Isolation and characterization of thermal degradation impurity in brimonidine tartrate by HPLC, LC-MS/MS, and 2DNMR

One potential unknown impurity was detected during the analysis of stability batches of brimonidine tartrate (BMT) in the level ranging from 0.03 % to 0.06 % by high-performance liquid chromatography (HPLC). Based on the liquid chromatography-mass spectrophotometry (LC-MS) analysis, the unknown impurity structure was presumed as 3,6,11,13,16-pentaazatetracyclo [8.6.0.0(2),(7).0(12),(16)] hexadeca-1,3,5,7,9,12-hexaene. The proposed structure was elucidated, after its isolation using preparative liquid chromatography from the impurity enriched reaction crude sample, using analytical applications such as 1D NMR (H-1,C-13 and DEPT-135), 2D NMR (HMBC and COSY), high-resolution mass spectrometry (HRMS) and infrared spectroscopy (IR). The unknown impurity was prepared from brimonidine by following Ull-man coupling reaction in the presence of CuBr2 in gram scale with optimum purity to use further in analytical developments. The identification, structural elucidation and synthesis of unknown degrada-tion impurity such as BMT-cyclized impurity, and HPLC method validation were reported for the first time in this paper. (C) 2021 Elsevier B.V. All rights reserved.