The objective of this study was to analyze the chemical profile of extracts from different fruit parts of the perennial arborescent cactus Opuntia leucotricha by applying ultra-high-performance liquid chromatography–high-resolution mass spectrometry (UHPLC-HRMS) and to evaluate their biological properties (antioxidant and enzyme inhibitory capacities). Extracts were obtained from the fruit pulp (without or with seeds), seeds, and peel by ultrasound-assisted extraction (UAE) using the conventional solvent methanol 50% and two Natural Deep Eutectic Solvents (NADESs) (glycerol:urea, 1:1 and citric acid:sorbitol, 1:2). A total of 33 compounds were identified, including phenolics, fatty acids, and others. Phenolic acids were the most abundant class of phenolics identified in all fruit parts, with the highest concentration observed in the methanol extracts from peel (593.02 µg/g_DW), followed by seed-containing pulp (69.03 µg/g_DW), pulp (57.83 µg/g_DW), and seeds (39.97 µg/g_DW). The second most effective extractant was NADES 1 (glycerol:urea, 1:1), which was also successful in extracting compounds with antioxidant capacity. Overall, the extracts demonstrated considerable enzyme inhibitory activities, with the greatest effects observed against α-amylase and α-glucosidase. The results indicate that O. leucotricha fruits could be a promising source of bioactive compounds, and NADES a viable alternative to organic solvents for their industrial exploitation.

This study evaluated the extraction efficiency of two Natural Deep Eutectic Solvents (NADESs), glycerol–urea (1:1) and citric acid–sorbitol (1:2), for recovering phenolic compounds from the different parts of the fruit (pulp, seed-containing pulp, seeds, and peel) of Opuntia robusta and Opuntia ficus-indica in comparison with 50% methanol. Phytochemical profiling was performed using ultra-high-performance liquid chromatography–high-resolution mass spectrometry, alongside antioxidant and enzyme inhibition assessments (acetylcholinesterase, butyrylcholinesterase, tyrosinase, α-glucosidase, and α-amylase). Glycerol–urea performed similarly to methanol in extracting phenolic compounds with notable antioxidant properties. Peel extracts contained the highest levels of bioactive compounds, particularly phenolic acids (525.49 in O. robusta and 362.96 µg/g_DW in O. ficus indica). Enzyme inhibition varied across species and fruit parts, with extracts from both species inhibiting all targeted enzymes. Notably, this study provides the first evidence of tyrosinase inhibitory activity in O. robusta, which exhibited the strongest inhibition. Overall, these results emphasize the potential of cactus fruit extracts, particularly from O. robusta, for valorization, and support the use of NADESs as a sustainable and medium for extracting antioxidant compounds. Furthermore, the potential of fruit peel as waste with nutraceutical applications was demonstrated.