13C NMR characterization of hydrated 13C labeled Bombyx mori silk fibroin sponges prepared using glycerin, poly(ethylene glycol diglycidyl ether) and poly(ethylene glycol) as porogens

There is a need to prepare softer and highly flexible Bombyx mori silk fibroin (SF) sponges for the development of biomaterials that are biodegradable and with stiffness that matches sponges and soft tissues. In this paper, we prepared SF sponges using glycerin (Glyc), poly(ethylene) glycol diglycidyl ether (PGDE) and poly(ethylene) glycol (PEG) as porogens. The detailed characterization of the hydrated SF sponges was done using three C-13 solid state NMR techniques, viz., C-13 refocused insensitive nuclei enhanced by polarization transfer (r-INEPT) NMR, C-13 cross polarization/magic angle spinning (CP/MAS) NMR, and C-13 dipolar decoupled-magic angle spinning (DD/MAS) NMR. These three NMR methods gave respective information on fast motion, slow motion, and both fast and slow motions for the local structure and dynamics of the hydrated SF sponges. There was no significant difference in the r-INEPT spectra of the three hydrated SF sponges. On the other hand, there were significant differences among the C-13 CP/MAS NMR spectra of the three sponges. The fractions of two kinds of beta-sheet structure, two kinds of random coil conformations with mobile and immobile motions, and the Silk I* (type II beta-turn) conformation were determined for the Ser residues in the C-13 DD/MAS NMR spectra. Similarly, the fractions of several conformations were also determined for Tyr, Ala and Gly residues in SF, which showed significant differences among the three hydrated sponges. The relationship between the local structure of these hydrated SF sponges and their mechanical properties was also briefly discussed.