Stability-Indicating Related Substances HPLC Method for Droxidopa and Characterization of Related Substances Using LC-MS and NMR.
Stress degradation studies using high-performance liquid chromatography (HPLC) was performed and validated for Droxidopa (L-DOPS). Droxidopa was susceptible to acid hydrolysis (0.1 N HCl), alkaline hydrolysis (0.15 N NaOH) and thermal degradation (105°C). It was found to be resistant to white light, oxidation and UV light exposure (72 h). The thermal, acid and alkali degradation impurities were detected with the retention time (RT) of 12.7, 19.25 and 22.95 min. Our HPLC method detected process impurities (2R,3R)-2-amino-3-(3,4-dihydroxyphenyl)-3-hydroxypropionic acid (Impurity H), N-Hydroxypthalimide (Impurity N), (2R,3S)-2-amino-3-(benzo[d][1,3]dioxol-5-yl)-3-hydroxypropionic acid (Impurity L) and L-threo n-phthaloyl-3-(3, 4-dihydroxyphenyl)-serine (Intermediate) with RTs of 3.48, 15.5, 25.76 and 28.0 min. The related substances were further characterized and confirmed by liquid chromatography-mass spectroscopy (LC-MS), and nuclear magnetic resonance spectroscopy analysis. Our HPLC method detected up to 0.05 µg/mL of Droxidopa with S/N > 3.0 and quantified up to 0.10 µg /mL of Droxidopa with S/N ratio > 10.0. Droxidopa was highly stable for 12 h after its preparation for HPLC analysis. Our newly developed HPLC method was highly precise, specific, reliable and accurate for the analysis of Droxidopa and its related substances.
Analytical Research and Development Division, Piramal Enterprises Limited, Chennai 600057, Tamil Nadu, India t.kumar@piramal.com.
Thangarathinam
T
Kumar
Department of Genetic Engineering, SRM University, Kattankulathur, Chennai 603203, Tamil Nadu, India.
Mohandass
M
Ramya
Department of Chemistry, St Josephs College, Tiruchirappalli 620001, Tamil Nadu, India.
S J
SJ
Arockiasamy Xavier
D002626:Chemistry, Pharmaceutical / Q000379:methods*; D002851:Chromatography, High Pressure Liquid; D002853:Chromatography, Liquid; D015103:Droxidopa / Q000737:chemistry*; D004340:Drug Contamination; D004355:Drug Stability; D009682:Magnetic Resonance Spectroscopy; D015203:Reproducibility of Results; D053719:Tandem Mass Spectrometry
D015103:Droxidopa