Assessment of in vitro cytotoxicity, anti-Alzheimer, and antidiabetic properties of fenugreek, white mulberry, and nettle leaves

Leafy plants are known for their rich bioactive profiles and have gained attention for their potential health benefits. This study evaluated the total phenolic content (TPC) using the Folin-Ciocalteu method and antioxidant properties, including ferric reducing antioxidant power (FRAP) and 2,2'-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) activities, of ethanolic extracts from fenugreek (FL), white mulberry (WBL), and nettle leaves (NL). It also investigated their inhibitory effects on alpha-amylase, alpha-glucosidase, acetylcholinesterase, and butyrylcholinesterase, and assessed their cytotoxicity on human embryonic kidney cells (HEK-293) and colorectal adenocarcinoma cells (CaCo-2) using MTT assays. The results revealed that the TPC was highest in NL (241.86 mg gallic acid equivalents (GAE) g-1 dry weight (DW)), followed by WBL (165.68 mg GAE g-1 DW) and FL (72.09 mg GAE g-1 DW), with NL also showing the highest FRAP (240.48 μmol Fe²⁺ g-1 extract) and ABTS antioxidant activities (19.26 mg trolox equivalents (TE) g-1 extract). Moreover, the inhibition of alpha-amylase ranged from 8.85% to 90.39% depending on the extract concentration (62.5–500 µg mL-1), with WBL and NL showing significant inhibitory effects on alpha-glucosidase within the same concentration range. Additionally, NL ethanolic extracts exhibited the highest butyrylcholinesterase inhibitory activity at 38.40% compared to FL (33.87%) and WBL (17.94%) at 2 mg mL-1, while acetylcholinesterase inhibition rates ranged from 23.14% for WBL to 53.35% for NL across all leaf samples. Furthermore, the ethanol extracts from FL, WBL, and NL yielded IC50 values of 1159.98, 1235.67, and 972.22 µg mL-1, respectively, on HEK-293 cells, while on CaCo-2 cells, the IC50 values were 897.41 µg mL-1 for FL, 754.11 µg mL-1 for WBL, and 648.80 µg mL-1 for NL. These findings underscore the potential of NL, FL, and WBL as valuable natural sources with diverse health benefits and significant therapeutic potential, making them promising candidates for industrial applications as functional ingredients.