Dynamic nuclear polarization-enhanced, double-quantum filtered 13C-13C dipolar correlation spectroscopy of natural 13C abundant bone- tissue biomaterial

Here, we describe a method for obtaining a dynamic nuclear polarization (DNP)-enhanced double quantum filtered (DQF) two-dimensional (2D) dipolar C-13 -C-13 correlation spectra of bone-tissue material at natural 13C abundance. DNP-enhanced DQF 2D dipolar( 13)C -C-13 spectra were obtained using a few different mixing times of the dipolar-assisted rotational resonance (DARR) scheme and these spectra were compared to a conventional 2D through-space double-quantum (DQ)-single-quantum (SQ) correlation spectrum. While this scheme can only be used for an assignment purpose to reveal the carbon-carbon connectivity within a residue, the DQF C-13 -C-13 dipolar correlation scheme introduced here can be used to obtain longer distance carbon-carbon constraints. A DQF pulse block is placed before the DARR mixing scheme for removing dominant C-13 single-quantum (SQ) signals because these SQ C-13 signals are overwhelmingly large compared to those C-13 -C-13 dipolar cross-peaks generated and therefore saturate the dynamic range of the NMR detection. This approach exhibits strong enough 2D cross-peaks in a dipolar( 13)C -C-13 correlation spectrum and potentially provides pairwise C-13 -C-13 dipolar constraints because the dipolar truncation effect as well as multi-step signal propagations involving a spin cluster that contains more than two spins can be ignored probabilistically. To obtain fast signal averaging, AsymPolPOK was used to provide a short 1H DNP signal build-up time (1.3 s) and to expedite our MAS DNP NMR acquisitions while still maintaining a satisfactory DNP enhancement factor (epsilon= 50). Under long DARR mixing, a t1-noise-like artifact was observed at a site that possesses a large chemical shift anisotropy (CSA) and a few different strategies to address this problem were discussed.(c) 2022 Elsevier Inc. All rights reserved.