Insights into Insulin Fibril Assembly at Physiological and Acidic pH and Related Amyloid Intrinsic Fluorescence

Human insulin is a widely used model protein for the study of amyloid formation as both associated to insulin injection amyloidosis in type II diabetes and highly prone to form amyloid fibrils in vitro. In this study, we aim to gain new structural insights into insulin fibril formation under two different aggregating conditions at neutral and acidic pH, using a combination of fluorescence, circular dichroism, Fourier-transform infrared spectroscopy, and transmission electron miscroscopy. We reveal that fibrils formed at neutral pH are morphologically different from those obtained at lower pH. Moreover, differences in FTIR spectra were also detected. In addition, only insulin fibrils formed at neutral pH showed the characteristic blue-green fluorescence generally associated to amyloid fibrils. So far, the molecular origin of this fluorescence phenomenon has not been clarified and different hypotheses have been proposed. In this respect, our data provide experimental evidence that allow identifying the molecular origin of such intrinsic property

Monitoring Production Process of Cisplatin-Loaded PLGA Nanoparticles by FT-IR Microspectroscopy and Univariate Data Analysis

Cisplatin-loaded PLGA nanoparticles for drug delivery have been prepared using a well-established water/oil/water double emulsion-solvent evaporation method. The production process has been monitored by using Fourier Transform Infrared (FT-IR) microspectroscopy without using KBr tablets and any preliminary sample preparation. Significant spectra have been obtained for all chemical compounds and for samples at different steps of production process. The use of a linear or univariate approach using a R2 determination coefficient has been proposed for discriminating among FT-IR spectra even when small differences are present. The obtained results confirm that new geometries of data acquisition contribute to make infrared microspectroscopy a very useful tool for a rapid and detailed monitoring of production processes of pharmacological interest. Moreover, morphological and physiological characterizations have been performed on cisplatin-loded samples showing results in good agreement with those reported in literatur

Visible micro-Raman spectroscopy for determining glucose content in beverage industry

The potential of Raman spectroscopy with excitation in the visible as a tool for quantitative determination of single components in food industry products was investigated by focusing the attention on glucose content in commercial sport drinks. At this aim, micro-Raman spectra in the 600–1600 cm1 wavenumber shift region of four sport drinks were recorded, showing well defined and separated vibrational fingerprints of the various contained sugars (glucose, fructose and sucrose). By profiting of the spectral separation of some peculiar peaks, glucose content was quantified by using a multivariate statistical analysis based on the interval Partial Least Square (iPLS) approach. The iPLS model needed for data analysis procedure was built by using glucose aqueous solutions at known sugar concentrations as calibration data. This model was then applied to sport drink spectra and gave predicted glucose concentrations in good agreement with the values obtained by using a biochemical assay. These results represent a significant step towards the development of a fast and simple method for the on-line glucose quantification in products of food and beverage industry

Preparation and characterization of 3D hyaluronic-acid-based scaffolds with controlled optical properties for biomedical applications

Optical techniques are increasingly employed for monitoring cell-matrix interactions and the availability of 3D scaffolds with controlled optical properties can be extremely useful in this field. Hyaluronic acid (HA) is a glycosaminoglycan found in extracellular tissue in many parts of the body which is becoming more and more popular in biomaterials science as key material for the creation of new tissues using scaffolds. In this frame, a crosslinked hyaluronic acid-lys scaffold has been prepared and characterized. The peculiar optical properties of this scaffold make it a good candidate for exploiting optical techniques for monitoring cell-matrix interactions. Its optical and physico-chemical properties have been investigated by means of UV-VIS and FT-IR spectroscopy. In particular, transmittance and fluorescence measurements have shown low levels of background signals coming from the scaffold itself. In addition the optical quality of 3D scaffolds has been preliminarily tested by two-photon microscopy imaging of embedded fluorescent microbeads and labelled human keratinocites cells at different depths from the scaffold surface