In vivo neuroprotective capacity of a Dunaliella salina extract - comprehensive transcriptomics and metabolomics study.

In this study, an exhaustive chemical characterization of a Dunaliella salina (DS) microalga extract obtained using supercritical fluids has been performed, and its neuroprotective capacity has been evaluated in vivo using an Alzheimer's disease (AD) transgenic model of Caenorhabditis elegans (strain CL4176). More than 350 compounds were annotated in the studied DS extract, with triacylglycerols, free fatty acids (FAs), carotenoids, apocarotenoids and glycerol being the most abundant. DS extract significantly protects C. elegans in a dose-dependent manner against Aß-peptide paralysis toxicity, after 32 h, 53% of treated worms at 50 µg/mL were not paralyzed. This concentration was selected to further evaluate the transcriptomics and metabolomics changes after 26 h by using advanced analytical methodologies. The RNA-Seq data showed an alteration of 150 genes, mainly related to the stress and detoxification responses, and the retinol and lipid metabolism. The comprehensive metabolomics and lipidomics analyses allowed the identification of 793 intracellular metabolites, of which 69 were significantly altered compared to non-treated control animals. Among them, different unsaturated FAs, lysophosphatidylethanolamines, nucleosides, dipeptides and modified amino acids that have been previously reported as beneficial during AD progression, were assigned. These compounds could explain the neuroprotective capacity observed, thus, providing with new evidences of the protection mechanisms of this promising extract.

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.

Towards green extraction of bioactive natural compounds.

The increasing interest in natural bioactive compounds is pushing the development of new extraction processes that may allow their recovery from a variety of different natural matrices and biomasses. These processes are clearly sought to be more environmentally friendly than the conventional alternatives that have traditionally been used and are closely related to the 6 principles of green extraction of natural products. In this trend article, the most critical aspects regarding the current state of this topic are described, showing the different lines followed to make extraction processes greener, illustrated by relevant examples. These include the implementation of new extraction technologies, the research on new bio-based solvents, and the development of new sequential process and biorefinery approaches to produce a full valorization of the natural sources. Moreover, the future outlook in the field is presented, in which the main areas of evolution are identified and discussed.

Evolution of anthocyanin content during grape ripening and characterization of the phenolic profile of the resulting wine by comprehensive two-dimensional liquid chromatography.

The typical phenolic profile in grapes is characterized by its complexity both in terms of number of diverse chemical structures and their variation during ripening. Besides, the specific phenolic composition of grapes directly influences the presence of those components in the resulting wine. In this contribution, a new method based on the application of comprehensive two-dimensional liquid chromatography coupled to a diode array detector and tandem mass spectrometry has been developed to obtain the typical phenolic profile of Malbec grapes cultivated in Brazil. Moreover, the method has been demonstrated to be useful to study how the phenolic composition in grapes evolved during a 10-week ripening period. Main detected compounds in grapes and in the wine derived from them were anthocyanins, although a good number of polymeric flavan-3-ols were also tentatively identified, among other compounds. Results show how the amount of anthocyanins present in grapes was increased during ripening up to 5-6 weeks and then decreased towards week 9. The two-dimensional approach applied was demonstrated to be useful for the characterization of the complex phenolic profile of these samples, involving more than 40 different structures and has the potential to be further applied to the study of this important fraction is different grapes and wines systematically.

COVID-19 stressor reduces risk taking: the role of trait interoception.

The Framing Effect (FE) demonstrated that the way two alternatives are displayed affects people's inclination to make a specific choice, showing a risk aversion when alternatives are displayed on positive frames and risk seeking in negative frames. Risk seeking in negative frames is closely linked to loss aversion. Moreover, classical research and the salience-of-losses hypothesis argues that stress may enhance the FE and loss aversion. Recent studies also suggest that the trait interoception and alexithymia could interact and moderate the framing susceptibility. However, experimental paradigms on stress could ignore variables such as threat perception. In this sense, COVID-19 pandemic has become a powerful real-life stressor in many countries. We aimed to study how real-life stressors influence decision-making under risk. A total of 97 participants were divided into a control (n = 48) and an experimental group (n = 49). The experimental group were exposed to a stressor manipulation, a 5 min COVID-19 lockdown documentary. Our results show that COVID-19-related stressors significantly decreased bet acceptance regardless of the frame, also reducing loss aversion. Moreover, interoception was a significant predictor of loss aversion under stress conditions. Our results do not support classical research on stress and FE.

The coordination of anti-phage immunity mechanisms in bacterial cells.

Bacterial cells are equipped with a variety of immune strategies to fight bacteriophage infections. Such strategies include unspecific mechanisms directed against any phage infecting the cell, ranging from the identification and cleavage of the viral DNA by restriction nucleases (restriction-modification systems) to the suicidal death of infected host cells (abortive infection, Abi). In addition, CRISPR-Cas systems generate an immune memory that targets specific phages in case of reinfection. However, the timing and coordination of different antiviral systems in bacterial cells are poorly understood. Here, we use simple mathematical models of immune responses in individual bacterial cells to propose that the intracellular dynamics of phage infections are key to addressing these questions. Our models suggest that the rates of viral DNA replication and cleavage inside host cells define functional categories of phages that differ in their susceptibility to bacterial anti-phage mechanisms, which could give raise to alternative phage strategies to escape bacterial immunity. From this viewpoint, the combined action of diverse bacterial defenses would be necessary to reduce the chances of phage immune evasion. The decision of individual infected cells to undergo suicidal cell death or to incorporate new phage sequences into their immune memory would be determined by dynamic interactions between the host's immune mechanisms and the phage DNA. Our work highlights the importance of within-cell dynamics to understand bacterial immunity, and formulates hypotheses that may inspire future research in this area.

Study of the potential neuroprotective effect of Dunaliella salina extract in SH-SY5Y cell model.

Alzheimer's disease (AD) is the most common form of dementia caused by a progressive loss of neurons from different regions of the brain. This multifactorial pathophysiology has been widely characterized by neuroinflammation, extensive oxidative damage, synaptic loss, and neuronal cell death. In this sense, the design of multi-target strategies to prevent or delay its progression is a challenging goal. In the present work, different in vitro assays including antioxidant, anti-inflammatory, and anti-cholinergic activities of a carotenoid-enriched extract from Dunaliella salina microalgae obtained by supercritical fluid extraction are studied. Moreover, its potential neuroprotective effect in the human neuron-like SH-SY5Y cell model against remarkable hallmarks of AD was also evaluated. In parallel, a comprehensive metabolomics study based on the use of charged-surface hybrid chromatography (CSH) and hydrophilic interaction liquid chromatography (HILIC) coupled to high-resolution tandem mass spectrometry (Q-TOF MS/MS) was applied to evaluate the effects of the extract on the metabolism of the treated cells. The use of advanced bioinformatics and statistical tools allowed the identification of more than 314 metabolites in SH-SY5Y cells, of which a great number of phosphatidylcholines, triacylglycerols, and fatty acids were significantly increased, while several phosphatidylglycerols were decreased, compared to controls. These lipidomic changes in cells along with the possible role exerted by carotenoids and other minor compounds on the cell membrane might explain the observed neuroprotective effect of the D. salina extract. However, future experiments using in vivo models to corroborate this hypothesis must be carried out.

Capillary electromigration methods for food analysis and Foodomics: Advances and applications in the period February 2019-February 2021.

This work presents a revision of the main applications of capillary electromigration methods in food analysis and Foodomics. Articles that were published during the period February 2019-February 2021 are included. The work shows the multiple CE methods that have been developed and applied to analyze different types of molecules in foods. Namely, CE methods have been applied to analyze amino acids, biogenic amines, carbohydrates, chiral compounds, contaminants, DNAs, food additives, heterocyclic amines, lipids, secondary metabolites, peptides, pesticides, phenols, pigments, polyphenols, proteins, residues, toxins, vitamins, small organic and inorganic compounds, as well as other minor compounds. The last results on the use of CE for monitoring food interactions and food processing, including recent microchips developments and new applications of CE in Foodomics, are discussed too. The new procedures of CE to investigate food quality and safety, nutritional value, storage and bioactivity are also included in the present review work.

In vitro Neuroprotective Potential and Lipidomics Study of Olive Leaves Extracts Enriched in Triterpenoids.

Alzheimer's Disease (AD) is the most common form of dementia that is associated with extracellular amyloid beta (Aß) plaque formation. Genetic, environmental, and nutrition factors have been suggested as contributors to oxidative stress and neuroinflammation events that are connected to AD etiology, and secondary metabolites, such as triterpenes, have shown promising results in AD prevention. In this work, the neuroprotective and anti-inflammatory potential of an olive leaves fraction enriched in triterpenoid compounds obtained using supercritical fluid extraction (SFE) and dynamic adsorption/desorption using sea sand as adsorbent has been performed. In addition, a comprehensive lipidomics study of the response of SH-SY5Y neuroblastoma cell line to this fraction was carried out using advanced analytical methodologies, namely, charged-surface hybrid chromatography-quadrupole-time of flight mass spectrometry (CSH-Q-TOF MS/MS). The use of freely available lipidomic annotation tools and databases, and stringent cut-off filters allowed the annotation of more than 250 intracellular lipids. Advanced bioinformatics and statistical tools showed a number of phosphatidylcholines and phosphatidylethanolamines significantly increased, which could explain the protection against the cell death caused by Aß1-42. Moreover, several triacylglycerols were found decreased. These results suggest triterpenoids from olive leaves as good neuroprotective candidates, and open a new gate for future experiments using in vivo models to corroborate this hypothesis.

Chemometric optimization of trypsin digestion method applying infrared, microwave and ultrasound energies for determination of caseins and ovalbumin in wines.

Caseins and ovalbumin are frequently used as wine fining agents to remove undesirable compounds like polymeric phenols. Their presence in wines is a subject of concern because may cause adverse effects on susceptible consumers, especially when their presence is not labeled. A key step for its determination is trypsin digestion, which is considered the bottleneck of bottom-up approach workflow because usually requires several hours. To reduce this time, the objective of this work was to carry out a chemometric optimization of trypsin digestion method applying infrared, microwave and ultrasound energies to determine caseins and ovalbumin in wines. The conditions of each accelerated digestion method were optimized using a Response Surface Methodology based on central composite design. The parameters optimized were digestion time and trypsin: protein ratio. The response variable evaluated was digestion yield, which was determined through the peak area of each protein transition determined by liquid chromatography-mass spectrometry. The most effective technique was microwave followed by ultrasound and infrared. Since optimal values of microwave and ultrasound-assisted digestion were the same, the later was chosen considering sample preparation and cost. Applying the proposed approach, a reduction of ca. 140 and 240-fold on digestion time was achieved compared with optimized and non-optimized conventional methods, respectively. With this workflow, both proteins were digested in a single 3 min process allowing its detection by liquid chromatography-mass spectrometry at µg L-1 level, which is ca. 60 times lower than the current limit of 0.25 mg L-1.

Mycotoxin extraction from edible insects with natural deep eutectic solvents: a green alternative to conventional methods.

Edible insects are widely consumed in Africa, Asia, Oceania and Latin America, but less commonly so in Western countries. Since the turn of the millennium, however, entomophagy has aroused growing interest worldwide in response to the increasing scarcity of food resources. In fact, edible insects can be a source of high-quality protein, and also of fat, energy, minerals and vitamins. However, the lack of regulatory guidelines for microbiologically or chemically hazardous agents potentially present in these new foods (e.g., mycotoxins) may make their consumption unsafe. In this work, we developed an environmentally friendly analytical method using natural deep eutectic solvents (NADES or natural DES) in combination with ultra-high performance liquid chromatography/tandem mass spectrometry (UHPLC-MS/MS) for the simultaneous determination of six mycotoxins of great concern owing to their toxic effects on humans and animals (namely, fumonisin B1, fumonisin B2, T-2 toxin, HT-2 toxin, ochratoxin A and mycophenolic acid) in insect-based food products. The target mycotoxins were co-extracted from cricket flour by using the optimum DES composition (namely, a mixture of choline chloride and urea, in a 1:2 mole ratio, containing 15% water which resulted in the highest extraction recoveries for all toxins). An experimental design method (Fractional Factorial Design (FFD) was used to examine the influence of the operational variables DES volume and water content, amount of sample, extraction time and extraction temperature on the extraction efficiency for each mycotoxin. Under optimum conditions, extraction recoveries were close to 100% except for fumonisin B2 (70%) and T-2 toxin (50%), with relative standard deviations (RSDs) below 13% in all cases. The proposed NADES-UHPLC-MS/MS method was validated in accordance with the European Commission 2002/657/EC and 2006/401/EC decisions, and used to determine the target compounds in cricket flour, silkworm pupae powder and black cricket powder.

Chemometric optimisation of pressurised liquid extraction for the determination of alliin and S-allyl-cysteine in giant garlic (Allium ampeloprasum L.) by liquid chromatography tandem mass spectrometry.

INTRODUCTION: Giant garlic is a functional food that contains different kinds of bioactive molecules with beneficial effects on chronic noncommunicable diseases like diabetes and cardiovascular conditions. Considering biosynthesis pathways, abundance, and biological activity, alliin and S-allyl-cysteine were used as chemical markers of organosulphur compounds present in giant garlic. OBJECTIVE: To establish a chemometric optimisation of pressurised liquid extraction for the determination of alliin and S-allyl-cysteine in giant garlic by liquid chromatography tandem mass spectrometry (LC-MS/MS). METHODOLOGY: Samples were blanched (ca. 90°C for 10 min) to inactivate alliinase and γ-glutamyl transpeptidase enzymes and then freeze-dried. Chemometric optimisation was performed via response surface methodology based on central composite design (CCD). Organosulphur compound yields were determined applying a validated LC-MS/MS method in multiple reaction monitoring (MRM) mode using the following transitions: for alliin m/z 178 → 74 and for S-allyl-cysteine m/z 162 → 41. RESULTS: According to CCD results, under constant conditions of pressure (1500 psi) and time (20 min), the optimal conditions for pressurised liquid extraction of alliin and S-allyl-cysteine were 70.75 and 68.97% v/v of ethanol in extraction solvent and 76.45 and 98.88°C as extraction temperature, respectively. Multiple response optimisation for the simultaneous extraction of both organosulphur compounds was established via desirability function. Under these conditions, 2.70 ± 0.27 mg g-1 dry weight (DW) of alliin and 2.79 ± 0.22 mg g-1 DW of S-allyl-cysteine were extracted. CONCLUSIONS: These results clearly demonstrated that pressurised liquid extraction is an efficient green technique to extract bioactive organosulphur compounds from giant garlic. Extraction yields were significantly (p < 0.05) higher than those obtained with conventional ultra-turrax extraction.

Sleep-Wake Cycle and EEG-Based Biomarkers during Late Neonate to Adult Transition.

During the transition from neonate to adulthood, brain maturation establishes coherence between behavioral states-wakefulness, non-rapid eye movement, and rapid eye movement sleep. In animal models few studies have characterized and analyzed cerebral rhythms and the sleep-wake cycle in early ages, in relation to adulthood. Since the analysis of sleep in early ages can be used as a predictive model of brain development and the subsequent emergence of neural disturbances in adults, we performed a study on late neonatal mice, an age not previously characterized. We acquired longitudinal 24 h electroencephalogram and electromyogram recordings and performed time and spectral analyses. We compared both age groups and found that late neonates: (i) spent more time in wakefulness and less time in non-rapid eye movement sleep, (ii) showed an increased relative band power in delta, which, however, reduced in theta during each behavioral state, (iii) showed a reduced relative band power in beta during wakefulness and non-rapid eye movement sleep, and (iv) manifested an increased total power over all frequencies. The data presented here might have implications expanding our knowledge of cerebral rhythms in early ages for identification of potential biomarkers in preclinical models of neurodegeneration.

Comprehensive two-dimensional liquid chromatography-based quali-quantitative screening of aqueous phases from pyrolysis bio-oils.

Pyrolysis processes are an alternative to minimize the environmental problem associated to agrifood industrial wastes. The main product resulting from these processes is a high-value liquid product, called bio-oil. Recently, the use of comprehensive two-dimensional liquid chromatography (LC × LC) has been demonstrated as a useful tool to improve the characterization of the water-soluble phases of bio-oils, considering their complexity and high water content. However, the precise composition of bio-oils from different agrifood byproducts is still unknown. In the present study, the qualitative and quantitative screening of eight aqueous phases from different biomasses, not yet reported in the literature, using LC × LC is presented. The two-dimensional approach was based on the use of two reverse phase separations. An amide column in the first dimension together with a C18 column in the second dimension were employed. Thanks to the use of diode array and mass spectrometry detection, 28 compounds were identified and quantified in the aqueous phase samples with good figures of merit. Samples showed a distinct quali-quantitative composition and a great predominance of compounds belonging to aldehydes, ketones and phenols, most of them with high polarity.

Preparative Separation of Procyanidins from Cocoa Polyphenolic Extract: Comparative Study of Different Fractionation Techniques.

To provide further insight into the antioxidant potential of procyanidins (PCs) from cocoa beans, PC extract was fractionated by several methodologies, including solid phase extraction, Sephadex LH-20 gel permeation, and preparative HPLC using C18 and diol stationary phases. All the isolated fractions were analyzed by UHPLC-QTOF-MS to determine their relative composition. According to our results, classical techniques allowed good separation of alkaloids, catechins, dimers, and trimers, but were inefficient for oligomeric PCs. Preparative C18-HPLC method allowed the attainment of high relative composition of fractions enriched with alkaloids, catechins, and PCs with degree of polymerization (DP) < 4. However, the best results were obtained by preparative diol-HPLC, providing a separation according to the increasing DP. According to the mass spectrometry fragmentation pattern, the nine isolated fractions (Fractions II-X) consisted of exclusively individual PCs and their corresponding isomers (same DP). In summary, an efficient, robust, and fast method using a preparative diol column for the isolation of PCs is proposed. Regarding DPPH• and ABTS•+ scavenging activity, it increases according to the DP; therefore, the highest activity was for cocoa extract > PCs > monomers. Thereby, cocoa procyanidins might be of interest to be used as alternative antioxidants.

Occurrence of allergen proteins in wines from Chilean market.

Casein and ovalbumin are proteins commonly used as wine fining agents that may trigger allergic reactions in susceptible individuals. Therefore, their occurrence in wines could become a health risk, moreover when their presence is not reported. The objective of this work was to determine the occurrence of casein and ovalbumin in Chilean wines. Proteins were extracted by ultrafiltration and digested applying an optimised ultrasound-assisted method. Peptides were quantified by validated tandem mass spectrometry method using stable isotope dilution analysis. Optimal digestion was achieved in 3 minutes at a 1:10 enzyme protein ratio. The method was validated following ICH guidelines showing determination coefficients R2 ≥ 0.99, repeatability, and intermediate precision with RSD values <1.95% and recoveries from 89.8% to 115.1% (RSD < 5.84%). The method was applied to analyse 60 wine samples. Fifty-six samples showed quantifiable levels, from which 28% presented a total casein and ovalbumin concentration equal or higher to the European limit (0.25 mg L-1).