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.     

液相色谱-质谱联用法测定大鼠血浆中木通皂苷D的浓度

目的建立液相色谱-质谱联用法（LC-MS/MS）测定大鼠血浆中木通皂苷D的浓度。方法选用XDB-C18色谱柱,以5 mmol/L乙酸铵水溶液（含0.1%甲酸）-甲醇溶液为流动相,采用梯度洗脱进行分离。样本经甲醇沉淀后进样,选用3200QTRAP型质谱仪的多重反应监测扫描方式进行检测。结果木通皂苷D线性范围为10～1000 ng/ml,最低定量限为10ng/ml。准确度与精密度结果显示方法日间、日内变异均小于15%,相对误差为-2.8%～4.6%,低、中、高3个浓度提取回收率为95.3%～108.1%。结论本研究所建立的方法快速、灵敏、专属性强、重现性好,可用于大鼠血浆中木通皂苷D浓度的测定和药代动力学研究。     

LC-MS/MS法测定大鼠血浆中吴茱萸碱的浓度及其药代动力学研究

目的：建立用于测定吴茱萸碱血药浓度的液相色谱-串联质谱联用分析方法,并研究吴茱萸碱在大鼠体内的药代动力学。方法：6只大鼠灌胃给药吴茱萸碱100mg/kg,眼底取血,LC-MS/MS法测定血药浓度,并用DAS药代动力学程序拟合计算药代动力学参数。结果：吴茱萸碱浓度在0.2～50ng/mL内,线性关系良好（r^2=0.9997）。提取回收率96.12%～99.46%,日内、日间RSD分别为4.61%～13.51%和5.65%～11.49%。主要药代动力学参数为：Cmax=（5.3±1.5）ng/mL;tmax=（22±8）min;t1/2=（451±176）min。结论：建立的LC-MS/MS方法专属性强,灵敏度高,可用于吴茱萸碱的体内定量分析。     

Rapid and simultaneous measurement of midazolam, 1'-hydroxymidazolam and digoxin by liquid chromatography/tandem mass spectrometry: application to an in vivo study to simultaneously measure P-glycoprotein and cytochrome P450 3A activity

In order to simultaneously determine in vivo P-glycoprotein (P-gp) and Cytochrome P450 3A (CYP3A) activity, a new, rapid and sensitive liquid chromatography/tandem mass spectrometry (LC-MS/MS) method has been developed and fully validated to simultaneously determine midazolam (MDZ, as CYP3A substrate), 1'-hydroxymidazolam (1'-OHMDZ) and digoxin (DG, as P-gp substrate) in rat plasma using digitoxin as the internal standard (IS). After a single step liquid-liquid extraction with tert-butyl methyl ether/dichloromethane (75:25, v/v), analytes were subjected to LC-MS/MS analysis using positive electro-spray ionization (ESI(+)) under selected reaction monitoring mode (SRM). Chromatographic separation was performed on an XTerra MS C18 column (50mm×2.1mm, i.d. 3.5μm). The MS/MS detection was conducted by monitoring the fragmentation of 326.05 → 244.00 (m/z) for MDZ, 342.02 →168.01 (m/z) for 1'-OHMDZ, 798.33 → 651.36(m/z) for DG and 782.67 → 635.24 (m/z) for IS. The method had a chromatographic running time of 3min and linear calibration curves over the concentrations of 2-400ng/mL for MDZ and 1'-OHMDZ and 0.5-100ng/mL for DG. The recoveries of the method were 86.8-96.3% for MDZ, 84.6-86.4% for 1'-OH MDZ, and 81.7-85.1% for DG. The lower limit of quantification (LLOQ) of the method was 2ng/mL for MDZ and 1'-OHMDZ and 0.5ng/mL for DG. The intra- and inter-batch precision were less than 15% for all quality control samples at concentrations of 5, 50 and 320ng/mL for MDZ and 1'-OHMDZ and 1, 10 and 80ng/mL for DG. The validated LC-MS/MS method has been successfully used to analyze the concentrations of MDZ, 1'-OH MDZ and DG in rat plasma for simultaneous measurement of in vivo P-gp and CYP 3A activity.     

液相色谱-串联质谱法测定人血浆中多潘立酮浓度

目的建立高效液相色谱-串联质谱测定人血浆中多潘立酮(第2代胃动力药)浓度的方法。方法用TC-C18色谱柱,流动相为甲醇(含0.025%甲酸)-1 mmol.L-1甲酸铵溶液(含0.05%甲酸),用梯度洗脱进行分离,流速为1.0 mL.min-1。用正离子化模式,多重反应监测(MRM)扫描方式进行检测定量。结果多潘立酮的线性范围为0.1～50.0 ng.mL-1,日内、日间精密度均小于15%,提取回收率大于80%。结论本方法灵敏、准确、快速,可用于人血浆中多潘立酮浓度的测定和药代动力学研究。     

LC-MS/MS 法测定人血浆中倍他米松

建立测定人血浆中倍他米松的LC-MS/MS方法。采用Venusil XBP C₈ (200 mm×3.9 mm ID, 5 μm)色谱柱，流动相为甲醇-水(含甲酸铵5 mmol·L^-1)(80∶20)，流速0.4 mL·min^-1；质谱仪离子源为电喷雾离子源(ESI)，正离子模式检测，监测离子为393.3→355.2(倍他米松)和361.3→343.2(泼尼松龙,内标)。血浆样本用乙酸乙酯处理。倍他米松在0.5~80.0 ng·mL^-1线性关系良好(r=0.999 2)， 血浆低、 中、 高3种浓度(1.0， 10.0， 60.0 ng·mL^-1)平均提取回收率为88.24%，定量限为0.5 ng·mL^-1。本方法操作简便、准确、灵敏，适用于复方倍他米松注射液人体药代动力学研究。     

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).     

Evaluation of the reinforcing properties and phencyclidine-like discriminative stimulus effects of dextromethorphan and dextrorphan in rats and rhesus monkeys

Rationale: Dextromethorphan (DXM) and its metabolite, dextrorphan (DXO) have neuroprotective and anticonvulsant properties through their activity as N-methyl-d-aspartate (NMDA) receptor channel blockers. Based on this receptor activity, coupled with reports of DXM abuse, both were evaluated for abuse potential and phencyclidine (PCP)-like behavioral effects in two animal models. Objectives and methods: The discriminative stimulus properties of DXO and DXM were tested in rats (3–56 mg/kg DXM, i.p. and 2.2–40.9 mg/kg DXO, i.p.) and rhesus monkeys (0.3–10 mg/kg DXM, i.m. and 0.25–8.0 mg/kg DXO, i.m.) trained to discriminate PCP from saline using a standard two-lever drug-discrimination paradigm under a fixed-ratio (FR) schedule of food reinforcement. In a second set of experiments, i.v. self-administration of DXO (10–100 μg/kg/infusion) and DXM (10–1000 μg/kg/infusion) were tested under a FR schedule of reinforcement in monkeys trained to lever press for infusions of PCP during daily 1-h sessions. Results: In rats, both DXM and DXO produced a dose-dependent substitution for PCP. When tested in monkeys, DXM yielded partial (1 monkey) and full (2 monkeys) substitution for PCP, while DXO substituted fully for PCP in all four subjects tested. In the self-administration study, in five of the six subjects, at least one dose of DXM served as a positive reinforcer, maintaining infusion rates above those for saline. For DXO, at least one dose maintained infusion numbers well above mean saline infusion numbers in all subjects. Conclusions: Taken together, these data show that DXM has some PCP-like effects in rats and monkeys, but that they are more reliably produced by its metabolite, DXO. Thus, high doses of DXM may have some PCP-like abuse potential in humans but this potential may be associated with, or enhanced by, metabolism of DXM to DXO. Received: 30 November 1998 / Final version: 25 March 1999     

LC-MS/MS法测定大鼠血浆中的利福布丁

目的用LC-MS/MS法测定大鼠血浆中利福布丁的浓度。方法以替米沙坦为内标,加入0.2 mL血浆,用1 mL乙酸乙酯萃取,高速离心后上清液氮气吹干并用流动相复溶,取20μL进样检测。结果利福布丁的最低检测限为0.5 ng.mL-1,在0.3～9.6μg.mL-1内线性良好,低、中、高浓度的提取回收率均大于85%,批内、批间RSD低于10%,成功检测了大鼠血浆中利福布丁的血药浓度。结论所用方法灵敏度好、准确度高、分析时间短、样品处理更简便,适用于测定利福布丁的血药浓度。     

Determination of free and total cortisol in plasma and urine by liquid chromatography-tandem mass spectrometry

Cortisol is an important adrenal steroid hormone involved in the regulation of metabolic homeostasis. A new liquid chromatography-mass spectrometry/mass spectrometry (LC-MS/MS) multiple reactant monitoring (MRM) procedure for the measurement of cortisol concentration in plasma ultrafiltrate, whole plasma, and urine was developed and validated. Plasma, plasma ultrafiltrate, or urine was extracted by ethyl acetate. The extract was subjected to liquid chromatography with an Inertsil ODS-3 column with an aqueous NH4Cl (1 mM, pH 9.0):methanol mobile phase. The presence of NH4Cl in the mobile phase induced the formation of [M+Cl] in the first quadrupole at m/z 397 and 409 for cortisol and 6alpha-methylprednisolone (internal standard), respectively. In the collision cell, the complex dissociated to the neutral parent and the chloride ion at m/z 35; the latter ion was used for quantification. The calibration curve was linear from 0.5 to 100 ng/mL. The lower limit of quantification was 0.50 ng/mL and the limit of detection was 0.25 ng/mL. For quality control samples prepared in water, the intrabatch assay precision was 5.6%, 9.6%, and 9.9% at 50, 10, and 1 ng/mL, respectively. The interbatch assay precision was 4.2%, 6.3%, and 7.5% at 50, 10, and 1 ng/mL, respectively. For measurement of endogenous cortisol in plasma and urine samples, the intra-assay and interassay precision was 10.8% and 4.8% for total plasma cortisol, 13.1% and 5.2% for free plasma cortisol, 10.9% and 13.1% for cortisol protein-binding free fraction, and 8.9% and 14.4% for urine cortisol, respectively. A simple procedure of ultrafiltration coupled with the highly sensitive LC-MS/MS quantification offered a rapid and reproducible assay for plasma free cortisol, which may be useful in the assessment of adrenal function in patients, especially critically ill patients with abnormal protein binding. It may also be useful for plasma and urinary cortisol measurements in pharmacodynamic studies of adrenocorticoid response.     

Liquid chromatographic tandem mass spectrometry method for the quantification of miglitol in human plasma

A rapid, sensitive and accurate liquid chromatography-tandem mass spectrometry (LC-MS/MS) method was developed and validated for the quantification of miglitol (CAS 72432-03-2), an alpha-glucosidase inhibitor, in human plasma using gabapentin (CAS 60142-96-3) as internal standard (IS). Following protein precipitation, the analytes were separated using an isocratic mobile phase on a reversed phase phenyl column and analyzed by MS in the multiple reaction monitoring mode using the respective [M+H]+ ions, m/z 208/146 for miglitol and m/z 172/154 for the IS. The assay exhibited a linear dynamic range of 100-6000 ng/mL for miglitol in human plasma. The lower limit of quantification was 100 ng/mL with a relative standard deviation of less than 5 %. Acceptable precision and accuracy were obtained for concentrations over the standard curve ranges. The average absolute recoveries of miglitol and the IS from spiked plasma samples were 40.5 +/- 2.7 and 47.1 +/- 2.9 %, respectively. A run time of 2.5 min for each sample made it possible to analyze a throughput of more than 400 human plasma samples per day. The validated method has been successfully used to analyze human plasma samples for application in pharmacokinetic, bioavailability or bioequivalence studies. The miglitol plasma concentration profile could be obtained for pharmacokinetic study. The observed maximum plasma concentration (Cmax) of miglitol (100 mg oral dose) is 1740 ng/mL, time to observed maximum plasma concentration (tmax) is 3.5 h and elimination half-life (t(1/2)) is 2.5 h.     

Liquid chromatography-tandem mass spectrometry assay for the quantitation of plagiochin E and its main metabolite plagiochin E glucuronides in rat plasma

Plagiochin E, a macrocyclic bisbibenzyl isolated from liverwort Marchantia polymorpha, was found to have antifungal activity. To evaluate the pharmacokinetics of plagiochin E in rats, a sensitive and specific liquid chromatography/tandem mass spectrometric (LC-MS/MS) method was developed and validated for the quantitation of plagiochin E and its total conjugated metabolites in rat plasma. For detection, a Sciex API 4000 LC-MS/MS with a TurboIonSpray ionization (ESI) inlet in the negative ion-multiple reaction monitoring (MRM) mode was used. The plasma samples were pretreated by a simple liquid-liquid extraction with ethyl acetate. The concentration of plagiochin E parent form was determined directly and the concentration of plagiochin E conjugated metabolites was assayed in the form of plagiochin E after treatment with beta-glucuronidase/sulfatase. The statistical evaluation for this method reveals excellent linearity, accuracy and precision for the range of concentrations 0.5-1000.0 ng/mL. The method had a lower limit of quantification (LLOQ) of 0.5 ng/mL for plagiochin E in 50 microL of plasma. The method was successfully applied to the characterization of the pharmacokinetic profile of plagiochin E in rats after an oral and an intravenous administration.     

Quantitative determination of total methamphetamine and active metabolites in rat tissue by liquid chromatography with tandem mass spectrometric detection

High-throughput liquid chromatography with tandem mass spectrometric detection (LC-MS/MS) methodology for the determination of methamphetamine (METH), amphetamine (AMP), 4-hydroxymethamphetamine (4-OH-METH), and 4-hydroxyamphetamine (4-OH-AMP) was developed and validated using simple trichloroacetic acid sample treatment. The method was validated in rat serum, brain, and testis. Lower limits-of-quantitation (LOQ) for METH and AMP were 1 ng x mL(-1) using positive ion electrospray tandem mass spectrometry (MS/MS). The accuracy of the method was within 25% of the actual values over a wide range of analyte concentrations. The within-assay precision was better than 12% (coefficient of variation). The method was linear over a wide dynamic range (0.3-1000 ng x mL(-1)). Quantitation was possible in all 3 matrices using only serum standards because of minimal matrix-associated ion effects or the use of an internal standard. Finally, the LC-MS/MS method was used to determine serum, brain, and testis METH and AMP concentrations during a subcutaneous infusion (5.6 mg kg(-1) day(-1)) of METH in rats. Concentrations of 4-OH-AMP and 4-OH-METH were below the LOQ in experimental samples. The bias introduced by using serum calibrators for the determination of METH and AMP concentrations in testis and brain was less than 8% and insignificant relative to the interanimal variability.     

Pharmacokinetics of gastrodin and its metabolite p-hydroxybenzyl alcohol in rat blood, brain and bile by microdialysis coupled to LC-MS/MS

Gastrodin is a pharmacologically active substance isolated from Gastrodia elata Blume with sedation, anti-convulsion and anti-epilepsy activities. A rapid and sensitive liquid chromatography technique coupled to tandem mass spectrometry (LC-MS/MS) system was developed to determine gastrodin and its metabolite p-hydroxybenzyl alcohol (HBA) in rat blood, brain and bile collected using microdialysis technique. The analytes were separated using a reversed phase column (4.6 mm x 150 mm, 5 microm). The mobile phase for column separation was 30% methanol with a flow rate of 0.6 mL/min. As a post-column addition, 1% ammonium hydroxide solution (in methanol) was additionally pumped via a T-connection using a chromatographic pump (BAS PM-80, USA) at a flow rate of 0.2 mL/min after the column separation. A LC-MS/MS system equipped with a negative electrospray ionization (ESI) source in multiple reaction monitoring (MRM) mode was used to monitor m/z 285.0-->122.9 and m/z 123.0-->105.0 transitions for gastrodin and HBA, respectively. The lower limit of quantification (LLoQ) for gastrodin and HBA were 0.5 and 2 ng/mL, respectively. The calibration curves were linear over the range of 0.5-5,000 ng/mL and 2-1,000 ng/mL for gastrodin and HBA with a coefficient of determination >0.995, respectively. This selective and sensitive method is useful for the determination of gastrodin and HBA and in the pharmacokinetic studies of these compounds.     

Specific,sensitive and accurate LC-MS/MS method for the measurement of levovirin in rat and monkey plasma

Levovirin is a guanosine nucleoside analogue and the L-enantiomer of ribavirin. Levovirin has a better safety profile than ribavirin, exerts similar immunomodulatory effects in a mouse efficacy model, and may provide a better therapeutic option than ribavirin in patients with chronic hepatitis C virus (HCV) infection. To facilitate pharmacokinetic studies, a LC-MS/MS method for the analysis of levovirin in rat and monkey plasma was developed and validated. The method involved adding ICN 10537 as an internal standard, protein precipitation with acetonitrile followed by separation on an Intersil Silica column, and quantification by a MS/MS system equipped with positive electrospray ionization (ESI) in the multiple reaction monitoring (MRM) mode. The MS/MS reaction was selected to monitor the 245-->113 and 259-->128 transitions for levovirin and internal standard, respectively. The calibration curve was linear over a concentration range of 10-5000 ng/ml. The limit of quantitation was 10 ng/ml, the coefficient of variation (CV) was 3-5%, and the bias was 3-6%. Intra- and inter-day analysis of QC samples at 30, 1500 and 3500 ng/ml indicated that the method was precise (CV<6%) and accurate (bias <9%). Levovirin in rat and monkey plasma was stable at 5 degrees C for at least 24 h, 0 degrees C for at least 4 h, and after three freeze-thaw cycles. This specific, accurate and precise assay is useful in the study the pharmacokinetic characteristics of this compound.     

Development and validation of a highly sensitive and robust LC-MS/MS with electrospray ionization method for quantification of rosuvastatin in small volume human plasma samples and its application to a clinical study

A high-throughput, simple, highly sensitive and specific LC-MS/MS method (liquid chromatography coupled with tandem mass spectrometry) has been developed for the estimation of rosuvastatin (CAS 287714-41-4, RST) with 100 microl human plasma using atorvastatin (CAS 134523-00-5) as an internal standard (IS). The API-4000 LC-MS/MS was operated under the multiple reaction-monitoring mode (MRM) using the electro spray ionization technique. The assay procedure involved direct precipitation of RST and IS from plasma with acetonitrile. Sample preparation with this method yielded clean extracts and consistent recoveries: 91.39% for RST and 99.28% for IS. The total chromatographic run time was 3.5 min and the elution of RST and IS occurred at 2.5 and 3.1 min, respectively; this was achieved with a mobile phase consisting of 0.05 mol/L formic acid: acetonitrile (20:80, v/v) at a flow rate of 0.50 ml/min on an Inertsil ODS-3 column (4.6 x 100 mm, 3.0 microm). The developed method was validated in human plasma with a limit of quantitation of 0.05 ng/ml. A linear response function was established for the range of concentrations of 0.05 to 50.0 ng/ml with a correlation coefficient (r) of 0.999. The inter- and intra-day precision in the measurement of RST quality control (QC) samples at 0.05, 0.15, 25 and 40 ng/ml were in the range of 6.55 to 11.40% relative standard deviation (RSD) and 1.76 to 11.17% RSD, respectively. Accuracy in the measurement of QC samples for RST was in the range of 95.02 to 101.37% of the nominal values. RST was stable in the battery of stability studies viz., bench-top, auto-sampler and freeze-thaw cycles. The stability of RST was established for 1 month at -80 degrees C. The application of the assay to a clinical study confirmed the utility of the assay to derive human pharmacokinetic parameters.     

Liquid chromatographic tandem mass spectrometric assay for simultaneous quantification of compound 97/78 and its in vivo metabolite 97/63, a novel trioxane antimalarial, in human plasma and its application to a protein binding study

A sensitive, selective and specific LC-MS/ MS assay for simultaneous quantification of compound 97/78 and its active in vivo metabolite 97/63, a novel 1,2,4-trioxane antimalarial, in human plasma has been developed and validated using alpha-arteether as internal standard (IS). Extraction from plasma involves a simple protein precipitation method. The analytes were chromatographed on a Columbus C18 column with guard by isocratic elution with acetonitrile:ammonium acetate buffer (10 mM, pH 4.0) (80:20 v/v) as mobile phase at a flow rate of 0.45 mL min(-1) and analyzed in multiple reaction-monitoring (MRM) positive ion mode. The chromatographic run time was 4.0 min. The weighted (1/x2) calibration curves were linear over a range of 1.56-200 ng mL(-1) with correlation coefficients > 0.998. For both analytes, the limit of detection (LOD) and lower limit of quantification (LLOQ) were 0.5 ng mL(-1) and 1.56 ng mL(-1), respectively. The recovery of 97/78, 97/63 and IS from spiked control samples were > 90% and their matrix suppression obtained were < 8 %. The accuracy (% bias) and precision (%RSD) for both analytes were < 6.78%. Both analytes were stable after three freeze-thaw cycles (% deviation < 12.80), long-term for 30 days in plasma at -60 degrees C (% deviation < 14.38), for 8 h on bench top in plasma at ambient temperature (% deviation < 1.52) and also in the auto-sampler for 12 h (% deviation < 3.9%). The validated method was successfully applied to a protein binding study of compound 97/78 and metabolite 97/63 in human plasma. Furthermore, the validated method will be applicable to pharmacokinetics, bioavailability and metabolism in various clinical phases and in drug interaction studies.     

Establishment of a liquid chromatographic/mass spectrometry method for quantification of tetrandrine in rat plasma and its application to pharmacokinetic study

A rapid and sensitive liquid chromatography-tandem mass spectrometric method (LC/MS/MS) for the determination of tetrandrine in rat plasma has been developed, fully validated and successfully applied to pharmacokinetic study in Sprague-Dawley (SD) rats after a single oral administration. Sample preparation involves a liquid-liquid extraction with n-hexane-dichlormethane (65:35, containing 1% 2-propanol isopropyl alcohol, v/v). Tetrandrine and brodimoprim (internal standard) were well separated by LC with a Dikma C(18) column using acetonitrile-methanol-ammonium formate aqueous solution (20mM) containing 0.3% formic acid (20:30:50, v/v/v) as mobile phase. Detection was performed on a triple quadrupole mass spectrometer in multiple reaction monitoring mode. The ionization was optimized using ESI(+) and selectivity was achieved using MS/MS analysis, m/z 623.0-->381.0 and m/z 339.0-->281.0 for tetrandrine and I.S., respectively. The present method exhibited good linearity over the concentration range of 5-2,000 ng/mL for tetrandrine in rat plasma with a lower limit of quantification of 5 ng/mL. The intra- and inter-day precision were 2.0-9.2% and 4.5-9.4%, and the intra- and inter-day accuracy ranged from -7.6 to 10.3% and -6.0 to 5.3%, respectively. No endogenous compounds were found to interfere with the analysis, and tetrandrine was stable during the whole assay period. The method was successfully applied to a pharmacokinetic study after an intragastric administration (i.g.) of tetrandrine to SD rats with a single dose of 50mg/kg. The results confirm that the assay is suitable for the pharmacokinetic study of tetrandrine.     

Pharmacokinetic study of the novel, synthetic trioxane antimalarial compound 97-78 in rats using an LC-MS/MS method for quantification

The present study has been designed to investigate the pharmacokinetic parameters of the novel trioxane antimalarial 97-78 (US Patent 6316493 B1, 2001) in male and female rats after single oral and intravenous administration. The pharmacokinetic profile of 97-78 was investigated in the form of its completely converted metabolite 97-63 after dose administration. Quantification of metabolite 97-63 in rat plasma was achieved using a simple and rapid LC-MS/MS method. The LC-MS/MS method has been validated in terms of accuracy, precision, sensitivity and recovery for metabolite 97-63 in rat plasma. The intra- and interday accuracy (% bias) and precision (% RSD) values of the assay were less than 10% for metabolite 97-63. The chromatographic run time was 4.0 min and the weighted (1/x2) calibration curves were linear over the range 1.56-200 ng/ml. This method was successfully applied for analysis of pharmacokinetic study samples. Maximum plasma concentrations of 97-63 at 47 mg/kg oral administration in male and female rats were 1986.6 ng/ml and 4086.7 ng/ml at time (Tmax) 0.92 h and 0.58 h, respectively. The area under the curve (AUC(0-infinity)), elimination half-life (t(1/2) beta) and mean residence time (MRT) were 4669.98 ng x h/ml, 2.8 h and 4.2 h in male and 11786.0 ng x h/ml, 4.52 h and 4.32 h in female rats respectively. After single oral and intravenous administration of 97-78 to male and female rats significant differences were observed in pharmacokinetic parameters (AUC and t (1/2) beta) for metabolite 97-63.     

Validation of rapid and simple LC-MS/MS method for determination of voriconazole in rat plasma

A rapid, simple and sensitive LC-MS/MS analytical method was developed and validated for the determination of voriconazole (VRC) in rat plasma, using ketoconazole as internal standard (IS). Analysis was performed on a Shimadzu HPLC system using a Shimadzu C18 column and isocratic elution with acetonitrile-water-formic acid (60:40:0.05, v/v/v), at a flow of 1.0 mL/min (split ratio 1:5), and a mass spectrometer Micromass, equipped with a double quadrupole and an electrospray ionization interface, operated in a positive mode. Plasma samples were deproteinized with methanol (1:2) and 30 microL of the supernatant was injected into the system. The retention times of VRC and IS were approximately 3.3 and 2.7 min, respectively. Calibration curves in spiked plasma were linear over the concentration range of 50-2500 ng/mL with determination coefficient >0.98. The lower limit of quantification was 50 ng/mL. The accuracy of the method was within 5%. Intra- and inter-day relative standard deviations were less or equal to 12.5 and 7.7%, respectively. The applicability of the LC-MS-MS method for pharmacokinetic studies was tested using plasma samples obtained after intravenous administration of VRC to male Wistar rats. The reported method provided the necessary sensitivity, linearity, precision, accuracy, and specificity to allow the determination of VRC in pre-clinical pharmacokinetic studies.