Browsing by Subject "freeze-drying"

Native nanofibrillated cellulose is wood-derived, animal-free biocompatible biomaterial which has proved the suitability of nanoscale cellulose fiber based hydrogels for 3D cell culturing and wound healing applications. The problem of freeze-drying nanofibrillated cellulose hydrogel (NFCh) has been the aggregation of the hydrophilic fibrils of the NFC during freeze-drying, which leads deformed freeze-dried cake and unsuccessful reconstitution of the sample. Molecular Dynamic (MD) simulations have been earlier applied in formulation design of NFCh for freeze-drying successfully by screening excipients based on their attraction to the surface of NFCh. The weakness of MD simulations is it can only model the fresh formulation system intend to freeze-dry, but not the actual freeze-drying process and the effect of it and the excipients to the material. To evaluate the protecting properties of excipients and therefore the accuracy of the MD simulations detailed information about changes in the physical state and molecular orientation of the formulation before and after freeze-drying is needed. Non-invasive and label-free Raman spectroscopy can be used to determine vibrational modes of molecules to investigate changes in molecular orientation of the material. The aim of this study was to investigate the possible molecular changes induced by freeze-drying of NFCh-based formulations utilizing Raman spectroscopy and evaluate the connection of the results to MD simulations. NFCh with different excipients was freeze-dried and physicochemical properties, rheology and Raman signal were measured before and after freeze-drying and compared to the literature of MD simulations. The principal component analysis (PCA) was done to the Raman spectra and differences evaluated. The spectra of all formulations differed before and after freeze-drying, and more detailed analysis was done to two most potential 0.8% NFCh based formulations, lactose 300 mM and lactose 250 mM + glycine 50 mM. They had great attraction to NFCh in MD simulations and very similar rheological properties before and after freeze-drying and reconstitution. The spectra of different state of both formulations different on areas between 400 - 500 cm-1 and 850 - 900 cm-1 based on PCA analysis contributing the mutarotation of lactose during freeze-drying and reconstitution. Freeze-drying and the absence of water molecules in NFCh formulation favor different ratios of β and α anomers than the fresh hydrated state which could be detected utilizing Raman spectroscopy. Therefore, Raman spectroscopy was confirmed to be a sensitive option to assess subtle changes in molecular orientation in fresh, freeze-dried, and reconstituted NFCh-based formulations, resulting in a detail knowledge of the molecular behavior of excipients which could be applied in MD simulations and design of better freeze-drying formulations in future.