UHPLC–MS/MS‐Based Identification of Active Constituents From Pinus yunnanensis Cones and the Multi‐Target Mechanism Exploration Against Breast Cancer

Pinus yunnanensis cones are a rich source of bioactive compounds; yet, the systemic exposure and mechanistic basis of their anti‐breast cancer activity remain unclear. Herein, we systematically identify the systemically absorbed components of P. yunnanensis cone extract and elucidate its anti‐breast cancer mechanisms using an integrated analytical and computational approach. An optimized ultra‐high performance liquid chromatography–tandem mass spectrometry (UHPLC–MS/MS) method was developed to overcome severe matrix interference and insufficient sensitivity, facilitating the rapid identification of active constituents in the crude extract. UHPLC–MS/MS‐based analysis of rat plasma and feces revealed 16 (14 prototypes and 2 metabolites) and 12 (10 prototypes and 2 metabolites) systemically absorbed components, respectively, derived from 28 identified compounds. Subsequent network pharmacology combined with molecular docking validation was used to screen the anti‐breast cancer active components, core targets, and key pathways. Integrated network pharmacology predicted coniferaldehyde, phyllanflexoid A, 3′,4′,5,5′,6,7‐hexamethoxyflavone, dihydrocaffeic acid, usnic acid, and 7‐oxodehydroabietic acid, as core components, targeting TP53, STAT3, PIK3R1, SRC, and PIK3CA and involving key pathways such as VEGF signaling pathway, aldosterone‐regulated sodium reabsorption, prolactin signaling pathway, PD‐L1/PD‐1 checkpoint pathway in cancer, and ErbB signaling pathway. This study establishes a robust UHPLC–MS/MS platform for the quality control of P. yunnanensis cones and provides critical insights into their translational potential for anti‐breast cancer drug development and advancing separation science applications in natural product research.