Development of an LC-MS method for ultra trace-level determination of 2,2,6,6-tetramethylpiperidine-1-oxl (TEMPO), a potential genotoxic impurity within active pharmaceutical ingredients.
TEMPO (2,2,6,6-tetramethylpiperidine-1-oxyl) is a stable free radical which has been widely used for various research and industrial applications, including the manufacture of many active pharmaceutical ingredients. TEMPO has been identified as a potential genotoxic impurity resulting in the need for analytical methodology to accurately determine its level at several orders of magnitude less than typical impurity quantitation limits. TEMPO can undergo disproportionation to form both oxidized and reduced TEMPO, making individual determination unreliable. To overcome this challenge, all TEMPO related species were converted to the reduced form through reduction with sodium ascorbate. Given the ultra-trace (0.5 ppm) level requirements and the lack of UV response in the reduced form, a single quadrupole mass spectrometer (MS) was utilized. In order to implement a highly sensitive MS method in a GMP environment, several approaches were employed to optimize accuracy and robustness including: internal standard correction for drift elimination, six-level standard addition to reduce matrix effects, and weighted linear regression to cover a broad analytical range. The method was fully validated according to ICH guidelines. The method is specific, linear, accurate, precise, and robust within a range of 0.5-100 ppm.
Merck Research Laboratories, 126 E. Lincoln Ave, Rahway, NJ 07065, USA. Electronic address: Justin.pennington@merck.com.
Justin
J
Pennington
Merck Research Laboratories, 126 E. Lincoln Ave, Rahway, NJ 07065, USA. Electronic address: Ryan.cohen@merck.com.
Ryan D
RD
Cohen
Merck Research Laboratories, 126 E. Lincoln Ave, Rahway, NJ 07065, USA. Electronic address: ye.tian5@merck.com.
Ye
Y
Tian
Merck Research Laboratories, 126 E. Lincoln Ave, Rahway, NJ 07065, USA. Electronic address: Fabien.boulineau@merck.com.
Fabien
F
Boulineau
D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D003497:Cyclic N-Oxides / Q000737:chemistry*; D005609:Free Radicals; D006863:Hydrogen-Ion Concentration; D057230:Limit of Detection; D016014:Linear Models; D013058:Mass Spectrometry / Q000379:methods*; D010100:Oxygen / Q000737:chemistry; D004364:Pharmaceutical Preparations / Q000032:analysis; D010880:Piperidines / Q000032:analysis; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013696:Temperature
D003497:Cyclic N-Oxides; D005609:Free Radicals; D004364:Pharmaceutical Preparations; D010880:Piperidines; C551336:2,2,6,6-tetramethylpiperidide; D010100:Oxygen; C003959:TEMPO
LC–MS; PGI; Potential genotoxic impurity; TEMPO; Trace-level analysis