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Objective: The main objective of this research work is to develop and validate a single reverse-phase high-performance liquid chromatography (RP-HPLC) method. This method should becapable of quantifying all the known, as well as other possible degradation impurities of sodium nitroprusside (SNP) in its injection formulation. Methods: Of allmethod development trails, we have observed better separations between known and degradation impuritiesin Inert sustain C18, (250 x 4.6) mm, 5 µm column at 30 °C temperature. Isocratic elution was carried out by using pH 8.6 phosphate buffer and acetonitrile in the ratio of 65:35 %v/v with a flow rate of 0.8 ml/min. The detection was carried out at 220 nm, with an injection volume of 10 µl. Results: In the proposed method, SNP was eluted at 22.5 min. Nitrite, nitrate, and ferrocyanide were linear from 0.25 to 37 μg/ml, ferricyanide was linear from 1.0 to 37 μg/ml, and SNP was linear from 0.75 to 37 μg/ml. The % RSD for six spiked samples (precision)was found to be less than 0.5 %. Accuracy was performed for known impurities from LOQ to 150 % for a 0.5 % specification level. The resultswere found to be in the acceptance range of 90-110 %. The LOQ concentration of nitrite, nitrate, and ferrocyanide was 0.25 μg/ml each,LOQ offerricyanide and SNP was found to be 1.0 μg/ml and 0.75 μg/ml, respectively. The SNP injection samples were exposed to different degradation conditions, and the results were found specific in the proposed methodology. Conclusion: The proposed RP-HPLC method is specific, precise, accurate, linear, stable, and robust for quantification of known and other possible degradation impurities in SNP injection formulation.

Objective: To propose a comprehensive, simple, and affordable RP-HPLC method for impurity profiling and characterization of unknown degradation products of thiamine hydrochloride injectable formulation. Methods: The chromatographic separation employs gradient mode using the octadecyl silane column using a mobile phase consisting of phosphate buffer with ion pair reagent, acetonitrile, and methanol delivered flow rate at 1.2 ml/min. The detection was carried out at 248 nm using empower software. LC-MS/MS/QTOF hyphenated technique was used for isolation and characterization of unknown degradation impurity. The performance of the method was systematically validated as per ICH Q2 (R1) guidelines. Results: Degradation product observed in accelerated stability was characterized by LC-MS/MS/QTOF hyphenated technique and found m/z value 351.1604 and postulated as an oxidative degradation product of thiamine due to excipient interaction. The validated method was sensitive, selective, and specific data proves the method is precise and accurate from LOQ to 150% level and results are within 95-108% and less than 4.5% RSD. The developed method is linear from 0.03-58.83 µg/ml with a correlation coefficient of more than 0.990 and LOD and LOQ value ranged from 0.03 to1.51 μg/ml. Conclusion: An efficient RP-HPLC method for impurity profiling of thiamine injectable formulation was successfully developed and unknown degradation product observed instability condition samples characterized by LC-MS/MS/QTOF technique. The validated method can be successfully employed for the impurity profiling of thiamine injectable in the quality control department.