Objective To establish and validate liquid chromatography-tandem mass spectrometry (LC-MS/MS) and gas chromatography-mass spectrometry (GC-MS) methods for the determination of potential mutagenic impurities, namely methyl p-toluenesulfonate, propyl p-toluenesulfonate, isopropyl p-toluenesulfonate, 2-(thiophen-2-yl) ethyl p-toluenesulfonate, and 1-iodopropane, in rotigotine active pharmaceutical ingredient (API).

Methods The LC-MS/MS method was used for the determination of four p-toluenesulfonate esters: methyl p-toluenesulfonate, propyl p-toluenesulfonate, isopropyl p-toluenesulfonate, and 2-(thiophen-2-yl) ethyl p-toluenesulfonate. The separation was performed on an InfinityLab Poroshell 120 EC-C₁₈ column (150 mm×3.0 mm, 2.7 μm) with a mobile phase consisting of 5 mmol/L ammonium acetate aqueous solution and methanol using gradient elution at a flow rate of 0.5 mL/min. The column temperature was maintained at 35 ℃. MS was performed in positive ion mode using an electrospray ionization (ESI) source with multiple reaction monitoring (MRM). The GC-MS method was used for the determination of 1-iodopropane. Headspace injection was employed, and the separation was carried out on a DB-624 column (30 m × 0.25 mm, 1.4 μm). MS was performed using an electron ionization (EI) source in selected ion monitoring (SIM) mode, with the quantification ion set at m/z 170.

Results The method validation results demonstrated good linearity for all impurities within the tested concentration ranges (r ≥ 0.995 9). The limits of detection (LOD) for the four p-toluenesulfonate esters ranged from 0.0398 to 0.0406 μg/mL and the LOD for 1-iodopropane was 0.1005 μg/mL. The recoveries of all impurities ranged from 80.7% to 122.0%, and the RSDs for precision were all below 5.0%. The test results of three batches of rotigotine API showed that all target mutagenic impurities were below the LOD, complying with the limit calculated based on the threshold of toxicological concern (TTC) (93 ppm).

Conclusion The two analytical methods established in this study, based on LC-MS and GC-MS technologies, are highly sensitive, specific, accurate, and reliable. They can be used for the trace-level determination of multiple potential mutagenic impurities in rotigotine API, providing an important technical means for its quality control.

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