2-CYANO-4-PYRIDINE CARBOXYLIC ACID METHYL ESTER is an intermediate of Topiroxostat CAS number is 94413-64-6. Isonicotinic acid Methyl ester and methyl isonicotinic acid N-oxide is major impurities during the synthesis, because it is similar to the main impurity Isonicotinic acid Methyl ester, there is a certain difficulty in the content detection. A high-performance liquid chromatography purity method for the complete separation of 2-CYANO-4-PYRIDINE CARBOXYLIC ACID METHYL ESTER CAS number is 94413-64-6 with adjacent impurity peaks by isocratic elution and good peak shape was invented. The following steps are as below:
1. Preparing a test solution: taking 2-CYANO-4-PYRIDINE CARBOXYLIC ACID METHYL ESTER and dissolving it in a mobile phase or methanol to prepare a solution having a concentration of 0.5 mg/ml to 1.2 mg/ml as a test solution;
2. Preparation of impurity reference solution: taking methyl isonicotinic acid and methyl isonicotinic acid N-oxide, and dissolving in mob…