RESUMEN
Plants and agri-food by-products represent a wide and renewable source of bioactive compounds with neuroprotective properties. In this research, various green extraction techniques were employed to recover bioactive molecules from Kalanchoe daigremontiana (kalanchoe), epicarp of Cyphomandra betacea (tamarillo), and cooperage woods from Robinia pseudoacacia (acacia) and Nothofagus pumilio (lenga), as well as a reference extract (positive control) from Rosmarinus officinalis L. (rosemary). The neuroprotective capacity of these plant extracts was evaluated in a set of in vitro assays, including enzymatic [acetylcholinesterase (AChE), butyrylcholinesterase (BChE), and lipoxygenase (LOX)] and antioxidant [ABTS, and reactive oxygen and nitrogen species (ROS and RNS)] bioactivity tests. Extracts were also submitted to a parallel artificial membrane permeability assay mimicking the blood-brain barrier (PAMPA-BBB) and to two cell viability assays in HK-2 and SH-SY5Y cell lines. Comprehensive phytochemical profiling based on liquid chromatography coupled to quadrupole-time-of-flight mass spectrometry (LC-Q-TOF-MS) analysis showed enriched content of phenolic and terpenoid compounds in the target extracts. Moreover, in vitro bioactivity tests showed promising neuroprotective capacity, particularly for supercritical-fluid extraction (SFE) extract from acacia (ABTS IC50 = 0.11 µg ml-1; ROS IC50 = 1.56 µg ml-1; AChE IC50 = 4.23 µg ml-1; BChE IC50 = 1.20 µg ml-1; and LOX IC50 = 4.37 µg ml-1), whereas PAMPA-BBB assays revealed high perfusion capacity of some representative compounds, such as phenolic acids or flavonoids. Regarding cytotoxic assays, tamarillo and rosemary SFE extracts can be considered as non-toxic, acacia SFE extract and lenga pressurized liquid extraction (PLE) extract as mild-cytotoxic, and kalanchoe as highly toxic extracts. The obtained results demonstrate the great potential of the studied biomass extracts to be transformed into valuable food additives, food supplements, or nutraceuticals with promising neuroprotective properties.
RESUMEN
Tamarillo (Cyphomandra betacea (Cav.) Sendt.), or tree tomato, is a tropical fruit from the Andean region of South America; it is highly rich in vitamins, minerals, and polyphenolic compounds. In this study, extracts from tamarillo epicarp (TE) were obtained by pressurized liquid extraction (PLE), and their in-vitro neuroprotective potential was assessed. A central composite design with response surface methodology was performed to optimize PLE as a function of solvent composition and temperature. Selected response variables were extraction yield, total phenolic content (TPC), total flavonoid content (TFC), total carotenoid content (TCC), antioxidant (ABTS), and anti-inflammatory (LOX) activities, and anti-acetylcholinesterase (AChE) inhibitory capacity. According to the desirability function, the optimal conditions were 100% ethanol and 180°C with a 0.87 desirability value. Next, the anti-butyrylcholinesterase enzyme (BChE), reactive oxygen species (ROS), and reactive nitrogen species (RNS) inhibition as well as cytotoxicity in HK-2, THP-1 monocytes, and SH-5YSY neuroblastoma cell lines were studied for the TE extract obtained under optimized conditions. The optimum TE extract provided the following results: extraction yield (36.25%), TPC (92.09 mg GAE/g extract), TFC (4.4 mg QE/g extract), TCC (107.15 mg CE/g extract), antioxidant capacity (ABTS, IC50 = 6.33 mg/ml extract), LOX (IC50 = 48.3 mg/ml extract), and AChE (IC50 = 97.46 mg/ml extract), and showed no toxicity at concentration up to 120 µg/ml extract for all the tested cell lines. Finally, chemical characterization by liquid chromatography-tandem mass spectrometry (UHPLC-q-TOF-MS/MS) of the optimum TE extract exhibited an important presence of hydroxycinnamic acid derivatives and other phenolic acids as well as quercetin hexoside and rutin, as main metabolites responsible for the observed biological properties. All these results suggested that TE, which represents between 8 and 15% of the total fruit, could become a promising natural by-product with a potential "multitarget" activity against Alzheimer's disease.