title: Quantitative analysis of ripasudil hydrochloride hydrate and its impurities by reversed-phase high-performance liquid chromatography after precolumn derivatization: Identification of four impurities.
abstract: We report the development and validation of a stability-indicating reversed-phase high-performance liquid chromatography method with precolumn derivatization for the separation and identification of the impurities of ripasudil hydrochloride hydrate, a novel protein kinase inhibitor. 2,3,4,6-Tetra-O-acetyl-β-d-glucopyranosyl isothiocyanate was chosen as the derivatizing reagent and triethylamine was added as catalyst. 200 μL sample solution (1 mg/mL), 600 μL derivatizing reagent (1 mg/mL), and 200 μL triethylamine solution (1%, v/v) were mixed and reacted at 40°C for 30 min. The separation was achieved on an Inertsil C18 ODS-3 (250 mm × 4.6 mm, 5 μm) column using mobile phases including 10 mmol monopotassium phosphate buffer (pH 3.0) and methanol in gradient mode. The column temperature was adjusted at 25°C and the flow rate at 1 mL/min. The detection was carried out at 220 nm. Different precolumn derivatization conditions as well as the high-performance liquid chromatography conditions were optimized. Ripasudil hydrochloride hydrate and its four impurities were detected and quantitated, among which two new compounds were characterized. The proposed method was validated and proven to be selective, accurate, and precise and suitable for the quantitative analysis of ripasudil hydrochloride hydrate.
authors.affiliation: School of Pharmaceutical Sciences, Nanjing Tech University, Nanjing, P. R. China.
authors.forename: Wenkai
authors.initials: W
authors.lastname: Hui
authors.affiliation: College of Biotechnology and Pharmaceutical Engineering, Nanjing Tech University, Nanjing, P. R. China.
authors.forename: Lili
authors.initials: L
authors.lastname: Sun
authors.affiliation: School of Pharmaceutical Sciences, Nanjing Tech University, Nanjing, P. R. China.
authors.forename: Hui
authors.initials: H
authors.lastname: Zhang
authors.affiliation: School of Pharmaceutical Sciences, Nanjing Tech University, Nanjing, P. R. China.
authors.forename: Liang
authors.initials: L
authors.lastname: Zou
authors.affiliation: School of Pharmaceutical Sciences, Nanjing Tech University, Nanjing, P. R. China. qiaogenzou@hotmail.com.
authors.forename: Qiaogen
authors.initials: Q
authors.lastname: Zou
authors.affiliation: College of Biotechnology and Pharmaceutical Engineering, Nanjing Tech University, Nanjing, P. R. China.
authors.forename: Pingkai
authors.initials: P
authors.lastname: Ouyang
mesh_terms: D002851:Chromatography, High Pressure Liquid / Q000379:methods; D056148:Chromatography, Reverse-Phase / Q000379:methods*; D007546:Isoquinolines / Q000737:chemistry*; D017879:Isothiocyanates / Q000737:chemistry*; D047428:Protein Kinase Inhibitors / Q000737:chemistry*; D013449:Sulfonamides / Q000737:chemistry*
chemical_list: D007546:Isoquinolines; D017879:Isothiocyanates; C584679:K-115; D047428:Protein Kinase Inhibitors; D013449:Sulfonamides; C041943:2,3,4,6-tetra-O-acetylglucopyranosylisothiocyanate
keywords: Impurities; Precolumn derivatization; Quantitative analysis; Ripasudil hydrochloride hydrate