Validated LC-MS/MS method for quantification of gabapentin in human plasma: application to pharmacokinetic and bioequivalence studies in Korean volunteers

A sensitive validated liquid chromatography-tandem mass spectrometric method (LC-MS/MS) for gabapentin (GB) in human plasma has been developed and applied to pharmacokinetic (PK) and bioequivalence (BE) studies in human. In a randomized crossover design with a 1-week period, each subject received a 300 mg GB capsule. The procedure involves a simple protein precipitation with acetonitrile and separated by LC with a Gemini C(18) column using acetonitrile-10 mm ammonium acetate (20:80, v/v, pH 3.2) as mobile phase. The GB and internal standard [(S)-(+)-alpha-aminocyclohexanepropionic acid hydrate] were analyzed using an LC-API 2000 MS/MS in multiple reaction monitoring mode. The ionization was optimized using ESI(+) and selectivity was achieved using MS/MS analysis, m/z 172.0 --> 154.0 and m/z 172.0 --> 126.0 for GB and IS, respectively. The assay exhibited good linearity over a working range of 20-5000 ng/mL for GB in human plasma with a lower limit of quantitation of 20 ng/mL. No endogenous compounds were found to interfere with the analysis. The accuracy and precision were shown for concentrations over the standard ranges. This method was successfully applied for the PK and BE studies by analysis of blood samples taken up to 36 h after an oral dose of 300 mg of GB in 24 healthy volunteers.     

Rapid and sensitive liquid chromatography tandem mass spectrometry method for the quantification of ambroxol in human plasma

A sensitive, specific and rapid high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was described and validated for the quantification of ambroxol in human plasma using enalaprilat as the internal standard (IS). Chromatographic separation was performed on a Lichrospher CN column with a mobile phase of methanol and water (containing 0.1% formic acid) (70:30, v/v). The total run time was 5.0 min for each sample. The analytes was detected by mass spectrometry with electrospray ionization source in positive selected reaction monitoring mode. The precursor-fragment ion reaction for ambroxol was m/z 378.9 --> 263.8, and for IS was m/z 349.0 --> 205.9. The linearity was established over the concentration range of 1.56-400.00 ng/mL. The inter-day and the intra-day precisions were all within 10%. A simple protein precipitation with methanol was adopted for sample preparation. The extraction recoveries of ambroxol and IS were higher than 90.80%. The validated method was successfully applied in pharmacokinetic study after oral administration of 90 mg ambroxol to 24 healthy volunteers.     

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.     

Development of a rapid and sensitive liquid chromatography/tandem mass spectrometry method for the determination of 1,2-[bis(1,2-benzisoselenazolone-3(2H)-ketone)]-ethane (BBSKE), a novel anti-cancer agent in rat plasma

A rapid and sensitive liquid chromatography/electrospray ionization tandem mass spectrometric (LC/ESI-MS/MS) method has been developed to determine 1, 2-[bis(1,2-benzisoselenazolone-3(2H)-ketone)]-ethane (BBSKE), a novel antineoplastic agent, in rat plasma. The analytes were separated on a C18 column with a mobile phase of methanol-water (75:25, v/v) and detected using a triple-quadrupole mass spectrometer in positive mode with the selective reaction monitoring. The characteristic ion dissociation transitions were m/z 603.0 --> 448.9 for derivatized BBSKE and m/z 631.0 --> 476.8 for derivatized internal standard. The assay was linear over a range of 1-1000 ng/mL with a lower limit of quantification of 1 ng/mL. Intra- and inter-day precisions were less than 9.6 and 5.0%, respectively, and the accuracy ranged from -5.2 to 4.0%. The validated method was successfully applied to the characterization of pharmacokinetic profile of BBSKE after oral administration in rats. Cop     

Development and validation of a liquid chromatographic/tandem mass spectrometric method for the determination of sertraline in human plasma

A sensitive and rapid liquid chromatographic/tandem mass spectrometric method was developed and validated for the determination of sertraline in human plasma. The analyte and internal standard (IS, diphenhydramine) were extracted with 3 mL of diethyl ether/dichloromethane (2:1, v/v) from 0.25 mL plasma, then separated on a Zorbax Eclipse XDB C18 column using methanol/water/formic acid (75:25:0.1, v/v/v) as the mobile phase. The triple quadrupole mass spectrometry was applied via an atmospheric pressure chemical ionization (APCI) source for detection. The fragmentation pattern of the protonated sertraline was elucidated with the aid of product mass spectra of isotopologous peaks. Quantification was performed using selected reaction monitoring of the transitions of m/z 306 --> 159 for sertraline and m/z 256 --> 167 for the IS. The method was linear over the concentration range of 0.10-100 ng/mL. The intra-day and inter-day precisions, expressed by relative standard deviation, were both less than 6.7%. Assay accuracies were within +/-6.9% as terms of relative error. The lower limit of quantification (LLOQ) was identifiable and reproducible at 0.10 ng/mL with a precision of 8.3% and an accuracy of 9.6%. The validated method has been successfully applied for the pharmacokinetic study and bioequivalence evaluation of sertraline in 18 healthy volunteers after a single oral administration of 50 mg sertraline hydrochloride tablets.     

Simultaneous determination of udenafil and its active metabolite, DA-8164, in human plasma and urine using ultra-performance liquid chromatography-tandem mass spectrometry: application to a pharmacokinetic study

A rapid, sensitive, and simple ultra-performance liquid chromatography-tandem mass spectrometry (UPLC/MS/MS) method for the determination of udenafil and its active metabolite, DA-8164, in human plasma and urine using sildenafil as an internal standard (IS) was developed and validated. Udenafil, DA-8164 and IS from a 100 microL aliquot of biological samples were extracted by protein precipitation using acetonitrile. Chromatographic separation was carried on an Acquity UPLC BEH C(18) column (50 x 2.1 mm, i.d., 1.7 microm) with an isocratic mobile phase consisting of acetonitrile and containing 0.1% formic acid (75:25, v/v) at flow rate of 0.4 mL/min, and total run time was within 1 min. Detection and quantification was performed by the mass spectrometer using multiple reaction-monitoring mode at m/z 517 --> 283 for udenafil, m/z 406 --> 364 for DA-8164 and m/z 475 --> 100 for IS. The assay was linear over a concentration range of 1-600 ng/mL with a lower limit of quantification of 1 ng/mL in both human plasma and urine. The coefficient of variation of this assay precision was less than 13.7%, and the accuracy exceeded 92.0%. This method was successfully applied for pharmacokinetic study after oral administration of udenafil 100 mg to healthy Korean male volunteers.     

超高效液相色谱-质谱/质谱联用法测定人血浆中的辛伐他汀

建立了超高效液相色谱-质谱/质谱联用法(UPLC-MS/MS)测定人血浆中辛伐他汀的浓度。血浆样品经乙醚-正己烷-异丙醇(体积比为80∶20∶3)提取,以洛伐他汀为内标,采用ACQUITY UPLCTM BEH C18柱(50 mm×2.1 mm,1.7 μm)分离,以乙腈-10 mmol/L乙酸铵水溶液(体积比为85∶15)为流动相,流速为0.25 mL/min,通过电喷雾离子化,采用多反应监测(MRM)方式进行正离子检测。线性范围为0.051～20.4 ng/mL,日内及日间测定的相对标准偏差不高于10%,平均回收率为91.6%。方法灵敏度高,分析速度快,操作简便,适用于辛伐他汀药物动力学和生物等效性研究。     

Determination of bergenin in human plasma after oral administration by HPLC-MS/MS method and its pharmacokinetic study

A highly sensitive, simple and selective high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) method was developed and applied to the determination of bergenin concentration in human plasma. Bergenin and the internal standard (IS) thiamphenicol in plasma were extracted with ethyl acetate, separated on a C(18 )reversed-phase column, eluted with mobile phase of acetonitrile-water, ionized by negative ion pneumatically assisted electrospray and detected in the multi-reaction monitoring mode using precursor --> product ions of m/z 327.1 --> 192 for bergenin and 354 --> 185.1 for the IS, respectively. The linear range of the calibration curve for bergenin was 0.25-60 ng mL(-1), with the lowest limit of quantification of 0.25 ng mL(-1), and the intra/inter-day relative standard deviation (RSD) was less than 10%. The method is suitable for the determination of low bergenin concentration in human plasma after therapeutic oral doses, and has been first and successfully used for its pharmacokinetic studies in healthy Chinese volunteers.     

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.     

Rapid and sensitive determination of donepezil in human plasma by liquid chromatography/tandem mass spectrometry: application to a pharmacokinetic study

A liquid chromatography/tandem mass spectrometry (LC/MS/MS) method was developed and validated for the determination of donepezil in human plasma samples. Diphenhydramine was used as the internal standard. The collision-induced transition m/z 380 --> 91 was used to analyze donepezil in selected reaction monitoring mode. The signal intensity of the m/z 380 --> 91 transition was found to relate linearly with donepezil concentrations in plasma from 0.1-20.0 ng/mL. The lower limit of quantification of the LC/MS/MS method was 0.1 ng/mL. The intra- and inter-day precisions were below 10.2% and the accuracy was between -2.3% and +2.8%. The validated LC/MS/MS method was applied to a pharmacokinetic study in which healthy Chinese volunteers each received a single oral dose of 5 mg donepezil hydrochloride. The non-compartmental pharmacokinetic model was used to fit the donepezil plasma concentration-time curve. Maximum plasma concentration was 12.3 +/- 2.73 ng/mL which occurred at 3.50 +/- 1.61 h post-dosing. The apparent elimination half-life and the area under the curve were, respectively, 60.86 +/- 12.05 h and 609.3 +/- 122.2 ng . h/mL. LC/MS/MS is a rapid, sensitive and specific method for determining donepezil in human plasma samples.     

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.     

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.     

Liquid chromatographic-electrospray tandem mass spectrometric determination of bisoprolol in human plasma

An analytical method for the determination of bisoprolol in human plasma has been developed based on liquid chromatography-tandem mass spectrometry (LC-MS/MS). The analyte and internal standard (IS) diphenhydramine were cleaned up by protein precipitation with acetonitrile, reconstituted in mobile phase and separated by reversed-phase high-performance liquid chromatography (HPLC) using methanol:10 mm ammonium acetate:formic acid (70:30:0.1 v/v/v) as mobile phase. Detection was carried out by multiple reaction monitoring (MRM) on an LC-MS/MS system and was completed within 2.5 min. The assay was linear over the range 0.5-100 ng/mL with a limit of quantitation (LOQ) of 0.5 ng/mL. The intra- and inter-day precision levels were within 5.54 and 9.95%, respectively, while the accuracy was in the range 89.4-113%. This method has been utilized in a pharmacokinetic study, where healthy volunteers were treated with an oral dose of 5 mg bisoprolol.     

Determination of clopidogrel in human plasma by liquid chromatography/tandem mass spectrometry: application to a clinical pharmacokinetic study

A rapid and sensitive LC/MS/MS assay was developed and validated for the determination of clopidogrel in human plasma. Clopidogrel was extracted by single liquid-liquid extraction with pentane, and chromatographic separations were achieved on a C(18) column. The method was validated to demonstrate the specificity, linearity, recovery, lower limit of quantification (LLOQ), stability, accuracy and precision. The multiple reaction monitoring was based on m/z transition of 322.2 --> 211.9 for clopidogrel and 264.1 --> 125.1 for ticlopidine (internal standard). The total analytical run time was relatively short (3 min), and the LLOQ was 10 pg/mL using 0.5 mL of human plasma. The assay was linear over a concentration range from 10 to 10,000 pg/mL (r > 0.999). The intra- and inter-day accuracies were 101.3-108.8 and 98.4-103.5%, respectively, and the intra- and inter-day assay precisions were 1.9-5.5 and 4.4-8.1%, respectively. The developed assay method was applied to a pharmacokinetic study in human volunteers after oral administration of clopidogrel at a dose of 150 mg.     

Determination and pharmacokinetic study of norcantharidin in human serum by HPLC-MS/MS method

A sensitive, simple and selective high-performance liquid chromatography-tandem mass spectrometry method was developed and applied to the determination of norcantharidin concentration in human serum. Norcantharidin (NCTD) and cyclophosphamide (IS) in serum were extracted with acetone, separated on a C18 reversed-phase column, gradiently eluted with a mobile phase of acetonitrile-water containing 2 mm ammonium acetate and 0.1% formic acid (pH 3), ionized by positive ion pneumatically assisted electrospray and detected in the multi-reaction monitoring mode using precursor-->product ions of m/z 169.3-->123.1 for NCTD and 261.2-->140.2 for IS, respectively. The linear range of the calibration curve for NCTD was 2.5-50 ng/mL, with a lowest limit of quantification of 2.5 ng/mL, and the intra/inter-day RSD was less than 10%. The method was suitable for determination of low NCTD concentration in human serum after therapeutic oral doses, and has been successfully used for pharmacokinetic studies in healthy Chinese volunteers.     

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.     

Development and validation of a hydrophilic interaction liquid chromatography-tandem mass spectrometric method for the analysis of paroxetine in human plasma

A sensitive, simple, fast and rugged hydrophilic interaction liquid chromatography-tandem mass spectrometry (HILIC-MS/MS) method for the determination of paroxetine was developed and validated over curve range 0.050-50 ng/mL using only 0.4 mL plasma. This is the first published LC-MS/MS method and the low limit of quantitation of this method is 10-fold lower than previously published methods. A simple liquid-liquid extraction method using methyl-tert butyl ether (MTBE) as the extraction solvent was used to extract paroxetine and the internal standard (IS) fentanyl-d(5) from plasma. The extract was evaporated to dryness, reconstituted and injected onto a silica column using a low aqueous-high organic mobile phase. The chromatographic run time was 2.0 min per injection, with retention times of 1.1 and 1.2 min for paroxetine and IS, respectively. The detection was by monitoring paroxetine at m/z 330 --> 192 and IS at m/z 342 --> 188, respectively. The inter-day precision and accuracy of the quality control (QC) samples were <5.0% relative standard deviation (RSD) and <2.9% relative error (RE). This method can be used for supporting therapeutical drug monitoring and pharmacokinetic or drug-drug interaction studies.     

Determination of lovastatin in human plasma by ultra-performance liquid chromatography/electrospray ionization tandem mass spectrometry

A fast and sensitive ultra-performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) method was developed for the determination of lovastatin in human plasma. With simvastatin as internal standard, sample pretreatment involved one-step extraction with n-hexane-methylene dichloride-isopropanol (20:10:1, v/v/v) of 0.5 mL plasma. Chromatographic separation was carried out on an Acquity UPLC BEH C(18) column with mobile phase consisting of acetonitrile-water (containing 5 mmol/L ammonium acetate; 85:15, v/v) at a flow-rate of 0.35 mL/min. The detection was performed on a triple-quadrupole tandem mass spectrometer by multiple reaction monitoring (MRM) via electrospray ionization source with positive mode. The analysis time was shorter than 1.7 min per sample. The standard curve was linear (r2>or=0.99) over the concentration range 0.025-50.0 ng/mL with a lower limit of quantification of 0.025 ng/mL. The intra- and inter-day precision values were below 11% and the accuracy (relative error) was within 6.0% at three quality control levels. This is the first method of MS with MRM coupled to UPLC for the determination of lovastatin, which showed great advantages of high sensitivity, selectivity and high sample throughput. It was fully validated and successfully applied to the pharmacokinetic study of lovastatin tablets in healthy Chinese male volunteers after oral administration.     

Rapid Determination of Ranitidine in Human Plasma by Liquid Chromatography-Tandem Mass Spectrometry and Its Application to a Clinical Pharmacokinetic Study

A rapid and sensitive assay based on high performance liquid chromatography-tandem mass spectrometry(LC-MS/MS) was developed for the determination of ranitidine(RAN) in human plasma with codeine as internal standard(IS).After protein precipitation with acetonitrile,the analyte and IS were separated on a Zorbax SB-Aq C18 column(150 mm×4.6 mm i.d.,5 μm) eluted with a mobile phase consisting of methanol/acetonitrile/10 mmol/L ammonium acetate containing 1% formic acid(pH=2.4)(volume ratio 12.5:12.5:75) at a flow rate of 1.0 mL/min.Detection was performed by electrospray ionization in the positive ion mode followed by the multiple reaction monito-ring(MRM) of the transitions of RAN at m/z 315.1→176.3 and of IS at m/z 300.1→165.1.The method was linear over a concentration range of 1―1000 ng/mL(r=0.9991) with a lower limit of quantitation(LLOQ) of 1 ng/mL and a limit of detection(LOD) of 0.3 ng/mL.Accuracy as relative error was from-0.01% to-1.7% and intra-day and inter-day precisions as relative standard deviation were ≤8.9% and ≤5.5%,respectively.The method was successfully applied to a pharmacokinetic study of ranitidine,getting a single oral dose(160 mg) to healthy volunteers.     

A rapid and specific approach for direct measurement of donepezil concentration in human plasma by LC-MS/MS employing solid-phase extraction

A selective, rapid and simple liquid chromatography-tandem mass spectrometry (LC-MS/MS) method is described for assay of donepezil in human plasma using escitalopram as an internal standard. Chromatographic separation was achieved on a Betabasic-C(8), 5 microm, 100 x 4.6 mm column using methanol:water:formic acid (90:9.97:0.03, v/v/v) as mobile phase. Detection of donepezil and internal standard was achieved by ESI MS/MS in positive ion mode using 380.20/91.10 and 325.13/262.00 transitions, respectively. The linearity over the concentration range of 0.15-50 ng/mL for donepezil was obtained and the lower limit of quantification was 0.15 ng/mL. For each level of quality control samples, inter-day and intra-day precisions (RSD) were < or =8.92 and 10.35% and accuracy (%RE) were < or =7.33% and 9.33%, respectively. The recovery was more than 88.50% for both donepezil and internal standard by solid-phase extraction, eliminating evaporation and reconstitution steps.