Anticancer Therapies Based on Oxidative Damage: Lycium barbarum Inhibits the Proliferation of MCF-7 Cells by Activating Pyroptosis through Endoplasmic Reticulum Stress.

Lycium barbarum, commonly recognized as goji berry or wolfberry, is highly appreciated not only for its organoleptic and nutritional properties but also as an important source of bioactive compounds such as polysaccharides, carotenoids, phenolics, and various other non-nutritive compounds. These constituents give it a multitude of health benefits, including antioxidant, anti-inflammatory, and anticancer properties. However, the precise biochemical mechanisms responsible for its anticancer effects remain unclear, and the comprehensive composition of goji berry extracts is often insufficiently explored. This study aimed to investigate the biochemical pathways modulated in breast cancer cells by an ethanolic extract of Lycium barbarum fruit (LBE). Following metabolomic profiling using UHPLC-HRMS/MS, we assessed the antitumoral properties of LBE on different breast cancer cell lines. This investigation revealed that LBE exhibited cytotoxic effects, inducing a pro-oxidant effect that triggered pyroptosis activation through endoplasmic reticulum (ER) stress and subsequent activation of the P-IRE1α/XBP1/NLRP3 axis in MCF-7 cells. In addition, LBE did not display cytotoxicity toward healthy human cells but demonstrated antioxidant properties by neutralizing ROS generated by doxorubicin. These findings underscore the potential of LBE as a highly promising natural extract in cancer therapy.

Biorefinery approach with green solvents for the valorization of Citrus reticulata leaves to obtain antioxidant and anticholinergic extracts.

Citrus reticulata L leaves are one of the main post-harvest byproduct, containing bioactive compounds, that are usually undervalued. This work describes the development of a biorefinery process based on the application of supercritical CO2 (SC-CO2) followed by ultrasonic-assisted extraction (UAE) combined with Natural Deep Eutectic Solvents (NaDES) to extract bioactive terpenoids and phenolic compounds from these leaves. Extraction temperature and pressure of SC-CO2 were optimized, obtaining the highest bioactive terpenoids content using 200 bar at 60 °C. A Box-Behnken experimental design showed that 57% of water in NaDES composed of Choline Chloride and Glycerol (1:2) as extraction solvent at 25 °C for 50 min were the optimal UAE-NaDES extraction conditions to obtain the highest bioactive phenolic content from the residue of the optimal SC-CO2 extraction. The optimum extract presented the highest bioactivity and polyphenol content determined by LC-DAD-MS compared with extracts obtained using only water or NaDES as solvent.

Pressurized liquid extraction of glucosinolates from Camelina sativa (L.) Crantz by-products: Process optimization and biological activities of green extract.

The cultivation of Camelina sativa (L.) Crantz is rapidly increasing due to oil production resulting in a substantial volume of by-products, which still have an interesting composition in secondary metabolites, especially glucosinolates. Therefore, a green extraction procedure of glucosinolates by Pressurised Liquid Extraction was developed and optimized using a chemometric approach. Furthermore, the glucosinolates were purified by solid phase extraction, and a preliminary study on bioaccessibility and bioavailability study was carried out to evaluate the resistance of the glucosinolates to the digestive process. The application of pressurised liquid extraction to the recovery of glucosinolates from camelina sativa by-product, is a green, automatic, and rapid method, representing a valid alternative to conventional extraction method to obtain ingredients for food industries.

Capillary electromigration methods for food analysis and Foodomics: Advances and applications in the period March 2021 to March 2023.

This work presents a revision of the main applications of capillary electromigration (CE) methods in food analysis and Foodomics. Papers that were published during the period March 2021 to March 2023 are included. The work shows the multiple CE methods that have been developed and applied to analyze different types of molecules in foods and beverages. Namely, CE methods have been applied to analyze amino acids, biogenic amines, heterocyclic amines, peptides, proteins, phenols, polyphenols, pigments, lipids, carbohydrates, vitamins, DNAs, contaminants, toxins, pesticides, additives, residues, small organic and inorganic compounds, and other minor compounds. In addition, new CE procedures to perform chiral separation and for evaluating the effects of food processing as well as the last developments of microchip CE and new applications in Foodomics will be also discussed. The new procedures of CE to investigate food quality and safety, nutritional value, storage, and bioactivity are also included in the present review work.

Ergosterol and polyphenol contents as rapid indicators of orchardgrass silage safety.

The ergosterol (ERG) has been proposed as a potential indicator of fungal contamination, along with polyphenol content analysis to predict silage safety. Despite efforts in controlling fungal growth in silage, mycotoxin co-contamination represents a possible risk for animal and human health. Modern analytical techniques determine a multitude of fungal metabolites contaminating feed. Nonetheless, these methods require sometimes arduous sample pre-treatment, long separation times, and expensive standard compounds to identified contaminants. Thus, the goal of this study was to suggest a rapid analysis of ERG and polyphenol contents to assess silage hygienic quality in ten orchardgrass varieties ensiled without and with biological and chemical additives. The determination of ERG on samples was performed by high-performance liquid chromatography using UV detection and UV/Vis spectrophotometry to determine the polyphenol content. Statistically significant differences (P < 0.05) between varieties, years and silage additives were found. Bepro was the unique variety that did not present ERG in the first cut in 2012. ERG content increased in the first cut in 2013 using biological additives as well as ERG and polyphenol contents in the first cut in 2013 using chemical additives compared with untreated silage. In addition, biological and chemical additives used in this study did not satisfactorily reduce the content of ERG and polyphenols in silage grass. Consequently, our results provide fast information about the progressive fungal contamination of grass silage. To our knowledge, it is the first time that the presence of ERG and polyphenols is determined in ten different orchardgrass varieties treated without and with additives. In general, ERG and polyphenol contents showed to be good indicators of orchardgrass silage safety.

Composition of Nonextractable Polyphenols from Sweet Cherry Pomace Determined by DART-Orbitrap-HRMS and Their In Vitro and In Vivo Potential Antioxidant, Antiaging, and Neuroprotective Activities.

Sweet cherry pomace is an important source of phenolic compounds with beneficial health properties. As after the extraction of phenolic compounds, a phenolic fraction called nonextractable polyphenols (NEPs) remains usually retained in the extraction residue, alkaline and acid hydrolyses and enzymatic-assisted extraction (EAE) were carried out in this work to recover NEPs from the residue of conventional extraction from sweet cherry pomace. In vitro and in vivo evaluation of the antioxidant, antihypertensive, antiaging, and neuroprotective capacities employing Caenorhabditis elegans was achieved for the first time. Extractable phenolic compounds and NEPs were separated and identified by families by high-performance thin-layer chromatography (HPTLC) with UV/Vis detection. A total of 39 phenolic compounds were tentatively identified in all extracts by direct analysis in real-time high-resolution mass spectrometry (DART-Orbitrap-HRMS). EAE extracts presented the highest in vitro and in vivo antioxidant capacity as well as the highest in vivo antiaging and neuroprotective capacities. These results showed that NEPs with interesting biological properties are retained in the extraction residue, being usually underestimated and discarded.

In vitro assessment of the bioavailability of bioactive non-extractable polyphenols obtained by pressurized liquid extraction combined with enzymatic-assisted extraction from sweet cherry (Prunus avium L.) pomace.

In vitro digestion and absorption simulation processes of non-extractable polyphenols (NEPs) obtained by pressurized liquid extraction combined with enzymatic-assisted extraction with Promod enzyme (PLE-EAE) from the residue of conventional extraction of sweet cherry pomace were studied. In general, total phenolic and proanthocyanidin contents decreased in each phase of the digestion. However, the antioxidant capacity increased when the digestion process progressed. In addition, the highest total phenolic and proanthocyanidin contents and antioxidant capacity were obtained in the absorbed fraction. NEPs from PLE-EAE extract, digestive fractions, absorbed and unabsorbed fractions were analyzed by ultra-high-performance liquid chromatography coupled to electrospray ionization quadrupole Exactive-Orbitrap mass spectrometry (UHPLC-ESI-Q-Orbitrap-MS). Fifteen NEPs were identified in the intestinal fraction and five in the absorbed fraction after the digestion process. Results obtained in this study define for the first time the bioavailability of antioxidant NEPs obtained from sweet cherry pomace.

Rapid fingerprinting of extractable and non-extractable polyphenols from tropical fruit peels using direct analysis in real time coupled to orbitrap mass spectrometry.

A simple and rapid direct analysis in real-time coupled to high-resolution mass spectrometry (DART-HRMS) methodology was developed to generate the extractable and non-extractable polyphenols (NEPs) fingerprint for four different passion fruits, G. mangostana, and A. squamosa peels as case-study to investigate the influence of alkaline hydrolysis and enzymatic-assisted extraction (EAE) on the recovery of NEPs. The extraction residue obtained after these treatments was also analyzed by DART-HRMS. Data compiled from DART-HRMS mass spectra were processed with principal component analysis to discriminate among the different treatments. EAE with Depol enzyme enabled to obtain NEPs with the highest signal intensity in DART-HRMS analysis from all peels except for P. edulis and A. squamosa peels. In these two cases, NEPs were better extracted by EAE with Promod enzyme and alkaline hydrolysis. Results showed that the applied treatments were efficient to extract NEPs since their signal intensities in the extraction residues were very low compared with their extracts.

Pressurized Liquid Extraction Combined with Enzymatic-Assisted Extraction to Obtain Bioactive Non-Extractable Polyphenols from Sweet Cherry (Prunus avium L.) Pomace.

Sweet cherry generates large amounts of by-products within which pomace can be a source of bioactive phenolic compounds. Commonly, phenolic compounds have been obtained by conventional extraction methodologies. However, a significant fraction, called non-extractable polyphenols (NEPs), stays held in the conventional extraction residues. Therefore, in the present work, the release of NEPs from cherry pomace using pressurized liquid extraction (PLE) combined with enzyme-assisted extraction (EAE) using PromodTM enzyme is investigated for the first time. In order to study the influence of temperature, time, and pH on the NEPs extraction, a response surface methodology was carried out. PLE-EAE extracts displayed higher TPC (75 ± 8 mg GAE/100 g sample) as well as, PA content, and antioxidant capacity than the extracts obtained by PLE (with a TPC value of 14 ± 1 mg GAE/100 g sample) under the same extraction conditions, and those obtained by conventional methods (TPC of 8.30 ± 0.05 mg GAE/100 g sample). Thus, PLE-EAE treatment was more selective and sustainable to release NEPs from sweet cherry pomace compared with PLE without EAE treatment. Besides, size-exclusion chromatography profiles showed that PLE-EAE allowed obtaining NEPs with higher molecular weight (>8000 Da) than PLE alone.

High-performance thin-layer chromatography and direct analysis in real time-high resolution mass spectrometry of non-extractable polyphenols from tropical fruit peels.

Passiflora species, mangosteen, and cherimoya peels are a source of bioactive phenolic compounds. Nevertheless, a significant fraction of polyphenols, called non-extractable polyphenols (NEPs), are retained in the extraction residue after a conventional extraction. Thus, alkaline, acid, and enzymatic-assisted extractions to recover high contents of antioxidant NEPs from the extraction residue of fruit peels, were compared in this work. A high-performance thin-layer chromatography method with UV/Vis detection was developed in order to obtain the phenolic profile for the extracts. The most intense bands were further analyzed by direct analysis in real-time-high-resolution mass spectrometry to tentatively identified NEPs in fruit peel extracts. Total phenolic and proanthocyanidin contents and antioxidant capacity of the extracts were measured to carry out a multivariate statistical analysis. Alkaline hydrolysis was the most efficient treatment to recover NEPs from fruit peels as well as a promising treatment to obtain antioxidant extracts along with EAE. Cherimoya peel extracts were the richest in antioxidant NEPs. This work highlights that many NEPs remain on the extraction residue of fruit peels after conventional extraction and are not usually taken into account.

Enzyme-assisted extraction of bioactive non-extractable polyphenols from sweet cherry (Prunus avium L.) pomace.

Sweet cherries processing produces big amounts of wastes mainly constituted by cherry pomace that can be a source of bioactive polyphenols. However, during the extraction process, an important fraction called non-extractable polyphenols (NEPs) remains retained in the extraction residue. This work describes the development of an enzyme-assisted extraction (EAE) method to obtain NEPs from sweet cherry pomace employing three different enzymes. Box-Behnken experimental designs were employed to select the optimal conditions of extraction time, temperature, enzyme concentration, and pH. The total phenolic and proanthocyanidin contents and the antioxidant and antihypertensive capacities were measured. Optimal EAE conditions extracted higher content of proanthocyanidins and with higher bioactivity from extraction residue than alkaline and acid hydrolysis. Moreover, there were higher amounts of bioactive phenolics in the extraction residue than in the sweet cherry pomace extract. The estimation of NEPs molecular weight distribution by HPLC-SEC demonstrated that EAE extracted NEPs with high molecular weight.

Strategies for the extraction and analysis of non-extractable polyphenols from plants.

The majority of studies based on phenolic compounds from plants are focused on the extractable fraction derived from an aqueous or aqueous-organic extraction. However, an important fraction of polyphenols is ignored due to the fact that they remain retained in the residue of extraction. They are the so-called non-extractable polyphenols (NEPs) which are high molecular weight polymeric polyphenols or individual low molecular weight phenolics associated to macromolecules. The scarce information available about NEPs shows that these compounds possess interesting biological activities. That is why the interest about the study of these compounds has been increasing in the last years. Furthermore, the extraction and characterization of NEPs are considered a challenge because the developed analytical methodologies present some limitations. Thus, the present literature review summarizes current knowledge of NEPs and the different methodologies for the extraction of these compounds, with a particular focus on hydrolysis treatments. Besides, this review provides information on the most recent developments in the purification, separation, identification and quantification of NEPs from plants.