Chemical Profile of Ocotea delicata (Lauraceae) Using Ultra High-Performance Liquid Chromatography-High-Resolution Mass Spectrometry-Global Natural Products Social Molecular Networking Workflow.

Ocotea, the largest genus in the Lauraceae family, encompasses numerous species of scientific interest. However, most Ocotea species have only been described morphologically. This study used an untargeted metabolomics workflow with UHPLC-HRMS and GNPS-FBMN to provide the first chemical evaluation of the polar specialized metabolites of O. delicata leaves. Leaves from three O. delicata specimens were extracted using ultrasound-assisted extraction with 70% ethanol. Among the examined samples, 44 metabolites, including alkaloids and flavonoids, were identified. In contrast to other Ocotea species, O. delicata has a wider diversity of kaempferol derivatives than quercetin. The biomass of the specimens showed a significant correlation with the chemical profile. The similarity among specimens was mostly determined by the concentrations of quinic acid, kaempferol glycosides, and boldine. The evaluated specimens exhibited chemical features similar to those of species classified as New World Ocotea, with the coexistence of aporphine and benzylisoquinoline alkaloids.

Chemophenetic study of Ocotea canaliculata (Lauraceae) by UHPLC-HRMS and GNPS.

The metabolic fingerprint of a non-volatile fraction of Ocotea canaliculata (Rich.) Mez (Lauraceae) leaves was determined by UHPLC-HRMS analysis. Twenty-four compounds were suggestively identified by GNPS-FBMN. The results revealed a large production of flavonoids, mainly flavones and flavanones, a chemical class poorly described in the Ocotea genus. Within the identified compounds, four are being described for the first time in this genus. The major metabolite detected was astilbin, with a concentration corresponding to 23.2 ± 1.58% of the extracts. The expressive content of astilbin also highlights it as a chemical marker for the species. As a species that is classified as a complex, qualitative and semi-quantitative features obtained through the O. canaliculata flavonoid fingerprint can be further used for a more precise circumscription and species-specific characterization.

Peptidogalactomannan from Histoplasma capsulatum yeast cell wall: role of the chemical structure in recognition and activation by peritoneal macrophages.

Histoplasma capsulatum is the causative agent of histoplasmosis, a systemic disease responsible for most reported causes of morbidity and mortality among immunosuppressed individuals. Peptidogalactomannan (pGM) was purified from the yeast cell wall of H. capsulatum isolated from bats, and its structure and involvement in modulating the host immune response were evaluated. Gas chromatography, methylation analysis, and two-dimensional nuclear magnetic resonance (2D-NMR) were used for the structural characterization of pGM. Methylation and 2D-NMR data revealed that pGM comprises a main chain containing α-D-Manp (1 → 6) residues substituted at O-2 by α-D-Manp (1 → 2)-linked side chains, non-reducing end units of α-D-Galf, or ß-D-Galp linked (1→ 6) to α-D-Manp side chains. The involvement of H. capsulatum pGM in antigenic reactivity and in interactions with macrophages was demonstrated by ELISA and phagocytosis assay, respectively. The importance of the carbohydrate and protein moieties of pGM in sera reactivity was evaluated. Periodate oxidation abolished much pGM antigenic reactivity, suggesting that the sugar moiety is the most immunogenic part of pGM. Reactivity slightly decreased in pGM treated with proteinase K, suggesting that the peptide moiety plays a minor role in pGM antigenicity. In vitro experiments suggested that pGM is involved in the phagocytosis of H. capsulatum yeast and induction of IL-10 and IFN-γ secretion by peritoneal macrophages from C57BL/6 mice. These findings demonstrated the role of pGM in the H. capsulatum-host interaction.

Chemosystematic evaluation of Amazonian freshwater sponges.

Freshwater sponges can be considered a promising new source of bioactive compounds for the pharmaceutical industry; however, the research on their chemical composition is still in the incipient stage. We evaluated the most endemic Amazonian freshwater sponge species from the Drulia and Metania genera by untargeted metabolomic approaches, based on UHPCL-HRMS, in order to identify chemical markers and explore the diversity of specialized metabolites. The use of untargeted approaches allowed us to observe subsets of metabolites that enabled the characterization of, not only each genus, but also, of each species. Freshwater sponge species presented themselves as rich sources of fatty acids and sterols, which were putatively identified. These metabolites were suggested as chemical markers for further targeted metabolomic studies.

Phytochemistry by design: a case study of the chemical composition of Ocotea guianensis optimized extracts focused on untargeted metabolomics analysis.

Untargeted metabolomics aim to provide a global chemical fingerprint of biological matrices. This research field can be used in phytochemical screenings for bioactive species or in the identification of species. Despite its importance in providing a global chemical profile, little research has focused on the optimization of the extraction methods, as each type of matrix requires a specific procedure. Therefore, we propose to evaluate the effect of different extraction features in an ultrasound-assisted extraction for the untargeted metabolomic study of an Ocotea species, a genus of great economical interest but little chemical exploitation. Method optimization was performed in a full factorial 2232 design, evaluating the solvent composition, extraction temperature, sample particle size and sample : solvent ratio effects in the metabolomic response. The effect of these parameters on the quality of the untargeted metabolomic profiles was studied by analysis of the extraction yield as well as the chromatographic and spectrometric profiles. Most substances identified were glycosylated flavonoids and aporphinic alkaloids. The application of 70% ethanol enhanced the extraction of several specialized metabolites. Statistical analysis of extraction yield and chemical profiles indicates that high temperatures and low proportion between sample and extracting solvent reduce the quality and modify the chemical profile, both qualitatively and quantitatively. The use of 70% ethanol as the extracting solvent, 1 : 12 sample : solvent ratio, 40 °C as the extraction temperature and particle size of 0.595 mm were the optimized conditions to produce a comprehensive chemical profile for Ocotea guianensis.

Zebrafish (Danio rerio) water tank model for the investigation of drug metabolism: Progress, outlook, and challenges.

Zebrafish (Danio rerio) water tank (ZWT) approach was investigated as an alternative model for metabolism studies based on six different experiments with four model compounds. Sibutramine was applied for the multivariate optimization of ZWT conditions, also for the comparison of the metabolism among ZWT, humans and mice, beyond for the role of CYP2B6 in ZWT. After the optimization, 18 fish and 168 hours of experiments is the minimum requirement for a relevant panel of biotransformation products. A comparison among the species resulted in the observation of the same hydroxylated metabolites, with differences in metabolites concentration ratio. However, the ZWT allowed tuning of the conditions to obtain a specific metabolic profile, depending on the need. In addition, by utilizing CYP2B6 inhibition, a relevant ZWT pathway for the demethylation of drugs was determined. The stereospecificity of the ZWT metabolism was investigated using selegiline and no racemization or inversion transformations were observed. Moreover, the investigation of metabolism of cannabimimetics was performed using JWH-073 and the metabolites observed are the same described for humans, except for the hydroxylation at the indol group, which was explained by the absence of CYP2C9 orthologs in zebrafish. Finally, hexarelin was used as a model to evaluate studies by ZWT for drugs with low stability. As a result, hexarelin displays a very fast metabolization in ZWT conditions and all the metabolites described for human were observed in ZWT. Therefore, the appropriate conditions, merits, and relevant limitations to conduct ZWT experiments for the investigation of drug metabolism are described.

Doping control analysis at the Rio 2016 Olympic and Paralympic Games.

This paper summarises the results obtained from the doping control analyses performed during the Summer XXXI Olympic Games (August 3-21, 2016) and the XV Paralympic Games (September 7-18, 2016). The analyses of all doping control samples were performed at the Brazilian Doping Control Laboratory (LBCD), a World Anti-Doping Agency (WADA)-accredited laboratory located in Rio de Janeiro, Brazil. A new facility at Rio de Janeiro Federal University (UFRJ) was built and fully operated by over 700 professionals, including Brazilian and international scientists, administrative staff, and volunteers. For the Olympic Games, 4913 samples were analysed. In 29 specimens, the presence of a prohibited substance was confirmed, resulting in adverse analytical findings (AAFs). For the Paralympic Games, 1687 samples were analysed, 12 of which were reported as AAFs. For both events, 82.8% of the samples were urine, and 17.2% were blood samples. In total, more than 31 000 analytical procedures were conducted. New WADA technical documents were fully implemented; consequently, state-of-the-art analytical toxicology instrumentation and strategies were applied during the Games, including different types of mass spectrometry (MS) analysers, peptide, and protein detection strategies, endogenous steroid profile measurements, and blood analysis. This enormous investment yielded one of the largest Olympic legacies in Brazil and South America. Copyright © 2017 John Wiley & Sons, Ltd.