Characterization of main degradation products from dendrobine under stress conditions by multistage cleavage of UPLC-ESI-IT-TOF

Dendrobine is a sesquiterpene alkaloid primarily used in the treatment of inflammatory diseases, immune system disorders, and conditions related to oxidative stress. To understand the possible degradation pathways of dendrobine for its quality control, we conducted an in-depth investigation of its degradation products using forced degradation methods. The separation of dendrobine and its degradation products was achieved on a Shim-pack XR-ODS III (75 mm × 2 mm, 1.6 µm) column with a methanol-water mixture as the mobile phase under isocratic conditions, the isolated compounds were examined in positive ion mode with an ion trap-time of flight mass spectrometer (IT-TOF). In order to obtain in-depth structural information about the degradation products, mass spectrometry was performed using a five-stage fragmentation approach. This method allowed for thorough structural clarification via several rounds of selective fragmentation and high-resolution detection. System control and data acquisition were managed using LCMSsolution 3.81 software. The results showed that dendrobine undergoes significant degradation under oxidative, acidic, hydrolytic and thermal conditions, resulting in the formation of several degradation products with notable structural changes. Under oxidative conditions, dendrobine primarily generates two degradation products with mass increases of 16 Da and 32 Da, indicating mono-oxidation and di-oxidation reactions. Acidic degradation led to the identification of three degradation products, including a novel compound with an 18 Da mass increase, suggesting potential hydrolysis or dehydration reactions. Hydrolytic and thermal conditions resulted in the formation of two and three degradation products, respectively, with structural changes indicating possible molecular cleavage and reorganization mechanisms. In contrast, dendrobine exhibited strong stability under alkaline and photolytic conditions, with no significant degradation products detected. Detailed characterization of the degradation products via multi-stage mass spectrometry revealed key reaction pathways and mechanisms involved in dendrobine's degradation, providing critical insights for assessing its chemical stability and optimizing storage conditions.