Determination of pinostrobin in rat plasma by LC-MS/MS: application to pharmacokinetics

A rapid and sensitive method for the determination of pinostrobin in rat plasma was developed using liquid chromatography tandem mass spectrometry (LC-MS/MS) for the first time. Isoliquiritigenin was used as an internal standard in rat plasma. Chromatographic separation was performed on an HiQ Sil C₁₈ column with isocratic elution at a flow rate of 1 mL/min. The mobile phase consisted of water and methanol (9:91, v/v) containing 0.1% formic acid. The quantification limit was 10 ng/mL within a linear range of 10-1000 ng/mL (R = 0.9984). The intra- and inter-day assay precision ranged from 3.8-5.3% to 3.2-5.2%, respectively, and the intra- and inter-day assay accuracy was between 93.2-95.1% and 95.5-104.3%, respectively. Our results indicated that the LC-MS/MS method is effective for pharmacokinetic study of pinostrobin in rat plasma.     

Determination of chamaechromone in rat plasma by liquid chromatography-tandem mass spectrometry: application to pharmacokinetic study

A rapid, simple and accurate method was developed for the determination of chamaechromone in rat plasma using liquid chromatography tandem mass spectrometry (LC–MS–MS). Rosuvastatin was used as the internal standard. The plasma samples were extracted by liquid–liquid extraction with ethyl acetate. Chromatographic separation was performed on Xbridge™ C₁₈ column (2.1 mm × 50 mm, 3.5 μm) with linear gradient elution using water and methanol, both of which were acidified with 0.1% aqueous formic acid. The flow rate was 0.4 mL/min and the total run time was 6 min. Detection was performed on a triple-quadrupole tandem mass spectrometer using positive ion mode electrospray ionization (ESI) in the multiple reaction monitoring (MRM) mode. The MS/MS ion transitions monitored were m/z 543.3 → 198.9 and 481.9 → 258.3 for chamaechromone and rosuvastatin, respectively. Good linearity was observed over the concentration range of 8–6400 ng/mL in 0.1 mL of rat plasma. The lowest concentration (8 ng/mL) in the calibration curve was estimated as LLOQ with both deviation of accuracy and RSD of precision <20% (n = 6). Intra-assay and inter-assay variability were less than 11% in plasma. This method was successfully applied to a pharmacokinetic study of chamaechromone in rats after intravenous (5 mg/kg) and oral (100 mg/kg) administration. Following oral administration the concentration–time curve of chamaechromone exhibited a biphasic absorption profile. The maximum mean concentration in plasma (C_max, 795.9 ± 14.6 ng/L) was achieved at 11.3 ± 0.8 h (T_max) and the area under curve (AUC_0–60) was 6976.7 ± 1026.9 ng h/L. After single intravenously administration of chamaechromone, the essential pharmacokinetic parameters C_max, AUC_0–48 were 4300.7 ± 113.6 ng/L and 3672.1 ± 225.4 ng h/L, respectively. The result showed that the compound was poorly absorbed with an absolute bioavailability being approximately 8.9%.     

Quantitative determination of spinosin in rat plasma by liquid chromatography-tandem mass spectrometry method

A sensitive method for the quantitative determination of spinosin in rat plasma was developed and validated using high-performance liquid chromatographic separation with tandem mass spectrometric detection. The analytes of interest were extracted from rat plasma samples by methyl tert-butyl ether (MTBE) after acidification with 1.0% acetic acid aqueous solution. Chromatographic separation was achieved on an Agilent Zorbax SB-C(18) (50 mm x 4.6 mm, 5 microm) using a isocratic mobile phase consisting of acetonitrile-water (30:70, v/v) with 1% isopropyl alcohol and 0.01% heptafluorobutyric acid. The flow rate was 0.2 ml/min. The column temperature was maintained at 25 degrees C. Detection was performed on a triple quadrupole tandem mass spectrometer by selected reaction monitoring (SRM) mode via electrospray ionization (ESI). The calibration curve was linear over the range of 1.00-400 ng/ml in rat plasma, with 1.00 ng/ml of the lower limit of quantification (LLOQ). The inter- and intra-day precisions and accuracy for all samples were satisfactory. The validated method was successfully applied for the pharmacokinetic study of spinosin in rat. After oral administration of 20mg/kg spinosin to rats, the main pharmacokinetic parameters of T(max), C(max), T(0.5) and AUC(0-T) were 5.33+/-0.58 h, 132.2+/-10.6 ng/ml, 4.89+/-0.37 h, 1.02+/-0.09 microg h/l, respectively.     

Determination of a novel low-voltage-activated calcium channel blocker (HYP-10) in rat plasma by liquid chromatography-mass spectrometry

A novel T-type calcium channel blocker, 4-amino-1-{4-[(4-chloro-phenyl)-phenyl-methyl]-piperazin-1-yl}-butan-1-one (HYP-10) has been synthesized, and the compound has shown promise as both a nociceptive and inflammatory pain reliever as well as an analgesic in a rat neuropathic pain model. A quantification method was developed for the determination of HYP-10 in rat plasma. After simple protein precipitation with methanol, HYP-10 and the internal standard, methaqualone were chromatographed on a reversed-phase column and detected by liquid chromatography/tandem mass spectrometry with electrospray ionization. The accuracy and precision of the assay were in accordance with FDA regulations for validation of bioanalytical methods. This method was applied to measure the plasma HYP-10 concentration after a single intravenous administration of the compound in rats.     

Quantitative assay for bradykinin in rat plasma by liquid chromatography coupled to tandem mass spectrometry

An assay to quantify bradykinin in rat plasma has been developed and validated, using liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). Sar-D-Phe(8)-des-Arg(9)-bradykinin was used as internal standard. Aprotinin was added to rat plasma to inhibit the activity of proteinases. Recoveries for solid-phase extraction (SPE) on Strata X reversed phase were greater than 80%. Multiple reaction monitoring (MRM) on a triple quadrupole mass spectrometer equipped with an electrospray source (ESI), operating in the positive ion-mode, was used for detection. The assay was validated and stability was explored. Bradykinin (10-500 ng/mL) was quantified with accuracy values (% RE) below 10% and intra- and inter-day precisions (% RSD) below 12 and 16%, respectively, for all concentrations. The method was successfully applied to several plasma samples from low levels kallikrein rats (LKRs) compared with normal kallikrein rats (NKRs).     

Determination and pharmacokinetics of DT-13 in rat plasma by LC-MS

A sensitive and specific LC-MS assay for DT-13 in rat plasma was developed. DT-13 is an active steroidal saponin present in Liriopes Radix and is developed as an anti-tumor drug candidate. The samples were extracted by acetonitrile-mediated plasma protein precipitation. The chromatographic separation was carried out using a Ultimate C₁₈ column (250 mm × 4.6 mm, i.d., 5 μm) with a mobile phase composed of acetonitrile: 5 mmol/L aqueous ammonium acetate (60:40, v:v). The method was validated and the specificity, linearity (r² = 0.9980 within 10-1000 ng/mL), lower limit of quantitation (LLOQ, 10 ng/mL), precision (intra- and inter-day <12.3%), accuracy (93.4-106.3%), recovery (91.0 ± 4.7%) and stability were determined. The method was applied to the pharmacokinetic study of DT-13 in rat plasma after intravenous and intragastric administration. The results showed DT-13 underwent a prolonged absorption and slow elimination with a low oral bioavailability (5.51%) in rats.     

Development of a rapid LC-MS/MS method for ribavirin determination in rat brain

A rapid and specific liquid chromatography tandem mass spectrometry (LC-MS/MS) method for the determination of ribavirin (RBV) in rat brain was developed. Sample preparation required only two centrifuge steps before LC-MS/MS analysis and the chromatographic separation was achieved in isocratic conditions using an Atlantis T3 column with a nearly totally aqueous (95%) mobile phase. The method showed a good linearity over a concentration range of 5-1000ppb and satisfactory results in terms of accuracy.     

Determination of arbidol in human plasma by LC-ESI-MS

A sensitive, specific and accurate method for determination of arbidol in human plasma was developed. Arbidol and internal standard were extracted from plasma samples by liquid-liquid extraction with diethyl ether. The chromatographic separation was accomplished on a Shiseido C18 3 microm analytical column (100 mm x 2.0 mm i.d.) at a flow rate of 0.3 mL/min isocratically. Detection was performed on a single quadrupole mass spectrometer by selected ion monitoring (SIM) mode via electrospray ionization (ESI) source. The method had a chromatographic run time of 6 min and a good linear relationship over the range 1-1000 ng/mL. The limit of quantitation for arbidol in plasma was 1 ng/mL. The intra-day and inter-day precision (R.S.D.%) was lower than 7% and accuracy ranged from 95 to 105%. The proposed method enables unambiguous identification and quantification of arbidol in vivo and has been successfully applied to study the pharmacokinetics of arbidol in healthy male Chinese volunteers.     

液相色谱-串联质谱法测定大鼠血浆中左旋泮托拉唑钠的浓度及其毒代动力学研究

目的 建立液相色谱-串联质谱法测定大鼠血浆中左旋泮托拉唑钠的浓度并研究其毒代动力学.方法 大鼠血浆样本以乙腈沉淀蛋白后,经Chiralcel OJ-RH色谱柱(4.6 mm×150 mm,5 μm);流动相为乙腈∶水=28∶72,流速为0.6 mL·min-1,柱温为35 ℃;采用Agilent 6430三重四极杆串联质谱仪,离子化方式:电喷雾-正离子(API-ES),监测方式:多反应监测,左旋泮托拉唑监测离子对384.0/199.8,非那西丁监测离子对180.0/110.0,用作内标.结果 本法专属性良好;左旋泮托拉唑钠在50～30 000 ng·mL-1范围内线性关系符合要求;准确度均在85%～115%,精密度在15%内.SD大鼠连续4周静脉注射给予注射用左旋泮托拉唑钠低、中、高3个剂量,左旋泮托拉唑钠动力学参数AUC0～4 h增长与剂量增长均呈正相关.首次和4周给药后测定无明显差异.结论 本法经过方法学验证,适用于大鼠血浆中左旋泮托拉唑钠浓度的测定,可用于左旋泮托拉唑钠大鼠体内毒代动力学研究.     

Determination of zabofloxacin in rat plasma by liquid chromatography with mass spectrometry and its application to pharmacokinetic study

The objective of the present study was to develop a rapid and sensitive method for the determination of zabofloxacin, a novel, broad-spectrum fluoroquinolone antibiotic, in rat plasma. Rat plasma samples were deproteinized with methanol, and then were injected into an LC-MS system for quantification. Zabofloxacin and enrofloxacin, which served as an internal standard, were analyzed by selected ion monitoring (SIM) at m/z transitions of 402 for zabofloxacin and 360 for the internal standard. The lower limit of quantification (LLOQ) was determined to be 10 ng/mL, with acceptable linearity ranging from 10 to 5000 ng/mL (R>0.999). The validation parameters for zabofloxacin, such as absolute matrix effect (107.7-116.0%), accuracy (92.5-101.1% for intra-day and 90.3-103.8% for inter-day), precision (7.7-10.2% for intra-day and 4.2-8.9% for inter-day), and stability in rat plasma (96.0-101.8%), were found to be acceptable according to the assay validation guidelines of the FDA (2001). Following oral administration of zabofloxacin to rats at a dose of 20mg/kg, the concentration of zabofloxacin in plasma was quantifiable in plasma samples collected up to 8h following zabofloxacin administration. The method described in the present study is applicable to routine pharmacokinetic studies in rats.     

Determination of the cardioactive prototype LASSBio-294 and its metabolites in dog plasma by LC-MS/MS: application for a pharmacokinetic study

In this work we describe the evaluation of the pharmacokinetics of a novel cardioactive compound of the N-acylhydrazone class, LASSBio-294, using high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS) in dog plasma for the first time. Separation was achieved on a ZORBAX Rapid Resolution High Definition (RRHD) SB-C18 (50mm×2.1mm, 1.8μm) reversed-phase column at 20°C with methanol-10mM ammonium acetate solution (65:35, v/v) at a flow rate of 1.0mL/min. Detection was performed using an electrospray ionization (ESI) operating in positive ion multiple reaction monitoring (MRM) mode by monitoring the ion transitions from m/z 275.2→149.1 (LASSBio-294) and m/z 152.0→110.0 (acetaminophen, internal standard). The calibration curve of LASSBio-294 in plasma showed good linearity over the concentration range of 1.25-800ng/mL. The validated method was successfully applied to a pre-clinical pharmacokinetic study of the cardioactive prototype LASSBio-294 in beagles after oral administration. The main pharmacokinetic parameters t(1/2), C(max) and AUC(0-24) were (5.74±0.55)h, (547.66±35.12)ng/mL and (1621.77±41.66)ngh/mL, respectively.     

Polymethoxylated flavones metabolites in rat plasma after the consumption of Fructus aurantii extract: analysis by liquid chromatography/electrospray ion trap mass spectrometry

A LC with full scan MS(n) method was developed in order to investigate the in vivo absorption and biotransformation of polymethoxylated flavones (PMFs) by analysis of plasma samples from rats after ingestion of Fructus aurantii extract. Four parent compounds and six metabolites with intact flavonoid structures were tentatively identified. The metabolites were either glucuronides of parent compounds or glucuronides of demethylated products of parent compounds.     

Determination of limonin in rat plasma by liquid chromatography-electrospray mass spectrometry

A simple, sensitive and selective liquid chromatography coupled with electrospray ionization mass spectrometry (LC/ESI/MS) method for determination of limonin (LM) in rat plasma has been developed and validated. The method had advantages of a single liquid–liquid extraction with ether and high sensitivity. Analyses were conducted at a flow rate of 0.2 ml/min by a gradient elution. The detection utilized selected ion monitoring in the negative ion mode at m/z 460.00 and 423.15 for the deprotonated molecular ions of LM and the internal standard, respectively. The quantitation limit for LM in rat plasma was 1.0 ng/ml. The linearity was also excellent over the concentration range of 1.9–500 ng/ml of LM. The intra- and inter-day precision (relative standard deviation (R.S.D.%)) was lower than 10% and accuracy ranged from 90 to 110%, showing a good reproducibility. This developed method was successfully applied to analysis of LM in biological fluids.     

Simultaneous determination and pharmacokinetics of five bufadienolides in rat plasma after oral administration of Chansu extract by SPE-HPLC method

A sensitive and rapid solid-phase extraction-high performance liquid chromatography (SPE-HPLC) method has been developed for the determination of five bufadienolides, arenobufagin, teliocinobufagin, cinobufotalin, cinobufagin and resibufogenin in rat plasma and applied to a pharmacokinetic study in rats after oral administration of Chansu extract (Venenum Bufonis). Plasma samples were pretreated with solid-phase extraction using Extract-Clean cartridges, and the extracts were analyzed by a reversed-phase C(18) column on a HPLC system with photodiode array detection (DAD). The calibration curves were linear over the range of 0.10-1.66 microg/ml for arenobufagin, 0.03-1.20 microg/ml for telocinobufagin, 0.01-0.62 microg/ml for cinobufotalin, 0.03-0.70 microg/ml for cinobufagin and 0.02-2.57 microg/ml for resibufogenin, respectively. The limit of quantification was 1.1 ng/ml for arenobufagin, 0.3 ng/ml for telocinobufagin, 9.7 ng/ml for cinobufotalin, 8.8 ng/ml for cinobufagin, 7.7 ng/ml for resibufogenin, respectively. The established method could be easily applied to the determination and pharmacokinetic studies of five bufadienolides in rat plasma after oral administration of Chansu extract.     

液-质联用法测定人体血浆中万古霉素的浓度

目的:运用液-质联用技术(LC-MS/MS),建立测定人体血浆中万古霉素浓度的方法,监测万古霉素血药浓度。方法:以替洛福韦为内标,乙腈沉淀蛋白后,经LC-MS/MS方法分析。采用色谱柱为安捷伦Poroshell 120 EC-C₁₈柱(4.6 mm×100 mm,2.7 μm),流动相为混合有机相(乙腈:甲醇-2:1)-0.1%甲酸水溶液(10:90);流速0.4 mL·min^-1;采用质谱电喷雾电离源(ESI),正离子模式,多重反应监测方式(MRM)检测;待测物质万古霉素用于定量的母子离子对:m/z 725.6→m/z 144.2,内标替洛福韦用于定量的母子离子对:m/z 288.0→m/z 176.2。结果:万古霉素在0.2~100 μg·mL^-1范围内线性关系良好,最低定量限为0.2 μg·mL^-1;日内RSD均在6%以下,日间RSD均在12%以下。提取回收率为58.01%~66.58%。结论:该分析方法快速、准确、灵敏度高、专属性强、重复性好,可用于医疗机构检测人体血浆中万古霉素的药物浓度。     

液相色谱-质谱法测定人血浆中的甲哌卡因

目的建立灵敏、快速液相色谱-串联质谱法测定人血浆中甲哌卡因的浓度,并用于人体药动学研究。方法 200μL的血浆样品经甲基叔丁基醚提取处理后,以10 mmol.L-1醋酸铵(含0.5%甲酸)水溶液-乙腈(60∶40,v/v)为流动相,Inertsil CN柱分离,通过电喷雾离子源四极杆串联质谱,以多反应监测(MRM)的方式进行检测,选择离子反应为m/z 247.2→98.0(甲哌卡因)和m/z 235.2→86.0(利多卡因)。结果甲哌卡因在0.500～1 500 ng.mL-1线性关系良好,定量下限为0.500 ng.mL-1,日内、日间精密度(RSD)均≤10.1%,甲哌卡因和内标利多卡因的提取回收率分别为64.6%～67.4%和64.5%。结论该法选择性强、灵敏度高,适用于人血浆中甲哌卡因的浓度测定。     

LC-MS/MS测定大鼠血浆中虎杖苷浓度

目的 建立测定大鼠血浆中虎杖苷浓度的LC-MS/MS方法。 方法 采用LC-MS/MS测定大鼠血浆中虎杖苷的浓度,以二苯乙烯苷为内标,血浆样品用乙腈沉淀蛋白,色谱柱为Agilent Zorbax SB-C₁₈ (100 mm×2.1 mm, 3.5 μm),流动相为甲醇-乙腈-0.1%甲酸水溶液(18∶15∶67),流速0.3 ml/min,柱温30 ℃。 结果 虎杖苷的线性范围为1.0~5 000.0 ng/ml (r=0.998 4),最低定量检测浓度为1.0     

HPLC-MS/MS测定人体血浆中克拉霉素浓度的方法及方法学确证

目的建立测定人体血浆中克拉霉素浓度的高效液相色谱-串联质谱联法（HPLC-MS/MS）法。方法血浆样本用甲醇沉淀蛋白后,选用色谱柱Agilent-XDB-C8柱（2.1mm×150mm,5μm）,以甲醇（A相）：水（含0.1%甲酸）=85：15（V：V）的比例为流动相,流速为0.2ml/min,选用岛津（SHIMADZU）高效液相色谱系统,串联API3200型三重四极杆质谱仪,采用多重反应监测（MRM）扫描方式进行监测,电喷雾离子化源（ESI源）,正离子方式,用于定量分析的离子反应对分别为质荷比m/z748.4→m/z158.2（克拉霉素）和m/z515.3→m/z276.2（替米沙坦,内标）。结果本方法血浆中克拉霉素的线性范围为10～3000ng/ml（r〉0.99）,定量下限为10ng/ml,批内和批间相对标准偏差均〈15%,准确度（相对回收率）相对标准偏差也均〈15%。稳定性试验中,血浆中克拉霉素在方法学要求的各种贮存条件下均较稳定。结论该方法快速、灵敏、专属性强、重现性好,适用于人体内克拉霉素的药代动力学研究。     

LC-MS/MS法测定人体内的血浆中蝙蝠葛碱质量浓度

目的建立人血浆中蝙蝠葛碱质量浓度的测定方法。方法采用LC-MS/MS。色谱柱为Agi-lent TC-C18柱(150 mm×4.6 mm,5μm),流动相为甲醇-水-冰醋酸(体积比为60∶40∶0.8);流速700μL.min-1;柱温30℃;进样量20μL。结果血浆中内源性物质不干扰蝙蝠葛碱和内标原阿片碱的测定,蝙蝠葛碱的线性范围为1.0～200μg.L-1,血浆中平均回收率为65.3%～73.4%,蝙蝠葛碱的定量下限为1.0μg.L-1。结论 LC-MS/MS方法简单,快速,专属性强,灵敏度高,可用于人血浆中蝙蝠葛碱质量浓度的测定。     

LC-MS／MS法测定人血浆中舒芬太尼的血药浓度

目的建立液质联用（LC—MS／MS）法测定血浆中舒芬太尼血药浓度。方法以芬太尼为内标,血浆样品经乙腈沉淀蛋白后,以Ultimate XB C18（100mm×2．1mm,3．0μm）柱为色谱柱,流动相为水-甲醇（15：85,V／V）,各含10mmol·L^-1乙酸铵,流速为0．2mL·min^-1,柱温加℃;质谱条件为电喷雾电离源（ESI）,检测方式为正离子电离、多离子反应监测（MIIM）,用于定量分析的离子为舒芬太尼m／z387．2→238．1,芬太尼m／z337．2→188．2。结果舒芬太尼线性范围为0．05～2μg·L^-1（r=0．9964）,线性关系良好。舒芬太尼的提取回收率为83．68％。88．06％。批内和批间精密度RSD均〈10％。结论本研究建立的测定舒芬太尼血药浓度的方法简单、快速、准确、灵敏,可用于临床上血药浓度监测和药动学研究。