Influence of Extraction Method on the Phytochemical Composition and Biofunctional Potential of Centaurea urvillei subsp. Stepposa : A Multi‐Assay and In Silico Perspective

ABSTRACT This study explores how extraction methods influence the phytochemical composition and bioactivity of Centaurea urvillei subsp. stepposa, an Anatolian endemic. Methanolic extracts were obtained via maceration (MAC‐ME), Soxhlet (SOE‐ME), and ultrasound‐assisted extraction (UAE‐ME). LC‐MS/MS profiling identified chlorogenic acid as the dominant phenolic (up to 2181 µg/g in MAC‐ME), while UAE‐ME enriched glycosylated flavones such as apigenin 7‐glucoside (862 µg/g). Although total phenolic and flavonoid contents were similar across extracts, antioxidant and enzyme inhibition assays revealed method‐specific differences. UAE‐ME showed the highest Relative Antioxidant Capacity Index (RACI = +0. 31), with strong metal‐chelating and phosphomolybdenum activities. MAC‐ME exhibited superior AChE inhibition (IC 50 = 1. 24 mg/mL), while UAE‐ME best inhibited α‐amylase (IC 50 = 1. 74 mg/mL). Molecular docking and MM/GBSA analyses confirmed strong interactions between chlorogenic acid, apigenin 7‐glucoside, and enzyme targets (Δ G MM/GBSA as low as –48. 54 kcal/mol). Correlation analyses highlighted flavone‐type compounds as major contributors to bioactivity. The results underscore the impact of extraction technique on biological potential and identify UAE as suitable for generating a relatively balanced bioactive profile. This work highlights C. urvillei subsp. stepposa as a botanically rich source of phenolic compounds with potential relevance for future nutraceutical and cosmeceutical research.