confocal Raman microscopy Dotaz Zobrazit nápovědu
In this study, we investigated the release mechanism of the poorly water soluble drug aprepitant from different amorphous solid dispersions using confocal Raman microscopy (CRM). Solid dispersions were fabricated based on either Soluplus®, as an amphiphilic copolymer and solubilizer, or on polyvinylpyrrolidone, as a hydrophilic polymer, in order to elucidate the influence of the polymer characteristics on the drug form and dissolution mechanisms. Aprepitant exhibited its amorphous form in both solid dispersions. However, the release differed depending on the polymer. The high complexation effect of Soluplus was shown to be a crucial factor for stabilization of the amorphous drug, resulting in continuous release without any recrystallization of aprepitant. In contrast, solid dispersions based on polyvinylpyrrolidone showed a different mechanism of dissolution; due to the good affinity of PVP and water, the polymer is dissolving fast, leading to phase separation and local recrystallization of the drug. The study highlights the complexity of release processes from solid dispersions and elucidates the influence of the polymer on drug release kinetics.
- MeSH
- hydrofobní a hydrofilní interakce MeSH
- kinetika MeSH
- konfokální mikroskopie metody MeSH
- krystalizace MeSH
- morfoliny chemie MeSH
- nosiče léků chemie MeSH
- polyethylenglykoly chemie MeSH
- polymery chemie MeSH
- polyvinyly chemie MeSH
- povidon chemie MeSH
- Ramanova spektroskopie metody MeSH
- rozpustnost MeSH
- stabilita léku MeSH
- uvolňování léčiv * MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Photosynthetic energy conversion and the resulting photoautotrophic growth of green algae can only occur in daylight, but DNA replication, nuclear and cellular divisions occur often during the night. With such a light/dark regime, an algal culture becomes synchronized. In this study, using synchronized cultures of the green alga Desmodesmus quadricauda, the dynamics of starch, lipid, polyphosphate, and guanine pools were investigated during the cell cycle by two independent methodologies; conventional biochemical analyzes of cell suspensions and confocal Raman microscopy of single algal cells. Raman microscopy reports not only on mean concentrations, but also on the distribution of pools within cells. This is more sensitive in detecting lipids than biochemical analysis, but both methods-as well as conventional fluorescence microscopy-were comparable in detecting polyphosphates. Discrepancies in the detection of starch by Raman microscopy are discussed. The power of Raman microscopy was proven to be particularly valuable in the detection of guanine, which was traceable by its unique vibrational signature. Guanine microcrystals occurred specifically at around the time of DNA replication and prior to nuclear division. Interestingly, guanine crystals co-localized with polyphosphates in the vicinity of nuclei around the time of nuclear division.
- MeSH
- buněčná stěna chemie MeSH
- buněčný cyklus * MeSH
- časové faktory MeSH
- Chlorophyta cytologie růst a vývoj MeSH
- guanin analýza MeSH
- lipidová tělíska metabolismus MeSH
- lipidy analýza MeSH
- mikroskopie * MeSH
- polyfosfáty analýza MeSH
- Ramanova spektroskopie * MeSH
- škrob analýza MeSH
- velikost buňky MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- srovnávací studie MeSH
- Publikační typ
- abstrakt z konference MeSH
The biochemical responses of rock-inhabiting cyanobacteria towards native environmental stresses were observed in vivo in one of the Earth's most challenging extreme climatic environments. The cryptoendolithic cyanobacterial colonization, dominated by Chroococcidiopsis sp., was studied in an ignimbrite at a high altitude volcanic area in the Atacama Desert, Chile. Change in the carotenoid composition (red-shift) within a transect through the cyanobacteria dominant microbial community (average thickness ~1 mm) was unambiguously revealed in their natural endolithic microhabitat. The amount of red shifted carotenoid, observed for the first time in a natural microbial ecosystem, is depth dependent, and increased with increasing proximity to the rock surface, as proven by resonance Raman imaging and point resonance Raman profiling. It is attributed to a light-dependent change in carotenoid conjugation, associated with the light-adaptation strategy of cyanobacteria. A hypothesis is proposed for the possible role of an orange carotenoid protein (OCP) mediated non-photochemical quenching (NPQ) mechanism that influences the observed spectral behavior. Simultaneously, information about the distribution of scytonemin and phycobiliproteins was obtained. Scytonemin was detected in the uppermost cyanobacteria aggregates. A reverse signal intensity gradient of phycobiliproteins was registered, increasing with deeper positions as a response of the cyanobacterial light harvesting complex to low-light conditions.
- MeSH
- biologické pigmenty MeSH
- ekosystém MeSH
- fluorescenční mikroskopie MeSH
- karotenoidy chemie metabolismus MeSH
- konfokální mikroskopie MeSH
- mikrobiologie životního prostředí MeSH
- pouštní klima * MeSH
- sinice * izolace a purifikace metabolismus MeSH
- spektrální analýza MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Enterococcus faecalis is a ubiquitous bacterium of the gut that is observed in persistent periradicular infections. Its pathogenicity is associated with biofilm formation and the ability to survive under nutrient-poor (starvation) conditions. However, characteristics of chemical composition of biofilm cells developed by starved E. faecalis cells remain poorly understood. In this study, E. faecalis cells in exponential, stationary, and starvation phases were prepared and separately cultured to form biofilms. Confocal laser scanning microscopy was performed to verify biofilm formation. Raman microscopy was used to investigate the chemical composition of cells within the biofilms. Compared to cells in exponential or stationary phase, starved cells developed biofilms with fewer culturable cells (P < 0.05). Raman analysis revealed that cells produced in the biofilms from starved planktonic cells contained more protein and less nucleic acids than either the corresponding planktonic cells or the cells in biofilms from planktonic cells in exponential or stationary phases, suggesting that biofilm-grown cells from the starvation phase were characterized by increased synthesis of proteins and decreased nucleic acids. This study provides an insight into the chemical composition of biofilm cells developed by starved E. faecalis.
The aim of this study was to investigate the molecular structures of tadalafil solid dispersions prepared by different techniques and further to relate them to surface free energy information indicating the final amorphousness of the product. Thus, we tried to complement the existing knowledge of solid dispersion formation. Poorly water-soluble tadalafil was combined with different polymers, i.e. Kollidon® 12 PF, Kollidon® VA 64 and Soluplus®, to form model systems. To assess the extent of drug-polymer miscibility, we studied model solid dispersion surface energy using inverse gas chromatography and phase micro-structure using confocal Raman microscopy. The selection of the preparation method was found to play a crucial role in the molecular arrangement of the incorporated drug and the polymer in resulting solid dispersion. Our results showed that a lower surface free energy indicated the formation of a more homogeneous solid dispersion. Conversely, a higher surface free energy corresponded to the heterogeneous systems containing tadalafil amorphous clusters that were captured by Raman mapping. Thus, we successfully introduced a novel evaluation approach of the drug molecular arrangement in solid dispersions that is especially useful for examining the miscibility of the components when the conventional characterizing techniques are inconclusive or yield variable results.
- MeSH
- chromatografie plynová MeSH
- polymery * chemie MeSH
- povidon * chemie MeSH
- rozpustnost MeSH
- tadalafil chemie MeSH
- Publikační typ
- časopisecké články MeSH
Avian (ortho)reovirus (ARV), which belongs to Reoviridae family, is a major domestic fowl pathogen and is the causative agent of viral tenosynovitis and chronic respiratory disease in chicken. ARV replicates within cytoplasmic inclusions, so-called viral factories, that form by phase separation and thus belong to a wider class of biological condensates. Here, we evaluate different optical imaging methods that have been developed or adapted to follow formation, fluidity and composition of viral factories and compare them with the complementary structural information obtained by well-established transmission electron microscopy and electron tomography. The molecular and cellular biology aspects for setting up and following virus infection in cells by imaging are described first. We then demonstrate that a wide-field version of fluorescence recovery after photobleaching is an effective tool to measure fluidity of mobile viral factories. A new technique, holotomographic phase microscopy, is then used for imaging of viral factory formation in live cells in three dimensions. Confocal Raman microscopy of infected cells provides "chemical" contrast for label-free segmentation of images and addresses important questions about biomolecular concentrations within viral factories and other biological condensates. Optical imaging is complemented by electron microscopy and tomography which supply higher resolution structural detail, including visualization of individual virions within the three-dimensional cellular context.
