miRNA and DNA analysis by negative ion electron transfer dissociation and infrared multiple-photon dissociation mass spectrometry.

BACKGROUND: The use of simple and hybrid fragmentation techniques for the identification of molecules in tandem mass spectrometry provides different and complementary information on the structure of molecules. Nevertheless, these techniques have not been as widely explored for oligonucleotides as for peptides or proteins. The analysis of microRNAs (miRNAs) warrants special attention, given their regulatory role and their relationship with several diseases. The application of different fragmentation techniques will be very interesting for their identification. RESULTS: Four synthetic miRNAs and a DNA sequence were fragmented in an ESI-FT-ICR mass spectrometer using both simple and hybrid fragmentation techniques: CID, nETD followed by CID, IRMPD, and, for the first time, nETD in combination with IRMPD. The main fragmentation channel was base loss. The use of nETD-IRMPD resulted in d/z, a/w, and c/y ions at higher intensities. Moreover, nETD-IRMPD provided high sequence coverage and low internal fragmentation. Native MS analysis revealed that only miR159 and the DNA sequence formed stable dimers under physiological ionic strength. The use of organic co-solvents or additives resulted in a lower sequence coverage due to lesser overall ionization efficiency. NOVELTY: This work demonstrates that the combination of nETD and IRMPD for miRNA fragmentation constitutes a suitable alternative to common fragmentation methods. This strategy resulted in efficient fragmentation of [miRNA]5- using low irradiation times and fewer internal fragments while ensuring a high sequence coverage. Moreover, given that such low charge states predominate upon spraying in physiological-like conditions, native MS can be applied for obtaining structural information at the same time.

Multifunctional and Collaborative Protection of Proteins, Peptides, Phenolic Compounds, and Other Molecules against Oxidation in Apricot Seeds Extracts.

Antioxidant activity studies usually focus on a single type of molecule and do not consider possible collaborations among different molecules. The purpose of this work was to obtain multicomponent extracts exerting protection against oxidation from apricot seeds and to study the individual role of these components in the whole protection. Pressurized liquid extraction was employed to obtain extracts, and a response surface methodology enabled exploration of the effect of extraction conditions on the composition and prevalence of the antioxidant mechanism. Extractions carried out at 170 °C, in up to 7% ethanol, and for up to 25 min guaranteed multifunctional protection against oxidation by the collaboration of different molecules. While phenolic compounds were the main contributors to radical-scavenging capacity (R2 = 90% for ABTS and 88% for DPPH), proteins and phenolic compounds showed similar roles in the whole reducing power (proteins (R2 = 86%) and TPC (R2 = 90%)), and other compounds inhibited the formation of hydroxyl radicals and, especially, the peroxidation of lipids. The presence of peptides modified the antioxidant protection of extracts. UHPLC-Q-Orbitrap-MS/MS confirmed the presence of phenolic compounds and other antioxidant molecules. The presence of different kinds of molecules led to a multifunctional and collaborative protection against oxidation that could not be exerted by individual molecules.

Immobilization of Alcalase on Silica Supports Modified with Carbosilane and PAMAM Dendrimers.

Enzyme immobilization is a powerful strategy for enzyme stabilization and recyclability. Materials covered with multipoint molecules are very attractive for this goal, since the number of active moieties to attach the enzyme increases with respect to monofunctional linkers. This work evaluates different dendrimers supported on silica to immobilize a protease enzyme, Alcalase. Five different dendrimers were employed: two carbosilane (CBS) dendrimers of different generations (SiO2-G0Si-NH2 and SiO2-G1Si-NH2), a CBS dendrimer with a polyphenoxo core (SiO2-G1O3-NH2), and two commercial polyamidoamine (PAMAM) dendrimers of different generations (SiO2-G0PAMAM-NH2 and SiO2-G1PAMAM-NH2). The results were compared with a silica support modified with a monofunctional molecule (2-aminoethanethiol). The effect of the dendrimer generation, the immobilization conditions (immobilization time, Alcalase/SiO2 ratio, and presence of Ca2+ ions), and the digestion conditions (temperature, time, amount of support, and stirring speed) on Alcalase activity has been evaluated. Enzyme immobilization and its activity were highly affected by the kind of dendrimer and its generation, observing the most favorable behavior with SiO2-G0PAMAM-NH2. The enzyme immobilized on this support was used in two consecutive digestions and, unlike CBS supports, it did not retain peptides released in the digestion.

Comparative study of factors affecting the recovery of proteins from malt rootlets using pressurized liquids and ultrasounds.

Malt rootlets (MR) are a waste from brewing with high protein content. This work proposes to study the impact of extracting parameters on the recovery of proteins and the characteristics of extracts from MR using ultrasound-assisted extraction (UAE) and pressurized liquid extraction (PLE). A Box-Behnken experimental design was employed to study the effect of extracting parameters on the protein yield, while characterization comprised the study of antioxidant properties, the identification of extracted proteins using high-resolution tandem mass spectrometry, and the evaluation of the co-extraction of phenolic compounds. Protein extraction was promoted at an ultrasounds amplitude of 68%, for 20 min at 52 °C in UAE, while adding 33% ethanol resulted in the highest yield in PLE. While UAE extracted 53 ± 5% of MR proteins, PLE reached a 73 ± 7%, using more sustainable conditions. Significant antioxidant activities were observed in the PLE extract, although undermined by gastrointestinal digestion. Proteomic analysis detected 68 proteins from Hordeum vulgare in the UAE extract and 9 in the PLE extract. Proteins in MR are very different to that from barley grains or brewer's spent grains. PLE also co-extracted phenolic compounds while this was not significant by UAE.

Protease and gag diversity and drug resistance mutations among treatment-naive Mexican people living with HIV.

INTRODUCTION: In Mexico, HIV genotyping is performed in people living with HIV (PLWH) failing their first-line antiretroviral (ARV) regimen; it is not routinely done for all treatment-naive PLWH before ARV initiation. The first nationally representative survey published in 2016 reported that the prevalence of pretreatment drug mutations in treatment-naive Mexican PLWH was 15.5% to any antiretroviral drug and 10.6% to non-nucleoside reverse transcriptase inhibitors (NNRTIs) using conventional Sanger sequencing. Most reports in Mexico focus on HIV pol gene and nucleoside and non-nucleoside reverse transcriptase inhibitor (NRTI and NNRTI) drug resistance mutations (DRMs) prevalence, using Sanger sequencing, next-generation sequencing (NGS) or both. To our knowledge, NGS has not be used to detect pretreatment drug resistance mutations (DRMs) in the HIV protease (PR) gene and its substrate the Gag polyprotein. METHODS: Treatment-naive adult Mexican PLWH were recruited between 2016 and 2019. HIV Gag and protease sequences were obtained by NGS and DRMs were identified using the WHO surveillance drug resistance mutation (SDRM) list. RESULTS: One hundred PLWH attending a public national reference hospital were included. The median age was 28 years-old, and most were male. The median HIV viral load was 4.99 [4.39-5.40] log copies/mL and median CD4 cell count was 150 [68.0-355.78] cells/mm3. As expected, most sequences clustered with HIV-1 subtype B (97.9%). Major PI resistance mutations were detected: 8 (8.3%) of 96 patients at a detection threshold of 1% and 3 (3.1%) at a detection threshold of 20%. A total of 1184 mutations in Gag were detected, of which 51 have been associated with resistance to PI, most of them were detected at a threshold of 20%. Follow-up clinical data was available for 79 PLWH at 6 months post-ART initiation, seven PLWH failed their first ART regimen; however no major PI mutations were identified in these individuals at baseline. CONCLUSIONS: The frequency of DRM in the HIV protease was 7.3% at a detection threshold of 1% and 3.1% at a detection threshold of 20%. NGS-based HIV drug resistance genotyping provide improved detection of DRMs. Viral load was used to monitor ARV response and treatment failure was 8.9%.

Does the Fetus Limit Antibiotic Treatment in Pregnant Patients with COVID-19?

During pregnancy, there is a state of immune tolerance that predisposes them to viral infection, causing maternal-fetal vulnerability to the adverse effects of COVID-19. Bacterial coinfections significantly increase the mortality rate for COVID-19. However, it is known that all drugs, including antibiotics, will enter the fetal circulation in a variable degree despite the role of the placenta as a protective barrier and can cause teratogenesis or other malformations depending on the timing of exposure to the drug. Also, it is important to consider the impact of the indiscriminate use of antibiotics during pregnancy can alter both the maternal and fetal-neonatal microbiota, generating future repercussions in both. In the present study, the literature for treating bacterial coinfections in pregnant women with COVID-19 is reviewed. In turn, we present the findings in 50 pregnant women hospitalized diagnosed with SARS-CoV-2 without previous treatment with antibiotics; moreover, a bacteriological culture of sample types was performed. Seven pregnant women had coinfection with Staphylococcus haemolyticus, Staphylococcus epidermidis, Streptococcus agalactiae, Escherichia coli ESBL +, biotype 1 and 2, Acinetobacter jahnsonii, Enterococcus faecium, and Clostridium difficile. When performing the antibiogram, resistance to multiple drugs was found, such as macrolides, aminoglycosides, sulfa, dihydrofolate reductase inhibitors, beta-lactams, etc. The purpose of this study was to generate more scientific evidence on the better use of antibiotics in these patients. Because of this, it is important to perform an antibiogram to prevent abuse of empirical antibiotic treatment with antibiotics in pregnant women diagnosed with SARS-CoV-2.

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.

Extraction and Characterization of Antioxidant Peptides from Fruit Residues.

Fruit residues with high protein contents are generated during the processing of some fruits. These sustainable sources of proteins are usually discarded and, in all cases, underused. In addition to proteins, these residues can also be sources of peptides with protective effects against oxidative damage. The revalorization of these residues, as sources of antioxidant peptides, requires the development of suitable methodologies for their extraction and the application of analytical techniques for their characterization. The exploitation of these residues involves two main steps: the extraction and purification of proteins and their hydrolysis to release peptides. The extraction of proteins is mainly carried out under alkaline conditions and, in some cases, denaturing reagents are also employed to improve protein solubilization. Alternatively, more sustainable strategies based on the use of high-intensity focused ultrasounds, microwaves, pressurized liquids, electric fields, or discharges, as well as deep eutectic solvents, are being implemented for the extraction of proteins. The scarce selectivity of these extraction methods usually makes the subsequent purification of proteins necessary. The purification of proteins based on their precipitation or the use of ultrafiltration has been the usual procedure, but new strategies based on nanomaterials are also being explored. The release of potential antioxidant peptides from proteins is the next step. Microbial fermentation and, especially, digestion with enzymes such as Alcalase, thermolysin, or flavourzyme have been the most common. Released peptides are next characterized by the evaluation of their antioxidant properties and the application of proteomic tools to identify their sequences.

Feasibility of cationic carbosilane dendrimers for sustainable protein sample preparation.

Protein sample preparation is the bottleneck in the analysis of proteins. The aim of this work is to evaluate the feasibility of carbosilane dendrimers functionalized with cationic groups to make easier this step. Anionic carbosilane dendrimers (sulphonate- and carboxylate-terminated) have already demonstrated their interaction with proteins and their potential in protein sample preparation. In this work, interactions between positively charged carbosilane dendrimers and different model proteins were studied when working under different pH conditions, dendrimer concentrations, and dendrimer generations. Amino- and trimethylammonium-terminated carbosilane dendrimers presented, in some cases, weak interactions with proteins. Unlike them, carbosilane dendrimers with terminal dimethylamino groups could interact, in many cases, with proteins and these interactions were affected by the pH, the dendrimer concentration, and the dendrimer generation. Moreover, dendrimer precipitation was observed at all pHs, although just second and fourth generation (2 G and 4 G) dendrimers resulted in the formation of complexes with proteins. Under experimental conditions promoting dendrimer-protein interactions, 2 G dimethylamino-terminated dendrimers were proposed as an alternative to other methods used in analytical chemistry or analysis in which an organic solvent or a resin are required to enrich/purify proteins in a complex sample.

A sustainable approach for the extraction of cholesterol-lowering compounds from an olive by-product based on CO2-expanded ethyl acetate.

Olive (Olea europaea) processing results in large amounts of by-products that contain valuable molecules such as phenolic compounds and phytosterols. These molecules have demonstrated to reduce blood cholesterol levels. This work proposes the development of a method to obtain simultaneously phenolic compounds and phytosterols from the olive stone using CO2-expanded liquid extraction. Hansen solubility parameters were employed for the theoretical prediction of the most suitable bio-based solvent to extract target compounds. The Box-Behnken experimental design was employed to select the optimal conditions of pressure (8-25 MPa), the molar fraction of CO2 in ethyl acetate (0.15-0.55), and the temperature (40-80 °C). Extracts showing the highest and the lowest reductions of micellar cholesterol solubility capacity were analyzed by gas chromatography coupled to mass spectrometry to find out the compounds responsible for this activity. Different phenolic compounds, free fatty acids, and phytosterols were identified in the extracts. ß-Sitosterol and, especially, tyrosol and hydroxytyrosol were the compounds that primarily contributed to the reduction of micellar cholesterol solubility capacity.

Multiple protective effect of peptides released from Olea europaea and Prunus persica seeds against oxidative damage and cancer cell proliferation.

The long exposition to reactive species results in oxidative stress which has been related with the development of cancer and other serious diseases. Olea europaea and Prunus persica seeds present a high protein content and preliminary results demonstrated their high potency to obtain bioactive peptides. The protective effect against oxidative damage exerted by peptides released from Olea europaea and Prunus persica seeds has been evaluated in this work. Seed hydrolysates showed protection against oxidation through four different mechanisms: inhibition of the formation of hydroxyl radicals, scavenging of free radicals, reduction of oxidizing compounds, and inhibition of lipid peroxidation. Moreover, seed hydrolysates also reduced the oxidative stress induced by an oxidizing agent on human cancer cells. Despite protection evaluated by individual mechanisms seemed to be significantly affected by the seed genotype, overall protection of seed hydrolysates was not so different. Seeds hydrolysates were not cytotoxic on normal cells but they demonstrated antiproliferative effect on human cancer cells (HeLa, PC-3, and HT-29). Peptides in all seed hydrolysates were sequenced by RP-HPLC-ESI-Q-TOF. Eighteen common peptides were observed among olive seed hydrolysates while a wider variability was observed among Prunus seed hydrolysates.

Capillary liquid chromatography-ion trap-mass spectrometry methodology for the simultaneous quantification of four angiotensin-converting enzyme-inhibitory peptides in Prunus seed hydrolysates.

Prunus genus fruit seeds are sources of highly angiotensin-I-converting enzyme (ACE)-inhibitory peptides. The presence of peptides IYSPH, IYTPH, IFSPR, and VAIP seems to be related to this activity but no previous work has demonstrated the direct relationship between the concentration of these peptides and the antihypertensive activity of hydrolysates. This work describes the development of a method for the quantification of these peptides in Prunus seeds hydrolysates based on capillary liquid chromatography-IT-MS/MS. The analytical characteristics of the method were evaluated through the study of the linearity, LOD, LOQ, presence of matrix interferences, precision, and recovery. The developed methodology was applied to the determination of the four peptides in seed hydrolysates from different Prunus genus fruits: peaches (7 varieties), plums (2 varieties), nectarines (3 varieties), apricots (2 varieties), cherry, and paraguayo. Peaches and plums seed hydrolysates yielded the highest concentrations of these peptides while paraguayo one showed the lowest concentrations. A high correlation between peptides concentrations was demonstrated suggesting that the four peptides could be released from the same seed proteins.

Identification of peptides with antioxidant and antihypertensive capacities by RP-HPLC-Q-TOF-MS in dry fermented camel sausages inoculated with different starter cultures and ripening times.

Low molecular weight peptides are produced during meat fermentation. They contribute to generate flavor compounds but they can also exert certain bioactivities. The aim of this work was to evaluate, for the first time, the generation of bioactive peptides during the preparation of dry fermented camel sausages and to study the influence of the ripening time and the starter culture on bacteria growing, peptide concentration and size, and antioxidant and antihypertensive capacities of peptides. Camel meat sausages inoculated with different starter bacteria and non-inoculated were ripened up to 28days. Results demonstrated that bacteria population, peptide concentration, and peptide size were affected by the ripening time and the inoculated bacteria. Moreover, the ripening process resulted in an increasing antioxidant and antihypertensive capacity showing the highest bioactivities in fractions with peptides below 3kDa. Peptides in these fractions were identified by RP-HPLC-Q-TOF-MS analysis. Identified peptides showed common features with peptides with antioxidant or antihypertensive activity.

Sulfonate-terminated carbosilane dendron-coated nanotubes: a greener point of view in protein sample preparation.

Reduction or removal of solvents and reagents in protein sample preparation is a requirement. Dendrimers can strongly interact with proteins and have great potential as a greener alternative to conventional methods used in protein sample preparation. This work proposes the use of single-walled carbon nanotubes (SWCNTs) functionalized with carbosilane dendrons with sulfonate groups for protein sample preparation and shows the successful application of the proposed methodology to extract proteins from a complex matrix. SEM images of nanotubes and mixtures of nanotubes and proteins were taken. Moreover, intrinsic fluorescence intensity of proteins was monitored to observe the most significant interactions at increasing dendron generations under neutral and basic pHs. Different conditions for the disruption of interactions between proteins and nanotubes after protein extraction and different concentrations of the disrupting reagent and the nanotube were also tried. Compatibility of extraction and disrupting conditions with the enzymatic digestion of proteins for obtaining bioactive peptides was also studied. Finally, sulfonate-terminated carbosilane dendron-coated SWCNTs enabled the extraction of proteins from a complex sample without using non-environmentally friendly solvents that were required so far. Graphical Abstract Green protein extraction from a complex sample employing carbosilane dendron coated nanotubes.

Synthesis of Curcuminoids and Evaluation of Their Cytotoxic and Antioxidant Properties.

Curcumin (1) and ten derivatives (2-11) were synthesized and evaluated as cytotoxic and antioxidant agents. The results of primary screening by Sulforhodamine B assay against five human cancer cell lines (U-251 MG, glioblastoma; PC-3, human prostatic; HCT-15, human colorectal; K562, human chronic myelogenous leukemia; and SKLU-1, non-small cell lung cancer) allowed us to calculate the half maximal inhibitory concentration (IC50) values for the more active compounds against HCT-15 and K562 cell lines. Compounds 2 and 10 were the most active against both cell lines and were more active than curcumin itself. Thiobarbituric acid reactive substances (TBARS) assay showed that 7 has potent activity; even stronger than curcumin, α-tocopherol, and quercetin.

Factors affecting interactions between sulphonate-terminated dendrimers and proteins: A three case study.

This work proposes a deep study on the interactions between sulphonate-terminated carbosilane dendrimers and proteins. Three different proteins with different molecular weights and isoelectric points were employed and different pHs, dendrimer concentrations and generations were tested. Variations in fluorescence intensity and emission wavelength were used as protein-dendrimer interaction probes. Interaction between dendrimers and proteins greatly depended on the protein itself and pH. Other important issues were the dendrimer concentration and generation. Protein-dendrimer interactions were favored under acidic working conditions when proteins were positively charged. Moreover, in general, high dendrimer generations promoted these interactions. Modeling of protein-dendrimer interactions allowed to understand the different behaviors observed for every protein.

Proof of concept of a "greener" protein purification/enrichment method based on carboxylate-terminated carbosilane dendrimer-protein interactions.

Protein sample preparation is a critical and an unsustainable step since it involves the use of tedious methods that usually require high amount of solvents. The development of new materials offers additional opportunities in protein sample preparation. This work explores, for the first time, the potential application of carboxylate-terminated carbosilane dendrimers to the purification/enrichment of proteins. Studies on dendrimer binding to proteins, based on protein fluorescence intensity and emission wavelengths measurements, demonstrated the interaction between carboxylate-terminated carbosilane dendrimers and proteins at all tested pH levels. Interactions were greatly affected by the protein itself, pH, and dendrimer concentration and generation. Especially interesting was the interaction at acidic pH since it resulted in a significant protein precipitation. Dendrimer-protein interactions were modeled observing stable complexes for all proteins. Carboxylate-terminated carbosilane dendrimers at acidic pH were successfully used in the purification/enrichment of proteins extracted from a complex sample. Graphical Abstract Images showing the growing turbidity of solutions containing a mixture of proteins (lysozyme, myoglobin, and BSA) at different protein:dendrimer ratios (1:0, 1:1, 1:8, and 1:20) at acidic pH and SDS-PAGE profiles of the corresponsing supernatants. Comparison of SDS-PAGE profiles for the pellets obtained during the purification of proteins present in a complex sample using a conventional "no-clean" method based on acetone precipitation and the proposed "greener" method using carboxylate-terminated carbosilane dendrimer at a 1:20 protein:dendrimer ratio.

Identification of plum and peach seed proteins by nLC-MS/MS via combinatorial peptide ligand libraries.

UNLABELLED: Plum (Prunus domestica L.) and peach (Prunus persica (L.) Batsch) seed proteins are a source of bioactive peptides. These seeds, though, are usual residues produced during canning and beverage preparation that, in most cases, are irreversibly lost. The recovery and identification of these proteins might be of importance in human nutrition. This work employs the combinatorial peptide ligand libraries (CPLLs) technology as a tool to reduce the proteins dynamic concentration range. The most suitable extraction and CPLL capture conditions have been obtained and applied for the comprehensive identification of seed proteins. The analysis of recovered species by nLC-MS/MS has allowed the identification of 141 and 97 unique gene products from plum and peach seeds, respectively. It was possible to identify 16 proteins belonging to the Prunus genus. Moreover, a high number of histones and seed storage proteins were identified. Additionally, 21 and 14 bioactive peptides previously identified were found within protein sequences in plum and peach seeds, respectively. SIGNIFICANCE: Plums and peaches seeds are cheap sources of proteins that are irretrievably lost after canning and beverage production. Although this kind of residues has been used in animal feed or production of biofuel, they are usually incinerated or sent to landfills, wasting their huge potential. In order to exploit this, it is important to comprehensively study proteins present in plum and peach seeds. Nevertheless, since proteomics analysis is in most cases handicapped by the presence of high-abundance proteins masking the detection of the low-abundance ones, it is important to overcome this challenge. In this sense, combinatorial peptide ligand libraries (CPLLs) have been used in this work to reduce the dynamic protein concentration range to enable the identification of a higher amount of proteins than employing conventional methods. In this work, the better extracting conditions have been optimized and up to 141 and 97 unique gene products from plum and peach seeds have been found, respectively. Moreover, 21 and 14 peptides previously identified as bioactive peptides were ascertained within protein sequences in plum and peach seeds, respectively. For that reason, this research takes the first step in the recovery of these valuable proteins and in the extraction of bioactive peptides, which could be successfully adopted in human nutrition.

Experimental evidence for curcumin and its analogs for management of diabetes mellitus and its associated complications.

Diabetes mellitus is a serious world health problem and one of the most studied diseases; a major concern about its treatment is that ß-cell mass and functionality is hard to restore. In addition, it is frequently associated with severe complications, such as diabetic nephropathy and cardiomyopathy. The anti-inflammatory, anti-oxidative and anti-apoptotic properties of curcumin have made it a promising molecule for the treatment of this pathology; however, its solubility and bioavailability problems are still the subject of multiple studies. To cope with those difficulties, several approaches have been evaluated, such as the development of pharmaceutical formulations and curcumin analogs. This review discusses some of the studied therapeutic targets for curcumin in diabetes as well as the structural characteristics and targets of its analogs. The shortening of the central seven-carbon chain of curcumin has given rise to compounds without glucose-lowering effects but potentially useful for the treatment of diabetes complications; whereas preserving this chain retains the glucose-lowering properties. Most of the analogs discussed here have been recently synthesized and tested in animal models of type 1 diabetes; more studies in models of type 2 diabetes are needed.

HPLC-Q-TOF-MS identification of antioxidant and antihypertensive peptides recovered from cherry (Prunus cerasus L.) subproducts.

The processing of fruits, such as cherries, is characterized by generating a lot of waste material such as fruit stones, skins, etc. To contribute to environmental sustainability, it is necessary to recover these residues. Cherry stones contain seeds with a significant amount of proteins that are underused and undervalued. The aim of this work was to extract cherry seed proteins, to evaluate the presence of bioactive peptides, and to identify them by mass spectrometry. The digestion of cherry seed proteins was optimized, and three different enzymes were employed: Alcalase, Thermolysin, and Flavourzyme. Peptide extracts obtained by the digestion of the cherry seed protein isolate with Alcalase and Thermolysin yielded the highest antioxidant and antihypertensive capacities. Ultrafiltration of hydrolysates allowed obtaining fractions with high antioxidant and antihypertensive capabilities. HPLC-Q-TOF-MS together with bioinformatics tools enabled one to identify peptides in these fractions.