RESUMO
The aim of this research work was to develop and validate a stability-indicating, single reversed-phase HPLC method for the separation of five impurities, including enantiomers, diastereomers, and degradation products in sacubitril-valsartan tablets. The method was developed using a Chiralcel OJ-RH column (150 × 4.6 mm, 5 µm) at 45°C with a gradient program of (T/%B) 0.01/25, 10.0/25, 25/38, 37.0/45, 39.0/25, and 45.0/25 at a flow rate of 0.8 ml/min. Mobile phase A consisted of 1 ml of trifluoroacetic acid in 1000 ml of Milli-Q water. Mobile phase B consisted of 1 ml of trifluoroacetic acid in a mixture of acetonitrile and methanol in the ratio of 950:50 (v/v). Sacubitril, valsartan, and their five impurities were monitored at 254 nm. Degradation was not observed when sacubitril-valsartan was subjected to heat, light, hydrolytic, and oxidation conditions. In acid degradation study (1 N HCl/60°C/2 h) impurity 1 (m/z 383.44) was formed, and in base degradation study (0.1 N NaOH/40°C/1 h) impurities 1 and 5 (m/z 265.35) were formed; both impurities were confirmed using LC-MS. The degradation products, enantiomers, and diastereomers were well separated from sacubitril and valsartan, proving the stability-indicating power of the method. The developed method was validated per the International Council for Harmonisation of Technical Requirements for Pharmaceuticals for Human Use guidelines. The inter- and intra-day percentage relative standard deviation for sacubitril, valsartan, and their five impurities was less than 5.2%, recovery of the five impurities was between 93 and 105%, and linearity was ≥0.999. The limit of detection was 0.030-0.048 µg/ml, and the limit of quantification was 0.100-0.160 µg/ml.
