Fast, Easy, and\nReproducible Fingerprint Methods for\nEndotoxin Characterization in Nanocellulose and Alginate-Based Hydrogel\nScaffolds

Nanocellulose- and\nalginate-based hydrogels have been\nsuggested\nas potential wound-healing materials, but their utilization is limited\nby the Food and Drug Administration (FDA) requirements regarding endotoxin\nlevels. Cytotoxicity and the presence of endotoxin were assessed after\ngel sterilization using an autoclave and UV treatment. A new fingerprinting\nmethod was developed to characterize the compounds detected in cellulose\nnanocrystal (CNC)- and cellulose-nanofiber (CNF)-based hydrogels using\nboth positive- and negative-ion mode electrospray ionization Fourier\ntransform ion cyclotron resonance mass spectroscopy (ESI FT-ICR MS).\nThese biobased hydrogels were used as scaffolds for the cultivation\nand growth of human dermal fibroblasts to test their biocompatibility.\nA resazurin-based assay was preferred over all other biocompatibility\nmethodologies since it allowed for the evaluation of viability over\ntime in the same sample without causing cell lysis. The CNF dispersion\nof 6 EU mL<sup>–1</sup> was slightly above the limits, and\nit did not affect the cell viability, whereas CNC hydrogels induced\na reduction of metabolic activity by the fibroblasts. The chemical\nstructure of the detected endotoxins did not contain phosphates, but\nit encompassed hydrophobic sulfonate groups, requiring the development\nof new high-pressure sterilization methods for the use of cellulose\nhydrogels in medicine.