Abstract: Peucedani Radix, the dried root of Peucedanum praeruptorum Dunn, is traditionally reputed for its efficacy in descending qi to resolve phlegm and dispelling wind to clear heat. It finds extensive clinical application in treating conditions, such as phlegm-heat cough and asthma, wind-heat cough, chronic bronchitis, and upper respiratory tract infections. Modern pharmacological investigations reveal that Qianhu is rich in diverse chemical constituents, including coumarins, organic acids, volatile oils, phenanthraquinones, and sterols. Among these, coumarins are considered the primary active components, exerting significant beneficial effects, such as antitussive, antiasthmatic, cardiotonic effects, as well as preventive and therapeutic effects on cardiovascular diseases. However, the inherent complexity of Qianhu, coupled with the structural similarity, numerous isomers, and often trace abundance of active coumarin constituents, poses significant analytical challenges for their rapid and comprehensive characterization by conventional analytical methods. In this study, an integrated data-processing strategy facilitating rapid, chemical homologs-focused data filtering via molecular weight imprinting (MWI) and mass defect filtering (MDF) was proposed for the rapid screening of coumarin chemical components in Qianhu. High-resolution mass spectrometry data (m/z 100~1β000), including MS¹, MS², and retention time, were acquired for Qianhu and authentic compounds in positive ion mode using UPLC-MS. A comprehensive in-house coumarin chemical library from Qianhu was constructed by collecting literature and data from some available databases (e.g., ChemSpider, PubChem, HMDB). Furthermore, the characteristic mass spectrometric fragmentation rules for different structural types of coumarins were summarized. A two-dimensional scattering plot was generated, using retention time and exact m/z as the x and y coordinates, respectively. Based on the in-house library, red rectangular frames, 20 mDa wide and parallel to the x-axis, were applied to the scattering plot for MWI, to enable the preliminary screening of known coumarin components. Subsequent identification of target compounds was performed by integrating secondary fragment ion information, chromatographic retention behavior, and the established mass spectrometry fragmentation rules. Concurrently, a“five-point”MDF strategy was applied for comprehensive screening of the mass spectrometry data. Each MS¹ signal was decomposed into integer and decimal parts to construct a two-dimensional scattering plot. To precisely define the screening range, five structurally representative coumarin compounds from the in-house library were selected as the vertices, including a (m/z 163.039β0, C₉H₆O₃), b (m/z 207.028β8, C₁₀H₆O₅), c (m/z 441.115β6, C₂₁H₂₂O₉), d (m/z 625.210β3, C₂₇H₃₈O₁₅), and e (m/z 329.174β7, C₂₀H₂₄O₄), thus establishing the pentagonal frame. This specifically designed pentagonal frame proved effective in filtering out matrix interference and capturing structurally similar potential target compounds. Structural elucidation of these target compounds was then completed by integrating diagnostic ions, characteristic fragmentation patterns, and comparison with authentic compounds. Utilizing this integrated approach, 105 coumarin compounds were rapidly screened and identified from Qianhu, including 30 simple coumarins, 30 furanocoumarins, and 45 pyranocoumarins. Significant isomerism was observed. Furthermore, for the characterization of isomers within Qianhu, a precise differentiation strategy for structurally similar angular pyranocoumarin isomers was achieved based on specific cleavage pathways of substituents at particular positions (e.g., C-3' and C-4' positions). For instance, in angular pyranocoumarins with substituents at both C-3' and C-4' positions, the C-4' side chain preferentially cleaves due to its instability. The combined strategy of MWI and MDF significantly enhanced the identification efficiency of analogous components within complex matrices. Above all, this study provides a promising approach for the rapid screening and precise identification of diverse chemical constituents in, but not limited to, medicinal plants, facilitating their quality control and pharmacological research.

Key words: Peucedani Radix, coumarins, molecular weight imprinting (MWI), mass defect filtering (MDF), ultra-performance liquid chromatography-high resolution mass spectrometry (UPLC-HRMS)