A highly sensitive LC-MS/MS method for the determination and quantification of four mutagenic nitrosamine impurities in bempedoic acid drug substance

This study reports the development of an enhanced liquid chromatography-tandem mass spectroscopy technique, which substantially contributes to advancements in analytical methodology. The developed method detects and quantifies four mutagenic nitrosamines N-nitroso-N-methyl-4-aminobutyric acid, N-nitrosodiethylamine, N-nitrosodiisopropylamine, and N-nitrosodibutylamine) in bempedoic acid (BA). The technique employs atmospheric pressure chemical ionization -tandem mass spectrometry, coupled with a gradient elution strategy, offering a significant advancement in the analysis of bempedoic acid. The mobile phase system comprises solvent A, which contains 0.1% formic acid in water, and solvent B, which contains 0.1% formic acid in methanol. The degassed mobile phase is pumped at a flow rate of 0.5 mL/min, while the column temperature is maintained at 40°C throughout the analytical run to ensure consistent chromatographic performance. Analytical separation was performed using an X-Bridge® Phenyl of dimension 150 × 4.6 mm, 3.5 µm high performance liquid chromatography (HPLC) column, while the detection and quantification of four nitrosamine impurities were carried out using positive-mode atmospheric pressure chemical ionization in conjunction with multiple reaction monitoring. The developed method was validated according to international council for harmonisation (ICH) guidelines to confirm its accuracy, precision, and overall reliability. The method demonstrated acceptable signal-to-noise ratios for both the limit of detection and limit of quantification, with a linear dynamic range spanning 4.42 to 106.92 ng mL- 1 and correlation coefficients exceeding 0.999 for all four nitrosamine impurities. The recoveries ranged from 83.7% to 107.4%, confirming the method’s suitability for detecting potential nitrosamine impurities in BA. It effectively supports quality control analysis, providing a reliable and robust method for the pharmaceutical and analytical chemistry fields.