UPLC-QTOF-MS/MS-based metabolomic approach and gastroprotective effect of two chemotypes of Egletes viscosa (L.) less. against ethanol-induced gastric ulcer in mice.

ETHNOPHARMACOLOGICAL RELEVANCE: Egletes viscosa (L.) (macela) is a native wild herb that can be found in different states of northeastern Brazil. The infusions of its flower buds are traditionally used for the treatment of gastrointestinal disorders. E. viscosa possesses two chemotypes (named A and B), distinguishable by the composition of the essential oil from the flower buds. Although there are previous studies of the gastroprotective effect of the isolated constituents of E. viscosa, its infusions have not been investigated yet. AIM OF THE STUDY: The present study aimed to evaluate and compare the chemical composition and the gastroprotective effect of flower bud infusions of E. viscosa from chemotype A (EVCA) and chemotype B (EVCB). MATERIALS AND METHODS: Sixteen infusions were brewed with flower buds according to the traditional preparation mode and were analyzed through a UPLC-QTOF-MS/MS based metabolomic approach for determination of their metabolic fingerprints and quantification of bioactive compounds. Afterward, these data were analyzed by chemometric methods (OPLS-DA) for discrimination of the two chemotypes. Additionally, infusions of EVCA and EVCB (50, 100 and 200 mg/kg, p.o.) were evaluated on gastric ulcers induced by absolute ethanol (96%, 0.2 mL, p.o.) in mice. To elucidate the gastroprotective mechanisms, the effect of EVCA and EVCB on gastric acid secretion and gastric wall mucus was determined and the role of TRPV1 channels, prostaglandins, nitric oxide and KATP channels were assessed. Moreover, the oxidative stress-related parameters and the histological aspects of the stomach tissue were analyzed. RESULTS: The chemotypes can be discriminated from each other using UPLC-QTOF-MS/MS chemical fingerprints. Both chemotypes presented similar chemical compositions, consisting basically of caffeic acid derivatives, flavonoids and diterpenes. The quantification of bioactive compounds demonstrated that chemotype A possesses more ternatin, tanabalin and centipedic than chemotype B. EVCA and EVCB (50, 100 and 200 mg/kg, p.o.) significantly decreased the severity of ethanol-induced gastric lesions, as shown by a reduction in histological alterations and leucocyte infiltration in gastric tissue. The gastroprotective mechanism of both infusions involves an antioxidant effect, maintenance of gastric mucus and reduction gastric secretion. Stimulation of endogenous prostaglandins and nitric oxide release, activation of TRPV1 channels, and KATP channels are also involved in the gastroprotection of the infusions. CONCLUSION: The gastroprotective effect of EVCA and EVCB was equivalent and mediated through antioxidant and antisecretory actions, including the activation of TRPV1 receptors, stimulation of endogenous prostaglandins and nitric oxide, and opening of KATP channels. The presence of caffeic acid derivatives, flavonoids and diterpenes in both infusions is involved in mediating this protective effect. Our findings support the traditional use of infusions of E. viscosa for gastric disorders regardless of the chemotype.

Untargeted metabolomic profile of recovered bioactive compounds by subcritical water extraction of acerola (Malpighia emarginata DC.) pomace.

The untargeted metabolomics approach was used to compare the chemical profiles of acerola (Malpighia emarginata DC.) pomace extracts. The effect of drying the raw material before subcritical water extraction (SWE) at different temperatures on the yield, phenolic content, and in vitro antioxidant activity was evaluated. The results were compared with those obtained via Soxhlet and the findings suggest that SWE saves time (15 min) and solvent for extracting valuable components as compared to Soxhlet (6 h). An increase in temperature significantly improved the extraction yield (23.9 to 33.4 %), phenolic content (119.1 to 362 mgGAEg-1), and antioxidant activity, and higher values were obtained with SWE as compared to Soxhlet. The most abundant compounds detected by UPLC-ESI-QTOF-MS were ascorbic acid, kaempferol, quercetin, and isorhamnetin. The investigation of different moisture contents in the SWE showed promising results for eliminating the drying operation, saving time and energy, and obtaining highly concentrated phenolic-rich by-products.

Anti-Candida Properties of Gossypium hirsutum L.: Enhancement of Fungal Growth, Biofilm Production and Antifungal Resistance.

(1) Background: Candida is a genus of yeasts with notable pathogenicity and significant ability to develop antimicrobial resistance. Gossypium hirsutum L., a medicinal plant that is traditionally used due to its antimicrobial properties, has demonstrated significant antifungal activity. Therefore, this study investigated the chemical composition and anti-Candida effects of aqueous (AELG) and hydroethanolic (HELG) extracts obtained from the leaves of this plant. (2) Methods: The extracts were chemically characterized by UPLC-QTOF-MS/MS, and their anti-Candida activities were investigated by analyzing cell viability, biofilm production, morphological transition, and enhancement of antifungal resistance. (3) Results: The UPLC-QTOF-MS/MS analysis revealed the presence of twenty-one compounds in both AELG and HELG, highlighting the predominance of flavonoids. The combination of the extracts with fluconazole significantly reduced its IC50 values against Candida albicans INCQS 40006, Candida tropicalis INCQS 40042, and C. tropicalis URM 4262 strains, indicating enhanced antifungal activity. About biofilm production, significant inhibition was observed only for the AELG-treated C. tropicalis URM 4262 strain in comparison with the untreated control. Accordingly, this extract showed more significant inhibitory effects on the morphological transition of the INCQS 40006 and URM 4387 strains of C. albicans (4) Conclusions: Gossypium hirsutum L. presents promising antifungal effects, that may be potentially linked to the combined activity of chemical constituents identified in its extracts.

UPLC-MS-ESI-QTOF Analysis and Antifungal Activity of Aqueous Extracts of Spondias tuberosa.

This study aimed to identify the chemical composition of the Spondias tuberosa aqueous leaf and root extracts (EALST and EARST) and to evaluate their effect, comparatively, against opportunistic pathogenic fungi. Ultra-Performance Liquid Chromatography Coupled to a Quadrupole/Time of Flight System (UPLC-MS-ESI-QTOF) was employed for chemical analysis. Candida albicans and C. tropicalis standard strains and clinical isolates were used (CA INCQS 40006, CT INCQS 40042, CA URM 5974, and CT URM 4262). The 50% Inhibitory Concentration for the fungal population (IC50) was determined for both the intrinsic action of the extracts and the extract/fluconazole (FCZ) associations. The determination of the Minimum Fungicidal Concentration (MFC) and the verification of effects over fungal morphological transitions were performed by subculture in Petri dishes and humid chambers, respectively, both based on micro-dilution. UPLC-MS-ESI-QTOF analysis revealed the presence of phenolic and flavonoid compounds. The association of the extracts with fluconazole, resulted in IC50 values from 2.62 µg/mL to 308.96 µg/mL. The MFC of the extracts was ≥16,384 µg/mL for all tested strains, while fluconazole obtained an MFC of 8192 µg/mL against C. albicans strains. A reduction in MFC against CA URM 5974 (EALST: 2048 µg/mL and EARST: 1024 µg/mL) occurred in the extract/fluconazole association.

From mango by-product to food packaging: Pectin-phenolic antioxidant films from mango peels.

The objective of the study was to prepare active films based on pectin and polyphenol-rich extracts from Tommy Atkins mango peels. Aqueous and methanolic extracts showed a variety of phenolic compounds that were identified by UPLC-MS analysis, and a high content of total phenolics that were quantified by the Folin-Ciocalteau method. The methanolic extract showed better results in antioxidant tests and was more effective in inhibiting the growth of Gram-positive and Gram-negative bacteria. The pectin extracted from mango peels showed good thermal stability and a degree of methoxylation of 58.3% by 1H NMR. The films containing the phenolic extracts showed lower water vapor permeability when compared to the control film (without any phenolic extracts). The incorporation of the extracts led to an increase in elongation (ε) and a decrease in tensile strength (σ) and modulus of elasticity (Y). The films with aqueous or methanolic extracts showed higher antioxidant activity in terms of inhibition of the DPPH radical. Therefore, the films developed in this work are presented as a promising alternative for food packaging and/or coating applications.

UPLC-HRMS and NMR applied in the evaluation of solid-phase extraction methods as a rational strategy of dereplication of Phyllanthus spp. aiming at the discovery of cytotoxic metabolites.

The classical approach to drug discovery from natural products (NP's) requires strenuous and complex purification steps for the isolation and structural elucidation. Modern strategies as dereplication aim to accelerate the identification of known compounds present in a crude or partially purified extract. In this work, we investigated the influence of the solid-phase extraction (Oasis, Plexa, and Agilent C18 cartridges with and without organic modifiers) chemical profile obtained by UPLC-QTOF-MS and NMR and cytotoxicities of aqueous extracts from Phyllanthus niruri and P. amarus. Our results showed differences between the SPE cartridges and the mass recovered. P. niruri showed higher mass recovery than P. amarus indicating a higher amount of secondary metabolites. The UPLC-QTOF-MS analysis revealed that P. niruri crude extract presents higher contents of phenolic compounds than P. amarus. According to NMR analysis, P. niruri contained more tyrosine, corilagin, and glycosidic residues while P. amarus, presented higher content of ellagic acid. The different stationary phases, as well as mobile phases for exploratory SPE, enabled the exploitation of the different chemical functionalities within the Phyllanthus species. The SPE (MeOH:H2O 70:30 with C18 cartridges) samples showed greater in vitro cytotoxicity than the crude extracts, with IC50 ranging from 8.01 to 94.92 µg mL-1 against the tumor lines tested. The solid phase extraction allowed the concentration of molecules with desirable physicochemical characteristics, which might increase the hit of therapeutically useful substances.

Endophytic Fungus Isolated From Achyrocline satureioides Exhibits Selective Antiglioma Activity-The Role of Sch-642305.

Glioblastoma is the most devastating primary brain tumor. Current treatment is palliative, making necessary the development of new therapeutic strategies to offer alternatives to patients. Therefore, endophytes represent an interesting source of natural metabolites with anticancer potential. These microorganisms reside in tissues of living plants and act to improve their growth. Evidence revealed that several medicinal plants are colonized by endophytic fungi producer of antitumor metabolites. Achyrocline satureioides is a Brazilian medicinal plant characterized by its properties against gastrointestinal disturbances, anticancer and antioxidant effects. However, there are no reports describing the endophytic composition of A. satureioides. The present study proposes the isolation of endophytic fungus from A. satureioides, extract preparation, phytochemical characterization and evaluation of its antiglioma potential. Our data showed that crude extracts of endophyte decreased glioma viability with IC50 values of 1.60-1.63 µg/mL to eDCM (dichloromethane extract) and 37.30-55.12 µg/mL to eEtAc (ethyl acetate extract), respectively. Crude extracts induced cell death by apoptosis with modulation of redox status. In order to bioprospect anticancer metabolites, endophytic fungus extracts were subjected to guided fractionation and purification yielded five fractions of each extract. Six of ten fractions showed selective antiproliferative activity against glioma cells, with IC50 values ranged from 0.95 to 131.3 µg/mL. F3DCM (from eDCM) and F3EtAc (from eEtAc) fractions promoted C6 glioma toxicity with IC50 of 1.0 and 27.05 µg/mL, respectively. F3EtAc fraction induced late apoptosis and arrest in G2/M stage, while F3DCM promoted apoptosis with arrest in Sub-G1 phase. Moreover, F3DCM increased antioxidant defense and decreased ROS production. Additionally, F3DCM showed no cytotoxic activity against astrocytes, revealing selective effect. Based on promising potential of F3DCM, we identified the production of Sch-642305, a lactone, which showed antiproliferative properties with IC50 values of 1.1 and 7.6 µg/mL to C6 and U138MG gliomas, respectively. Sch-642305 promoted arrest on cell cycle in G2/M inducing apoptosis. Furthermore, this lactone decreased glioma cell migration and modulated redox status, increasing superoxide dismutase and catalase activities and enhancing sulfhydryl content, consequently suppressing reactive species of oxygen generation. Taken together, these results indicate that metabolites produced by endophytic fungus isolated from A. satureioides have therapeutic potential as antiglioma agent.

Metabolomic profile of Schinopsis brasiliensis via UPLC-QTOF-MS for identification of biomarkers and evaluation of its cytotoxic potential.

Schinopsis brasiliensis is a plant typically found in the caatinga biome (northeastern Brazil). Its leaves and bark have been used for the treatment of health dysfunctions such as cough, influenza, diarrhea, throat inflammation, and sexual impotence. However, there is a lack of knowledge regarding the chemical composition and pharmacological activities of this plant. High-performance liquid chromatography coupled to high-resolution mass spectrometry (UPLC-QTOF-MSE) allowed the partial identification of 33 compounds, including isomers from leaf, branch, and bark samples, with 16 compounds reported for the first time (corilagin, chlorogenic acid, and quercetin derivatives) in S. brasiliensis. Principal component analysis efficiently distinguished the respective parts of the plant. Orthogonal partial least squares discriminatory analysis, together with the variable importance in projection and S-Plot graphs were used to identify 23 biomarker compounds associated with cytotoxic activity against a colorectal cancer cell line.

Chemical composition, antifungal activity and potential anti-virulence evaluation of the Eugenia uniflora essential oil against Candida spp.

The development of fungal resistance to antifungal drugs has been worsening over the years and as a result research on new antifungal agents derived from plants has intensified. Eugenia uniflora L. (pitanga) has been studied for its various biological actions. In this study the chemical composition and antifungal effects of the E. uniflora essential oil (EULEO) were investigated against Candida albicans (CA), Candida krusei (CK) and Candida tropicalis (CT) standard strains. The essential oil obtained through hydro-distillation was analyzed by gas chromatography coupled to mass spectrometry (GC-MS). To determine the IC50 of the oil, the cellular viability curve and the inhibitory effects were measured by means of the oil's association with Fluconazole in a broth microdilution assay with spectrophotometric readings. The Minimum Fungicidal Concentration (MFC) was determined by solid medium subculture with the aid of a guide plate while the assays used to verify morphological changes emerging from the action of the fractions were performed in microculture chambers at concentrations based on the microdilution. Two major oil constituents stand out from the chemical analysis: selina-1,3,7(11)-trien-8-one (36.37%) and selina-1,3,7(11)-trien-8-one epoxide (27.32%). The concentration that reduced microorganismal growth was ≥8,192 µg/mL while the IC50 varied, this being between 1892.47 and 12491.80 µg/mL (oil), 10.07 - 80.78 µg/mL (fluconazole) and 18.53 - 295.60 µg/mL (fluconazole + oil). The combined activity (fluconazole + oil) resulted in indifference and antagonism. A MFC of the oil in association with fluconazole was recorded at the concentration of 8,192 µg/mL against CA and CK. The oil caused the inhibition of CA and CT morphological transition. In view of the results obtained, additional research is needed to elucidate the activity of the E. uniflora oil over genetic and biochemical processes regarding its effect on Candida spp. virulence.

Analysis by UPLC-MS-QTOF and antifungal activity of guava (Psidium guajava L.).

Psidium guajava L. is a plant widely used for food and in folk medicine all over the world. Studies have shown that guava leaves have antifungal properties. In this study, Flavonoid and Tannic fractions were tested to investigate their chemical composition and antifungal potential in vitro.21 compounds in the two fractions, presenting a higher content of phenolic compounds. The antifungal assays were performed against Candida albicans, Candida tropicalis and Candida krusei by microdilution to determine the IC50 and the cell viability curve. Minimal Fungicidal Concentration(MFC) and the inhibitory effects of the association of the fractions with Fluconazole, as well as the assays used to verify any morphological changes were performed in microculture chambers based on the concentrations from the microdilution. The IC50 of the isolated fractions and the fractions associated with each other were calculated, varying from 69.29 to 3444.62 µg/mL and the fractions associated with fluconazole varied from 925.56 to 1.57 µg/mL, it was clear that the association of the natural product with the antifungal presented a synergism. The fractions affected pleomorphism capacity and have a potential antifungal activity as they caused fungal inhibition in isolated use, potentiated the action of Fluconazole, reducing its concentration and impeding morphological transition, one of the virulence factors of the genus.

Chemical profiling of guarana seeds (Paullinia cupana) from different geographical origins using UPLC-QTOF-MS combined with chemometrics.

Paullinia cupana, commonly known as guarana, is an Amazonian fruit whose seeds are used to produce the powdered guarana, which is rich in caffeine and consumed for its stimulating activity. The metabolic profile of guarana from the two largest producing regions was investigated using UPLC-MS combined with multivariate statistical analysis. The principal component analysis (PCA) showed significant differences between samples produced in the states of Bahia and Amazonas. The metabolites responsible for the differentiation were identified by orthogonal partial least squares discriminant analysis (OPLS-DA). Fourteen phenolic compounds were characterized in guarana powder samples, and catechin, epicatechin, B-type procyanidin dimer, A-type procyanidin trimer and A-type procyanidin dimer were the main compounds responsible for the geographical variation of the samples.

Impact of bioaccessibility and bioavailability of phenolic compounds in biological systems upon the antioxidant activity of the ethanolic extract of Triplaris gardneriana seeds.

The most studied bioactive potential of phenolic compounds corresponds to antioxidant activity, which in turn, is associated with a reduction in the incidence of various human diseases. However, the total quantity of these bioactive substances in foods and medicinal preparations does not reflect the amount absorbed and metabolized by the body. The present study aimed to investigate the bioaccessibility of Triplaris gardneriana seeds ethanolic extract (EETg) by determination of phenolic composition and antioxidant activities before and after in vitro digestion as well as to estimate its bioavailability by chemical analysis of plasma and urine in animal models after oral administration. The bioaccessibility indexes of phenolic compounds in EETg were 48.65 and 69.28% in the presence and absence of enzymes, respectively. Among the identified phenolics classes, flavonoids, represented by galloylated procyanidins type B, proved to be more bioaccessible, 81.48 and 96.29% in the post-intestinal phase with and without enzymes, respectively. The oral administration in Wistar rats resulted in a significant decrease in plasma of the total antioxidant capacity, TAC, by FRAP assay 4h after beginning the experiment. For urine samples, an increase in TAC by DPPH and FRAP was observed from 1 and 4h after administration, respectively. UPLC-QTOF analysis of urine detected 2 metabolites originated from the degradation of phenolic compounds, i.e. hippuric acid and phenylacetil glycine. These results suggest that phenolic compounds in T. gardneriana are unstable under gastrointestinal conditions, being flavonoids the components with higher bioaccessibility; besides that, they showed limited bioavailability due to their rapid biotransformation and urinary elimination.

Antioxidant metabolism during fruit development of different acerola (Malpighia emarginata D.C) clones.

The present research work describes the major changes in the antioxidant properties during development of acerola from five different clones. Ripening improved fruit physicochemical quality parameters; however, total vitamin C and total soluble phenols (TSP) contents declined during development, which resulted in a lower total antioxidant activity (TAA). Despite the decline in TSP, at ripening, the anthocyanin and yellow flavonoid content increased and was mainly constituted of cyanidin 3-rhamnoside and quercetin 3-rhamnoside, respectively. The activities of oxygen-scavenging enzymes also decreased with ripening; furthermore, the reduction in vitamin C was inversely correlated to membrane lipid peroxidation, indicating that acerola ripening is characterized by a progressive oxidative stress. Among the studied clones, II47/1, BRS 237, and BRS 236 presented outstanding results for vitamin C, phenols, and antioxidant enzyme activity. If antioxidants were to be used in the food supplement industry, immature green would be the most suitable harvest stage; for the consumer's market, fruit should be eaten ripe.