-3-O-methy-beta-d-cymaropyranoside (CDMC) in rat plasma. This method involves a plasma clean-up step using liquid-liquid extraction, followed by LC separation and positive electrospray ionization mass spectrometry detection (LC/ESI-MS/MS). Chromatographic separation of the analytes was achieved using a C(18) column with a mobile phase of acetonitrile and water (70:30, v/v) at a flow rate of 1.0 mL/min. Low energy collision tandem mass spectrometric analysis (CID-MS/MS) using the multiple reaction monitoring (MRM) mode was used for analyte quantification. For the MRM analysis of CDMC, the following transition at m/z 658.4 --> 529.6 derived from the protonated molecule [M + Na](+). A calibration curve was linear in the 5-500 ng/mL range for CDMC, and the limit of detection was 5 ng/mL. The inter- and intra-day precisions (RSD) were 相似文献   

Development and validation of a highly sensitive LC-MS/MS method for quantitation of dexlansoprazole in human plasma: application to a human pharmacokinetic study

A highly sensitive, specific and simple LC-MS/MS method was developed for the simultaneous estimation of dexlansoprazole (DEX) with 50 μL of human plasma using omeprazole as an internal standard (IS). The API-4000 LC-MS/MS was operated under multiple reaction-monitoring mode using electrospray ionization. A simple liquid-liquid extraction process was used to extract DEX and IS from human plasma. The total run time was 2.00 min and the elution of DEX and IS occurred at 1.20 min. This was achieved with a mobile phase consisting of 0.2% ammonia-acetonitrile (20:80, v/v) at a flow rate of 0.50 mL/min on an X-terra RP 18 (50 × 4.6 mm, 5 μm) column. The developed method was validated in human plasma with a lower limit of quantitation of 2 ng/mL for DEX. A linear response function was established for the range of concentrations 2.00-2500.0 ng/mL (r > 0.998) for DEX. The intra- and inter-day precision values for DEX met the acceptance criteria as per FDA guidelines. DEX was stable in the battery of stability studies, viz. bench-top, auto-sampler and freeze-thaw cycles. The developed assay method was applied to an oral bioequivalence study in humans.     

Development and validation of LC‐MS/MS method for the estimation of β‐hydroxy‐β‐methylbutyrate in rat plasma and its application to pharmacokinetic studies

A simple, sensitive and specific high‐performance liquid chromatography mass spectrometry (LC‐MS/MS) method was developed and validated for the quantification of β‐hydroxy‐β‐methyl butyrate (HMB) in small volumes of rat plasma using warfarin as an internal standard (IS). The API‐4000 LC‐MS/MS was operated under the multiple reaction‐monitoring mode using the electrospray ionization technique. A simple liquid–liquid extraction process was used to extract HMB and IS from rat plasma. The total run time was 3 min and the elution of HMB and IS occurred at 1.48 and 1.75 min respectively; this was achieved with a mobile phase consisting of 0.1% formic acid in a water–acetonitrile mixture (15:85, v/v) at a flow rate of 1.0 mL/min on a Agilent Eclipse XDB C₈ (150 × 4.6, 5 µm) column. The developed method was validated in rat plasma with a lower limit of quantitation of 30.0 ng/mL for HMB. A linear response function was established for the range of concentrations 30–4600 ng/mL (r > 0.998) for HMB. The intra‐ and inter‐day precision values for HMB were acceptable as per Food and Drug Administration guidelines. HMB was stable in the battery of stability studies, viz. bench‐top, autosampler freeze–thaw cycles and long‐term stability for 30 days in plasma. The developed assay method was applied to a bioavailability study in rats. Copyright © 2012 John Wiley & Sons, Ltd.     

Rapid quantification of lisinopril in human plasma by liquid chromatography/tandem mass spectrometry

An assay based on protein precipitation and liquid chromatography/tandem mass spectrometry (LC-MS/MS) has been developed and validated for the quantitative analysis of lisinopril in human plasma. After the addition of enalaprilat as internal standard (IS), plasma samples were prepared by one-step protein precipitation using perchloric acid followed by an isocratic elution with 10 mm ammonium acetate buffer (pH adjusted to 5.0 with acetic acid)-methanol (70:30, v/v) on a Phenomenex Luna 5 mu C(18) (2) column. Detection was performed on a triple-quadrupole mass spectrometer utilizing an electrospray ionization (ESI) interface operating in positive ion and selected reaction monitoring (SRM) mode with the precursor to product ion transitions m/z 406 --> 246 for lisinopril and m/z 349 --> 206 for enalaprilat. Calibration curves of lisinopril in human plasma were linear (r = 0.9973-0.9998) over the concentration range 2-200 ng/mL with acceptable accuracy and precision. The limit of detection and lower limit of quantification in human plasma were 1 and 2 ng/mL, respectively. The validated LC-MS/MS method has been successfully applied to a preliminary pharmacokinetic study of lisinopril in Chinese healthy male volunteers.     

Determination and pharmacokinetics of manidipine in human plasma by HPLC/ESIMS

A sensitive HPLC/ESIMS method was established for the determination of manidipine in human plasma and pharmacokinetics study. After basified plasma with ammonia, manidipine and the internal standard (IS) (felodipine) were extracted with n-hexane and separated on a Hypersil ODS2 column with a mobile phase of methanol-5 mm ammonium acetate solution containing 0.1% acetic acid (85:15, v/v). MS determination was performed by electrospray ionization in the selected ion monitoring mode. Manidipine was monitored at m/z 611.4 and IS at m/z 384. The assay had a calibration range from 0.2 to 20 ng/mL and a lower limit of quantification of 0.1 ng/mL. The method has been successfully applied to the pharmacokinetic study in healthy volunteers.     

Simultaneous determination of 18alpha- and 18beta-glycyrrhetic acid in human plasma by LC-ESI-MS and its application to pharmacokinetics

A highly sensitive and selective LC-ESI-MS was developed, validated for the simultaneous determination of 18alpha-glycyrrhetic acid (alpha-GA) and 18beta-glycyrrhetic acid (beta-GA) for pharmacokinetic studies in healthy subjects. Sample preparation was performed by liquid-liquid extraction with ethyl acetate and the separations were achieved using a C(18) column with the mobile phase composed of 10 mmol/L ammonium acetate solution-methanol-acetonitrile (40:36:24, v/v/v) at a flow rate of 1 mL/min. The internal standard was honokiol and the epimers were quantified using a single quadrupole mass spectrometer employing ESI in the negative ion mode. The separation factor, alpha, was 1.512 for alpha- and beta-GA. The standard curves were linear for both epimers with coefficients of determination (r >or= 0.9998) over the concentration range of 1-150 ng/mL. The precision and accuracy were 相似文献   

Delapril and manidipine measurements by liquid chromatography-tandem mass spectrometry in a pharmaceutical formulation

A simple, specific, fast and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the simultaneous analysis of delapril (DEL) and manidipine (MAN) from their combination formulation was developed and validated using fesoterodine as the internal standard (IS). The LC-MS/MS method was carried out on a Luna C8 column (50 × 3.0 mm i.d., 3 μm) with a mobile phase consisting of methanol and 10 mmol L(-1) ammonium acetate (90 : 0, v/v), run at a flow rate of 0.25 mL min(-1). The mass spectrometry method was performed employing positive electrospray ionization operating in multiple reaction monitoring mode, monitoring the transitions of m/z 453.1 → 234.1 for DEL, m/z 611.1 → 167.0 for MAN and m/z 412.2 → 223.0 for IS. The total analysis time was 3 min and the method was linear in the concentration range of 6-1080 ng mL(-1) and 2-360 ng mL(-1) for DEL and MAN, respectively. Parameters investigated for the method validation, such as the specificity, linearity, precision, accuracy and robustness, gave results within the acceptable range. Moreover, the proposed method was successfully applied for the simultaneous determination of DEL and MAN and the results were compared to validated liquid chromatography and capillary electrophoresis methods showing non-significant differences (P = 0.9).     

Selective and sensitive determination of bis(7)-tacrine, a high erythrocyte binding acetylcholinesterase inhibitor, in rat plasma by high-performance liquid chromatography-tandem mass spectrometry

The current study aims to develop a specific and sensitive LC-MS/MS method for determination of bis(7)-tacrine (B7T) in rat plasma. A 100 microL plasma sample was extracted with ethyl acetate. B7T and the internal standard (IS), pimozide, in the samples were then analyzed with LC-MS/MS in positive electrospray ionization condition. Chromatographic separation of B7T and IS was achieved in a C(18) reversed-phase HPLC column (150 x 2.1 mm i.d.) by isocratic elution with a mobile phase consisting of 0.05% formic acid in water and acetonitrile (1:1, v/v) at a flow rate of 0.35 mL/min. Multiple-reaction monitoring (MRM) mode was employed to measure the ion transitions: m/z 247 to 197 for B7T and m/z 462 to m/z 328 for IS, respectively. The method was linear over the studied ranges of 100-5000 and 10-100 ng/mL. The intra-day and inter-day variations of the analysis were less than 6.8% with standard errors less than 9.0%. The detection limit of B7T in rat plasma was 1 ng/mL. The developed method was successfully applied to the pharmacokinetic study of B7T after intravenous administration of 1 mg/kg B7T and further proved to be readily utilized for determination of B7T in rat plasma samples.     

A simple and sensitive assay for the quantitative analysis of paclitaxel and metabolites in human plasma using liquid chromatography/tandem mass spectrometry

A sensitive and specific LC-MS/MS assay for the determination of paclitaxel and its 3'p- and 6-alpha-hydroxy metabolites is presented. A 200 microL plasma aliquot was spiked with a 13C6-labeled paclitaxel internal standard and extracted with 1.0 mL tert-butylmethylether. Dried extracts were reconstituted in 0.1 M ammonium acetate-acetonitrile (1:1, v/v) and 25 microL volumes were injected onto the HPLC system. Separation was performed on a 150 x 2.1 mm C18 column using an alkaline eluent (10 mm ammonium hydroxide-methanol, 30:70, v/v). Detection was performed by positive ion electrospray followed by tandem mass spectrometry. The assay quantifies a range for paclitaxel from 0.25 to 1000 ng/mL and metabolites from 0.25 to 100 ng/mL using 200 microL human plasma samples. Validation results demonstrate that paclitaxel and metabolite concentrations can be accurately and precisely quantified in human plasma. This assay is now used to support clinical pharmacologic studies with paclitaxel.     

Liquid chromatography/electrospray ionization tandem mass spectrometry for the quantification of mitiglinide in human plasma: validation and its application to pharmacokinetic studies

A sensitive and specific method was developed and validated for the determination of mitiglinide in human plasma using liquid chromatographic separation with electrospray ionization tandem mass spectrometric detection. Acidified plasma samples were extracted with ethyl acetate. The chromatographic separation was performed on an Agilent Zorbax Eclipse Plus C(18) column with a mobile phase of methanol-10 mm ammonium acetate solution at a flow rate of 0.3 mL/min. Analytes were detected with an Agilent 6410 Triple qudrupole mass spectrometer equipped with an electrospray ionization source in positive multiple reaction monitoring mode: m/z 316.2 (precursor ion) to 298.2 (product ion) for mitiglinide and m/z 318.2 (precursor ion) to 120.2 (product ion) for the internal standard. This method was validated over a linear range of 0.5-4000 ng/mL for mitiglinide in human plasma. The lower limit of quantification (LLOQ) was 0.5 ng/mL, while a relative standard deviation (RSD) was less than 3.9%. The intra- and inter-run precision (as RSD, %) obtained from three validation runs were all less than 15%. The validated method was successfully used to analyze human plasma samples for application in pharmacokinetic studies.     

A simple,rapid and sensitive liquid chromatography–tandem mass spectrometry method for the determination of dienogest in human plasma and its pharmacokinetic applications under fasting

A simple, rapid and sensitive analytical method using liquid chromatography coupled to tandem mass spectrometry (LC‐MS/MS) detection with positive ion electrospray ionization was developed for the determination of dienogest in human K₂EDTA plasma using levonorgestrel d₆ as an internal standard (IS). Dienogest and IS were extracted from human plasma using simple liquid–liquid extraction. Chromatographic separation was achieved on a Zorbax XDB‐Phenyl column (4.6 × 75 mm, 3.5 µm) under isocratic conditions using acetonitrile–5 mm ammonium acetate (70:30, v/v) at a flow rate of 0.60 mL/min. The protonated precursor to product ion transitions monitored for dienogest and IS were at m/z 312.30 → 135.30 and 319.00 → 251.30, respectively. The method was validated with a linearity range of 1.003–200.896 ng/mL having a total analysis time for each chromatograph of 3.0 min. The method has shown tremendous reproducibility with intra‐ and inter‐day precision (coefficient of variation) <3.97 and 6.10%, respectively, and accuracy within ±4.0% of nominal values. The validated method was applied to a pharmacokinetic study in human plasma samples generated after administration of a single oral dose of 2.0 mg dienogest tablets to healthy female volunteers and was proved to be highly reliable for the analysis of clinical samples. Copyright © 2014 John Wiley & Sons, Ltd.     

A rapid and sensitive LC‐MS/MS method for quantification of 3,29‐dibenzoyl rarounitriol in rat plasma: application to a pharmacokinetic study

A rapid, sensitive and high‐throughput liquid chromatography–tandem mass spectrometry was established and validated to assay the concentrations of 3,29‐dibenzoyl rarounitriol in rat plasma. Plasma samples were processed by liquid–liquid extraction with ethyl acetate and separated on a Hypersil Gold C₁₈ column (50 × 4.6 mm, 3 µm) at an isocratic flow rate of 0.5 mL/min using methanol–10 mm ammonium acetate–formic acid (90:10:0.1, v/v/v) as mobile phase. The total run time was 5 min for each sample. MS/MS detection was accomplished in selected reaction monitoring mode with positive electrospray ionization. The calibration curve was linear over the concentration range of 0.125–50 ng/mL with lower limit of quantification of 0.125 ng/mL. The intra‐ and inter‐day precisions were <10.1% in terms of coefficient of variation, and the accuracy was within ±11.7% in terms of relative error. The developed method was successfully applied to a pharmacokinetic study of 3,29‐dibenzoyl rarounitriol following intragastric administration of 3.65 mg/kg to Wistar rats. Copyright © 2015 John Wiley & Sons, Ltd.     

A highly sensitive LC-MS/MS method capable of simultaneously quantitating celiprolol and atenolol in human plasma for a cassette cold-microdosing study

A highly sensitive simultaneous quantitative method for a cassette cold-microdosing study on celiprolol and atenolol was developed with liquid chromatography-tandem mass spectrometry. The method utilizes a combination of solid-phase extraction (SPE) with strong cation exchange (SCX) cartridge columns and reversed-phase chromatography with an ODS analytical column. SCX-SPE cartridge columns (100 mg sorbent) were used for a selective extraction of celiprolol, atenolol and metoprolol (internal standard) from 500 μL of human plasma samples. Turbo-ion spray at positive mode was employed for the ionization of the drug compounds. Quantitation was performed on a triple quadrupole mass spectrometer by selected reaction monitoring with the transitions of m/z 380 to m/z 251 for celiprolol and m/z 267 to m/z 145 for atenolol. Separation of analytes was achieved on an ODS column (100 mm length × 2.1 mm id, 3 μm) by a gradient elution with 10 mM formic acid and methanol by varying their proportion at a flow rate of 0.2 mL/min. The method was validated in the range of 1-250 pg/mL for celiprolol and 2.5-250 pg/mL for atenolol and was successfully applied to the elucidation of pharmacokinetic profiling in a cold cassette microdosing study of the β-blockers.     

Simultaneous determination of amiloride and hydrochlorothiazide in human plasma by liquid chromatography/tandem mass spectrometry with positive/negative ion-switching electrospray ionisation

A new method for simultaneous determination of amiloride and hydrochlorothiazide by liquid chromatography/electrospray tandem mass spectrometry (LC/MS/MS) operated in positive and negative ionization switching mode was developed and validated. Protein precipitation with acetonitrile was selected for sample preparation. The analytes were separated on a Phenomenex Curosil-PFP (250x4.6 mm, 5 microm) column by a gradient elution with a mobile phase consisting of 0.15% formic acid solution containing 0.23% ammonium acetate and methanol pumped at a flow rate of 1.0 mL.min(-1). Rizatriptan was used as the internal standard (IS) for quantification. The determination was carried out on a Waters Quattro-micro triple-quadrupole mass spectrometer operated in multiple reaction monitoring (MRM) mode using the following transitions monitored simultaneously: positive m/z 230-->171 for amiloride, m/z 270-->158 for rizatriptan, and negative m/z 296-->205 for hydrochlorothiazide. The lower limits of quantification (LLOQs) were 0.1 and 1.0 ng.mL(-1) for amiloride and hydrochlorothiazide, respectively, which were lower than other published methods by using ultraviolet (UV), fluorimetric or mass spectrometric detection. The intra- and inter-day precision and accuracy were studied at three different concentration levels and were always better than 15% (n=5). This simple and robust LC/MS/MS method was successfully applied to the pharmacokinetic study of compound amiloride and hydrochlorothiazide tablets in healthy male Chinese volunteers.     

Simultaneous quantification of oxysophocarpine and its active metabolite sophocarpine in rat plasma by liquid chromatography/mass spectrometry for a pharmacokinetic study

A sensitive, rapid and specific method for the simultaneous quantification of oxysophocarpine (OSC) and its active metabolite sophocarpine (SC) in rat plasma was developed and validated, using a liquid-liquid extraction procedure followed by liquid chromatography/electrospray ionization mass spectrometric (LC/ESI-MS) analysis. The separation was performed on a Zorbax Extend-C(18) column (2.1 mm i.d. x 50 mm, 5 microm) with a C(18) guard column using methanol-water containing 5 mm ammonium acetate (15:85, v/v) as mobile phase. Analysis was performed in selected ion monitoring (SIM) mode with an electrospray ionization (ESI) interface. [M + H](+) at m/z 263 for OSC, [M + H](+) at m/z 247 for SC and [M + H](+) at m/z 249 for matrine (internal standard) were selected as detecting ions, respectively. The method was linear in the concentration ranges 10-1000 ng/mL for OSC and 5-500 ng/mL for SC. The intra- and inter-day precisions (coefficient of variation) were within 7% for both analytes. Their accuracy (relative error) ranged from -6.4 to 1.5%. The limits of detection for OSC and SC were 3 and 1.5 ng/mL, respectively. The limits of quantitation for OSC and SC were 10 and 5 ng/mL, respectively. Recoveries of both analytes were greater than 85% at the low, medium and high concentrations. Both analytes were stable during all sample storage, preparation and analytic procedures. The method was successfully applied to a pharmacokinetic study after an oral administration of OSC to rats with a dose of 15 mg/kg.     

Development and validation of a sensitive LC-MS/MS method with electrospray ionization for quantitation of zafirlukast, a selective leukotriene antagonist in human plasma: application to a clinical pharmacokinetic study

A highly sensitive and specific LC-MS/MS method has been developed and validated for the estimation of zafirlukast (ZFK) with 500 microL human plasma using valdecoxib as an internal standard (IS). The API-4,000 LC-MS/MS was operated under multiple reaction-monitoring mode using the electrospray ionization technique. The assay procedure involved extraction of ZFK and IS from human plasma with ethyl acetate. The resolution of peaks was achieved with 10 mm ammonium acetate (pH 6.4):acetonitrile (20:80, v/v) on a Hypersil BDS C(18) column. The total chromatographic run time was 2.0 min and the elution of ZFK and IS occurred at approximately 1.11 and 1.58 min, respectively. The MS/MS ion transitions monitored were 574.2 --> 462.1 for ZFK and 313.3 --> 118.1 for IS. The method was proved to be accurate and precise at a linearity range of 0.15-600 ng/mL with a correlation coefficient (r) of >or=0.999. The method was rugged with 0.15 ng/mL as lower limit of quantitation. The intra- and inter-day precision and accuracy values were found to be within the assay variability limits as per the FDA guidelines. The developed assay method was applied to a pharmacokinetic study in human volunteers following oral administration of 20 mg ZFK tablet.     

Development of a fast LC‐MS/MS assay for the determination of deferiprone in human plasma and application to pharmacokinetics

A fast and accurate liquid chromatography/tandem mass spectrometric (LC‐MS/MS) assay was first developed and validated for the determination of deferiprone in human plasma. The analytes were extracted with acetonitrile from only 50 μL aliquots of human plasma to achieve the protein precipitation. After extraction, chromatographic separation of analytes in human plasma was performed using a Synergi Fusion‐RP 80A column at 30 °C. The mobile phase consisted of methanol and 0.2% formic acid containing 0.2 mM EDTA (60:40, v/v). The flow rate of the mobile phase was 0.8 mL/min. The total run time for each sample analysis was 4 min. Detection was performed using electrospray ionization in positive ion multiple reaction monitoring mode by monitoring the precursor‐to‐parent ion transitions m/z 140.1 → 53.1 for deferiprone and m/z 143.1 → 98.1 for internal standard. A linear range was established from 0.1 to 20 µg/mL. The limit of detection was determined as 0.05 µg/mL. The validated method was estimated for linearity, recovery, stability, precision and accuracy. Intraday and interday precisions were 4.3–5.5 and 4.6–7.3%, respectively. The recovery of deferiprone was in the range of 80.1–86.8%. The method was successfully applied to a pharmacokinetic study of deferiprone in six thalassemia patients. Copyright © 2012 John Wiley & Sons, Ltd.