Assessment of Cellulose Nanofiber-Based Elastase Biosensors to Inflammatory Disease as a Function of Spacer Length and Fluorescence Response

Authors: Easson, Michael; Jordan, Jacobs; Prevost, Nicolette; Edwards, Judson; DUPRE, REBECCA - Oak Ridge Institute For Science And Education (ORISE); Hillyer, Matthew; Lima, Isabel; Nam, Sunghyun

Submitted to: ACS Applied Bio Materials
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/23/2024
Publication Date: 2/20/2024
Citation: Easson, M.W., Jordan, J.H., Prevost, N.T., Edwards, J.V., Dupre, R.A., Hillyer, M.B., Lima, I.M., Nam, S. 2024. Assessment of Cellulose Nanofiber-Based Elastase Biosensors to Inflammatory Disease as a Function of Spacer Length and Fluorescence Response. ACS Applied Bio Materials. https://doi.org/10.1021/acsabm.3c00885.
DOI: https://doi.org/10.1021/acsabm.3c00885

Interpretive Summary: Inflammatory disease biomarker detection has become a high priority in point-of-care diagnostic research in relation to chronic wounds with a variety of sensor-based designs becoming available. Herein, two primary aspects of biosensor design are examined: 1) assessment of the cellulose nanofiber (CNF) matrix derived from ginning by-products as the sensor transducer surface; 2) assessment of the relation of spacer length between the CNF cellulose backbone and peptide fluorophore as a function of sensor activity for porcine pancreatic and human neutrophil elastases. X-ray crystallography, specific surface area and pore size analyses confirmed the suitability of CNF as a matrix for wound care diagnostics. Based upon the normalized degree of substitution, a pegylated-linker connecting CNF transducer substrate to peptide fluorophore showed the greatest fluorescence response, compared to short and long-chain alkylated linkers.

Technical Abstract: Six fluorophore biosensors were each synthesized in three steps by combining a cellulose nanofiber substrate, a hydrophobic or hydrophilic linker and a tripeptide-coumarin synthetic motif. Intermediates and final products were confirmed by Raman and LC/MS spectroscopy. Results showed that well-solvated, hydrophilic biosensors containing pegylated units in their respective linkers interacted better with PPE and HNE elastases and released higher concentrations of AMC fluorophore than poorly solvated aliphatic biosensors when the fluorescence response was normalized for degree of substitution. Increasing aliphatic linker length contributed to poorer fluorescence response in both elastase assays. Fits of the fluorescence response and the calculated concentration of liberated AMC fluorophore analyte gave approximately equal values for the first-order rate expression. Cellulose nanofibers offer a promising transducer substrate for appending point-of-care diagnostic biosensor molecules. X-ray crystallography, specific surface area and pore size analyses confirm that CNFs are a highly crystalline medium with considerable surface area and pore size volume that enable opportunities for biosensor application.