Quinn A. Best

The Feasibility of Formation and Kinetics of NMR Signal Amplification by Reversible Exchange (SABRE) at High Magnetic Field (9.4 T)

1H NMR signal amplification by reversible exchange (SABRE) was observed for pyridine and pyridine-d5 at 9.4 T, a field that is orders of magnitude higher than what is typically utilized to achieve the conventional low-field SABRE effect. In addition to emissive peaks for the hydrogen spins at the ortho positions of the pyridine substrate (both free and bound to the metal center), absorptive signals are observed from hyperpolarized orthohydrogen and Ir-complex dihydride. Real-time kinetics studies show that the polarization build-up rates for these three species are in close agreement with their respective 1H T1 relaxation rates at 9.4 T. The results suggest that the mechanism of the substrate polarization involves cross-relaxation with hyperpolarized species in a manner similar to the spin-polarization induced nuclear Overhauser effect. Experiments utilizing pyridine-d5 as the substrate exhibited larger enhancements as well as partial H/D exchange for the hydrogen atom in the ortho position of pyridine and concomitant formation of HD molecules. While the mechanism of polarization enhancement does not explicitly require chemical exchange of hydrogen atoms of parahydrogen and the substrate, the partial chemical modification of the substrate via hydrogen exchange means that SABRE under these conditions cannot rigorously be referred to as a non-hydrogenative parahydrogen induced polarization process.

In Situ and Ex Situ Low-Field NMR Spectroscopy and MRI Endowed by SABRE Hyperpolarization†

By using 5.75 and 47.5 mT nuclear magnetic resonance (NMR) spectroscopy, up to 10(5)-fold sensitivity enhancement through signal amplification by reversible exchange (SABRE) was enabled, and subsecond temporal resolution was used to monitor an exchange reaction that resulted in the buildup and decay of hyperpolarized species after parahydrogen bubbling. We demonstrated the high-resolution low-field proton magnetic resonance imaging (MRI) of pyridine in a 47.5 mT magnetic field endowed by SABRE. Molecular imaging (i.e. imaging of dilute hyperpolarized substances rather than the bulk medium) was conducted in two regimes: in situ real-time MRI of the reaction mixture (in which pyridine was hyperpolarized), and ex situ MRI (in which hyperpolarization decays) of the liquid hyperpolarized product. Low-field (milli-Tesla range, e.g. 5.75 and 47.5 mT used in this study) parahydrogen-enhanced NMR and MRI, which are free from the limitations of high-field magnetic resonance (including susceptibility-induced gradients of the static magnetic field at phase interfaces), potentially enables new imaging applications as well as differentiation of hyperpolarized chemical species on demand by exploiting spin manipulations with static and alternating magnetic fields.

High-Resolution Low-Field Molecular Magnetic Resonance Imaging of Hyperpolarized Liquids

We demonstrate the feasibility of microscale molecular imaging using hyperpolarized proton and carbon-13 MRI contrast media and low-field (47.5 mT) preclinical scale (38 mm i.d.) 2D magnetic resonance imaging (MRI). Hyperpolarized proton images with 94 × 94 μm2 spatial resolution and hyperpolarized carbon-13 images with 250 × 250 μm2 in-plane spatial resolution were recorded in 4–8 s (largely limited by the electronics response), surpassing the in-plane spatial resolution (i.e., pixel size) achievable with micro-positron emission tomography (PET). These hyperpolarized proton and 13C images were recorded using large imaging matrices of up to 256 × 256 pixels and relatively large fields of view of up to 6.4 × 6.4 cm2. 13C images were recorded using hyperpolarized 1-13C-succinate-d2 (30 mM in water, %P13C = 25.8 ± 5.1% (when produced) and %P13C = 14.2 ± 0.7% (when imaged), T1 = 74 ± 3 s), and proton images were recorded using 1H hyperpolarized pyridine (100 mM in methanol-d4, %PH = 0.1 ± 0.02% (when imaged), T1 = 11 ± 0.1 s). Both contrast agents were hyperpolarized using parahydrogen (>90% para-fraction) in an automated 5.75 mT parahydrogen induced polarization (PHIP) hyperpolarizer. A magnetized path was demonstrated for successful transportation of a 13C hyperpolarized contrast agent (1-13C-succinate-d2, sensitive to fast depolarization when at the Earth’s magnetic field) from the PHIP polarizer to the 47.5 mT low-field MRI. While future polarizing and low-field MRI hardware and imaging sequence developments can further improve the low-field detection sensitivity, the current results demonstrate that microscale molecular imaging in vivo is already feasible at low (<50 mT) fields and potentially at low (∼1 mM) metabolite concentrations.

Anilinomethylrhodamines: pH sensitive probes with tunable photophysical properties by substituent effect.

A series of pH dependent rhodamine analogues possessing an anilino-methyl moiety was developed and shown to exhibit a unique photophysical response to pH. These anilinomethylrhodamines (AnMR) maintain a colorless, nonfluorescent spirocyclic structure at high pH. The spirocyclic structures open in mildly acidic conditions and are weakly fluorescent; however, at very low pH, the fluorescence is greatly enhanced. The equilibrium constants of these processes show a linear response to substituent effects, which was demonstrated by the Hammett equation.

Environmentally Robust Rhodamine Reporters for Probe-based Cellular Detection of the Cancer-linked Oxidoreductase hNQO1

UNLABELLED We successfully synthesized a fluorescent probe capable of detecting the cancer-associated NAD(P)H quinoneoxidoreductase isozyme-1 within human cells, based on results from an investigation of the stability of various rhodamines and seminaphthorhodamines toward the biological reductant NADH, present at ∼100-200 μM within cells. While rhodamines are generally known for their chemical stability, we observe that NADH causes significant and sometimes rapid modification of numerous rhodamine analogues, including those oftentimes used in imaging applications. Results from mechanistic studies lead us to rule out a radical-based reduction pathway, suggesting rhodamine reduction by NADH proceeds by a hydride transfer process to yield the reduced leuco form of the rhodamine and oxidized NAD(+). A relationship between the structural features of the rhodamines and their reactivity with NADH is observed. Rhodamines with increased alkylation on the N3- and N6-nitrogens, as well as the xanthene core, react the least with NADH; whereas, nonalkylated variants or analogues with electron-withdrawing substituents have the fastest rates of reaction. These outcomes allowed us to judiciously construct a seminaphthorhodamine-based, turn-on fluorescent probe that is capable of selectively detecting the cancer-associated, NADH-dependent enzyme NAD(P)H quinoneoxidoreductase isozyme-1 in human cancer cells, without the issue of NADH-induced deactivation of the seminaphthorhodamine reporter.