Based on offline two-dimensional liquid chromatography-mass spectrometry and mass defect filtering techniques: Comprehensive characterization of flavonoids in Scutellaria baicalensis Georgi

High-resolution mass spectrometry (HRMS) has become a core technology in the analysis of traditional Chinese medicine (TCM) components due to its high mass accuracy and abundant fragmentation information. Flavonoid components in TCM are characterized by structural diversity and significant content variations. However, HRMS still faces challenges such as difficulty in comprehensive capture, low precision in targeted screening and insufficient detection sensitivity, which restrict the elucidation of the chemical material basis of TCM. In this study, an offline two-dimensional liquid chromatography-high-resolution mass spectrometry (offline 2D-LC-HRMS) analysis platform was established. It integrated a quadrilateral mass defect filtering model to screen potential flavonoid components, and simultaneously proposed a data acquisition strategy of iterative precursor ion list combined with active exclusion for the comprehensive characterization of flavonoids in Scutellaria baicalensis Georgi. The quality of data annotation was enhanced via neutral losses, diagnostic ions and mass spectral fragmentation rules, combined with standard substances and database comparison. Finally, 138 flavonoid components were initially identified, providing reliable technical support for the analysis of complex flavonoids in TCM.