Exploring the phytochemical profile, antioxidant activity, and anticancer potential of Achillea millefolium extracts: In-vitro and in-silico investigation

Abstract

Recently, research has been focused on exploring bioactive compounds from natural resources for cancer treatment and management. We investigated the possible anticancer activity of Achillea millefolium L. (A. millefolium) extracts on liver cancer (HepG2) and non-cancerous cells (BHK) as an alternative anticancer therapy. In the current research, AME (ethanolic extract of A. millefolium) extract has higher phenolics, flavonoids, and tannins content, i.e. (232.99 mgGAE/g, 168.25 mgQE/g, and 46.05 mgTAE/g), respectively, than AMH (n-hexane extract of A. millefolium) extract. The same extract exhibited higher antioxidant activity with less IC₅₀ values of 33.64, 52.15, 68.38, and 70.71 μg/mL using 2,2-diphenyl-picryl hydrazyl radical, nitric oxide, hydrogen peroxide_, and superoxide anion protocols, respectively, in comparison to another extract. GC/MS investigation of A. millefollium extracts identified a wide array of bioactive compounds. Moreover, the antiproliferative activity of both extracts was determined using MTT, crystal violet, and annexin V/PI staining assays on HepG2 and BHK cell lines. AME extract showed substantially higher cytotoxicity with lower percentages of cell survival and maximal activation of apoptosis in HepG2 cells without harming BHK cells compared to untreated cells and AMH extract. After following Lipinski's guidelines, only sixteen bioactive constituents from both extracts were chosen for molecular docking against cancer-causing target proteins. In silico analysis revealed that Thiophene-2-carbonitrile, 5-tert-butyl-3-(4-chlorobenzylidenamino) indicated best binding affinities of -7.4, -8.0, -8.5 and -8.0 kcal/mol against TNF-α, TGF-β R1, VEGFR2, and IL-17A, respectively, with favorable ADMET characteristics. In conclusion, the active constituent of A. millefolium might be a potential source of alternative drugs with a multitargeted approach for HCC treatment.