NOVEL RP-HPLC METHOD FOR THE ... - Ijsidonline.info

Thamma Narendra Kumar et al., IJSID, 2011, 1 (3), 395-421MATERIALS AND METHODSChemicals and reagentsTablets and Standards components were supplied by Medreich India limited. All solvents were HPLC gradeand High purity water was prepared by using Millipore Milli-Q plus water purification system (Millipore, Milford,MA, USA).Equipment:The Shimadzu and agilent make HPLC systems were used for this study and these instruments consists of apump, auto sampler and a PDA detector. The output signal was monitored and processed by using WatersEmpower software.Chromatographic conditionsThe method was developed using an Atlantis dC18, 250 × 4.6 mm; 5.0 µm column. An isocratic elution ofmobile phase is used, which is a mixture of buffer (dissolved 2.2 g of 1-Heptane sulphonic acid sodium salt in 1000ml of Milli-Q water and adjusted to pH 5.0 with diluted Orthophosphoric acid) and Acetonitrile in the ratio of 750and 250 respectively. Then filtered the mobile phase through 0.45 µm PVDF filter and degassed. The flow rate ofthe mobile phase was 1.0 ml/min. The column temperature was maintained at 25 o C and the wavelength wasmonitored at 233 nm. The injection volume was 10 µl. The run time is fixed to 10 minutes.Preparation of standard solutionsA stock solution was prepared by transferring accurately weighed about 85 mg of MetforminHydrochloride working standard into a 100 ml volumetric flask, added 40 ml of Methanol, sonicated for 5 min todissolve, then made up to the volume with purified water and mixed well. Transferred 5 ml of above standardstock solution into a 50 ml volumetric flask and made up to the volume with purified water.Preparation of sample solutionsTwenty tablets were weighed, and then crushed in to a fine powder using a mortar and pestle. Weighedabout 1650 mg of tablet powder (equivalent to 850 mg of Metformin Hydrochloride) then transferred in to a 500ml volumetric flask, 200 ml of Methanol was added, sonicated for 20 min, then added 150 ml of purified water,sonicated for 15 minutes with intermittent shaking by maintaining bath temperature 20°C to 28°C, cooled it, thenmade up to the volume with purified water and mixed well. This solution was centrifuged at 3000 rpm for 10 minand collected the supernatant clear solution. Transferred 5 ml of supernatant solution into a 100 ml volumetricflask and made up to the volume with purified water.Calculation of % analyte:Calculation for % Metformin Hydrochloride:TR W 5 500 100 P-----------x-----------x-----------x-----------x-----------x-----------x ASR 100 50 TW 5 LInternational Journal of Science Innovations and Discoveries Vol 1, Issue 3, November-December 2011397