Diplotaxis tenuifolia (L.) DC.: phytochemical profiling and multifunctional biological evaluation of extracts obtained by maceration and ultrasound-assisted extraction

Diplotaxis tenuifolia is an edible Brassicaceae species recognized for its phytochemical richness and health-promoting potential. This study evaluated six extracts obtained by maceration (MAC) and ultrasound-assisted extraction (UAE) using ethyl acetate (EtOAc), methanol (MeOH), and water. LC-MS/MS profiling revealed solvent- and method-dependent metabolite occurrence, including glucosinolates, flavonol glycosides, phenolic acids, and long-chain fatty acids, with representative compounds such as glucoraphanin, quercetin- and kaempferol-derived glycosides, p-coumaric and ferulic acids, and α-linolenic, linoleic, and erucic acids. UAE enhanced phytochemical diversity, particularly in water extracts, increasing detected compounds from 16 to 30. Total phenolic and flavonoid contents ranged from 15.80 to 24.07 mg GAE per g and from 2.06 to 31.53 mg RE per g, respectively. In antioxidant assays, MeOH extracts showed the highest DPPH activity (8.55 mg TE per g), whereas EtOAc-UAE exhibited the strongest CUPRAC and metal-chelating activities (45.67 mg TE per g and 22.14 mg EDTAE per g). In enzyme inhibition assays, MeOH-UAE showed the highest acetylcholinesterase inhibition (1.99 mg GALAE per g), while EtOAc-UAE was most active against butyrylcholinesterase, α-amylase, and α-glucosidase (3.97 mg GALAE per g, 0.89 mmol ACAE per g, and 0.64 mmol ACAE per g). EtOAc extracts were the only fractions active against Bacillus subtilis (6.7 and 8.7 mm for EtOAc-MAC and EtOAc-UAE), while EtOAc-UAE reduced Staphylococcus aureus biofilm formation to 13.6% of the control. EtOAc-UAE also showed the strongest cytotoxicity in MDA-MB-231 and PC-3 cells (IC50: 306.3 and 389.6 µg mL−1), with RT-qPCR confirming approximately 2-fold bax and 3-fold LC3B upregulation. These findings demonstrate that solvent polarity and extraction method critically determine the phytochemical and biological profiles of D. tenuifolia extracts.