Determination of Pregabalin in Human Plasma Using LC-MS-MS |
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Authors: | Uttam Mandal Amlan Kanti Sarkar Kadagi Veeran Gowda Sangita Agarwal Anirbandeep Bose Uttam Bhaumik Debotri Ghosh Tapan Kumar Pal |
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Institution: | (1) Bioequivalence Study Centre, Department of Pharmaceutical Technology, Jadavpur University, Kolkata, 32, India |
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Abstract: | A bioanalytical method has been developed and validated for determination of pregabalin in human plasma. The analytical method
consists in the precipitation of plasma sample with trichloro acetic acid (20% v/v solution in water), followed by the determination of pregabalin by an LC-MS-MS method using gabapentin as internal standard.
Separation was achieved on a Gemini C18 50 mm × 2.0 mm (3 μm) column with an isocratic mobile phase consisting of methanol–water (98:2, v/v) with 0.5% v/v formic acid. Protonated ions formed by a turbo ionspray in positive mode were used to detect analyte and internal standard.
The MS-MS detection was by monitoring the fragmentation of 160.2→55.1 (m/z) for pregabalin and 172.2→67.1 (m/z) for gabapentin on a triple quadrupole mass spectrometer. The assay was calibrated over the range 0.1–15.0 μg mL−1 with correlation coefficient of 0.9998. Validation data showed intra-batch (n = 6) CV% ≤ 6.89 and RE (%) between −4.17 and +3.08 and inter-batch (n = 18) CV% < 9.09 and RE (%) between −3.0 and +10.00. Mean extraction recovery were 80.45–89.12% for three QC samples and 87.56%
for IS. Plasma samples were stable for three freeze–thaw cycles, or 24 h ambient storage, or 1 and 3 months storage at −20 °C.
Processed sample (ready for injection) were stable up to 72 h at autosampler (4 °C). This method has been used for analyzing
plasma samples from a bioequivalence study with 18 volunteers. |
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Keywords: | Column liquid chromatography– mass spectrometry Bioequivalence study Turbo ionspray Pregabalin |
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