Structure identification of related substances in doxycycline hyclate by 2D-LC-IT-TOF/MS / 中国药学杂志

ABSTRACT

OBJECTIVE: To rapidly identify the related substances in doxycycline hyclate tablets and investigate the possible degradation pathways of doxycycline hyclate solution by 2D-LC-IT-TOF/MS. METHODS: Firstly, the chromatography was carried out using the 1stD InertSustain ™C18 (4.6 mm×150 mm, 5 μm) column and a mobile phase containing a mixture of buffer solution (0.25 mol·L-1 ammonium acetate solution-0.1 mol·L-1 ethyldiaminetetraacetic acid disodium; triethyiamine=100101)-acetonitrile (8515). Solution pH was adjusted to 8.8 with glacial acetic acid. Then a InertSustain C18 (2.1 mm×50 mm, 2 μm) column was used with 0.1% formic acid-water as mobile A and 0.1% formic acid-acetonitrile as mobile B in a gradient elution mode in 2ndD. Electrospray ionization (ESI) source was tested in both positive and negative ion modes. Nebulized gas flow was 1.5 L·min-1. Dry gas flow was 10 L·min-1. The desolvation tube temperature was kept at 200℃. Related substances were characterized according to multi-level MS behaviors. The 2D-LC-IT-TOF/MS method was employed to identify the structures of impurities in forced degradation solutions and illuminate the degradation pathways of doxycycline hyclate solution. RESULTS: A total of eight related substances were detected in doxycycline hyclate tablets. Four of them had a content of more than 0.1%, and their structures were identified to be 4-epidoxycycline, metacycline, β-epidoxycycline and 2-acetyl-2-decar-bamoyldoxycycline. The solution of doxycycline hyclate easily degraded under alkaline condition and generated an open loop compound taken off the keto group. The solution was sensitive to heat, and generated 4-epidoxycycline with a little amount of 4-epi-6-epidoxycycline; but the solution was stable under illumination and acidic condition. CONCLUSION: The established method is suitable for rapid identification of impurities in doxycycline hyclate, which can be applied as a useful analytical tool for quality control and drug process optimization of doxycycline hyclate.