Dd2 600 nmr spectrometer

Wood
delignification and IL-facilitated partial dissolution were verified
with a Thermo Scientific Nicolet iS50 FTIR spectrometer with an attenuated
total reflectance (ATR) diamond (Thermo Scientific, USA). All spectra
were collected in the absorption mode in the 400–4000 cm^–1 wavelength range from 32 scans with a resolution
of 4 cm^–1. The ¹³C cross polarization
(CP) magic angle spinning (MAS) NMR measurements were performed using
an Agilent DD2 600 NMR spectrometer with a magnetic flux density of
14.1 T, equipped with a 3.2 mm T3 MAS NMR probe operating in a double-resonance
mode. Samples were packed in ZrO₂ rotors, and the MAS rate
in experiments was set to 10 kHz. A total of 14,000 scans were accumulated
using a 1.3 ms contact time and a 5.0 s delay between successive scans.
Protons were decoupled during acquisition using SPINAL-64 proton decoupling
with a field strength of 80 kHz. 90° pulse durations and Hartmann–Hahn
match for CP were calibrated using a-glycine. The spectra were processed
using TopSpin 3.5 software. The changes in the crystallinity of the
wood after delignification and partial dissolution with the IL were
estimated from signal areas of the cellulose C₄ carbon
signals originating from crystalline (89.0–84.4 ppm) and noncrystalline
(84.4–77.3 ppm) regions. Signal areas were determined by integration.

In a 500 μL reaction mixture, 50 mM potassium phosphate pH 8.0, 10 mM MgCl₂, 2 mM L-arginine, 4 mM S-(5′-Adenosyl)-L-methionine (SAM), and 5 μM of SznE were mixed and incubated overnight at room temperature. Negative control assays omitting SAM, L-arginine, or SznE in the reaction were also performed. The reaction mixtures were flash frozen with liquid N₂ and lyophilized. The residues were resuspended in D₂O (Cambridge Isotope Inc.) and analyzed with proton nuclear magnetic resonance (¹H NMR) spectroscopy using an Agilent DD2–600 NMR spectrometer (600 MHz). Chemical shifts (δ) are reported in parts per million (ppm) downfield from tetramethylsilane using the solvent resonance as an internal standard for ¹H (D₂O = 4.79 ppm). This experiment was performed in triplicate.