Toward Next-Generation Molecular Imaging with a Clinical Low-Field (0.064 T) Point-of-Care MRI Scanner.

Low-field (LF) MRI promises soft-tissue imaging without the expensive, immobile magnets of clinical scanners but generally suffers from limited detection sensitivity and contrast. The sensitivity boost provided by hyperpolarization can thus be highly synergistic with LF MRI. Initial efforts to integrate a continuous-bubbling SABRE (signal amplification by reversible exchange) hyperpolarization setup with a portable, point-of-care 64 mT clinical MRI scanner are reported. Results from 1H SABRE MRI of pyrazine and nicotinamide are compared with those of benchtop NMR spectroscopy. Comparison with MRI signals from samples with known H2O/D2O ratios allowed quantification of the SABRE enhancements of imaged samples with various substrate concentrations (down to 3 mM). Respective limits of detection and quantification of 3.3 and 10.1 mM were determined with pyrazine 1H polarization (PH) enhancements of ∼1900 (PH ∼0.04%), supporting ongoing and envisioned efforts to realize SABRE-enabled MRI-based molecular imaging.

Single Nucleotide Polymorphism in Cell Adhesion Molecule L1 Affects Learning and Memory in a Mouse Model of Traumatic Brain Injury.

The L1 cell adhesion molecule (L1) has demonstrated a range of beneficial effects in animal models of spinal cord injury, neurodegenerative disease, and ischemia; however, the role of L1 in TBI has not been fully examined. Mutations in the L1 gene affecting the extracellular domain of this type 1 transmembrane glycoprotein have been identified in patients with L1 syndrome. These patients suffer from hydrocephalus, MASA (mental retardation, adducted thumbs, shuffling gait, aphasia) symptoms, and corpus callosum agenesis. Clinicians have observed that recovery post-traumatic brain injury (TBI) varies among the population. This variability may be explained by the genetic differences present in the general population. In this study, we utilized a novel mouse model of L1 syndrome with a mutation at aspartic acid position 201 in the extracellular domain of L1 (L1-201). We assessed the impact of this specific single nucleotide polymorphism (SNP) localized to the X-chromosome L1 gene on recovery outcomes following TBI by comparing the L1-201 mouse mutants with their wild-type littermates. We demonstrate that male L1-201 mice exhibit significantly worse learning and memory outcomes in the Morris water maze after lateral fluid percussion (LFP) injury compared to male wild-type mice and a trend to worse motor function on the rotarod. However, no significant changes were observed in markers for inflammatory responses or apoptosis after TBI.

Toward Ultra-high-quality-factor Wireless Masing Magnetic Resonance Sensing.

It has been recently shown that a bolus of hyperpolarized nuclear spins can yield stimulated emission signals similar in nature to that of maser, potentially enabling new ways of sensing of hyperpolarized contrast media, including most notably [1-13C]pyruvate that is under evaluation in over 50 clinical trials for metabolic imaging of cancer. The stimulated NMR signal emissions lasting for minutes do not require radio-frequency excitation, offering unprecedented advantages compared to conventional MR sensing. However, creating nuclear spin maser emission is challenging in practice due to stringent fundamental requirements, making practical in vivo applications hardly possible using conventional passive MR detectors. Here, we demonstrate the utility of a wireless NMR maser detector, the quality factor of which was enhanced 22-fold (to 1,670) via parametric pumping. This active-feedback technique breaks the intrinsic fundamental limit of NMR detector circuit quality factor. We show the use of parametric pumping to reduce the threshold requirement for inducing nuclear spin masing at 300 MHz resonance frequency in preclinical MRI scanner. Indeed, stimulated emission from hyperpolarized protons was obtained under highly unfavorable conditions of low magnetic field homogeneity (T2* of 3 ms). Greater gains of the quality factor of MR detector (up to 1 million) were demonstrated.

13C Radiofrequency Amplification by Stimulated Emission of Radiation Threshold Sensing of Chemical Reactions.

Conventional nuclear magnetic resonance (NMR) enables detection of chemicals and their transformations by exciting nuclear spin ensembles with a radio-frequency pulse followed by detection of the precessing spins at their characteristic frequencies. The detected frequencies report on chemical reactions in real time and the signal amplitudes scale with concentrations of products and reactants. Here, we employ Radiofrequency Amplification by Stimulated Emission of Radiation (RASER), a quantum phenomenon producing coherent emission of 13C signals, to detect chemical transformations. The 13C signals are emitted by the negatively hyperpolarized biomolecules without external radio frequency pulses and without any background signal from other, nonhyperpolarized spins in the ensemble. Here, we studied the hydrolysis of hyperpolarized ethyl-[1-13C]acetate to hyperpolarized [1-13C]acetate, which was analyzed as a model system by conventional NMR and 13C RASER. The chemical transformation of 13C RASER-active species leads to complete and abrupt disappearance of reactant signals and delayed, abrupt reappearance of a frequency-shifted RASER signal without destroying 13C polarization. The experimentally observed "quantum" RASER threshold is supported by simulations.

Parahydrogen in Reversible Exchange Induces Long-Lived 15N Hyperpolarization of Anticancer Drugs Anastrozole and Letrozole.

Hyperpolarization modalities overcome the sensitivity limitations of NMR and unlock new applications. Signal amplification by reversible exchange (SABRE) is a particularly cheap, quick, and robust hyperpolarization modality. Here, we employ SABRE for simultaneous chemical exchange of parahydrogen and nitrile-containing anticancer drugs (letrozole or anastrozole) to enhance 15N polarization. Distinct substrates require unique optimal parameter sets, including temperature, magnetic field, or a shaped magnetic field profile. The fine tuning of these parameters for individual substrates is demonstrated here to maximize 15N polarization. After optimization, including the usage of pulsed µT fields, the 15N nuclei on common anticancer drugs, letrozole and anastrozole, can be polarized within 1-2 min. The hyperpolarization can exceed 10%, corresponding to 15N signal enhancement of over 280,000-fold at a clinically relevant magnetic field of 1 T. This sensitivity gain enables polarization studies at naturally abundant 15N enrichment level (0.4%). Moreover, the nitrile 15N sites enable long-lasting polarization storage with [15N]T1 over 9 min, enabling signal detection from a single hyperpolarization cycle for over 30 min.

Effects of L1 adhesion molecule agonistic mimetics on signal transduction in neuronal functions.

The L1 cell adhesion molecule plays an essential role in neural development and repair. It is not only a 'lock and key' recognition molecule, but an important signal transducer that stimulates regenerative-beneficial cellular functions such as neurite outgrowth, neuronal cell migration, survival, myelination, and synapse formation. Triggering L1 functions after neurotrauma improves functional recovery. In addition, loss-of-function mutations in the L1 gene lead to the L1 syndrome, a rare, X-linked neurodevelopmental disorder with an incidence of approximately 1:30,000 in newborn males. To use L1 for beneficial functions, we screened small compound libraries for L1 agonistic mimetics that trigger L1 functions and improve conditions in animal models of neurotrauma and the L1 syndrome. To understand the mechanisms underlying these functions, it is important to gain a better understanding of L1-dependent cellular signaling that is triggered by the L1 agonistic mimetics. We tested the cell signaling features of L1 agonistic mimetics that contribute to neurite outgrowth and neuronal migration. Our findings indicates that L1 agonistic mimetics trigger the same cell signaling pathways underlying neurite outgrowth, but only the L1 mimetics tacrine, polydatin, trimebutine and honokiol trigger neuronal migration. In contrast, the mimetics crotamiton and duloxetine did not affect neuronal migration, thus limiting their use in increasing neuronal migration, leaving open the question of whether this is a desired or not desired feature in the adult.

Spin dynamics of [1,2-13C2]pyruvate hyperpolarization by parahydrogen in reversible exchange at micro Tesla fields.

Hyperpolarization of 13C-pyruvate via Signal Amplificaton By Reversibble Exchange (SABRE) is an important recent discovery because of both the relative simplicity of hyperpolarization and the central biological relevance of pyruvate as a biomolecular probe for in vitro or in vivo studies. Here, we analyze the [1,2-13C2]pyruvate-SABRE spin system and its field dependence theoretically and experimentally. We provide first-principles analysis of the governing 4-spin dihydride-13C2 Hamiltonian and numerical spin dynamics simulations of the 7-spin dihydride-13C2-CH3 system. The analytical and the numerical results are compared to matching systematic experiments. With these methods we unravel the observed spin state mixing of singlet states and triplet states at microTesla fields and we also analyze the dynamics during transfer from micro-Tesla field to high field for detection to understand the resulting spectra from the [1,2-13C2]pyruvate-SABRE system.

SABRE Hyperpolarization with up to 200 bar Parahydrogen in Standard and Quickly Removable Solvents.

Parahydrogen (p-H2)-based techniques are known to drastically enhance NMR signals but are usually limited by p-H2 supply. This work reports p-H2-based SABRE hyperpolarization at p-H2 pressures of hundreds of bar, far beyond the typical ten bar currently reported in the literature. A recently designed high-pressure setup was utilized to compress p-H2 gas up to 200 bar. The measurements were conducted using a sapphire high-pressure NMR tube and a 43 MHz benchtop NMR spectrometer. In standard methanol solutions, it could be shown that the signal intensities increased with pressure until they eventually reached a plateau. A polarization of about 2%, equal to a molar polarization of 1.2 mmol L-1, could be achieved for the sample with the highest substrate concentration. While the signal plateaued, the H2 solubility increased linearly with pressure from 1 to 200 bar, indicating that p-H2 availability is not the limiting factor in signal enhancement beyond a certain pressure, depending on sample composition. Furthermore, the possibility of using liquefied ethane and compressed CO2 as removable solvents for hyperpolarization was demonstrated. The use of high pressures together with quickly removable organic/non-organic solvents represents an important breakthrough in the field of hyperpolarization, advancing SABRE as a promising tool for materials science, biophysics, and molecular imaging.

Small Organic Compounds Mimicking the Effector Domain of Myristoylated Alanine-Rich C-Kinase Substrate Stimulate Female-Specific Neurite Outgrowth.

Myristoylated alanine-rich C-kinase substrate (MARCKS) is a critical member of a signaling cascade that influences disease-relevant neural functions such as neural growth and plasticity. The effector domain (ED) of MARCKS interacts with the extracellular glycan polysialic acid (PSA) through the cell membrane to stimulate neurite outgrowth in cell culture. We have shown that a synthetic ED peptide improves functional recovery after spinal cord injury in female but not male mice. However, peptides themselves are unstable in therapeutic applications, so we investigated more pharmacologically relevant small organic compounds that mimic the ED peptide to maximize therapeutic potential. Using competition ELISAs, we screened small organic compound libraries to identify molecules that structurally and functionally mimic the ED peptide of MARCKS. Since we had shown sex-specific effects of MARCKS on spinal cord injury recovery, we assayed neuronal viability as well as neurite outgrowth from cultured cerebellar granule cells of female and male mice separately. We found that epigallocatechin, amiodarone, sertraline, tegaserod, and nonyloxytryptamine bind to a monoclonal antibody against the ED peptide, and compounds stimulate neurite outgrowth in cultured cerebellar granule cells of female mice only. Therefore, a search for compounds that act in males appears warranted.

Parahydrogen-Induced Carbon-13 Radiofrequency Amplification by Stimulated Emission of Radiation.

The feasibility of Carbon-13 Radiofrequency (RF) Amplification by Stimulated Emission of Radiation (C-13 RASER) is demonstrated on a bolus of liquid hyperpolarized ethyl [1-13 C]acetate. Hyperpolarized ethyl [1-13 C]acetate was prepared via pairwise addition of parahydrogen to vinyl [1-13 C]acetate and polarization transfer from nascent parahydrogen-derived protons to the carbon-13 nucleus via magnetic field cycling yielding C-13 nuclear spin polarization of approximately 6 %. RASER signals were detected from samples with concentration ranging from 0.12 to 1 M concentration using a non-cryogenic 1.4T NMR spectrometer equipped with a radio-frequency detection coil with a quality factor (Q) of 32 without any modifications. C-13 RASER signals were observed for several minutes on a single bolus of hyperpolarized substrate to achieve 21 mHz NMR linewidths. The feasibility of creating long-lasting C-13 RASER on biomolecular carriers opens a wide range of new opportunities for the rapidly expanding field of C-13 magnetic resonance hyperpolarization.

Temperature Cycling Enables Efficient 13C SABRE-SHEATH Hyperpolarization and Imaging of [1-13C]-Pyruvate.

Molecular metabolic imaging in humans is dominated by positron emission tomography (PET). An emerging nonionizing alternative is hyperpolarized MRI of 13C-pyruvate, which is innocuous and has a central role in metabolism. However, similar to PET, hyperpolarized MRI with dissolution dynamic nuclear polarization (d-DNP) is complex costly, and requires significant infrastructure. In contrast, Signal Amplification By Reversible Exchange (SABRE) is a fast, cheap, and scalable hyperpolarization technique. SABRE in SHield Enables Alignment Transfer to Heteronuclei (SABRE-SHEATH) can transfer polarization from parahydrogen to 13C in pyruvate; however, polarization levels remained low relative to d-DNP (1.7% with SABRE-SHEATH versus ≈60% with DNP). Here we introduce a temperature cycling method for SABRE-SHEATH that enables >10% polarization on [1-13C]-pyruvate, sufficient for successful in vivo experiments. First, at lower temperatures, ≈20% polarization is accumulated on SABRE catalyst-bound pyruvate, which is released into free pyruvate at elevated temperatures. A kinetic model of differential equations is developed that explains this effect and characterizes critical relaxation and buildup parameters. With the large polarization, we demonstrate the first 13C pyruvate images with a cryogen-free MRI system operated at 1.5 T, illustrating that inexpensive hyperpolarization methods can be combined with low-cost MRI systems to obtain a broadly available, yet highly sensitive metabolic imaging platform.

Rapid 13C Hyperpolarization of the TCA Cycle Intermediate α-Ketoglutarate via SABRE-SHEATH.

α-Ketoglutarate is a key biomolecule involved in a number of metabolic pathways─most notably the TCA cycle. Abnormal α-ketoglutarate metabolism has also been linked with cancer. Here, isotopic labeling was employed to synthesize [1-13C,5-12C,D4]α-ketoglutarate with the future goal of utilizing its [1-13C]-hyperpolarized state for real-time metabolic imaging of α-ketoglutarate analytes and its downstream metabolites in vivo. The signal amplification by reversible exchange in shield enables alignment transfer to heteronuclei (SABRE-SHEATH) hyperpolarization technique was used to create 9.7% [1-13C] polarization in 1 minute in this isotopologue. The efficient 13C hyperpolarization, which utilizes parahydrogen as the source of nuclear spin order, is also supported by favorable relaxation dynamics at 0.4 µT field (the optimal polarization transfer field): the exponential 13C polarization buildup constant Tb is 11.0 ± 0.4 s whereas the 13C polarization decay constant T1 is 18.5 ± 0.7 s. An even higher 13C polarization value of 17.3% was achieved using natural-abundance α-ketoglutarate disodium salt, with overall similar relaxation dynamics at 0.4 µT field, indicating that substrate deuteration leads only to a slight increase (∼1.2-fold) in the relaxation rates for 13C nuclei separated by three chemical bonds. Instead, the gain in polarization (natural abundance versus [1-13C]-labeled) is rationalized through the smaller heat capacity of the "spin bath" comprising available 13C spins that must be hyperpolarized by the same number of parahydrogen present in each sample, in line with previous 15N SABRE-SHEATH studies. Remarkably, the C-2 carbon was not hyperpolarized in both α-ketoglutarate isotopologues studied; this observation is in sharp contrast with previously reported SABRE-SHEATH pyruvate studies, indicating that the catalyst-binding dynamics of C-2 in α-ketoglutarate differ from that in pyruvate. We also demonstrate that 13C spectroscopic characterization of α-ketoglutarate and pyruvate analytes can be performed at natural 13C abundance with an estimated detection limit of 80 micromolar concentration × *%P13C. All in all, the fundamental studies reported here enable a wide range of research communities with a new hyperpolarized contrast agent potentially useful for metabolic imaging of brain function, cancer, and other metabolically challenging diseases.

Order-Unity 13 C Nuclear Polarization of [1-13 C]Pyruvate in Seconds and the Interplay of Water and SABRE Enhancement.

Signal Amplification By Reversible Exchange in SHield Enabled Alignment Transfer (SABRE-SHEATH) is investigated to achieve rapid hyperpolarization of 13 C1 spins of [1-13 C]pyruvate, using parahydrogen as the source of nuclear spin order. Pyruvate exchange with an iridium polarization transfer complex can be modulated via a sensitive interplay between temperature and co-ligation of DMSO and H2 O. Order-unity 13 C (>50 %) polarization of catalyst-bound [1-13 C]pyruvate is achieved in less than 30 s by restricting the chemical exchange of [1-13 C]pyruvate at lower temperatures. On the catalyst bound pyruvate, 39 % polarization is measured using a 1.4 T NMR spectrometer, and extrapolated to >50 % at the end of build-up in situ. The highest measured polarization of a 30-mM pyruvate sample, including free and bound pyruvate is 13 % when using 20 mM DMSO and 0.5 M water in CD3 OD. Efficient 13 C polarization is also enabled by favorable relaxation dynamics in sub-microtesla magnetic fields, as indicated by fast polarization buildup rates compared to the T1 spin-relaxation rates (e. g., ∼0.2 s-1 versus ∼0.1 s-1 , respectively, for a 6 mM catalyst-[1-13 C]pyruvate sample). Finally, the catalyst-bound hyperpolarized [1-13 C]pyruvate can be released rapidly by cycling the temperature and/or by optimizing the amount of water, paving the way to future biomedical applications of hyperpolarized [1-13 C]pyruvate produced via comparatively fast and simple SABRE-SHEATH-based approaches.

Interplay of Near-Zero-Field Dephasing, Rephasing, and Relaxation Dynamics and [1-13C]Pyruvate Polarization Transfer Efficiency in Pulsed SABRE-SHEATH.

Hyperpolarized [1-13C]pyruvate is a revolutionary molecular probe enabling ultrafast metabolic MRI scans in 1 min. This technology is now under evaluation in over 30 clinical trials, which employ dissolution Dynamic Nuclear Polarization (d-DNP) to prepare a batch of the contrast agent; however, d-DNP technology is slow and expensive. The emerging SABRE-SHEATH hyperpolarization technique enables fast (under 1 min) and robust production of hyperpolarized [1-13C]pyruvate via simultaneous chemical exchange of parahydrogen and pyruvate on IrIMes hexacoordinate complexes. Here, we study the application of microtesla pulses to investigate their effect on C-13 polarization efficiency, compared to that of conventional SABRE-SHEATH employing a static field (∼0.4 µT), to provide the matching conditions of polarization transfer from parahydrogen-derived hydrides to the 13C-1 nucleus. Our results demonstrate that using square-microtesla pulses with optimized parameters can produce 13C-1 polarization levels of up to 14.8% (when detected, averaging over all resonances), corresponding to signal enhancement by over 122,000-fold at the clinically relevant field of 1.4 T. We anticipate that our results can be directly translated to other structurally similar biomolecules such as [1-13C]α-ketoglutarate and [1-13C]α-ketoisocaproate. Moreover, other more advanced pulse shapes can potentially further boost heteronuclear polarization attainable via pulsed SABRE-SHEATH.

Interactions between the Polysialylated Neural Cell Adhesion Molecule and the Transient Receptor Potential Canonical Channels 1, 4, and 5 Induce Entry of Ca2+ into Neurons.

The neural cell adhesion molecule (NCAM) plays important functional roles in the developing and mature nervous systems. Here, we show that the transient receptor potential canonical (TRPC) ion channels TRPC1, -4, and -5 not only interact with the intracellular domains of the transmembrane isoforms NCAM140 and NCAM180, but also with the glycan polysialic acid (PSA) covalently attached to the NCAM protein backbone. NCAM antibody treatment leads to the opening of TRPC1, -4, and -5 hetero- or homomers at the plasma membrane and to the influx of Ca2+ into cultured cortical neurons and CHO cells expressing NCAM, PSA, and TRPC1 and -4 or TRPC1 and -5. NCAM-stimulated Ca2+ entry was blocked by the TRPC inhibitor Pico145 or the bacterial PSA homolog colominic acid. NCAM-stimulated Ca2+ influx was detectable neither in NCAM-deficient cortical neurons nor in TRPC1/4- or TRPC1/5-expressing CHO cells that express NCAM, but not PSA. NCAM-induced neurite outgrowth was reduced by TRPC inhibitors and a function-blocking TRPC1 antibody. A characteristic signaling feature was that extracellular signal-regulated kinase 1/2 phosphorylation was also reduced by TRPC inhibitors. Our findings indicate that the interaction of NCAM with TRPC1, -4, and -5 contributes to the NCAM-stimulated and PSA-dependent Ca2+ entry into neurons thereby influencing essential neural functions.

Myristoylated alanine-rich C-kinase substrate effector domain peptide improves sex-specific recovery and axonal regrowth after spinal cord injury.

Myristoylated alanine-rich C-kinase substrate (MARCKS) is an intracellular receptor for polysialic acid. MARCKS supports development, synaptic plasticity, and regeneration after injury. MARCKS binds with its functionally essential effector domain (ED) to polysialic acid. A 25-mer peptide comprising the ED of MARCKS stimulates neuritogenesis of primary hippocampal neurons after addition to the culture. This motivated us to investigate whether ED peptide has similar effects in spinal cord injury. ED peptide supported recovery and regrowth of monoaminergic axons in female, but not in male mice. Sex-specific differences in response to ED peptide application also occurred in cultured neurons. In female but not male neurons, the ED peptide enhanced neurite outgrowth that could be suppressed by inhibitors of the estrogen receptors α and ß, fibroblast growth factor receptor-1, protein kinase C, and matrix metalloproteinase 2. In addition, we observed female-specific elevation of phosphorylated MARCKS levels after ED peptide treatment. In male neurons, the ED peptide enhanced neuritogenesis in the presence of an androgen receptor inhibitor to the extent seen in ED peptide-treated female neurons. However, inhibition of androgen receptor did not lead to increased phosphorylation of MARCKS. These results provide insights into the functions of a novel compound contributing to gender-dependent regeneration.

Density Functional Theory Study of Reaction Equilibria in Signal Amplification by Reversible Exchange.

The front cover artwork is provided by the groups of Prof. Thomas Theis (North Carolina State University) Prof. Volker Blum (Duke University). The image shows the reaction network of Signal Amplification by Reversible Exchange (SABRE), elucidated by density functional theory (DFT). Read the full text of the Review at 10.1002/cphc.202100204.

Density Functional Theory Study of Reaction Equilibria in Signal Amplification by Reversible Exchange.

An in-depth theoretical analysis of key chemical equilibria in Signal Amplification by Reversible Exchange (SABRE) is provided, employing density functional theory calculations to characterize the likely reaction network. For all reactions in the network, the potential energy surface is probed to identify minimum energy pathways. Energy barriers and transition states are calculated, and harmonic transition state theory is applied to calculate exchange rates that approximate experimental values. The reaction network energy surface can be modulated by chemical potentials that account for the dependence on concentration, temperature, and partial pressure of molecular constituents (hydrogen, methanol, pyridine) supplied to the experiment under equilibrium conditions. We show that, under typical experimental conditions, the Gibbs free energies of the two key states involved in pyridine-hydrogen exchange at the common Ir-IMes catalyst system in methanol are essentially the same, i. e., nearly optimal for SABRE. We also show that a methanol-containing intermediate is plausible as a transient species in the process.

A Versatile Compact Parahydrogen Membrane Reactor.

We introduce a Spin Transfer Automated Reactor (STAR) that produces continuous parahydrogen induced polarization (PHIP), which is stable for hours to days. We use the PHIP variant called signal amplification by reversible exchange (SABRE), which is particularly well suited to produce continuous hyperpolarization. The STAR is operated in conjunction with benchtop (1.1 T) and high field (9.4 T) NMR magnets, highlighting the versatility of this system to operate with any NMR or MRI system. The STAR uses semipermeable membranes to efficiently deliver parahydrogen into solutions at nano to milli Tesla fields, which enables 1 H, 13 C, and 15 N hyperpolarization on a large range of substrates including drugs and metabolites. The unique features of the STAR are leveraged for important applications, including continuous hyperpolarization of metabolites, desirable for examining steady-state metabolism in vivo, as well as for continuous RASER signals suitable for the investigation of new physics.

Hyperpolarization of common antifungal agents with SABRE.

Signal amplification by reversible exchange (SABRE) is a robust and inexpensive hyperpolarization (HP) technique to enhance nuclear magnetic resonance (NMR) spectroscopy and magnetic resonance imaging (MRI) signals using parahydrogen (pH2 ). The substrate scope of SABRE is continually expanding. Here, we present the polarization of three antifungal drugs (voriconazole, clotrimazole, and fluconazole) and elicit the detailed HP mechanisms for 1 H and 15 N nuclei. In this exploratory work, 15 N polarization values of ~1% were achieved using 50% pH2 in solution of 3-mM catalyst and 60-mM substrate in perdeuterated methanol. All hyperpolarized 15 N sites exhibited long T1 in excess of 1 min at a clinically relevant field of 1 T. Hyperpolarizing common drugs is of interest due to their potential biomedical applications as MRI contrast agents or to enable studies on protein dynamics at physiological concentrations. We optimize the polarization with respect to temperature and the polarization transfer field (PTF) for 1 H nuclei in the millitesla regime and for 15 N nuclei in the microtesla regime, which provides detailed insights into exchange kinetics and spin evolution. This work broadens the SABRE substrate scope and provides mechanistic and kinetic insights into the HP process.