Isolation and identification of a major impurity in a new bulk drug candidate by preparative LC, ESI-MS(n), LC-MS-MS, and NMR.
A simple preparative liquid chromatography (LC) method is developed to isolate a major impurity in a new bulk drug candidate, 6-bromo-4-(carbamidinemethyl)-5-hydroxy-1-methyl-2-(phenylthiomethyl)-1H-indole-3-carboxylic acid ethyl ester hydrochloride monohydrate (carmidole). The carmidole solution for preparation is exposed to daylight before isolation. Based on the electrospray ionization (ESI)-mass spectroscopy (MS(n)) spectral data of the impurity fraction and carmidole, the impurity is preliminarily characterized as 6-bromo-4-(carbamidinemethyl)-5-hydroxy-1-methyl-2-methyl-1H-indole-3-carboxylic acid ethyl ester. LC-MS-MS is used to analyze a carmidole sample. The impurity, lyophilate, is obtained from the fraction of preparative LC, and the impurity standard is synthesized. By comparison of the retention times of high-performance liquid chromatography, ESI-MS(n), and (1)H-nuclear magnetic resonance of the impurity lyophilate with impurity standard and carmidole itself, the structure of the impurity is confirmed and its formation is discussed.
Li
Ning
N
Department of Analytical Chemistry, Shenyang Pharmaceutical University, Shenyang 110016, China.
Yang
Jing
J
Qin
Fang
F
Li
Famei
F
Gong
Ping
P
D000998:Antiviral Agents / Q000592:standards*; D002853:Chromatography, Liquid / Q000379:methods*; D004340:Drug Contamination; D006146:Guanidines / Q000737:chemistry / Q000302:isolation & purification* / Q000592:standards*; D007211:Indoles / Q000737:chemistry / Q000302:isolation & purification* / Q000592:standards*; D009682:Magnetic Resonance Spectroscopy; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods*
C518741:6-bromo-4-(carbamidinemethyl)-5-hydroxy-1,2-dimethyl-1H-indole-3-carboxylic acid ethyl ester; D000998:Antiviral Agents; D006146:Guanidines; D007211:Indoles; C518740:carmidole