Literature reveals that potentiometric,[2] spectrophotometric,[6,7] and chromatographic methods[8�C15] have been reported for their individual analysis, along with other combinations in pharmaceutical formulation and biological fluids. However, no method has been www.selleckchem.com/products/Rapamycin.html reported for their simultaneous determination in their combined fixed dose tablet formulation, to the best of our knowledge. The aim of the present study is to develop a feasible, rapid, sensitive, and specific HPLC method for the analysis of the investigated drugs. Figure 1 Structure of tolperisone hydrochloride Figure 2 Structure of etodolac MATERIALS AND METHODS Chemical The TOLP and ETD standard was obtained from Lupin Pharmaceuticals Ltd., Mumbai. The tablet formulation was obtained as a gift sample from Zydus Cadila Health Care Ltd.

All the chemicals used were of Analytical Reagent grade and the solvents were of HPLC grade. HPLC grade water, methanol, acetonitrile, orthophosphoric acid, and tri-ethylamine (TEA) were purchased from S.D Fine Chemicals, Mumbai, India. Apparatus Separation was performed with a Shimadzu LC_10_AT, equipped with a Rheodyne injector valve with a 20.0 ��l loop and a UV / VIS detector, and the PDA (SPD_M_10_A VP) detector operated at 257 nm. The Class-VP software was applied for data collection and processing. A Chemline digital pH-meter was used for pH measurements. Chromatographic conditions A Phenomenex C18 column (150 mm �� 4.6 mm, 5 ��) was used in this study. The mobile phase was a phosphate buffer (KH2PO4) of pH 5.5 : Methanol : Acetonitrile : Tri-ethylamine (40 : 40 : 20 : 1.

5) adjusted to pH with orthophosphoric acid. The flow rate was 1.0 mL / minute and UV detection was performed at 257 nm by UV detector at 257 nm. The mobile phase was shaken on an ultrasonic bath for 30 minutes. The resulting transparent mobile phase was filtered through a 0.45-��m membrane filter (Millipore, Ireland). Preparation of standard stock solutions Stock solutions containing 150 ��g / ml of TOLP and 400 ��g / ml ETD were prepared in methanol and were used as working solutions. The solutions were kept in tight closed containers and were found to be stable for at least one week, when kept in the refrigerator. Study of experimental parameters Different experimental parameters including, mobile phase composition, detection wavelength, and flow rate were intensively studied, in order to specify the optimum conditions for the assay procedure.

The variables were optimized by changing each, in turn, while keeping all others constant. Construction of the calibration curve Aliquots of the standard solutions covering the final working concentration range of 3.0 �C 21.0 ��g / ml for TOLP and 8 �C 56 ��g / ml ETD were transferred into a series Drug_discovery of 10 ml volumetric flasks and diluted with the de-gassed mobile phase up to the mark. Aliquots of 20 ��l were injected (n = 6) and eluted with the mobile phase under the reported chromatographic conditions.