Unveiling the skin-protective material basis of Coreopsis tinctoria Nutt.: Ethnopharmacology-Guided discovery of multi-target enzyme inhibitors using a ligand-fishing approach.

Coreopsis tinctoria Nutt. (commonly known as Kunlun snow chrysanthemum) is a traditional medicinal and edible plant distributed mainly in the high-altitude regions of Xinjiang, China. In traditional medical practices, the dried flower heads (capitula) of C. tinctoria have been widely consumed as an herbal tea for clearing heat, detoxifying, and alleviating inflammatory conditions. In traditional medical theory, the accumulation of "heat and toxins" is closely associated with skin disorders, oxidative damage, and premature aging. However, direct ethnobotanical evidence for its skin-specific or topical use remains limited. Therefore, the present study investigated the skin-protective potential of the dried flower heads of C. tinctoria as an ethnopharmacology-guided extension of its documented heat-clearing, detoxifying, and inflammation-related traditional uses. This study aimed to screen and identify active constituents from the dried flower heads of C. tinctoria and to evaluate their multi-target skin-protective activities related to melanogenesis, oxidative stress, and extracellular-matrix degradation, thereby providing pharmacological evidence for its potential skin-protective applications. A lysozyme-assisted ligand-fishing approach was established by immobilizing tyrosinase, elastase, and hyaluronidase onto lysozyme-modified filter paper to screen potential enzyme inhibitors from the dried flower extract. The isolated compounds were evaluated using α-MSH-induced B16 melanoma cells, UVB-induced HaCaT keratinocytes, and human dermal fibroblasts (HDFs). Western blot analysis, molecular docking, and network pharmacology were used to explore possible mechanisms and structure-related interactions. Fourteen compounds were successfully isolated and identified. In cell-free enzyme assays, quercetagetin-7-O-β-glucoside (2) and okanin (8) showed measurable tyrosinase inhibitory activity, with IC values of 78.3 ± 0.0 μM and 66.2 ± 0.4 μM, respectively. Naringenin (12) and luteolin (14) showed elastase inhibitory activity, with IC values of 162 μM and 143 μM, respectively. Compound 2 reduced melanin content in the α-MSH-treated B16 cells from 167% to 91.5%, compared to 100% of the untreated normal cells set as control group. Compound 8 exhibited strong radical-scavenging activity (for DPPH· IC = 4.23 μM). Several flavonoids also attenuated UVB-induced oxidative-stress-related changes in HaCaT cells and modulated photo-damage-related protein expression in HDFs. The identified multi-target enzyme inhibitors and bioactive flavonoids provide a material basis for the anti-photoaging properties of the flowers of C. tinctoria. These findings support an ethnopharmacology-guided extension of its traditional heat-clearing and detoxifying use into modern skin-protection research, while further studies are required to clarify compound uptake, skin permeability, and in vivo efficacy.