A fast, validated UPLC method coupled with PDA-QDa detectors for impurity profiling in betamethasone acetate and betamethasone phosphate injectable suspension and isolation, identification, characterization of two thermal impurities.

For impurity profiling of betamethasone acetate and betamethasone phosphate injectable suspensions, a quick, verified stability indicating UPLC technique incorporating the detectors PDA-QDa had been established. This method with an analysis time of 12min could able to separate all possible degradation impurities. Two of the thermal impurities have been identified in positive mode of detection by using QDa detector and isolated by using preparative HPLC. The method works at a flow rate of 0.5mL/min in column: Poroshell 120 EC C18 (100×2.1)mm, 1.9µm, maintained temperature precisely at 40°C. The M/Z values in ESI positive mode for the two new degradation impurities have been identified (M+H) as 393.22 (DP1), 363.17 (DP2) and confirmed by 1H NMR. The approach was also verified in accordance with the rules of ICH Q2 (R1). From LOQ quantity value to 150% quantity of specified concentration (2% for betamethasone and 0.5% for other impurities), the technique of UPLC-PDA-QDa was proven to be linear and accurate. Precision and ruggedness results showed˂5% RSD. Accuracy results showed more than 95% recovery from LOQ till 150% of impurity specification. This UPLC-PDA-QDa methodology was found specific, precise, stable and robust for quantification of all possible degradation impurities. The proposed method has been transferred to quality control laboratories to access the impurity profile during product storage.

Assay of tirofiban and identification of oxidative impurity in aqueous injection by using UPLC-PDA/QDa detectors.

The basic objective of this study is to propose a short, reliable, mass compatible ultra-performance liquid chromatography (UPLC) method to confirm the identity of impurities and to estimate the assay and purity of Tirofiban simultaneously in aqueous injection (5mg/100mL bag). Aqueous formulations are susceptible to oxidation, hence the possible oxidative degradation impurities of Tirofiban were studied in this experiment by using UPLC coupled with photodiode array/Quadrupole Dalton Analyzer (PDA/QDa) detectors. The required separations were achieved in the column: ACQUITY HSS T3 (100×2.1) mm, 1.7µm, operated at 30°C by using 0.02% Triethyl amine (TEA) in water, pH 2.8 with formic acid as solution-A and 0.1% formic acid in 9:1 acetonitrile, water as solution-B. Binary gradient flow is delivered at the rate of 0.5mL/min and the detection of impurities specifically carried out at 227nm using empower3 software. RP-UPLC/PDA with QDa detector was used for the experiment. The method was linear and accurate from the concentrations: 0.04 to 0.38µg/mL for impurity-A and 0.04 to 75µg/mL for Tirofiban. The major unknown degradation impurity generated during the oxidative degradation has been identified as N-oxide derivative (Impurity-B) [(M+H)+ 455.1] by using QDa detector operated in an electro spray positive ion mode by applying a voltage of 0.8kV. This method was further validated as per ICH Q2 (R2) guidelines. Hence, the proposed method is said to be a fast, sensitive and comprehensive technique, which could give a clear idea about the assay and impurity profile of Tirofiban injection.