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DEVELOPMENT AND VALIDATION OF A HPLC METHOD FOR IN-VIVO STUDY OF DICLOFENAC POTASSIUM

Keywords: Diclofenac Potassium , HPLC , Mixed column , Method Validation , In-vivo study

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Abstract:

A High Performance Liquid Chromatographic method was developed for determination of Diclofenac Potassium in in-vivo study during bioavailability study. The chromatographic separation was conducted on Shimadzu Prominence LC 20 connected with PDA detector; using mixed column ODS/Cyano; ACE, (100 x 4.6 mm, 5 μm). The mobile phase was isocratic consisted of Methanol: 50 mM potassium dihydrogen phosphate buffer, in ratio of (50 : 50 v/v) and was delivered to the system at a flow rate of 1.5 ml/min, with an injection volume of 20 μl and the detection wavelength (λ max) was 280 nm. Diazepam was used as internal standard. All assays were performed at ambient conditions. The calibration curve of Diclofenac Potassium in plasma was linear with correlation coefficient (r2) = 0.9996; over a concentration range of 0.25 - 3 ug/ml. The retention times for Diclofenac Potassium and the internal standard (Diazepam) were found to be 5.95 and 7.90 minutes, respectively. The mean recovery was found to be 90.57%.The relative standard deviation (RSD) was found to be < 2. The proposed method was validated and successfully applied to the determination of Diclofenac Potassium in human plasma for bioavailability studies.

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