Endometriosis is a chronic disorder and the present study investigated the therapeutic potential of quercetin, derived from Paeoniae Radix Alba and Hedysarum multijugum Maxim (BSHQ), by identifying bioactive ingredients and key biomarkers with a focus on their molecular interactions in the context of endometriosis. The present study identified 32 bioactive ingredients from BSHQ, linked to 231 target genes. Gene Expression Omnibus datasets and Weighted Gene Co-expression Network Analysis were used to analyze differentially expressed genes and key co-expression modules. In vitro assays, including half-maximal inhibitory concentration, Cell Counting Kit-8 and 5-ethynyl-2'-deoxyuridine assays, assessed the impact of quercetin on cell proliferation in endometriosis cell lines. Molecular docking, cellular thermal shift assay (CETSA) and surface plasmon resonance (SPR) analyzed the interaction of quercetin with biomarkers. The present study identified 16 BSHQ-related genes. Single-cell RNA sequencing analysis showed distinct expression patterns of these genes in various cell types. Molecular docking confirmed strong binding between quercetin and these biomarkers. In vitro assays revealed quercetin markedly inhibited the proliferation of endometriosis cell lines, reducing cell proliferation and serpin family E member 1 (SERPINE1) expression. CETSA showed that quercetin can directly bind to SERPINE1 protein and enhance its thermal stability against temperature-induced degradation, a post-translational effect independent of expression regulation, and SPR analysis further supported the direct interaction between quercetin and SERPINE1. Furthermore, SERPINE1 overexpression rescued the inhibitory effects of quercetin on proliferation, highlighting its critical role in mediating the effects of quercetin in endometriosis. The present study underscored SERPINE1 as a pivotal biomarker in endometriosis and revealed that quercetin effectively modulated key disease mechanisms, offering promising therapeutic potential. These findings provide a solid basis for future research aimed at exploring the role of quercetin in the treatment of endometriosis.