Rapid Determination of Fingolimod Hydrochloride-Related Substances and Degradation Products in API and Pharmaceutical Dosage Forms by Use of a Stability-Indicating UPLC Method

Abstract

Ultra-performance liquid chromatography coupled with photodiode-array detection has been used to develop a simple, sensitive, and reproducible reversed-phase method for quantitative determination of fingolimod hydrochloride and all possible process-related impurities. Chromatographic separation was achieved on a Waters Acquity BEH C18 (100 mm × 2.1 mm, 1.7 µm) column. The mobile phase was a gradient prepared from potassium dihydrogen phosphate (20 mM) containing 0.1 % (v/v) triethylamine and adjusted to pH 6.5 with trifluoroacetic acid (component A) and 85:15 (v/v) acetonitrile–water (component B); the gradient program (time (min)/% B) was: 0.01/20, 2.0/20, 6.0/75, 9.0/90, 12.0/90, 14.0/20, 16.0/20; the run time was 16 min and fingolimod hydrochloride and its six impurities were well separated. Eluting compounds were monitored at 220 nm. The method was validated for precision, specificity, linearity, limit of detection, limit of quantification, accuracy, and robustness in accordance with International Conference on Harmonization guidelines. Fingolimod hydrochloride was subjected to oxidative, acid, base, hydrolytic, thermal, and photolytic stress, and analysis was conducted to determine the amounts of related impurities formed.