Doxorubicin, an anthracyline medication, is regularly used to treat a number of malignancies, including sarcoma, lung, breast, gastric, thyroid, non-Hodgkin's lymphoma, ovarian, Hodgkin's lymphoma, multiple myeloma, and childhood cancers. One unidentified doxorubicin impurity with an RRT of 1.23 and a concentration higher than 0.1% was discovered during the quality inspection of doxorubicin in bulk forms. By thoroughly analysing the 1D-NMR (1H) spectroscopic data and the LC-MS mass ionization spectrum data, the structure and molecular mass of the unknown doxorubicin impurity with an RRT of 1.23 were determined. From spectroscopic data and the LC-MS mass ionization spectrum data, it was also observed that doxorubicin and unknown doxorubicin impurity with an RRT of 1.23 are interconvertible.