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Vijaya Bhaskar V.,DMPK Laboratory Biology Division | Middha A.,University of Rajasthan | Tiwari S.,DMPK Laboratory Biology Division | Shivakumar S.,DMPK Laboratory Biology Division
Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences | Year: 2013

A rapid sensitive and selective MRM based method for the determination of polyethylene glycol 400 (PEG 400) in rat plasma was developed using liquid chromatography/tandem mass spectrometry (LC-MS/MS). PEG 400 and telmisartan (Internal standard) were extracted from rat plasma with acetonitrile and analysed on C18 column (Waters Xbridge, 50. ×. 4.6. mm, 3.5. μm) with the mobile phase (A - 0.1% formic acid in water; B - methanol). A generic gradient method with a short run time of 3.5. min was developed for the analysis of PEG 400. A total of nine oligomers were identified for PEG 400. The most abundant ions corresponding to PEG 400 oligomers at m/. z 327, 371, 432, 476, 520, 564, 608, 652 and 696 with daughter ion at m/. z 89 were selected for multiple reaction monitoring (MRM) in electrospray mode of ionisation. Analyte peak area of the oligomers was summed up to calculate the plasma concentrations of total PEG 400. The standard curve was linear (0.9954) over the concentration range of 1.01-1013.40. μg/mL. The lower limit of quantitation for PEG 400 was 1.01. μg/mL using 50. μL plasma. The coefficient of variation and relative error for inter and intraassay at three QC levels were 2.31-13.34 and -7.99 to 0.37, respectively. The method was validated for various parameters such as extraction recovery, matrix effect, autosampler stability, benchtop stability, freeze thaw stability, long term stability and was proved to be consistent across three QC levels with overall %CV less than 15. The developed method was successfully applied to the absolute bioavailability study of PEG 400 in male Sprague Dawley rats. Plasma concentrations of PEG 400 was measured after administration through oral and intravenous routes in male Sprague Dawley rats at a dose of 3.38. g/kg. Pharmacokinetic (PK) parameters were characterised by performing the analysis using Phoenix Winnonlin software (v 6.3). PEG 400 has good oral bioavailability with mean absolute bioavailability of 47.23%. Plasma concentration profile/PK parameters of PEG 400 was established in both intravenous and oral routes, which helps to qualify the analytical batch of NCEs having spiky plasma concentration profiles/erratic results. Purity of the PEG 400 oligomers was estimated using ELSD detection. Differences in pharmacokinetics of oligomers was studied. It was found that with increase in molecular weight of the oligomer, a decrease in absolute bioavailability was observed. © 2013. Source


Bhaskar V.V.,DMPK Laboratory Biology Division | Bhaskar V.V.,University of Rajasthan | Middha A.,University of Rajasthan | Srivastava P.,DMPK Laboratory Biology Division | Rajagopal S.,DMPK Laboratory Biology Division
Journal of Pharmaceutical Analysis | Year: 2015

A rapid, sensitive and selective pseudoMRM (pMRM)-based method for the determination of solutol HS15 (SHS15) in rat plasma was developed using liquid chromatography/tandem mass spectrometry (LC-MS/MS). The most abundant ions corresponding to SHS15 free polyethyleneglycol (PEG) oligomers at m/z 481, 525, 569, 613, 657, 701, 745, 789, 833, 877, 921 and 965 were selected for pMRM in electrospray mode of ionization. Purity of the lipophilic and hydrophilic components of SHS15 was estimated using evaporative light scattering detector (ELSD). Plasma concentrations of SHS15 were measured after oral administration at 2.50 g/kg dose and intravenous administration at 1.00 g/kg dose in male Sprague Dawley rats. SHS15 has poor oral bioavailability of 13.74% in rats. Differences in pharmacokinetics of oligomers were studied. A novel proposal was conveyed to the scientific community, where formulation excipient could be analyzed as a qualifier in the analysis of new chemical entities (NCEs) to address the spiky plasma concentration profiles. © 2014 Xi'an Jiaotong University. Source


Vijayabhaskar V.,DMPK Laboratory Biology Division | Srivastava P.,DMPK Laboratory Biology Division | Rajagopal S.,DMPK Laboratory Biology Division
Journal of Chromatography B: Analytical Technologies in the Biomedical and Life Sciences | Year: 2015

A rapid selective and sensitive liquid chromatography/tandem mass spectrometry (LC-MS/MS) method was developed for the quantitative determination of derivatised cytochrome P450-2C19 oxidation product (dansyl-4-OH mephenytoin) and its underivatised form (4-OH mephenytoin). Samples were anaysed on C18 column (Waters Xbridge, 50mm×4.6mm, 3.5μm particle size) with the mobile phase consisting of 0.1% formic acid in water and 0.1% formic acid in acetonitrile. A gradient method with a short run time of 2.5min and 3.5min was developed for the analysis of dansyl-4-OH mephenytoin and 4-OH mephenytoin, respectively. The standard curve was linear (r2=0.9972 for 4-OH mephenytoin; r2=0.9946 for dansyl-4-OH mephenytoin) over the concentration range of 0.16 to 40ng/mL for both derivatised and underivatised forms. The CV (%) and relative error (RE) for inter and intraassay at three QC levels for dansyl-4-OH mephenytoin was 0.97-5.85% and -9.80 to 2.51%, respectively. Whereas, for 4-OH mephenytoin the CV (%) and RE (%) at three QC levels was 0.82-3.47% and -6.69 to -0.01%, respectively. The developed method was validated for various parameters such as linearity, precision & accuracy, extraction recovery, matrix effect, autosampler stability and was proved to be consistent across three QC levels with overall CV (%) less than 15. Dansylation helped in increasing the sensitivity of hydroxy mephenytoin by 100-200 fold. Given the simplicity involved in derivatisation process, we believe that this novel methodology will change the current approaches used for the enhancing the detection sensitivity of 4-OH mephenytoin. © 2015 Elsevier B.V. Source

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