Emergent Glycerophospholipid Fluorescent Probes: Synthesis and Applications.

Fluorescent labeling through bioconjugation is the preferred tool for the investigation of biological functions involving lipids, namely for clarifying metabolic pathways and molecular mechanisms of diseases. The lack of functionalized lipid probes with biological and physicochemical properties suitable for these studies is still a major limitation. Moreover, the synthesis of these probes is challenging and strongly dependent on the application envisioned. The objective of this Review is to highlight advances in the application of fluorescent glycerophospholipid probes through innovative approaches in the synthesis reported in the past decade. The reaction pathways, choice of fluorophore, and location of fluorophore in the glycerophospholipid structure are critically addressed. The relevance of these bioconjugates is exemplified with applications using advanced analysis by fluorescence enhancement or quenching to unravel biomembrane structure features and phospholipase activity. Finally, this Review reinforces the need for innovative and more efficient routes for the synthesis of tailored glycerophospholipids fluorescent conjugates.

Mental health professionals' attitudes towards mental illness: professional and cultural factors in the INTER NOS study.

BACKGROUND: Research shows that personnel working in mental health facilities may share some of the societal prejudices towards mental illness. This might result in stigmatizing behaviours towards people suffering from mental disorders, undermining the quality of their care. AIMS: To describe and compare attitudes towards mental illness across a sample of professionals working in a wide range of mental health facilities in Spain, Portugal and Italy. METHOD: We administered a survey to personnel including two questionnaires related to stigmatizing attitudes: The Community Attitudes toward the Mentally Ill (CAMI) and the Attribution Questionnaire (AQ-27). Data were compared according to professional category, work setting and country. RESULTS: 34.06% (1525) professionals of the surveyed population responded adequately. Psychologists and social therapists had the most positive attitudes, and nursing assistants the most negative, on most factors of CAMI and AQ-27. Community staff had more positive attitudes than hospital-based professionals in most factors on CAMI and in discriminatory responses on AQ-27. CONCLUSIONS: Globally, mental health professionals showed a positive attitude towards mental illness, but also a relative support to coercive treatments. There are differences in attitudes modulated by professional category and setting. Results can guide preventive strategies, particularly for the hospital-based and nursing staff.

Flow-based bioconjugation of coumarin phosphatidylethanolamine probes: Optimised synthesis and membrane molecular dynamics studies.

A series of phosphatidylethanolamine fluorescent probes head-labelled with 3-carboxycoumarin was prepared by an improved bioconjugation approach through continuous flow synthesis. The established procedure, supported by a design of experiment (DoE) set-up, resulted in a significant reduction in the reaction time compared to the conventional batch method, in addition to a minor yield increase. The characterization of these probes was enhanced by an in-depth molecular dynamics (MD) study of the behaviour of a representative probe of this family, 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine labelled with 3-carboxycoumarin (POPE-COUM), in bilayers of 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC)/1-stearoyl-2-linoleoyl-sn-glycero-3-phosphocholine (SLPC) 2:1, mimicking the composition of the egg yolk lecithin membranes recently used experimentally by our group to study POPE-COUM as a biomarker of the oxidation state and integrity of large unilamellar vesicles (LUVs). The MD simulations revealed that the coumarin group is oriented towards the bilayer interior, leading to a relatively internal location, in agreement with what is observed in the nitrobenzoxadiazole fluorophore of commercial head-labelled NBD-PE probes. This behaviour is consistent with the previously stated hypothesis that POPE-COUM is entirely located within the LUVs structure. Hence, the delay on the oxidation of the probe in the oxygen radical absorbance capacity (ORAC) assays performed is related with the inaccessibility of the probe until alteration of the LUV structure occurs. Furthermore, our simulations show that POPE-COUM exerts very little global and local perturbation on the host bilayer, as evaluated by key properties of the unlabelled lipids. Together, our findings establish PE-COUM as suitable fluorescent lipid analogue probes.

Gas-phase structural characterization of neuropeptides Y Y1 receptor antagonists using mass spectrometry: Orbitrap vs triple quadrupole.

Collision induced dissociation of triple quadrupole mass spectrometer (CID-QqQ) and high-energy collision dissociation (HCD) of Orbitrap were compared for four neuropeptides Y Y1 (NPY Y1) receptor antagonists and showed similar qualitative fragmentation and structural information. Orbitrap high resolution and high mass accuracy HCD fragmentation spectra allowed unambiguous identification of product ions in the range 0.04-4.25 ppm. Orbitrap mass spectrometry showed abundant analyte-specific product ions also observed on CID-QqQ. These results show the suitability of these product ions for use in quantitative analysis by MRM mode. In addition, it was found that all compounds could be determined at levels >1 µg L-1 using the QqQ instrument and that the detection limits for this analyzer ranged from 0.02 to 0.6 µg L-1. Overall, the results obtained from experiments acquired in QqQ show a good agreement with those acquired from the Orbitrap instrument allowing the use of this relatively inexpensive technique (QqQ) for accurate quantification of these compounds in clinical and academic applications.