Identification of the related substances in fasudil hydrochloride with hyphenated techniques

The study aims to identify the related substances in fasudil hydrochloride by hyphenated techniques.  A WondaSil C₁₈ (250 mm × 4.6 mm, 5 μm) column was used for the separation of the related substances with a mixture of methanol and ammonium acetate buffer solution as the mobile phase by gradient elution.  The structures of the related substances were speculated by electrospray positive ionization LC-TOF/ MS accurate ion mass and MS/MS determination and elucidation, and verified further through synthesis and spectroscopic analysis.  Fasudil hydrochloride and the related substances were separated under the established HPLC condition.  Three related substances in fasudil hydrochloride were characterized by hyphenated techniques.  The hyphenated LC-MS method is useful for the identification of related substances in fasudil hydrochloride and the results obtained are valuable for its manufacturing process and quality control.