Pharmacokinetic study of ardisiacrispin A in rat plasma after intravenous administration by UPLC–MS/MS

In this work, a sensitive and selective UPLC‐MS/MS method for determination of ardisiacrispin A in rat plasma was developed. Cyasterone used as an internal standard (IS) and protein precipitation by acetonitrile–methanol (9:1, v /v) was used to prepare samples. Chromatographic separation was achieved on a UPLC BEH C₁₈ column (2.1 × 100 mm, 1.7 μm) with 0.1% formic acid and acetonitrile as the mobile phase with gradient elution. An electrospray ionization source was applied and operated in positive ion mode; multiple reaction monitoring mode was used for quantification using target fragment ions m /z 1083.5 → 407.1 for ardisiacrispin A and m /z 521.3 → 485.2 for IS. Calibration plots were linear throughout the range 5–2000 ng/mL for ardisiacrispin A in rat plasma. Mean recoveries of ardisiacrispin A in rat plasma ranged from 80.4 to 92.6%. The values of RSD of intra‐ and inter‐day precision were both <11%. The accuracy of the method was between 97.3 and 105.6%. The method was successfully applied to pharmacokinetic study of ardisiacrispin A after intravenous administration in rats.     

An LC–MS/MS assay for the quantitative determination of 2‐pyridyl acetic acid,a major metabolite and key surrogate for betahistine,using low‐volume human K2EDTA plasma

Betahistine is widely used for the treatment of vertigo. Owing to first‐pass metabolism, 2‐pyridyl acetic acid (2PAA, major metabolite of betahistine) was considered as surrogate for quantitation. A specific and sensitive LC–MS/MS method was developed and validated for quantitation of 2PAA using turbo‐ion spray in a positive ion mode. A solid‐phase extraction was employed for the extraction of 2PAA and 2PAA d₆ (IS) from human plasma. Chromatographic separation of analytes was achieved using an ACE CN, 5 μm (50 × 4.6 mm) column with a gradient mobile phase comprising acetonitrile–methanol (90:10% v /v) and 0.7% v/v formic acid in 0.5 mm ammonium trifluoroacetate in purified water (100% v/v). The retention times of 1.15 and 1.17 min for 2PAA and internal standard, respectively, were achieved. Quantitation of 2PAA and internal standard was achieved by monitoring multiple reaction monitoring transition pairs (m /z 138.1 to m /z 92.0 and m /z 142.1 to m /z 96.1, respectively). The developed method was validated for various parameters. The calibration curves of 2PAA showed linearity from 5.0 to 1500 ng/mL, with a lower limit of quantitation of 5.0 ng/mL. The bias and precision for inter‐ and intra‐batch assays were <10%. The developed method was used to support clinical sample analysis.     

Validation of an LC–MS/MS method for simultaneous detection of four HDAC inhibitors – belinostat,panobinostat, rocilinostat and vorinostat in mouse plasma and its application to a mouse pharmacokinetic study

A sensitive and rapid LC–MS/MS method was developed and validated for the simultaneous quantitation of four HDAC inhibitors, namely belinostat (BST), panobinostat (PST), rocilinostat (RST) and vorinostat (VST), in mouse plasma as per regulatory guidelines. The analytes and internal standard were extracted from 50 μL mouse plasma by protein precipitation, followed by chromatographic separation using an Atlantis C₁₈ column with an isocratic mobile phase comprising 0.1% formic acid–acetonitrile (25:75, v /v) at a flow rate of 0.5 mL/min within 2.5 min. Detection and quantitation were done by multiple reaction monitoring on a triple quadrupole mass spectrometer following the transitions: m/z 319 → 93, 350 → 158, 434 → 274 and 265 → 232 for BST, PST, RST and VST, respectively, in the positive ionization mode. The calibration curves were linear from 2.92 to 2921 ng/mL for BST and PST and from 1.01 to 1008 ng/mL for RST and VST with r ² ≥ 0.99 for all of the analytes. The intra‐ and inter‐batch accuracy and precision (CV) across quality controls varied from 85.5 to 112% and from 2.30 to 12.5, respectively, for all of the analytes. Analytes were found to be stable under different stability conditions. The method was applied to an i.v. pharmacokinetic study in mice.     

Highly sensitive LC‐MS/MS method for determination of galantamine in rat plasma: application to pharmacokinetic studies in rats

A rapid and highly sensitive assay method has been developed and validated for the estimation of galantamine (GLM) in rat plasma using liquid chromatography coupled to tandem mass spectrometry with electrospray ionization in the positive‐ion mode. The assay procedure involves a simple liquid–liquid extraction of GLM and phenacetin (internal standard, IS) from rat plasma using acetonitrile. Chromatographic separation was achieved with 0.2% formic acid:acetonitrile (50:50, v/v) at a flow rate of 0.60 mL/min on an Atlantis dC₁₈ column with a total run time 2.5 min. The MS/MS ion transitions monitored were 288.10 → 213.10 for GLM and 180.10 → 110.10 for IS. Method validation was performed as per United States Food and Drug Administration guidelines and the results met the acceptance criteria. The lower limit of quantitation achieved was 0.12 ng/mL and linearity was observed from 0.12 to 525 ng/mL. The intra‐ and inter‐day precision were in the ranges of 4.73–11.7 and 5.83–8.64%, respectively. This novel method has been applied to a pharmacokinetic study in rats. Copyright © 2014 John Wiley & Sons, Ltd.     

Determination of a natural DNMT1 inhibitor,peperomin E,in rat plasma by UFLC‐MS/MS and method application in a pharmacokinetic study

Peperomin E (PepE), a naturally occurring secolignan isolated from Peperomia dindygulensis , has drawn much attention recently owing to its anticancer and DNA methyltransferase 1 (DNMT1) inhibitory activity. Here, a simple and sensitive ultra‐fast liquid chromatography–tandem mass spectrometry method was developed and validated for the determination of PepE in rat plasma for the first time. Samples were prepared by simple protein precipitation. Separation was performed on an XBridge™ C₁₈ column using a mobile phase of acetonitrile and 0.1% (v /v) aqueous formic acid. PepE and the internal standard arctigenin were detected in a positive‐ion mode using multiple reaction monitoring of the transitions at m/z 413.2 → 261.0 and 373.2 → 137.2, respectively. The calibration curve for PepE was linear over the range of concentrations of 1.46–6000 ng/mL, with a lower limit of quantitation of 1.46 ng/mL. Both intra‐ and interday precisions were within 11.05%, and the accuracy ranged from −11.5 to 5.51%. The extraction recovery and matrix effect were within acceptable limits. Stability tests showed that PepE remained stable throughout the analytical procedure. The validated method was then used to analyze the pharmacokinetics of PepE administered to rats orally (12.5 and 25 mg/kg) or intravenously (6.25 and 12.5 mg/kg).     

Validated LC–MS/MS method for the quantification of sciadopitysin in rat plasma and its application to pharmacokinetic and bioavailability studies in vivo

A sensitive and rapid LC–MS/MS method was developed and validated for quantitation of sciadopitysin in rat plasma using amentoflavone as an internal standard. Sample processing was accomplished after deproteinization with 150 μL aliquot of acetonitrile. Chromatographic separation was achieved using an Agela C₁₈ column with an isocratic mobile phase comprising 2 mm ammonium acetate–acetonitrile (35:65, v/v) at a flow rate of 0.4 mL/min. Detection was performed by selection reaction monitoring on a triple‐quadrupole mass spectrometer following the transitions m/z 579 → 547 and 537 → 375 for sciadopitysin and internal standard, respectively, in the negative ionization mode. The calibration curve was linear from 2.90 to 1160 ng/mL for sciadopitysin. Intra‐ and inter‐day precisions were in the ranges 4.1–11.4 and 5.7–9.1% for sciadopitysin. Sciadopitysin was stable under different stability conditions. The validated assay was applied to pharmacokinetic and bioavailability studies in rats.     

Quantification of β‐eudesmol in rat plasma using LC–MS/MS and its application to a pharmacokinetic study

A sensitive and specific LC–MS/MS assay for determination of β ‐eudesmol in rat plasma was developed and validated. After liquid–liquid extraction with ethyl ether , the analyte and IS were separated on a Capcell Pak C₁₈ column (50 × 2.0 mm, 5 μm) by isocratic elution with acetonitrile—water–formic acid (77.5:22.5:0.1, v /v/v) as the mobile phase at a flow rate of 0.4 mL/min. An ESI source was applied and operated in positive ion mode; a selected reaction monitoring scan was used for quantification by monitoring the precursor–product ion transitions of m/z 245.1 → 163.1 for β ‐eudesmol and m/z 273.4 → 81.2 for IS. Good linearity was observed in the concentration range of 3–900 ng/mL for β ‐eudesmol in rat plasma. Intra‐ and inter‐day precision and accuracy were both within ±14.3%. This method was applied for pharmacokinetic studies after intravenous bolus of 2.0 mg/kg or intragastric administration of 50 mg/kg β ‐eudesmol in rats.     

Assessment of pharmacokinetics,bioavailability and protein binding of anacetrapib in rats by a simple high‐performance liquid chromatography–tandem mass spectrometry method

Anacetrapib is a potent and selective CETP inhibitor and is undergoing phase III clinical trials for the treatment of dyslipidemia. A simple and sensitive high‐performance liquid chromatography–tandem mass spectrometry (HPLC–MS/MS) method for the quantification of anacetrapib in rat plasma was developed and validated using an easily purchasable compound, chlorpropamide, as an internal standard (IS). A minimal volume of rat plasma sample (20 μL) was prepared by a single‐step deproteinization procedure with 80 μL of acetonitrile. Chromatographic separation was performed using Kinetex C₁₈ column with a gradient mobile phase consisting of water and acetonitrile containing 0.1% formic acid at a flow rate of 0.3 mL/min. Mass spectrometric detection was performed using selected reaction monitoring modes at the mass/charge transitions m/z 638 → 283 for anacetrapib and m/z 277 → 175 for IS. The assay was validated to demonstrate the selectivity, linearity, precision, accuracy, recovery, matrix effect and stability. The lower limit of quantification was 5 ng/mL. This LC‐MS/MS assay was successfully applied in the rat plasma protein binding and pharmacokinetic studies of anacetrapib. The fraction of unbound anacetrapib was determined to be low (ranging from 5.66 to 12.3%), and the absolute oral bioavailability of anacetrapib was 32.7%.     

Development of a targeted method for quantification of gypenoside XLIX in rat plasma,using SPE and LC–MS/MS

A sensitive, selective and rapid liquid chromatography–tandem mass spectrometry (LC–MS/MS) method was developed for the quantification of gypenoside XLIX, a naturally occurring gypenoside of Gynostemma pentaphyllum in rat plasma and then validated according to the US Food and Drug Administration's Guidance for Industry: Bioanalytical Method Validation . Plasma samples were prepared by a simple solid‐phase extraction. Separation was performed on a Waters XBridgeTM BEH C₁₈ chromatography column (4.6 × 50 mm, 2.5 μm) using a mobile phase of acetonitrile and water (62.5:37.5, v /v). Gypenoside XLIX and the internal standard gypenoside A were detected in the negative ion mode using selection reaction monitoring of the transitions at m/z 1045.6 → 913.5 and 897.5 → 765.4, respectively. The calibration curve was linear (R ² > 0.990) over a concentration range of 10–7500 ng/mL with the lower quantification limit of 10 ng/mL. Intra‐ and inter‐day precision was within 8.6% and accuracy was ≤10.2%. Stability results proved that gypenoside XLIX and the IS remained stable throughout the analytical procedure. The validated LC–MS/MS method was then applied to analyze the pharmacokinetics of gypenoside XLIX after intravenous administration to rats (1.0, 2.0 and 4.0 mg/kg).     

A simple and selective UHPLC–MS/MS method for quantification of plantagoguanidinic acid in rat plasma and its application to a pharmacokinetic study

A simple, sensitive and specific UHPLC–MS/MS method for quantification of plantagoguanidinic acid (PGA) in rat plasma was applied to investigate the pharmacokinetic behavior in vivo , using protopine as internal standard. The chromatography was separated on a Phenomenex® Luna‐C₁₈ column (2.1 × 150 mm, 3.0 μm) within 7.0 min using a mobile phase consisting of acetonitrile–0.1% formic acid solution under gradient elution at a flow rate of 0.4 mL/min. Prepared samples were monitored by multiple reaction monitoring mode, with the target fragmentions m/z 226.2 → 84.2 for PGA and m/z 354.2 → 188.9 for IS in positive electrospray ionization. The calibration curve of PGA was linear throughout the range 1–1000 ng/mL (r = 0.9962). The lower limit of quantitation in plasma for PGA was 0.1 ng/mL, and the recovery was >88.6%. Intra‐ and interday accuracy ranged from −8.6 to 4.9%. Furthermore, this validated method was successfully used for a pre‐clinical pharmacokinetic study of PGA at a single dose of 20 and 5 mg/kg in rats via oral and intravenous administration. The study showed that PGA was absorpted rapidly and eliminated gradually with a greater absolute oral bioavailability of 70.1% in rats.     

A rapid and sensitive determination of hypoxic radiosensitizer agent nimorazole in rat plasma by LC‐MS/MS and its application to a pharmacokinetic study

A highly sensitive, accurate and robust LC‐MS/MS method was developed and validated for determination of nimorazole (NMZ) in rat plasma using metronidazole (MNZ) as internal standard (IS). The analyte and IS were extracted from plasma by precipitating protein with acetonitrile and were chromatographed using an Agilent Poroshell 120, EC‐C₁₈ column. The mobile phase was composed of a mixture of acetonitrile and 0.1 % formic acid (85:15 v/v). The total run time was 1.5 min and injection volume was 5 μL. Multiple reaction monitoring mode using the transitions of m/z 227.1 → m/z 114.0 for MNZ and m/z 172.10 → m/z 128.1 for IS were monitored on a triple quadrupole mass spectrometer, operating in positive ion mode. The calibration curve was linear in the range of 0.25–200 ng/mL (r² > 0.9996) and the lower limit of quantification was 0.25 ng/mL in the rat plasma samples. Recoveries of NMZ ranged between 88.05 and 95.25%. The precision (intra‐day and inter‐day) and accuracy of the quality control samples were 1.25–8.20% and ?2.50–3.10, respectively. The analyte and IS were found to be stable during all sample storage and analysis procedures. The LC‐MS/MS method described here was validated and successfully applied to pharmacokinetic study in rats. Copyright © 2015 John Wiley & Sons, Ltd.     

Development and validation of an LC‐MS/MS method for the determination of a novel thienoquinolin urea transporter inhibitor PU‐48 in rat plasma and its application to a pharmacokinetic study

A specific, sensitive and stable high‐performance liquid chromatographic–tandem mass spectrometry (LC‐MS/MS) method was developed and validated for the quantitative determination of methyl 3‐amino‐6‐methoxythieno [2,3‐b]quinoline‐2‐carboxylate (PU‐48), a novel diuretic thienoquinolin urea transporter inhibitor in rat plasma. In this method, the chromatographic separation of PU‐48 was achieved with a reversed‐phase C₁₈ column (100 × 2.1 mm, 3 μm) at 35°C. The mobile phase consisted of acetonitrile and water with 0.05% formic acid added with a gradient elution at flow rate of 0.3 mL/min. Samples were detected with the triple‐quadrupole tandem mass spectrometer with multiple reaction monitoring mode via electrospray ionization source in positive mode. The retention time were 6.2 min for PU‐48 and 7.2 min for megestrol acetate (internal standard, IS). The monitored ion transitions were mass‐to‐charge ratio (m/z) 289.1 → 229.2 for PU‐48 and m/z 385.3 → 267.1 for the internal standard. The calibration curve for PU‐48 was linear over the concentration range of 0.1–1000 ng/mL (r² > 0.99), and the lower limit of quantitation was 0.1 ng/mL. The precision, accuracy and stability of the method were validated adequately. The developed and validated method was successfully applied to the pharmacokinetic study of PU‐48 in rats.     

Quantitative determination of euphol in rat plasma by LC‐MS/MS and its application to a pharmacokinetic study

Euphol is a potential pharmacologically active ingredient isolated from Euphorbia kansui. A simple, rapid, and sensitive method to determine euphol in rat plasma was developed based on liquid chromatography‐tandem mass spectrometry (LC‐MS/MS) for the first time. The analyte and internal standard (IS), oleanic acid, were extracted from plasma with methanol and chromatographied on a C₁₈ short column eluted with a mobile phase of methanol–water–formic acid (95:5:0.1, v/v/v). Detection was performed by positive ion atmospheric pressure chemical ionization in selective reaction monitoring mode. This method monitored the transitions m/z 409.0 → 109.2 and m/z 439.4 → 203.2 for euphol and IS, respectively. The assay was linear over the concentration range 27–9000 ng/mL, with a limit of quantitation of 27 ng/mL. The accuracy was between –7.04 and 4.11%, and the precision was <10.83%. This LC‐MS/MS method was successfully applied to investigate the pharmacokinetic study of euphol in rats after intravenous (6 mg/kg) and oral (48 mg/kg) administration. Results showed that the absolute bioavailability of euphol was approximately 46.01%. Copyright © 2014 John Wiley & Sons, Ltd.     

Simultaneous determination of glaucocalyxin A and glaucocalyxin B in rat plasma by LC‐MS/MS and its application to a pharmacokinetic study after oral administration of Rabdosia japonica extract

A specific and sensitive liquid chromatography tandem mass spectrometry (LC‐MS/MS) method was developed and validated for the analysis of glaucocalyxin A and glaucocalyxin B in rat plasma using praeruptorin A as an internal standard. Separation was performed on a Hypurity C₁₈ column (2.1 × 50 mm, 5 μm) with isocratic elution using 0.2% formic acid in water–acetonitrile (20:80, v/v). Mass spectrometric detection was conducted using selected reaction monitoring via an electrospray ionization source. Both analytes exhibited good linearity within their concentration ranges (r² > 0.9932). The lower limit of quantitation of glaucocalyxin A and glaucocalyxin B was 1.10 ng/mL. Intra‐ and inter‐day precision exhibited an RSD within 14.5%, and the accuracy (RE) ranged from –12.1 to 15.0% at the lower limit of quantitation and three quality control levels. The developed assay was successfully applied to a pharmacokinetic study of glaucocalyxin A and glaucocalyxin B in rats after oral administration of Rabdosia japonica extract.     

A validated LC‐MS/MS assay for simultaneous quantification of methotrexate and tofacitinib in rat plasma: application to a pharmacokinetic study

A highly sensitive, specific and rapid LC‐ESI‐MS/MS method has been developed and validated for simultaneous quantification of methotrexate (MTX) and tofacitinib (TFB) in rat plasma (50 μL) using phenacetin as an internal standard (IS), as per the US Food and Drug Administration guidelines. After a solid‐phase extraction procedure, the separation of the analytes and IS was performed on a Chromolith RP_18e column using an isocratic mobile phase of 5 m m ammonium acetate (pH 5.0) and acetonitrile at a ratio of 25:75 (v/v) using flow‐gradient with a total run time of 3.5 min. The detection was performed in multiple reaction monitoring mode, using the transitions of m/z 455.2 → 308.3, m/z 313.2 → 149.2 and m/z 180.3 → 110.2 for MTX, TFB and IS, respectively. The calibration curves were linear over the range of 0.49–91.0 and 0.40–74.4 ng/mL for MTX and TFB, respectively. The intra‐ and interday accuracy and precision values for MTX and TFB were <15% at low quality control (QC), medium QC and high QC and <20% at lower limit of quantification. The validated assay was applied to derive the pharmacokinetic parameters for MTX and TFB post‐dosing of MTX and TFB orally and intravenously to rats. Copyright © 2014 John Wiley & Sons, Ltd.     

A highly sensitive LC‐MS/MS method for the determination of S‐citalopram in rat plasma: application to a pharmacokinetic study in rats

A highly sensitive, rapid assay method has been developed and validated for the estimation of S‐citalopram (S‐CPM) in rat plasma with liquid chromatography coupled to tandem mass spectrometry with electrospray ionization in the positive‐ion mode. The assay procedure involves a simple liquid–liquid extraction of S‐CPM and phenacetin (internal standard, IS) from rat plasma with t‐butyl methyl ether. Chromatographic separation was operated with 0.2% formic acid:acetonitrile (20:80, v/v) at a flow rate of 0.50 mL/min on a Symmetry Shield RP₁₈ column with a total run time of 3.0 min. The MS/MS ion transitions monitored were 325.26 → 109.10 for S‐CPM and 180.10 → 110.10 for IS. Method validation and pre‐clinical sample analysis were performed as per FDA guidelines and the results met the acceptance criteria. The lower limit of quantitation achieved was 0.5 ng/mL and the linearity was observed from 0.5 to 5000 ng/mL. The intra‐ and inter‐day precisions were in the range of 1.14–5.56 and 0.25–12.3%, respectively. This novel method has been applied to a pharmacokinetic study and to estimate brain‐to‐plasma ratio of S‐CPM in rats. Copyright © 2010 John Wiley & Sons, Ltd.     

Development and validation of HPLC‐MS/MS method for the determination of lixivaptan in mouse plasma and its application in a pharmacokinetic study

The aim of the present study was to develop a simple, sensitive and accurate liquid chromatography–electrospray ionization tandem mass spectrometry (ESI‐MS/MS) method for the determination of lixivaptan (LIX) in mouse plasma using vildagliptin as the internal standard (IS). A precipitation procedure was used for the extraction of LIX and vildagliptin from mouse plasma. Chromatographic separation of LIX was achieved using a C₁₈ analytical column (50 × 2.1 mm, 1.8 μm) at 25°C. The mobile phase comprised acetonitrile and ammonium formate (10 mm , pH 3.1; 40:60, v /v) pumped at a flow rate of 0.3 mL min⁻¹. A tandem mass spectrometer with an electrospray ionization source was used to perform the assay. Quantification of LIX at m/z 290 → 137 and IS at 154 → 97 was attained through multiple reaction monitoring. The investigated method was authenticated following the bio‐analytical method of validation guidelines of the US Food and Drug Administration. The developed method showed a good linearity over the concentration range from 5 to 500 ng mL⁻¹, and the calibration curve was linear (r = 0.9998). The mean recovery of LIX from mouse plasma was 99.2 ± 0.68%. All validation parameters for LIX were within the levels required for acceptance. The proposed method was effectively used for a pharmacokinetic study of LIX in mouse plasma.     

A chiral LC‐MS/MS method for the enantioselective determination of R‐(+)‐ and S‐(–)‐pantoprazole in human plasma and its application to a pharmacokinetic study of S‐(–)‐pantoprazole sodium injection

Pantoprazole, a proton pump inhibitor, is clinically used for the treatment of peptic diseases. An enantioselective LC‐MS/MS method was developed and validated for the simultaneous determination of pantoprazole enantiomers in human plasma. Pantoprazole enantiomers and the internal standard were extracted from plasma using acetonitrile. Chiral separation was carried on a Chiralpak IE column using the mobile phase consisted of 10 mm ammonium acetate solution containing 0.1% acetic acid–acetonitrile (28 : 72, v /v). MS analysis was performed on an API 4000 mass spectrometer. Multiple reactions monitoring transitions of m /z 384.1→200.1 and 390.1→206.0 were used to quantify pantoprazole enantiomers and internal standard, respectively. For each enantiomer, no apparent matrix effect was found, the calibration curve was linear over 5.00–10,000 ng/mL, the intra‐ and inter‐day precisions were below 10.0%, and the accuracy was within the range of –5.6% to 0.6%. This method was applied to the stereoselective pharmacokinetic studies in human after intravenous administration of S ‐(–)‐pantoprazole sodium injections. No chiral inversion was observed during sample storage, preparation procedure and analysis. While R ‐(+)‐pantoprazole was detected in human plasma with a slightly high concentration, which implied that S ‐(–)‐pantoprazole may convert to R ‐(+)‐pantoprazole in some subjects.     

Validated LC-MS/MS assay for the quantitative determination of nalbuphine in human plasma and its application to a pharmacokinetic study

A solid‐phase extraction–liquid chromatographic–tandem mass spectrometry method for the determination of nalbuphine concentrations in human plasma has been developed. Samples (1 mL) were extracted using a Strata™‐X solid phase extraction cartridges. Chromatographic separation of nalbuphine and naloxone (internal standard) was achieved on a Phenomenex Kinetex PFP (2.6 μm, 100 A, 100 × 2.1 mm) column using a mobile phase consisting of 0.1% formic acid, 15 mM ammonium acetate in deionized water and acetonitrile (60:40, v/v). The flow rate was 0.3 mL/min and the total run time was 2 min. Detection of the analytes was achieved using positive ion electrospray ionization via multiple reactions monitoring mode. The mass transitions were m/z 358 → 340 for nalbuphine and m/z 328 → 310 for naloxone. The assay was linear over the concentration range 0.50–500.00 ng/mL, with correlation coefficients ≥0.995. The lower limit of quantitation was set at 0.5 ng/mL plasma based on an average signal‐to‐noise ratio of 44.79. The intra‐ and inter‐day precision was less than 8.07% in terms of relative standard deviation and accuracy ranged from 94.97 to 106.29% at all quality control levels. The method was applied successfully to determine nalbuphine concentrations in human plasma samples obtained from subjects receiving intravenous administration of nalbuphine. The method is rapid, sensitive, selective and directly applicable to human pharmacokinetic studies involving nalbuphine. Copyright © 2011 John Wiley & Sons, Ltd.     

Simultaneous determination of the bioactive components in rat plasma by UPLC‐MS/MS and application in pharmacokinetic studies after oral administration of radix Scutellariae extract

A highly sensitive and rapid ultraperformance liquid chromatography–tandem mass spectrometry (UPLC‐MS/MS) method has been developed and validated for simultaneous quantification of the four main bioactive compounds, i.e. baicalin, baicalein, wogonoside and wogonin, in rat plasma after oral administration of Radix Scutellariae extract. Clarithromycin was used as an internal standard (IS). Plasma samples were processed by protein precipitation with methanol. The separation was performed on an Acquity BEH C₁₈ column (100 × 2.1 mm, 1.7 μm) at a flow rate of 0.4 mL/min, using 0.1% formic acid–acetonitrile as mobile phase. The MS/MS ion transit ions monitored were 447.5 → 270.1 for baicalin, 270.1 → 168.1 for baicalein, 461.2 → 284.0 for wogonoside, 284.2 → 168.1 for wogonin and 748.5 → 158.1 for IS. Method validation was performed according to US Food and Drug Administration guidelines and the results met the acceptance criteria. The lower limit of quantification (LLOQ) achieved was 1.13 ng/mL for baicalin, 1.23 ng/mL for baicalein, 0.82 ng/mL for wogonoside and 0.36 ng/mL for wogonin. The calibration curves obtained were linear (r > 0.99) over the concentration range ~ 1–1000 ng/mL. The intra‐ and inter‐day precision was <15% and the accuracy was within ±14.7%. After validation, this method was successfully applied to a pharmacokinetic study of Radix Scutellariae extract.