Systems and methods for producing hyperpolarized materials

Abstract

Systems and methods are disclosed for generating highly polarized molecules with high molar polarization. The systems and methods generally reduce or suppress strong demagnetizing fields that may otherwise result from polarizing high concentrations of molecules. Because such demagnetizing fields can hinder polarization transfer, reducing such fields increases molar polarization. The systems and methods generally use novel pulse sequences that counteract the effects of the demagnetizing fields.

Description

[Technical Field]

[0001] cross reference This application claims priority to U.S. Provisional Patent Application No. 63 / 375,390, entitled "SYSTEMS AND METHODS FOR GENERATION OF HYPERPOLARIZED MATERIALS," filed September 13, 2022, and U.S. Provisional Patent Application No. 63 / 460,629, entitled "SYSTEMS AND METHODS FOR GENERATION OF HYPERPOLARIZED MATERIALS," filed April 20, 2023, each of which is incorporated by reference in its entirety for all purposes.

[0002] The disclosed embodiments relate generally to the production of hyperpolarized materials for use in nuclear magnetic resonance (NMR), magnetic resonance imaging (MRI), or similar applications. [Background technology]

[0003] Nuclear magnetic resonance (NMR) and magnetic resonance imaging (MRI) are techniques with important applications in chemistry, biology, and medical imaging. Despite their success, magnetic resonance applications require minute nuclear polarization (typically 10 s) of analytes in thermal equilibrium. -5 It is recognized that there is often a limitation due to the small amount of nuclear polarization (approximately 100 keV). This small amount of nuclear polarization can limit sensitivity compared to other analytical techniques such as mass spectrometry.

[0004] Increasing the nuclear spin polarization beyond the thermal equilibrium value can improve magnetic resonance sensitivity. Nuclear spin polarization can be increased using known techniques such as dynamic nuclear polarization (DNP), parahydrogen induced polarization (PHIP), PHIP-sidearm hydrogenation (PHIP-SAH), PHIP relayed via proton exchange (PHIP-X), and PHIP nuclear Overhauser effect systems (PHIPNOESYS). Using these techniques, the nuclear spin polarization of a material can often be increased by a factor of more than 100, and in some cases, by more than 10,000. Enhanced nuclear spin polarization can proportionally increase the NMR / MRI signal.

[0005] These techniques can achieve high polarization at moderate concentrations, making them attractive approaches for a variety of applications. For example, polarized molecules prepared via PHIP or PHIP-SAH can be used directly in NMR or MRI experiments. Alternatively, polarized molecules can be used as a source to transfer polarization to other molecules via procedures such as PHIPNOESYS and PHIP-X. However, when polarized molecules are present at high concentrations, the strong demagnetizing field resulting from the dipole field associated with the magnetization of polarized molecules in a sample can interfere with polarization transfer and limit the achievable product of polarization and concentration (i.e., molar polarization). Thus, PHIP, PHIP-SAH, PHIPNOESYS, and PHIP-X have thus far been limited in the achievable molar polarization. Summary of the Invention

[0006] According to the present disclosure, a solution can be obtained in which hyperpolarized molecules are dissolved. The hyperpolarized molecules can include at least one nucleus with a molar polarization of at least 50 millimolar (mM). Prior to obtaining the solution, a nuclear spin hyperpolarization protocol can be performed on the hyperpolarized molecules, thereby imparting molar polarization to at least one nucleus. The nuclear spin hyperpolarization procedure includes obtaining a solution containing a derivative of the hyperpolarized molecule, the derivative containing at least one unsaturated carbon-carbon double bond or unsaturated carbon-carbon triple bond and having the form R1-C=C-R2 or R1-C≡C-R2, where R1 and R2 comprise a side chain, and hydrogenating the double or triple bond with para-hydrogen to form a para-hydrogenated derivative of the hyperpolarized molecule, the para-hydrogenated derivative having the form R1-CH * -CH * -R2 or R1-CH * =CH * -R2, H * The method may include forming a parahydrogen-derived hydrogen atom (B1) attached across a double or triple bond, and applying a polarization transfer waveform to transfer nuclear spin order from at least one of the parahydrogen-derived hydrogen atoms to at least one nucleus, thereby imparting nuclear spin hyperpolarization to the at least one nucleus. The polarization transfer waveform may be configured to suppress a dipolar field associated with magnetization generated during the accumulation of nuclear spin hyperpolarization. The polarization transfer waveform may include a dipolar decoupling sequence. The polarization transfer waveform may further include a coupling driving field based on a parameter sweep. The parameter sweep may further include a transverse magnetic field (B1) sweep. For example, the polarization transfer waveform may include a B1 sweep in the Lee-Goldberg frame. The polarization transfer waveform may include a pulse sequence selected from the group consisting of polarized MREV-8, polarized BLEW-12, and polarized BR-24. Hyperpolarized molecules can be used in PHIP, PHIP-SAH, PHIPNOESYS, or PHIP-X experiments. Polarization transfer waveforms can enable molar polarization that would not otherwise be obtainable due to the accumulation of magnetization during the polarization procedure.

[0007] It is to be understood that both the foregoing general description and the following detailed description are exemplary and explanatory only and are not restrictive of embodiments of the disclosure as claimed. [Brief explanation of the drawings]

[0008] The accompanying drawings, which form a part of this specification, illustrate some embodiments and, together with the description, serve to explain the principles and features of the disclosed embodiments.

[0009] [Figure 1] 1 illustrates an exemplary method for generating high molar polarization in hyperpolarized molecules via a PHIP nuclear spin hyperpolarization protocol, according to disclosed embodiments. [Figure 2] 2 illustrates an exemplary method 200 for generating high molar polarization in hyperpolarized molecules via a PHIP-SAH nuclear spin hyperpolarization protocol, according to disclosed embodiments. [Figure 3] 10 shows exemplary numerical simulations of molar polarization versus concentration for polarized MREV-8, polarized BLEW-12, polarized BR-24, and transverse field sweep in Lee-Goldberg frame pulse sequences for (1-C,d)-dimethylmaleate, according to disclosed embodiments. [Figure 4] 1 shows exemplary numerical simulations of molar polarization versus concentration for polarized MREV-8, polarized BLEW-12, polarized BR-24, and transverse field sweep in Lee-Goldberg frame pulse sequences for (1-C)-fumaric acid, according to disclosed embodiments. [Figure 5] 10 shows exemplary numerical simulations of molar polarization versus concentration for polarized MREV-8, polarized BLEW-12, polarized BR-24, and transverse field sweep in Lee-Goldberg frame pulse sequences for (1,2-d2)-ethyl acetate, according to disclosed embodiments. [Figure 6] 1 shows an exemplary 1H molar polarization of hyperpolarized (1- 13 C,d 6 )-dimethyl maleate as a function of concentration, according to disclosed embodiments. [Figure 7]1 shows an exemplary 1H spin polarization of highly polarized (1-13C,d6)-dimethylmaleate as a function of the effective angle of Lee-Goldberg decoupling, according to disclosed embodiments. [Figure 8] 1 shows an exemplary hyperpolarized 1H spectrum of 1- 13 C-d6-dimethylmaleate acquired in a 9.41 T magnetic field after polarization transfer using a polarization MREV-8 sequence, according to disclosed embodiments. DETAILED DESCRIPTION OF THE INVENTION

[0010] Exemplary embodiments will now be described in detail and discussed with reference to the accompanying drawings. In some instances, the same reference numbers are used throughout the drawings, and the following description refers to the same or similar parts. Unless otherwise defined, technical and / or scientific terms have the meanings commonly understood by those skilled in the art. The disclosed embodiments are described in sufficient detail to enable those skilled in the art to practice the disclosed embodiments. It will be understood that other embodiments may be utilized, and changes may be made without departing from the scope of the disclosed embodiments. Accordingly, the materials, methods, and examples are illustrative only and are not intended to be necessarily limiting.

[0011] NMR and MRI can be used in a wide range of applications, including, but not limited to, determining chemical structure in synthetic intermediates, determining atomic-level structure and dynamics in proteins and nucleic acids, minimally invasive imaging of biological tissues or organisms, and even metabolic analysis of biological tissues or organisms. However, NMR and MRI can be limited in sensitivity due to the small size of nuclear magnetic moments, combined with the corresponding small polarization at thermal equilibrium. This limited sensitivity may prevent the use of NMR and MRI in some applications and can make other applications of NMR and MRI impractically time- or material-intensive.

[0012] NMR and MRI can be improved by using higher magnetic fields and optimized detection systems. However, an alternative approach is to enhance NMR and MRI by increasing the nuclear spin polarization to a level significantly higher than thermal equilibrium. Such hyperpolarization techniques can often increase NMR and MRI sensitivity by a factor significantly greater than increasing the magnetic field or using optimized detection systems.

[0013] Nuclear spin polarization can be increased using a variety of techniques, including dynamic nuclear polarization (DNP), parahydrogen-induced polarization (PHIP), PHIP-sidearm hydrolysis (PHIP-SAH), signal amplification by reversible exchange (SABRE), PHIP relayed via proton exchange (PHIP-X), PHIP nuclear Overhauser effect system (PHIPNOESYS), spin-exchange optical pumping (SEOP), optically initialized electronic triplet states (also called photoexcited triplet states, PETS), and other suitable methods. Among these techniques, parahydrogen-based methods such as PHIP, PHIP-SAH, PHIP-X, and PHIPNOESYS are particularly promising because they can be implemented with high throughput using relatively low-cost equipment.

[0014] For example, recent research in NMR and MRI has demonstrated that NMR and MRI signals associated with various biocontrast agents can be enhanced by several orders of magnitude using PHIP or PHIP-SAH. Such dramatic signal enhancement enables spectroscopic analysis of biocontrast agents as they are metabolized by various tissues at different locations within the body. Analysis of metabolic information determined by such spectroscopic imaging can enable noninvasive determination of the health of tissues within the body. For example, abnormal metabolism of biocontrast agents can indicate diseases such as cancer at several locations within the body.

[0015] In PHIP and PHIP-SAH, a derivative (e.g., precursor) of a molecule of interest is reacted with para-hydrogen to form a para-hydrogenated form of the derivative. Spin order is then transferred from the proton added via the para-hydrogenation reaction to a nucleus of interest (e.g., carbon-13 nucleus) contained within the molecule of interest. In PHIP, the para-hydrogenated form of the derivative is chemically identical to the molecule of interest and is distinguished from it only by the spin order resulting from the para-hydrogenation reaction. In PHIP-SAH, the para-hydrogenated form of the derivative is cleaved (e.g., hydrolyzed) to yield a hyperpolarized molecule of interest. In SABRE, the molecule of interest itself forms a coordination complex with a polarization transfer catalyst and para-hydrogen. Spin order is then transferred from para-hydrogen to a nucleus of interest within the molecule of interest via the coordination complex. The molecule of interest is then optionally purified and used in NMR or MRI procedures. PHIP-X and PHIPNOESYS utilize PHIP or PHIP-SAH to generate hyperpolarized material (e.g., a source compound) and transfer polarization from the source compound to a material (e.g., a target compound, target molecule, or molecule of interest) used in NMR spectroscopy. In PHIP-X, the transfer or polarization from the source compound to the target compound proceeds via proton exchange from the source compound to the target compound. Polarization can then be transferred internally within the target compound via an intramolecular Nuclear Overhauser Effect (NOE). In PHIPNOESYS, the transfer of polarization from the source compound to the target compound proceeds via an intermolecular NOE. PHIPNOESYS has been shown to increase NMR spectroscopy signals by up to approximately 2,000-fold, allowing NMR spectroscopy to be applied at significantly reduced concentrations than would otherwise be achievable.

[0016] The ultimate goal of hyperpolarization techniques such as PHIP, PHIP-SAH, PHIPNOESYS, and PHIP-X is to produce high concentrations of hyperpolarized molecules. However, when polarized molecules are present at high concentrations, strong demagnetizing fields from polarized molecules in a sample can interfere with polarization transfer and limit the achievable product of polarization and concentration (i.e., molar polarization). Thus, PHIP, PHIP-SAH, PHIPNOESYS, and PHIP-X have been limited in the molar polarization they can achieve. Therefore, there is a need for systems and methods that counteract the effects of the large demagnetizing fields generated when polarized molecules are present at high concentrations and enable the production of large molar polarizations.

[0017] As described herein, the challenges posed by the strong depolarization fields described above can be mitigated using novel pulse sequences that counteract the effects of the depolarization fields. The systems and methods described herein enable efficient polarization transfer, even at high concentrations, without any molar polarization limitations. Such novel pulse sequences offer exciting opportunities for using hyperpolarization techniques in a wide range of applications, including MRI and drug discovery, with potential in molecular imaging, materials science, and beyond.

[0018] Disclosed embodiments generate hyperpolarized molecules dissolved in a solution. The hyperpolarized molecules generally include at least one nucleus with a large molar polarization. The large molar polarization is generated by applying a polarization transfer waveform during polarization accumulation on the at least one nucleus. The polarization transfer waveform is generally configured to suppress the dipolar field associated with the magnetization generated during polarization accumulation. For example, the polarization transfer waveform may include any one or more of a dipolar decoupling sequence, such as a transverse magnetic field sweep in a Lee-Goldberg frame, a polarized MREV-8 pulse sequence, a polarized BLEW-12 pulse sequence, a polarized BR-24 pulse sequence, or the like. Following the polarization transfer waveform, the at least one nucleus may be associated with a relatively high molar polarization. The hyperpolarized molecules may then be used by an end user, such as a hospital or clinic, in an NMR or MRI experiment.

[0019] Hyperpolarization and parahydrogen As used in this disclosure, "polarization" refers to an imbalance in electronic or nuclear spin orientation. In some embodiments, polarization can be the normalized, approximate difference between the number of spins in a first direction minus the number of spins in the opposite direction. As a non-limiting example, 200,000 1 Assuming a nuclear spin of H, a polarization of 2% can correspond to 102,000 spins in one direction and 98,000 spins in the opposite direction. In some embodiments, "hyperpolarization" can include polarization of a species (e.g., nuclear, selective, or the like) beyond the typical polarization level of that species observed at thermal equilibrium upon exposure to a particular magnetic field. As a non-limiting example, 1 A sample in a magnetic field of 1 Tesla (T) at thermal equilibrium where the H nuclear spin polarization exceeds 0.000341% is substantially higher (e.g., at least one order of magnitude higher) than the 0.000341% thermal equilibrium polarization. 1 As a further non-limiting example, the ion beam may be hyperpolarized to have a H nuclear spin polarization of greater than 0.000257%. 13 A sample in a 3 T magnetic field at thermal equilibrium with a spin polarization of C can be hyperpolarized. 15 A sample in a 3 T magnetic field at thermal equilibrium with a spin polarization of N can be hyperpolarized.

[0020] As used in this disclosure, "hyperpolarization" describes a state in which the absolute value of the difference between a population of spin states (e.g., nuclear spin states, proton spin states, etc.) in one state (e.g., spin up) and a population of spin states in another state (e.g., spin down) exceeds the absolute value of the corresponding difference at thermal equilibrium.

[0021] Parahydrogen can be used as a polarization source consistent with disclosed embodiments. Parahydrogen is a form of molecular hydrogen in which two proton spins are in a singlet state, as described herein. The disclosed embodiments are not limited to a particular method of producing parahydrogen. Parahydrogen can be formed in gaseous or liquid form. In some embodiments, parahydrogen is produced in gaseous form by flowing hydrogen gas at low temperature through a chamber using a catalyst (e.g., iron oxide or another suitable catalyst). The hydrogen gas can contain both parahydrogen and orthohydrogen. The low temperature can bring the hydrogen gas to thermodynamic equilibrium within the chamber, increasing the population of parahydrogen.

[0022] As used in this disclosure, the population difference between two spin states is the difference between the populations of the two spin states divided by the total population of the two spin states. The population difference may be expressed as a fractional population difference or a percentage population difference. In some embodiments, the fractional population difference is at least about 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, or more, up to about 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, 0.1, or less, or within a range defined by any two of the foregoing values.

[0023] At thermal equilibrium, hydrogen gas can exhibit a population difference between proton spin states that significantly exceeds the population difference between the proton spin states. Hydrogen gas containing a high concentration of parahydrogen can have a large population difference between either the singlet or triplet spin state. In the case of Iz1Iz2 ordering, for example, there is a large population difference between the spin states |↑>|↓> and |↑>|↑>. The population difference between the proton spin states can be at least about 0.1 (e.g., a 10% difference in spin states or 55% of the parahydrogen molecules in a sample are in the singlet state and 45% are in the triplet state), 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, or more, up to about 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, 0.1, or less, or within a range defined by any two of the foregoing values.

[0024] As used in this disclosure, "molar polarization" refers to the product of the polarization and concentration of a particular nucleus. As used in this disclosure, molar polarization is typically quoted in concentration units, such as millimolar (mM), and is obtained by multiplying the concentration of a particular nucleus by its polarization, expressed as a fractional value. For example, a molecule may be present in a solution at a concentration of 100 mM. The molecule may contain a single carbon-13 nucleus with a 40% nuclear spin polarization (i.e., a polarization of 0.4). In this disclosure, such a molecule would have a carbon-13 molar polarization of 100 mM x 0.4 = 40 mM. Similarly, a molecule present in a solution at a concentration of 100 mM containing two identical protons, each with a nuclear spin polarization of 30%, would have a proton polarization of 60 mM. Thus, different molar polarizations can be obtained for different nuclei within a molecule. theoretical background

[0025] In conventional hyperpolarization experiments utilizing dynamic nuclear polarization (DNP), the concentration of target nuclei is typically low enough that the magnetic field generated by the target nuclei during polarization accumulation is negligible. The same is true for PHIP, PHIP-SAH, PHIP-X, and PHIPNOESYS experiments at moderate concentrations. However, numerical studies of PHIP at high concentrations suggest that this is no longer the case once the molar polarization exceeds a certain value. At this point, the magnetic field generated by the target nuclei can become strong enough to exceed the vibrational frequency associated with the splitting of pseudospin hydrogen. This can cause the nonlinear equations, which include nuclear backaction as a mean-field contribution, to become chaotic.

[0026] Although it seems possible to eliminate the presence of demagnetizing fields within a spherical reaction chamber, this can only be achieved with a perfectly uniform spatial distribution of nuclei and their polarization (i.e., polarization density). In reality, even within a spherical chamber, these quantities can fluctuate randomly, making it difficult to completely avoid demagnetizing fields. In the chaotic regime, these random fluctuations can be amplified very quickly (e.g., exponentially in time), further exacerbating the problem. Furthermore, in parahydrogen-based polarization procedures, the parahydrogenation reaction is subject to random fluctuations, making it difficult to avoid demagnetizing fields through geometric considerations alone. Therefore, it is necessary to develop approaches to suppress the effect of demagnetizing fields on polarization dynamics.

[0027] The systems and methods described herein solve this problem by combining a dipolar decoupling scheme (including but not limited to Lee-Goldberg decoupling, MREV-8 decoupling, BLEW-12 decoupling, or BR-24 decoupling) with a coupled driving field to develop novel polarization schemes (referred to herein as transverse field sweep in the Lee-Goldberg frame, polarized MREV-8, polarized BLEW-12, or polarized BR-24, respectively) based on a parameter sweep that is highly robust and not limited by the concentration of molecules undergoing the hyperpolarization procedure.

[0028] Considered systems Regarding the explanation of PHIP and hyperpolarization at high concentrations, we refer to M. Korzeczek et al., "Towards a unified picture of polarization transfer-equivalence of DNP and PHIP," arXiv:2303.07478 (2023) (hereafter, Korzeczek 2023) and J. Eills et al., "Singlet order conversion and parahydrogen-induced hyperpolarization of 13We used a model similar to that presented in "C nuclei in near-equivalent spin systems," J. Magn. Reson. 274, 163-172 (2017), each of which is incorporated herein by reference in its entirety for all purposes. We combined this model with a semiclassical mean-field description of intermolecular dipole-dipole coupling and effects from dipole fields (see, e.g., M.H. Levitt, "Demagnetization field effects in two-dimensional solution NMR," Conc. Magn. Reson. 8, 77-103 (1996)), which is incorporated herein by reference in its entirety for all purposes). For the derivation of the transfer sequence, we consider a two-spin system as in Korzeczek 2023.

[0029] The complete dynamics is calculated using i) the single-molecule Hamiltonian H0, ii) the effect of the magnetic field H B , and iii) intermolecular coupling as described by the dipole field H dip It is done by.

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[0030] Molecules are described by two (pseudo) spins, S and I, the former of which can be driven by a drive (e.g., radio frequency (RF) drive, transverse magnetic field (B1 drive)). The spins can be expressed in the form

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[0031] where ω S =γ S B0 and ω I =γ H B0 is a magnetic field of magnitude B0

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[0032] To account for the effects of dipole-dipole coupling between molecules in highly concentrated samples, we use a semiclassical mean-field description of the dipole field. Note, however, that all of the transfer schemes that suppress the dipole field contribution also suppress contributions from the full quantum description of dipole-dipole coupling.

[0033] For the mean-field description, we first consider the position across the sample

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[0034] where:

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[0035] therefore,

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[0036] Before entering the frame that corotates with the Larmor precession of S, we detune

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[0037] We now consider the (detuned) Larmor precession of S

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[0038] where:

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[0039] The effect of dipole fields on non-adaptive sequences For all typical polarization sequences, the (1) frame

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[0040] Here, we consider H dip from

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[0041] Amplitude Sweep For the amplitude sweep, we assume an initial π / 2 pulse with phase -Y followed by a continuous wave pulse with amplitude Ω(t) and phase X. We will later choose this to be a linear amplitude sweep.

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[0042] Inserting into all Hamiltonians and using the same arguments as before, we get

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[0043] Here, we use the fact that S spins give rise only to z magnetization to derive the second equation: the remaining dipole field term

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[0044] From this, we see that reliable transfer via adiabatic amplitude sweeps can be possible up to demagnetization fields significantly higher than those at which the untuned sequence is effective. Next, we consider how dynamical decoupling can lead to further improvements.

[0045] Lee-Goldberg Transfer During Decoupling In this section, we consider the driving field

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[0046] To account for the LG drive, we use the constant

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[0047] where:

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[0048] To maintain equation (19), the state

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[0049] We now define the following:

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[0050] Equations (22) to (24) are

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[0051] this is,

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[0052] General dipolar decoupling sequences as a basis for polarization transfer. Continuous wave Lee-Goldberg decoupling,

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[0053] These properties are sufficient to ensure, in a rough Hamiltonian theory approximation, that

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[0054] This corresponds to the situation without a dipole field.

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[0055] This leads us to the following:

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[0056] in principle,

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[0057] Without loss of generality, we now consider the preferred (0) frame

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[0058]

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[0059] Dipolar decoupling polarization sequence Linearization of bipolar decoupling sequences

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[0060] Polarized MREV-8:

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[0061] Here, the total duration is

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[0062] Polarized BLEW-12:

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[0063] Here, the total duration is

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[0064] Polarized BR-24:

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[0065] Here, the total duration is

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[0066] Using the same principles used in the construction of the polarized MREV-8, BLEW-12, and BR-24 sequences,

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[0067] Similarly, the principles described herein can be used to construct pulse sequences that allow for the generation of pulse sequences that enhance molar polarization in hyperpolarized molecules prepared via alternative hyperpolarization techniques such as DNP or dissolution DNP.

[0068] Methods for producing high molar polarization 1 illustrates an exemplary method 100 for generating high molar polarization in a hyperpolarized molecule via a PHIP nuclear spin hyperpolarization protocol, according to disclosed embodiments. In some embodiments, the hyperpolarized molecule comprises at least one nucleus. In some embodiments, the at least one nucleus comprises at least one NMR-active nucleus, such as at least one proton, deuterium nucleus, carbon-13 nucleus, nitrogen-15 nucleus, oxygen-17 nucleus, fluorine-19 nucleus, or phosphorus-31 nucleus.

[0069] In some embodiments, method 100 imparts a molar polarization to at least one nucleus. In some embodiments, the molar polarization is at least about 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, 100 mM, 110 mM, 120 mM, 130 mM, 140 mM, 150 mM, 160 mM, 170 mM, 180 mM, 190 mM, 200 mM, 210 mM, 220 mM, 230 mM, 240 mM, 250 mM, 260 mM, 270 mM, 280 mM, 290 mM, 300 mM, 310 mM, 320 mM, 330 mM, 340 mM, 350 mM, 360 mM, 370 mM, 380 mM, 390 mM, 400 mM, 410 mM, 420 mM, 430 mM, 440 mM, 450 mM, 460 mM, 470 mM, 480 mM, 490 mM, 500 mM, 510 mM, 520 mM, 530 mM, 540 mM, 550 mM, 560 mM, 570 mM, 580 mM, 590 mM, 600 mM, 610 mM, 620 mM, 630 mM, 640 mM, 650 mM, 660 mM, 670 mM, 680 mM, 690 mM, 700 mM, 710 mM, 720 mM, 730 mM, 740 mM, 750 mM, 760 mM, 770 mM, 780 mM, 790 mM, 800 mM, 810 mM, 820 mM, 830 mM, 840 mM 0mM, 280mM, 290mM, 300mM, 310mM, 320mM, 330mM, 340mM, 350mM, 360mM, 370mM, 380mM, 390mM , 400mM, 410mM, 420mM, 430mM, 440mM, 450mM, 460mM, 470mM, 480mM, 490mM, 500mM, 510mM, 520 mM, 530mM, 540mM, 550mM, 560mM, 570mM, 580mM, 590mM, 600mM, 610mM, 620mM, 630mM, 640mM, 650mM, 660mM, 670mM, 680mM, 690mM, 700mM, 710mM, 720mM, 730mM, 740mM, 750mM, 760mM, 770m 800 mM, 810 mM, 820 mM, 830 mM, 840 mM, 850 mM, 860 mM, 870 mM, 880 mM, 890 mM, 900 mM, 910 mM, 920 mM, 930 mM, 940 mM, 950 mM, 960 mM, 970 mM, 980 mM, 990 mM, 1,000 mM, or more.In some embodiments, the molar polarization is up to about 1,000 mM, 990 mM, 980 mM, 970 mM, 960 mM, 950 mM, 940 mM, 930 mM, 920 mM, 910 mM, 900 mM, 890 mM, 880 mM, 870 mM, 860 mM, 850 mM, 840 mM, 830 mM, 820 mM, 810 mM, 800 mM, 79 ... mM, 780mM, 770mM, 760mM, 750mM, 740mM, 730mM, 720mM, 710mM, 700mM, 690mM, 680mM, 670mM, 660mM, 650mM, 640mM, 630mM, 620mM, 610mM, 600mM, 590mM, 580mM, 570mM, 560mM, 550mM, 540m M, 530mM, 520mM, 510mM, 500mM, 490mM, 480mM, 470mM, 460mM, 450mM, 440mM, 430mM, 420mM, 4 10mM, 400mM, 390mM, 380mM, 370mM, 360mM, 350mM, 340mM, 330mM, 320mM, 310mM, 300mM, 290m 20 mM, 280 mM, 270 mM, 260 mM, 250 mM, 240 mM, 230 mM, 220 mM, 210 mM, 200 mM, 190 mM, 180 mM, 170 mM, 160 mM, 150 mM, 140 mM, 130 mM, 120 mM, 110 mM, 100 mM, 90 mM, 80 mM, 70 mM, 60 mM, 50 mM, or less. In some embodiments, the molar polarization is within a range defined by any two of the foregoing values.

[0070] In the example shown, a solution is obtained at 110. In some embodiments, the solution comprises a derivative of the hyperpolarized molecule. In some embodiments, the derivative comprises at least one unsaturated carbon-carbon double bond or at least one unsaturated carbon-carbon triple bond. In some embodiments, the derivative has the form R1-C=C-R2 or R1-C≡C-R2, where R1 and R2 represent side chains, = represents a carbon-carbon double bond, and ≡ represents a carbon-carbon triple bond.

[0071] In some embodiments, the unsaturated carbon-carbon double bond or the unsaturated carbon-carbon triple bond is configured to undergo a hydrogenation reaction with para-hydrogen. Thus, at 120, the unsaturated carbon-carbon double bond or the unsaturated carbon-carbon triple bond is hydrogenated with para-hydrogen to form a para-hydrogenated derivative of the hyperpolarized molecule. In some embodiments, the para-hydrogenated derivative has the form R-CH * -CH * -R2 or R1-CH * =CH * -R2, where R1 and R2 represent side chains, - represents a carbon-carbon single bond, = represents a carbon-carbon double bond, and H * indicates a para-hydrogen-derived hydrogen atom added across a double or triple bond during the hydrogenation reaction.

[0072] In some embodiments, the hydrogenation reaction is carried out by mixing parahydrogen gas with a solution such that the parahydrogen gas mixes with the derivative. In some embodiments, the solution contains a hydrogenation catalyst. In some embodiments, the parahydrogen gas is mixed with the derivative in the presence of the hydrogenation catalyst. In some embodiments, the mixture of parahydrogen gas and the derivative molecule in the presence of the hydrogenation catalyst induces a parahydrogenation reaction between the parahydrogen gas and the derivative.

[0073] At 130, a polarization transfer waveform is applied. In some embodiments, the polarization transfer waveform transfers nuclear spin order from at least one of the parahydrogen-derived hydrogen atoms to at least one nucleus. In some embodiments, the polarization transfer waveform thereby imparts nuclear spin hyperpolarization to at least one nucleus. In some embodiments, operation 130 is applied after operation 120. In some embodiments, the polarization transfer waveform is configured to suppress a dipolar field associated with magnetization generated during the accumulation of nuclear spin hyperpolarization. In some embodiments, the polarization transfer waveform includes any one or more of a dipolar decoupling sequence, a transverse field (B1) sweep, e.g., a sweep in a Lee-Goldberg frame, and a pulse sequence selected from the group consisting of polarized MREV-8, polarized BLEW-12, and polarized BR-24.

[0074] In some embodiments, operations 110, 120, and 130 produce hyperpolarized molecules, i.e., in some embodiments, operations 110, 120, and 130 form a PHIP nuclear spin polarization protocol.

[0075] In some embodiments, the hyperpolarized molecule comprises any molecule of interest described herein. In some embodiments, the hyperpolarized molecule is used in an NMR or MRI experiment.

[0076] In other embodiments, the nuclear spin polarization from the hyperpolarized molecule is transferred to any molecule of interest described herein, and the molecule of interest is used in an NMR or MRI experiment. In some embodiments, the nuclear spin polarization is transferred from the hyperpolarized molecule to the molecule of interest by a PHIPNOESYS or PHIP-X procedure. That is, in some embodiments, the nuclear spin polarization is transferred from the hyperpolarized molecule to the molecule of interest by an intermolecular NOE between the hyperpolarized molecule and the molecule of interest, or by proton exchange between the hyperpolarized molecule and the molecule of interest.

[0077] In some embodiments, method 100 further comprises performing at least one purification protocol on the hyperpolarized molecule or molecule of interest. Example purification protocols are described, for example, in WO2022 / 018514 and WO2022 / 269350, each of which is incorporated by reference herein in its entirety for all purposes.

[0078] 2 illustrates an exemplary method 200 for generating high molar polarization in a hyperpolarized molecule via a PHIP-SAH nuclear spin hyperpolarization protocol, according to disclosed embodiments. In some embodiments, the hyperpolarized molecule comprises at least one nucleus. In some embodiments, the at least one nucleus comprises an NMR-active nucleus as described herein with respect to FIG. 1.

[0079] In some embodiments, method 200 imparts a molar polarization to at least one nucleus. In some embodiments, the molar polarization is any molar polarization described herein with respect to FIG.

[0080] In the example shown, a solution is obtained at 210. In some embodiments, the solution comprises a derivative of the hyperpolarized molecule. In some embodiments, the derivative comprises at least one unsaturated carbon-carbon double bond or at least one unsaturated carbon-carbon triple bond. In some embodiments, the derivative has the form R1-C=C-R2 or R1-C≡C-R2, where R1 and R2 represent side chains, = represents a carbon-carbon double bond, and ≡ represents a carbon-carbon triple bond.

[0081] In some embodiments, the unsaturated carbon-carbon double bond or the unsaturated carbon-carbon triple bond is configured to undergo a hydrogenation reaction with para-hydrogen. Thus, at 220, the unsaturated carbon-carbon double bond or the unsaturated carbon-carbon triple bond is hydrogenated with para-hydrogen to form a para-hydrogenated derivative of the hyperpolarized molecule. In some embodiments, the para-hydrogenated derivative has the form R-CH * -CH * -R2 or R1-CH * =CH * -R2, where R1 and R2 represent side chains, - represents a carbon-carbon single bond, = represents a carbon-carbon double bond, and H * indicates a para-hydrogen-derived hydrogen atom added across a double or triple bond during the hydrogenation reaction.

[0082] In some embodiments, the hydrogenation reaction is carried out by mixing parahydrogen gas with a solution such that the parahydrogen gas mixes with the derivative. In some embodiments, the solution contains a hydrogenation catalyst. In some embodiments, the parahydrogen gas is mixed with the derivative in the presence of the hydrogenation catalyst. In some embodiments, the mixture of the parahydrogen gas and the derivative in the presence of the hydrogenation catalyst induces a parahydrogenation reaction between the parahydrogen gas and the derivative.

[0083] At 230, a polarization transfer waveform is applied. In some embodiments, the polarization transfer waveform transfers nuclear spin order from at least one of the para-hydrogen-derived hydrogen atoms to at least one nucleus. In some embodiments, the polarization transfer waveform thereby imparts nuclear spin hyperpolarization to at least one nucleus. In some embodiments, operation 230 is applied after operation 220. In some embodiments, the polarization transfer waveform is configured to suppress a dipole field associated with magnetization generated during the accumulation of nuclear spin hyperpolarization. In some embodiments, the polarization transfer waveform includes a polarization transfer waveform described herein with respect to FIG. 1 .

[0084] At 240, the para-hydrogenated derivative is hydrolyzed. In some embodiments, hydrolyzing the para-hydrogenated derivative forms a hyperpolarized molecule. In some embodiments, the para-hydrogenated derivative is mixed with a hydrolysis agent, such as sodium hydroxide (NaOH) or potassium hydroxide (KOH). In some embodiments, the hydrolysis agent hydrolyzes the para-hydrogenated derivative of the molecule of interest, forming a hydrolyzed side arm and a hyperpolarized molecule via a PHIP-SAH interaction. Examples of PHIP-SAH interactions can be found, for example, in WO2022 / 157534, WO2022 / 018514, and WO2021 / 198776, each of which is incorporated by reference in its entirety for all purposes.

[0085] In some embodiments, operations 210, 220, 230, and 240 generate hyperpolarized molecules, i.e., in some embodiments, operations 210, 220, 230, and 240 form a PHIP-SAH nuclear spin polarization protocol.

[0086] In some embodiments, the hyperpolarized molecule comprises any molecule of interest described herein. In some embodiments, the hyperpolarized molecule is used in an NMR or MRI experiment.

[0087] In other embodiments, the nuclear spin polarization from the hyperpolarized molecule is transferred to any molecule of interest described herein, and the molecule of interest is used in an NMR or MRI experiment. In some embodiments, the nuclear spin polarization is transferred from the hyperpolarized molecule to the molecule of interest by a PHIPNOESYS or PHIP-X procedure. That is, in some embodiments, the nuclear spin polarization is transferred from the hyperpolarized molecule to the molecule of interest by an intermolecular NOE between the hyperpolarized molecule and the molecule of interest, or by proton exchange between the hyperpolarized molecule and the molecule of interest.

[0088] In some embodiments, method 200 further comprises performing at least one purification protocol on the hyperpolarized molecule or molecule of interest, hi some embodiments, the at least one purification protocol comprises any of the purification protocols described herein with respect to FIG.

[0089] Targeted molecules and bio-related imaging agents Disclosed embodiments include systems and methods for producing and utilizing molecules of interest with clinically relevant polarization, concentration, volume, or purity. In some embodiments, the method is for preparing NMR materials (also referred to herein as "molecules of interest"). In some embodiments, the NMR materials are suitable for use in NMR or MRI operations. In some embodiments, the NMR materials increase NMR or MRI signals and signal-to-noise ratios (SNRs). In some embodiments, the NMR materials are suitable for use in solution NMR spectroscopy. In some embodiments, the NMR materials are chemical compounds. In some embodiments, the NMR materials are metabolites (e.g., biologically relevant molecules such as amino acids, sugars, and derivatives thereof), such as metabolites suitable for use in NMR metabolomics applications. In some embodiments, the NMR materials are suitable for in vitro probing of the metabolism of cell cultures or other biological tissues. In some embodiments, the NMR materials are used in NMR probes to investigate transient effects where high signal enhancement due to hyperpolarization is required, such as proton exchange between water and biomolecules. In some embodiments, the NMR material is a small molecule or metabolite suitable for injection into a cell, tissue, or organism for detection in an MRI scan. In some embodiments, the NMR material is introduced into a chamber for further analysis by NMR or MRI operation. In some embodiments, the NMR material is one or more deuterium ( 2 H) or carbon-13 ( 13 C) concentrated in atoms.

[0090] Consistent with disclosed embodiments, the NMR material may include a biorelevant contrast agent. In some embodiments, the biorelevant contrast agent may be suitable for use in NMR or MRI operations. In some embodiments, the biorelevant contrast agent may increase the NMR or MRI signal or signal-to-noise ratio (SNR). In some embodiments, the biorelevant contrast agent may be suitable for use in solution NMR spectroscopy. In some embodiments, the biorelevant contrast agent may be a metabolite (e.g., a molecule with biorelevance, such as an amino acid, sugar, or derivative thereof), such as a metabolite suitable for use in NMR metabolomics applications. In some embodiments, the biorelevant contrast agent is used for perfusion or contrast-enhanced imaging in MRI scans. In some embodiments, the biorelevant contrast agent may be suitable for in vitro probing of the metabolism of cell cultures or other biological tissues. In some embodiments, the biorelevant contrast agent is used for in vitro probing of the metabolism of cell cultures or other biological tissues. In some embodiments, the biorelevant contrast agent may be used in NMR probes to investigate transient effects where high signal enhancement due to hyperpolarization is required, such as proton exchange between water and biomolecules. In some embodiments, the biorelevant imaging agent may be a small molecule or metabolite suitable for injection into a cell, tissue, or organism for detection in an MRI scan. In some embodiments, the biorelevant imaging agent may be introduced into a chamber for further analysis by NMR or MRI operation. In some embodiments, the biorelevant imaging agent may be one or more 2 H or 13 It is enriched in C atoms.

[0091] In some embodiments, the biologically relevant imaging agent is pyruvate, lactate, alpha-ketoglutarate, bicarbonate, fumarate, urea, dehydroascorbate, glutamic acid, glutamine, acetate, dihydroxyacetone, acetoacetate, glucose, ascorbate, zymonate, alanine, fructose, imidazole, nicotinamide, nitroimidazole, pyrazinamide, isoniazid, conjugate acids of any of the above, natural and unnatural amino acids, esters thereof, or any of the foregoing. 2 H, 13C, or nitrogen-15( 15 In some embodiments, the biologically relevant imaging agent includes pyruvate, lactate, alpha-ketoglutarate. In some embodiments, the biologically relevant imaging agent includes pyruvate. In some embodiments, the biologically relevant imaging agent includes lactate. In some embodiments, the biologically relevant imaging agent includes alpha-ketoglutarate (e.g., ethyl alpha-ketoglutarate).

[0092] In some embodiments, the biologically relevant imaging agent comprises at least one non-hydrogen nuclear spin. In some embodiments, the non-hydrogen nucleus comprises at least one spin 1 / 2 atom. In some embodiments, the non-hydrogen nuclear spin is 13 C or 15 N. In some embodiments, the biologically relevant imaging agent is at least partially isotopically labeled with non-hydrogen nuclear spins. In some embodiments, the biologically relevant imaging agent is at least partially enriched in non-hydrogen nuclear spins compared to an analog of the biologically relevant imaging agent characterized by non-hydrogen nuclear spins at their natural abundance. In some embodiments, the biologically relevant imaging agent is at least partially enriched in non-hydrogen nuclear spins compared to an analog of the biologically relevant imaging agent characterized by non-hydrogen nuclear spins at their natural abundance. In some embodiments, the biologically relevant imaging agent is at least about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or more, up to about 99%, 98%, 97%, 96%, Enriched to characterize non-hydrogen nuclear spins at abundances of 95%, 94%, 93%, 92%, 91%, 90%, 85%, 80%, 75%, 70%, 65%, 60%, 55%, 50%, 45%, 40%, 35%, 30%, 25%, 20%, 15%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less, or abundances within a range defined by any two of the foregoing values.

[0093] In some embodiments, the non-hydrogen nuclear spins are NMR inactive (i.e., spin 0) nuclei of an analog of a biologically relevant imaging agent that characterizes the non-hydrogen nuclear spins at their natural abundance (e.g., 12C or quadrupolar (i.e., spin >1 / 2) nuclei (e.g., nitrogen-14, 14 N) at their natural abundance. 13 The analogue of pyruvate that characterizes C has the structure H3C-C * (=O)-C * OOH either C * So, about 98.9% 12 C and about 1.1% 13 As a biorelevant imaging agent, pyruvate may contain one or both of the C * at any abundance described herein 13 To include C, instead 13 As used herein, * C and C * teeth, 12 C or 13 C describes carbon, which can be any of the carbon isotopes. 15 The analogue of urea that characterizes N is the structure H2N * -C(=O)- * Any N in NH2 * Approximately 99.6% 14 N and about 0.4% 15 As a biorelevant imaging agent, urea may contain one or both N * at any abundance described herein 15 To include N, instead 15 As used herein, * N and N * teeth, 14 N or 15 Describes nitrogen, which can be any of the nitrogen isotopes N.

[0094] precipitation In various embodiments, hyperpolarized molecules or molecules of interest can be crystallized or precipitated from the solutions described herein. The disclosed embodiments are not limited to any particular method of inducing such precipitation. For example, such precipitation can be induced via a change in temperature or pH, application of an electromagnetic stimulus (e.g., optical radiation such as ultraviolet radiation or optical radiation at another suitable wavelength or wavelengths), application of a mechanical stimulus (e.g., ultrasound, agitation, or another suitable mechanical stimulus), addition of another solute or solvent to the solution, or another suitable method, or any combination thereof. In some embodiments, after precipitation, the molecule of interest can be separated from the solution (e.g., using a filter or another suitable method). In some embodiments, the molecule of interest can then be combined with another solution or redissolved. This solution can have desirable properties (e.g., biocompatibility, concentration, volume, temperature, pH, polarity, or other relevant properties, or any combination thereof) for the intended NMR or MRI application.

[0095] Use of targeted molecular and biologically relevant imaging agents In some embodiments, at least a portion of the molecule of interest can be injected into a subject or patient for use in an MRI experiment. In various embodiments, at least a portion of the molecule of interest can be used in NMR spectroscopy. At least one NMR or MRI pulse sequence can be applied to the molecule of interest.

[0096] The foregoing description has been presented for purposes of illustration. It is not exhaustive and is not limited to the precise forms or embodiments disclosed. Modifications and adaptations of the embodiments will be apparent from consideration of the specification and practice of the disclosed embodiments. For example, while the described implementations include hardware, systems and methods consistent with the present disclosure can be implemented using both hardware and software. Further, while certain components are described as being coupled to each other, such components may be integrated with each other or distributed in any suitable manner.

[0097] Moreover, while exemplary embodiments are described herein, the scope includes any and all embodiments having equivalent elements, modifications, omissions, combinations (e.g., of aspects across various embodiments), adaptations, or alterations based on this disclosure. Claim elements should be construed broadly based on the language used in the claims and not limited to the examples described herein or during prosecution of the application, which examples should be construed as non-exclusive. Furthermore, the steps of the methods of the present disclosure can be modified in any manner, including rearranging steps or inserting or deleting steps.

[0098] The features and advantages of the present disclosure will be apparent from the detailed specification, and accordingly, the appended claims are intended to cover all systems and methods that fall within the true spirit and scope of the present disclosure. As used herein, the indefinite articles "a" and "an" mean "one or more." Similarly, the use of a plural term does not necessarily imply a plurality, unless the use is ambiguous in a given context. Furthermore, since numerous modifications and variations will readily occur from a study of this disclosure, it is not desired to limit the disclosure to the exact construction and operation illustrated and described. Accordingly, all suitable modifications and equivalents may be resorted to, falling within the scope of the present disclosure.

[0099] As used herein, unless specifically stated otherwise, the term "or" includes all possible combinations, including both conjunctions and disjunctions, unless impracticable. For example, if a component is stated to include A or B, the component may include only A, or only B, or A and B, unless specifically stated otherwise or impracticable. As a second example, if a component is stated to include A, B, or C, the component may include only A, or only B, or only C, or A and B, or A and C, or B and C, or A, B, and C. [Example]

[0100] Example 1: Numerical simulation of polarization transfer dynamics The systems and methods herein were used to numerically evaluate the polarization transfer dynamics of various molecules using a variety of untuned and tuned pulse sequences. The Hamiltonian used corresponded to a two-spin system to describe homonuclear PHIP with the following correspondence:

number

number

[0101] For the numerical simulations, parameters from three different molecules (carbon-13 labeled deuterated dimethyl maleate, carbon-13 labeled fumaric acid, and deuterated ethyl acetate) were used. For fumaric acid, a molecule without a carbon-13 spin label was used. Therefore, to induce a chemical shift, fumaric acid was simulated using a magnetic field amplitude of 100 millitesla (mT) instead of the 100 microtesla (μT) used for the other two molecules. The numerical simulations explicitly rotate all three (two for ethyl acetate) instead of the representative Hamiltonian from the previous section. The J coupling between hydrogens, J, and (where applicable) the two hydrogen and carbon spins, J, are used. (1) Heteronuclear J coupling between, and J (2) Then, the relevant parameters for different molecules are as follows: (1- 13C,d6)-dimethylmaleate: J = (2π) 12.2 Hz, J (1) =(2π)13.1Hz, J (2) =(2π)2.7Hz;(1- 13 C)-fumaric acid: J=(2π)7.1Hz, J (1) =(2π)3.2Hz, J (2) = (2π) 0.4 Hz; (1,2-d2)-ethyl acetate: J = (2π) 15.9 Hz, A = 100 mT,

number

[0102] Example 2: (1- 13 Numerical simulations of polarized MREV-8, polarized BLEW-12, polarized BR-24, and transverse field sweeps in the Lee-Goldberg frame for C,d6)-dimethylmaleate Figure 3 shows the (1- 13 Figure 3 shows exemplary numerical simulations of molar polarization versus concentration for polarized MREV-8 ("MREV-8 adj" in Figure 3), polarized BLEW-12 ("BLEW-12 adj" in Figure 3), polarized BR-24 ("BR-24 adj" in Figure 3), and transverse field sweeps with Lee-Goldberg frame ("LG+Sweep" in Figure 3) pulse sequences for (C,d6)-dimethylmaleate. Numerical simulations were performed as described herein with respect to Example 1. Simulations were also performed for SLIC and pure transverse field sweeps ("Sweep" in Figure 3) to allow for appropriate comparisons between the pulse sequences described herein and previous pulse sequences. Concentrations are conveniently expressed in units of (2π) Hz in performing the simulations. As discussed herein, concentrations can be converted to units of mM by multiplying by a proportionality constant. Similarly, molar polarization is expressed in units of (fraction) polarization time (2π) Hz and can be converted to molar polarization in units of mM by multiplying by a proportionality constant. As shown in Figure 3, the pulse sequence described herein is clearly superior to SLIC and pure transverse field sweep approaches, and the transverse field sweep in the Lee-Goldberg frame achieves the highest molar polarization.

[0103] Example 3: (1- 13 C) Numerical simulations of polarized MREV-8, polarized BLEW-12, polarized BR-24, and transverse field sweep in the Lee-Goldberg frame of fumaric acid. Figure 4 shows the (1- 13 Figure 4 shows exemplary numerical simulations of molar polarization versus concentration for the Lee-Goldberg frame ("LG+Sweep" in Figure 4) pulse sequence for C-fumaric acid with polarized MREV-8 ("MREV-8 adj" in Figure 4), polarized BLEW-12 ("BLEW-12 adj" in Figure 4), polarized BR-24 ("BR-24 adj" in Figure 4), and transverse field sweep. Numerical simulations were performed as described herein with respect to Example 1. Simulations were also performed for SLIC and pure transverse field sweep ("Sweep" in Figure 4) to allow for appropriate comparison between the pulse sequences described herein and previous pulse sequences. Concentrations are conveniently expressed in units of (2π) Hz in performing the simulations. As discussed herein, concentrations can be converted to units of mM by multiplying by a proportionality constant. Similarly, molar polarization is expressed in units of (fraction) polarization time (2π) Hz and can be converted to molar polarization in units of mM by multiplying by a proportionality constant. As shown in Figure 4, the pulse sequence described herein is clearly superior to SLIC and pure transverse field sweep approaches, and the transverse field sweep in the Lee-Goldberg frame achieves the highest molar polarization.

[0104] Example 4: Numerical simulations of polarized MREV-8, polarized BLEW-12, polarized BR-24, and transverse field sweep in the Lee-Goldberg frame of (1,2-,d2)-ethyl acetate Figure 5 shows exemplary numerical simulations of molar polarization versus concentration for the Lee-Goldberg frame (LG+Sweep in Figure 5) pulse sequence for (1,2-d2)-ethyl acetate with polarized MREV-8 (MREV-8 adj in Figure 5), polarized BLEW-12 (BLEW-12 adj in Figure 5), polarized BR-24 (BR-24 adj in Figure 5), and transverse field sweep. Numerical simulations were performed as described herein with respect to Example 1. Simulations were also performed for SLIC and pure transverse field sweep (Sweep in Figure 5) to allow for appropriate comparison between the pulse sequences described herein and previous pulse sequences. Concentrations are conveniently expressed in (2π) Hz units in the simulation runs. As discussed herein, concentrations can be converted to units of mM by multiplying by a proportionality constant. Similarly, molar polarization is expressed in units of (fraction) polarization time (2π) Hz and can be converted to molar polarization in units of mM by multiplying by a proportionality constant. As shown in Figure 5, the pulse sequence described herein clearly outperforms SLIC and pure transverse field sweep approaches, with transverse field sweeps in the Lee-Goldberg frame achieving the highest molar polarization.

[0105] Example 5: Using a transverse magnetic field sweep in the Lee-Goldberg frame, (1- 13 Experimental demonstration of high molar polarization of (c,d6)-dimethylmaleate Using the systems and methods described herein, 13 C,d6)-dimethylmaleate, imparting high molar polarization up to 450 mM. 13 Precursor solutions of (1-(c,d6)-dimethylmaleate) were prepared by dissolving 5 mM of [Rh(dppb)(COD)]BF4 catalyst (CAS number: 79255-71-3) in acetone-d6. 13C,d6)-dimethylacetylenedicarboxylate was mixed for different experiments: 20, 40, 80, 160, 320, 640, and 1080 mM for each concentration point in the data series provided herein. Parahydrogen was produced by an ARS parahydrogen generator packed with iron monohydrate catalyst and run at a temperature of 22 K, producing gas with a para enrichment level of approximately 93%.

[0106] Each experiment was initiated by injecting 500 microliters (μL) of solution into the tube and bubbling para-enriched hydrogen gas through the solution at 10 bar pressure in a 96 μT bias field. This was followed by bubbling nitrogen at 10 bar to stop further reaction progress. The total bubbling period was fixed at 30 seconds (s) in all experiments to avoid rapid singlet order decay. 1 H decoupling was provided.

[0107] The polarization transfer was performed in two different ways. The first method was to use the amplitude ( 1 The transverse magnetic field consisted of a swept frequency from 0 Hz to 25 Hz at 1000 kJ / s (relative to H) followed by an adiabatic pulse. The pulses were arranged by ramping down the transverse magnetic field amplitude over 1 s with a stepwise carrier frequency shift of -200 Hz.

[0108] The second method is to calculate the effective magnetic field B e is at an angle θ to the bias field e As shown in Figure 6, off-resonance driving was included during polarization transfer. The effective field amplitude was set to 600 Hz and 400 Hz for the experiments described with respect to Figures 6 and 7, respectively. The transfer flip-back pulse was performed by ramping down the transverse magnetic field amplitude in 1 s with a step carrier frequency shift of -200 Hz. The polarization transfer was performed with a frequency of 0 Hz to 25 Hz ( 1 This was achieved by ramping modulation of the off-resonance driving field amplitude (relative to H). The modulation frequency was set to match the effective field amplitude. To implement an adiabatic pulse along the effective field, the modulation amplitude was ramped down with a step modulation frequency shift of -200 Hz over 1 s.

[0109] 1 H free induction decay was initiated by a small flip-angle hard pulse of 20 kHz radio frequency (RF) amplitude and recorded at a 131-point density with a spectral width of 400 ppm. Additional H decoupling was used in all experiments. Thermal equilibrium 1 H spectra were recorded at room temperature with a 90 s recycle delay and a 90 degree flip angle pulse. The flip angle scaling factor was taken into account when estimating the polarization level.

[0110] Figure 6 shows the high polarization (1- 13 C,d6)-dimethyl maleate 1 Figure 6 shows the molar polarization of H. Data points were acquired using transverse field sweeps with Lee-Goldberg frame (black dots) and amplitude-swept SLIC (gray dots) pulse sequences. The amplitude-swept SLIC duration was set to 2 seconds, and the Lee-Goldberg effective field amplitude was set to 600 Hz. The dashed line represents the linear dependence at a fixed polarization level of 47%. Note that the molar polarization is calculated as concentration × 2 to account for the presence of two polarized protons in dimethylmaleate after PHIP polarization. Rescaled inserts are provided for clarity. As shown in Figure 6, transverse field sweeps with Lee-Goldberg frame pulse sequences achieved molar polarization levels of up to 450 mM, compared to approximately 50 mM using a standard amplitude-swept SLIC pulse sequence.

[0111] Figure 7 shows the high polarization (1- 13 C,d6)-dimethyl maleate 1The H spin polarization is shown. Data points acquired at concentrations of 17 mM and 223 mM are shown in gray and black, respectively. The amplitude-swept SLIC duration was set to 4 seconds, and the Lee-Goldberg effective field amplitude was set to 400 Hz. The dashed lines indicate the polarization levels acquired with transverse field sweeps of the Lee-Goldberg frame pulse sequences derived in this work at high and low concentrations. The magic angle, approximately 54.7°, is shown separately. As shown in Figure 7, polarization was maximized at the magic angle.

[0112] Example 6: Polarized MREV-8 was used to 13 Experimental demonstration of high molar polarization of (c,d6)-dimethylmaleate Figure 8 shows the 1- 13 Exemplary hyperpolarization of C-d6-dimethylmaleate 1 The H spectrum is shown. The following thermally polarized spectrum was acquired at room temperature after the hyperpolarization experiment. The MREV-8 sequence is 1 Hydrogenation was carried out in a 200 μT bias field with the pulse amplitude set to 400 Hz, the free evolution time set to 0.625 ms, and 30 loops with a total duration of 0.3 s. Hydrogenation was carried out by bubbling parahydrogen at 10 bar for 25 s with an amplitude of 3 μT. 1 This was achieved by bubbling nitrogen at 10 bar for 5 seconds under H continuous wave decoupling. As shown in Figure 8, approximately 20% polarization was achieved with approximately 1 molar (M) = 1,000 mM 1- 13 This was achieved with a C-d6-dimethylmaleate concentration, which corresponds to a molar polarization of approximately 400 mM, since dimethylmaleate contains two polarized protons after PHIP polarization.

[0113] Enumeration of Embodiments Embodiment 1. A method comprising: (a) obtaining a solution having hyperpolarized molecules dissolved therein, the hyperpolarized molecules comprising at least one nucleus having a molar polarization of at least 50 millimolar (mM).

[0114] Embodiment 2. The method of embodiment 1, further comprising, prior to (a), performing a nuclear spin hyperpolarization protocol on the hyperpolarized molecule, thereby imparting molar polarization to at least one nucleus.

[0115] Embodiment 3. The nuclear spin hyperpolarization protocol comprises: (b) obtaining a solution containing a derivative of a hyperpolarized molecule, the derivative containing at least one unsaturated carbon-carbon double bond or unsaturated carbon-carbon triple bond and having the form R1-C=C-R2 or R1-C≡C-R2, wherein R1 and R2 contain side chains; (c) hydrogenating the double or triple bond with para-hydrogen to form a para-hydrogenated derivative of the hyperpolarized molecule, wherein the para-hydrogenated derivative has the form R-CH * -CH * -R2 or R1-CH * =CH * -R2, H * represents a para-hydrogen-derived hydrogen atom attached across a double or triple bond; (d) applying a polarization transfer waveform to transfer nuclear spin order from at least one of the para-hydrogen-derived hydrogen atoms to at least one nucleus, thereby imparting nuclear spin hyperpolarization to the at least one nucleus.

[0116] Embodiment 4. The method of embodiment 3, wherein the polarization transfer waveform is configured to suppress the dipolar field associated with the magnetization generated during the accumulation of nuclear spin hyperpolarization.

[0117] Embodiment 5. The method of embodiment 3 or 4, wherein the polarization transfer waveform comprises a bipolar decoupling sequence.

[0118] Embodiment 6. The method of embodiment 4 or 5, wherein the polarization transfer waveform further comprises a coupled driving field based on a parameter sweep.

[0119] Embodiment 7. The embodiment of claim 6, wherein the parameter sweep includes a transverse magnetic field (B1) sweep.

[0120] Embodiment 8. The method of any one of embodiments 3 to 7, wherein the polarization transfer waveform comprises a B1 sweep in the Lee-Goldberg frame.

[0121] Embodiment 9. The method of any one of embodiments 3 to 7, wherein the polarization transfer waveform comprises a pulse sequence selected from the group consisting of polarized MREV-8, polarized BLEW-12, and polarized BR-24.

[0122] Embodiment 10. The method of any one of embodiments 3 to 9, wherein hyperpolarized molecules are produced by (b) to (d).

[0123] Embodiment 11. The method of any one of embodiments 3-9, further comprising: (e) hydrolyzing the para-hydrogenated derivative, thereby forming a hyperpolarized molecule.

[0124] Embodiment 12. The method of embodiment 10 or 11, further comprising using the hyperpolarized molecule in a nuclear magnetic resonance (NMR) or magnetic resonance imaging (MRI) experiment, wherein the hyperpolarized molecule comprises a molecule of interest.

[0125] Embodiment 13. The method of embodiment 10 or 11, further comprising: (f) transferring nuclear spin polarization from the hyperpolarized molecule to the molecule of interest; and (g) using the molecule of interest in an NMR or MRI experiment.

[0126] Embodiment 14. The method of embodiment 13, wherein (f) comprises transferring nuclear spin polarization from a highly polarized molecule to the molecule of interest by a Parahydrogen Induced Polarization Nuclear Overhauser Effect System (PHIPNOESYS) procedure.

[0127] Embodiment 15. (f) The method of embodiment 13, comprising transferring nuclear spin polarization from a highly polarized molecule to a molecule of interest by relayed parahydrogen-induced polarization via a proton exchange (PHIP-X) procedure.

[0128] Embodiment 16. The method of embodiment 12 or 13, wherein the molecule of interest comprises a biologically relevant imaging agent.

[0129] Embodiment 17. The method of embodiment 16, wherein the biologically relevant imaging agent is selected from the group consisting of pyruvate, glutamate, glutamine, lactate, acetate, acetoacetate, zymonate, alanine, fructose, fumarate, bicarbonate, urea, dehydroascorbate, alpha-ketoglutarate, dihydroxyacetone, glucose, ascorbate, and their conjugate acids.

[0130] Embodiment 18. The method of any one of embodiments 1 to 17, wherein the molar polarization is composed of the product of the concentration of at least one nucleus and the nuclear spin polarization of at least one nucleus.

[0131] Embodiment 19. The molar polarization is at least 60 mM, 70 mM, 80 mM, 90 mM, 100 mM, 110 mM, 120 mM, 130 mM, 140 mM, 150 mM, 160 mM, 170 mM, 180 mM, 190 mM, 200 mM, 210 mM, 220 mM, 230 mM, 240 mM, 250 mM, 260 mM, 270 mM, 280 mM, 290 mM, 300 mM, 310 mM, 320 mM, 330 mM, 340 mM, 350 mM, 360 mM, 370 mM, 380 mM, 390 mM, 400 mM, 410 mM, 420 mM, 430 mM, 440 mM, 450 mM, 460 mM, 470 mM, 480 mM, 490 mM, 500 mM, 510 mM, 520 mM, 530 mM, 540 mM, 550 mM, 560 mM, 570 mM, 580 mM, 590 mM, 600 mM, 610 mM, 620 mM, 630 mM, 640 mM, 650 mM, 660 mM, 670 mM, 680 mM, 690 mM, 700 mM, 710 mM, 720 mM, 730 mM, 740 mM, 750 mM, 760 mM, 770 mM, 780 mM, 790 mM, 800 mM, 810 mM, 820 mM, 830 mM, 840 mM, 850 mM, 860 mM, 870 mM, 880 mM, 890 mM 90mM, 300mM, 310mM, 320mM, 330mM, 340mM, 350mM, 360mM, 370mM, 380mM, 390mM, 400mM, 410mM , 420mM, 430mM, 440mM, 450mM, 460mM, 470mM, 480mM, 490mM, 500mM, 510mM, 520mM, 530mM, 540m M, 550mM, 560mM, 570mM, 580mM, 590mM, 600mM, 610mM, 620mM, 630mM, 640mM, 650mM, 660mM, 67 0mM, 680mM, 690mM, 700mM, 710mM, 720mM, 730mM, 740mM, 750mM, 760mM, 770mM, 780mM, 790mM, 8 19. The method of any one of embodiments 1 to 18, wherein the concentration of HCl is 800 mM, 810 mM, 820 mM, 830 mM, 840 mM, 850 mM, 860 mM, 870 mM, 880 mM, 890 mM, 900 mM, 910 mM, 920 mM, 930 mM, 940 mM, 950 mM, 960 mM, 970 mM, 980 mM, 990 mM, or 1,000 mM.

[0132] Embodiment 20. At least one nucleus has at least one proton ( 1 H), carbon-13( 13 C), nitrogen-15( 15 N), fluorine-19( 19 F), or phosphate-31( 31 P) the method of any one of embodiments 1 to 19, comprising a nucleus.

[0133] Embodiment 21. A composition comprising: A composition comprising a solution having hyperpolarized molecules dissolved therein, the hyperpolarized molecules comprising at least one nucleus having a molar polarization of at least 50 millimolar (mM).

[0134] Embodiment 22. The composition of embodiment 21, wherein the hyperpolarized molecule is produced by performing a nuclear spin hyperpolarization protocol on the hyperpolarized molecule, thereby imparting molar polarization to at least one nucleus.

[0135] Embodiment 23. The nuclear spin hyperpolarization protocol comprises: (a) obtaining a solution containing a derivative of a hyperpolarized molecule, the derivative containing at least one unsaturated carbon-carbon double bond or unsaturated carbon-carbon triple bond and having the form R1-C=C-R2 or R1-C≡C-R2, wherein R1 and R2 contain side chains; (b) hydrogenating the double or triple bond with para-hydrogen to form a para-hydrogenated derivative of the hyperpolarized molecule, the para-hydrogenated derivative having the formula R-CH * -CH * -R2 or R1-CH * =CH * -R2, H * represents a para-hydrogen-derived hydrogen atom attached across a double or triple bond; (c) applying a polarization transfer waveform to transfer nuclear spin order from at least one of the para-hydrogen-derived hydrogen atoms to at least one nucleus, thereby imparting nuclear spin hyperpolarization to the at least one nucleus.

[0136] Embodiment 24. The composition of embodiment 23, wherein the polarization transfer waveform is configured to suppress the dipolar field associated with the magnetization generated during the accumulation of nuclear spin hyperpolarization.

[0137] Embodiment 25. The composition of embodiment 23 or 24, wherein the polarization transfer waveform comprises a bipolar decoupling sequence.

[0138] Embodiment 26. The composition of embodiment 24 or 25, wherein the polarization transfer waveform further comprises a coupled driving field based on a parameter sweep.

[0139] Embodiment 27. The composition of embodiment 26, wherein the parameter sweep comprises a transverse magnetic field (B1) sweep.

[0140] Embodiment 28. The composition of any one of embodiments 23 to 27, wherein the polarization transfer waveform comprises a B1 sweep in the Lee Goldberg frame.

[0141] Embodiment 29. The composition of any one of embodiments 23 to 27, wherein the polarization transfer waveform comprises a pulse sequence selected from the group consisting of polarized MREV-8, polarized BLEW-12, and polarized BR-24.

[0142] Embodiment 30. The composition of any one of embodiments 23 to 29, wherein hyperpolarized molecules are produced by (a) to (c).

[0143] Embodiment 31. The composition of any one of embodiments 23-29, wherein the hyperpolarized molecule is further generated by (d) hydrolyzing the para-hydrogenated derivative, thereby forming the hyperpolarized molecule.

[0144] Embodiment 32. The composition of embodiment 30 or 31, wherein the hyperpolarized molecule is for use in a nuclear magnetic resonance (NMR) or magnetic resonance imaging (MRI) experiment, and the hyperpolarized molecule comprises a molecule of interest.

[0145] Embodiment 33. The composition of embodiment 31 or 32, wherein the hyperpolarized molecule is for use in (e) transferring nuclear spin polarization from the hyperpolarized molecule to a molecule of interest, and (f) using the molecule of interest in an NMR or MRI experiment.

[0146] Embodiment 34. (e) The composition of embodiment 33, comprising transferring nuclear spin polarization from a highly polarized molecule to a molecule of interest by a Parahydrogen Induced Polarization Nuclear Overhauser Effect System (PHIPNOESYS) procedure.

[0147] Embodiment 35.(e) The composition of embodiment 33, comprising transferring nuclear spin polarization from a highly polarized molecule to a molecule of interest by parahydrogen-induced polarization relayed via a proton exchange (PHIP-X) procedure.

[0148] Embodiment 36. The composition of embodiment 34 or 35, wherein the molecule of interest comprises a biologically relevant imaging agent.

[0149] Embodiment 37. The composition of embodiment 36, wherein the biologically relevant imaging agent is selected from the group consisting of pyruvate, glutamate, glutamine, lactate, acetate, acetoacetate, zymonate, alanine, fructose, fumarate, bicarbonate, urea, dehydroascorbate, alpha-ketoglutarate, dihydroxyacetone, glucose, ascorbate, and their conjugate acids.

[0150] Embodiment 38. The composition of any one of embodiments 21 to 37, wherein the molar polarization is composed of the product of the concentration of at least one nucleus and the nuclear spin polarization of at least one nucleus.

[0151] Embodiment 39. The molar polarization is at least 60 mM, 70 mM, 80 mM, 90 mM, 100 mM, 110 mM, 120 mM, 130 mM, 140 mM, 150 mM, 160 mM, 170 mM, 180 mM, 190 mM, 200 mM, 210 mM, 220 mM, 230 mM, 240 mM, 250 mM, 260 mM, 270 mM, 280 mM, 290 mM, 300 mM, 310 mM, 320 mM, 330 mM, 340 mM, 350 mM, 360 mM, 370 mM, 380 mM, 390 mM, 400 mM, 410 mM, 420 mM, 430 mM, 440 mM, 450 mM, 460 mM, 470 mM, 480 mM, 490 mM, 500 mM, 510 mM, 520 mM, 530 mM, 540 mM, 550 mM, 560 mM, 570 mM, 580 mM, 590 mM, 600 mM, 610 mM, 620 mM, 630 mM, 640 mM, 650 mM, 660 mM, 670 mM, 680 mM, 690 mM, 700 mM, 710 mM, 720 mM, 730 mM, 740 mM, 750 mM, 760 mM, 770 mM, 780 mM, 790 mM, 800 mM, 810 mM, 820 mM, 830 mM, 840 mM, 850 mM, 860 mM, 870 mM, 880 mM, 890 mM 90mM, 300mM, 310mM, 320mM, 330mM, 340mM, 350mM, 360mM, 370mM, 380mM, 390mM, 400mM, 410mM, 420mM, 430mM, 440mM, 450mM, 460mM, 470mM, 480mM, 490mM, 500mM, 510mM, 520mM, 530mM, 540mM , 550mM, 560mM, 570mM, 580mM, 590mM, 600mM, 610mM, 620mM, 630mM, 640mM, 650mM, 660mM, 670m M, 680mM, 690mM, 700mM, 710mM, 720mM, 730mM, 740mM, 750mM, 760mM, 770mM, 780mM, 790mM, 800 39. The composition of any one of embodiments 21-38, wherein the HCl concentration is 810 mM, 820 mM, 830 mM, 840 mM, 850 mM, 860 mM, 870 mM, 880 mM, 890 mM, 900 mM, 910 mM, 920 mM, 930 mM, 940 mM, 950 mM, 960 mM, 970 mM, 980 mM, 990 mM, or 1,000 mM.

[0152] Embodiment 40. At least one nucleus has at least one proton ( 1 H), carbon-13( 13 C), nitrogen-15( 15 N), fluorine-19( 19 F), or phosphate-31( 31 P) a composition according to any one of embodiments 21 to 39, comprising a core.

Claims

1. 1. A method comprising: (a) obtaining a solution having hyperpolarized molecules dissolved therein, the hyperpolarized molecules comprising at least one nucleus having a molar polarization of at least 50 millimolar (mM).

2. 10. The method of claim 1, further comprising, prior to (a), performing a nuclear spin hyperpolarization protocol on the hyperpolarized molecule, thereby imparting the molar polarization to the at least one nucleus.

3. the nuclear spin hyperpolarization protocol comprises: (b) obtaining a solution containing a derivative of said hyperpolarized molecule, said derivative containing at least one unsaturated carbon-carbon double bond or unsaturated carbon-carbon triple bond and having the form R1-C=C-R2 or R1-C≡C-R2, wherein R1 and R2 contain side chains; (c) hydrogenating the double or triple bond with para-hydrogen to form a para-hydrogenated derivative of the hyperpolarized molecule, wherein the para-hydrogenated derivative is of the form R1-CH ＊ -CH ＊ -R2 or R1-CH ＊ =CH ＊ -R2, H ＊ represents a para-hydrogen-derived hydrogen atom attached across said double or triple bond; (d) applying a polarization transfer waveform to transfer nuclear spin order from at least one of the para-hydrogen-derived hydrogen atoms to the at least one nucleus, thereby imparting nuclear spin hyperpolarization to the at least one nucleus.

4. The method of claim 3 , wherein the polarization transfer waveform is configured to suppress a dipolar field associated with magnetization generated during the accumulation of the nuclear spin hyperpolarization.

5. 5. The method of claim 3 or 4, wherein the polarization transfer waveform comprises a bipolar decoupling sequence.

6. 6. The method of claim 4 or 5, wherein the polarization transfer waveform further comprises a coupled driving field based on a parameter sweep.

7. The parameter sweep is performed by measuring the transverse magnetic field (B １ ) sweep.

8. The polarization transfer waveform is B in the Lee Goldberg frame. １ The method of any one of claims 3 to 7, comprising a sweep.

9. The method of any one of claims 3 to 7, wherein the polarization transfer waveform comprises a pulse sequence selected from the group consisting of polarized MREV-8, polarized BLEW-12, and polarized BR-24.

10. 10. The method of claim 3, wherein the hyperpolarized molecules are generated by (b) to (d).

11. 10. The method of any one of claims 3 to 9, further comprising: (e) hydrolyzing the para-hydrogenated derivative, thereby forming the hyperpolarized molecule.

12. 12. The method of claim 10 or 11, further comprising using the hyperpolarized molecules in a nuclear magnetic resonance (NMR) or magnetic resonance imaging (MRI) experiment, wherein the hyperpolarized molecules comprise a molecule of interest.

13. 12. The method of claim 10 or 11, further comprising: (f) transferring nuclear spin polarization from the hyperpolarized molecule to a molecule of interest; and (g) using the molecule of interest in an NMR or MRI experiment.

14. 14. The method of claim 13, wherein (f) comprises transferring the nuclear spin polarization from the highly polarized molecule to the molecule of interest by a Parahydrogen Induced Polarization Nuclear Overhauser Effect System (PHIPNOESYS) procedure.

15. 14. The method of claim 13, wherein (f) comprises transferring the nuclear spin polarization from the highly polarized molecule to the molecule of interest by relayed parahydrogen induced polarization via a proton exchange (PHIP-X) procedure.

16. 1. A composition comprising: A composition comprising a solution having hyperpolarized molecules dissolved therein, the hyperpolarized molecules comprising at least one nucleus having a molar polarization of at least 50 millimolar (mM).

17. 17. The composition of claim 16, wherein the hyperpolarized molecule is generated by performing a nuclear spin hyperpolarization protocol on the hyperpolarized molecule, thereby imparting the molar polarization to the at least one nucleus.

18. the nuclear spin hyperpolarization protocol comprises: (a) obtaining a solution comprising a derivative of said hyperpolarized molecule, said derivative comprising at least one unsaturated carbon-carbon double bond or unsaturated carbon-carbon triple bond and having the form R1-C=C-R2 or R1-C≡C-R2, wherein R1 and R2 comprise side chains; (b) hydrogenating the double or triple bond with para-hydrogen to form a para-hydrogenated derivative of the hyperpolarized molecule, wherein the para-hydrogenated derivative is of the form R1-CH ＊ -CH ＊ -R2 or R1-CH ＊ =CH ＊ -R2, H ＊ represents a para-hydrogen-derived hydrogen atom attached across said double or triple bond; (c) applying a polarization transfer waveform to transfer nuclear spin order from at least one of the para-hydrogen-derived hydrogen atoms to the at least one nucleus, thereby imparting nuclear spin hyperpolarization to the at least one nucleus.

19. 20. The composition of claim 18, wherein the polarization transfer waveform is configured to suppress a dipolar field associated with magnetization generated during the accumulation of the nuclear spin hyperpolarization.

20. 20. The composition of claim 18 or 19, wherein the polarization transfer waveform comprises a bipolar decoupling sequence.

21. 21. The composition of claim 19 or 20, wherein the polarization transfer waveform further comprises a coupled driving field based on a parameter sweep.

22. The parameter sweep is performed by measuring the transverse magnetic field (B １ 22. The composition of claim 21, comprising a .) sweep.

23. The polarization transfer waveform is B in the Lee Goldberg frame. １ The composition of any one of claims 18 to 22, comprising a sweep.

24. 23. The composition of any one of claims 18 to 22, wherein the polarization transfer waveform comprises a pulse sequence selected from the group consisting of polarized MREV-8, polarized BLEW-12, and polarized BR-24.

25. 25. The composition of any one of claims 18 to 24, wherein the hyperpolarized molecule is generated by (a) to (c).

26. 25. The composition of any one of claims 18 to 24, wherein the hyperpolarized molecule is further produced by (d) hydrolyzing the para-hydrogenated derivative, thereby forming the hyperpolarized molecule.

27. 27. The composition of claim 25 or 26, wherein the hyperpolarized molecule is for use in a nuclear magnetic resonance (NMR) or magnetic resonance imaging (MRI) experiment, and the hyperpolarized molecule comprises a molecule of interest.

28. 27. The composition of claim 25 or 26, wherein the hyperpolarized molecule is for use in (e) transferring nuclear spin polarization from the hyperpolarized molecule to a molecule of interest, and (f) using the molecule of interest in an NMR or MRI experiment.

29. 30. The composition of claim 28, wherein (e) comprises transferring the nuclear spin polarization from the highly polarized molecule to the molecule of interest by a Parahydrogen Induced Polarization Nuclear Overhauser Effect System (PHIPNOESYS) procedure.

30. 30. The composition of claim 28, wherein (e) comprises transferring the nuclear spin polarization from the highly polarized molecule to the molecule of interest by relayed parahydrogen induced polarization via a proton exchange (PHIP-X) procedure.

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