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Propolis is a resinous bee product with a very complex composition, which is dependent upon the plant sources that bees visit. Due to the promising antimicrobial activities of red Brazilian propolis, it is paramount to identify the compounds responsible for it, which, in most of the cases, are not commercially available. The aim of this study was to develop a quick and clean preparative-scale methodology for preparing fractions of red propolis directly from a complex crude ethanol extract by combining the extractive capacity of counter-current chromatography (CCC) with preparative HPLC. The CCC method development included step gradient elution for the removal of waxes (which can bind to and block HPLC columns), sample injection in a single solvent to improve stationary phase stability, and a change in the mobile phase flow pattern, resulting in the loading of 2.5 g of the Brazilian red propolis crude extract on a 912.5 mL Midi CCC column. Three compounds were subsequently isolated from the concentrated fractions by preparative HPLC and identified by NMR and high-resolution MS: red pigment, retusapurpurin A; the isoflavan 3(R)-7-O-methylvestitol; and the prenylated benzophenone isomers xanthochymol/isoxanthochymol. These compounds are markers of red propolis that contribute to its therapeutic properties, and the amount isolated allows for further biological activities testing and for their use as chromatographic standards.

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Caffeine is present in a large number of beverages and is an additive used in dietary supplements. Therefore, the concern about its quality and safety for consumers has been increasing and hence requires faster and simpler analytical methods to determine the caffeine amount. The high-throughput analysis is an appropriate solution to pharmaceuticals, bioanalysis, forensic and food laboratory routines. In this sense, Venturi easy ambient sonic-spray ionization mass spectrometry (V-EASI-MS), a specific ambient ionization source, is suitable to enable direct analysis of sample solutions in real time and is appropriate to be coupled to liquid chromatography (LC). The development of an on-line solid phase extraction system coupled to V-EASI-MS optimizes the advantages of LC-MS hyphenation by enhancing the figures of merit of the analytical method according to AOAC guidelines and simultaneously minimizing the runtime analysis to 1.5 min per sample, as well as sample preparation steps and solvent consumption, which is currently a challenge for quantitative applications of ambient ionization MS.

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Phytophthora parasitica is an oomycete pathogen that infects a broad range of crops of worldwide economic interest; among them are citrus species. In general, some Citrus and the rootstocks of related genera offer considerable resistance against P. parasitica; therefore, understanding the mechanisms involved in the virulence of this pathogen is crucial. In this work, P. parasitica secondary metabolite production was studied using matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) and ultrahigh-performance liquid chromatography coupled with electrospray ionization quadrupole time-of-flight tandem mass spectrometry (UHPLC/ESI-Q-TOF-MS) combined with chemometric tools, and its metabolic profile was evaluated under the influence of Citrus sunki (a highly susceptible host) and Poncirus trifoliata (a resistant genotype) extracts. The root extracts of Citrus sunki had an influence on the growth and hyphae morphology, and the root extracts of P. trifoliata had an influence on the zoospore behavior. In parallel, the spatial distribution of several metabolites was revealed in P. parasitica colonies using MALDI-MSI, and the metabolite ion of m/z 246 was identified as the protonated molecule of Arg-Ala. The MALDI-MSI showed variations in the surface metabolite profile of P. parasitica under the influence of the P. trifoliata extract. The P. parasitica metabolome analysis using UHPLC-ESI-Q-TOF-MS resulted in the detection of Arg-Gln (m/z 303.1775), as well as L-arginine (m/z 175.1191) and other unidentified metabolites. Significant variations in this metabolome were detected under the influence of the plant extracts when evaluated using UHPLC-ESI-Q-TOF-MS. Both techniques proved to be complementary, offering valuable insights at the molecular level when used to assess the impact of the plant extracts on microbial physiology in vitro. The metabolites identified in this study may play significant roles in the interaction or virulence of P. parasitica, but their functional characterization remains to be analyzed. Overall, these data confirm our initial hypotheses, demonstrating that P. parasitica has the capabilities of (i) recognizing host signals and altering its reproductive programing and (ii) distinguishing between hosts with varying responses in terms of reproduction and the production of secondary metabolites.

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Full-thickness cutaneous trauma, due to the lack of dermis, leads to difficulty in epithelialization by keratinocytes, developing a fibrotic scar, with less elasticity than the original skin, which may have disorders in predisposed individuals, resulting in hypertrophic scar and keloids. Biomedical materials have excellent characteristics, such as good biocompatibility and low immunogenicity, which can temporarily replace traditional materials used as primary dressings. In this work, we developed two dermal matrices based on Nile tilapia collagen, with (M_GAG) and without (M) glycosaminoglycans, using a sugarcane polymer membrane as a matrix support. To assess the molecular mechanisms driving wound healing, we performed qualitative proteomic analysis on the wound bed in an in vivo study involving immunocompetent murine models at 14 and 21 days post-full-thickness skin injury. Gene Ontology and Pathway analysis revealed that both skins were markedly represented by modulation of the immune system, emphasizing controlling the acute inflammation response at 14 and 21 days post-injury. Furthermore, both groups showed significant enrichment of pathways related to RNA and protein metabolism, suggesting an increase in protein synthesis required for tissue repair and proper wound closure. Other pathways, such as keratinization and vitamin D3 metabolism, were also enriched in the groups treated with M matrix. Finally, both matrices improved wound healing in a full post-thick skin lesion. However, our preliminary molecular data reveals that the collagen-mediated healing matrix lacking glycosaminoglycan (M) exhibited a phenotype more favorable to tissue repair, making it more suitable for use before skin grafts.

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Araticum is a native species of the Brazilian Cerrado with a high potential for exploitation. Several studies have stated that araticum is a rich source of phytochemicals with multifaceted biological actions. However, little information is available regarding the characterization of phytochemicals found in the pulp of this fruit. In this context, this study aimed to carry out a comprehensive characterization of phytochemicals present in the araticum pulp using ultra-high-performance liquid chromatography coupled to a quadrupole time-of-flight mass spectrometer (UHPLC-ESI-QTOF-MS/MS). The antioxidant potential of araticum pulp was also evaluated. UHPLC-ESI-QTOF-MS/MS profiling of the phytochemicals allowed for the identification and annotation of 139 phytochemicals, including organic acids, jasmonates, iridoids, phenolic compounds, alkaloids, annonaceous acetogenins, fatty acid derivatives, and other compounds. Among them, 116 compounds have been found for the first time in araticum pulp. Phenolic compounds and their derivatives represented about 59% of the phytochemicals identified in the extract. Moreover, araticum pulp showed high total phenolic compound content and antioxidant activity. The majority of identified phytochemicals have been associated with key roles in the plant's defense mechanisms against biotic and abiotic stress factors in the Cerrado environment. Furthermore, many of these phytochemicals found in the araticum pulp are already widely recognized for their beneficial effects on human health. Our findings showed that the araticum fruit contains different classes of phytochemicals that exert various biological activities, both in the plant itself and in humans.

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Capsiate and phenolics present in the free, esterified, glycosylated, and insoluble-bound forms of BRS Moema peppers were characterized and quantified using UHPLC-ESI-MS/MS. Additionally, the in vitro antiproliferative activity of BRS Moema extract was evaluated. The peppers showed considerable quantities of capsiate and phenolic compounds. Esterified phenolics were the main fraction, followed by the insoluble-bound fraction, indicating that relying solely on the extraction of soluble phenolics may underestimate the total phenolic content. Among the fourteen phenolics identified in extract fractions, gallic acid was the major constituent. Phenolic fractions displayed high antioxidant capacity by TEAC and ORAC assays. Nevertheless, the correlation between phenolic compounds and antioxidant activity suggested that other bioactive or phenolic compounds may contribute to the overall phenolic compounds and antioxidant capacity of the obtained fractions. Concerning the antiproliferative activity, the extract did not exhibit any effect on cell proliferation within the evaluated concentration range. These findings indicated that BRS Moema peppers can serve as a rich source of phenolic compounds. Therefore, fully utilizing them could bring advantages to the food and pharmaceutical industries, as well as to consumers and producers.

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The effect of L-165041 (PPARδ-agonist) on decreasing apoptosis and intracellular lipid content was assessed in fresh and vitrified-warmed in vitro -produced bovine embryos. It was hypothesised that the addition of L-165041 to the culture medium enhances development and cryopreservation. Oocytes were allocated to one of two treatments: control-standard culture medium, or L-165041 added to the medium on day1 with no media change. Ultrastructure, cleavage, and blastocyst rates were evaluated in fresh, and in post-vitrification cultured embryos by optical and electronic microscopy. A subset of fresh embryos were fixed for TUNEL assay and for Sudan-Black-B histochemical staining. Vitrified-warmed embryos were assessed using MALDI-MS technique. Cleavage and blastocyst rates (control 49.4±5.2, L-165041 51.8±4.3) were not influenced by L-165041. The proportion of inner cell mass cells (ICM) was higher in fresh embryos, and the rate of total and ICM apoptosis was lower in L-165041. In warmed-embryos, total and ICM apoptosis was lower in L-165041. The overall hatching rate was higher in L-165041 (66.62±2.83% vs 53.19±2.90%). There was less lipid accumulation in fresh L-165041-embryos. In conclusion, the use of L-165041 is recommended to improve the viability of in vitro -derived bovine embryos.

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The indiscriminate use of chemical pesticides increasingly harms the health of living beings and the environment. Thus, biological control carried out by microorganisms has gained prominence, since it consists of an environmentally friendly alternative to the use of pesticides for controlling plant diseases. Herein, we evaluated the potential role of endophytic Trichoderma strains isolated from forest species of the Cerrado-Caatinga ecotone as biological control agents of crop pathogenic fungi. Nineteen Trichoderma strains were used to assess the antagonistic activity by in vitro bioassays against the plant pathogens Colletotrichum truncatum, Lasiodiplodia theobromae, Macrophomina phaseolina, and Sclerotium delphinii isolated from soybean, cacao, fava bean, and black pepper crops, respectively. All Trichoderma strains demonstrated inhibitory activity on pathogen mycelial growth, with maximum percent inhibition of 70% against C. truncatum, 78% against L. theobromae, 78% against M. phaseolina, and 69% against S. delphinii. Crude methanol extracts (0.5 to 2.0 mg mL-1) of Trichoderma strains were able to inhibit the growth of C. truncatum, except Trichoderma sp. T3 (UFPIT06) and T. orientale (UFPIT09 and UFPIT17) at 0.5 mg mL-1, indicating that the endophytes employ a biocontrol mechanism related to antibiosis, together with multiple mechanisms. Discriminant metabolites of Trichoderma extracts were unveiled by liquid chromatography-tandem mass spectrometry-based metabolomics combined with principal component analysis (PCA), which included antifungal metabolites and molecules with other bioactivities. These results highlight the biocontrol potential of Trichoderma strains isolated from the Cerrado-Caatinga ecotone against crop pathogenic fungi, providing support for ongoing research on disease control in agriculture.

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This work aimed to carry out phytochemical prospecting and evaluate the antioxidant potential of Diplopterys pubipetala, a species of Malpighiaceae family that has not yet been studied.In qualitative analyses of hydroethanolic extracts of leaves and stems were detected the presence of flavonoids, alkaloidsand terpenes. The histochemical evaluation evidenced a greater distribution of these compounds in the tissues of leaf when compared with those of stem. The analysis by mass spectrometry allowed the identification of prenylated xanthones and glycoside flavonoids that have not yet been reported in the literature. The antioxidant activity of the stem extract was considered moderate (IAA = 0.79), but the leaves presented a strong antioxidant activity (IAA = 1.6). In this work we present information about the phytochemicals of D. pubipetala, showing that the species is promising in obtaining compounds with medicinal potential mainly antioxidant potential.

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The strong odour of faeces and excessive production of gases in some dog breeds have long been a concern of owners. The pet food industry uses nutritional alternatives, such as high-quality ingredients and additives, to improve the odour of faeces. However, there are still some dog breeds, such as the French Bulldog, that present this problem due to the presence into the large intestine of indigested protein. Therefore, a deeper understanding of the volatile compounds that influence the odour of dog faeces is important. This study aimed to identify changes of faecal odour compounds that are most prevalent in French Bulldogs based on food containing different high-quality protein sources and their effect in sensory analysis. Four maintenance foods with different protein sources were formulated: P, poultry meal food; W, wheat gluten food; PW, poultry meal and wheat gluten food; and PWH, poultry meal, wheat gluten, and hydrolysed protein food. Eight adult French Bulldogs were arranged in a 4x4 Latin square design and adapted to foods for 28 days. Fresh faeces were collected for analysis of volatile organic compounds (VOCs) and sensory analysis. The means were compared by SAS, and statistical significance was indicated by p ≤ 0.05. No adverse effects were observed in the animals regarding VOCs, and a significant difference was observed in two of the 68 compounds identified. The animals fed a P food had higher concentrations of phenol in the faeces, whereas the indole compound was present at higher concentrations in animals fed the W food. P food was associated with higher odour perception during sensory evaluation. In summary, the source of protein in the foods had little impact on the composition of VOCs, and a greater perception of the odour was determined by sensory analysis when foods containing animal protein were administered.

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