Clinical development of imatinib: an anticancer drug.

BACKGROUND: A novel and accurate high-throughput tandem mass spectroscopic method has been developed and validated for determination of imatinib, a protein-tyrosine kinase inhibitor against chronic myeloid leukemia. MATERIALS & METHODS: Chromatographic separation was carried on XTerra® RP18 column (150 mm × 4.6 mm, 5 µm particle size) manufactured by Waters Corporation, MA, USA. The detection was performed on a triple quadruple tandem mass spectrometer by multiple reactions monitoring mode via electrospray ionization source. RESULTS: The selective and sensitive method was linear in the concentration range of 9.57-4513.29 ng/ml and reported no matrix effect. CONCLUSION: The mean Cmax was found to be 10-15% lower in European subjects as compared with Indian subjects.

Development and Validation of an LC-MS-MS Method for Determination of Simvastatin and Simvastatin Acid in Human Plasma: Application to a Pharmacokinetic Study.

A liquid chromatography-tandem mass spectrometry method was developed and validated for the simultaneous determination of simvastatin (SV) and simvastatin acid (SVA) in human plasma. To improve assay sensitivity and achieve simultaneous analysis, SVA monitored in (-)ESI (electrospray ionization) mode within the first 4.5 min and SV thereafter in (+)ESI mode. The separation of all compounds was achieved in about 6.2 min using a C18 reverse-phase fused-core(®) column (Ascentis(®) Express C18) and a mobile phase, which was composed of 2.00 ± 0.05 mM ammonium acetate buffer titrated to pH 3.8 with glacial acetic acid-acetonitrile (25:75, v/v), in isocratic mode at a flow rate of 0.500 mL/min. Additionally, a solid-phase extraction step was performed to reduce any ion-suppression and/or enhancement effects. The developed method was linear in the concentration range of 0.100-74.626 ng/mL for SV, and 0.100-48.971 ng/mL for SVA, with correlation coefficient greater than 0.99 for both analytes. The method has shown tremendous reproducibility, with intra- and inter-day precision <7.6%, and intra- and interday accuracy within ±10.9% of nominal values, for the both analytes. The method was successfully applied to characterize the pharmacokinetic profiles of SV and SVA following an oral administration of 40 mg SV tablet to healthy human volunteers.

A Multi-centric Bioequivalence Trial in Ph+ Chronic Myeloid Leukemia Patients to Assess Bioequivalence and Safety Evaluation of Generic Imatinib Mesylate 400 mg Tablets.

PURPOSE: This study was designed to characterize the pharmacokinetic profile and to assess bioequivalence of the sponsor's test formulation (imatinib mesylate 400 mg tablets) with an innovator product (Gleevec 400 mg tablets, Novartis Pharmaceuticals) under fed conditions, in adult patients of Philadelphia chromosome positive chronic myeloid leukemia (Ph+ CML) stabilized on imatinib mesylate 400 mg. In addition, the aim of this study was to monitor the safety profile of investigational medicinal products (IMPs). MATERIALS AND METHODS: A multicenter, randomized, open label, two-period, crossover, single dose bioequivalence study was designed for conduct under fed conditions in 42 adult Ph+ CML patients already stabilized on imatinib 400 mg tablets. Pharmacokinetic parameters Tmax, Cmax, and AUC0-24 were calculated using a non-compartmental model on validated WinNonlin software. Validated SAS software was used for statistical evaluation of data. The safety profile of investigational products was monitored during the course of study by applying a clinical process for recording observed untoward effects postadministration of investigational products. RESULTS: The 90% confidence intervals for the test/reference mean ratios of the ln-transformed PK variables Cmax (99.0%) and AUC0-24 (99.2%) were within an acceptable range of 80%-125%, as per bioequivalence assumptions. Both formulations were well tolerated after oral administration of IMPs. CONCLUSION: The test product was found to be bioequivalent and safe, and thus can be used interchangeably in clinical practice.

Liquid chromatography tandem mass spectrometry method for the quantitation of mycophenolate mofetil in human plasma: Application to a bioequivalence study and metabolite identification.

We established a sensitive, selective, and rapid analytical method for the quantitation and pharmacokinetic investigation of mycophenolate mofetil in human plasma. To our knowledge, this is the first method that characterizes presence of mycophenolate mofetil glucuronide in clinical samples through tandem mass spectrometry detection and resolves mycophenolate mofetil from its glucuronide metabolite. Liquid chromatography coupled to tandem mass spectrometry detection in positive ion mode was selected to provide optimal selectivity and sensitivity. Due to the ionizable characteristics of the mycophenolate mofetil, a mixed-mode cation-exchange disposable extraction cartridge was prudently chosen. The chromatographic separation was achieved on Luna(®) C18(2) (100×4.60 mm) column using mobile phase consisting of a mixture of 1±0.05 mM ammonium formate in water, titrated to pH 3.1±0.1 with formic acid, and methanol (20:80, v/v), at a flow rate of 0.7 mL/min. The detection was led at m/z ratios of 434.4→ 114.2 and 438.4→ 118.3, for mycophenolate mofetil and mycophenolate mofetil-D4, respectively. The developed method was linear between 40.2-4986.0 pg/mL. All validation parameters were within the defined limits. The validated method was then successfully applied for the evaluation of bioequivalence parameters of mycophenolate mofetil after an oral administration of 500 mg mycophenolate mofetil tablet to healthy male Indian volunteers.

Liquid chromatography-tandem mass spectrometry method for the estimation of adefovir in human plasma: Application to a pharmacokinetic study.

An analytical method based on solid phase extraction was developed and validated for analysis of adefovir in human plasma. Adefovir-d4 was used as an internal standard and Synergi MAX RP80A (150 mm×4.6 mm, 4 µm) column provided the desired chromatographic separation of compounds followed by detection with mass spectrometry. The method used simple isocratic chromatographic condition and mass spectrometric detection in the positive ionization mode. The calibration curves were linear over the range of 0.50-42.47 ng/mL with the lower limit of quantitation validated at 0.50 ng/mL. Matrix effect was assessed by post-column infusion experiment to monitor phospholipids and post-extraction addition experiment was performed. The degree of matrix effect for adefovir was determined as 7.5% and ion-enhancement in five different lots of human plasma was 7.1% and had no impact on study samples analysis with 4.5 min run time. The intra- and inter-day precision values were within 7.7% and 7.8%, respectively, for adefovir at the lower limit of quantification level. Validated bioanalytical method was successfully applied to clinical sample analysis.

Development and Validation of a LC-MS/MS Method for the Simultaneous Estimation of Amlodipine and Valsartan in Human Plasma: Application to a Bioequivalence Study.

A reliable, simple, and robust liquid chromatography-tandem mass spectro-metric (LC-MS/MS) method has been developed and validated that employs solid-phase extraction for the simultaneous estimation of amlodipine and valsartan in human K3EDTA plasma using amlodipine-d4 and valsartan-d9 as internal standards. Chromatographic separation of amlodipine and valsartan was achieved on the Luna C18 (2)100A (150 × 4.6 mm, 5 µm) column using acetonitrile: 5 mM ammonium formate solution (80:20, v/v) as the mobile phase at a flow rate of 0.8 mL/min in isocratic mode. Quantification was achieved using an electrospray ion interface operating in positive mode, under multiple reaction monitoring (MRM) conditions. The assay was found to be linear over the range of 0.302-20.725 ng/mL for amlodipine and 6.062-18060.792 ng/mL for valsartan. The method has shown good reproducibility, as intra- and interday precisions were within 10% and accuracies were within 8% of nominal values for both analytes. The method was successfully applied for the bioequivalence study of amlodipine and valsartan after oral administration of a fixed dose of the combination. Additionally, as required by the current regulatory bodies, incurred sample reanalysis was performed and found to be acceptable.

Simultaneous quantitation of atorvastatin and its two active metabolites in human plasma by liquid chromatography/(-) electrospray tandem mass spectrometry.

A sensitive, accurate and selective liquid chromatography-tandem mass spectrometry method (LC-MS/MS) was developed and validated for the simultaneous quantitation of atorvastatin (AT) and its equipotent hydroxyl metabolites, 2-hydroxy atorvastatin (2-AT) and 4-hydroxy atorvastatin (4-AT), in human plasma. Electrospray ionization (ESI) interface in negative ion mode was selected to improve the selectivity and the sensitivity required for this application. Additionally, a solid phase extraction (SPE) step was performed to reduce any ion-suppression and/or enhancement effects. The separation of all compounds was achieved in less than 6 min using a C18 reverse-phase fused-core® column and a mobile phase, composed of a mixture of 0.005% formic acid in water:acetonitrile:methanol (35:25:40, v/v/v), in isocratic mode at a flow rate of 0.6 mL/min. The method has lower limit of quantitation (LLOQ) of 0.050 ng/mL for all analytes. The method has shown tremendous reproducibility, with intra- and inter-day precision less than 6.6%, and intra- and inter-day accuracy within ±4.3% of nominal values, for all analytes, and has proved to be highly reliable for the analysis of clinical samples.

Pharmacokinetic comparison and bioequivalence evaluation of losartan/ hydrochlorothiazide tablet between Asian Indian and Japanese volunteers.

OBJECTIVES: To demonstrate the bioequivalence between the test and reference formulations of losartan/hydrochlorothiazide 50 + 12.5 mg tablet and evaluate the effect of ethnicity on pharmacokinetics properties of losartan, losartan carboxylic acid and hydrochlorothiazide on healthy Asian Indian and Japanese volunteers. METHODS: Randomized, open-label, crossover, bioavailability studies were conducted separately in healthy Asian Indian and Japanese volunteers. One tablet either of test or of reference product was administered after 10 hours of overnight fasting. After dosing, serial blood samples were collected for a period of 48 hours for both the studies. Plasma samples were analyzed for losartan, losartan carboxylic acid and hydrochlorothiazide by a validated liquid chromatographic and mass spectrometric method (LC-MS/MS). The pharmacokinetic parameters AUC0-t, AUC0-∞, Cmax, tmax, and other pharmacokinetics parameters were determined from plasma concentration-time profiles for both test and reference formulations of losartan/hydrochlorothiazide 50 + 12.5 mg tablets. Statistical evaluations were done to evaluate bioequivalence between generic test formulation (EPR0001) and Japanese reference product (Preminent®). RESULTS: Losartan, losartan carboxylic acid and hydrochlorothiazide were well tolerated by subjects in all periods of each study under fasted conditions. No serious adverse events were observed. The ratios of least square means for AUC0-t and Cmax and the affiliated 90% confidence intervals were within acceptance range recommended by PMDA. Marginal differences were observed in pharmacokinetic values of Asian Indian and Japanese volunteers. CONCLUSIONS: The results of these bioavailability studies indicate that the test formulation of losartan/hydrochlorothiazide 50 + 12.5 mg (EPR0001) tablets is bioequivalent to marketed Preminent® reference formulation in Asian Indian and Japanese volunteers, when administered under fasting conditions. Both test and reference formulations were well tolerated as a single oral dose when administered to healthy adult subjects under fasted conditions. Although Asian Indian and Japanese volunteers are ethnically different, results of these studies indicate that pharmacokinetic parameters of Asian Indian and Japanese volunteers are comparable to each other in terms of bioavailability of losartan, losartan carboxylic acid and hydrochlorothiazide. Similar least square means ratios were obtained in Asian Indian and Japanese volunteers demonstrating that a bioequivalence study conducted on Japanese volunteers seems to be substituted by Asian Indian volunteers' studies.

Simultaneous quantitation of atorvastatin and its two active metabolites in human plasma by liquid chromatography/(-) electrospray tandem mass spectrometry / 药物分析学报

A sensitive, accurate and selective liquid chromatography-tandem mass spectrometry method (LC-MS/MS) was developed and validated for the simultaneous quantitation of atorvastatin (AT) and its equipotent hydroxyl metabolites, 2-hydroxy atorvastatin (2-AT) and 4-hydroxy atorvastatin (4-AT), in human plasma. Electrospray ionization (ESI) interface in negative ion mode was selected to improve the selectivity and the sensitivity required for this application. Additionally, a solid phase extraction (SPE) step was performed to reduce any ion-suppression and/or enhancement effects. The separation of all compounds was achieved in less than 6 min using a C18 reverse-phase fused-cores column and a mobile phase, composed of a mixture of 0.005%formic acid in water:acetonitrile:methanol (35:25:40, v/v/v), in isocratic mode at a flow rate of 0.6 mL/min. The method has lower limit of quantitation (LLOQ) of 0.050 ng/mL for all analytes. The method has shown tremendous reproducibility, with intra-and inter-day precision less than 6.6%, and intra- and inter-day accuracy within 74.3% of nominal values, for all analytes, and has proved to be highly reliable for the analysis of clinical samples.

Combining benefits of an adrenergic and a muscarinic blocker in a single formulation - a pharmacokinetic evaluation.

A pharmacokinetic bioequivalence study was conducted in Asian subjects, to compare a fixed dose combination capsule single oral dose of alpha adrenoceptor blocker-Alfuzosin hydrochloride 10mg extended release and muscarinic antagonists-Solifenacin succinate 5mg against individually administered Xatral XL 10mg tablets (Alfuzosin) of Sanofi Synthelabo Limited, United Kingdom (UK) and Vesicare 5mg tablets (Solifenacin) of Astellas Pharma Limited, UK under fed conditions. Blood samples were collected pre-dose up to 72 h post dose for determination of plasma Alfuzosin and Solifenacin concentrations and calculation of the pharmacokinetic parameters. ANOVA was performed on the log (natural)-transformed pharmacokinetic parameters. A 90% confidence interval for the ratios of the test and reference product averages (least square means) were calculated for alfuzosin and solifenacin. The 90% confidence intervals obtained for alfuzosin for Cmax, AUC0-t and AUC0-∞ were 102.74-122.75%, 95.84-116.96% and 95.82-116.76%, respectively. The 90% confidence intervals obtained for Solifenacin for Cmax, and AUC0-72 were 89.55-97.91% and 90.47-99.38%, respectively. Based on the results, the fixed dose combination was concluded to be bioequivalent to individually administered products.

Bioequivalence of oxycodone hydrochoride extended release tablets to marketed reference products OxyContin® in Canada and US.

INTRODUCTION: Oxycodone is a semisynthetic opioid agonist used for the relief of moderate to severe pain. A new generic oxycodone hydrochloride (HCl) extended release (ER) tablet is currently being developed by Ranbaxy Pharmaceutical Inc., New Brunswick, NJ, USA. OBJECTIVE: To assess relative bioavailability of a new generic (test) formulation of oxycodone hydrochloride (HCl) extended release (ER) tablets with that of marketed reference products, OxyContin®, in Canada and USA, in healthy adult subjects under fasting and fed conditions. METHODS: Five studies were conducted in all, three of which were designed to comply with the regulatory criteria for marketing a new generic formulation of OxyContin® in Canada and the remaining two to comply with regulatory criteria for marketing a new generic formulation of OxyContin® in the USA. Each study was a balanced, randomized two-period, two-treatment, two-sequence, crossover design. A single oral dose of test or reference product was given in Period 1, followed by a 7-day washout period, after which subjects received the alternative product in Period 2. In order to block the pharmacological effects of oxycodone, subjects were administered naltrexone HCl (1 × 50 mg tablet) 12 hours prior to oxycodone HCl administration, concurrent with oxycodone HCl administration, and 12 hours after oxycodone HCl administration. Throughout the confinement portion of the study, adverse events were closely monitored. Serial blood samples were collected, following which oxycodone in plasma was estimated using a validated analytical procedure. RESULTS: Oxycodone was well tolerated by subjects in both periods of each study under both fed and fasted conditions. No serious adverse events were observed. The ratios of geometric means for AUC0-t and Cmax and the affiliated 90% confidence intervals for AUC were within acceptance range recommended by Health Canada. These criteria were met for both the raw data as well as data corrected for measured drug content (potency). The ratios of geometric means and the 90% confidence intervals for AUC0-t, AUC0-∞ and Cmax were within acceptance range recommended by United States Food and Drug Administration (FDA). CONCLUSIONS: Results demonstrate that the test formulation of oxycodone HCl ER tablets is bioequivalent to marketed OxyContin® reference formulations in Canada and USA, when administered both under fasted and fed conditions. Additionally, oxycodone HCl ER tablets were well tolerated as a single oral dose when administered to healthy adult subjects under fasted and fed conditions.

Development and validation of high performance liquid chromatographic method for analysis of clozapine.

In this study a rapid, simple and sensitive assay to quantify clozapine in human plasma by using reverse phase high performance liquid chromatographic method has been developed. Clozapine was extracted from human plasma using a mixture of chloroform: n-hexane 50:50 employing liquid-liquid extraction method. The calibration curve was found to be linear in the concentration range of 25-800 ng/ml. The inter day and intra day assay accuracy and precision fulfilled the criteria specified by USFDA, Guidance for industry: bioanalytical method validation. Clozapine was found to be stable in human plasma after 6 h incubation at room temperature, 50 days storage at -27°C and freeze thaw cycles, as well as after reconstitution with mobile phase after 24 h of storage in refrigerator. The validated method offers the advantage of using minimum injection volume (25µl) and plasma sample volume (300µl). The extraction method is simple and single step with no back extraction step, thus, making this method applicable to determination of pharmacokinetic profiles and parameters.

Liquid chromatography tandem mass spectrometry method for the simultaneous stereoselective determination of venlafaxine and its major metabolite, O-desmethylvenlafaxine, in human plasma.

A sensitive, accurate and highly stereoselective assay for the simultaneous determination of venlafaxine (VEN) and its equipotent metabolite, O-desmethyl venlafaxine (ODV), in human plasma was developed and validated. Analytes were simultaneously extracted from plasma using solid-phase extraction and detected by tandem mass spectrometry in positive ion mode with a turbo ion spray interface. Deuterium-labeled VEN and ODV were used as internal standards. Chromatographic separation was performed on a Chiral AGP column, using a time programmed gradient flow with a total run time of 16 min. The method has a lower limit of quantitation of 0.60 ng/mL. The assay was linear over a range 0.60-300.00 ng/mL for both the enantiomers of VEN and ODV, respectively, with coefficient of correlation > 0.99. The extraction recoveries were >77.0% on an average for all the four analytes. The analytes were found stable in plasma through three freeze (-15 °C) and thaw cycles and under storage at room temperature for 8 h, and also in mobile phase at 10 °C for 54 h. The method has shown good reproducibility, with intra- and inter-day variation coefficients < 9%, for all the analytes, and has proved to be very reliable for analysis of VEN and its metabolite in clinical study samples.

Liquid chromatography tandem mass spectrometry method for the estimation of lamotrigine in human plasma: Application to a pharmacokinetic study.

A reliable, selective and sensitive liquid chromatography tandem mass spectrometry method was developed and validated for the quantification of lamotrigine in human plasma using lamotrigine-13C3, d3 as an internal standard. Analyte and internal standard were extracted from human plasma by solid-phase extraction and detected in positive ion mode by tandem mass spectrometry with electrospray ionization (ESI) interface. Chromatographic separation was performed on a Chromolith® SpeedROD; RP-18e column (50-4.6 mm i.d.) using acetonitrile: 5±0.1 mM ammonium formate solution (90:10, v/v) as the mobile phase at a flow rate of 0.500 mL/min. The calibration curves were linear over the range of 5.02-1226.47 ng/mL with the lower limit of quantitation validated at 5.02 ng/mL. The analytes were found stable in human plasma through three freeze (-20 °C)-thaw (ice-cold water bath) cycles and under storage on bench-top in ice-cold water bath for at least 6.8 h, and also in the mobile phase at 10 °C for at least 57 h. The method has shown good reproducibility, as the intra- and inter-day precisions were within 3.0%, while the accuracies were within ±6.0% of nominal values. The validated LC-MS/MS method was applied for the evaluation of pharmacokinetic and bioequivalence parameters of lamotrigine after an oral administration of 50 mg lamotrigine tablet to thirty-two healthy adult male volunteers.

Metaxalone estimation in biological matrix using high-throughput LC-MS/MS bioanalytical method.

Metaxalone is a skeletal muscle relaxant, an approved drug for pain relief. Published bioanalytical methods lacked detailed stability evaluation in blood and plasma. An accurate, precise, high-throughput tandem mass spectroscopic method has been developed and validated. Following solid phase extraction (SPE), metaxalone and the internal standard metaxalone-d(3) were extracted from an aliquot of 200 µL of human plasma. Chromatographic separation achieved on an Ascentis Express C18 column (50 mm × 4.6 mm i.d., 2.7 µm particle size) with mobile phase is a mixture of 10mM ammonium acetate buffer (pH 4.5)-methanol-acetonitrile (20:50:30, v/v/v), at an isocratic flow rate of 0.7 mL/min. The detection was performed on a triple quadrupole tandem mass spectrometer by multiple reaction monitoring (MRM) mode via electrospray ionization (ESI) source. The mass transitions of metaxalone and metaxalone-d(3) were m/z 222.3→161.2 and m/z 225.3→163.3, respectively. The linear calibration curves were obtained in the concentration range of 0.105-10.081 µg/mL (r(2)≥0.99) with a lower limit of quantification (LLOQ) of 0.105 µg/mL. The intra- and inter-day precisions and relative error were all within 6%. Despite achieving high mean recovery (>78%), no interference peaks or matrix effects were observed. Detailed stability exercises including drug stability in blood, hemolyzed, lipemic and normal plasma were conducted to extend the method applicability in vast majority of clinical studies using 800 mg metaxalone extended release oral dosage form.

ESI-MS/MS stability-indicating bioanalytical method development and validation for simultaneous estimation of donepezil, 5-desmethyl donepezil and 6-desmethyl donepezil in human plasma.

A bioanalytical method was developed and validated to estimate donepezil, 6-desmethyl donepezil and 5-desmethyl donepezil simultaneously in human plasma using galantamine as an internal standard (IS). The chromatographic separation was achieved on a reverse-phase XTerra RP (150 × 4.6 mm, 5 µm) column without affecting recovery (mean recovery > 60% with CV < 10%) for all analytes. ESI-MS/MS multiple reaction monitoring in positive polarity was used to detect mass pairs for donepezil (m/z 380.3 → 91.3), 6-desmethyl donepezil (m/z 366.4 → 91.3), 5-desmethyl donepezil (m/z 366.4 → 91.3) and galantamine m/z (288.1 → 213.0). The linearity was established over a dynamic range of 0.339-51.870, 0.100-15.380 and 0.103-15.763 ng/mL for donepezil, 6-desmethyl donepezil and 5-desmethyl donepezil, respectively. The current method shows that minimal conversion of labile metabolites to parent donepezil in plasma as stability was successfully achieved for 211 days at -15 °C storage temperature. The method was successfully applied to a clinical study after administration of 10 mg donepezil tablets to healthy male Indian volunteers.

Pharmacokinetics of a fixed-dose combination of atorvastatin and metformin extended release versus concurrent administration of individual formulations: a randomized, open-label, two-treatment, two-period, two-sequence, single-dose, crossover, bioequivalence study.

BACKGROUND: Type 2 diabetes mellitus is associated with a 2- to 4-fold increased risk of coronary heart disease (CHD). Combined therapy with an antihyperglycaemic agent and an HMG-CoA reductase inhibitor (statin) is indicated for the treatment of diabetic patients at risk of CHD. Patients with type 2 diabetes are generally considered to be at equivalent cardiovascular disease risk to patients with established CHD, and should have low-density lipoprotein (LDL) cholesterol levels reduced to <100 mg/dL or by 30-40%. Atorvastatin is the drug of choice for lowering LDL cholesterol levels. Metformin is the first therapeutic option in type 2 diabetes patients who are overweight or obese because it may also prevent vascular complications and mortality. Hence, a fixed-dose combination (FDC) of atorvastatin 10 mg and metformin 500 mg extended release (ER) was developed for patients with type 2 diabetes with or without hyperlipidaemia. OBJECTIVES: This study set out to establish bioequivalence between treatment 1 (test) - atorvastatin/metformin ER 10 mg/500 mg FDC, and treatment 2 (reference) - atorvastatin 10 mg (Lipitor®) and metformin 500 mg (Glucophage® XR) administered concurrently as individual tablets. METHODS: The study was a randomized, open-label, two-treatment, two-period, two-sequence, single-dose, crossover, bioequivalence study in 40 male subjects of Asian origin aged 18-45 years. The order of receiving the test and reference treatments for each subject during both the periods of the study was determined according to an SAS®-generated randomization schedule. The two treatments were separated by a washout period of 11 days. Blood samples were collected pre-dose and up to 72 hours post-dose in each period for determination of plasma atorvastatin/metformin concentrations and calculation of the respective pharmacokinetic parameters. ANOVA was performed on the lognormal-transformed pharmacokinetic parameters. A 90% confidence interval (CI) for the ratios of the test and reference product averages (least squares means) was calculated for atorvastatin and metformin to establish bioequivalence. RESULTS: The 90% CIs for atorvastatin and metformin were within the bioequivalence acceptance criteria of 80-125%. The 90% CIs obtained for atorvastatin for maximum plasma concentration (C(max))(,) area under the plasma concentration-time curve (AUC) from time zero to time of last measurable concentration (AUC(last)) and AUC from time zero to infinity (AUC(∞)) were (88.11, 106.93), (91.18, 107.94) and (89.25, 106.60), respectively. The 90% CIs observed for metformin for C(max,) AUC(last,) AUC(∞) and AUC from time zero to 24 hours (AUC(24)) were (113.3, 124.0), (102.65, 117.97), (101.87, 116.82) and (102.44, 117.53), respectively. The two treatments were well tolerated by the study subjects. CONCLUSION: Atorvastatin/metformin ER 10 mg/500 mg FDC has similar bioavailability to the co-administration of separate atorvastatin 10 mg and metformin 500 mg tablets. The FDC tablets show similar safety and tolerability profiles to their individual components. Therefore, atorvastatin/metformin ER 10 mg/500 mg FDC tablets can be used safely in clinical settings to decrease the pill burden and increase patient compliance with therapy.

Establishing bioequivalence of racemic venlafaxine formulations using stereoselective assay method: Is it necessary?

A bioequivalence study for venlafaxine generic formulation was conducted as an open label, balanced, randomized, two-way crossover, single-dose study. In this study, a comparison of various pharmacokinetic parameters of venlafaxine hydrochloride 150 mg modified release capsules of Ranbaxy and EFEXOR®-XR 150 mg capsules of Wyeth, in healthy, adult, male, human subjects under fasting condition was performed to conclude bioequivalence. Venlafaxine and its major active metabolite O-desmethylvenlafaxine (ODV) are racemates. The "(S)-(+)" and "(R)-(-)" enantiomers of venlafaxine and ODV are established as being active. Hence, subject samples were analyzed using nonstereoselective and stereoselective assay methods. Both (S)-(+) and (R)-(-) enantiomers of venlafaxine and ODV showed similar absorption and disposition. The 90% confidence intervals for venlafaxine, (R)-(-)-venlafaxine as well as (S)-(+)-venlafaxine were within acceptance range concluding bioequivalence. The results obtained by stereoselective assay were comparable to the nonstereoselective analysis, as sum of concentrations of (S)-(+)- and (R)-(-)-enantiomers of venlafaxine and ODV. The mean (S)-(+)/(R)-(-) ratios of the enantiomers of venlafaxine and ODV at various time points were consistent in the study subjects. Therefore, the estimation of venlafaxine and ODV using nonstereoselective assay method is effective in distinguishing formulation differences (if any) in bioequivalence studies in a cost-effective manner.

Determination of valganciclovir and ganciclovir in human plasma by liquid chromatography tandem mass spectrometric detection.

OBJECTIVES: The aim of this work was to develop a simple, sensitive and selective LC/MS/MS method for the assay of valganciclovir and ganciclovir in human plasma. DESIGN AND METHODS: Sample preparation involved solid phase extraction on mix mode cation exchanger. Separation was performed on Chromolith RP18e column using water, trifluoroacetic acid (1M, pH 4.4) and methanol (29.9:0.1:70, v/v) as mobile phase. Both analytes were detected by electro spray ionization mass spectrometry in positive ion multiple reaction monitoring mode. RESULTS: CCs with good linearties having r≥0.9990 and ≥0.9992 were obtained in the range of 5-800ng/mL and 70-11,200ng/mL for valganciclovir and ganciclovir, respectively. The extraction recoveries were around 85% for both the analytes. CONCLUSION: The method provided a simple and selective procedure that can be easily used for the evaluation of the pharmacokinetic profile of valganciclovir and ganciclovir in human plasma.

Controlled ex-vivo plasma hydrolysis of valaciclovir to acyclovir demonstration using tandem mass spectrometry.

Plasma estimation of valaciclovir, an antiviral drug, is challenging due to both in-vivo and ex-vivo hydrolysis to active metabolite acyclovir. A simultaneous method is described involving the solid-phase ion-exchange extraction procedure requiring 100 µL of plasma volume, a reverse-phase Lichrosphere RP Select B (125 × 6 mm, 5 µm) column and isocratic mobile phase to achieve the desired chromatographic separation. ESI-MS/MS multiple reaction monitoring in positive polarity, detected mass pairs for valaciclovir (m/z 325.5 → 152.2), acyclovir (m/z 226.3 → 152.2) and respective internal standards valganciclovir (m/z 307.1 → 220.3) and acyclovir-d4 (m/z 230.2 → 152.0). Fully fledged method validation was evaluated as per current regulatory requirements and results were deemed acceptable. The plasma samples showed extensive hydrolysis of valaciclovir when collected or processed at room temperature, without buffer stabilization prior to storage at -15°C. Our results showed that using prechilled K3 EDTA vacutainers immersed in an iced-water bath during blood sample collection, and addition of 50% orthophosphoric acid solution to plasma samples prior to storage at -50°C for at least 120 days controlled the hydrolysis of valaciclovir to acyclovir. While monitoring drug absorption into systematic circulation, the valaciclovir to acyclovir formation ratio was improved to 1:20 in healthy volunteers for the first time.