Rationale: Characterization of unknown impurities in degraded paromomycin 15% topical cream samples by ultra-high-performance liquid chromatography/tandem mass spectrometry (UHPLC/MS/MS) gas-phase fragmentation is described.
Methods: A volatile reversed-phase high-performance liquid chromatography/charged aerosol detection (RP-HPLC/CAD) method was developed and validated for fast and consistent analysis of the paromomycin potency and impurity values in drug substance and drug product samples. A Veo charged aerosol detector was chosen for routine analysis. The method shows separation of two paromomycin isomers as well as twelve unknown impurity peaks. The identity of the unknown impurities was investigated by LC/MS/MS using a quadrupole time-of-flight (QTOF) mass spectrometer.
Results: Six of the HPLC/CAD impurity peaks met the ICH Q3B(R2) threshold for identification and were investigated by coupling the developed LC method with a QTOF instrument. Structures for the impurities were proposed based on gas-phase fragmentation patterns of the paromomycin reference standard. Impurity fragmentation pathways were proposed based on literature of other structurally related aminoglycoside compounds.
Conclusions: Nine unknown paromomycin impurities were identified and the observed modifications included acetylation, carbamylation, gain of a hexose, and loss of glucosamine. The most abundant paromomycin modification was carbamylation by urea (a cream excipient) characterized by a 43 Da mass increase.
© 2019 John Wiley & Sons, Ltd.