液相色谱-串联质谱法测定犬血浆中布地奈德

建立液相色谱-串联质谱法测定犬血浆中布地奈德。血浆样品碱化后，经乙酸乙酯液-液萃取，以乙腈-5 mmol·L^-1醋酸铵(60∶40，v/v)为流动相，Capcell Pak C₁₈ MG柱分离；采用电喷雾电离源，以多反应监测(MRM)方式进行负离子检测，用于定量分析的离子反应分别为m/z 489→m/z 357(布地奈德)和m/z 493→m/z 413(内标，曲安奈德)。测定血浆中布地奈德方法的线性范围为25.0～2 000 pg·mL^-1，定量下限为25.0 pg·mL^-1，日内、日间精密度(RSD)均小于15%，准确度(RE)在-8.1%～-1.7%。应用本法研究6只比格犬单次和多次给予布地奈德缓释胶囊9 mg后的药代动力学结果显示：单次给药后T_max为(3.5±3.3) h，C_max为(786±498) pg·mL^-1；多次给药后C_max为(2 142±1 515) pg·mL^-1。该法选择性强、灵敏度高、操作简便，适用于布地奈德缓释制剂的药代动力学研究。     

LC-MS/MS法测定人血浆中加巴喷丁的浓度及其药动学研究

建立液相色谱串联质谱(LC-MS/MS)法测定人血浆中加巴喷丁的浓度并将其应用于人体药动学研究。取血浆样品经甲醇沉淀蛋白后,以甲醇0.2%甲酸水溶液(80∶20)为流动相,用Inertsil ODS-3 C18柱(50 mm×2.1 mm ID,3μm)分离,采用电喷雾离子源,以多反应监测(MRM)方式进行正离子检测,定量分析的离子反应分别为m/z 172→m/z 154(加巴喷丁)和m/z 130→m/z 71(内标二甲双胍)。加巴喷丁线性范围为40.8～8.16×103 ng.mL 1,定量限为40.8 ng.mL 1,每个样品测试时间仅2.2 min,日内、日间精密度(RSD)均小于12%,准确度(RE)在±6.4%范围内。应用此法研究了20名健康志愿者单剂量口服加巴喷丁胶囊600 mg后的药动学特点。该方法快速、专属、灵敏、适用性强,可应用于加巴喷丁的人体药动学研究。     

Quantification of vincristine and its major metabolite in human plasma by high-performance liquid chromatography/tandem mass spectrometry

An analytical method using electrospray ionization and high-performance liquid chromatography/tandem mass spectrometry (LC/ESI-MS/MS) was developed to quantify vincristine and M1, the CYP3A-mediated metabolite of vincristine, in human plasma. Vinblastine (internal standard), vincristine, and M1 in plasma were extracted in methylene chloride after acidification with TCAA. The analytes were separated on an Inertsil ODS-3 C18 column (2.1 x 150 mm) with a 5-mum particle size using a gradient elution with a run time of 20 min. The initial mobile phase composition was 0.2% formic acid/water (80:20, v/v) with a final composition of 0.2% formic acid/water (20:80, v/v). Detection was accomplished with multiple reaction monitoring for vinblastine (m/z 406.3--> 271.7), vincristine (m/z 413.2--> 362.2), and M1 (m/z 397.3 --> 376.2). At three concentrations of vincristine and M1, the inter-day and intra-day accuracy and precision were within the acceptable limits for validation (106.8 +/- 9.6% for intra-day, n = 5 each concentration; 90.9 +/- 10.9% for inter-day, n = 4 each concentration). For both vincristine and M1, the concentration limits of quantification and detection were 12 pg/mL and 6 pg/mL, respectively. Stability studies indicated that 80% of M1 degraded in plasma after 15 hours at room temperature (n = 3, high and low QC concentrations). Therefore, short plasma processing times (<30 min) are recommended. The assay was used successfully to quantify vincristine and M1 in pediatric plasma samples up to 24 hours after vincristine administration. Vincristine and M1 concentrations were within the limits of quantification for all patient plasma samples.     

Development and validation of a sensitive and robust LC-tandem MS method for the analysis of warfarin enantiomers in human plasma

A liquid chromatography-tandem mass spectrometry method (LC-MS-MS) was developed and validated for measuring warfarin (WAR) enantiomers (R-WAR and S-WAR) in human EDTA plasma. Liquid-liquid extraction using ethyl ether was used to extract the analytes from the plasma. Baseline resolution of S- and R-WAR as well as the internal standard enantiomers (S- and R-p-ClWAR, S-IS and R-IS) was achieved on a beta-cyclodextrin column with a mobile phase of acetonitrile-acetic acid-triethylamine (1000:3:2.5, v/v/v). The retention times are 6.9, 8.0, 7.0, and 7.9 min for S-WAR, R-WAR, S-IS and R-IS, respectively. The detection was by monitoring S- and R-WAR at m/z 307-->161 and S- and R-IS at m/z 341-->161 using (-) ESI. The standard curve range was 1-100 ng ml(-1) for both S- and R-WAR. The inter-day precision and accuracy of the quality control (QC) samples were <7.3% relative standard deviation (RSD) and <7.3% bias for S-WAR, and <6.5% RSD and <5.8% bias for R-WAR, respectively. Analyte stability during sample processing and storage were established. Method ruggedness was demonstrated by the reproducible performance from analysis of clinical samples.     

Simultaneous determination of loratadine and pseudoephedrine sulfate in human plasma by liquid chromatography-electrospray mass spectrometry for pharmacokinetic studies

To support the pharmacokinetic and bioavailability study of an extended-release loratadine (LOR)/pseudoephedrine sulfate (PES) tablet, a high performance liquid chromatographic-electrospray ionisation-mass spectrometric method (LC-MS) was developed for the simultaneous determination of LOR and PES in human plasma. Diazepam (DP) and phenylpropanolamine (PPA) were used as internal standards for LOR and PES, respectively. Analytes were extracted from alkalized human plasma by liquid/liquid extraction using ethyl ether. Chromatographic separation was performed on an ODS column at flow rate of 0.2 ml/min. The total chromatographic run time was 10.5 min with the retention time of 7.1 min and 6.2 min for LOR and DP, respectively, and 2.2 min for both of PES and PPA. The LOQ was 10 pg/ml and 50 pg/ml for LOR and PES, respectively. The method is accurate and precise enough for its intended purpose.     

Simultaneous determination of pioglitazone and its two active metabolites in human plasma by LC-MS/MS

A liquid chromatography/tandem mass spectrometry (LC-MS/MS) method was developed and validated for the simultaneous determination of pioglitazone (PIO) and its two metabolites: M-III (keto-derivative) and M-IV (hydroxy-derivative) in human plasma. Human plasma samples of 0.2 ml were extracted by a single step liquid-liquid extraction procedure and analyzed using a high performance liquid chromatography (HPLC) electrospray tandem mass spectrometer system. The compounds were eluted isocratically on a C-18 column, ionized using a positive ion atmospheric pressure electrospray ionization source and analyzed using multiple reaction monitoring mode. The ion transitions monitored were m/z 357-->134 for PIO, m/z 371-->148 for M-III, m/z 373-->150 for M-IV and m/z 413-->178 for the internal standard. The chromatographic run time was 2.5 min per injection, with retention times of 1.45, 1.02 and 0.95 min for PIO, M-III and M-IV, respectively. The calibration curves of pioglitazone, M-III and M-IV were well fit over the range of 0.5-2000 ng/ml (r(2)>0.998759) by using a weighted (1/x(2)) quadratic regression. The inter-day precisions of the quality control samples (QCs) were 相似文献   

在线固相萃取-液质联用法定量分析生物基质中丁香醛及其在大鼠体内药代动力学研究中的应用

目的建立测定SD大鼠血浆中丁香醛的在线固相萃取-液质联用法(on-line SPE LC-MS/MS)。方法采用乙腈沉淀蛋白后,取上清经on-line SPE系统进行血浆样品预处理,选用香兰素为内标,以LC-MS/MS法测定大鼠血浆中丁香醛的含量。色谱柱为GRACE Alltima HP C18(50 mm×2.1 mm,5μm),流动相为乙腈-0.1%甲酸(60∶40,V∶V),流速为0.4 ml·min-1。电喷雾离子化(ESI)方式,采用多反应监测,检测离子为正离子,分别选择m/z 182.3→m/z123.1和m/z 153.1→m/z 93.0作为丁香醛和内标物香兰素的检测离子对。结果丁香醛在10.0～2 000μg·L-1范围内线性关系良好(r=0.9997)。方法的准确度(相对误差,RE)范围为-8.5%～5.9%;日内精密度(相对标准偏差,RSD)<16.3%,日间精密度RSD<13.8%。SD大鼠灌胃给予丁香醛(17.5 mg.kg-1和70 mg.kg-1)后,主要药动学参数:T12分别为41.9 min和54.0 min、Cmax分别为361.0μg·L-1和944.0μg·L-1、Tmax分别为10.0 min和27.5 min、CL分别为848.1 ml·min-1.kg-1和683.9 ml·min-1·kg-1。结论建立的基质中丁香醛的测定方法快速、准确、特异性好,适用于丁香醛在SD大鼠体内的药代动力学研究。     

Liquid chromatography/tandem mass spectrometry for the determination of carbamazepine and its main metabolite in rat plasma utilizing an automated blood sampling system

A liquid chromatography/tandem mass spectrometry (LC/MS/MS) method for the simultaneous determination of carbamazepine and its main metabolite carbamazepine 10,11-epoxide in rat plasma is described. The method consists of a liquid-liquid extraction procedure and electrospray LC/MS/MS analysis. The chromatographic separation was achieved within 5 min using a C(8) (150 mm x 2.1mm) 5 microm column with a mobile phase composed of water/acetonitrile/acetic acid (69.5:30:0.5, v/v/v) at a flow rate of 0.4 ml/min. D(10)-carbamazepine is used as the internal standard for all compounds. Analytes were determined by electrospray ionization tandem mass spectrometry in the positive ion mode using selected reaction monitoring (SRM). Carbamazepine was monitored by scanning m/z 237-->194, carbamazepine 10,11-epoxide by m/z 253-->210 and d(10)-carbamazepine by m/z 247-->204. The lower limit of quantitation (LLOQ) is 5 ng/ml for each analyte, based on 0.1 ml aliquots of rat plasma. The extraction recovery of analytes from rat plasma was over 87%. Intra-day and inter-day assay coefficients of variations were in the range of 2.6-9.5 and 4.0-9.6%, respectively. Linearity is observed over the range of 5-2000 ng/ml. This method was used for pharmacokinetic studies of carbamazepine and carbamazepine 10,11-epoxide in response to two different blood sampling techniques (i.e., manual sampling versus automated sampling) in the rat. Several differences between the two sampling techniques suggest that the method of blood collection needs to be considered in the evaluation of pharmacokinetic data.     

Development of a rapid and sensitive LC-MS/MS assay for the determination of sorafenib in human plasma

A rapid and sensitive liquid chromatography/tandem mass spectrometric (LC/MS/MS) assay was developed for the quantitative determination of sorafenib in human plasma. Sample pretreatment involved simple protein precipitation by the addition of 0.5 mL acetonitrile, containing internal standard ([2H3, 15N] sorafenib), to 50 microL of plasma sample volume. Separation was achieved on a Waters SymmetryShield RP8 (2.1 mm x 50 mm, 3.5 microm) column at room temperature using an isocratic elution method with acetonitrile/0.1% formic acid in water: 65/35 (v/v) at a flow rate of 0.25 mL/min. Detection was performed using electrospray ionization in positive ion multiple reaction monitoring (MRM) mode by monitoring the ion transitions from m/z 464.9 --> 252.0 (sorafenib) and m/z 469.0 --> 259.0 (internal standard). Calibration curves were linear in the concentration range of 5-2000 ng/mL. The accuracy and precision values, calculated from three different sets of quality control samples analyzed in quintuplicate on six different days, ranged from 92.86% to 99.88% and from 1.19% to 4.53%, respectively.     

Simple, sensitive and rapid LC-MS/MS method for the quantitation of cerivastatin in human plasma--application to pharmacokinetic studies

A simple and sensitive liquid chromatography-tandem mass spectrometry method was developed and validated for estimation of cerivastatin (I) in human plasma, a potent hydroxy-methylglutaryl-coenzyme A reductase inhibitor. The analyte and internal standard (atorvastatin, II) were extracted by liquid/liquid extraction with diethyl ether/dichloromethane (70/30, v/v). The chromatographic separation was performed on reverse phase Xterra ODS column with a mobile phase of water/acetonitrile (30/70, v/v) with 0.03% formic acid. The protonated analyte was quantitated in positive ionization by multiple reaction monitoring with a mass spectrometer. The mass transitions m/z 460.4 --> 356.3 and 559.2 --> 440.3 were used to measure I and II, respectively. The lower limit of quantitation was 10pg/mL with a relative standard deviation of less than 15%. Acceptable precision and accuracy were obtained for concentrations over the calibration curve ranges (0.01-10ng/mL). Sample analysis time of 2min for each sample made it possible to analyze a throughput of more than 400 human plasma samples per day. The assay can be used to analyze human plasma samples to support phase I and II clinical studies.     

LC-MS/MS方法测定人血浆中多西环素浓度

目的建立迅速、简单、灵敏的液相色谱-串联质谱法测定人血浆中多西环素的浓度。方法血浆样品采用固相萃取的方法,以含0.04%TFA的混合溶液(甲醇-乙腈-水=45∶45∶10,v/v/v)为流动相;采用Dikma C18柱(30 mm×4.6 mm,5μm)分离,通过电喷雾电离源(ESI),以选择性反应监测(SRM)方式进行正离子检测,用于定量分析的离子反应分别为m/z 444.7→428.1(多西环素)和m/z 464.6→448.0(内标,去甲金霉素)。结果建立的人血浆内多西环素测定方法线性范围为50⁵ 000 ng·mL-1,定量下限可达50 ng·mL-1。日内、日间精密度(RSD)均<12%。结论该方法预处理简洁,灵敏,专属性强,可方便用于人血浆中多西环素的测定。     

Development and validation of a sensitive LC-MS/MS method for the determination of adefovir in human serum and urine

A sensitive and selective liquid chromatography-tandem mass spectrometric (LC-MS/MS) method was developed and validated for the determination of adefovir (PMEA) in human serum and urine. The analyte was separated on a Diamonsil C(18) column (250 mm x 4.6 mm i.d., 5 microm particle size) by isocratic elution with methanol-water-formic acid (20:80:0.1, v/v/v) at a flow rate of 0.6 ml/min, and analyzed by mass spectrometry in multiple reaction-monitoring mode. The precursor-to-product ion transitions of m/z 274-->162 and m/z 226-->135 were used to measure and quantify the analyte and internal standard (I.S.), respectively. The weighted (1/x(2)) calibration curve was linear over serum concentration range 1.25-160.00 ng/ml and urine concentration range 0.05-8.00 microg/ml, with a correlation coefficient (r) of 0.9992 and 0.9978, respectively. The lower limit of quantification in human serum was 1.25 ng/ml. The inter- and intra-day precisions (R.S.D.%) in both serum and urine were lower than 8.64%, the mean method accuracies and recoveries from spiked serum samples at three concentrations ranged from 96.3 to 102.0% and 56.5 to 59.3%, respectively. The serum extract was stable when stored for 24h. The developed method was successfully applied to determine PMEA in human serum and urine, and proved suitable to clinical pharmacokinetic study.     

Determination of loratadine and its active metabolite in human plasma by high-performance liquid chromatography with mass spectrometry detection

A new sensitive and selective liquid chromatography coupled with mass spectrometry (LC/MS/MS) method for quantification of loratadine (LOR) and its active metabolite descarboethoxyloratadine (DSL) in human plasma was validated. After addition of the internal standard, metoclopramide, the human plasma samples (0.3 ml) were precipitated using acetonitrile (0.75 ml) and the centrifuged supernatants were partially evaporated under nitrogen at 37 degrees C at approximately 0.3 ml volume. The LOR, DSL and internal standard were separated on a reversed phase column (Zorbax SB-C18, 100 mmx3.0 mm i.d., 3.5 microm) under isocratic conditions using a mobile phase of an 8:92(v/v) mixture of acetonitrile and 0.4% (v/v) formic acid in water. The flow rate was 1 ml/min and the column temperature 45 degrees C. The detection of LOR, DSL and internal standard was in MRM mode using an ion trap mass spectrometer with electrospray positive ionisation. The ion transitions were monitored as follows: 383-->337 for LOR, 311-->(259+294+282) for DSL and 300-->226.8 for internal standard. Calibration curves were generated over the range of 0.52-52.3 ng/ml for both LOR and DSL with values for coefficient of determination greater than 0.994 by using a weighted (1/y) quadratic regression. The lower limits of quantification were established at 0.52 ng/ml LOR and DSL, respectively, with an accuracy and precision less than 20%. Both analytes demonstrated good short-term, long-term, post-preparative and freeze-thaw stability. Besides its simplicity, the sample treatment allows obtaining a very good recovery of both analytes, around 100%. The validated LC/MS/MS method has been applied to a pharmacokinetic study of loratadine tablets on healthy volunteers.     

LC-MS/MS方法测定Beagle犬血浆中比卡鲁胺的浓度

目的 建立灵敏的液相色谱-串联质谱法测定狗血浆中比卡鲁胺的浓度.方法 血浆样品采用乙腈蛋白沉淀方法,以0.2％甲酸-乙腈(35∶65,v/v)为流动相;采用Zorbax SB C18柱(150 mm×4.6mm,5 μm)分离,通过电喷雾电离源,以选择多反应监测(MRM)方式进行负离子检测,用于定量分析的离子反应分别为m/z428.9→254.7(比卡鲁胺)和m/z 269→169.6(内标,甲苯磺丁脲).结果 建立的体内比卡鲁胺测定方法线性范围为5～2 000 ng·mL-1,定量下限可达1 ng·mL-1.日内、日间精密度(RSD)均＜10％.结论 该方法预处理操作简洁、灵敏、专属性强,可方便用于比卡鲁胺药物的体内药动学研究.     

液质联用法测定裸鼠血浆中吉西他滨浓度

目的:建立高效灵敏的液质联用法(LC-MS/MS)测定裸鼠血浆中吉西他滨浓度。方法:选用0.01%醋酸水和乙腈(80∶20)作为流动相;质谱检测选择正离子方式,多离子反应监测(MRM)扫描,用于吉西他滨和内标(头孢克洛)定量分析的离子反应对分别为m/z 264.1→m/z 112.0和m/z 368.1→m/z 174.1。结果:该方法线性范围为5～500 ng.mL-1,定量下限为5 ng.mL-1。日内日间精密度均<12.9%,准确度<3.4%。结论:本方法操作简便、快速,灵敏度高,能够满足裸鼠血浆中吉西他滨检测的要求。     

Development and validation of an automated SPE-LC-MS/MS assay for valdecoxib and its hydroxylated metabolite in human plasma

A sensitive and specific liquid chromatography-tandem mass spectrometry assay was developed to quantitate valdecoxib (I) and its hydroxylated metabolite (II) in human plasma. The analytes (I and II) and a structurally analogue internal standard (IS) were extracted on a C(18) solid phase extraction (SPE) cartridge using a Zymark RapidTrace automation system. The chromatographic separation was performed on a narrow-bore reverse phase Zorbax XDB-C(8) HPLC column with a mobile phase of acetonitrile:water (50:50, v/v) containing 10 mM ammonium acetate. The analytes were ionized using negative electrospray mass spectrometry, then detected by multiple reaction monitoring (MRM) with a tandem mass spectrometer. The precursor to product ion transitions of m/z 313-->118 and m/z 329-->196 were used to measure I and II, respectively. The assay exhibited a linear dynamic range of 0.5-200 ng/ml of I and II in human plasma with absolute recoveries from plasma at 91 and 86%, respectively. The lower limit of quantitation was 0.5 ng/ml for I and II. Acceptable precision and accuracy were obtained for concentrations over the calibration curve ranges (0.5-200 ng/ml). Sample analysis time for each injection was 5 min, a throughput of 70 human plasma standards and samples per run was achieved. The assay has been successfully used to analyze human plasma samples to support clinical phase I and II studies.     

Development and validation of a sensitive LC/MS-MS method for the determination of letrozole in nude mice plasma and its application to a pharmacokinetic study

A sensitive, rapid and simple liquid chromatography-tandem mass spectrometric (LC-MS/MS) method was developed and validated for the determination of letrozole (LTZ) in nude mouse plasma in the current study, which was successfully applied to a pharmacokinetic study. Using anastrozole as internal standard (IS), plasma samples went through a one-step protein precipitation with acetonitrile before determination. The analyte and IS were analyzed on a reversed-phase ZORBAX-SB-C₁₈column (4.6 mm×250 mm, 5 μm) with an isocratic mobile phase consisting of acetonitrile and water containing 0.1% formic acid (v/v) at a flow rate of 1.0 mL/min. The analyte and IS were detected by a triple-quadrupole tandem mass spectrometer, and electrospray and multiple reaction monitoring (MRM) were employed to select LTZ at m/z 286.4/217.1 and IS at m/z 294.1/225.3 simultaneously in the positive ion mode. The calibration curve showed good linearity ranging from 0.8–2000.0 ng/mL (r>0.99). The intra-day and inter-day precisions of LTZ were 4.0%–8.4%, with an accuracy of 98.6%–104.9%. Using this method, we successfully characterized the pharmacokinetics (PK) of LTZ by a one-compartment model with first-order absorption in female BALB/c nude mice.     

大鼠血浆中奥昔布宁及其N-去乙基代谢物的UPLC-MS/MS法测定

建立了超高效液相色谱-串联质谱法同时测定大鼠血浆中的奥昔布宁及其主要代谢物N-去乙基奥昔布宁.用液-液萃取法处理血样,以盐酸奥昔布宁-D11为内标,采用ESI源正离子模式、多反应监测进行测定.检测离子对分别为m/z 358.2→124.1(奥昔布宁)、m/z 330.2→96.1 (N-去乙基奥昔布宁)和m/z 369.3→142.1(奥昔布宁-D11).奥昔布宁及N-去乙基奥昔布宁在0.5～100 ng/ml和0.2～40 ng/ml浓度范围内线性关系良好,日内、日间RSD均小于8％.     

Liquid chromatography-tandem mass spectrometry (LC-MS/MS) method for simultaneous determination of tenofovir and emtricitabine in human plasma and its application to a bioequivalence study

A rapid and sensitive liquid chromatography-tandem mass spectrometry (LC-MS/MS) method has been developed and validated for simultaneous quantification of Tenofovir (TEN) and Emtricitabine (EMT) in human plasma using Chromolith Speed Rod RP18. The mass transition ion-pair has been followed as m/z 288.10-->176.10 for TEN, m/z 248.20-->130.20 for EMT and m/z 230.10-->112.10 for Lamivudine (LAM). The method involves solid phase extraction from plasma, simple isocratic chromatographic conditions and mass spectrometric detection using an API 5000 instrument that enables detection at nanogram levels. Lamivudine was used as the internal standard. The proposed method has been validated with a linear range of 10-600 ng/ml for TEN and 25-2,500 ng/ml for EMT. The intrarun and interrun precision values are within 12.0% for TEN and 15.6% for EMT at their respective LOQ levels. The overall recoveries for TEN and EMT were 84.3% and 68.5%, respectively. Total elution time was as low as 2 min.