Composição de fenólicos, flavonoides, antocianinas, determinação da atividade antioxidante e comparação de métodos extrativos da folha e da casca de Caryocar brasiliense

Introduction: Caryocar brasiliense is a medicinal plant used in the treatment of respiratory diseases, gastric ulcers and muscle pain. Objective: to compare the extraction of natural compounds using two extractive methods, magnetic stirring and ultrasonic bath. In addition, to evaluate the antioxidant activity, the content of phenolic compounds, flavonoids and anthocyanins in the leaves and bark of Pequi (C. brasiliense), with a view to adding value through its functional properties. Methodology: the phenolic content was determined by the Folin-Ciocalteu method, flavonoids and anthocyanins by the Lima and Melo method. Antioxidant activity was measured using the 2.2-diphenyl-1-picrilhhydrazyl (DPPH) method. Results: the extract of the Pequi leaves exhibited a higher phenolic content in relation to the bark for both extraction methods. The Pequi leaf in an ultrasonic bath showed strong antioxidant activity with a value of 72.2%. Conclusion: Pequi extracts demonstrated a promising phytochemical profile that should be investigated in the future for pharmacological application as an adjuvant or precursor in the synthesis of a new cosmetic or medicine with antioxidant function.Introdução: Caryocar brasiliense é conhecida popularmente como Pequi ou Pequizeiro, sendo uma planta com propriedades medicinais para tratamento de doenças respiratórias, úlceras gástricas e dores musculares. Objetivo: comparar a extração de biocompostos por meio de dois métodos extrativos, agitação magnética e banho ultrassônico. Além disso, avaliar a atividade antioxidante, o teor de compostos fenólicos, flavonoides e antocianinas em extratos das folhas e da casca de Pequi (C. brasiliense), com vistas a agregar valor quanto às suas propriedades funcionais. Metodologia: o conteúdo de compostos fenólicos foi determinado pelo método de Folin-Ciocalteu, flavonoides e antocianinas pelo método de Lima e Melo. A atividade antioxidante foi medida pelo método 2,2-difenil-1-picrilhidrazil (DPPH). Resultados: o extrato das folhas de Pequi exibiu maior teor de fenólicos em relação à casca, independente do método de extração. O extrato das folhas obtido em banho ultrassônico apresentou forte atividade antioxidante com valor de 72,2%. Conclusão: os extratos de Pequi demonstraram um perfil fitoquímico promissor que deve ser investigado no futuro para aplicação farmacológica como adjuvante ou precursor na síntese de novos cosméticos ou medicamentos com propriedades antioxidante

Identification of metabolic pathways influenced by the G-protein coupled receptors GprB and GprD in Aspergillus nidulans

Heterotrimeric G-protein-mediated signaling pathways play a pivotal role in transmembrane signaling in eukaryotes. Our main aim was to identify signaling pathways regulated by A. nidulans GprB and GprD G-protein coupled receptors (GPCRs). When these two null mutant strains were compared to the wild-type strain, the DeltagprB mutant showed an increased protein kinase A (PKA) activity while growing in glucose 1% and during starvation. In contrast, the DeltagprD has a much lower PKA activity upon starvation. Transcriptomics and (1)H NMR-based metabolomics were performed on two single null mutants grown on glucose. We noted modulation in the expression of 11 secondary metabolism gene clusters when the DeltagprB and DeltagprD mutant strains were grown in 1% glucose. Several members of the sterigmatocystin-aflatoxin gene cluster presented down-regulation in both mutant strains. The genes of the NR-PKS monodictyphenone biosynthesis cluster had overall increased mRNA accumulation in DeltagprB, while in the DeltagprD mutant strain the genes had decreased mRNA accumulation. Principal component analysis of the metabolomic data demonstrated that there was a significant metabolite shift in the DeltagprD strain. The (1)H NMR analysis revealed significant expression of essential amino acids with elevated levels in the DeltagprD strain, compared to the wild-type and DeltagprB strains. With the results, we demonstrated the differential expression of a variety of genes related mainly to secondary metabolism, sexual development, stress signaling, and amino acid metabolism. We propose that the absence of GPCRs triggered stress responses at the genetic level. The data suggested an intimate relationship among different G-protein coupled receptors, fine-tune regulation of secondary and amino acid metabolisms, and fungal development

In vitro schistosomicidal activity of curcumin against Schistosoma mansoni adult worms

The in vitro schistosomicidal activity of curcumin (doses ranging from 5 to 100 mu M) was carried out against Schistosoma mansoni adult worms. Curcumin (at 50 and 100 mu M) caused death of all worms. When tested at the doses of 5 and 20 mu M, it decreased the worm viability in comparison with negative (Roswell Memorial Park Institute (RPMI) 1640 medium alone or RPMI 1640 medium with 10% dimethyl sulfoxide) and positive (heat-killed worms at 56A degrees C or praziquantel 10 mu M) control groups. All pairs of coupled adult worms were separated into individual male and female by the action of curcumin at the doses of 20 to 100 mu M. When tested at 5 and 10 mu M, curcumin reduced egg production by 50% in comparison with the positive control group. It is the first time that the schistosomicidal activity has been reported for curcumin.CAPESFAPES

Uncovering Notch pathway in the parasitic flatworm Schistosoma mansoni.

Several signaling molecules that govern development in higher animals have been identified in the parasite Schistosoma mansoni, including the transforming growth factor ?, protein tyrosine kinases, nuclear hormone receptors, among others. The Notch pathway is a highly conserved signaling mechanism which is involved in a wide variety of developmental processes including embryogenesis and oogenesis in worms and flies. Here we aimed to provide the molecular reconstitution of the Notch pathway in S. mansoni using the available transcriptome and genome databases. Our results also revealed the presence of the transcripts coded for SmNotch, SmSu(H), SmHes, and the gamma-secretase complex (SmNicastrin, SmAph-1, and SmPen-2), throughout all the life stages analyzed. Besides, it was observed that the viability and separation of adult worm pairs were not affected by treatment with N-[N(3,5)-difluorophenacetyl)-L-Alanyl]- S-phenylglycine t-butyl ester (DAPT), a Notch pathway inhibitor. Moreover, DAPT treatment decreased the production of phenotypically normal eggs and arrested their development in culture. Our results also showed a significant decrease in SmHes transcript levels in both adult worms and eggs treated withDAPT. These results provide, for the first time, functional validation of the Notch pathway in S. mansoni and suggest its involvement in parasite oogenesis and embryogenesis. Given the complexity of the Notch pathway, further experiments shall highlight the full repertoire of Notch-mediated cellular processes throughout the S. mansoni life cycle

Curcumin Generates Oxidative Stress and Induces Apoptosis in Adult Schistosoma mansoni Worms.

Inducing apoptosis is an interesting therapeutic approach to develop drugs that act against helminthic parasites. Researchers have investigated how curcumin (CUR), a biologically active compound extracted from rhizomes of Curcuma longa, affects Schistosoma mansoni and several cancer cell lines. This study evaluates how CUR influences the induction of apoptosis and oxidative stress in couples of adult S. mansoni worms. CUR decreased the viability of adult worms and killed them. The tegument of the parasite suffered morphological changes, the mitochondria underwent alterations, and chromatin condensed. Different apoptotic parameters were determined in an attempt to understand how CUR affected adult S. mansoni worms. CUR induced DNA damage and fragmentation and increased the expression of SmCASP3/7 transcripts and the activity of Caspase 3 in female and male worms. However, CUR did not intensify the activity of Caspase 8 in female or male worms. Evaluation of the superoxide anion and different antioxidant enzymes helped to explore the mechanism of parasite death further. The level of superoxide anion and the activity of Superoxide Dismutase (SOD) increased, whereas the activity of Glutathione-S-Transferase (GST), Glutathione reductase (GR), and Glutathione peroxidase (GPX) decreased, which culminated in the oxidation of proteins in adult female and male worms incubated with CUR. In conclusion, CUR generated oxidative stress followed by apoptotic-like-events in both adult female and male S. mansoni worms, ultimately killing them

Genome-wide identification, characterisation and expression profiling of the ubiquitin-proteasome genes in Biomphalaria glabrata

Submitted by Nuzia Santos ([email protected]) on 2019-08-27T16:39:37Z No. of bitstreams: 1 Genome-wide identification.pdf: 12612955 bytes, checksum: fd4068e8355f870120d20ad7ff2c2fc6 (MD5)Approved for entry into archive by Nuzia Santos ([email protected]) on 2019-08-27T16:43:34Z (GMT) No. of bitstreams: 1 Genome-wide identification.pdf: 12612955 bytes, checksum: fd4068e8355f870120d20ad7ff2c2fc6 (MD5)Made available in DSpace on 2019-08-27T16:43:34Z (GMT). No. of bitstreams: 1 Genome-wide identification.pdf: 12612955 bytes, checksum: fd4068e8355f870120d20ad7ff2c2fc6 (MD5) Previous issue date: 2019Universidade Federal de Uberlândia. Laboratório de Bioinformática e Análises Moleculares. Patos de Minas, MG, Brasil.Universidade Federal de Uberlândia. Laboratório de Bioinformática e Análises Moleculares. Patos de Minas, MG, BrasilFundação Oswaldo Cruz. Instituto René Rachou. Grupo de Pesquisa em Biologia do Schistosoma mansoni e sua Interação com o Hospedeiro. Belo Horizonte, MG, Brasil.Universidade Federal de Lavras. Departamento de Biologia. Seção de Fisiologia de Plantas. Laboratório de Fisiologia Molecular de Plantas. Lavras, MG, Brasil.Universidade Federal de Ouro Preto. Escola de Farmácia. Departamento de Farmácia. Ouro Preto, MG, Brasil.Fundação Oswaldo Cruz. Instituto René Rachou. Grupo de Pesquisa em Biologia do Schistosoma mansoni e sua Interação com o Hospedeiro. Belo Horizonte, MG, Brasil.Fundação Oswaldo Cruz. Instituto René Rachou. Grupo de Pesquisa em Biologia do Schistosoma mansoni e sua Interação com o Hospedeiro. Belo Horizonte, MG, Brasil.Universidade Federal de Uberlândia. Laboratório de Bioquímica e Biologia Molecular. Patos de Minas, MG, Brasil.Universidade Estadual de Campinas. Instituto de Biologia. Departamento de Biologia Animal. Campinas, SP, Brasil.Fundação Oswaldo Cruz. Instituto René Rachou. Grupo de Pesquisa em Helmintologia e Malacologia Médica. Belo Horizonte, MG, Brasil.Universidade Federal de Uberlândia. Laboratório de Bioinformática e Análises Moleculares. Patos de Minas, MG, Brasil.BACKGROUND: Biomphalaria glabrata is the major species used for the study of schistosomiasis-related parasite-host relationships, and understanding its gene regulation may aid in this endeavor. The ubiquitin-proteasome system (UPS) performs post-translational regulation in order to maintain cellular protein homeostasis and is related to several mechanisms, including immune responses. OBJECTIVE: The aims of this work were to identify and characterise the putative genes and proteins involved in UPS using bioinformatic tools and also their expression on different tissues of B. glabrata. METHODS: The putative genes and proteins of UPS in B. glabrata were predicted using BLASTp and as queries reference proteins from model organism. We characterised these putative proteins using PFAM and CDD software describing the conserved domains and active sites. The phylogenetic analysis was performed using ClustalX2 and MEGA5.2. Expression evaluation was performed from 12 snail tissues using RPKM. FINDINGS: 119 sequences involved in the UPS in B. glabrata were identified, which 86 have been related to the ubiquitination pathway and 33 to proteasome. In addition, the conserved domains found were associated with the ubiquitin family, UQ_con, HECT, U-box and proteasome. The main active sites were lysine and cysteine residues. Lysines are responsible and the starting point for the formation of polyubiquitin chains, while the cysteine residues of the enzymes are responsible for binding to ubiquitin. The phylogenetic analysis showed an organised distribution between the organisms and the clades of the sequences, corresponding to the tree of life of the animals, for all groups of sequences analysed. The ubiquitin sequence was the only one with a high expression profile found in all libraries, inferring its wide range of performance. MAIN CONCLUSIONS: Our results show the presence, conservation and expression profile of the UPS in this mollusk, providing a basis and new knowledge for other studies involving this system. Due to the importance of the UPS and B. glabrata, this work may influence the search for new methodologies for the control of schistosomiasis

ChIP-seq reveals a role for CrzA in the Aspergillus fumigatus high-osmolarity glycerol response (HOG) signalling pathway

Aspergillus fumigatus is an opportunistic pathogen and allergen of mammals. Calcium signalling is essential for A. fumigatus pathogenicity and is regulated by the CrzA transcription factor. We used ChIP-seq (Chromatin Immunoprecipitation DNA sequencing) to explore CrzA gene targets in A. fumigatus. In total, 165 potential binding peaks including 102 directly regulated genes were identified, resulting in the prediction of the A[GT][CG]CA[AC][AG] CrzA-binding motif. The 102 CrzA putatively regulated genes exhibited a diverse array of functions. The phkB (Afu3g12530) histidine kinase and the sskB (Afu1g10940) MAP kinase kinase kinase of the HOG (high-osmolarity glycerol response) pathway were regulated by CrzA. Several members of the two-component system (TCS) and the HOG pathway were more sensitive to calcium. CrzAConselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP

Evaluation of the Antioxidant, Antimicrobial, and Anti-Biofilm Effects of the Stem Bark, Leaf, and Seed Extracts from <i>Hymenaea courbaril</i> and Characterization by UPLC-ESI-QTOF-MS/MS Analysis

Currently, biofilm-forming bacteria are difficult to treat by conventional antibiotic therapy and are, thus, becoming a clinical and epidemiological problem worldwide. Medicinal plants have been identified as novel alternative treatments due to their therapeutic and antimicrobial effects. In this context, the present study aimed to determine the total phenolic content, antioxidant capacity, and antimicrobial and anti-biofilm potential of nine extracts of Hymenaea courbaril (Fabaceae), popularly known as Jatobá. Furthermore, extracts that exhibited biofilm inhibitory activity against S. aureus (ATCC 25923) were selected for UPLC-HRMS/MS chemical analysis. Our results showed a high total phenolic content, mainly in the stem bark extract, and that the plant is rich in compounds with antioxidant activity. In the anti-biofilm analysis, leaf extracts stood out in comparison with chloramphenicol, with inhibition percentages of 78.29% and 78.85%, respectively. Through chemical analysis by UPLC-HRMS/MS, chrysoeriol-7-O-neohesperidoside, isorhamnetin-3-O-glucoside, and 3,7-di-O-methylquercetin were annotated for the first time in the leaves of H. courbaril. Therefore, these results showed the potential use of H. courbaril as an antioxidant and point to its use in antimicrobial therapy with an anti-biofilm effect

Characterization and mRNA expression analysis of PI31, an endogenous proteasome inhibitor from Schistosoma mansoni.

The proline-rich inhibitor of 31 kDa (PI31) is highly conserved through metazoan evolution, and its activity in the proteasome inhibition is well-established although the precise mechanism of inhibition is unclear. The coding DNA sequence of Schistosoma mansoni PI31 (SmPI31) was cloned, and the recombinant protein was expressed in bacterial system. The correct amino acid sequence was confirmed by mass spectrometry and circular dichroism suggests that SmPI31 contains both ?-helix and non-structured regions. Inhibition assays, using the SucLeu-Leu-Val-Tyr-4-MCA substrate for proteasome degradation, showed that the S. mansoni proteasome may be regulated by the inhibitory activity of SmPI31. A gene expression assay using qRT-PCR at various stages during the S. mansoni life cycle has shown that SmPI31 transcripts are expressed in all studied stages, suggesting that PI31 plays an important role during the developmental processes of the parasite. In this study first evidence is presented that PI31 has a conserved structure and plays a role as proteasome inhibitor in adult worms and it is expressed through life cycle