Simultaneous quantitation of dexamethasone palmitate and dexamethasone in human plasma by liquid chromatography/tandem mass spectrometry

A rapid and sensitive liquid chromatography/tandem mass spectrometry (LC/MS/MS) method for simultaneous quantitation of dexamethasone palmitate and dexamethasone in human plasma was developed. After sample preparation by protein precipitation and liquid-liquid extraction, the analytes and internal standard (IS) were separated on a Venusil XBP-C8 column using gradient elution. Multiple reaction monitoring of dexamethasone palmitate, dexamethasone and IS used the precursor to product ion transitions at m/z 631.8-->373.1, m/z 393.2-->147.1 and m/z 264.2-->58.1, respectively. The method was linear over the ranges 1.5-1000ng/mL for dexamethasone palmitate and 2.5-250ng/mL for dexamethasone with intra- and inter-day precisions of <10% and accuracies of 100+/-7%. The assay was applied to a clinical pharmacokinetic study involving the injection of dexamethasone palmitate to healthy volunteers.     

Simultaneous quantitation of aconitine, mesaconitine, hypaconitine, benzoylaconine, benzoylmesaconine and benzoylhypaconine in human plasma by liquid chromatography-tandem mass spectrometry and pharmacokinetics evaluation of "SHEN-FU" injectable powder

A rapid, specific and sensitive liquid chromatography-tandem mass spectrometry (LC/MS/MS) method was developed for simultaneous quantitation of six Aconitum alkaloids, i.e. aconitine (AC), mesaconitine (MA), hypaconitine (HA), benzoylaconine (BAC), benzoylmesaconine (BMA) and benzoylhypaconine (BHA) in human plasma collected from 18 healthy volunteers after intravenous drop infusion of "SHEN-FU" injectable powder in three different dosages. Lappaconitine was selected as the internal standard (IS). LC/MS/MS system coupled with an electrospray ionization (ESI) source was performed in multiple-reaction monitoring (MRM) mode. The transitions of the Aconitum alkaloids executed as following: m/z 646.3-->586.0 for AC; m/z 632.4-->573.1 for MA; m/z 616.2-->556.1 for HA; m/z 604.2-->104.8 for BAC; m/z 590.1-->104.8 for BMA; m/z 574.1-->104.8 for BHA; m/z 585.2-->161.8 for IS. Sample preparation was performed with solid-phase extraction (SPE) on a 1 mL HLB cartridge prior to analysis. The separation was applied on a Waters C(18) column (1.7 microm, 2.1 mm x 100 mm) and a gradient elution of methanol and 0.1% formic acid-water was used as mobile phase. The retention time was less than 4.5 min. The concentrations ranged from 0.1 to 1000 ng/mL for all six Aconitum alkaloids and showed a good linearity with the correlation coefficient (r(2)) >0.995. The validated method was employed to simultaneous quantitation and successfully used for the first time for the pharmacokinetic evaluation of the six Aconitum alkaloids after intravenous drop administration of "SHEN-FU" injectable powder in phase I clinical trial.     

Simultaneous determination of three isomeric metabolites of tacrolimus (FK506) in human whole blood and plasma using high performance liquid chromatography-tandem mass spectrometry

An ammonium-adduct based liquid chromatography-tandem mass spectrometry (LC-MS/MS) method has been developed and validated for the simultaneous determination of three isomeric metabolites of tacrolimus (FK506), 13-O-demethylated (M1), 31-O-demethylated (M2) and 15-O-demethylated (M3) tacrolimus in human whole blood and plasma. These metabolites and the internal standards were extracted from biological matrix by methylbutyl ether (MTBE). Separation was achieved on a Genesis C(18) column with a gradient mobile phase elution. Ammonium-adduct ions formed by a Turbo Ionspray in positive ion mode were used to detect each analyte and internal standard. The MS/MS detection was by monitoring the fragmentation of 807.5-->772.4 (m/z) for M1, 807.5-->754.5 (m/z) for both M2 and M3, 795.5-->760.5 (m/z) for IS1 (FR298701) and 961.5-->908.5 (m/z) for IS2 (FR290198) on a triple quadrupole mass spectrometer (Sciex API 3000). The retention times were approximately 4.1 min for M1, 6.8 min for M2, 6.0 min for M3, and 3.9 min for IS1 and 6.4 min for IS2, respectively. The validated dynamic range was 0.2-20 ng/ml for all three metabolites based on a sample volume of 0.25-ml. The linearity of calibration curves for M1, M2, and M3 in both matrices had a correlation coefficient of >/=0.9984. In whole blood, validation data showed intra-batch (n=6) CVs of 相似文献   

Sensitive and selective liquid chromatography-tandem mass spectrometry method for the quantification of aniracetam in human plasma

A rapid, sensitive and selective LC-MS/MS method was developed and validated for the quantification of aniracetam in human plasma using estazolam as internal standard (IS). Following liquid-liquid extraction, the analytes were separated using a mobile phase of methanol-water (60:40, v/v) on a reverse phase C18 column and analyzed by a triple-quadrupole mass spectrometer in the selected reaction monitoring (SRM) mode using the respective [M+H]+ ions, m/z 220-->135 for aniracetam and m/z 295-->205 for the IS. The assay exhibited a linear dynamic range of 0.2-100 ng/mL for aniracetam in human plasma. The lower limit of quantification (LLOQ) was 0.2 ng/mL with a relative standard deviation of less than 15%. Acceptable precision and accuracy were obtained for concentrations over the standard curve range. The validated LC-MS/MS method has been successfully applied to study the pharmacokinetics of aniracetam in healthy male Chinese volunteers.     

Determination of HS270, a new histone deacetylase inhibitor, in rat plasma by LC-MS/MS--application to a preclinical pharmacokinetic study

A rapid and sensitive liquid chromatography/tandem mass spectrometry (LC-MS/MS) method has been developed and fully validated to determine HS270, a new histone deacetylase (HDAC) inhibitor, in rat plasma using SAHA as the internal standard (IS). After a single step liquid-liquid extraction with acetoacetate, analytes were subjected to LC-MS/MS analysis using positive electro-spray ionization (ESI(+)) under selected reaction monitoring mode (SRM). The chromatographic separation was achieved on a Hypurity C(18) column (50 mm × 2.1 mm, i.d., 5 μm). The MS/MS detection was conducted by monitoring the fragmentation of m/z 392.3→100.1 for HS270, m/z 265.1→232.1 for IS. The method had a chromatographic running time of 2.5 min and linear calibration curves over the concentrations of 0.5-1000 ng/mL. The recovery of the method was 70.8-82.5% and the lower limit of quanti?cation (LLOQ) was 0.5 ng/mL. The intra- and inter-batch precisions were less than 15% for all quality control samples at concentrations of 1.0, 100.0, and 750.0 ng/mL. The validated LC-MS/MS method has successfully applied to a HS270 pharmacokinetic study after oral doses of 25, 50, 100, 200 mg/kg, and i.v. dose of 5 mg/kg to rats.     

Determination of glabridin in human plasma by solid-phase extraction and LC-MS/MS

Glabridin is a major flavonoid included specifically in licorice (Glycyrrhiza glabra L.), and has various physiological activities including antioxidant and anti-inflammatory effects. We have developed and validated an analytical method for determination of glabridin in human plasma by solid-phase extraction (SPE) and LC-MS/MS. Glabridin was extracted from plasma by SPE using a C8 cartridge and analyzed by LC-MS/MS using mefenamic acid as an internal standard (IS). The analyte were separated by a C18 column on LC, and monitored with a fragment ion of m/z 201 formed from a molecular ion of m/z 323 for glabridin and that of m/z 196 from m/z 240 for IS during negative ion mode with tandem MS detection. The lower limit of quantitation (LLOQ) of glabridin was 0.1 ng/mL in plasma, corresponding to 1.25 pg injected on-column. The calibration curves exhibited excellent linearity (r>0.997) between 0.1 and 50 ng/mL. Precision and accuracy were <17 and <+/-7% at LLOQ, and <11 and <+/-5% at other concentrations. Glabridin was recovered >90%, and was stable when kept at 10 degrees C for 72 h, at -20 degrees C until 12 weeks, and after three freeze-thaw cycles. This is the first report on determination of glabridin in body fluids by the selective, sensitive, and reproducible method.     

Rapid and simultaneous determination of nifedipine and dehydronifedipine in human plasma by liquid chromatography-tandem mass spectrometry: Application to a clinical herb-drug interaction study

Nifedipine (NIF), a calcium channel antagonist, is metabolized primarily by cytochrome P450 (CYP3A4) to dehydronifedipine (DNIF). As such, NIF is often used as a probe drug for determining CYP3A4 activity in human studies. A rapid and sensitive liquid chromatography/tandem mass spectrometry (LC/MS/MS) method was developed and validated to simultaneously determine NIF and DNIF in human plasma using nitrendipine as the internal standard (IS). After extraction of the plasma samples by ether-n-hexane (3:1, v/v), NIF, DNIF and the IS were subjected to LC/MS/MS analysis using electro-spray ionization (ESI). Chromatographic separation was performed on a Hypersil BDS C(18) column (50 mm x 2.1 mm, i.d., 3 microm). The method had a chromatographic running time of approximately 2.5 min and linear calibration curves over the concentrations of 0.5-100 ng/mL for NIF and DNIF. The recoveries of the one-step liquid extraction method were 81.3-89.1% for NIF and 71.6-80.4% for DNIF. The lower limit of quantification (LLOQ) of the analytical method was 0.5 ng/mL for both analytes. The intra- and inter-day precision was less than 15% for all quality control samples at concentrations of 2, 10, and 50 ng/mL. The validated LC/MS/MS method has been successfully used to study pharmacokinetic interactions of NIF with the herbal antidepressant St. John's wort in healthy volunteers. These results indicated that the developed LC/MS/MS method was efficient with a significantly shorter running time (2.5 min) for NIF and DNIF compared to those methods previously reported in the literature. The presented LC/MS/MS method had acceptable accuracy, precision and sensitivity and was used in a clinical pharmacokinetic interaction study of NIF with St. John's wort, a known herbal inducer of CYP3A4. St. John's wort was shown to induce NIF metabolism with increased plasma concentrations of DNIF.     

Sensitive and rapid liquid chromatography/tandem mass spectrometric assay for the quantification of chloroquine in dog plasma

A simple, sensitive and rapid liquid chromatography/tandem mass spectrometric (LC-MS/MS) method was developed and validated for quantification of chloroquine, an antimalarial drug, in plasma using its structural analogue, piperazine bis chloroquinoline as internal standard (IS). The method is based on simple protein precipitation with methanol followed by a rapid isocratic elution with 10 mM ammonium acetate buffer/methanol (25/75, v/v, pH 4.6) on Chromolith SpeedROD RP-18e reversed phase chromatographic column and subsequent analysis by mass spectrometry in the multiple reaction monitoring mode (MRM). The precursor to product ion transitions of m/z 320.3-->247.2 and m/z 409.1-->205.2 were used to measure the analyte and the IS, respectively. The assay exhibited a linear dynamic range of 2.0-489.1 ng/mL for chloroquine in dog plasma. The limit of detection (LOD) and lower limit of quantification (LLOQ) were 0.4 and 2.0 ng/mL, respectively in 0.05 mL plasma. Acceptable precision and accuracy were obtained for concentrations over the standard curve range of 2.0-489.1 ng/mL. A run time of 2.0 min for a sample made it possible to achieve a throughput of more than 400 plasma samples analyzed per day. The validated method was successfully used to analyze samples of dog plasma during non-clinical study of chloroquine.     

A capillary liquid chromatographic/tandem mass spectrometric method for the quantification of gamma-aminobutyric acid in human plasma and cerebrospinal fluid

A sensitive and reliable method for the determination of gamma-aminobutyric acid (GABA), a major inhibitory neurotransmitter, in human plasma and cerebrospinal fluid (CSF) has been developed. The method is based on capillary liquid chromatography (LC)/tandem mass spectrometry (MS/MS) using deuterium-labeled GABA (gamma-aminobutyric acid-2,2-D(2), GABA-d(2)) as internal standard. Pre-column derivatization with 7-fluoro-4-nitrobenzoxadiazole (NBD-F) was deployed, allowing both effective in-line pre-concentration and sensitive tandem MS detection of the analyte. An extraction column (10 mm x 0.25 mm, 7 microm, C(18)) was used for preconcentrating and stacking the sample. Separation was carried out on an analytical column (50 mm x 0.25 mm, 5 microm, C(18)). Characteristic precursor-to-product ion transitions, m/z 267--> 249 (for NBD-GABA) and m/z 269--> 251 (for NBD-GABA-d(2)) were monitored for the quantification. A linear calibration curve from 10 to 250 ng/mL GABA with an r(2) value of 0.9994 was obtained. Detection limit was estimated to be 5.00 ng/mL GABA (S/N = 3). Human plasma and CSF samples were analyzed. The concentrations of GABA were found to be 98.6 +/- 33.9 ng/mL (mean +/- S.D., n = 12), and 44.3 +/- 10.0 ng/mL (n = 6) in plasma and CSF, respectively.     

Simultaneous determination of triazolam and its metabolites in human plasma by liquid chromatography-tandem mass spectrometry

A sensitive and selective liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for the determination of triazolam and its metabolites, alpha-hydroxytriazolam (alpha-OHTRZ) and 4-hydroxytriazolam (4-OHTRZ), was developed and validated. Triazolam-D4 was used as the internal standard (IS). This analysis was carried out on a Thermo((R)) C(18) column and the mobile phase was composed of acetonitrile:H(2)O:formic acid (35:65:0.2, v/v/v). Detection was performed on a triple-quadrupole tandem mass spectrometer using positive ion mode electrospray ionization (ESI) and quantification was performed by multiple reaction monitoring (MRM) mode. The MS/MS ion transitions monitored were m/z 343.1-->308.3, 359.0-->308.3, 359.0-->111.2 and 347.0-->312.0 for triazolam, alpha-OHTRZ, 4-OHTRZ and triazolam-D4, respectively. LLOQ of the analytical method was 0.05ng/mL for triazolam and 0.1ng/mL for alpha-OHTRZ and 4-OHTRZ. The within- and between-run precisions were less than 15.26% and accuracy was -8.08% to 13.33%. The method proved to be accurate and specific, and was applied to the pharmacokinetic study of triazolam in healthy Chinese volunteers.     

Development and validation of highly sensitive method for determination of misoprostol free acid in human plasma by liquid chromatography-electrospray ionization tandem mass spectrometry: application to a clinical pharmacokinetic study

A highly sensitive, selective and evaporation free SPE extraction, ESI-LC-MS/MS method has been developed for estimation of misoprostol free acid in human plasma using misoprostol acid-d(5) as an internal standard (IS). The analyte was separated using isocratic mobile phase on reverse phase column and analyzed by MS/MS in the multiple reaction monitoring mode using the respective [M-H] anions, m/z 367-249 for misoprostol acid and m/z 372-249 for the IS. The total run time was 5.0 min and the elution of misoprostol acid and misoprostol acid-d(5) (IS) occurred at 3.6 min. The developed method was validated in human plasma with a lower limit of quantification of 2.5 pg/mL. A linear response function was established for the range of concentrations 2.5-1200 pg/mL (r>0.998) for misoprostol acid in human plasma. The intra and inter-day precision values for misoprostol acid met the acceptance as per FDA guidelines. Misoprostol acid 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 pharmacokinetic study in humans.     

Ultra-performance liquid chromatography-tandem mass spectrometric method for the determination of Artemisinin in rat serum and its application in pharmacokinetics

A rapid and sensitive ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) method was developed and validated to quantify artemisinin in rat serum. The lower limit of quantification (LLOQ) was 4 ng/mL. The calibration curve was linear from 4 ng/mL to 10,000 ng/mL (R=0.998). The assay was based on the selected reaction monitoring (SRM) transitions at m/z 305.4-151.10 for artemisinin and m/z 335.2-163.10 for arteether (internal standard). The artemisinin and internal standard can be separated from endogenous interferences in rat serum. Inter- and intra-day assay variation was less than 15%. The extraction recoveries ranged from 80.0 to 107.3% at the three concentrations (5000, 2000, and 200 ng/mL). This method was successfully applied to pharmacokinetic studies of artemisinin after intravenous and oral administration to rats.     

Development and validation of a LC-ESI-MS/MS method for the determination of swertiamarin in rat plasma and its application in pharmacokinetics

A new LC-ESI-MS/MS assay method has been developed and validated for the quantification of swertiamarin, a representative bioactive substance of Swertia plants, in rat plasma using gentiopicroside, an analog of swertiamarin on chemical structure and chromatographic action, as the internal standard (IS). The swertiamarin and IS were extracted from rat plasma using solid-phase extraction (SPE) as the sample clean-up procedure, and they were chromatographed on a narrow internal diameter column (Agilent ZORBAX ECLIPSE XDB-C(18) 100 mm × 2.1 mm, 1.8 μm) with the mobile phase consisting of methanol and water containing 0.1% acetic acid (25:75, v/v) at a flow rate of 0.2 mL/min. The detection was performed on an Agilent G6410B tandem mass spectrometer by negative ion electrospray ionisation in multiple-reaction monitoring mode while monitoring the transitions of m/z 433 [M+CH(3)COO](-)→179 and m/z 415 [M+CH(3)COO](-)→179 for swertiamarin and IS, respectively. The lower limit of quantification (LLOQ) was 5 ng/mL within a linear range of 5-1000 ng/mL (n=7, r(2)≥0.994), and the limit of detection (LOD) was demonstrated as 1.25 ng/mL (S/N≥3). The method also afforded satisfactory results in terms of sensitivity, specificity, precision (intra- and inter-day), accuracy, recovery, freeze/thaw, long-time stability and dilution integrity. This method was successfully applied to determination of the pharmacokinetic properties of swertiamarin in rats after oral administration at a dose of 20 mg/kg. The following pharmacokinetic parameters were obtained (mean): maximum plasma concentration, 1920.1 ng/mL; time to reach maximum plasma concentration, 0.945 h; elimination half-time, 1.10h; apparent total clearance, 5.638 L/h/kg; and apparent volume of distribution, 9.637 L/kg.     

A liquid chromatography/tandem mass spectrometry method for the simultaneous quantification of valsartan and hydrochlorothiazide in human plasma

A rapid and sensitive liquid chromatography/tandem mass spectrometry (LC/MS/MS) method was developed and validated for simultaneous quantification of valsartan and hydrochlorothiazide in human plasma. After a simple protein precipitation using acetonitrile, the analytes were separated on a Zorbax SB-Aq C18 column using acetonitrile-10mM ammonium acetate (60:40, v/v, pH 4.5) as mobile phase at a flow rate of 1.2 mL/min. Valsartan and hydrochlorothiazide were eluted at 2.08 min and 1.50 min, respectively, ionized using ESI source, and then detected by multiple reaction monitoring (MRM) mode. The precursor to product ion transitions of m/z 434.2-350.2 and m/z 295.9-268.9 were used to quantify valsartan and hydrochlorothiazide, respectively. The method was linear in the concentration range of 4-3600 ng/mL for valsartan and 1-900 ng/mL for hydrochlorothiazide. The method was successfully employed in a pharmacokinetic study after an oral administration of a dispersible tablet containing 80 mg valsartan and 12.5 mg hydrochlorothiazide to each of the 20 healthy volunteers.     

HPLC-MS/MS法测定人血浆中草乌甲素的浓度及生物等效性研究(英文)

本文建立了一种快速、高灵敏的HPLC-MS/MS法用于检测人血浆中的草乌甲素浓度。血浆样品采用沃特斯HLB小柱进行固相萃取,汉邦C18色谱柱(150 mm×4.6 mm,5μm)进行分离,流动相为甲醇∶水(85∶15,v/v),水相含10 mmol/L的醋酸铵和0.1%的甲酸。采用ESI源和多反应监测(MRM)的方式进行检测,草乌甲素及内标的反应离子对分别为644.4/584.4和237.2/194.2,草乌甲素血药浓度在0.010～1.0 ng/mL范围内线性关系良好,最低定量限为0.010 ng/mL可以满足口服0.4 mg草乌甲素后血药浓度的检测,日内日间及质控样品精密度及准确度均在允许范围内。本检测方法被成功的应用在中国健康志愿者生物等效性研究中,20名志愿者口服0.4 mg草乌甲素试验制剂和参比制剂后主要药代动力学参数分别如下:Cmax(0.325±0.110),(0.323±0.115)ng/mL;AUC0-16(1.627±0.489),(1.732±0.556)ng.h/mL;AUC0-∞(1.730±0.498),(1.831±0.562)ng.h/mL;t1/2(4.26±0.95),(3.80±0.90)h;Tmax(1.34±0.54),(1.83±0.99)h。     

Development and validation of a liquid chromatography/tandem mass spectrometry assay for the simultaneous determination of D-amphetamine and diphenhydramine in beagle dog plasma and its application to a pharmacokinetic study

A new drug, quick-acting anti-motion capsule (QAAMC) composed of d-amphetamine sulfate, dimenhydrinate and ginger extraction has been studied for anti-motion-sickness use. We have developed a sensitive, specific liquid chromatography-tandem mass spectrometry (LC-MS/MS) for the quantitative determination of d-amphetamine and diphenhydramine, the main effective components of the QAAMC, using pseudoephedrine as the internal standard. The analytes and internal standard were isolated from 200 microL plasma samples by a simple liquid-liquid extraction (LLE). Reverse-phase HPLC separation was accomplished on a Zorbax SB-C18 column (100 mm x 3.0 mm, 3.5 microm) with a mobile phase composed of methanol-water-formic acid (65:35:0.5, v/v/v) at a flow rate of 0.2 mL/min. The method had a chromatographic total run time of 5 min. A Varian 1200 L electrospray tandem mass spectrometer equipped with an electrospray ionization source was operated in selected reaction monitoring (SRM) mode with the precursor-to-product ion transitions m/z 136.0-->91.0 (D-amphetamine), 256.0-->167.0 (diphenhydramine) and 166.1-->148.0 (IS) used for quantitation. The method was sensitive with a lower limit of quantitation (LLOQ) of 0.5 ng/mL for d-amphetamine and 1 ng/mL for diphenhydramine, with good linearity in the range 0.5-200 ng/mL for D-amphetamine and 1-500 ng/mL for diphenhydramine (r(2)> or =0.9990). All the validation data, such as accuracy, precision, and inter-day repeatability, were within the required limits. The method was successfully applied to pharmacokinetic study of the QAAMC in beagle dogs.     

Determination of salirasib (S-trans,trans-farnesylthiosalicylic acid) in human plasma using liquid chromatography-tandem mass spectrometry

A liquid chromatography/tandem mass spectrometric (LC/MS/MS) assay was developed for the quantitative determination of salirasib (S-trans,trans-farnesylthiosalicylic acid, FTS) in human plasma. Sample pretreatment involved liquid-liquid extraction with methyl t-butyl ether of 0.5-mL aliquots of lithium heparin plasma spiked with the internal standard, S-trans,trans-5-fluoro-farnesylthiosalicylic acid (5-F-FTS). Separation was achieved on Waters X-Terra C(18) (50 mm x 2.1 mm i.d., 3.5 microm) at room temperature using isocratic elution with acetonitrile/10 mM ammonium acetate buffer mobile phase (80:20, v/v) containing 0.1% formic acid at a flow rate of 0.20 mL/min. Detection was performed using electrospray MS/MS by monitoring the ion transitions from m/z 357.2-->153.0 (salirasib) and m/z 375.1-->138.8 (5-F-FTS). Calibration curves were linear in the concentration range of 1-1000 ng/mL. A 5000 ng/mL sample that was diluted 1:10 (v/v) with plasma was accurately quantitated. The values for both within day and between day precision and accuracy were well within the generally accepted criteria for analytical method (<8.0%). This assay was subsequently used for the determination of salirasib concentrations in plasma of cancer patients after oral administration of salirasib at a dose of 400 mg.     

Determination of the quaternary ammonium compound trospium in human plasma by LC–MS/MS: Application to a pharmacokinetic study

A highly sensitive, specific and evaporation free SPE extraction, LC–MS/MS method has been developed for the estimation of trospium in human plasma using trospium-d8 as an internal standard (IS). The analyte was separated using isocratic mobile phase on reverse phase column and analyzed by MS/MS in the multiple reaction monitoring mode using the respective [M⁺] cations, m/z 392–164 for trospium and m/z 400–172 for the IS. The total run time was 3.50 min and the elution of trospium and trospium-d8 (IS) occurred at 2.8 min. The developed method was validated in human plasma with a lower limit of quantification of 0.05 ng/mL. A linear response function was established for the range of concentrations 0.05–10 ng/mL (r > 0.998) for trospium in human plasma. The intra- and inter-day precision values for trospium met the acceptance as per FDA guidelines. Trospium was stable in the battery of stability studies viz., bench-top, auto-sampler, dry extracts and freeze/thaw cycles. The developed assay method was applied to an oral pharmacokinetic study in humans.     

Determination of perospirone by liquid chromatography/electrospray mass spectrometry: application to a pharmacokinetic study in healthy Chinese volunteers

Perospirone is a novel atypical antipsychotic with a unique combination of 5-HT(1A) receptor agonism as well as 5-HT(2A) and D(2) receptor antagonism. A simple rapid and selective LC-MS method utilizing a single quadrupole mass spectrometer was developed and validated for the determination of perospirone hydrochloride in human plasma. N-hexane was used to extract perospirone hydrochloride and amlodipine benzenesulfonate (internal standard (IS)) from an alkaline plasma sample. LC separation was performed on a XTerra MS C(18) column (100mmx2.1mm, i.d. 3.5microm) using methanol -10mM ammonium acetate (84:16, v/v) as a mobile phase. The quantification of target compounds was obtained by using a selected ion monitoring (SIM) at m/z 427.5 [M+H](+) for perospirone hydrochloride, and at m/z 431.4 [M+Na](+) for IS (amlodipine benzenesulfonate). Perospirone and IS eluted as sharp, symmetrical peaks with retention times of 3.11+/-0.01min and 4.15+/-0.2min, respectively. Calibration curves of perospirone hydrochloride in human plasma at concentrations ranging from 0.10 to 21.1ng/mL exhibited excellent linearity (r(2)=0.9997). The mean absolute recovery of the drug from plasma was more than 85%. Intra- and inter-day relative standard deviations were less than 6.43% and 11.9% for perospirone hydrochloride at the range from 0.32 to 10.6ng/mL. Stability characteristics of the drug-containing plasma were thoroughly evaluated to establish appropriate conditions to process, store and prepare for chromatographic analysis without inducing significant chemical degradation. The following pharmacokinetic parameters were elucidated after administering a single dose of 8mg perospirone hydrochloride. The area under the plasma concentration versus time curve from time 0 to 24h (AUC(0-24)) was 15.48+/-4.23microg/Lh; peak plasma concentration (C(max)) was 2.79+/-0.78microg/L; time to C(max) (T(max)) was 1.79+/-0.45h; and elimination half-life (t(1/2)) 6.78+/-1.38h. The described assay method showed acceptable precision, accuracy, linearity, stability, and specificity and can be used for pharmacokinetic studies, therapeutic drug monitoring, and drug abuse screening.     

Quantification of 5-azacytidine in plasma by electrospray tandem mass spectrometry coupled with high-performance liquid chromatography

5-Azacytidine (5AC), a nucleoside analogue and hypomethylating agent, has anticancer properties and has been utilized in the treatment of various malignancies. 5AC is unstable and rapidly hydrolyzed to several by-products, including 5-azacytosine and 5-azauracil. A sensitive, reliable method was developed to quantitate 5AC using LC/MS/MS to perform pharmacokinetic and pharmacodynamic studies on 5AC combination therapy trials. Blood samples were collected in a heparinized tube and immediately processed for storage. To increase the stability of 5AC in plasma, 25 ng/mL tetrahydrouridine was added to the plasma and snap frozen. Plasma samples were extracted using acetonitrile then cleaned up by Oasis MCX ion exchange solid-phase extraction cartridges. 5AC was separated on an YMC Jsphr M80 C(18) column with gradient elution of ammonium acetate (2 mM) with 0.1% formic acid and methanol mobile phase. 5AC elutes at 5.0 +/- 0.2 min with a total run time of 30 min. Identification was through positive-ion mode and multiple reaction monitoring mode at m/z+ 244.9-->113.0 for 5AC and m/z+ 242.0-->126.0 for 5-methyl-2'-deoxycytidine, the internal standard. The lower limit of quantitation of 5AC was 5 ng/mL in human plasma, and linearity was observed from 5 to 500 ng/mL fitted by linear regression with 1/x weight. This method is 50 times more sensitive than previously published assays and successfully allows studies to characterize the pharmacokinetics and pharmacodynamics of 5AC.