Chiroptical properties of bioactive molecules: sensitivity to conformation and solvation

Chiroptical spectroscopies play a fundamental role in pharmaceutical analysis for the stereochemical characterisation of bioactive molecules, due to the close relationship between chirality and optical activity and the increasing evidence of stereoselectivity in the pharmacological and toxicological profiles of chiral drugs. The correlation between chiroptical properties and absolute stereochemistry, however, requires the development of accurate and reliable theoretical models. The present thesis will report the application of theoretical chiroptical spectroscopies in the field of drug analysis, with particular emphasis on the huge influence of conformational flexibility and solvation on chiroptical properties and on the main computational strategies available to describe their effects by means of electronic circular dichroism (ECD) spectroscopy and time-dependent density functional theory (TD-DFT) calculations. The combination of experimental chiroptical spectroscopies with state-of-the-art computational methods proved to be very efficient at predicting the absolute configuration of a wide range of bioactive molecules (fluorinated 2-arylpropionic acids, β-lactam derivatives, difenoconazole, fenoterol, mycoleptones, austdiol). The results obtained for the investigated systems showed that great care must be taken in describing the molecular system in the most accurate fashion, since chiroptical properties are very sensitive to small electronic and conformational perturbations. In the future, the improvement of theoretical models and methods, such as ab initio molecular dynamics, will benefit pharmaceutical analysis in the investigation of non-trivial effects on the chiroptical properties of solvated systems and in the characterisation of the stereochemistry of complex chiral drugs

Molecular Packing and Photoluminescence Efficiency in Odd-Membered Oligothiophene S,S-Dioxides

The single-crystal X-ray structures of three odd-membered thiophene oligomers bearing one central thienyl-S,S-dioxide moietytrimer, pentamer, and heptamerare reported. Absolute photoluminescence quantum yields in microcrystalline powders are given for all compounds. The solid-state photoluminescence efficiencies of the trimer (45%) and the pentamer (12%) were up to 1 order of magnitude higher than those generally measured in conventional oligothiophenes, while that of the heptamer amounted to only 2%. These results are accounted for in terms of molecular packing characteristics, which, owing to the competing effects of dipolar intermolecular interactions between the sulfonyl groups and intra- and intermolecular C−H···O hydrogen bondings and S···S interactions, change dramatically on changing the oligomer size. While the trimer is highly distorted and crystallizes in a chiral orthorhombic space group with the molecular long axes markedly tilted with respect to one another, the heptamer displays a coplanar ..

Microsampling and enantioselective liquid chromatography coupled to mass spectrometry for chiral bioanalysis of novel psychoactive substances

In this paper, the development of efficient enantioselective HPLC methods for the analysis of five benzofuran-substituted phenethylamines, two substituted tryptamines, and three substituted cathinones is described. For the first time, reversed-phase (eluents made up with acidic water-methanol solutions) and polar-ionic (eluent made up with an acetonitrile-methanol solution incorporating both an acidic and a basic additive) conditions fully compatible with mass spectrometry (MS) detectors were applied with a chiral stationary phase (CSP) incorporating the (+)-(18-crown-6)-tetracarboxylic acid chiral selector. Enantioresolution was achieved for nine compounds with α and RS factors up to 1.32 and 5.12, respectively. Circular dichroism (CD) detection, CD spectroscopy in stopped-flow mode and quantum mechanical (QM) calculations were successfully employed to investigate the absolute stereochemistry of mephedrone, methylone and butylone and allowed to establish a (R)<(S) enantiomeric elution order for these compounds on the chosen CSP. Whole blood miniaturized samples collected by means of volumetric absorptive microsampling (VAMS) technology and fortified with the target analytes were extracted following an optimized protocol and effectively analysed by means of an ultra-high performance liquid chromatography-MS system. By this way a proof-of-concept procedure was applied, demonstrating the suitability of the method for quali-quantitative enantioselective assessment of the selected psychoactive substances in advanced biological microsamples. VAMS microsamplers including a polypropylene handle topped with a small tip of a polymeric porous material were used and allowed to volumetrically collect small aliquots of whole blood (10 μL) independently from its density. Highly appreciable volumetric accuracy (bias, in the -8.7-8.1% range) and precision (% CV, in the 2.8-5.9% range) turned out

Investigating in vitro amyloid peptide 1-42 aggregation: impact of higher molecular weight stable adducts

The self-assembly of amyloid peptides (Aβ), in particular Aβ1–42, into oligomers and fibrils is one of the main pathological events related to Alzheimer’s Disease (AD). Recent studies have demonstrated the ability of carbon monoxide releasing molecules (CORMs) to protect neurons and astrocytes from Aβ1-42 toxicity. In fact, CORMs are able to carry and release controlled levels of CO and are known to exert a wide range of anti-inflammatory and anti-apoptotic activities at physiologically relevant concentrations. In order to investigate the direct effects of CORMs on Aβ1–42, we studied the reactivity of CORM-2 and CORM-3 with Aβ1–42 in vitro and the potential inhibition of its aggregation by mass spectrometry (MS), as well as fluorescence and circular dichroism (CD) spectroscopies. The application of an electrospray ionisation-mass spectrometry (ESI-MS) method allowed the detection of stable Aβ1–42/CORMs adducts, involving the addition of the Ru(CO)2 portion of CORMs at histidine residues on the Aβ1-42 skeleton. Moreover, CORMs showed anti-aggregating properties through formation of stable adducts with Aβ1–42 as demonstrated by a thioflavin T (ThT) fluorescence assay and MS analysis. As a further proof, comparison of the CD spectra of Aβ1–42 recorded in the absence and in the presence of CORM-3 at 1:1 molar ratio showed the ability of CORM-3 to stabilize the peptide in its soluble, unordered conformation, thereby preventing its misfolding and aggregation. This multi-methodological investigation revealed novel interactions between Aβ1–42 and CORMs, contributing new insights into the proposed neuroprotective mechanisms mediated by CORMs and disclosing a new strategy to divert amyloid aggregation and toxicity

Evidence of training influence on infant manual behavior: a systematic review

Introduction: Researchers have widely investigated how interventions by means of training can improve manual behaviors in infants. However, no systematic review has been found on this topic. Objective: To analyze the quality of scientific evidence considering the methodological quality and level of evidence by type of study in research on training of object-directed manual behaviors in infants in the first 18 months of life. Methods: National Library of Medicine (PubMed/MEDLINE), Latin American and Caribbean Health Sciences (LILACS), Virtual Health Library (BIREME/BVS), Science Direct, SciELO, and Physiotherapy Evidence Database (PEDro) databases were used. Only clinical trials that assessed the benefits of manual object-directed training in infants and were published up to February 2018, in English, were included. The Cochrane Collaboration Model was adapted to extract bibliographical data from the articles and their methodological quality was assessed using the PEDro scale and the Oxford Centre for Evidence-Based Medicine’s Levels of Evidence. Results: Twenty one clinical trials were included. Studies investigated typically developing full-term infants, preterm infants, and infants at risk for autism spectrum disorders. Trainings were administered to infants by means of “sticky mittens” paradigm, task-specific practice, or contingency reinforcement. Most of the studies presented fair or poor methodological quality. Only studies that used task-specific active practice presented high methodological quality. Conclusions: The results indicate there is high quality evidence that task-specific training improves object-directed manual behaviors in typically developing infants and preterm infants in the first 2-4 months of life. Studies addressing infants with established diagnoses of developmental dysfunction are lacking.Introdução: Pesquisadores tem amplamente investigado como o treino pode melhorar comportamentos manuais em lactentes. No entanto, nenhuma revisão sistemática foi encontrada sobre este tópico. Objetivo: Analisar a qualidade da evidência científica considerando a qualidade metodológica e o nível de evidência por tipo de estudo em pesquisas sobre treino de comportamentos manuais direcionados a objetos em lactentes nos primeiros 18 meses de vida. Método: Foram utilizadas as bases de dados da Biblioteca Nacional de Medicina (PubMed / MEDLINE), Literatura Latino-Americana e do Caribe (LILACS), Biblioteca Virtual em Saúde (BIREME / BVS), Science Direct, SciELO e Physiotherapy Evidence Database (PEDro). Apenas ensaios clínicos que avaliaram os benefícios do treino manual orientado a objetos em lactentes e que foram publicados até fevereiro de 2018, em inglês, foram incluídos. O Modelo de Colaboração Cochrane foi adaptado para extrair dados bibliográficos dos artigos e sua qualidade metodológica foi avaliada pela escala PEDro e pelo Nível de Evidência Científica por Tipo de Estudo de Oxford. Resultados: Vinte e um ensaios clínicos foram incluídos. Os estudos investigaram lactentes nascidos a termo, prematuros e lactentes de risco para distúrbios do espectro autista. Os treinos foram administrados aos lactentes por meio das abordagens do paradigma de “luvas aderentes”, prática específica da tarefa ou reforço de contingência. A maioria dos estudos apresentou qualidade metodológica razoável ou fraca. Apenas os estudos que utilizaram a prática ativa específica da tarefa apresentaram qualidade metodológica alta. Conclusão: Há evidência de alta qualidade de que o treino específico da tarefa aprimora comportamentos manuais orientados a objetos em lactentes com desenvolvimento típico e lactentes prematuros nos primeiros 2-4 meses de vida. Estudos abordando lactentes com diagnósticos estabelecidos de disfunção do desenvolvimento não foram encontrados

Targeted delivery of neutralizing anti-C5 antibody to renal endothelium prevents complement- dependent tissue damage

Complement activation is largely implicated in the pathogenesis of several clinical conditions and its therapeutic neutralization has proven effective in preventing tissue and organ damage. A problem that still needs to be solved in the therapeutic control of complement-mediated diseases is how to avoid side effects associated with chronic neutralization of the complement system, in particular, the increased risk of infections. We addressed this issue developing a strategy based on the preferential delivery of a C5 complement inhibitor to the organ involved in the pathologic process. To this end, we generated Ergidina, a neutralizing recombinant anti-C5 human antibody coupled with a cyclic-RGD peptide, with a distinctive homing property for ischemic endothelial cells and effective in controlling tissue damage in a rat model of renal ischemia/reperfusion injury (IRI). As a result of its preferential localization on renal endothelium, the molecule induced complete inhibition of complement activation at tissue level, and local protection from complement-mediated tissue damage without affecting circulating C5. The ex vivo binding of Ergidina to surgically removed kidney exposed to cold ischemia supports its therapeutic use to prevent posttransplant IRI leading to delay of graft function. Moreover, the finding that the ex vivo binding of Ergidina was not restricted to the kidney, but was also seen on ischemic heart, suggests that this RGD-targeted anti-C5 antibody may represent a useful tool to treat organs prior to transplantation. Based on this evidence, we propose preliminary data showing that Ergidina is a novel targeted drug to prevent complement activation on the endothelium of ischemic kidney

Structural basis for the magnesium-dependent activation of transketolase from Chlamydomonas reinhardtii

Background In photosynthetic organisms, transketolase (TK) is involved in the Calvin-Benson cycle and participates to the regeneration of ribulose-5-phosphate. Previous studies demonstrated that TK catalysis is strictly dependent on thiamine pyrophosphate (TPP) and divalent ions such as Mg2 +. Methods TK from the unicellular green alga Chlamydomonas reinhardtii (CrTK) was recombinantly produced and purified to homogeneity. Biochemical properties of the CrTK enzyme were delineated by activity assays and its structural features determined by CD analysis and X-ray crystallography. Results CrTK is homodimeric and its catalysis depends on the reconstitution of the holo-enzyme in the presence of both TPP and Mg2 +. Activity measurements and CD analysis revealed that the formation of fully active holo-CrTK is Mg2 +-dependent and proceeds with a slow kinetics. The 3Dâstructure of CrTK without cofactors (CrTKapo) shows that two portions of the active site are flexible and disordered while they adopt an ordered conformation in the holo-form. Oxidative treatments revealed that Mg2 +participates in the redox control of CrTK by changing its propensity to be inactivated by oxidation. Indeed, the activity of holo-form is unaffected by oxidation whereas CrTK in the apo-form or reconstituted with the sole TPP show a strong sensitivity to oxidative inactivation. Conclusion These evidences indicate that Mg2 +is fundamental to allow gradual conformational arrangements suited for optimal catalysis. Moreover, Mg2 +is involved in the control of redox sensitivity of CrTK. General significance The importance of Mg2 +in the functionality and redox sensitivity of CrTK is correlated to light-dependent fluctuations of Mg2 +in chloroplasts

Validação da estrutura interna da Escala Brasileira de Vulnerabilidade Odontológica

OBJETIVO: Avaliar evidências de validade da estrutura interna da Escala Brasileira de Vulnerabilidade Odontológica (EVO-BR) quando aplicada no Brasil. MÉTODOS: Trata-se de um estudo de natureza psicométrica, que busca validar uma escala construída através de evidências de estrutura interna. A coleta de dados foi realizada em 18 unidades básicas de saúde que executam a metodologia da Planificação da Atenção à Saúde, distribuídas nas cinco regiões do Brasil. A versão inicial da EVO-BR continha 41 itens que mediam vulnerabilidade odontológica e foram aplicados por em usuários com 18 anos ou mais, usuários do Sistema Único de Saúde, que estivessem nas unidades básicas de saúde para consulta por profissionais de nível superior. Para avaliação das evidências foram utilizadas as seguintes análises estatísticas: análise fatorial exploratória, confirmatória e network analysis. RESULTADOS: Participaram do estudo 1.753 usuários. Para adequação da amostra considerou-se a fatorabilidade obtida de Kaiser-Meyer-Olkin (KMO) = 0,65, Bartlett sphericity = 8019,7 e determinante da matriz de 0,008. A análise paralela inicial indicou um modelo de 4 dimensões e teve os itens ajustados conforme cargas fatorais (variaram de 0,38 a 0,99), comunalidades (0,13 a 0,89) e Pratt’s measure até que o modelo tivesse congruência nos princípios estatístico e interpretativo simultaneamente. O modelo final apresentou 15 itens, manteve a indicação de quatro dimensões pela análise paralela e com uma variância explicada de 68,56%. CONCLUSÕES: A EVO-BR é uma escala validada para mensurar vulnerabilidade odontológica, e pode contribuir para organização do acesso a equipe de saúde bucal na atenção primária à saúde por meio da estratificação da população, como recomendado na planificação

Targeting CD34(+) cells of the inflamed synovial endothelium by guided nanoparticles for the treatment of rheumatoid arthritis

Despite the advances in the treatment of rheumatoid arthritis (RA) achieved in the last few years, several patients are diagnosed late, do not respond to or have to stop therapy because of inefficacy and/or toxicity, leaving still a huge unmet need. Tissue-specific strategies have the potential to address some of these issues. The aim of the study is the development of a safe nanotechnology approach for tissue-specific delivery of drugs and diagnostic probes. CD34 + endothelial precursors were addressed in inflamed synovium using targeted biodegradable nanoparticles (tBNPs). These nanostructures were made of poly-lactic acid, poly-caprolactone, and PEG and then coated with a synovial homing peptide. Immunofluorescence analysis clearly demonstrated their capacity to selectively address CD34 + endothelial cells in synovial tissue obtained from human, mouse, and rat. Biodistribution studies in two different animal models of rheumatoid arthritis (antigen-induced arthritis/AIA and collagen-induced arthritis/CIA) confirmed the selective accumulation in inflamed joints but also evidenced the capacity of tBNP to detect early phases of the disease and the preferential liver elimination. The therapeutic effect of methotrexate (MTX)-loaded tBNPs were studied in comparison with conventional MTX doses. MTX-loaded tBNPs prevented and treated CIA and AIA at a lower dose and reduced administration frequency than MTX. Moreover, MTX-loaded tBNP showed a novel mechanism of action, in which the particles target and kill CD34 + endothelial progenitors, preventing neo-angiogenesis and, consequently, synovial inflammation. tBNPs represent a stable and safe platform to develop highly-sensitive imaging and therapeutic approaches in RA targeting specifically synovial neo-angiogenesis to reduce local inflammation