高效液相色谱-串联质谱法测定比格犬血浆中的莫诺苷浓度及其药代动力学

运用高效液相色谱-串联质谱联用技术,建立了简单、快速、灵敏的比格犬灌胃莫诺苷后血药浓度的检测方法。血浆样品采用蛋白质沉淀法处理,以芍药苷作为内标,色谱柱为Inertsil ODS-SP色谱柱(50 mm×2.1 mm,5μm),流动相为水(含1 mmol/L甲酸钠)-乙腈,梯度洗脱,流速0.4 mL/min。采用电喷雾离子源(ESI),正离子多反应监测(MRM)模式。绘制血药浓度-时间曲线,并采用DAS 2.0软件计算药代动力学参数。方法学实验结果表明内源性杂质不干扰莫诺苷和内标的测定,线性范围为2~5 000μg/L(r=0.996 6),定量限为2μg/L。方法精密度、准确度、回收率和基质效应均符合生物样品测定的要求,适合比格犬血浆中莫诺苷浓度的测定,可以应用该方法进行莫诺苷的药代动力学研究。比格犬灌胃莫诺苷3个剂量(5、15、45 mg/kg)后的血药浓度-时间曲线下面积(AUC(0-∞))分别为(1 631.20±238.50)、(3 984.05±750.38)、(10 397.64±3 156.34)μg/L·h,与给药剂量之间呈现良好的线性关系。     

Simultaneous determination of piperaquine and its N‐oxidated metabolite in rat plasma using LC‐MS/MS

A sensitive and efficient liquid chromatography tandem mass spectrometry method was developed and validated for the simultaneous determination of piperaquine (PQ) and its N ‐oxidated metabolite (PQ‐M) in plasma. A simple protein precipitation procedure was used for sample preparation. Adequate chromatographic retention was achieved on a C₁₈ column under gradient elution with acetonitrile and 2 mm aqueous ammonium acetate containing 0.15% formic acid and 0.05% trifluoroacetic acid. A triple‐quadrupole mass spectrometer equipped with an electrospray source was set up in the positive ion mode and multiple reaction monitoring mode. The method was linear in the range of 2.0–400.0 ng/mL for PQ and 1.0–50.0 ng/mL for PQ‐M with suitable accuracy, precision and extraction recovery. The lower limits of detection (LLOD) were established at 0.4 and 0.2 ng/mL for PQ and PQ‐M, respectively, using 40 μL of plasma sample. The matrix effect was negligible under the current conditions. No effect was found for co‐administrated artemisinin drugs or hemolysis on the quantification of PQ and PQ‐M. Stability testing showed that two analytes remained stable under all relevant analytical conditions. The validated method was successfully applied to a pharmacokinetic study performed in rats after a single oral administration of PQ (60 mg/kg).     

Concurrent determination of olanzapine,risperidone and 9‐hydroxyrisperidone in human plasma by ultra performance liquid chromatography with diode array detection method: application to pharmacokinetic study

A simple and sensitive ultra‐performance liquid chromatography (UPLC) method has been developed and validated for simultaneous estimation of olanzapine (OLZ), risperidone (RIS) and 9‐hydroxyrisperidone (9‐OHRIS) in human plasma in vitro. The sample preparation was performed by simple liquid–liquid extraction technique. The analytes were chromatographed on a Waters Acquity H class UPLC system using isocratic mobile phase conditions at a flow rate of 0.3 mL/min and Acquity UPLC BEH shield RP₁₈ column maintained at 40°C. Quantification was performed on a photodiode array detector set at 277 nm and clozapine was used as internal standard (IS). OLZ, RIS, 9‐OHRIS and IS retention times were found to be 0.9, 1.4, .1.8 and 3.1 min, respectively, and the total run time was 4 min. The method was validated for selectivity, specificity, recovery, linearity, accuracy, precision and sample stability. The calibration curve was linear over the concentration range 1–100 ng/mL for OLZ, RIS and 9‐OHRIS. Intra‐ and inter‐day precisions for OLZ, RIS and 9‐OHRIS were found to be good with the coefficient of variation <6.96%, and the accuracy ranging from 97.55 to 105.41%, in human plasma. The validated UPLC method was successfully applied to the pharmacokinetic study of RIS and 9‐OHRIS in human plasma. Copyright © 2015 John Wiley & Sons, Ltd.     

A UPLC–MS/MS approach for simultaneous determination of eight flavonoids in rat plasma,and its application to pharmacokinetic studies of Fu‐Zhu‐Jiang‐Tang tablet in rats

The purpose of this study is to establish and validate a UPLC–MS/MS approach to determine eight flavonoids in biological samples and apply the method to pharmacokinetic study of Fu‐Zhu‐Jiang‐Tang tablet. A Waters BEH C₁₈ UPLC column was employed with methanol/0.1% formic acid–water as mobile phases. The mass analysis was carried out in a triple quadrupole mass spectrometer using multiple reaction monitoring with negative scan mode. A one‐step protein precipitation by methanol was used to extract the analytes from blood. Eight major flavonoids were selected as markers. Our results showed that calibration curves for 3′‐hydroxypuerarin, mirificin, puerarin, 3′‐methoxypuerarin, daidzin, rutin, astragalin and daidzein displayed good linear regression (r ² > 0.9986). The intra‐day and inter‐day precisions (RSD) of the eight flavonoids at high, medium and low levels were <8.03% and the bias of the accuracies ranged from −5.20 to 6.75%.The extraction recoveries of the eight flavonoids were from 91.4 to 100.5% and the matrix effects ranged from 89.8 to 103.8%. The validated approach was successfully applied to a pharmacokinetic study in Sprague–Dawley rats after oral administration of FZJT tablet. Double peaks were emerged in curves of mean plasma concentration for 3′‐methoxypuerarin, which was reported for the first time.     

Simultaneous determination of cucurbitacin IIa and cucurbitacin IIb of Hemsleya amabilis by HPLC–MS/MS and their pharmacokinetic study in normal and indomethacin‐induced rats

A selective and sensitive HPLC–MS/MS method was developed for the simultaneous determination of cucurbitacin IIa (cuIIa) and cucurbitacin IIb (cuIIb), the major bioactive cucurbitacins of Hemsleya amabilis, in rat plasma using euphadienol as internal standard (IS). After liquid–liquid extraction with dichloromethane, separation was achieved on a Syncronis HPLC C₁₈ column (150 mm × 4.6 mm, 5 μm) using an isocratic mobile phase system consisting of acetonitrile–water (85:15, v/v) at a flow rate of 0.6 mL/min with a split ratio of 1:2. Detection was performed on a TSQ Quantum Ultra mass spectrometer equipped with an positive‐ion electrospray ionization source. The lower limits of quantification (LLOQs) were 0.25 and 0.15 ng/mL for cuIIa and cuIIb, respectively. The intra‐ and inter‐day precision was <11.5% for the LLOQs and each quality control level of the analytes, and accuracy was between ?9.1 and 7.6%. The extraction recoveries of the analytes and IS from rat plasma were all >87.1%. The method was fully validated and applied to compare the pharmacokinetic profiles of the two cucurbitacins in rat plasma after oral administration of H. amabilis extract between normal and indomethacin‐induced rats. Copyright © 2016 John Wiley & Sons, Ltd.     

Application of a liquid chromatography–tandem mass spectrometry method to the pharmacokinetics,tissue distribution and excretion in the study of anemoside B4, a novel antiviral agent candidate,in rats

A simple, sensitive and reliable LC–MS/MS method was developed and validated for the quantification of anemoside B4, a potential antiviral constituent isolated from Pulsatilla chinensis in rat plasma, tissue, bile, urine and feces. All biological samples were prepared by protein precipitation method, and ginsenoside‐Rg1 was chosen as the internal standard (IS). The analyte and IS were separated using a C₁₈ column (2.1 × 50 mm, 1.8 μm) and a mobile phase consisting of 0.1% formic acid in water (v /v) and acetonitrile running at a flow rate of 0.2 mL/min for 5 min. The multiple reaction monitoring transitions were monitored at m /z 1219.5–749.5 for anemoside B4 and 845.4–637.4 for ginsenoside‐Rg1 in electrospray ionization negative mode. The calibration curve was linear in the range of 10–2000 ng/mL for all biological matrices with a lower limit of quantification of 10 ng/mL. The validated method was successfully applied to a pharmacokinetics, tissue distribution and excretion study. These preclinical data will be beneficial for further development of anemoside B4 in future studies.     

Analysis of the pharmacokinetics and metabolism of aloe‐emodin following intravenous and oral administrations in rats

Aloe‐emodin, a natural polyphenolic anthraquinone, has shown various beneficial bioactivities in vitro. The aim of this study was to investigate the pharmacokinetics and metabolism of aloe‐emodin. Aloe‐emodin was intravenously and orally administered to rats. The concentrations of aloe‐emodin and rhein, a metabolite of aloe‐emodin, were determined by HPLC method prior to and after hydrolysis with β‐glucuronidase and sulfatase/β‐glucuronidase. The results showed that the systemic exposures of aloe‐emodin and its metabolites were ranked as aloe‐emodin glucuronides (G) > rhein sulfates (S) > aloe‐emodin > rhein and rhein G when aloe‐emodin was given intravenously. In contrast, when aloe‐emodin was administered orally, the parent form of aloe‐emodin was not absorbed per se, and the systemic exposures of its metabolites were ranked as aloe‐emodin G > rhein G > rhein. In conclusion, the metabolites of aloe‐emodin are more important than the parent form for the bioactivities in vivo. Copyright © 2016 John Wiley & Sons, Ltd.     

Validation and clinical application of an LC‐ESI‐MS/MS method for simultaneous determination of tolmetin and MED5, the metabolites of amtolmetin guacil in human plasma

A highly sensitive, rapid assay method has been developed and validated for the simultaneous estimation of tolmetin (TMT) and MED5 in human plasma with liquid chromatography coupled to tandem mass spectrometry with electrospray ionization in the positive‐ion mode. A simple solid‐phase extraction process was used to extract TMT and MED5 along with mycophenolic acid (internal standard, IS) from human plasma. Chromatographic separation was achieved with 0.2% formic acid–acetonitrile (25:75, v/v) at a flow rate of 0.50 mL/min on an X‐Terra RP₁₈ column with a total run time of 2.5 min. The MS/MS ion transitions monitored were 258.1 → 119.0 for TMT, 315.1 → 119.0 for MED5 and 321.2 → 207.0 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 20 ng/mL and the linearity was observed from 20 to 2000 ng/mL, for both the anlaytes. The intra‐day and inter‐day precisions were in the range 3.27–4.50 and 5.32–8.18%, respectively for TMT and 4.27–5.68 and 5.32–8.85%, respectively for MED5. This novel method has been applied to a clinical pharmacokinetic study. Copyright © 2010 John Wiley & Sons, Ltd.     

Determination of piracetam in rat plasma by LC–MS/MS and its application to pharmacokinetics

A sensitive and selective liquid chromatography–tandem mass spectrometry method for the determination of piracetam in rat plasma was developed and validated over the concentration range of 0.1–20 µg/mL. After addition of oxiracetam as internal standard, a simplified protein precipitation with trichloroacetic acid (5%) was employed for the sample preparation. Chromatographic separation was performed by a Zorbax SB‐Aq column (150 × 2.1 mm, 3.5 µm). The mobile phase was acetonitrile–1% formic acid in water (10:90 v/v) delivered at a flow rate of 0.3 mL/min. The MS data acquisition was accomplished in multiple reaction monitoring mode with a positive electrospray ionization interface. The lower limit of quantification was 0.1 µg/mL. For inter‐day and intra‐day tests, the precision (RSD) for the entire validation was less than 9%, and the accuracy was within the 94.6–103.2% range. The developed method was successfully applied to pharmacokinetic studies of piracetam in rats following single oral administration dose of 50 mg/kg. Copyright © 2010 John Wiley & Sons, Ltd.     

Simultaneous determination of eight bioactive components of Qishen Yiqi dripping pills in rat plasma using UFLC–MS/MS and its application to a pharmacokinetic study

In this study, a rapid and reliable ultra‐fast liquid chromatography–tandem mass spectrometry method was developed and validated for the simultaneous determination of eight active ingredients, including astragaloside IV, ononin, tanshinol, protocatechualdehyde, protocatechuic acid, salvianolic acid D, rosmarinic acid and ginsenoside Rg₁, in rat plasma. The plasma samples were pretreated by protein precipitation with acetonitrile. Chromatographic separation was performed on a Waters Acquity UPLC® BEH C₁₈ column (1.7 μm particles, 2.1 × 100 mm). The mobile phase consisted of 0.1% aqueous formic acid (A)–acetonitrile with 0.1% formic acid (B) at a flow rate of 0.4 mL/min. Quantification was performed on a triple quadruple tandem mass spectrometry with electrospray ionization by multiple reaction monitoring both in the negative and in the positive ion mode. The lower limit of quantification of tanshinol was 2.0 ng/mL and the others were 5.0 ng/mL. The extraction recoveries, matrix effects, intra‐ and inter‐day precision and accuracy of eight tested components were all within acceptable limits. The validated method was successfully applied to the pharmacokinetic study of the eight active constituents after intragastric administration of three doses (1.0, 3.0, 6.0 g/kg body weight) of Qishen Yiqi Dripping Pills to rats.