Chemical Physics and Applications of Dynamic Nuclear Polarization-enhanced Nuclear Magnetic Resonance

Nuclear magnetic resonance (NMR) spectroscopy of nuclei with low magnetic moments such as 13C spins can be quite challenging and time-consuming. Dynamic nuclear polarization (DNP) via the dissolution method greatly alleviates this sensitivity problem by enhancing the NMR signals of these insensitive nuclei by several thousand-fold. Dissolution DNP thus allows 13C NMR tracking of cellular metabolism in living cells in real-time with superb sensitivity and high specificity. Herein, the bulk of my PhD dissertation work has been devoted to the elucidation and optimization of chemical physics of DNP technology in pursuit of attaining the highest NMR signal enhancements. One finding highlighted in this dissertation is the confirmation that the solidstate 13C DNP efficiency is affected by the isotopic location of the 13C label within the target molecule. Such can be explained via the thermal mixing model of DNP. Another major work in this dissertation is the investigation of the effects of 2H enrichment of the glassing solvents on the solid-state 13C spin-lattice T1 relaxation times of hyperpolarized 13C acetate. It is reported herein that glassing solvent deuteration elongates the 13C T1 relaxation times significantly, indicative of reduced intermolecular dipolar interaction of 13C spins with 2H spins compared to coupling with 1H spins. Next, this dissertation also encompasses two studies regarding the effect upon DNP of doping samples with mixtures of two different free radicals as opposed to doping them with one type of free radical. In one of the two studies, it was determined that a mixture of the wide EPR width 4-oxo-TEMPO and narrow EPR width trityl OX063 yields interesting 13C DNP results. There appears to be competing effects when the microwave irradiation frequency was set to the negative polarization peak of trityl OX063 which coincides with the positive polarization peak of 4-oxo-TEMPO. On the other hand, a mixture of both narrow EPR widths trityl and BDPA free radicals yields an additive effect. Finally, this dissertation also details the use of 13C NMR in the characterization of 13C-labelled amino acids and their application in investigating cancer cell metabolism. 13C-labeled amino acids are potential hyperpolarized 13C NMR spectroscopy and imaging (MRI) metabolic probes for cancer because a number of metabolic pathways that involve these biomolecules are abnormal in tumors. For instance, the enzyme branched chain amino acid transferase (BCAT), which catalyzes the conversion of branched chain amino acids (BCAA) to their ketoacid counterparts or vice versa, is overexpressed in several cancers. In this project, [1- 13C] L-leucine and [1- 13C] alpha-ketoisocaproate (KIC) were used to study the aberrant BCAT metabolic activity in glioblastoma. SfXL glioblastoma cells appear to preferentially convert 13CKIC to 13C leucine rather than vice versa. Western blot experiments confirmed that BCAT expression is higher in SfXL cells than in normal astrocytes. Overall, this dissertation details the chemical tuning methods in DNP that I have unraveled in pursuit of attaining the highest 13C NMR signal enhancements. These optimized DNP signals are crucial to the success of in vivo NMR or MRI studies, particularly in probing the hyperactive metabolism of cancer.