Encapsulation of biomacromolecules in metal–organic frameworks (MOFs) can preserve biological functionality in harsh environments. Despite the success of this approach, termed biomimietic ...mineralization, limited consideration has been given to the chemistry of the MOF coating. Here, we show that enzymes encapsulated within hydrophilic MAF-7 or ZIF-90 retain enzymatic activity upon encapsulation and when exposed to high temperatures, denaturing or proteolytic agents, and organic solvents, whereas hydrophobic ZIF-8 affords inactive catalase and negligible protection to urease.
Fish intestine plays various physiological functions and its integrity is essential to guarantee fish growth and welfare. Several laboratory techniques have been proposed for studying ...morpho-physiology of fish intestine, but classical histology still represents one of the gold standards. Nevertheless, this technique is time consuming and does not provide comprehensive information on the biochemical composition of tissues. The present study, introduces a new approach for studying fish intestine, the Fourier Transform Infrared (FTIR) spectroscopy. This is a fast, label-free analytical technique, which analyses the vibrational transitions induced on tissues by the interaction with the electromagnetic radiation. On the base of vibrational features of some functional groups classes and molecular bonds, FTIR allows the analysis of the macromolecular composition of the sample. Rainbow trout (O. mykiss) represents an important species for modern aquaculture and deeper knowledge about its intestine anatomy and macromolecular composition is desirable. In the present study, besides the traditional histological analysis, the biochemical building of the intestine mucosa was analyzed for the first time by FTIR. Spectral data can be considered a complement to traditional histological analysis.
•A new method for fish intestine analysis was tested and verified.•A comparison with classical histology was performed.•FTIR spectroscopy can be considered a complementary tool to histology.
Label-free spectromicroscopy methods offer the capability to examine complex cellular phenomena. Electron and X-ray based spectromicroscopy methods, though powerful, have been hard to implement with ...hydrated objects due to the vacuum incompatibility of the samples and due to the parasitic signals from (or drastic attenuation by) the liquid matrix surrounding the biological object of interest. Similarly, for many techniques that operate at ambient pressure, such as Fourier transform infrared spectromicroscopy (FTIRM), the aqueous environment imposes severe limitations due to the strong absorption of liquid water in the infrared regime. Here we propose a microfabricated multi-compartmental and reusable hydrated sample platform suitable for use with several analytical techniques, which employs the conformal encapsulation of biological specimens by a few layers of atomically thin graphene. Such an electron, X-ray, and infrared transparent, molecularly impermeable and mechanically robust enclosure preserves the hydrated environment around the object for a sufficient time to allow
examination of hydrated bio-objects with techniques operating under both ambient and high vacuum conditions. An additional hydration source, provided by hydrogel pads lithographically patterned in the liquid state near/around the specimen and co-encapsulated, has been added to further extend the hydration lifetime. Note that the in-liquid lithographic electron beam-induced gelation procedure allows for addressable capture and immobilization of the biological cells from the solution. Scanning electron microscopy and optical fluorescence microscopy, as well as synchrotron radiation based FTIR and X-ray fluorescence microscopy, have been used to test the applicability of the platform and for its validation with yeast, A549 human carcinoma lung cells and micropatterned gels as biological object phantoms.
Biofilms are communities of bacteria living embedded in a highly hydrated matrix composed of polysaccharides, proteins, and extracellular DNA. This life style confers numerous advantages to bacteria ...including protection against external threats. However, they also contribute to increase bacterial resistance against antimicrobials, an issue particularly relevant in dangerous infections. Due to the complexity of the matrix, few information is present in the literature on details of its architecture including the spatial distribution of the macromolecular components which might give hints on the way the biofilm scaffold is built up by bacteria. In this study, we investigated the possibility to combine well-established microbiological procedures with advanced microscopies to get information on composition and distribution of the macromolecular components of biofilm matrices. To this, confocal microscopy, diffraction-limited infrared (IR) spectral imaging, and atomic force microscopy (AFM) were used to explore biofilm produced by a clinical strain of
Klebsiella pneumoniae
. IR imaging permitted to have clues on how the biofilm grows and spreads on surfaces, and the local distribution of the components within it. Through the analysis of the pure component spectra, it was possible to assess the chemical and structural composition of the saccaridic matrix, confirming the data obtained by NMR. It was also possible to follow the time course of biofilm from 6 up to 48 h when the biofilm grew into a 3-dimensional multi-layered structure, characteristic of colonies of bacteria linked together by a complex matrix. In addition, nanoFTIR and AFM investigations allowed the estimation of biofilm growth in the vertical direction and the morphological analysis of bacterial colonies at different time points and the evaluation of the chemical composition at the nanoscale.
Magnetic nanoparticles (MNPs) have shown exceptional potential for several biological and clinical applications. However, MNPs must be coated by a biocompatible shell for such applications. The aim ...of this study is to understand if and how the surface charge and coating can affect the electronic and magnetic properties of CoFe2O4 MNPs. The role of the surface on the total magnetic moment of MNPs is a controversial issue, and several effects can contribute to make it deviate from the bulk value, including the charge, the nature of the coating, and also the synthetic technique. Positively and negatively charged uncoated CoFe2O4 NPs as well as citrate-coated NPs were prepared by soft chemistry synthesis. The electronic properties and cationic distribution of CoFe2O4 NPs were probed by X-ray absorption spectroscopy (XAS), X-ray magnetic circular dichroism (XMCD), and X-ray photoemission spectroscopy (XPS) techniques and confirmed by theoretical simulations. The overall magnetic behavior and the hyperthermic properties were evaluated by magnetometry and calorimetric measurements, respectively. The results show that all of the investigated CoFe2O4 NPs have high magnetic anisotropy energy, and the surface charge and coating do not influence appreciably their electronic and magnetic properties. In addition, the citrate shell improves the stability of the NPs in aqueous environment, making CoFe2O4 NPs suitable for biomedical applications.
Technical limitations regarding bulk analysis of phytoplankton biomass limit our comprehension of carbon fluxes in natural populations and, therefore, of carbon, nutrients and energy cycling in ...aquatic ecosystems. In this study, we took advantage of Synchrotron FTIR micro-spectroscopy and the partial least square regression (PLSr) algorithm to simultaneously quantify the protein, lipid and carbohydrate content at the single-cell level in a mock phytoplankton community (composed by a cyanobacterium, a green-alga and a diatom) grown at two temperatures (15 °C and 25 °C).
The PLSr models generated to quantify cell macromolecules presented high quality fit (R
≥ 0.90) and low error of prediction (RMSEP 2-6% of dry weight). The regression coefficients revealed that the prediction of each macromolecule was not exclusively dependent on spectral features corresponding to that compound, but rather on all major macromolecular pools, reflecting adjustments in the overall cell carbon balance. The single-cell analysis, studied by means of Kernel density estimators, showed that the modes of density distribution of macromolecules were different at 15 °C and 25 °C. However, a substantial proportion of cells was biochemically identical at the two temperatures because of population heterogeneity.
The spectroscopic approach presented in this study allows the quantification of macromolecules in single phytoplankton cells. This method showed that population heterogeneity most likely ensures a backup of non-acclimated cells that may rapidly exploit new favourable niches. This finding may have important consequences for the ecology of phytoplankton populations and shows that the "average cell" concept might substantially limit our comprehension of population dynamics and biogeochemical cycles in aquatic ecosystems.
Here we present a new bonding protocol for SU-8 negative tone photoresist that exploits the chemical modifications induced in the resin by exposure to 254 nm (UVC) light. Fourier Transform Infrared ...microspectroscopy (μ-FTIR) was used to carry out a thorough study on the chemical processes and modifications occurring within the epoxy resin by exposure to 365 nm and 254 nm light. In particular, we established that UVC light promotes the opening of the epoxy rings bypassing the post-exposure bake. The possibility to promote a further activation of the resin, already patterned with standard UV lithography, was exploited to produce closed microfluidic devices. Specifically, we were able to fabricate fluidic chips, characterized by broadband transparency from mid-IR to UV and long term stability in continuous flow conditions. CaF2 was used as substrate, coated by sputtering with a nanometric silicon film, in order to make surface properties of this material more suitable for standard fabrication processes with respect to the original substrate. The fabricated microfluidic chips were used to study by μ-FTIR the biochemical response of live breast cancer MCF-7 cells to osmotic stress and their subsequent lysis induced by the injection of deionized water in the device. μ-FTIR analyses detected fast changes in protein, lipid and nucleic acid content as well as cytosol acidification.
The mesenchymal state in cancer is usually associated with poor prognosis due to the metastatic predisposition and the hyper-activated metabolism. Exploiting cell glucose metabolism we propose a new ...method to detect mesenchymal-like cancer cells. We demonstrate that the uptake of glucose-coated magnetic nanoparticles (MNPs) by mesenchymal-like cells remains constant when the glucose in the medium is increased from low (5.5 mM) to high (25 mM) concentration, while the MNPs uptake by epithelial-like cells is significantly reduced. These findings reveal that the glucose-shell of MNPs plays a major role in recognition of cells with high-metabolic activity. By selectively blocking the glucose transporter 1 channels we showed its involvement in the internalization process of glucose-coated MNPs. Our results suggest that glucose-coated MNPs can be used for metabolic-based assays aimed at detecting cancer cells and that can be used to selectively target cancer cells taking advantage, for instance, of the magnetic-thermotherapy.
Superparamagnetic iron oxide nanoparticles (SPIONs) have great potential for use in medicine, but they may cause side effects due to oxidative stress. In our study, we investigated the effects of ...silica-coated SPIONs on endothelial cells and whether oleic acid (OA) can protect the cells from their harmful effects. We used viability assays, flow cytometry, infrared spectroscopy, fluorescence microscopy, and transmission electron microscopy. Our results show that silica-coated SPIONs are internalized by endothelial cells, where they increase the amount of reactive oxygen species (ROS) and cause cell death. Exposure to silica-coated SPIONs induced accumulation of lipid droplets (LD) that was not dependent on diacylglycerol acyltransferase (DGAT)-mediated LD biogenesis, suggesting that silica-coated SPIONs suppress LD degradation. Addition of exogenous OA promoted LD biogenesis and reduced SPION-dependent increases in oxidative stress and cell death. However, exogenous OA protected cells from SPION-induced cell damage even in the presence of DGAT inhibitors, implying that LDs are not required for the protective effect of exogenous OA. The molecular phenotype of the cells determined by Fourier transform infrared spectroscopy confirmed the destructive effect of silica-coated SPIONs and the ameliorative role of OA in the case of oxidative stress. Thus, exogenous OA protects endothelial cells from SPION-induced oxidative stress and cell death independent of its incorporation into triglycerides.
Tannin-furanic rigid foams are bio-based copolymers of tannin plant extract and furfuryl alcohol, promising candidates to replace synthetic insulation foams, as for example polyurethanes and ...phenolics, in eco-sustainable buildings thanks to their functional properties, such as lightness of the material and fire resistance. Despite their relevance as environmental-friendly alternatives to petroleum derivatives, many aspects of the polymerization chemistry still remain unclear. One of the open issues is on the spatial heterogeneity of the foam, i.e., whether the foam constituents prevalently polymerize in spatially segregated blocks or distribute almost homogenously in the foam volume. To address this matter, here we propose a multiscale FTIR study encompassing 1D FTIR spectroscopy, 2D FTIR imaging and 3D FTIR micro-tomography (FTIR-μCT) on tannin-furanic rigid foams obtained by varying the synthesis parameters in a controlled way. Thanks to the implementation of the acquisition and processing pipeline of FTIR-μCT, we were able for the first time to demonstrate that the polymer formulations influence the spatial organization of the foam at the microscale and, at the same time, prove the reliability of FTIR-μCT data by comparing 2D FTIR images and the projection of the 3D chemical images on the same plane.
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