1H NMR and UPLC-HRMS-based metabolomic approach for evaluation of the grape maturity and maceration time of Touriga Nacional wines and their correlation with the chemical stability.

Touriga Nacional is a well-adapted Portuguese grape variety in São Francisco River Valley (northeastern Brazil). Nevertheless, it has only been indicated to short-term consumption because of the lack of chemical stability, which is attributed to low grape acidity and incomplete phenolic maturity. Therefore, we used Ultra-Performance Liquid Chromatography coupled High-resolution Mass Spectrometry, Nuclear Magnetic Resonance and chemometrics (PCA and PLS-DA) to evaluate the grape maturity and maceration time on chemical composition of wines from two harvest seasons. Moreover, we investigated how these experimental factors could affect their chemical stability. Grapes maturity showed to be the main effect. Overall, phenolic acids and short-chain organic acids were found to be at higher levels in wines produced with unripe grapes from February and shorter maceration time (p < 0.05). Proanthocyanidins and other flavonoids were increased in wines macerated for longer time using overripe grapes harvested in July. Furthermore, stable wines were made from overripe grapes, which contained more galacturonic acid.

Insecticidal Compound from Himatanthus drasticus Latex against Cowpea Infestation by Callosobruchus maculatus (Coleoptera: Chrysomelidae).

Vigna unguiculata is an important source of proteins and energy for humans and animals. However, postharvest losses caused by Callosobruchus maculatus can reach from 20 to 100% of stored seeds. In this study, the insecticide potential of compounds extracted from Himatanthus drasticus latex was assessed. The latex was extracted with ethanol (70%) and then partitioned through sequential use of hexane and chloroform. These fractions were investigated by chromatography to determine their chemical composition. Plumieride, identified in a hydroalcoholic subfraction, was tested for insecticidal activity against C. maculatus. The ethanolic fraction (LC50 = 0.109; LC90 = 0.106%) and the plumieride (LC50 = 0.166; LC90 = 0.167%) were lethal to larvae. Plumieride (0.25%) delayed larval development, and mortality reached 100%. Its inhibitory action on intestinal α-amylase from larvae was higher (89.12%) than that of acarbose (63.82%). Plumieride (0.1%) inhibited the enzyme α-amylase in vivo in the larval intestine. This result was confirmed by a zymogram test performed by SDS-PAGE when the enzyme electrophoresed on gel copolymerized with starch. When spread on seeds, the hydroalcoholic fraction (1.0%) reduced infestation. The loss of seed mass was 5.26% compared to the control (44.97%). The results confirm the effect of latex compounds in protecting stored seeds against weevil infestation.

Chemical profiling of secondary metabolites from Himatanthus drasticus (Mart.) Plumel latex with inhibitory action against the enzymes α-amylase and α-glucosidase: In vitro and in silico assays.

ETHNOPHARMACOLOGICAL RELEVANCE: Himatanthus drasticus is an important medicinal plant whose latex is traditionally used in Northeast Brazil to treat various diseases, including diabetes. The use of α-amylase and α-glucosidase inhibitors can be an effective strategy to modulate levels of postprandial hyperglycemia via control of starch metabolism. AIMS OF THE STUDY: This study aimed to verify if H. drasticus latex has inhibitory activity against enzymes linked to type 2 diabetes, besides chemically characterizing the metabolites responsible for such activities. In addition, in silico analysis was performed to support the traditional claim of possible antidiabetic activity of this latex. MATERIALS AND METHODS: Latex from H. drasticus stems was sequentially partitioned with n-hexane (FHDH), CHCl3 (FHDC) and EtOH (FHDHA). Wash extraction of the FHDHA fraction was performed to obtain the other extract fractions. The FHDHA was submitted to chromatography in a SPE C18 cartridge using gradient elution with MeOH/H2O to produce five fractions: FHDHA1, FHDHA2, FHDHA3, FHDHA4 and FHDHA5. The FHDHA1 was subjected to semi-preparative reverse phase HPLC. Lineweaver-Burk plots were used to investigate the kinetic parameters of α-amylase and α-glucosidase inhibitory activity. The interactions between plumieride and porcine pancreatic α-amylase and α-glucosidase were analyzed through an in silico molecular docking study. RESULTS: Phytochemical identification of compounds present in the FHDHA fraction of H. drasticus latex was possible by 1H, 13C NMR analysis and mass spectrometry, and the results were compared with the literature. The identified compounds were α-ethyl glucoside, protocatechuic acid, 3-O-caffeoylquinic acid, 15-demethylplumieride acid, 5-O-caffeoylquinic acid, caffeic acid, vanillic acid, plumieride, and catechin. The inhibition results of the fractions tested against α-amylase and α-glucosidase showed inhibitory activities dependent on the increase of fractions and compound concentrations. The IC50 results obtained from FHDHA, FHDHA1 and plumieride fractions against α-amylase were 36.46, 72.61, 33.87 µg/mL respectively. The IC50 of plumieride was the closest to that of acarbose (22.52 µg/mL), a result similar to that obtained for α-glucosidase. The type of inhibition was competitive for both enzymes. CONCLUSIONS: There was strong inhibition of α-amylase and α-glucosidase by FHDHA, FHDHA1 and plumieride, suggesting that these enzymes slow glucose absorption.

1H NMR and LC-MS-based metabolomic approach for evaluation of the seasonality and viticultural practices in wines from São Francisco River Valley, a Brazilian semi-arid region.

São Francisco River Valley (SFRV) is a wine-producing semi-arid region in Brazil. Therefore, we used a 1H NMR and UPLC-MS-based metabolomic approach coupled to chemometrics to evaluate the variability in Chenin Blanc and Syrah wines for two harvest seasons, two vine training system and six rootstocks. Overall, the secondary metabolites were influenced by the three factors studied, whereas the primary metabolites were only by the seasonality. Chenin Blanc wines made in December presented higher content of an unidentified carbohydrate. In Syrah wines, glycerol, tartaric acid, succinic acid and 2,3-butanediol were greater in December, while proline and lactic acid were more abundant in July. For training system, caffeic acid derivatives were increased in wines produced from espalier. Lyre system increased phenolic compounds, organic acids and apocarotenoids. The effect of the rootstocks was less pronounced, affecting basically caffeic acid derivatives. Thus, we expect that our results may assist the winemakers to improve the SFRV wine quality.

The intake of dry cashew apple fiber reduced fecal egg counts in Haemonchus contortus-infected sheep.

Helminthiasis is a common disease in which parasite resistance is frequently caused by inadequate administration of anthelmintics in small ruminant production. Since phytotherapy may be an adjuvant for parasite control, we assessed whether the ingestion of cashew apple fiber (Anacardium occidentale) would reduce Haemonchus contortus infection in Santa Inês sheep. Twenty-one male sheep with mean age of 240 ±â€¯9.7 days were dewormed, infected with 4000 L3 of H. contortus Embrapa2010 (day 0 - D0) and on D28 were divided into three equally sized experimental groups: 1) control (no treatment), 2) treated with anthelmintic (monepantel, 2.5 mg/kg PV) and 3) cashew apple fiber (0.3% BW) for 7 days of adaptation plus 28 days (D63). The animals were weighed weekly for diet adjustment and individual EPGs were performed twice a week. Corn silage was given ad libitum after each animal had eaten all the cashew apple fiber, which always occurred due to its palatable flavor. The silage, cashew apple fiber and leftovers were weighed daily and the samples were analyzed for dry matter. In cashew apple fiber, the total polyphenol contents were determined spectrophotometrically and the phenol compounds were identified by LC-MS. Cashew apple fiber contained 93.6% DM, 13.0% CP, 68.7% NDF, 47.6% FDA, 1.3% MM, 1.9% EE and 22.3% LIG. Twenty phenolic compounds were detected, among them phenolic acids and flavonoids, including glycosylated ones. The general EPG averages were statistically different among control, anthelmintic and cashew groups (3449, 14 and 2070, respectively), while the mean total weight gain did not differ (3.21, 3.20 and 1.94 kg, respectively) (p < 0.05). In relation to the control group, the anthelmintic showed efficacy of 99.6% and the cashew apple fiber 40.8%. Phenolic compounds appear to play an important role in the anthelmintic activity of cashew apple fiber. Thus, its use as an adjuvant in the control of H. contortus can be encouraged in regions where it is available at low cost, mitigating the use of veterinary drugs, reducing environmental contamination by agroindustrial residues and promoting the more sustainable production of small ruminants.

New flavone and other compounds from Tephrosia egregia: assessing the cytotoxic effect on human tumor cell lines

ABSTRACT The organic extracts from stems, roots and leaves of Tephrosia egregia Sandwith, Fabaceae, provided a new flavone, 5-hydroxy-8-(1",2"-epoxy-3"-hydroxy-3"-methylbutyl)-7-methoxyflavone (1), in addition to eleven known compounds: pongaflavone (2), praecansone B (3), 12a-hydroxyrotenone (4), praecansone A, 2',6'-dimethoxy-4',5'-(2",2"-dimethyl)-pyranochalcone, pongachalcone, maackiain, β-sistosterol and its glucoside, p-cumaric acid and cinnamic acid. The structures of all compounds were established on the basis of spectroscopic methods, mainly 1D and 2D NMR and HRESIMS, involving comparison with literature data. Cytotoxicity of compounds 1-4 was evaluated against AGP-01 (cancerous ascitic fluid), HCT-116 (colon adenocarcinoma), HL-60 (leukemia), PC-3 (prostate carcinoma), SF-295 (glioblastoma) and SKMEL 28 (melanoma) cell lines.

Stabilizing effect of montmorillonite on acerola juice anthocyanins.

This study was conducted to evaluate color and anthocyanin stability of clarified acerola juice (CAJ) as affected by montmorillonite (Mnt) at different concentrations (0-6 wt%, dry basis). While non-complexed CAJ suffered noticeable color degradation with time and pH variations, the presence of Mnt (especially at 4-6 wt%) not only changed the initial color of CAJ but also made it more stable with time and pH changes. CAJ/Mnt mixtures were ultracentrifuged in order to separate them into supernatants and anthocyanin-complexed Mnt precipitates. The supernatants presented decreasing anthocyanin contents with increasing Mnt concentrations, indicating pigment retention by the precipitates. X-ray diffraction of precipitates showed that Mnt interlayer spacing was increased by increasing anthocyanin/Mnt ratios, corroborating anthocyanin intercalation. FTIR revealed a band at 1530 cm-1 ascribed to formation of anthocyanin-Mnt complexes. Moreover, chromatograms indicated the selective adsorption of two compounds by Mnt, which were identified by LC-MS as cyanidin-3-O-rhamnoside and pelargonidin-3-O-rhamnoside.

UPLC-qTOF-MS/MS-based phenolic profile and their biosynthetic enzyme activity used to discriminate between cashew apple (Anacardium occidentale L.) maturation stages.

Cashew immature and ripe peduncles (Anacardium occidentale L.) from orange- and red-colored clones CCP 76 and BRS 189, respectively, were prepared as juice or fibrous fraction and submitted to UPLC-MS analyses, while the soluble fraction was also submitted to enzymatic evaluation. Cinnamoyl glucoside was present in ripe juice samples from both cashew clones, while monogalloyl diglucoside and digalloyl glucoside were present in immature juice samples from both cashew clones. Four compounds were found at immature fiber of both clones, anacardic acids (1, 2, 3) and GA19. The phenolic biosynthetic pathway was evaluated in juice samples and phenylalanine ammonia-lyase activity decreased significantly during the development, although it was much higher in ripe CCP 76. UDP-glycosyltransferases activity differed between clones, however its product cinnamoyl glucoside was a possible chemical marker of ripe juice samples from both clones. Flavonol synthase showed the highest specific activity in both cashew clones and its product, flavonols were identified in cashew apple at immature and ripe stages.

New Cytotoxic Bibenzyl and Other Constituents from Bauhinia ungulata L. (Fabaceae).

A new bibenzyl, 2'-hydroxy-3,5-dimethoxy-4-methylbibenzyl (1) and four known compounds identified as 2'-hydroxy-3,5-dimethoxybibenzyl (2), liquiritigenin (3), guibourtinidol (4) and fisetinidol (5) were isolated from the roots of Bauhinia ungulata L. Phytochemical investigations of the stems of B. ungulata led to the isolation of the known compounds identified as liquiritigenin (3), guibourtinidol (4), fisetinidol (5), taraxerol (6), betulinic acid (7), taraxerone (8), glutinol (9), a mixture of sitosterol (10) and stigmasterol (11), pacharin (12), naringenin (13) and eriodictyol (14). The structures of these compounds were elucidated on the basis of their spectral data (IR, MS, 1D- and 2D-NMR). The cytotoxicity of the bibenzyl 1 has been evaluated against four human cancer cell lines, showing the IC50 values of 4.3 and 6.5 µg ml-1 against pro-myelocytic leukemia (HL-60) and cervical adenocarcinoma (HEP-2) cell lines, respectively. This article also registers for the first time the 13 C-NMR data of the known bibenzyl 2.

Synthesis, antibacterial and cytotoxic activities of new biflorin-based hydrazones and oximes.

Biflorin 1 is a biologically active quinone, isolated from Capraria biflora. Five new biflorin-based nitrogen derivatives were synthesized, of which two were mixtures of (E)- and (Z)- isomers: (Z)-2a, (Z)-2b, (Z)-3a, (Z)- and (E)-3b, (Z)- and (E)-3c. The antibacterial activity was investigated using the microdilution method for determining the minimum inhibitory concentration (MIC) against six bacterial strains. Tests have shown that these derivatives have potential against all bacterial strains. The cytotoxic activity was also evaluated against three strains of cancer cells, but none of the derivatives showed activity.