Stilbenes, phenanthrenes and antiproliferative activity of Cattleya intermedia.

The phytochemical study of Cattleya intermedia (Orchidaceae) led to the isolation of two new stilbenoids and one new 9,10-dihydrophenanthrene, 4',5-dihydroxy-2',3-dimethoxy-dihydrostilbene (1), 3,6'-dihydroxy-4'-methoxy-dihydrostilbene (2) and 1,2,6-trihydroxy-3,8-dimethoxy-9,10-dihydrophenanthrene (3), named cattleymediol, cattleyol and phenanmediol, respectively, in addition to other five known compounds (4-8). The structural elucidations of the isolated compounds were carried out through the analyses of the one-dimensional 1H,1³C and NOE NMR spectra, and the 2D HSQC, HMBC, COSY and NOESY spectra, besides high-resolution mass spectrometry. In addition to this, the crude extract and its main fractions were analysed by ultra-high performance liquid chromatography coupled to high resolution mass spectrometry (UHPLC-QTOF-MS/MS), leading to the putative identification of several other compounds, including flavonoids and organic acids derivatives. Finally, the main fractions of the crude extract, and the pure compounds cattleymediol (1) and lusiantridine (7), had their antiproliferative activities evaluated against human cancerous HeLa and non-cancerous VERO cells.

Bioactive compounds by UHPLC-MS/MS and molecular networking from Myrceugenia foveolata.

Myrceugenia foveolata, commonly known as 'guamirim', was analysed using UHPLC-MS/MS and molecular networking. Hydroalcoholic extracts of leaves and stem bark were obtained using ethanol:water (70:30) as solvent. Chemical composition of the extracts was identified using UHPLC-MS/MS and tested for antioxidant activity and growth inhibition against E. coli, S. aureus, L. monocytogenes and S. enterica. Ten compounds were tentatively identified in the extracts including myricitrin, quercitrin, and betulin (in leave extracts), and avicularin, kaemferol-3-O-arabinopyroniside, 2"-O-galloylquercitrin and the acid triterpenes ursolic, sumaresinolic, asiatic and maslinic (in both extracts). Both extracts showed similar antioxidant activities, phenolic composition, and growth inhibition. The most pronounced response was observed against L. monocytogenes, with a growth inhibition rate of 73% to leaves extract and 65% do stem bark extract.

Ultrasonication and thermosonication applied in the processing of jalapeno pepper (Capsicum annuum var. annuum) sauce.

Ultrasonication is one of the non-thermal physical methods that can be used on foods and when used in synergy with temperature (thermosonication), this technique proves to be more effective, thus reducing the duration and intensity of heat treatment and the consequent damage to the foods. This work aimed to use the technique of ultrasonication and thermosonication in the processing of jalapeno pepper sauces in comparison with pasteurization. Two types of sauces were produced, one with pre-cooking (a) and the other without cooking (b), and the influence of time and temperature was analyzed by applying ultrasonication and thermosonication. Times of 15 and 30 min and temperatures of 25 and 65 °C were used. Both treatments stood out for their effectiveness when compared to the traditional method (pasteurization 65 °C and 30 min). The results demonstrate that, in general, the sauces are good sources of phenolic compounds (141.83 ± 0.10 mg gallic acid equivalent/100 g), flavonoids (50.40 ± 0.30 mg quercetin equivalent/100 g) and carotenoids (2.39 ± 0.07 mg ß-carotene/100 g). The sauces had an increase in carotenoids by about 25% (thermosonicated at 15 and 30 min and pre-cooked) and in antioxidant activity (ferric reducing antioxidant power) with about 12% and 13% (thermosonicated at 30 min with and without cooking, respectively) in relation to control (pasteurization). On comparing thermosonication with ultrasound process total phenolics had improved by around 14% and flavonoids by 55%. At the first time, capsantin, capsaicin, dihydrocapsaicin, and nordihydrocapsaicin were identified by ultra-high performance liquid chromatography-MS/MS (UHPLC-MS/MS). Finally, as both treatments demonstrate efficiency (thermosonication at 15 and 30 min), the use of 15 min is indicated as feasible by the reduced process time and in preventing the loss of bioactive compounds in the sauces when compared to the pasteurization treatment.

2,5-Diketopiperazines via Intramolecular N-Alkylation of Ugi Adducts: A Contribution to the Synthesis, Density Functional Theory Study, X-ray Characterization, and Potential Herbicide Application.

To investigate the herbicidal potential of 2,5-diketopiperazines (2,5-DKPs), we applied a known protocol to produce a series of 2,5-DKPs through intramolecular N-alkylation of Ugi adducts. However, the method was not successful for the cyclization of adducts presenting aromatic rings with some substituents at the ortho position. Results from DFT calculations showed that the presence of voluminous groups at the ortho position of a benzene ring results in destabilization of the transition structure. Lower activation enthalpies for the SN2-type cyclization of Ugi adducts were obtained when bromine, instead of a chlorine anion, is the leaving group, indicating that the activation enthalpy for the cyclization step controls the formation of the 2,5-DKP. Some Ugi adducts and 2,5-DKPs formed crystals with suitable qualities for single-crystal X-ray diffraction data collection. Phytotoxic damage of some 2,5-DKPs on leaves of the weed Euphorbia heterophylla did not differ from those caused by the commercial herbicide diquat.

Metabolite profiles of energy cane and sugarcane reveal different strategies during the axillary bud outgrowth.

Commercial cultivation of sugarcane is usually carried out by planting culm segments (sett) carrying buds in their internodes. However, this is an inefficient practice due to high sprouting irregularity. In this work, we inspect the first stages of the physiological preparation of the culm for sprouting, trying to identify compounds that actively participate in this process. We compared, during the first 48 h, the metabolic profile of sugarcane against energy cane, a cultivar known to have higher sprouting speed and consistency. In fact, during this short period it was possible to observe that energy cane already had a higher physiological activity than sugarcane, with significant changes in the catabolism of amino acids, increased levels of reducing sugars, lipids and metabolic activity in the phenylpropanoid pathway. On the other hand, sugarcane samples had just begun their activity during this same period, with an increase in the level of glutamate as the most significant change, which may be linked to the strategy of these cultivars to develop their roots before leaves, opposite of what is seen for energy cane. These results contribute to the development of strategies for increasing the efficiency of sprouting in sugarcane.

Chemical profile, antioxidant and anti-inflammatory properties of Miconia albicans (Sw.) Triana (Melastomataceae) fruits extract.

ETHNOPHARMACOLOGICAL RELEVANCE: Miconia albicans (Sw.) Triana has been widely used in Brazilian popular medicine for the treatment of several diseases. Aerial parts are used as an infusion to treat arthrosis and arthritis, to relieve rheumatic and stomach pains, and intestinal disorders due to its anti-inflammatory, anti-mutagenic anti-nociceptive, digestive and hepatoprotective properties. AIM OF THE STUDY: This study aimed to characterize the of M. albicans (Sw.) Triana fruits extract (MAFRE) chemical profile and to evaluate its antioxidant, anti-inflammatory and antitumor activities, as well as its toxicity. MATERIALS AND METHODS: Maceration with methanol as liquid extractor was used to prepare MAFRE. M. albicans (Sw.) Triana fruits chemical composition was characterized by UHPLC-QTOF-MS/MS and GC-FID (fatty acid methyl esters composition from lyophilized fruits). MAFRE antioxidant potential was evaluated in vitro using a combination of assays: Folin-Ciocalteu reducing capacity, DPPH• and ABTS radical scavenging ability and ferric reducing antioxidant power (FRAP). In vitro antiproliferative activity was investigated in four human tumor cell lines (U251, 786-0, HT29 and MDA-MB-231) while the effect on the non-tumor cell viability was assessed in the VERO cell line using the on-step MTT assay. In addition, in vivo anti-inflammatory effect was assessed by Croton oil-induced ear edema in mice followed by myeloperoxidase (MPO) activity evaluation. RESULTS: Thirty-five compounds were identified by UHPLC-QTOF-MS/MS. Among it flavonoids derived from quercetin (8), myricetin (1), kaempferol (2), terpenoids (6) and other compounds (18). GC-FID analysis identified and quantified nine fatty acids: palmitic, stearic, arachidic, behenic, elaidic, oleic, eicosenoic, and linoleic acids. The most abundant fatty acids were polyunsaturated fatty acids (5.33 ± 0.17 mg g-1), followed by saturated fatty acids (2.38 ± 0.07 mg g-1) and monounsaturated fatty acids (1.74 ± 0.09 mg g-1). The extract revealed high content of phenolic compounds (43.68 ± 0.50 mg GAE/g of extract), potent antioxidant, and ferrous chelating capacities. Morever, it proved to be non-toxic to the VERO cells, not affecting cells viability (95% of viable cells). No antiproliferative effect against human tumor cell lines were found. Furthermore, MAFRE significantly (p<0.05) reduced ear edema (≈35%) and MPO activity (84.5%) having a statistical effect similar to traditional steroidal and non-steroidal anti-inflammatory drugs. CONCLUSIONS: Taken together, the results evidenced that M. albicans fruit extract has antioxidant properties, a higher concentration of phenolic compounds, flavonoids, fatty acids, and also topical anti-inflammatory activity with low toxicity of extract on VERO cells. Through the ethnomedicinal study, these findings supporting the popular use of M. albicans, but also highlight that not only aerial parts and leaves deserve attention, but the fruits also have anti-inflammatory proprieties and can be a source of phenolic compounds and other substances with potential health benefices.

Phytotoxic property of metabolites isolated from Garcinia gardneriana.

Garcinia gardneriana is a medicinal tree species used in Brazil in the treatment of hepatitis and gastritis. This use is attributed to phenolic compounds, mainly 7-epiclusianone, guttiferone-A and fukugetin, which present several proven biological activities. However, to the best of our knowledge, no study on the phytotoxic activity of G. gardneriana extracts has been conducted until now. This research proposed to isolate and quantify by high-performance liquid chromatography (HPLC) the major compounds from G. gardneriana seed extracts in ethyl acetate and to evaluate their phytotoxic activities. The natural products 7-epiclusianone, guttiferone-A and fukugetin were quantified at concentrations varying from 0.46 to 1.13 mg mL-1 and were isolated with yields ranging from 7% to 23% (w/w). The phytotoxic assay indicated that the crude extract showed no action on the dry matter of Sorghum bicolor plants, but the isolated compounds fukugetin and 7-epiclusianone had moderate activity. On the other hand, guttiferone-A displayed a greater herbicide activity than glyphosate, a positive control, even in almost three times lower concentrations, causing severe intoxication in the plants. This work is the first report on this activity by the extract of G. gardneriana and its isolated compounds. Besides that, guttiferone-A showed up as a scaffold for the development of new agrochemicals. Complementing these findings, computational studies suggested that this benzophenone can interact effectively with transferase enzymes type, in special caffeic acid O-methyltransferase from S. bicolor (PDB code: 4PGH), indicating a possible mechanism of action in this plant.

Spray drying encapsulation of stevia extract with maltodextrin and evaluation of the physicochemical and functional properties of produced powders.

This work aimed to formulate and perform physicochemical and functional characterization of maltodextrin microcapsules containing ethanolic extract of stevia, rich in antioxidant compounds, encapsulated by a spray-drying process with two maltodextrins (DE10 and DE19). The powders were named M10 and M19, respectively. We analyzed the physicochemical parameters, antidiabetic activity, cytotoxicity, bioaccessibility of the compounds by in vitro digestion, as well as the structure of the microcapsules by scanning electron microscopy. Microcapsules showed higher solubility (∼35%), lower moisture content (∼29%), and the maltodextrin DE10 had higher efficiency as an encapsulating agent (87%) when compared to DE19 (76%) and showed well-defined spherical structures. The microencapsulation preserved the content of phenolic compounds and antioxidant activity present in the extract (7.2% and 87.5%, respectively). The bioaccessibility of these microencapsulated compounds and antioxidant activity were higher under different conditions of in vitro digestion (mouth, gastric, and intestinal conditions) and showed no cytotoxic effects. We identified 41 compounds (by UHPLC-MS/MS-Qtof) related to the nutritional benefits offered by stevia and the microencapsulation technique can be recommended to preserve bioactive compounds. PRACTICAL APPLICATION: Ethanol extract from stevia leaves contains antioxidant phytochemicals related to the nutritional benefits of stevia. However, this extract presents low solubility and consequently low bioaccessibility under in vitro digestion. The microencapsulation process protects the bioactive compounds of the different pH from digestion and improves the physical-chemical parameters of the extract, increasing its applicability as a possible food additive.

Tyrosol 1,2,3-triazole analogues as new acetylcholinesterase (AChE) inhibitors.

The present work proposed the preparation of triazolic analogues of tyrosol, a biophenol found in olive oil and whose wide range of bioactivities has been the target of many studies. We obtained fifteen novel tyrosol derivatives and the compounds of the series were later evaluated as acetylcholinesterase (AChE) inhibitors. The study of AChE inhibition is important for the development of new drugs and pesticides, and especially the research for managing Alzheimer's disease. The most active compound, namely 7-({1-[2-(4-hydroxyphenyl)ethyl]-1H-1,2,3-triazol-4-yl}methoxy)-4-methyl-2H-chromen-2-one (30), showed IC50 value of 14.66 ±â€¯2.29 µmol L-1. Docking experiments corroborated by kinetic assay are suggestive of a competitive inhibition mechanism. Derivatives interacted with amino acids from the AChE active site associated to the development of Alzheimer's disease. The results indicate that the compounds synthesized have a high potential as prototypes for the development of new acetylcholinesterase inhibitors.

Addressing Global Ruminant Agricultural Challenges Through Understanding the Rumen Microbiome: Past, Present, and Future.

The rumen is a complex ecosystem composed of anaerobic bacteria, protozoa, fungi, methanogenic archaea and phages. These microbes interact closely to breakdown plant material that cannot be digested by humans, whilst providing metabolic energy to the host and, in the case of archaea, producing methane. Consequently, ruminants produce meat and milk, which are rich in high-quality protein, vitamins and minerals, and therefore contribute to food security. As the world population is predicted to reach approximately 9.7 billion by 2050, an increase in ruminant production to satisfy global protein demand is necessary, despite limited land availability, and whilst ensuring environmental impact is minimized. Although challenging, these goals can be met, but depend on our understanding of the rumen microbiome. Attempts to manipulate the rumen microbiome to benefit global agricultural challenges have been ongoing for decades with limited success, mostly due to the lack of a detailed understanding of this microbiome and our limited ability to culture most of these microbes outside the rumen. The potential to manipulate the rumen microbiome and meet global livestock challenges through animal breeding and introduction of dietary interventions during early life have recently emerged as promising new technologies. Our inability to phenotype ruminants in a high-throughput manner has also hampered progress, although the recent increase in "omic" data may allow further development of mathematical models and rumen microbial gene biomarkers as proxies. Advances in computational tools, high-throughput sequencing technologies and cultivation-independent "omics" approaches continue to revolutionize our understanding of the rumen microbiome. This will ultimately provide the knowledge framework needed to solve current and future ruminant livestock challenges.

Identification and ultra-high-performance liquid chromatography coupled with high-resolution mass spectrometry characterization of biosurfactants, including a new surfactin, isolated from oil-contaminated environments.

Biosurfactant-producing bacteria were isolated from samples collected in areas contaminated with crude oil. The isolates were screened for biosurfactant production using qualitative drop-collapse test, oil-spreading and emulsification assays, and measurement of their tensoactive properties. Five isolates tested positive for in the screening experiments and displayed decrease in the surface tension below 30 mN m-1 . The biosurfactants produced by these isolates were further investigated and their molecular identification revealed that they are bacteria related to the Bacillus genus. Additionally, the biosurfactants produced were chemically characterized via UHPLC-HRMS experiments, indicating the production of surfactin homologues, including a new class of these molecules.

Development of an analytical method for identification of Aspergillus flavus based on chemical markers using HPLC-MS.

Aspergillus flavus is a filamentous fungus found in nature and characterized by the production of bright and colourful colonies. It grows on different substrates, producing secondary metabolites and, if present in foodstuffs, can be a source of health problems for humans and animals, as well as causing economic losses. Traditional methods for fungal identification are based on morphological characteristics, requiring specialists and being very time-consuming. The development of analytical alternatives might have advantages such as greater efficiency, more reproducibility and be less time-consuming. Thus, a qualitative analytical method to detect Aspergillus flavus in food samples, based on the identification of fungal chemical markers by HPLC-MS, was developed. The method comprises methanol extraction followed by HPLC-MS analysis, and was able to identify 14 fungus secondary metabolites, namely aflatoxin B1, aflatoxin G1, aspergillic acid, aspyrone, betaine, chrysogine, deacetyl parasiticolide A, flufuran, gregatin B, hydroxysydonic acid, nicotinic acid, phomaligin A, spinulosin and terrein.

Phytochemical analysis of Pfaffia glomerata inflorescences by LC-ESI-MS/MS.

Pfaffia glomerata contains high levels of ß-ecdysone, which has shown a range of beneficial pharmacological effects. The present study demonstrated that inflorescences of P. glomerata contain other important bioactive compounds in addition to ß-ecdysone. The identification of compounds from inflorescences using liquid chromatography coupled with electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS) was performed for the first time. The eight compounds identified were ß-ecdysone, flavonoid glycosides such as quercetin-3-O-glucoside, kaempferol-3-O-glucoside and kaempferol-3-O-(6-p-coumaroyl)-glucoside, oleanane-type triterpenoid saponins such as ginsenoside Ro and chikusetsusaponin IV, in addition to oleanonic acid and gluconic acid. This study provided information on the phytochemicals contained in P. glomerata inflorescences revealing the potential application of this plant part as raw material for the phytotherapeutic and cosmetic industries.

Effects of cadmium and copper biosorption on Chlorella vulgaris.

Changes in protein levels and lipid compositions in algal cells indicate the severity of stress related to toxic concentrations of heavy metals. In this study, the effects of exposure to cadmium and copper on Chlorella vulgaris and its capacity to remove metals were evaluated. The data revealed ion removal activity by microalgae under all treatments and different levels of protein expression after 48 h of exposure. Furthermore, we analyzed lipids contents to characterize them.

A new label-free approach for the determination of reaction rates in oxidative footprinting experiments.

Hydroxyl radical-mediated oxidative footprinting coupled to mass spectrometric analysis is an attractive technique for protein surface mapping, conformational changes monitoring, and protein-ligand interfaces mapping in solution. In this technique, a protein is oxidized by in situ-generated hydroxy radicals and the site and rate of oxidation can be determined by proteolysis followed by mass spectrometric analysis. Changes in peptide oxidation rate can then be correlated to the changes in solvent exposure, and information about conformational changes or interaction domains can be obtained. The method relies, therefore, on the accurate measurements of peptide oxidation rate. Here, we describe a new label-free method to determine the oxidation rate of peptides that is based on the consumption of the unoxidized peptide instead of measuring the formation of oxidized peptides. The reaction rate thus obtained presents a better linearity and lower variation when compared to the traditional method. The label-free method is also simpler to implement and automation can be achieved through label-free quantitation software.

Traveling-wave ion mobility mass spectrometry analysis of isomeric modified peptides arising from chemical cross-linking.

Traveling-wave ion mobility (TWIM) coupled to mass spectrometry (MS) has emerged as a powerful tool for structural and conformational analysis of proteins and peptides, allowing the analysis of isomeric peptides (or proteins) with the same sequence but modified at different residues. This work demonstrates the use of the novel TWIM-MS technique to separate isomeric peptide ions derived from chemical cross-linking experiments, which enables the acquisition of distinct product ion spectra for each isomer, clearly indicating modification on different sites. Experiments were performed with four synthetic peptides, for which variable degrees of mobility separation were achieved. In cases of partially overlapping mobility arrival time distributions (ATDs), extracting the ATDs of fragment ions belonging to each individual isomer allowed their separation into two distinct ATDs. Accumulation over regions from the specific ATDs generates the product ion spectrum of each isomer, or a spectrum highly enriched in their fragments. The population of both modified peptide isomers was correlated with the intrinsic reactivities of different Lys residues from reactions conducted at different pH conditions.

HPLC based method using sample precolumn cleanup for the determination of triazines and thiolcarbamates in hemodialysis saline solutions.

Solid-phase extraction (SPE) procedures for cleanup and preconcentration followed by HPLC-UV method were investigated for the simultaneous determination of seven low-dosed pesticides in saline concentrates for hemodialysis. The target compounds were ametryn, desmetryn, prometryn, terbutryn, molinate, triallate and butylate. Polyethylene (three different types), teflon, polyurethane and polystyrene, in powder form, were investigated as adsorbents for solid-phase extraction of the analytes from the saline samples. Quantification was performed at 222nm and the analytes were separated on a LiChrosorb RP-18 (5mum, 125mm x 4mm i.d.) column using gradient elution with water/acetonitrile as mobile phase. The duration each chromatographic run was 18min including column reconditioning. The efficiency of the different SPE substrates for retaining the analytes from the highly concentrated saline (HCS) samples was discussed. The best performance was achieved with polystyrene as SPE material considering preconcentration factor, precolumn clogging, reusing capability and similarity between the mobile phases for SPE and HPLC procedures. Analyte concentrations as low as 1mugL(-1) could be determined in spiked HCS samples after preconcentration on polystyrene SPE precolumns. Recoveries between 98.7 and 102.2% were obtained from commercial spiked samples. Detection limits ranging from 4.8 (for prometryn) to 46mugL(-1) (for butylate) were calculated (without preconcentration). The within-day relative standard deviations (n = 9) ranged from 2.3 to 4.8%.