Spinlab hyperpolarizer

Dynamic nuclear polarization was performed with a 5.0-T SpinLab Hyperpolarizer (GE Healthcare). A 2.0 M solution of 3 dissolved in water containing 15 mM OX063 trityl radical (Oxford Instruments) was polarized in a 5.0-T SpinLab Hyperpolarizer for 3 hours. The hyperpolarized substrate was ejected from the polarizer via dissolution with a large excess of superheated D₂O to a final concentration of 25 mM with a resultant pH of 7.2. Polarization quantification was carried immediately after dissolution. One milliliter of the hyperpolarized dissolution was added to an NMR tube and transferred to a 1-T Spinsolve ¹³C NMR spectrometer (Magritek, NZ). NMR spectra were acquired every 3 s with a 5° excitation over 3 min. Apparent relaxation time (T₁) was fitted to a mono-exponential curve and corrected for flip angle. Thermal polarization was determined from the average spectrum of 1024 scans acquired with 90° flip angle every 10 s. Final polarization values after correcting for flip angles and the apparent T₁ were calculated to be 5 to 10% for all HP fructose dissolutions. Substrate concentration was measured by ¹³C NMR at 14.1 T in the presence of 1 mM Gd-DOTA and 15 mM [¹³C]-acetate standard. The integral of the C₂ carbon of fructose was compared to that of the acetate standard to calculate concentrations.