“DEVELOPMENT AND VALIDATION OF STABILITY INDICATING METHOD FOR SIMULTANEOUS ESTIMATION OF CEFTRIAXONE AND TAZOBACTAM INJECTION USING RP-UPLC METHOD”
2.1 OBJECTIVE
To develop and Validate Stability Indicating Method for simultaneous estimation of Ceftriaxone and Tazobactam in their combined dosage form by RP-UPLC method in various stress condition like, alkaline, acidic, oxidative, thermal and photo degradation.
2.4 METHODOLOGY
2.4.1 Chromatographic Condition
Column Separation was achieved by using Acquity UPLC BEH C-18 (1.7 μm), 2.1 X 100 mm as stationary phase. Flow rate is 0.3 ml/min. Quantification was achieved of of Ceftriaxone and Tazobactam at 230 nm with PDA detector. Column Temperature at 25 ± 2°C temperature condition and 1 μl injection volume. 2.4.2 Experimental work Mobile Phase: Solution A: Accurately weighed and dissolved about 3.5 gm of Potassium di-hydrogen ortho-phosphate anhydrous and 14.5 gm of Disodium hydrogen phosphate anhydrous in 1000 ml of water for pH 6.5 Buffer solution. pH of 6.5±0.2 was adjusted by using diluted orthophosphoric acid. Accurately weighed and dissolved about 20.5 gm of Citric acid in 1000 ml of water for pH 5.0 Buffer solution. pH of 5.0±0.2 was adjusted by using NaOH solution. Water, pH 6.5 buffer, pH 5.0 buffer and acetonitrile taken in the ratio 600:180:20:200 (v/v) and mix well. Solution B: Accurately weighed and dissolved 4.0 gm of Tetradecyl ammonium bromide and 4.0 gm of Tetraheptyl ammonium bromide in 500 ml of acetonitrile sonicated to dissolve and made up to 1000 ml with acetonitrile and mix well. Volume of solution (A) and Solution (B) taken in the ratio 65:35 (v/v) and mixed well and filter through 0.45 µm membrane filter and degas for 10 minutes. Standard Preparation: An accurately weighed Ceftriaxone (40 mg) and Tazobactam (5 mg) were transferred to 100 ml volumetric flask, dissolved in 50 ml with Mobile phase and diluted up to mark with Mobile phase to get 400 μg/ml solution of Ceftriaxone and 50 μg/ml solution of Tazobactam. Sample Preparation: Take about 55 mg (Ceftriaxone sodium equivalent to Ceftriaxone 40 mg and Tazobactam sodium equivalent to Tazobactam 5 mg) of powder and transferred to 100 ml volumetric flask, dissolved in Mobile phase (50 ml) sonicated for 30 min and dilute up to the mark with Mobile phase. The solution was filtered through Whatmann filter paper No. 41. The solution was diluted up to the mark with Mobile phase to get final working concentration of Ceftriaxone sodium equivalent to Ceftriaxone (400 μg/ml) …show more content…
• The % RSD values for Ceftriaxone and Tazobactam were found to be < 2 %, which indicates that the proposed method is repeatable. The low % RSD values of repeatability of assay (0.258-0.210 %), inter day (0.080-0.345 % and 0.045-0.321 %) and intraday (0.055-0.136 % and 0.055-0.090 %) variations for Ceftriaxone and Tazobactam, respectively (Table 2.03), reveal that the proposed method is precise.
• LOD values for Ceftriaxone and Tazobactam were found to be 0.005 μg/ml and 0.125 μg/ml, respectively and LOQ values for Ceftriaxone and Tazobactam were found to be 0.017 μg/ml and 0.416 μg/ml, respectively (Table 2.03). The results of system suitability testing are given in (Table 2.04).
• Peak of Degraded products were not interfering with the main drug peak of Ceftriaxone and Tazobactam. Thus, these degradation products have not been identified. The results of Degradation of Ceftriaxone and Tazobactam in different conditions are given in (Table
2.1 OBJECTIVE
To develop and Validate Stability Indicating Method for simultaneous estimation of Ceftriaxone and Tazobactam in their combined dosage form by RP-UPLC method in various stress condition like, alkaline, acidic, oxidative, thermal and photo degradation.
2.4 METHODOLOGY
2.4.1 Chromatographic Condition
Column Separation was achieved by using Acquity UPLC BEH C-18 (1.7 μm), 2.1 X 100 mm as stationary phase. Flow rate is 0.3 ml/min. Quantification was achieved of of Ceftriaxone and Tazobactam at 230 nm with PDA detector. Column Temperature at 25 ± 2°C temperature condition and 1 μl injection volume. 2.4.2 Experimental work Mobile Phase: Solution A: Accurately weighed and dissolved about 3.5 gm of Potassium di-hydrogen ortho-phosphate anhydrous and 14.5 gm of Disodium hydrogen phosphate anhydrous in 1000 ml of water for pH 6.5 Buffer solution. pH of 6.5±0.2 was adjusted by using diluted orthophosphoric acid. Accurately weighed and dissolved about 20.5 gm of Citric acid in 1000 ml of water for pH 5.0 Buffer solution. pH of 5.0±0.2 was adjusted by using NaOH solution. Water, pH 6.5 buffer, pH 5.0 buffer and acetonitrile taken in the ratio 600:180:20:200 (v/v) and mix well. Solution B: Accurately weighed and dissolved 4.0 gm of Tetradecyl ammonium bromide and 4.0 gm of Tetraheptyl ammonium bromide in 500 ml of acetonitrile sonicated to dissolve and made up to 1000 ml with acetonitrile and mix well. Volume of solution (A) and Solution (B) taken in the ratio 65:35 (v/v) and mixed well and filter through 0.45 µm membrane filter and degas for 10 minutes. Standard Preparation: An accurately weighed Ceftriaxone (40 mg) and Tazobactam (5 mg) were transferred to 100 ml volumetric flask, dissolved in 50 ml with Mobile phase and diluted up to mark with Mobile phase to get 400 μg/ml solution of Ceftriaxone and 50 μg/ml solution of Tazobactam. Sample Preparation: Take about 55 mg (Ceftriaxone sodium equivalent to Ceftriaxone 40 mg and Tazobactam sodium equivalent to Tazobactam 5 mg) of powder and transferred to 100 ml volumetric flask, dissolved in Mobile phase (50 ml) sonicated for 30 min and dilute up to the mark with Mobile phase. The solution was filtered through Whatmann filter paper No. 41. The solution was diluted up to the mark with Mobile phase to get final working concentration of Ceftriaxone sodium equivalent to Ceftriaxone (400 μg/ml) …show more content…
• The % RSD values for Ceftriaxone and Tazobactam were found to be < 2 %, which indicates that the proposed method is repeatable. The low % RSD values of repeatability of assay (0.258-0.210 %), inter day (0.080-0.345 % and 0.045-0.321 %) and intraday (0.055-0.136 % and 0.055-0.090 %) variations for Ceftriaxone and Tazobactam, respectively (Table 2.03), reveal that the proposed method is precise.
• LOD values for Ceftriaxone and Tazobactam were found to be 0.005 μg/ml and 0.125 μg/ml, respectively and LOQ values for Ceftriaxone and Tazobactam were found to be 0.017 μg/ml and 0.416 μg/ml, respectively (Table 2.03). The results of system suitability testing are given in (Table 2.04).
• Peak of Degraded products were not interfering with the main drug peak of Ceftriaxone and Tazobactam. Thus, these degradation products have not been identified. The results of Degradation of Ceftriaxone and Tazobactam in different conditions are given in (Table