Determination of acteoside in Cistanche deserticola and Boschniakia rossica and its pharmacokinetics in freely-moving rats using LC-MS/MS

A sensitive LC-MS/MS method with a simple solid-phase extraction for the determination of acteoside in rat plasma and tissue homogenates was established for the investigation of bioavailability and brain distribution in freely-moving rats. Acteoside in Cistanche deserticola and Boschniakia rossica was also determined. Acteoside and internal standard were separated on a RP-select B column (125mmx4.6mm i.d., particle size 5microm). The mobile phase consisted of 35% methanol and 65% acetic acid-water (1:100, v/v) at a flow-rate of 1mL/min. Acteoside and the internal standard were monitored using the multiple-reaction monitoring (MRM) mode at m/z transitions of 623-->161 and 609-->301, respectively. The acteoside content was 38.4+/-2.4mg/kg (n=3) for B. rossica, which is obviously lower than 21134.2+/-805.5mg/kg (n=3) of C. deserticola. The protein binding in rat plasma was 75.5+/-1.8%. The brain distribution result indicated that acteoside was evenly distributed in brain tissues (brain stem, cerebellum, the rest of the brain, cortex, hippocampus and striatum) which was about 0.45-0.68% of that in plasma (4.5+/-0.5microg/mL) after 15min of acteoside administration (10mg/kg, i.v.). After acteoside was given (3mg/kg, i.v.; 100mg/kg, p.o.), the oral bioavailability (AUC(p.o.)/dose(p.o.))/(AUC(i.v.)/dose(i.v.)) was only 0.12%.     

顺式藁苯内酯口服给药在大鼠血浆和脑内的药动学特性(英文)

建立大鼠血浆和脑中Z-槀苯内酯(LIG)浓度测定的高效液相色谱法。采用Agilent Hypersil ODS C18色谱柱(150mm×4.6mm,5μm),流动相为甲醇-5%异丙醇水溶液(60:40,v/v),流速为1.0mL/min,检测波长为280nm。血浆与脑中槀苯内酯浓度线性检测范围分别为93.75~3750ng/m(r=0.9999)和93.75~3750ng/g(r=0.9997),日内及日间精密度RSD10%。本法适用于大鼠口服LIG后血浆及脑中药物浓度的研究。     

Development and validation of a sensitive liquid chromatography-tandem mass spectrometry method for the determination of paeoniflorin in rat brain and its application to pharmacokinetic study

A sensitive and specific method was developed and validated for the determination of paeoniflorin in rat brain with liquid chromatography-tandem mass spectrometry. Sample pretreatment involved protein precipitation following solid-phase extraction. Paeoniflorin and geniposide (internal standard) were separated isocratically on a Waters Symmetry C18 column (150 mm x 2.1 mm i.d., 5 microm), using a mobile phase of methanol/water with 0.1% formic acid (50:50, v/v) at a flow-rate of 200-300 microL/min in 4min. A Finngan LTQ tandem mass spectrometer equipped with electrospray ionization source was operated in the positive ion mode. Selective reaction monitoring was performed to quantify paeoniflorin and the internal standard at m/z transitions of 503-->381 and 411-->231, respectively. A good linearity was found in the range of 2-500 ng/mL (R(2)=0.9939). The intra- and inter-batch assay precisions (coefficient of variation, CV) at 5, 50 and 400 ng/mL (n=5) ranged from 6.3% to 9.7% and 1.2% to 7.2%, respectively, and the accuracies were from 95.9% to 101.6% and 99.4% to 102.9%, respectively. The mean recoveries of paeoniflorin were 81.2%, 80.9% and 82.3% at 5, 50 and 400 ng/mL (n=5), respectively, and the mean recovery of the internal standard was 76.7% with a concentration of 50 ng/mL (n=5). Stability studies showed that paeoniflorin was stable in different conditions. Finally, the method was successfully applied to the pharmacokinetic study of paeoniflorin in rat brain following a single subcutaneous administration (10 mg/kg) to rats.     

Development and validation of a highly sensitive and robust LC-MS/MS with electrospray ionization method for simultaneous quantitation of itraconazole and hydroxyitraconazole in human plasma: application to a bioequivalence study

A highly sensitive and specific LC-MS/MS method has been developed for simultaneous estimation of itraconazole (ITZ) and hydroxyitraconazole (OH-ITZ) with 500 microL of human plasma using fluconazole as an internal standard (IS). The API-4000 LC-MS/MS was operated under the multiple reaction-monitoring mode (MRM) using the electrospray ionization technique. Solid phase extraction process was used to extract ITZ, OH-ITZ and IS from human plasma. The total run time was 3.0 min and the elution of ITZ, OH-ITZ and IS occurred at 2.08 min, 1.85 min and 1.29 min, respectively; this was achieved with a mobile phase consisting of 0.2% (v/v) ammonia solution:acetonitrile (20:80, v/v) at a flow rate of 0.50 mL/min on a HyPurity C(18) (50 mm x 4.6 mm, 5 microm) column. The developed method was validated in human plasma with a lower limit of quantitation of 0.50 ng/mL for both ITZ and OH-ITZ. A linear response function was established for the range of concentrations 0.5-263 ng/mL (r>0.998) for both ITZ and OH-ITZ. The intra- and inter-day precision values for ITZ and OH-ITZ met the acceptance as per FDA guidelines. ITZ and OH-ITZ were stable in the battery of stability studies, viz., bench-top, auto-sampler, dry extract and freeze/thaw cycles. The developed assay method was applied to an oral bioequivalence study in humans.     

HPLC-UV method development for fentanyl determination in rat plasma and its application to elucidate pharmacokinetic behavior after i.p. administration to rats

A simple, rapid and validated high performance liquid chromatography method with UV detection for the quantification of an opioid agonist, fentanyl (FEN), in rat plasma was developed. The assay procedure involved chromatographic separation using a ZIC-HILIC SeQUANT column (250 mm × 4.6 mm, i.d., 5 μm) and a mobile phase of acetonitrile and acetate buffer (pH 3.4, 20mM) of ratio (=65:35, v/v) at a flow rate of 1.2 mL/min and detection wavelength of 201 nm. Plasma sample (100 μL) pretreatment was based on simple deprotienization by acetonitrile spiked with clonidine as an internal standard (I.S.) of 20 ng/mL followed by extraction with tert-butyl methyl ether and centrifugation. The organic layer was evaporated under N(2) gas and reconstituted with 100 μL of acetate buffer (pH 3.4, 20mM), and 50-μL portions of reconstituted sample were injected onto the column. Sample analysis including sample pretreatment was achieved within 35 min. Calibration curve was linear (r ≥ 0.998) from 5 to 100 ng/mL. Both intra- and inter-day assay precisions that are presented through RSD were lower than 12.6% for intra-day and lower than 12.0% for inter-day assessment. Limit of detection was 0.8 ng/mL at S/N of 3. This method was omitting the use of expensive solid phase extraction and time consuming liquid extraction procedures. Moreover, the present method was successfully applied to study pharmacokinetic parameters of FEN after intraperitoneal administration to male Wistar rat. Pharmacokinetic parameters estimated by using moment analysis were T(1/2) 198.3 ± 44.7 min, T(max) 28.3 ± 2.9 min and AUC(0-180) 15.6 ± 2.9(× 10(2))ngmin/mL.     

Development of a liquid chromatography/tandem mass spectrometry assay for the determination of bestatin in rat plasma and its application to a pharmacokinetic study

Bestatin is a low molecular weight aminopeptidase inhibitor originally isolated from culture filtrates of Streptomyces olivoreticuli. We have developed a sensitive, specific liquid chromatography-tandem mass spectrometry (LC-MS/MS) for the quantitative determination of bestatin in rat plasma using granisetron as the internal standard. The analyte and internal standard were isolated from 50 microL plasma samples by solid phase extraction (SPE). Reverse-phase HPLC separation was accomplished on a Lichrospher C18 column (4.6 mm x 50 mm, 5 microm) with a mobile phase composed of methanol-water-formic acid (70:30:0.5, v/v/v) at a flow rate of 0.8 mL/min. The method had a chromatographic total run time of 3 min. A Varian 1200L electrospray tandem mass spectrometer equipped with an electrospray ionization source was operated in selected reaction monitoring (SRM) mode with the precursor-to-product ion transitions m/z 309.2-->120.0 (bestatin) and 313.4-->138.0 (granisetron) used for quantitation. The method was sensitive with a lower limit of quantitation (LLOQ) of 5 ng/mL, with good linearity (r2 >or= 0.999) over the linear range of 5-2000 ng/mL. All the validation data, such as accuracy, precision, and inter-day repeatability, were within the required limits. The method was successfully applied to pharmacokinetic study of bestatin in rats.     

Felodipine quantification in human plasma by high-performance liquid chromatography coupled to tandem mass spectrometry

A rapid, sensitive, robust and specific method was developed for the determination and quantitation of felodipine, in human blood plasma by liquid chromatography coupled with tandem mass spectrometry using nimodipine as internal standard. Felodipine was extracted from 0.5 mL human plasma by use of a liquid/liquid procedure using diethyl ether/hexane (80/20, v/v) as eluent. The method included a chromatographic run of 5 min using a C(18) analytical column (100 mm x 4.6 mm i.d.) and the calibration curve was linear over the range from 0.02 to 10 ng mL(-1) (r(2) > 0.994). The between-run precision, determined as relative standard deviation of replicate quality controls, was 5.7% (0.06 ng mL(-1)), 7.1% (0.6 ng mL(-1)) and 6.8% (7.5 ng mL(-1)). The between-run accuracy was +/- 0.0, 2.1 and 3.1% for the above-mentioned concentrations, respectively.     

Simultaneous blood and brain sampling of cephalexin in the rat by microdialysis and microbore liquid chromatography: application to pharmacokinetics studies

To circumvent the need for laborious sample clean-up and multiple blood sampling, a system was developed consisting of on-line microdialysis coupled to microbore liquid chromatography and ultraviolet detection. The system was designed for the simultaneous and continuous monitoring of unbound blood and brain cephalexin in the rat following single bolus intravenous administrations (10 mg/kg, n=6). Microdialysis probes were inserted into the jugular vein and brain striatum, respectively, for blood and brain sampling. Chromatographic conditions consisted of a mobile phase of methanol–100 mM monosodium phosphoric acid (20:80, v/v, pH 5.0) pumped through a microbore reversed-phase column at a flow-rate of 0.05 ml/min. Detection wavelength was set at 260 nm. The method was validated for response linearity as well as intra- and inter-day variabilities. Rapid appearance of cephalexin in the striatal dialysate suggested good blood–brain barrier penetration. This study provided pharmacokinetics information for cephalexin as well as demonstrated the applicability of this continuous sampling method for pharmacokinetics studies.     

Development and validation of a liquid chromatography/tandem mass spectrometry assay for the simultaneous determination of D-amphetamine and diphenhydramine in beagle dog plasma and its application to a pharmacokinetic study

A new drug, quick-acting anti-motion capsule (QAAMC) composed of d-amphetamine sulfate, dimenhydrinate and ginger extraction has been studied for anti-motion-sickness use. We have developed a sensitive, specific liquid chromatography-tandem mass spectrometry (LC-MS/MS) for the quantitative determination of d-amphetamine and diphenhydramine, the main effective components of the QAAMC, using pseudoephedrine as the internal standard. The analytes and internal standard were isolated from 200 microL plasma samples by a simple liquid-liquid extraction (LLE). Reverse-phase HPLC separation was accomplished on a Zorbax SB-C18 column (100 mm x 3.0 mm, 3.5 microm) with a mobile phase composed of methanol-water-formic acid (65:35:0.5, v/v/v) at a flow rate of 0.2 mL/min. The method had a chromatographic total run time of 5 min. A Varian 1200 L electrospray tandem mass spectrometer equipped with an electrospray ionization source was operated in selected reaction monitoring (SRM) mode with the precursor-to-product ion transitions m/z 136.0-->91.0 (D-amphetamine), 256.0-->167.0 (diphenhydramine) and 166.1-->148.0 (IS) used for quantitation. The method was sensitive with a lower limit of quantitation (LLOQ) of 0.5 ng/mL for d-amphetamine and 1 ng/mL for diphenhydramine, with good linearity in the range 0.5-200 ng/mL for D-amphetamine and 1-500 ng/mL for diphenhydramine (r(2)> or =0.9990). All the validation data, such as accuracy, precision, and inter-day repeatability, were within the required limits. The method was successfully applied to pharmacokinetic study of the QAAMC in beagle dogs.     

Application of liquid chromatographic method with fluorescence detection for the determination of triclabendazole in tablets and biological fluids

A simple and rapid liquid chromatographic method was developed and validated for the determination of triclabendazole with high accuracy and precision within 6 min. Good chromatographic separation was achieved using a CLC Shim‐pack C₈ (250 × 4.6 mm, 5 µm particle size) using the mobile phase containing a mixture of 0.02 m phosphate buffer and methanol with a ratio of (20 : 80 v/v) at pH 4.0 was pumped at a flow rate of 1.2 mL/min with fluorescence detection for the first time at 338 nm after excitation at 298 nm. Losartan potassium was used as an internal standard. The method showed good linearity in the ranges of 0.05–2.0 µg/mL with limits of detection and quantification of 14.1 and 42.6 ng/mL, respectively. The suggested method was successfully applied for the analysis of triclabendazole in tablets. The high sensitivity of the method enabled the determination of the studied drug in spiked human plasma with mean percentage of recoveries of 99.79 ± 5.09. Statistical evaluation of the data was performed according to ICH Guidelines. Copyright © 2013 John Wiley & Sons, Ltd.     

Development and validation of a highly rapid and sensitive LC-MS/MS method for determination of SZ-685C, an investigational marine anticancer agent, in rat plasma--application to a pharmacokinetic study in rats

A sensitive and rapid method was developed and validated for the quantitative analysis of the novel anticancer agent SZ-685C in rat plasma using high-performance liquid chromatography/tandem mass spectrometry (LC/MS/MS) in negative ion mode in order to support the following pre-clinical and clinical studies. SZ-685C and the internal standard (IS, emodin) were extracted from rat plasma by a simple liquid-liquid extraction technique using ethyl acetate as extraction solvent. Chromatographic separation was performed on an Elite Hypersil BDS C18 column (100 mm × 2.1 mm i.d., 3 μm). Elution was carried out using methanol/acetonitrile/2mM ammonium formate (pH 4) (80:15:5 (v/v/v)) at a flow-rate of 0.3 mL/min with a run time of 2.5 min. This assay was linear over a concentration range of 50-10,000 ng/mL with a lower limit of quantification of 50 ng/mL. The intra- and inter-batch precision was less than 15% for all quality control samples at concentrations of 100, 1000 and 7500 ng/mL. These results indicate that the method was efficient with a short run time and acceptable accuracy, precision and sensitivity. This method was successfully applied to explore pharmacokinetics of SZ-685C in rats after oral and intravenous administration of this agent. The absolute bioavailability is about 54.8-66.8% and the t(1/2) is 5.7-9.2h, these results provide basic information for further comprehensive pre-clinical research.     

Liquid chromatography studies on the pharmacokinetics of phentermine and fenfluramine in brain and blood microdialysates after intraperitoneal administration to rats

A highly sensitive and simple HPLC method with fluorescence detection for the determination of phentermine (Phen), fenfluramine (Fen) and norfenfluramine (Norf, the active metabolite of Fen) in rat brain and blood microdialysates has been developed. The brain and blood microdialysates were directly subjected to derivatization with 4-(4,5-diphenyl-1H-imidazol-2-yl) benzoyl chloride (DIB-Cl) in the presence of carbonate buffer (0.1 M, pH 9.0) at room temperature. The chromatographic conditions consisted of an ODS column and mobile phase composition of acetonitrile and water (65:35, v/v) with flow rate set at 1.0 ml/min. The detection was performed at excitation and emission wavelengths of 325 and 430 nm, respectively. Under these conditions, the DIB-derivatives of Phen, Fen and Norf were well separated and showed good linearities in the studied ranges (5-2000 nM for Phen and 10-2000 nM for Norf and Fen) with correlation coefficients greater than 0.999. The obtained detection limits were less than 23 fmol on column (for the three compounds) in both brain and blood microdialysates at a signal-to-noise ratio of 3 (S/N=3). The intra- and the inter-assay precisions were lower than 10%. The method coupled with microdialysis was applied for a pharmacokinetic drug-drug interaction study of Phen and Fen following individual and combined intraperitoneal administration to rats. In addition, since the role of protein binding in drug interactions can be quite involved, the method was applied for the determination of total and free Phen and Fen in rat plasma and ultrafiltrate, respectively. The results showed that Fen and/or Norf significantly altered the pharmacokinetic parameters of Phen in both blood and brain but did not alter its protein binding. On the other hand, there was no significant difference in the pharmacokinetics of Fen when administered with Phen.     

An HPLC method for determination of oridonin in rabbits using isopsoralen as an internal standard and its application to pharmacokinetic studies for oridonin-loaded nanoparticles

A simple and sensitive HPLC method has been developed and validated for the determination of oridonin (ORI) in rabbit plasma. A simple liquid-liquid extraction (LLE) method was applied to extract ORI and the internal standard (IS), isopsoralen, from rabbit plasma. Chromatographic separation of ORI and the IS was achieved with a Kromasil C18 5-mum column (250 mm x 4.6 mm) using methanol-water (50:50, v/v) as mobile phase at a flow rate of 1 mL/min. The ultraviolet (UV) detection wavelength was set at 241 nm. The lower limit of quantification (LLOQ) was 0.02 microg/mL. The calibration curves were linear over a concentration range of 0.02-10 microg/mL. The assay accuracy and precision were within the range of 95.1-113.5% and 5.4-8.6%, respectively. This HPLC method was applied successfully to the pharmacokinetic study of ORI-loaded poly(caprolactone)-poly(ethylene oxide)-poly(caprolactone) copolymer nanoparticles (ORI-PCL-PEO-PCL-NP) in rabbits, given as a single intravenous injection at the dose equivalent to 2mg of ORI/kg, and the pharmacokinetic parameters for ORI were compared with a single intravenous injection of a ORI solution at the same dose.     

Plasma pharmacokinetics and tissue distribution study of physalin D in rats by ultra-pressure liquid chromatography with tandem mass spectrometry

Physalin D is an important constituent of some traditional Chinese medicines, and has several known bioactivities. An UPLC-MS/MS method for the determination of physalin D in rat plasma and tissues was developed and the pharmacokinetic and tissue distribution characteristics of physalin D after intravenous administrations were investigated. The bio-samples were prepared by a simple protein precipitation, and the separation of physalin D was achieved on a UPLC HSS T3 column with a mobile phase consisting of methanol/acetonitrile (70:30, v/v) and water (containing 0.1% formic acid and 10 mM ammonium acetate) at a flow rate of 0.3 mL/min. The MS/MS detection was carried out by monitoring the fragmentation of m/z 544.9→508.8 for physalin D and m/z 286.7→152.8 for luteolin (internal standard; IS) on a triple-quadrupole mass spectrometer. The total run time was only 3.6 min. The analyte showed good linearity over a wide concentration range (R(2)>0.995) and its lower limit of quantification was 2 ng/mL. The pharmacokinetic study found that physalin D was distributed and eliminated rapidly in rats (t(1/2)<10 min). Tissue distribution showed the highest level was observed in kidney, then in liver, but no physalin D was detected in brain, which indicated that kidney was the major distribution tissue for physalin D in rats and that physalin D does not cross the blood-brain barrier.     

Simple and rapid method determination for metformin in human plasma using high performance liquid chromatography tandem mass spectrometry: application to pharmacokinetic studies

A rapid and simple method for quantitation of metformin (MET) in human plasma by HPLC-MS/MS was developed and validated. The sample preparation consists of plasma deproteinization using acetonitrile. The mobile phase consisted of water-acetonitrile and formic acid (55/45/0.048, v/v/%) and the run time was 3 min. A pursuit C(18) (100 mm x 2.0 mm i.d., 3 microm) column connected to a guard column MS-pursuit (0.20 mm x 0.20 mm i.d., 5 microm) was used. The range of the calibration curve was from 20 to 5000 ng/mL, the limit of quantitation being 20 ng/mL. The detection was performed on a mass spectrometer (ESI+), using metoprolol as internal standard. The calibration curves have r(2) values of 0.995 (CV=0.24%, n=10). The accuracy and precision were between 90.74 and 106.7% and coefficients of variations (CV) of 1.10 and 4.35%, respectively. The method was applied to determine the pharmacokinetic parameters: C(max) (1667.25 ng/mL) and T(max) (3.89 h).     

Selective and rapid liquid chromatography/negative-ion electrospray ionization mass spectrometry method for the quantification of valacyclovir and its metabolite in human plasma

A rapid, sensitive and specific method was developed for the quantification of valacyclovir and acyclovir in human plasma. Sample preparation was performed by protein precipitation with acetonitrile followed by filtration. Valacyclovir, acyclovir and ganciclovir (internal standard) were separated isocratically on a reversed-phase porous graphitized carbon analytical column (2.1 mm x 125.0 mm i.d., particle size 5 microm), using a mobile phase of acetonitrile/water with 0.05% (v/v) diethylamine (50:50, v/v) at a flow rate of 0.15 mL min(-1) in 4.0 min. Detection was performed by negative electrospray ionization using the selected ion monitoring mode of the deprotonated molecular ions at m/z 323.0 for valacyclovir, 224.0 for acyclovir and 254.0 for ganciclovir. The assay had linear calibration curves over the range 0.020-0.800 microg mL(-1) for valacyclovir and 0.100-20.00 microg mL(-1) for acyclovir. Accuracy and precision were within the acceptance limit of 15%. The method was successfully applied to the analysis of plasma samples obtained from patients after oral administration of valacyclovir.     

Simultaneous determination of metolazone and losartan potassium in their binary mixtures using high‐performance liquid chromatography with fluorimetric detection: application to combined tablets and spiked human plasma

A new, specific and sensitive reversed‐phase high‐performance liquid chromatography method was developed for the simultaneous determination of metolazone (MET) and losartan potassium (LOS). Good chromatographic separation was achieved within 6.0 min on a 150 × 4.6 mm i.d., 5 µm Waters, Ireland and ProDIGY 5 ODS 3 100 A column. A mobile phase containing a mixture of methanol and 0.02 M phosphate buffer (65:35, v/v) at pH 3.0 was used. The analysis was performed at a flow rate of 1 mL/min with fluorescence detection at 410 nm after excitation at 230 nm. Aspirin (ASP) was used as an internal standard. The proposed method was rectilinear over 2.0–40.0 (MET) and 40.0–800.0 ng/mL (LOS), with limits of detection of 0.22 and 4.52 ng/mL and limits of quantification of 0.68 and 13.70 ng/mL for MET and LOS, respectively. The method was successfully applied for the simultaneous analysis of the studied drugs in their laboratory‐prepared mixtures, single tablets and co‐formulated tablets. Moreover, the method was applied to an in vitro drug release (dissolution) test. The method was further extended to the determination of LOS in spiked human plasma. Statistical evaluation and comparison of data obtained using the proposed and comparison methods revealed no significant difference between the two methods in addition to good accuracy and precision for the proposed method. Copyright © 2013 John Wiley & Sons, Ltd.     

Development and validation of a liquid chromatography-tandem mass spectrometry for the determination of BPR0L075, a novel antimicrotuble agent, in rat plasma: application to a pharmacokinetic study

A rapid and sensitive liquid chromatography-tandem mass spectrometric method (LC-MS/MS) had been developed and validated to determine the concentrations of BPR0L075 in rat plasma. After a simple protein precipitation of plasma samples by acetonitrile, BPR0L075 was analyzed on a C(8) column at a flow rate of 0.5 mL/min. The mobile phase consisted of a mixture of 10 mM ammonium acetate containing 0.1% formic acid and acetonitrile (20:80, v/v). Both BPR0L075 (analyte) and the internal standard (BPR0L092) were determined using electro-spray ionization and the MS data acquisition was via multiple reactions monitoring (MRM) in positive scanning model. The MS/MS ion transitions monitored are m/z 342.2/195.2 and 312.5/165.2 for BPR0L075 and BPR0L092, respectively. The low limit of quantitation was 0.5 ng/mL. Each plasma sample was chromatographed within 5 min. The method was validated with respect to linearity, accuracy, precision, recovery, and stability. A good linear relationship was observed over the concentration range of 0.5-1000 ng/mL (r>0.9994). Absolute recoveries ranged from 63.45 to 68.34% in plasma at the concentrations of 2, 40, 400, and 800 ng/mL. The intra- and inter-day accuracy ranged from 92.04 to 111.80%. Intra- and inter-day relative standard deviations were 1.08-3.29% and 1.96-5.46%, respectively. This developed and validated assay method had been successfully applied to a pharmacokinetic study after intravenous injection of BPR0L075 in rats at a dose of 5mg/kg.     

Validation and implementation of a liquid chromatography/tandem mass spectrometry assay to quantitate ON 01910.Na, a mitotic progression modulator, in human plasma

A reverse-phase high performance liquid chromatographic method with tandem mass spectrometry (LC-MS/MS) was developed and validated for the quantitation of ON 01910.Na, a novel synthetic benzyl styryl sulfone, in human plasma. The assay involved a simple sample preparation with acetonitrile protein precipitation. ON 01910.Na and the internal standard temazepam were separated on a Waters X-Terra MS C(18) column with mobile phase of acetonitrile containing 0.1% formic acid /10mM ammonium acetate (55:45, v/v) using isocratic flow at 0.2 mL/min for 5 min. The analytes were monitored by tandem-mass spectrometry with electrospray positive ionization. Two calibration curves were generated over the range of 10-2000 ng/mL and 100-20000 ng/mL. The lower limit of quantitation (LLOQ) was 10 ng/mL for ON 01910.Na in human plasma. The accuracy and within- and between-day precisions were within the acceptance criteria for bioanalytical assays. ON 01910.Na was found stable in plasma at -70 degrees C for at least 1 year. The method was successfully applied to characterize the plasma concentration-time profiles of ON 01910.Na in the cancer patients in the Phase I study.     

Development and validation of a liquid chromatography-mass spectrometry method for the quantitation of naltrexone and 6beta-naltrexol in guinea pig plasma

A quantitative liquid chromatographic-electrospray ionization mass spectrometry method for the determination of naltrexone and 6beta-naltrexol in guinea pig plasma has been developed and validated using naloxone as an internal standard. A single step precipitation-extraction technique was carried out to extract the plasma samples using acetonitrile:ethyl acetate (1:1, v/v). The chromatographic separation was performed on a C(18) column using a mobile phase consisting of 35:65 (v/v) acetonitrile:2 mM ammonium acetate with 0.01 mM ammonium citrate at a flow rate of 0.25 mL/min. The analyte was detected after positive electrospray ionization using selected ion monitoring (SIM) mode. The mean recoveries for naltrexone, naltrexol, and naloxone were 91.7, 89.3, and 99.0%, respectively. The lower limit of quantification (LLOQ) for naltrexone and 6beta-naltrexol was 1.25 ng/mL, and the limit of detection (LOD) was 0.75 ng/mL. The method was applied to a pharmacokinetic study in order to assess the drug disposition of naltrexone in guinea pigs.