Immobilized bacteria are being extensively used for metabolite production, biocatalysts, and biosensor construction. However, long-term viability and metabolic activity of entrapped bacteria is affected by several conditions such as their physiological state, the presence of high-osmolarity environments, porous structure and shrinkage of the matrix. The aim of this work was to evaluate the effect of various parameters on bacteria immobilized in sol-gel-derived silica matrices. With this purpose, we evaluated the stress of immobilization over bacteria cultures obtained from different growing states, the effect of cell density and bacteria capability to proliferate inside matrices. Best results to attain longer preservation times were obtained when we immobilized suspensions with an optimized bacterial number of 1 x 10(7) cfu/gel in the presence of LB medium using aqueous silica precursors. Furthermore, the impact of osmotic stress with the subsequent intracellular trehalose accumulation and the addition of osmolites were investigated. Shorter preservation times were found for bacteria immobilized in the presence of osmolites while trehalose accumulation in stressed cells did not produce changes on entrapped bacteria viability. Finally, nutrient addition in silica matrices was studied indicating that the presence of a carbon source without the simultaneous addition of nitrogen was detrimental for immobilized E. coli. However, when both carbon and nitrogen sources were present, bacteria were able to survive longer periods of time.
Silica-collagen scaffolds were obtained by covalent binding of an aminosilane to glutaraldehyde fixed collagen hydrogels, rendering a three dimensional network of silicon coated collagen fibrils. When compared to non-silicified collagen, silica containing matrices exhibited a 60 fold increment in the rheological properties. Moreover, acellular degradation by collagenase type I indicated that enzymatic digestion occurred at a slower rate for silica modified hydrogels, hence enabling a controlled degradation of the obtained material. In addition, fibroblastic cells seeded on silicified collagen matrices were able to adhere, proliferate and migrate within the scaffold for over 3 weeks as shown by MTT tests and hematoxylin-eosin staining. These results suggest that the herein described method could be useful in the design of materials for tissue engineering purposes.
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