Challenges in optimizing sample preparation and LC‐MS/MS conditions for the analysis of carglumic acid,an N‐acetyl glutamate derivative in human plasma

This paper describes a systematic approach to overcoming challenges in developing a robust and selective liquid chromatography‐tandem mass spectrometry (LC‐MS/MS) method for reliable and precise determination of carglumic acid in human plasma. Sample extraction was tested on several reversed‐phase solid‐phase extraction (SPE) sorbents with different chemistries, such as hydrophobic C18, hydrophilic‐lipophilic balance, and mixed‐mode cation and anion exchange. The best recovery under the optimized extraction conditions was obtained with Oasis MAX (30 mg, 1cc) mixed‐mode anion exchange (~ 50%) cartridge, compared to other sorbents from 100 μL plasma sample. Complete analytical separation of carglumic acid and carglumic acid‐13C5 15N as an internal standard (IS) from endogenous plasma components was achieved on ACE 5CN (150 × 4.6 mm, 5 µm) column under isocratic conditions using acetonitrile:methanol (50:50, v/v) ? 0.1% acetic acid in water [80:20, v/v] as the mobile phase. The deprotonated precursor → product ion transitions for carglumic acid (189/146) and IS (195/152) were monitored in the negative ionization mode on a triple quadrupole mass spectrometer. The regression curves were linear over a concentration range of 6.00‐6000 ng/mL (r² ≥ 0.9987). Matrix effect was evaluated in terms of IS‐normalized matrix factors, which ranged from 0.95 to 1.01 across four quality control levels. Intra‐ and inter‐batch accuracy and precision, and the stability of carglumic acid in spiked plasma samples were assessed under different conditions. The method was applied to assess the pharmacokinetics of 100 mg/kg body weight carglumic acid in a healthy Indian subject. Copyright © 2015 John Wiley & Sons, Ltd.     

液相色谱-串联质谱法测定苯妥英在健康人体的血药浓度并研究其生物等效性

目的建立快速、灵敏的液相色谱-串联质谱法测定健康人血浆中的苯妥英(抗癫痫药),并进行生物等效性研究。方法血浆样品50μL经液- 液萃取后,以甲醇-水-甲酸(90:10:0．2)为流动相,Zorbax SB-C18柱分离; 样品经大气压化学电离源(APCI)正离子化后,通过三重四极杆串联质谱仪, 用选择反应监测(SRM)对苯妥英(m／z 253→m／z 182)和柳胺酚(m／z 230→ m／z 121,内标)进行测定。用此法测定了20名受试者单剂量口服受试和参比制剂后苯妥英的血药浓度。结果线性范围为2．5—3 000 ng·mL-1,定量下限为2．5 ng·mL-1;日内、日问精密度(RSD)均<7．0％,准确度(RE)在 -0．5％～2．3％。2种制剂的Cmax、AUC0-t均无显著性差异。结论该法专属、灵敏、快速,适用于复方制剂中苯妥英的生物等效性评价。     

LC-MS/MS法测定伊曲康唑的血浆浓度及在生物等效性试验中的应用(英文)

目的建立液相色谱-串联质谱(LC MS/MS)法测定人血浆中伊曲康唑浓度方法,并评价伊曲康唑受试制剂和参比制剂是否具有生物等效性。方法采用蛋白沉淀的前处理方法,以10μL进样分析。色谱柱为CAPCELLPAKC18柱,流动相为V(乙腈)∶V(水)∶V(甲酸)=75∶25∶0.15),流速为0.20 mL.min-1。串联质谱API 3000采用正电喷雾离子源和多反应监测的扫描模式。结果伊曲康唑在质量浓度2～500μg.L-1内线性良好,定量下限为2μg.L-1,批内、批间的RSD均<10%。伊曲康唑受试制剂和参比制剂的相对生物利用度为(112.71±59.58)%,经方差分析及双单侧t检验表明两者具有生物等效性。结论该方法可以准确、快速和灵敏地应用于伊曲康唑的药动学与生物等效性研究,结果显示受试制剂和参比制剂生物等效。     

LC-MS/MS法同时测定厄贝沙坦和氢氯噻嗪血药浓度及厄贝沙坦氢氯噻嗪片的生物等效性研究

目的建立同时测定人血浆中厄贝沙坦和氢氯噻嗪血药浓度的液相色谱-质谱方法,并评价2种厄贝沙坦氢氯噻嗪片的生物等效性。方法以氯沙坦为内标,血浆样品经固相萃取后检测。Agilent Zorbax SB-C18(4.6 mm×50mm,1.8μm)为分析柱,以甲醇-乙腈-0.15%甲酸水溶液(40∶25∶35)为流动相,流速:0.3 m L·min^-1,用电喷雾离子化源(ESI),以负离子多离子反应监测(MRM)扫描方式进行监测。22例健康男性受试者随机于2个周期交叉口服相同剂量的厄贝沙坦氢氯噻嗪片受试药物或参比药物,考察其生物等效性。结果血浆中厄贝沙坦和氢氯噻嗪浓度分别在20.0~4000.0 ng·m L^-1和1.0~200.0 ng·m L^-1线性关系良好,线性回归方程分别为y=9.28×10^-2x-7.50×10^-2(r=0.999 2)和y=5.27×10-2x-2.99×10-2(r=0.998 8),日内和日间RSD均小于15%。受试药物相对于参比药物的厄贝沙坦和氢氯噻嗪的生物利用度分别为(107.4±19.9)%和(105.6±16.2)%。结论本方法简便、快速、灵敏度高,适用于人血浆中厄贝沙坦、氢氯噻嗪的浓度测定,可用于厄贝沙坦氢氯噻嗪片的人体生物等效性研究;2种药物生物等效。     

Development and validation of an LC‐MS/MS method after chiral derivatization for the simultaneous stereoselective determination of methylenedioxy‐methamphetamine (MDMA) and its phase I and II metabolites in human blood plasma

3,4‐Methylenedioxymethamphetamine (MDMA, ecstasy) is a racemic drug of abuse and its two enantiomers are known to differ in their dose‐response curves. The S‐enantiomer was shown to be eliminated at a higher rate than the R‐enantiomer. The most likely explanation for this is a stereoselective metabolism also claimed in in vitro studies. Urinary excretion studies showed that the main metabolites in humans are 4‐hydroxy 3‐methoxymethamphetamine (HMMA) 4‐sulfate, HMMA 4‐glucuronide and 3,4‐dihydroxymethamphetamine (DHMA) 3‐sulfate. For stereoselective pharmacokinetic analysis of phase I and phase II metabolites in human blood plasma useful analytical methods are needed. Therefore the aim of the presented study was the development and validation of a stereoselective liquid chromatography‐tandem mass spectrometry (LC‐MS/MS) method for the simultaneous quantification of MDMA, 3,4‐methylenedioxyamphetamine, DHMA, DHMA 3‐sulfate, HMMA, HMMA 4‐glucuronide, HMMA 4‐sulfate, and 4‐hydroxy 3‐methoxyamphetamine in blood plasma for evaluation of the stereoselective pharmacokinetics in humans. Blood plasma samples were prepared by simple protein precipitation and afterwards all analytes were derivatized using N‐(2,4‐dinitro‐5‐fluorophenyl) L‐valinamide resulting in the formation of diastereomers which were easily separable on standard reverse phase stationary phases. This simple and fast method was validated according to international guidelines including specificity, recovery, matrix effects, accuracy and precision, stabilities, and limits of quantification. The method proved to be selective, sensitive, accurate and precise for all tested analytes except for DHMA. Copyright © 2014 John Wiley & Sons, Ltd.     

Development and validation of a multi‐analyte LC‐MS/MS approach for quantification of neuroleptics in whole blood,plasma, and serum

Based on a similar approach for quantification of antidepressants, benzodiazepines, and z‐drugs, a liquid chromatography‐tandem mass spectrometry (LC‐MS/MS) multi‐analyte approach with simple liquid‐liquid extraction was extended for fast target screening and quantification of neuroleptics in whole blood, plasma, and serum. As this method is part of a multi‐analyte procedure for over 100 analytes from different drug classes and as the extracts were additionally used in the authors' laboratory for gas chromatography‐mass spectrometry (GC‐MS) analysis, one universal stable‐isotope‐labelled internal standard (SIL‐IS) was used to save time and resource. The method was validated with respect to international guidelines. For accuracy and precision, full calibration was performed with ranges from subtherapeutic to toxic concentrations. Selectivity problems could not be observed, but matrix effects ranged from 68 to 211% in all samples. For the low quality control (QC), recovery ranged from 32 to 112%, process efficiency from 31 to 165% and for the high QC recovery from 42 to 141%, process efficiency from 29 to 154%. In addition statistical data evaluation of the variances of the recovery, matrix effects, and process efficiency data between whole blood vs. plasma, whole blood vs. serum, and plasma vs. serum were done. The presented LC‐MS/MS approach was applicable for selective detection of 33 neuroleptics as well as accurate and precise quantification of 25 neuroleptics in whole blood, 19 in plasma, and 17 in serum. More significant matrix effects (ME) for neuropletic drugs overall in plasma and serum as compared with whole blood were detected. Copyright © 2015 John Wiley & Sons, Ltd.     

LC−MS/MS assay for the determination of a novel anti‐fibrotic candidate mefunidone in monkey plasma and its application to a pharmacokinetics study

Mefunidone (MFD) is a promising anti‐fibrotic candidate molecule with greater anti‐fibrotic activity than pirfenidone (PFD). However, there has been no report on the methodology used for the quantification of MFD or on any investigation of its pharmacokinetics. In this study, an efficient and reliable liquid chromatography–tandem mass spectrometry (LC–MS/MS) method was developed to assay MFD in monkey plasma. This assay method was validated and applied to a pharmacokinetics study in monkeys. The lower limit of quantification of this assay was 0.1 μg·mL^?1, and the linear calibration curve was acquired with R² > 0.99 between 0.1 and 60 μg·mL^?1. The intra‐day and inter‐day precision were evaluated with coefficient of variations of 1.5%–5.8%, whereas the mean accuracy ranged from 91.7% to 106.9%. A negligible matrix effect and good recovery were obtained using this assay, with average extraction recoveries of MFD and the internal standard (IS) in the range of 85.5%–124.8% and 84.1%–94.0%, respectively. The precision of the absolute matrix effect of MFD and the IS was 1.2–3.0% and 1.2–7.3%, respectively. The samples were stable under all experimental conditions. Linear pharmacokinetics were observed for MFD in monkeys, where the exposures of MFD increased proportionally with increasing MFD doses at the range of 10–90 mg·kg^?1. Moderate elimination of MFD from the body was observed, with t_1/2 of 5–7 h, and the elimination rate of MFD was stable during multiple dosing. In conclusion, this method provides an reliable analytical approach for quantification of MFD in plasma and was successfully applied to a pharmacokinetics study in monkeys.     

LC／MS／MS法测定人血中硫普罗宁的浓度及药物动力学研究

目的建立LC／MS／MS法测定人血中硫普罗宁浓度的方法,并研究其在健康男性受试者体内的药物动力学。方法采用LC／MS／MS（ESI源）测定硫普罗宁的血浆浓度,计算药物动力学参数。结果硫普罗宁线性范围为25．0-5000ng·mL^-1,定量下限为25．0ng·mL^-1日内、日间精密度（RSD）均〈15％,准确度（RE）在15％以内。应用本法测得20名健康男性受试者口服200mg硫普罗宁胶囊后主要药代动力学参数为：tmax,为（4．20±1．01）h,t1／2为（5．61±4．42）h,Cmax为（4456±2447）ng·mL^-1,AU C0-24h为（20566±9902）ng·mL^-1,Ke为（0．173±0．094）h^-1。结论该法操作简便、快速、灵敏,可用于测定血浆中硫普罗宁浓度。     

基于超高效液相色谱-串联质谱快速检测索非布韦及其代谢物在大鼠体内的生物等效性分析

目的：建立超高效液相色谱-串联质谱（UPLC-MS/MS）快速并同时测定大鼠血浆中抗丙肝药索非布韦及其代谢物GS-331007的含量,探讨索非布韦代谢产物作为标记物测定药时曲线的可能性,研究不同厂家抗丙肝药索非布韦在大鼠体内的生物等效性。方法：通过液质联用检测原研药A和仿制药B以36 mg·kg^-1灌胃大鼠各时间点索非布韦和GS-331007的血药浓度。用DAS 2.1.1和SPSS 17.0软件计算药动学参数并比较原研药A和仿制药B的一致性。结果：原研药A和仿制药B中索非布韦药动学参数C_max分别为（1 376.08±174.95）ng·mL^-1和（1 297.58±164.93）ng·mL^-1,t_max分别为（0.75±0.08）h和（0.72±0.16）h,t_1/2分别为（1.57±0.20）h和（1.73±0.45）h,AUC_（0→t）分别为（2 691.67±280.85）ng·mL^-1·h和（2 851.20±199.54）ng·mL^-1·h,AUC_{（0→∞）}分别为（2 748.51±258.91）ng·mL^-1·h和（3 007.75±364.02）ng·mL^-1·h,原研药A和仿制药B代谢物GS-331007 C_max分别为（1 302.52±163.73）ng·mL^-1和（1 430.88±107.52）ng·mL^-1,t_max分别为（3.97±0.74）h和（3.95±1.38）h,t_1/2分别为（5.56±2.55）h和（5.44±1.38）h,AUC_（0→t）分别为（9 723.24±1170.38）ng·mL^-1·h和（9 032.31±1 037.76）ng·mL^-1·h,AUC_{（0→∞）}分别为（9 893.26±1 251.89）和（9 316.90±1 293.44）ng·mL^-1·h。结论：本实验建立的UPLC-MS/MS方法可在3.5 min内准确测定大鼠血浆中索非布韦及其代谢物GS-331007含量。根据索非布韦和其代谢物GS-331007药时曲线得出原研药A和仿制药B的药动学参数一致性较好（P>0.05）。本工作发现用代谢物GS-331007作为索非布韦生物等效性研究的可能性。     

液相色谱-串联质谱法测定人血浆中二甲双胍的浓度

目的:建立LC-MS/MS法测定人血浆中二甲双胍的浓度。方法:人血浆样本以乙腈沉淀蛋白后,选用Zorbax SB-C18Narrow-Bore色谱柱(150 mm×2.1 mm,5μm),以甲醇-10 mmol.L-1乙酸铵(含1%甲酸)(5:95)为流动相,流速为0.3 mL.min-1;选用API3200型三重四极杆串联质谱仪的多重反应监测(MRM)扫描方式进行监测,电喷雾离子化源,正离子方式,选择监测离子反应分别为m/z130.1→m/z71.0(二甲双胍)和m/z147.1→m/z58.2(内标米曲肼)。结果:二甲双胍和米曲肼的保留时间分别为1.27 min和1.26 min;血浆中二甲双胍的线性范围为0.010～3.000 mg.L-1(r>0.99),定量下限为0.010mg.L-1;日内、日间RSD均小于6%;相对偏差(RE)均在±6%的范围以内;平均提取回收率为(86.6±5.4)%;稳定性试验中,在各种贮存条件下血浆中二甲双胍均较稳定。结论:该方法快速、灵敏,专属性强,重现性好,适用于人血浆中二甲双胍浓度的测定,可应用于盐酸二甲双胍肠溶片的人体生物等效性研究。     

Elucidation of the metabolites of the novel psychoactive substance 4‐methyl‐N‐ethyl‐cathinone (4‐MEC) in human urine and pooled liver microsomes by GC‐MS and LC‐HR‐MS/MS techniques and of its detectability by GC‐MS or LC‐MSn standard screening approaches

4‐methyl‐N‐ethcathinone (4‐MEC), the N‐ethyl homologue of mephedrone, is a novel psychoactive substance of the beta‐keto amphetamine (cathinone) group. The aim of the present work was to study the phase I and phase II metabolism of 4‐MEC in human urine as well as in pooled human liver microsome (pHLM) incubations. The urine samples were worked up with and without enzymatic cleavage, the pHLM incubations by simple deproteinization. The metabolites were separated and identified by gas chromatography‐mass spectrometry (GC‐MS) and liquid chromatography‐high resolution‐tandem mass spectrometry (LC‐HR‐MS/MS). Based on the metabolites identified in urine and/or pHLM, the following metabolic pathways could be proposed: reduction of the keto group, N‐deethylation, hydroxylation of the 4‐methyl group followed by further oxidation to the corresponding 4‐carboxy metabolite, and combinations of these steps. Glucuronidation could only be observed for the hydroxy metabolite. These pathways were similar to those described for the N‐methyl homologue mephedrone and other related drugs. In pHLM, all phase I metabolites with the exception of the N‐deethyl‐dihydro isomers and the 4‐carboxy‐dihydro metabolite could be confirmed. Glucuronides could not be formed under the applied conditions. Although the taken dose was not clear, an intake of 4‐MEC should be detectable in urine by the GC‐MS and LC‐MSⁿ standard urine screening approaches at least after overdose. Copyright © 2014 John Wiley & Sons, Ltd.     

液相色谱-质谱联用法测定人血浆双氢青蒿素浓度

目的:建立液相色谱-质谱联用法测定健康人血浆中双氢青蒿素浓度的方法。方法:以青蒿素为内标,血浆样品采用液-液萃取法处理。用电喷雾离子化和正离子多离子反应监测方式检测双氢青蒿素。结果:该方法双氢青蒿素线性范围为1.01~2020 ng.ml-1;定量下限为1.001±0.072 ng.ml-1;方法回收率在93.0%~98.2%;批内、批间变异系数均<10%。结论:该方法准确、灵敏、特异、简便,适用于健康人血浆双氢青蒿素浓度的测定。     

LC‐MS/MS detection of unaltered glucuronoconjugated metabolites of metandienone

The aim of this study was to evaluate the direct detection of glucuronoconjugated metabolites of metandienone (MTD) and their detection times. Metabolites resistant to enzymatic hydrolysis were also evaluated. Based on the common mass spectrometric behaviour of steroid glucuronides, three liquid chromatography‐tandem mass spectrometry (LC‐MS/MS) strategies were applied for the detection of unpredicted and predicted metabolites: precursor ion scan (PI), neutral loss scan (NL), and theoretical selected reaction monitoring (SRM) methods. Samples from four excretion studies of MTD were analyzed for both the detection of metabolites and the establishment of their detection times. Using PI and NL methods, seven metabolites were observed in post‐administration samples. SRM methods allowed for the detection of 13 glucuronide metabolites. The detection times, measured by analysis with an SRM method, were between 1 and 22 days. The metabolite detected for the longest time was 18‐nor‐17β‐hydroxymethyl‐17α‐methyl‐5β‐androsta‐1,4,13‐triene‐3‐one‐17‐glucuronide. One metabolite was resistant to hydrolysis with β ‐glucuronidase; however it was only detected in urine up to four days after administration. The three glucuronide metabolites with the highest retrospectivity were identified by chemical synthesis or mass spectrometric data, and although they were previously reported, this is the first time that analytical data of the intact phase II metabolites are presented for some of them. The LC‐MS/MS strategies applied have demonstrated to be useful for detecting glucuronoconjugated metabolites of MTD, including glucuronides resistant to enzymatic hydrolysis which cannot be detected by conventional approaches. Copyright © 2016 John Wiley & Sons, Ltd.     

Toxicokinetics of new psychoactive substances: plasma protein binding,metabolic stability,and human phase I metabolism of the synthetic cannabinoid WIN 55,212‐2 studied using in vitro tools and LC‐HR‐MS/MS

The new psychoactive substance WIN 55,212‐2 ((R)‐(+)‐[2,3‐dihydro‐5‐methyl‐3‐(4‐morpholinylmethyl)pyrrolo‐[1,2,3‐de]‐1,4‐benzoxazin‐6‐yl]‐1‐napthalenylmethanone) is a potent synthetic cannabinoid receptor agonist. The metabolism of WIN 55,212‐2 in man has never been reported. Therefore, the aim of this study was to identify the human in vitro metabolites of WIN 55,212‐2 using pooled human liver microsomes and liquid chromatography‐high resolution‐tandem mass spectrometry (LC‐HR‐MS/MS) to provide targets for toxicological, doping, and environmental screening procedures. Moreover, a metabolic stability study in pooled human liver microsomes (pHLM) was carried out. In total, 19 metabolites were identified and the following partly overlapping metabolic steps were deduced: degradation of the morpholine ring via hydroxylation, N‐ and O‐dealkylation, and oxidative deamination, hydroxylations on either the naphthalene or morpholine ring or the alkyl spacer with subsequent oxidation, epoxide formation with subsequent hydrolysis, or combinations. In conclusion, WIN 55,212‐2 was extensively metabolized in human liver microsomes incubations and the calculated hepatic clearance was comparably high, indicating a fast and nearly complete metabolism in vivo. This is in line with previous findings on other synthetic cannabinoids. Copyright © 2016 John Wiley & Sons, Ltd.     

HPLC-MS/MS测定人血浆中羧甲司坦的浓度及其生物等效性研究

目的 建立HPLC-MS/MS方法测定人血浆中的羧甲司坦体内浓度,用于人体生物等效性分析.方法 以直接沉淀法处理血浆,色谱柱为XTerra MS C18柱(150 mm×2.1mm,5μm);流动相为甲醇-20 mmol,L-1甲酸铵(含0.5％甲酸)(75∶25,v/v);流速为0.3 mL· min-1;柱温为40℃.采用电喷雾离子源(ESI)负离子模式,用多反应监测模式(MRM)进行定量分析.结果 羧甲司坦线性范围为0.210 9～54 μg,mL-1,定量下限为0.210 9 μg·mL-1,绝对回收率＞80％,日内和日间RSD均＜15％.结论 此方法能够简便、高效、快速和灵敏的检测健康志愿者血浆中的羧甲司坦,可用于羧甲司坦药物代谢动力学及生物等效性研究.     

Development and validation of an LC‐MS/MS method for simultaneous determination of piperaquine and 97‐63, the active metabolite of CDRI 97‐78, in rat plasma and its application in interaction study

Piperaquine‐dihydroartemisinin combination is the latest addition to the repertoire of ACTs recommended by the World Health Organization (WHO) for treatment of falciparum malaria. Due to the increasing resistance to artemisinin derivatives, CSIR‐CDRI has developed a prospective short acting, trioxane antimalarial derivative, CDRI 97‐78. In the present study, a liquid chromatography‐electrospray ionization‐tandem mass spectrometry (LC–ESI‐MS/MS) method for the simultaneous quantification of piperaquine (PPQ) and 97‐63, the active metabolite of CDRI 97‐78 found in vivo, was developed and validated in 100 μL rat plasma using halofantrine as internal standard. PPQ and 97‐63 were separated using acetonitrile:methanol (50:50, v/v) and ammonium formate buffer (10 mM, pH 4.5) in the ratio of 95:5(v/v) as mobile phase under isocratic conditions at a flow rate of 0.65 mL/min on Waters Atlantis C₁₈ (4.6 × 50 mm, 5.0 µm) column. The extraction recoveries of PPQ and 97‐63 ranged from 90.58 to 105.48%, while for the internal standard, it was 94.27%. The method was accurate and precise in the linearity range 3.9–250 ng/mL for both the analytes, with a correlation coefficient (r) of ≥ 0.998. The intra‐ and inter‐day assay precision ranged from 2.91 to 8.45% and; intra‐ and inter‐day assay accuracy was between 92.50 and 110.20% for both the analytes. The method was successfully applied to study the effect of oral co‐administration of PPQ on the pharmacokinetics of CDRI 97‐78 in Sprague‐dawley rats and vice versa. The co‐administration of CDRI 97‐78 caused significant decrease in AUC_0–∞ of PPQ from 31.52 ± 2.68 to 14.84 ± 4.33 h*µg/mL. However, co‐administration of PPQ did not have any significant effect on the pharmacokinetics of CDRI 97‐78. Copyright © 2015 John Wiley & Sons, Ltd.     

UHPLC-MS/MS测定人血浆中依匹哌唑浓度及其生物等效性研究

目的 建立简便灵敏的超高效液相色谱-串联质谱法(ultra performance liquid chromatography tandem mass spectrometry,UHPLC-MS/MS)测定人血浆中依匹哌唑浓度,并应用于2种片剂的生物等效性研究。方法 采用Waters Acquity UPLC BEH C₁₈色谱柱(2.1 mm×50 mm,1.7μm),以0.1%甲酸水溶液(A)-乙腈-甲醇(50∶50,含0.1%甲酸)(B)梯度洗脱,流速0.4 mL·min^–1,进样量为2μL,柱温40℃,以正离子MRM模式测定依匹哌唑(m/z 434.2→273.2)的浓度,依匹哌唑-d₈(m/z 442.4→281.3)作为内标,离子源为ESI源。血浆样本加入内标,加入甲醇后进行蛋白沉淀,取上清液稀释后进样检测。结果 依匹哌唑在0.2～50 ng·mL^–1呈线性关系,定量下限为0.2 ng·mL^–1,质控样品批内、批间精密度CV≤5.0%,准确度相对偏差在标示值–1.7...     

Method development and validation of montelukast in human plasma by HPLC coupled with ESI-MS/MS: application to a bioequivalence study

A simple, sensitive, and specific LC-ESI–MS/MS method for quantification of Montelukast (MO) in human plasma using Montelukast-d₆ (MOD6) as an internal standard (IS) is discussed here. Chromatographic separation was performed on YMC-pack pro C₁₈, 50 x 4.6 mm, S-3 μm column with an isocratic mobile phase composed of 10mM ammonium formate (pH 4.0):acetonitrile (20:80 v/v), at a flow-rate of 0.8 mL min⁻¹. MO and MOD6 were detected with proton adducts at m/z 586.2→568.2 and 592.3→574.2 in multiple reaction monitoring (MRM) positive mode respectively. MO and MOD6 were extracted using acetonitrile as precipitating agent. The method was validated over a linear concentration range of 1.0–800.0 ng mL⁻¹ with correlation coefficient (r²) ≥ 0.9996. The intraday precision and accuracy were within 1.91–7.10 and 98.32–99.17. The inter-day precision and accuracy were within 3.42–4.41% and 98.14–99.27% for MO. Both analytes were found to be stable throughout three freeze-thawing cycles, bench top, and autosampler stability studies. This method was utilized successfully for the analysis of plasma samples following oral administration of MO (5 mg) in 31 healthy Indian male human volunteers under fasting conditions.     

液相色谱-质谱联用法测定人参皂苷Re在健康人血浆的浓度

目的建立液相色谱-质谱联用法测定人参皂苷Re在健康人血浆中浓 度的方法。方法血浆样品用固相萃取法处理。用电喷雾离子化和正离子多 离子反应监测方式检测人参皂苷Re。结果该方法人参皂苷Re线性范围为 1．05～1 050 ng·mL-1;定量下限为1．05 ng·mL-1;方法回收率在99．3％～ 104．3％;日内、日间变异系数(RSD)均<15％。结论该法准确、灵敏、特异, 适用于健康人血浆人参皂苷Re浓度测定。     

Application of an LC‐MS/MS method to the pharmacokinetics of TM‐2, a potential antitumour agent,in rats

TM‐2 is a novel semi‐synthetic taxane derivative, selected for preclinical development based on its greater anticancer activity and lower toxicity compared with docetaxel. In this study, a rapid and sensitive analytical method based on ultra performance liquid chromatography‐tandem mass spectrometry (LC‐MS/MS) has been developed for the determination of TM‐2 in rat plasma. The biological samples were extracted with methyl tert‐butyl ether and separated on a C₁₈ column (50 mm × 2.1 mm, 1.7 µm) using a mobile phase consisting of acetonitrile and 2 mM ammonium acetate. The standard curves were linear over the range 5–1000 ng/mL in rat plasma. The precision (relative standard deviation, RSD, %) were within 14.5%, and the accuracy (relative error, RE, %) ranged from ?1.56 to 2.36%. Recovery and matrix effect were satisfactory in rat plasma. The validated method was successfully applied to pharmacokinetic studies after intravenous administration of TM‐2 to rats. The pharmacokinetics of TM‐2 in rats were characterized by a large volume of distribution and a long half‐life of elimination after single dose (4, 8, and 16 mg/kg), and a good correlation was observed between AUC and dose. The preclinical data will be useful for the design of subsequent trials of TM‐2. Copyright © 2014 John Wiley & Sons, Ltd.