System and method for generating hyperpolarized materials

Abstract

This disclosure describes hyperpolarized materials for use in nuclear magnetic resonance, magnetic resonance imaging, or similar applications.This disclosure describes methods for producing hyperpolarized materials for use in nuclear magnetic resonance, magnetic resonance imaging, or similar applications.This disclosure describes precursor compounds for use in producing hyperpolarized materials for use in nuclear magnetic resonance, magnetic resonance imaging, or similar applications.

Description

[Technical Field]

[0001] Reference to related applications This application claims priority to U.S. Provisional Patent Application No. 63 / 164,585 filed on 23 March 2021, U.S. Provisional Patent Application No. 63 / 260,631 filed on 27 August 2021, and U.S. Provisional Patent Application No. 63 / 266,986 filed on 21 January 2022, each of which is incorporated herein by reference in whole for all purposes.

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

[0003] Para-hydrogen-induced polarization (PHIP) is a low-cost, high-throughput method for polarization of metabolites for hyperpolarized (HP) magnetic resonance imaging (MRI). Para-hydrogen-induced polarization with a sidearm (PHIP-SAH) can be used to polarize metabolites, such as acetic acid molecules. However, existing PHIP-SAH polarization approaches may not be suitable for preclinical or clinical HP MRI applications. [Overview of the project]

[0004] In some embodiments, the present disclosure relates to a composition comprising a compound of formula (I), [ka] In the formula, Z is (i) 1 H (proton), 2A composition is described that comprises (i) a carbon-carbon double bond (-C=C-) substituted to include H (deuterium), or combinations thereof, or (ii) a carbon-carbon triple bond (-C≡C-), wherein R1 comprises a parahydrogen-induced polarization (PHIP) transfer moiety, R2 comprises an optionally substituted hydrocarbon, alkoxy group, primary amine, secondary amine, or tertiary amine, and R3 comprises a bio-related contrast agent that contains a non-hydrogen nuclear spin. [[ID=]1]

[0005] In some embodiments, the disclosure is a composition comprising a compound of formula (II),

Chemical formula

[0006] In some embodiments, the disclosure is a composition comprising (i) a bio-related contrast agent that contains a non-hydrogen nuclear spin and (ii) a compound of formula (III),

Chemical formula

[0007] In some embodiments, the present disclosure relates to a composition comprising (i) a hyperpolarized bio-relevant contrast agent containing non-hydrogen nuclear spins and (ii) a compound of formula (IV), [ka] In the formula, Z is (i) 1 H (proton), 2 The following describes a composition comprising (ii) a carbon-carbon double bond (-C=C-) substituted to include H (deuterium) or a combination thereof, or (ii) a carbon-carbon triple bond (-C≡C-), wherein R1' comprises a para-hydrogen-induced polarization (PHIP) transfer moiety, and R2 comprises an optionally substituted hydrocarbon, alkoxy group, primary amine, secondary amine, or tertiary amine.

[0008] In some embodiments, this disclosure describes compounds of formula I, II, III, or IV that include a PHIP transfer moiety. In some embodiments, the PHIP transfer moiety includes an optionally substituted C1 hydrocarbon or an optionally substituted C2 hydrocarbon. In some embodiments, the PHIP transfer moiety includes *CR4R5, *CR4Y, *C=Y, or any deuterated version thereof, where *C is 12 C or 13 R4 and R5 are carbon isotopes of C, and they are independent of each other. 1 H, 2 H, 3H is selected from linear, branched, or cyclic C1-C10 alkyl hydrocarbons, C6 aryl, benzyl, phenyl, heteroaryl, and haloalkyl groups, and Y is selected from a spin 1 / 2 atom and a spin 1 / 2 atom covalently bonded to one or more chemical parts selected from linear, branched, or cyclic C1-C10 alkyl hydrocarbons, C6 aryl, benzyl, phenyl, heteroaryl, halogen, or haloalkyl groups, or from heteroatoms such as N, O, S that are optionally substituted with linear, branched, or cyclic C1-C10 alkyl hydrocarbons, C6 aryl, benzyl, phenyl, heteroaryl, halogen, or haloalkyl groups. In some embodiments, the PHIP transfer part includes *CR6R7-*CR8R9, or any deuterated version thereof, and *C is 12 C or 13 It is a carbon isotope of C,

[0009] R6, R7, R8, and R9 are each independent of each other. 1 H, 2 H, 3 H is selected from linear, branched, or cyclic C1-C10 alkyl hydrocarbons, C6 aryls, benzyls, phenyls, heteroaryls, and haloalkyl groups. In some embodiments, the PHIP transfer portion includes *CH2, *CH2-*CH2, *CHY, *C=Y, or any deuterated version thereof, where *C is 12 C or 13 The carbon isotope of C, where Y is selected from a spin-1 / 2 atom and a spin-1 / 2 atom covalently bonded to one or more chemical parts selected from linear, branched, or cyclic C1-C10 alkyl hydrocarbons, C6 aryl, benzyl, phenyl, heteroaryl, halogen, or haloalkyl groups, or from heteroatoms such as N, O, and S that are optionally substituted with linear, branched, or cyclic C1-C10 alkyl hydrocarbons, C6 aryl, benzyl, phenyl, heteroaryl, halogen, or haloalkyl groups. In some embodiments, the spin-1 / 2 atom is 1 H, 13 C, 15 N, 19 F, or 31Selected from P. In some embodiments, the PHIP moving portion includes at least one atom that is J-coupled with a non-hydrogen nuclear spin of at least 0.1 Hz. In some embodiments, Z includes at least one atom that is J-coupled with a non-hydrogen nuclear spin of at least 0.1 Hz.

[0010] In some embodiments, this disclosure describes compounds of formula I, II, III, or IV in which R2 comprises a solubilizing moiety. In some embodiments, R2 comprises a hydrophobic and / or organic affinity moiety. In some embodiments, R2 comprises an organic solubilizing moiety. In some embodiments, R2 comprises a hydrophilic and / or organic affinity moiety. In some embodiments, R2 comprises an optionally substituted hydrocarbon, alkoxy group, primary amine, secondary amine, or tertiary amine. In some embodiments, R2 includes or is selected from a methyl group, an ethyl group, a propyl group, an isopropyl group, an n-butyl group, an s-butyl group, a t-butyl group, an isobutyl group, a hydroxyl group, a methyl alcohol group, an ethyl alcohol group, an n-propanol group, an isopropyl alcohol group, an alcohol propionic acid group, an n-butyl alcohol group, an s-butyl alcohol group, a t-butyl alcohol group, an isobutyl alcohol group, a methoxy group, an ethoxy group, a propoxy group, an isopropoxy group, an alcohol propionic acid group, a butoxy group, a t-butoxy group, an s-butoxy group, an ester group, a phenyl group, a substituted phenyl group, a primary amine group, a secondary amine group, a tertiary amine group, a primary amide group, a secondary amide group, and a tertiary amide group.

[0011] In some embodiments, this disclosure describes compounds of formula I, formula II, formula III, or formula IV, including bio-reactive contrast agents. In some embodiments, the bio-reactive contrast agent is formula R 10 It contains a compound C(=O)X-, in which R 10 X is selected from a linear, branched, or cyclic C1-C10 alkyl group, and one or more C atoms are optionally substituted with C=C, CO, COH, CNH2, COOH, CH2COOH, CONH2, OC(=O), and X is NR 11 Selected from , S, and O, in the formula, R11 teeth, 1 H, 2 H, 3 H is selected from an amino protecting group optionally selected from trifluoroacetyl, acetyl, benzoyl, carbobenzoxy, tert-butyl carbonate, and benzyl. In some embodiments, the bio-contrast agent is selected from pyruvate, glutamate, glutamine, lactate, acetic acid, acetoacetic acid, zymonate, alanine, fructose, fumaric acid, bicarbonate, urea, dehydroascorbic acid, α-ketoglutarate, dihydroxyacetone, glucose, ascorbic acid, and their conjugate acids. In some embodiments, the bio-contrast agent contains pyruvate. In some embodiments, the bio-contrast agent contains lactate. In some embodiments, the bio-contrast agent contains α-ketoglutarate.

[0012] In some embodiments, compositions of formula I, II, III, or IV have a solubility in water of less than 50 mmol (mM). In some embodiments, compositions of formula I, II, III, or IV have a solubility in organic solvents (e.g., acetone, ethanol, chloroform, and toluene) of less than 50 mmol (mM).

[0013] In some embodiments, a composition of formula I reacted with parahydrogen yields a chemical yield of at least 30% of the parahydrogenated product.

[0014] In some embodiments, the compositions of this disclosure are intended for use in para-hydrogen-induced polarization (PHIP) processes.

[0015] In some embodiments, the disclosure describes a method for preparing a hyperpolarized bio-relevant contrast agent or a pharmaceutically acceptable salt thereof. In some embodiments, the method provides (a) a composition comprising a compound of formula (I), [ka] In the formula, Z is (i) 1 H (proton),2 (ii) a carbon-carbon double bond (-C=C-) or a carbon-carbon triple bond (-C≡C-) substituted to include H (deuterium) or a combination thereof, wherein R1 comprises a parahydrogen-induced polarization (PHIP) transfer moiety, R2 comprises an optionally substituted hydrocarbon, alkoxy group, primary amine, secondary amine, or tertiary amine, and R3 comprises a bio-related contrast agent containing a non-hydrogen nuclear spin, and (b) a double bond or triple bond in a compound of formula I is hydrogenated with parahydrogen to form a parahydrogenated derivative of the compound of formula I, wherein the parahydrogenated derivative has the structure of formula (II), [ka] In the formula, Z' is (i) 1 H (proton), 2 (ii) Parahydrogenated carbon-carbon single bonds (-CH*-CH*-) substituted to include H (deuterium) or a combination thereof, or 1 H (proton), 2 (c) A parahydrogenated carbon-carbon double bond (-CH*=CH*-) substituted to include H (deuterium) or a combination thereof, wherein H* is a hydrogen having a spin order derived from parahydrogen, R1 includes a parahydrogen-induced polarization (PHIP) transfer portion, R2 includes an optionally substituted hydrocarbon, alkoxy group, primary amine, secondary amine, or tertiary amine, and R3 includes a bio-relevant contrast agent containing a non-hydrogen nuclear spin; and (c) a polarization transfer waveform is applied to the non-hydrogen nuclear spin to transfer the nuclear spin order from at least one H* in the compound of formula II, thereby forming a derivative of formula II having a hyperpolarized bio-relevant contrast agent.

[0016] In some embodiments, the present disclosure provides a method for preparing a hyperpolarized bio-relevant contrast agent or a pharmaceutically acceptable salt thereof, wherein the method provides a composition comprising (a) formula (II) of a compound, [ka] In the formula, Z' is (i) 1 H (proton), 2 (ii) Parahydrogenated carbon-carbon single bonds (-CH*-CH*-) substituted to include H (deuterium) or a combination thereof, or 1 H (proton), 2 The present invention provides a parahydrogenated carbon-carbon double bond (-CH*=CH*-) substituted to include H (deuterium) or a combination thereof, wherein H* is a hydrogen having a spin order derived from parahydrogen, R1 comprises a parahydrogen-induced polarization (PHIP) transfer portion, R2 comprises an optionally substituted hydrocarbon, alkoxy group, primary amine, secondary amine, or tertiary amine, and R3 comprises a bio-relevant contrast agent containing a non-hydrogen nuclear spin; and the present invention describes a method comprising (b) applying a polarization transfer waveform to the non-hydrogen nuclear spin to transfer the nuclear spin order from at least one H* in the compound of formula II, thereby forming a derivative of formula II having a hyperpolarized bio-relevant contrast agent.

[0017] In some embodiments, the method involves hydrolyzing a derivative of formula II to obtain (i) a hyperpolarized bio-relevant contrast agent containing non-hydrogen nuclear spins, and (ii) a compound of formula (III), [ka] In the formula, Z'' is (i) 1 H (proton), 2 (ii) Parahydrogenated carbon-carbon single bonds (-CH*-CH*-) substituted to include H (deuterium) or a combination thereof, or 1 H (proton), 2The present invention further provides a composition comprising a compound comprising a parahydrogenated carbon-carbon double bond (-CH*=CH*-) substituted to include H (deuterium) or a combination thereof, wherein R1' comprises a parahydrogen-induced polarization (PHIP) transfer moiety, and R2 comprises an optionally substituted hydrocarbon, alkoxy group, primary amine, secondary amine, or tertiary amine. In some embodiments, the method further comprises washing the hyperpolarized bio-related contrast agent once or twice with an organic solvent. In some embodiments, the non-hydrogen nuclear spins have more than 10% non-hydrogen nuclear spin polarization after the washing step.

[0018] In some embodiments, the Disclosure describes hyperpolarized bio-relevant contrast agents or pharmaceutically acceptable salts thereof produced by the methods of the Disclosure.

[0019] It should be understood that both the general description above and the detailed description below are illustrative and explanatory, and do not limit the disclosed embodiments as requested. [Brief explanation of the drawing]

[0020] The accompanying drawings, including portions of this specification, illustrate some embodiments and, together with this specification, serve to illustrate certain principles and features of the disclosed embodiments. In the drawings,

[0021] [Figure 1] A first exemplary process for generating polarized bio-related contrast agents is shown, according to various embodiments. [Figure 2] A second exemplary process for generating polarized bio-related contrast agents is shown, according to various embodiments. [Figure 3A] Exemplary proton (1H) nuclear magnetic resonance (NMR) spectra corresponding to 4-((2-oxopropanoyl)oxy)buta-2-inoatemethyl are shown in various embodiments. [Figure 3B] Exemplary 13C NMR spectra corresponding to 4-((2-oxopropanoyl)oxy)buta-2-inoatemethyl in various embodiments are shown. [Figure 4A] Exemplary 1H NMR spectra corresponding to isopropyl 4-((2-oxopropanoyl)oxy)buta-2-inoate in various embodiments are shown. [Figure 4B] Exemplary 13C NMR spectra corresponding to isopropyl 4-((2-oxopropanoyl)oxy)buta-2-inoate in various embodiments are shown. [Figure 5A] Exemplary 1H NMR spectra corresponding to tert-butyl 4-((2-oxopropanoyl)oxy)buta-2-inoate in various embodiments are shown. [Figure 5B] Exemplary 13C NMR spectra corresponding to tert-butyl 4-((2-oxopropanoyl)oxy)buta-2-inoate in various embodiments are shown. [Figure 6A] Exemplary 1H NMR spectra corresponding to tert-butyl 4-((2-oxopropanoyl-1-13C)oxy)buta-2-inoate in various embodiments are shown. [Figure 6B] Exemplary 13C NMR spectra corresponding to tert-butyl 4-((2-oxopropanoyl-1-13C)oxy)buta-2-inoate in various embodiments are shown. [Figure 7A] Exemplary 1H NMR spectra corresponding to 2-(methyl-d3)propan-2-yl-1,1,1,3,3,3-d64-((2-oxopropanoyl-1-13C)oxy)buta-2-inoate in various embodiments are shown. [Figure 7B] Exemplary 13C NMR spectra corresponding to 2-(methyl-d3)propan-2-yl-1,1,1,3,3,3-d64-((2-oxopropanoyl-1-13C)oxy)buta-2-inoate in various embodiments are shown. [Figure 8A] Exemplary 1H NMR spectra corresponding to tert-butyl 4-((2-oxopropanoyl)oxy)buta-2-inoate-4-d in various embodiments are shown. [Figure 8B]Exemplary 13C NMR spectra corresponding to tert-butyl 4-((2-oxopropanoyl)oxy)buta-2-inoate-4-d in various embodiments are shown. [Figure 9A] Exemplary 1H NMR spectra corresponding to tert-butyl 4-((2-oxopropanoyl)oxy)penta-2-inoate in various embodiments are shown. [Figure 9B] Exemplary 13C NMR spectra corresponding to tert-butyl 4-((2-oxopropanoyl)oxy)penta-2-inoate in various embodiments are shown. [Figure 10A] Exemplary 1H NMR spectra corresponding to tert-butyl 4-((2-oxopropanoyl)oxy)-4-phenylbuta-2-inoate in various embodiments are shown. [Figure 10B] Exemplary 13C NMR spectra corresponding to tert-butyl 4-((2-oxopropanoyl)oxy)-4-phenylbuta-2-inoate in various embodiments are shown. [Figure 11] Exemplary 1H NMR spectra corresponding to benzhydryl 4-((2-oxopropanoyl)oxy)buta-2-inoate in various embodiments are shown. [Figure 12A] Exemplary 1H NMR spectra corresponding to 4-oxo-4-phenylbuta-2-in-1-yl-2-oxopropanoate in various embodiments are shown. [Figure 12B] Exemplary 13C NMR spectra corresponding to 4-oxo-4-phenylbuta-2-in-1-yl-2-oxopropanoate in various embodiments are shown. [Figure 13A] Exemplary 1H NMR spectra corresponding to 4-oxo-4-(phenyl-d5)buta-2-in-1-yl-2-oxopropanoate in various embodiments are shown. [Figure 13B] Exemplary carbon-13 (13C) NMR spectra corresponding to 4-oxo-4-(phenyl-d5)buta-2-in-1-yl-2-oxopropanoate in various embodiments are shown. [Figure 14A]Exemplary 1H NMR spectra corresponding to 1-(4-(tert-butoxy)-4-oxobuta-2-in-1-yl)5-ethyl 2-oxopentanedioate in various embodiments are shown. [Figure 14B] Exemplary 13C NMR spectra corresponding to 1-(4-(tert-butoxy)-4-oxobuta-2-in-1-yl)5-ethyl 2-oxopentanedioate in various embodiments are shown. [Figure 15A] Exemplary 1H NMR spectra corresponding to 4-(2,2-dichloroacetoxy)buta-2-inoatemethyl are shown for various embodiments. [Figure 15B] Exemplary 13C NMR spectra corresponding to various embodiments of 4-(2,2-dichloroacetoxy)buta-2-inoatemethyl are shown. [Figure 16A] Exemplary 1H NMR spectra corresponding to tert-butyl 4-acetoxybuta-2-inoate in various embodiments are shown. [Figure 16B] Exemplary 13C NMR spectra corresponding to tert-butyl 4-acetoxybuta-2-inoate in various embodiments are shown. [Figure 17A] Exemplary 1H NMR spectra corresponding to parahydrogenated 3-phenylpropa-2-in-1-yl 2-oxopropanoates (i.e., 3-phenylallyl 2-oxopropanoates having two protons H* with spin order derived from parahydrogen) are shown for various embodiments. [Figure 17B] Exemplary 1H NMR spectra corresponding to para-hydrogenated tert-butyl 4-((2-oxopropanoyl)oxy)buta-2-inoate (i.e., tert-butyl 4-((2-oxopropanoyl)oxy)buta-2-enoate having two H* are shown in various embodiments. [Figure 18A] Exemplary 1H NMR spectra corresponding to para-hydrogenated 3-phenylpropa-2-in-1-yl 2-oxopropanoates (i.e., 3-phenylallyl 2-oxopropanoates having two H*) are shown for various embodiments. [Figure 18B] Exemplary 1H NMR spectra corresponding to para-hydrogenated 4-oxo-4-phenylbuta-2-in-1-yl 2-oxopropanoates (i.e., 4-oxo-4-phenylbuta-2-en-1-yl 2-oxopropanoates having two H*) are shown in various embodiments. [Figure 19A] Exemplary 13C NMR spectra corresponding to 200 mM para-hydrogenated 3-phenylpropa-2-in-1-yl 2-oxopropanoate (i.e., 3-phenylallyl 2-oxopropanoate having two H*) are shown for various embodiments. [Figure 19B] Exemplary 13C NMR spectra corresponding to 200 mM para-hydrogenated tert-butyl 4-((2-oxopropanoyl)oxy)buta-2-inoate (i.e., tert-butyl 4-((2-oxopropanoyl)oxy)buta-2-enoate having two H* are shown in various embodiments. [Figure 20A] Exemplary 13C NMR spectra corresponding to 200 mM para-hydrogenated 3-phenylpropa-2-in-1-yl 2-oxopropanoate (i.e., 3-phenylallyl 2-oxopropanoate having two H*) are shown for various embodiments. [Figure 20B] Exemplary 13C NMR spectra corresponding to 200 mM para-hydrogenated 4-oxo-4-phenylbuta-2-in-1-yl 2-oxopropanoate (i.e., 4-oxo-4-phenylbuta-2-en-1-yl 2-oxopropanoate having 2 H*) are shown for various embodiments. [Figure 21] Exemplary 13C NMR spectra corresponding to 133 mM para-hydrogenated methyl 4-((2-oxopropanoyl)oxy)buta-2-inoate (i.e., methyl 4-((2-oxopropanoyl)oxy)buta-2-enoate having 2 H* are shown in various embodiments. [Figure 22A]Exemplary 1H NMR spectra corresponding to para-hydrogenated 1-(4-(tert-butoxy)-4-oxobuta-2-in-1-yl)5-ethyl 2-oxopentanedioate (i.e., 1-(4-(tert-butoxy)-4-oxobuta-2-in-1-yl)5-ethyl 2-oxopentanedioate having 2 H* are shown in various embodiments. [Figure 22B] Exemplary 13C NMR spectra corresponding to 70 mM para-hydrogenated 1-(4-(tert-butoxy)-4-oxobuta-2-en-1-yl)5-ethyl 2-oxopentanedioate (i.e., 1-(4-(tert-butoxy)-4-oxobuta-2-en-1-yl)5-ethyl 2-oxopentanedioate having 2 H* are shown in various embodiments. [Modes for carrying out the invention]

[0022] Herein, exemplary embodiments are described in detail and discussed with respect to the accompanying drawings. Unless otherwise defined, technical and / or scientific terms have the meanings generally 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. Naturally, other embodiments may be used and modifications may be made without departing from the scope of the disclosed embodiments. Thus, the materials, methods and examples are illustrative and not necessarily intended to be limiting.

[0023] Recent research in the fields of nuclear magnetic resonance (NMR) and magnetic resonance imaging (MRI) has demonstrated that NMR and MRI signals associated with various bio-reactive contrast agents can be dramatically enhanced using various so-called hyperpolarization techniques. This signal enhancement allows for improved spectroscopic analysis of bio-reactive contrast 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 non-invasive determination of the health status of tissues within the body. For example, abnormal metabolism of bio-reactive contrast agents may indicate diseases such as cancer at several locations within the body.

[0024] Existing techniques for hyperpolarized bio-relevant contrast agents include dissolution-dynamic nuclear polarization (DNP), para-hydrogen-induced polarization (PHIP), PHIP-SAH side-chain hydrogenation (PHIP-SAH), and reversible exchange-mediated signal amplification (SABRE). In PHIP-SAH, a precursor of the bio-relevant contrast agent is reacted with para-hydrogen to form a para-hydrogenated derivative of the precursor. The spin order is then transferred from the added proton via the para-hydrogenation reaction to the target nucleus (e.g., a carbon-13 nucleus) contained within the bio-relevant contrast agent. The para-hydrogenated derivative of the precursor is cleaved (e.g., hydrolyzed) to obtain the hyperpolarized bio-relevant contrast agent. The bio-relevant contrast agent is then purified and used in NMR or MRI procedures. In some embodiments, the precursor may include a bio-relevant contrast agent coupled to a side arm containing at least one unsaturated bond (e.g., at least one carbon-carbon double bond or at least one carbon-carbon triple bond) suitable for reaction with para-hydrogen. However, previous precursors have used side effects that may not enable the production of biocompatible contrast agents with clinically relevant polarization, concentration, volume, or purity. Such behavior may be related to poor solubility of the precursor in organic solvents (when para-hydrogen is highly soluble), poor yield in the reaction between the unsaturated bond and para-hydrogen, or various other factors. Therefore, there is a need for novel PHIP-SAH precursors that produce hyperpolarized biocompatible contrast agents with clinically relevant polarization, concentration, volume, or purity.

[0025] The disclosed embodiments include systems and methods for producing bio-related contrast agents with clinically significant polarization, concentration, volume, and purity. The disclosed embodiments provide technical improvements to polarized bio-related contrast agents in solution. These technical improvements support increases in bio-related contrast agent concentration and degree of bio-related polarization.

[0026] Hyperpolarization and parahydrogen As used in this disclosure, hyperpolarization describes a state in which the absolute difference between a spin state (e.g., a nuclear spin state, a proton spin state, etc.) in one state (e.g., spin-up) and a group of spin states (e.g., spin-down) exceeds the absolute difference of the corresponding difference in thermal equilibrium.

[0027] Parahydrogen can be used as a polarization source, consistent with the 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 specific method for producing parahydrogen. Parahydrogen can be formed in gaseous or liquid form. In some embodiments, parahydrogen is produced in gaseous form by flowing hydrogen gas through a chamber at a low temperature using a catalyst (e.g., iron oxide or another suitable catalyst). The hydrogen gas may contain both parahydrogen and orthohydrogen. The low temperature can cause the hydrogen gas to reach thermodynamic equilibrium in the chamber, increasing the population of parahydrogen.

[0028] The disclosed embodiments are not limited to a specific parahydrogen production or use location. Parahydrogen may be produced at a first location and then transported to a second location for use. In some embodiments, the first location may be a chamber, which may be part of a container, bottle, holder, or other area capable of holding a gas or liquid. Such a chamber may be maintained at a suitable pressure or temperature. In some embodiments, the first location may be a physical location such as a room, laboratory, a specific warehouse, a hospital, or other location where parahydrogen is produced.

[0029] The disclosed embodiments are not limited to any particular parahydrogen transport method. The generated parahydrogen may be transported in a chamber that may be different from the chamber in which the parahydrogen was generated. The chamber in which the parahydrogen gas is transported may be maintained at a suitable pressure or temperature that can be transported by vehicle or by person. Transporting parahydrogen may involve moving parahydrogen from one container to a different container. Transporting parahydrogen may involve moving parahydrogen within the same location, such as from one part of a room to another part of a room. Transporting parahydrogen may involve moving parahydrogen from one room in a building to a different room in the same building, or to a nearby building. Transporting parahydrogen may involve moving parahydrogen to a different location in a different part of the same city, or to a different city. Transporting parahydrogen may involve transporting parahydrogen near a polarizer, NMR device, or MRI device. Transporting parahydrogen may involve packaging or transporting parahydrogen in a suitable container.

[0030] In some embodiments, the collective difference between two spin states is the difference between the two spin states divided by the total collective of the two spin states. The collective difference can be expressed as a fractional collective difference or a percentage collective difference. In some embodiments, the fractional collective difference is at least about 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9 or higher, and at most about 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, 0.1 or lower, or within the range defined by any two of the aforementioned values.

[0031] Hydrogen gas can exhibit a collective difference between proton spin states that significantly exceeds the collective difference between proton spin states in thermal equilibrium. Parahydrogen can have a large collective difference between either the singlet spin state or the triplet spin state. For example, in the case of Iz1Iz2, there is a large collective difference between the spin states |↑>|↓> and |↑>|↑>. The collective difference of proton spin states can be at least about 0.1 (e.g., a 10% difference in spin states - 55% of 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 higher, and at most about 0.9, 0.8, 0.7, 0.6, 0.5, 0.4, 0.3, 0.2, 0.1, or lower, or within the range defined by any two of the aforementioned values.

[0032] Biomedical contrast agents The disclosed embodiments include systems and methods for producing and utilizing bio-relevant contrast agents with clinically significant polarization, concentration, volume, or purity. In some embodiments, the methods are for preparing NMR materials. In some embodiments, the NMR materials are suitable for use in NMR or MRI operations. In some embodiments, the NMR materials increase the NMR or MRI signal and signal-to-noise ratio (SNR). 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., bio-relevant molecules such as amino acids, sugars, and their derivatives) 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 requiring high signal enhancement due to hyperpolarization, such as proton exchange between water and biomolecules. In some embodiments, the NMR materials are small molecules or metabolites suitable for injection into cells, tissues, or organisms for detection in MRI scans. In some embodiments, the NMR material is introduced into the chamber for further analysis by NMR or MRI operation. In some embodiments, the NMR material contains one or more deuterium ( 2 H) or carbon-13 ( 13 It is concentrated by C atoms.

[0033] In accordance with the disclosed embodiments, the NMR material may include a bio-related contrast agent. In some embodiments, the bio-related contrast agent may be suitable for use in NMR or MRI operations. In some embodiments, the bio-related contrast agent may increase the NMR or MRI signal or signal-to-noise ratio (SNR). In some embodiments, the bio-related contrast agent may be suitable for use in solution NMR spectroscopy. In some embodiments, the bio-related contrast agent may be a metabolite (e.g., a bio-related molecule such as an amino acid, sugar, or its derivative) suitable for use in NMR metabolomics applications. In some embodiments, the bio-related contrast agent is used for perfusion imaging or contrast enhancement imaging in MRI scans. In some embodiments, the bio-related contrast agent may be suitable for in vitro probing of the metabolism of cell cultures or other biological tissues. In some embodiments, the bio-related contrast agent is used for in vitro probing of the metabolism of cell cultures or other biological tissues. In some embodiments, the bio-related contrast agent may be used in NMR probes to investigate transient effects requiring high signal enhancement due to hyperpolarization, such as proton exchange between water and biomolecules. In some embodiments, the bio-relevant contrast agent may be a small molecule or metabolite suitable for injection into cells, tissues, or organisms for detection in MRI scans. In some embodiments, the bio-relevant contrast agent may be introduced into a chamber for further analysis by NMR or MRI operation. In some embodiments, the bio-relevant contrast agent may be one or more 2 H or 13 It is concentrated by carbon atoms.

[0034] In some embodiments, the bio-relevant contrast agent is pyruvate, lactic acid, α-ketoglutaric acid, bicarbonate, fumaric acid, urea, dehydroascorbic acid, glutamic acid, glutamine, acetic acid, dihydroxyacetone, acetoacetic acid, glucose, ascorbic acid, zymonate, alanine, fructose, imidazole, nicotinamide, nitroimidazole, pyrazinamide, isoniazid, any of the above conjugate acids, natural and unnatural amino acids, their esters, or any of the above. 2 H, 13 C, or nitrogen-15 ( 15N) includes rich versions. In some embodiments, the bio-reactive contrast agent comprises pyruvate, lactate, and α-ketoglutaric acid. In some embodiments, the bio-reactive contrast agent comprises pyruvate. In some embodiments, the bio-reactive contrast agent comprises lactate. In some embodiments, the bio-reactive contrast agent comprises α-ketoglutaric acid (e.g., ethyl α-ketoglutaric acid).

[0035] In some embodiments, the bio-relevant contrast agent includes at least one non-hydrogen nucleus spin. In some embodiments, the non-hydrogen nucleus includes at least one spin 1 / 2 atom. In some embodiments, the non-hydrogen nucleus spin is 13 C or 15 Contains N. In some embodiments, the bio-related contrast agent is at least partially isotope-labeled with non-hydrogen nuclear spin. In some embodiments, the bio-related contrast agent is at least partially enriched with non-hydrogen nuclear spin compared to analogs of the bio-related contrast agent characterized by its natural abundance of non-hydrogen nuclear spin. In some embodiments, the bio-related contrast agent is enriched with 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%, The non-hydrogen nucleus spins are enriched to characterize them 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 within the range defined by any two of the aforementioned values.

[0036] In some embodiments, non-hydrogen nuclear spins are NMR-inactive (i.e., spin 0) nuclei of bio-related contrast agent analogs characterized by their natural abundance (e.g., 12 C or quadropolar (i.e., spin > 1 / 2) nuclei (e.g., nitrogen-14) 14Substitute N). For example, with its natural abundance 13 The pyruvate analogs that characterize C are those with C* in any of the structures H3C-C*(=O)-C*OOH, which account for approximately 98.9%. 12 C and approximately 1.1% 13 It may contain C. As a bio-relevant contrast agent, pyruvate contains one or both C* in any amount described herein. 13 To include C, instead 13 It can be isotope enriched with C. When used herein, *C and C* are 12 C or 13 This describes carbon that can be any of the carbon isotopes. As another example, in its natural abundance... 15 The urea analogues that characterize N are N* in one of the structures H2N*-C(=O)-*NH2, which accounts for approximately 99.6%. 14 N and approximately 0.4% 15 It may contain N. As a bio-relevant contrast agent, urea may contain one or both N* in any amount described herein. 15 Instead, include N 15 It can be isotope enriched with N. Where used herein, *N and N* are, 14 N or 15 Describes nitrogen, which can be any of the nitrogen isotopes of N.

[0037] Biomedical contrast agent precursors In some embodiments, this disclosure describes a precursor (i.e., a precursor compound) comprising a bio-relevant contrast agent and a side arm. In some embodiments, the bio-relevant contrast agent is covalently bonded to the side arm. In some embodiments, the bio-relevant contrast agent is attached to the side arm via a moving part, such as a PHIP moving part, which is part of the side arm. The side arm may be para-hydrogenated using para-hydrogen (e.g., by mixing the precursor and para-hydrogen). In some embodiments, the hydrogenation produces an Iz1Iz2 order, which is a lower energy state between |↑>|↓> and |↓>|↑>, or a singlet spin order of two hydrogen spins, depending on whether the hydrogenation is performed in a low or high magnetic field.

[0038] In some embodiments, the precursor is selected after hydrogenation and other optional chemical reactions to make the bio-relevant contrast agent suitable for use in NMR or MRI applications where it is hyperpolarized. In some embodiments, additional chemical reactions after hydrogenation can be used to separate the bio-relevant contrast agent from the precursor. Such additional chemical reactions may include, for example, cleavage of the side arms of the precursor by hydrolysis. For example, the bio-relevant contrast agent can be a metabolite molecule such that the precursor can be a derivative of a metabolite molecule, the derivative having the general chemical structure of formula I. The bio-relevant contrast agent can be polarized using the PHIP-SAH method (i.e., para-hydrogenation of the side arms and subsequent polarization transfer to the bio-relevant contrast agent). After hydrogenation and polarization transfer, the bonds in the precursor (e.g., ester bonds) may be hydrolyzed to produce the polarized bio-relevant contrast agent and separate side arm elements.

[0039] As used herein, hydrolysis is defined as the cleavage of a molecule via a nucleophilic substitution reaction involving the addition of the element water. Hydrolysis may also be carried out under anhydrous conditions in the presence of hydroxide ions.

[0040] In accordance with the disclosed embodiments, precursors in the general chemical form presented in Formula I can be used as precursors for PHIP-SAH. After hydrogenation of such precursors, two spin-ordered compounds 1 The H spin is close to the target carbon or nitrogen on the metabolite (e.g., only 3, 4, or 5 bonds away), as described herein. 13 C concentration or 15 N can be enriched. In some embodiments, 13 C or 15 N-spin and parahydrogen derived 1High J-coupling is achieved between at least one of the H spins. In some embodiments, J-coupling is achieved at least about 0.1 Hz, 0.2 Hz, 0.3 Hz, 0.4 Hz, 0.5 Hz, 0.6 Hz, 0.7 Hz, 0.8 Hz, 0.9 Hz, 1 Hz, 2 Hz, 3 Hz, 4 Hz, 5 Hz, 6 Hz, 7 Hz, 8 Hz, 9 Hz, 10 Hz or higher, and at most about 10 Hz, 9 Hz, 8 Hz, 7 Hz, 6 Hz, 5 Hz, 4 Hz, 3 Hz, 2 Hz, 1 Hz, 0.9 Hz, 0.8 Hz, 0.7 Hz, 0.6 Hz, 0.5 Hz, 0.4 Hz, 0.3 Hz, 0.2 Hz, 0.1 Hz or lower, or within the range defined by any two of the aforementioned values. For example, in some embodiments, J-coupling is used for frequencies of 1Hz~2Hz, 1Hz~3Hz, 1Hz~4Hz, 1Hz~5Hz, 1Hz~6Hz, 1Hz~7Hz, 1Hz~8Hz, 1Hz~9Hz, 1Hz~10Hz, 2Hz~3Hz, 2Hz~4Hz, 2Hz~5Hz, 2Hz~6Hz, 2Hz~7Hz, 2Hz~8Hz, 2Hz~9Hz, 2Hz~10Hz, 3Hz~4Hz, 3Hz~5Hz, 3Hz~6Hz, 3Hz~7Hz, 3Hz~8Hz These are Hz, 3Hz~9Hz, 3Hz~10Hz, 4Hz~5Hz, 4Hz~6Hz, 4Hz~7Hz, 4Hz~8Hz, 4Hz~9Hz, 4Hz~10Hz, 5Hz~6Hz, 5Hz~7Hz, 5Hz~8Hz, 5Hz~9Hz, 5Hz~10Hz, 6Hz~7Hz, 6Hz~8Hz, 6Hz~9Hz, 6Hz~10Hz, 7Hz~8Hz, 7Hz~9Hz, 7Hz~10Hz, 8Hz~9Hz, 8Hz~10Hz, or 9Hz~10Hz. Such J-coupling is, 13 This could enable efficient polarization of C-spins.

[0041] This specification refers to the compounds of formulas I, II, III, and IV, their tautomers, deuterated derivatives of their compounds, and their tautomers, their salts, and at one or more sites within the molecule. 13 C or 15N-enriched derivatives (which can then be subjected to hyperpolarization), and novel precursors are disclosed, including the subsequent generation of precursors given by general formulas I, II, III, and IV.

[0042] Precursors of formula I In some embodiments, the precursor comprises a compound of formula I. Formula I encompasses the following structure.

Chemical formula

[0043] In some embodiments, the compound of formula I has a solubility in water of at least about 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 6 mM, 7 mM, 8 mM, 9 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, 100 mM, 150 mM, 200 mM, 250 mM, 300 mM, 350 mM, 400 mM, 450 mM, 500 mM, 550 mM, 600 mM, 650 mM, 700 mM, 750 mM, 800 mM, 850 mM, 900 mM, 950 mM, 1,000 mM, or more, up to about 1,000 mM. It has a solubility in water of 0mM, 950mM, 900mM, 850mM, 800mM, 750mM, 700mM, 650mM, 600mM, 550mM, 500mM, 450mM, 400mM, 350mM, 300mM, 250mM, 200mM, 150mM, 100mM, 90mM, 80mM, 70mM, 60mM, 50mM, 40mM, 30mM, 20mM, 10mM, 9mM, 8mM, 7mM, 6mM, 5mM, 4mM, 3mM, 2mM, 1mM or less, or a solubility in water that is within the range defined by any two of the aforementioned values.

[0044] In some embodiments, the compound of formula I is used in an organic solvent (e.g., acetone, ethanol, chloroform, toluene) at a concentration of at least about 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 6 mM, 7 mM, 8 mM, 9 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, 100 mM, 150 mM, 200 mM, 250 mM, 300 mM, 350 mM, 400 mM, 450 mM, 500 mM, 550 mM, 600 mM, 650 mM, 700 mM, 750 mM, 800 mM, 850 mM, 900 mM, 950 mM, 1,000 mM, or more. It has a solubility in organic solvents of up to approximately 1,000 mM, 950 mM, 900 mM, 850 mM, 800 mM, 750 mM, 700 mM, 650 mM, 600 mM, 550 mM, 500 mM, 450 mM, 400 mM, 350 mM, 300 mM, 250 mM, 200 mM, 150 mM, 100 mM, 90 mM, 80 mM, 70 mM, 60 mM, 50 mM, 40 mM, 30 mM, 20 mM, 10 mM, 9 mM, 8 mM, 7 mM, 6 mM, 5 mM, 4 mM, 3 mM, 2 mM, 1 mM, or less, or a solubility in organic solvents defined by any two of the aforementioned values.

[0045] In some embodiments, the compound of formula I comprises 4-((2-oxopropanoyl)oxy)buta-2-inoatemethyl. In some embodiments, the compound of formula I comprises 4-((2-hydroxypropanoyl)oxy)buta-2-inoatemethyl. In some embodiments, the compound of formula I comprises 5-((4-methoxy-4-oxobuta-2-in-1-yl)oxy)-4,5-dioxopentanoic acid.

[0046] In some embodiments, the compound of formula I comprises isopropyl 4-((2-oxopropanoyl)oxy)buta-2-inoate. In some embodiments, the compound of formula I comprises isopropyl 4-((2-hydroxypropanoyl)oxy)buta-2-inoate. In some embodiments, the compound of formula I comprises 5-((4-isopropoxy-4-oxobuta-2-in-1-yl)oxy)-4,5-dioxopentanoic acid.

[0047] In some embodiments, the compound of formula I comprises tert-butyl 4-((2-oxopropanoyl)oxy)but-2-ynoate. In some embodiments, the compound of formula I comprises tert-butyl 4-((2-hydroxypropanoyl)oxy)but-2-ynoate. In some embodiments, the compound of formula I comprises 5-((4-(tert-butoxy)-4-oxobut-2-yn-1-yl)oxy)-4,5-dioxopentanoic acid.

[0048] In some embodiments, the compound of formula I comprises tert-butyl 4-((2-oxopropanoyl-1- 13 C)oxy)but-2-ynoate. In some embodiments, the compound of formula I comprises tert-butyl 4-((2-hydroxypropane-1- 13 C l)oxy)but-2-ynoate. In some embodiments, the compound of formula I comprises 5-((4-(tert-butoxy)-4-oxobut-2-yn-1-yl)oxy)-4,5-dioxopentanoic acid 13 C acid.

[0049] In some embodiments, the compound of formula I comprises 2-(methyl-d3)propan-2-yl-1,1,1,3,3,3-d6 4-((2-oxopropanoyl-1-13C)oxy)but-2-ynoate. In some embodiments, the compound of formula I comprises 2-(methyl-d3)propan-2-yl-1,1,1,3,3,3-d6 4-((2-hydroxypropanoyl-1-13C)oxy)but-2-ynoate. In some embodiments, the compound of formula I comprises 5-((4-((2-(methyl-d3)propan-2-yl-1,1,1,3,3,3-d6)oxy)-4-oxobut-2-yn-1-yl)oxy)-4,5-dioxopentanoic acid.

[0050] In some embodiments, the compound of formula I comprises tert-butyl4-((2-oxopropanoyl)oxy)buta-2-inoate-4-d. In some embodiments, the compound of formula I comprises tert-butyl4-((2-hydroxypropanoyl)oxy)buta-2-inoate-4-d. In some embodiments, the compound of formula I comprises 5-((4-(tert-butoxy)-4-oxobuta-2-in-1-yl-1-d)oxy)-4,5-dioxopentanoic acid.

[0051] In some embodiments, the compound of formula I comprises tert-butyl 4-((2-oxopropanoyl)oxy)penta-2-inoate. In some embodiments, the compound of formula I comprises tert-butyl 4-((2-hydroxypropanoyl)oxy)penta-2-inoate. In some embodiments, the compound of formula I comprises 5-((5-(tert-butoxy)-5-oxopenta-3-in-2-yl)oxy)-4,5-dioxopentanoic acid.

[0052] In some embodiments, the compound of formula I comprises tert-butyl 4-((2-oxopropanoyl)oxy)-4-phenylbuta-2-inoate. In some embodiments, the compound of formula I comprises tert-butyl 4-((2-hydroxypropanoyl)oxy)-4-phenylbuta-2-inoate. In some embodiments, the compound of formula I comprises 5-((4-(tert-butoxy)-4-oxo-1-phenylbuta-2-in-1-yl)oxy)-4,5-dioxopentanoic acid.

[0053] In some embodiments, the compound of formula I comprises benzhydryl 4-((2-oxopropanoyl)oxy)buta-2-inoate. In some embodiments, the compound of formula I comprises benzhydryl 4-((2-hydroxypropanoyl)oxy)buta-2-inoate. In some embodiments, the compound of formula I comprises 5-((4-(benzhydryloxy)-4-oxobuta-2-in-1-yl)oxy)-4,5-dioxopentanoic acid.

[0054] In some embodiments, the compound of formula I comprises 4-oxo-4-phenylbuta-2-in-1-yl 2-oxopropanoate. In some embodiments, the compound of formula I comprises 4-oxo-4-phenylbuta-2-in-1-yl 2-hydroxypropanoate. In some embodiments, the compound of formula I comprises 4,5-dioxo-5-((4-oxo-4-phenylbuta-2-in-1-yl)oxy)pentanoic acid.

[0055] In some embodiments, the compound of formula I contains 4-oxo-4-(phenyl-d5)buta-2-in-1-yl2-oxopropanoate. In some embodiments, the compound of formula I contains 4-oxo-4-(phenyl-d5)buta-2-in-1-yl2-hydroxypropanoate. In some embodiments, the compound of formula I contains 4,5-dioxo-5-((4-oxo-4-(phenyl-d5)buta-2-in-1-yl)oxy)pentanoic acid.

[0056] In some embodiments, the compound of formula I comprises tert-butyl 4-acetoxybuta-2-inoate. In some embodiments, the compound of formula I comprises 1-(4-(Tert-butoxy)-4-oxobuta-2-in-1-yl)5-ethyl 2-oxopentanedioate.

[0057] In some embodiments, the compound of formula I includes 4-(2,2-dichloroacetoxy)buta-2-inoatemethyl.

[0058] In some embodiments, the compound of formula I contains trityl 4-((2-oxopropanoyl)oxy)buta-2-inoate. In some embodiments, the compound of formula I contains trityl 4-((2-hydroxypropanoyl)oxy)buta-2-inoate. In some embodiments, the compound of formula I contains 4,5-dioxo-5-((4-oxo-4-(trityloxy)buta-2-in-1-yl)oxy)pentanoic acid.

[0059] In some embodiments, the compound of formula I comprises 4-(diphenylamino)-4-oxobuta-2-in-1-yl2-oxopropanoate. In some embodiments, the compound of formula I comprises 4-(diphenylamino)-4-oxobuta-2-in-1-yl2-hydroxypropanoate. In some embodiments, the compound of formula I comprises 5-((4-(diphenylamino)-4-oxobuta-2-in-1-yl)oxy)-4,5-dioxopentanoic acid.

[0060] In some embodiments, the compound of formula I comprises 4-(diisopropylamino)-4-oxobuta-2-in-1-yl 2-oxopropanoate. In some embodiments, the compound of formula I comprises 4-(diisopropylamino)-4-oxobuta-2-in-1-yl 2-hydroxypropanoate. In some embodiments, the compound of formula I comprises 5-((4-(diisopropylamino)-4-oxobuta-2-in-1-yl)oxy)-4,5-dioxopentanoic acid.

[0061] In some embodiments, the compound of formula I comprises 4-oxopenta-2-in-1-yl 2-oxopropanoate. In some embodiments, the compound of formula I comprises 4-oxopenta-2-in-1-yl 2-hydroxypropanoate. In some embodiments, the compound of formula I comprises 4,5-dioxo-5-((4-oxopenta-2-in-1-yl)oxy)pentanoic acid.

[0062] In some embodiments, the compound of formula I comprises 4-oxo-4-(pyridine-2-yl)buta-2-in-1-yl 2-oxopropanoate. In some embodiments, the compound of formula I comprises 4-oxo-4-(pyridine-2-yl)buta-2-in-1-yl 2-hydroxypropanoate. In some embodiments, the compound of formula I comprises 4,5-dioxo-5-((4-oxo-4-(pyridine(27yridine)-2-yl)buta-2-in-1-yl)oxy)pentanoic acid.

[0063] In some embodiments, the compound of formula I includes 4-(1-methyl-1H-imidazole-2-yl)-4-oxobuta-2-in-1-yl2-oxopropanoate. In some embodiments, the compound of formula I includes 4-(1-methyl-1H-imidazole-2-yl)-4-oxobuta-2-in-1-yl2-hydroxypropanoate. In some embodiments, the compound of formula I includes 5-((4-(1-methyl-1H-imidazole-2-yl)-4-oxobuta-2-in-1-yl)oxy)-4,5-dioxopentanoic acid.

[0064] Hydrogenation precursor of formula II In some embodiments, the compound of formula I is parahydrogenated (i.e., modified via the addition of a parahydrogenated proton across Z via a hydrogenation reaction between formula I and parahydrogen) as described herein. In some embodiments, the parahydrogenation of the compound of formula I yields the compound of formula II. Formula II encompasses the following structure: [ka] The tautomers thereof, deuterated derivatives of those compounds and their tautomers, pharmaceutically acceptable salts thereof, and at one or more sites 13 C or 15 Contains N-enriched derivatives. In some embodiments, Z' is (i) 1 H (proton), 2 (ii) Para-hydrogenated carbon-carbon single bonds (-CH*-CH*-) substituted to include H (deuterium) or a combination thereof (e.g., -CH2H*-CH2H*-, -CHDH*-CH2H*-, -CD2H*-CH2H*-), or 1 H (proton), 2The parahydrogenated carbon-carbon double bond (-CH*=CH*-) is substituted to include H (deuterium) or a combination thereof. In some embodiments, H* represents a spin-ordered hydrogen derived from parahydrogen (i.e., a hydrogen atom or proton added across the carbon-carbon double or carbon-carbon triple bond Z via a hydrogenation reaction between the compound of formula I and parahydrogen as described herein). In some embodiments, H* represents a spin-ordered hydrogen derived from parahydrogen (e.g., before polarization transfer). In some embodiments, R1 includes a PHIP transfer portion as described herein. In some embodiments, R2 includes an optionally substituted hydrocarbon, alkoxy group, primary amine, secondary amine, or tertiary amine as described herein. In some embodiments, R3 includes a bio-related contrast agent as described herein. In formula II, all parts to the right of the R3-R1 bond (i.e., -R1-Z'-(C=O)-R2) can be collectively referred to as a parahydrogenated sidearm.

[0065] In some embodiments, the compound of formula II has a solubility in water of at least about 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 6 mM, 7 mM, 8 mM, 9 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, 100 mM, 150 mM, 200 mM, 250 mM, 300 mM, 350 mM, 400 mM, 450 mM, 500 mM, 550 mM, 600 mM, 650 mM, 700 mM, 750 mM, 800 mM, 850 mM, 900 mM, 950 mM, 1,000 mM, or more, up to about 1.0 It has a solubility in water of 00mM, 950mM, 900mM, 850mM, 800mM, 750mM, 700mM, 650mM, 600mM, 550mM, 500mM, 450mM, 400mM, 350mM, 300mM, 250mM, 200mM, 150mM, 100mM, 90mM, 80mM, 70mM, 60mM, 50mM, 40mM, 30mM, 20mM, 10mM, 9mM, 8mM, 7mM, 6mM, 5mM, 4mM, 3mM, 2mM, 1mM or less, or a solubility in water that is within the range defined by any two of the aforementioned values.

[0066] In some embodiments, the compound of formula II is used in an organic solvent (e.g., acetone, ethanol, chloroform, toluene) at a concentration of at least about 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 6 mM, 7 mM, 8 mM, 9 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, 100 mM, 150 mM, 200 mM, 250 mM, 300 mM, 350 mM, 400 mM, 450 mM, 500 mM, 550 mM, 600 mM, 650 mM, 700 mM, 750 mM, 800 mM, 850 mM, 900 mM, 950 mM, 1,000 mM, or higher in an organic solvent (e.g., acetone, ethanol, chloroform, toluene). It has a solubility in organic solvents of up to approximately 1,000 mM, 950 mM, 900 mM, 850 mM, 800 mM, 750 mM, 700 mM, 650 mM, 600 mM, 550 mM, 500 mM, 450 mM, 400 mM, 350 mM, 300 mM, 250 mM, 200 mM, 150 mM, 100 mM, 90 mM, 80 mM, 70 mM, 60 mM, 50 mM, 40 mM, 30 mM, 20 mM, 10 mM, 9 mM, 8 mM, 7 mM, 6 mM, 5 mM, 4 mM, 3 mM, 2 mM, 1 mM, or less, or a solubility in organic solvents defined by any two of the aforementioned values.

[0067] In some embodiments, when the composition of formula I reacts with parahydrogen, the chemical yield (e.g., the chemical yield of the compound of formula II) is at least about 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, or more, and at most about 95%, 90%, 85%, 80%, 75%, 70%, 65%, 60%, 55%, 50%, 45%, 40%, 35%, 30%, or less, or within the range defined by any two of the aforementioned values. For example, in some embodiments, when the composition of formula I reacts with parahydrogen, the chemical yield is 30%~35%, 30%~40%, 30%~45%, 30%~50%, 30%~55%, 30%~60%, 30%~65%, 30%~70%, 30%~75%, 30%~80%, 30%~85%, 30%~90%, 30%~95%, 35%~40%, 35%~45%, 35%~50%, 35%~55%, 35%~60%, 35%~65%, 35%~70%, 35%~75%, 35%~80%, 35%~85%, 35%~90%, 35%~95%, 40%~45%, 40%~50%, 40%~55%, 40%~60%, 40%~65%, 40%~70%, 40%~75%, 40%~80%, 40%~85%, 40%~90%, 40%~95%, 45%~50%, 45%~55%, 45%~60%, 45%~65%, 45%~70%, 45%~75%, 45%~80%, 45 %~85%, 45%~90%, 45%~95%, 50%~55%, 50%~60%, 50%~65%, 50%~70%, 50%~75%, 50%~80%, 50%~85%, 50%~90%, 50%~95%, 55%~60%, 55%~65%, 55%~70%, 55%~75%, 55%~80%, 55%~85%, 55%~90%, 55%~95%, 60%~65%, 60%~70%, 60%~75%, 60%~80%, 60% The percentages are ~85%, 60%~90%, 60%~95%, 65%~70%, 65%~75%, 65%~80%, 65%~85%, 65%~90%, 65%~95%, 70%~75%, 70%~80%, 70%~85%, 70%~90%, 70%~95%, 75%~80%, 75%~85%, 75%~90%, 75%~95%, 80%~85%, 80%~90%, 80%~95%, 85%~90%, 85%~95%, or 90%~95%.

[0068] The cleaved precursor of Equation III In some embodiments, the compound of formula II is cleaved (e.g., hydrolyzed) as described herein. In some embodiments, the compound of formula II is cleaved (e.g., hydrolyzed) as described herein to provide a sidearm compound and a corresponding bio-relevant contrast agent. In some embodiments, cleavage of the compound of formula II results in a compound of formula III and a corresponding bio-relevant contrast agent as described herein. Formula III encompasses the following structure: [ka] The tautomers thereof, deuterated derivatives of those compounds and their tautomers, pharmaceutically acceptable salts thereof, and at one or more sites 13 C or 15 Contains N-enriched derivatives. In some embodiments, Z'' is (i) 1 H (proton), 2 (ii) Parahydrogenated carbon-carbon single bonds (-CH*-CH*-) substituted to include H (deuterium) or a combination thereof, or 1 H (proton), 2 The parahydrogenated carbon-carbon double bond (-CH*=CH*-) is substituted to include H (deuterium) or a combination thereof. In some embodiments, R1' includes a PHIP transfer moiety as described herein. In some embodiments, R2 includes optionally substituted hydrocarbons, alkoxy groups, primary amines, secondary amines, or tertiary amines as described herein. In Formula III, all of the R1-Z''-(C=O)-R2 portion may be referred to as collectively cleaved sidearms or hydrolyzed sidearms.

[0069] In some embodiments, the compound of formula III has a solubility in water of at least about 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 6 mM, 7 mM, 8 mM, 9 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, 100 mM, 150 mM, 200 mM, 250 mM, 300 mM, 350 mM, 400 mM, 450 mM, 500 mM, 550 mM, 600 mM, 650 mM, 700 mM, 750 mM, 800 mM, 850 mM, 900 mM, 950 mM, 1,000 mM, or more, up to about 1.0 It has a solubility in water of 00mM, 950mM, 900mM, 850mM, 800mM, 750mM, 700mM, 650mM, 600mM, 550mM, 500mM, 450mM, 400mM, 350mM, 300mM, 250mM, 200mM, 150mM, 100mM, 90mM, 80mM, 70mM, 60mM, 50mM, 40mM, 30mM, 20mM, 10mM, 9mM, 8mM, 7mM, 6mM, 5mM, 4mM, 3mM, 2mM, 1mM or less, or a solubility in water that is within the range defined by any two of the aforementioned values.

[0070] In some embodiments, the compound of formula III is dissolved in an organic solvent (e.g., acetone, ethanol, chloroform, toluene) in concentrations of at least about 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 6 mM, 7 mM, 8 mM, 9 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, 100 mM, 150 mM, 200 mM, 250 mM, 300 mM, 350 mM, 400 mM, 450 mM, 500 mM, 550 mM, 600 mM, 650 mM, 700 mM, 750 mM, 800 mM, 850 mM, 900 mM, 950 mM, 1,000 mM, or more. The solubility in organic solvents is approximately 1,000 mM, 950 mM, 900 mM, 850 mM, 800 mM, 750 mM, 700 mM, 650 mM, 600 mM, 550 mM, 500 mM, 450 mM, 400 mM, 350 mM, 300 mM, 250 mM, 200 mM, 150 mM, 100 mM, 90 mM, 80 mM, 70 mM, 60 mM, 50 mM, 40 mM, 30 mM, 20 mM, 10 mM, 9 mM, 8 mM, 7 mM, 6 mM, 5 mM, 4 mM, 3 mM, 2 mM, 1 mM, or less, or within the range defined by any two of the aforementioned values.

[0071] Sidearm of Formula IV In some embodiments, a bio-related contrast agent and side arm, such as a side arm compound of formula IV, are conjugated as described herein to form a precursor compound, such as a compound of formula I. Formula IV encompasses the following structure: [ka] The tautomers thereof, deuterated derivatives of those compounds and their tautomers, pharmaceutically acceptable salts thereof, and at one or more sites 13 C or 15 Contains N-enriched derivatives. In some embodiments, Z is (i) 1 H (proton), 2 H (deuterium), or combinations thereof (for example, -C) 1 H=C 1 H-, -C1 H=C 2 H-, -C 2 H=C 2 (ii) carbon-carbon double bonds (-C=C-) or carbon-carbon triple bonds (-C≡C-) are described below. In some embodiments, R1 comprises a para-hydrogen-induced polarization (PHIP) transfer moiety as described herein. In some embodiments, R2 comprises a solubilizing moiety as described herein. In some embodiments, R2 comprises optionally substituted hydrocarbons, alkoxy groups, primary amines, secondary amines, or tertiary amines. In some embodiments, complexation of a compound of formula IV with a bio-related contrast agent results in a compound of formula I as described herein.

[0072] PHIP moving part In some embodiments, the PHIP moving portion described herein moves from one or more parahydrogenated protons H* (e.g., H* in the side arms) to one or more nonhydrogen nuclear spins of a bio-relevant contrast agent (one or more bio-relevant contrast agents as described herein). 13 C or 15 The compound includes a chemical portion configured to allow or enhance polarization transfer to an atom (such as a N atom). In some embodiments, the PHIP transfer portion enables or enhances polarization transfer from a parahydrogenated proton H* in the sidearm of the compound of formula II to the non-hydrogen nuclear spin of the corresponding bio-relevant contrast agent of the compound of formula II. In some embodiments, the PHIP transfer portion enables or enhances polarization transfer from a parahydrogenated proton H* in the sidearm of the compound of formula II to the non-hydrogen nuclear spin of the corresponding bio-relevant contrast agent of the compound of formula II after a parahydrogenation reaction between formula I and parahydrogen.

[0073] In some embodiments, the PHIP transfer portion includes optionally substituted C1 hydrocarbons or optionally substituted C2 hydrocarbons.

[0074] In some embodiments, the PHIP transfer portion includes a chemical portion in the form of *CR4R5, *CR4Y, *C=Y, or any deuterated version thereof. In some embodiments, *C is 12 C or 13 It is a carbon isotope of C. In some embodiments, R4 and R5 are each independently selected from the following: 1 H, 2 H, 3 H, linear, branched, or cyclic C1-C10 alkyl hydrocarbons, C6 aryl, benzyl, phenyl, heteroaryl, and haloalkyl groups. In some embodiments, Y is selected from a spin 1 / 2 atom and a spin 1 / 2 atom covalently bonded to one or more chemical parts selected from a linear, branched, or cyclic C1-C10 alkyl hydrocarbon, C6 aryl, benzyl, phenyl, heteroaryl, halogen, or haloalkyl group, or from heteroatoms such as N, O, S, which are optionally substituted with linear, branched, or cyclic C1-C10 alkyl hydrocarbon, C6 aryl, benzyl, phenyl, heteroaryl, halogen, or haloalkyl group. In some embodiments, the spin 1 / 2 atom is selected from: 1 H, 13 C, 15 N, 19 F, and 31 P. In some embodiments, 15 N may be substituted with a nitro group, an amine group, an amide group, or an imine group. In some embodiments, 31 P may be substituted with one or more keto groups, one or more nitro groups, one or more amine groups, one or more amide groups, or one or more imine groups.

[0075] In some embodiments, the PHIP transfer portion includes a chemical portion in the form of *CR6R7-*CR8R9, or any deuterated version thereof. In some embodiments, *C is 12 C or 13 These are carbon isotopes of C. In some embodiments, R6, R7, R8, and R9 are each independently selected from the following: 1 H, 2 H, 3H, a straight-chain, branched-chain, or cyclic C1-C10 alkyl hydrocarbon, C6 aryl, benzyl, phenyl, heteroaryl, and haloalkyl group.

[0076] In some embodiments, the PHIP transfer moiety comprises a chemical moiety in the form of *CH2, *CH2-*CH2, *CHY, *C=Y, or any deuterated version thereof. In some embodiments, *C is 12 C or 13 a carbon isotope of C. In some embodiments, Y is a spin 1 / 2 atom and a spin 1 / 2 atom covalently bonded to one or more chemical moieties selected from heteroatoms such as N, O, S, or a straight-chain, branched-chain, or cyclic C1-C10 alkyl hydrocarbon, C6 aryl, benzyl, phenyl, heteroaryl, halogen or haloalkyl group, or a straight-chain, branched-chain, or cyclic C1-C10 alkyl hydrocarbon, C6 aryl, benzyl, phenyl, heteroaryl, halogen or haloalkyl group optionally substituted with. In some embodiments, the spin 1 / 2 atom is selected from: 1 H, 13 C, 15 N, 19 F, and 31 P.

[0077] In some embodiments, the compositions described herein have a first J-coupling J 12 between the spin 1 / 2 atom described herein and the non-hydrogen nuclear spin described herein. In some embodiments, the compositions described herein have a second J-coupling J 13 between the spin 1 / 2 atom described herein and the parahydrogenated proton H* described herein. In some embodiments, the compositions described herein have a third J-coupling J 23 between the non-hydrogen nuclear spin described herein and the parahydrogenated proton H* described herein. In some embodiments, J 12 and / or J 13 is greater than J 23 . In such cases, the PHIP transfer moiety may permit or enhance polarization transfer.

[0078] In some embodiments, the PHIP moving portion induces J-coupling between one or both of *H nuclear spins having non-hydrogen nuclear spins of at least about 0.1Hz, 0.2Hz, 0.3Hz, 0.4Hz, 0.5Hz, 0.6Hz, 0.7Hz, 0.8Hz, 0.9Hz, 1Hz, 2Hz, 3Hz, 4Hz, 5Hz, 6Hz, 7Hz, 8Hz, 9Hz, 10Hz or higher, and at most about 10Hz, 9Hz, 8Hz, 7Hz, 6Hz, 5Hz, 4Hz, 3Hz, 2Hz, 1Hz, 0.9Hz, 0.8Hz, 0.7Hz, 0.6Hz, 0.5Hz, 0.4Hz, 0.3Hz, 0.2Hz, 0.1Hz or lower, or induces J-coupling with non-hydrogen nuclear spins within the range defined by any two of the aforementioned values. For example, in some embodiments, J-coupling is used for frequencies of 1Hz~2Hz, 1Hz~3Hz, 1Hz~4Hz, 1Hz~5Hz, 1Hz~6Hz, 1Hz~7Hz, 1Hz~8Hz, 1Hz~9Hz, 1Hz~10Hz, 2Hz~3Hz, 2Hz~4Hz, 2Hz~5Hz, 2Hz~6Hz, 2Hz~7Hz, 2Hz~8Hz, 2Hz~9Hz, 2Hz~10Hz, 3Hz~4Hz, 3Hz~5Hz, 3Hz~6Hz, 3Hz~7Hz, 3Hz~8Hz The frequencies are Hz, 3Hz~9Hz, 3Hz~10Hz, 4Hz~5Hz, 4Hz~6Hz, 4Hz~7Hz, 4Hz~8Hz, 4Hz~9Hz, 4Hz~10Hz, 5Hz~6Hz, 5Hz~7Hz, 5Hz~8Hz, 5Hz~9Hz, 5Hz~10Hz, 6Hz~7Hz, 6Hz~8Hz, 6Hz~9Hz, 6Hz~10Hz, 7Hz~8Hz, 7Hz~9Hz, 7Hz~10Hz, 8Hz~9Hz, 8Hz~10Hz, or 9Hz~10Hz.

[0079] R2 units In some embodiments, the R2 group described herein includes an optionally substituted hydrocarbon, alkoxy group, primary amine, secondary amine, or tertiary amine. In some embodiments, the R2 group described herein includes an optionally substituted hydrocarbon, alkoxy group, primary amine, secondary amine, or tertiary amine that functions as a solubilizing moiety. In some embodiments, the R2 group described herein includes a solubilizing moiety. In some embodiments, the solubilizing moiety includes any chemical moiety configured to permit or enhance the solubility of a compound such as any of the compounds of Formulas I, II, III, and / or IV in a solution in which a para-hydrogenation reaction or a cleavage (e.g., hydrolysis) reaction occurs. In some embodiments, the enhancement of solubility is measured with respect to a variant of a compound of Formula I, II, III, or IV that utilizes one or more protons in place of the R2 group. In some embodiments, the enhancement of solubility is measured with respect to a variant of a compound of Formula I, II, III, or IV that utilizes a methyl group as the R2 group.

[0080] In some embodiments, the solubilizing moiety includes a hydrophobic moiety or an organic affinity moiety. In some embodiments, the solubilizing moiety includes an organic solubilizing moiety. For example, in some embodiments, the solubilizing moiety includes a hydrophobic moiety, an organic affinity moiety, or an organic solubilizing moiety. In some embodiments, the solubilizing moiety includes a hydrophilic moiety or an organic affinity moiety.

[0081] In some embodiments, the R2 group includes, or is selected from, a methyl group, an ethyl group, a propyl group, an isopropyl group, an n-butyl group, an s-butyl group, a t-butyl group, an isobutyl group, a hydroxyl group, a methyl alcohol group, an ethyl alcohol group, an n-propanol group, an isopropyl alcohol group, an alcohol propionic acid group, an n-butyl alcohol group, an s-butyl alcohol group, a t-butyl alcohol group, an isobutyl alcohol group, a methoxy group, an ethoxy group, an ethoxy group, an isopropoxy group, an alcohol propionic acid group, an ethoxy group, a t-butoxy group, an s-butoxy group, an ester group, a phenyl group, a substituted phenyl group, a primary amine group, a secondary amine group, a tertiary amine group, a primary amide group, a secondary amide group, and a tertiary amide group. In some embodiments, the substituted phenyl group is selected from fluorobenzene, chlorobenzene, bromobenzene, iodobenzene, toluene, cumene, ethylbenzene, styrene, orthoxylene, metaxylene, paraxylene, phenol, benzoic acid, benzaldehyde, acetophenone, methyl benzoate, anisole, aniline, nitrobenzene, benzonitrile, benzamide, benzenesulfonic acid, naphthalene, and anthracene.

[0082] R3 groups In some embodiments, the R3 group described herein includes a bio-relevant contrast agent. In some embodiments, the bio-relevant contrast agent has the formula R4C(=O)X-. In some embodiments, R4 is selected from a linear, branched, or cyclic C1-C10 alkyl group, where one or more C atoms are optionally substituted with CO, COOH, CH2COOH, CONH2, OH, amino(NR'R''), one or more halogen atoms, one or more haloalkyl groups, or one or more carbocyclic rings, where the carbocyclic rings are optionally substituted with one or more aliphatic or aromatic rings optionally substituted with one or more functional groups. In some embodiments, X is selected from NR''' and O. In some embodiments, R', R'', and R''' are each independently 1 H, 2 H, 3H is selected from an amino protecting group optionally selected from trifluoroacetyl, acetyl, benzoyl, carbobenzoxy, tert-butyl carbonate, and benzyl. In some embodiments, the R3 group comprises any bio-related contrast agent described herein.

[0083] In some embodiments, the R3 group includes at least one non-hydrogen nucleus spin. In some embodiments, the non-hydrogen nucleus includes at least one spin 1 / 2 atom. In some embodiments, the non-hydrogen nucleus spin is 13 C or 15 Contains N. In some embodiments, the R3 group is at least partially isotope-labeled with non-hydrogen nuclear spin. In some embodiments, the R3 group is at least partially enriched with non-hydrogen nuclear spin compared to analogs of the R3 group that characterize non-hydrogen nuclear spin by its natural abundance. In some embodiments, the R3 group 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%, 95% The non-hydrogen nucleus spins are enriched to characterize them at abundances of %, 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 within the range defined by any two of the aforementioned values.

[0084] In some embodiments, the non-hydrogen nuclear spin is an NMR-inactive (i.e., spin 0) nucleus of an analog of the R3 group that characterizes the non-hydrogen nuclear spin by its natural abundance, as described herein (e.g., 12 C or quadropolar (i.e., spin > 1 / 2) nuclei (e.g., 14Substitute N). In some embodiments, the non-hydrogen nucleus spin is located about one or fewer chemical bonds from the carbonyl (C=O) carbon in the R3 group.

[0085] Parahydrogenation In accordance with the disclosed embodiments, precursors of bio-relevant contrast agents (such as compounds of formula I as described herein) can be parahydrogenated by combining the precursor, parahydrogen, and a hydrogenation catalyst. The disclosed embodiments are not limited to a specific method for producing the parahydrogenated precursor. In some embodiments, the precursor is added to a mixture containing parahydrogen. In some embodiments, the parahydrogen gas is added to a solution containing the precursor (for example, the parahydrogen gas may be bubbled in such a solution). In the hydrogenation of the precursor, the parahydrogen can create a singlet spin order on the two hydrogen spins in the precursor, with respect to the two hydrogen spins in the precursor, a preferred group of lower energy states between Iz1Iz2 order, |↑>|↓>, and |↓>|↑>.

[0086] The precursor may have unsaturated bonds (such as unsaturated carbon-carbon double bonds or unsaturated carbon-carbon triple bonds) that can be hydrogenated by parahydrogen gas. After combining the precursor with parahydrogen, a proportion of the precursor can be hydrogenated, which is defined as at least about 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, or more of the precursor, and at most about 90%, 80%, 70%, 60%, 50%, 40%, 30%, 20%, 10%, or less of the precursor, or within the range defined by any two of the aforementioned values.

[0087] In some embodiments, the parahydrogenated precursor has a gregacy difference of at least about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 40%, 45%, 50%, or more in the parahydrogenated proton spin state, and a maximum gregacy difference of about 50%, 45%, 40%, 35%, 30%, 25%, 20%, 15%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less, or within the range defined by any two of the aforementioned values. For example, in some embodiments, the group differences are 10%~15%, 10%~20%, 10%~25%, 10%~30%, 10%~35%, 10%~40%, 10%~45%, 10%~50%, 15%~20%, 15%~25%, 15%~30%, 15%~35%, 15%~40%, 15%~45%, 15%~50%, 20%~25%, 20%~30%, 2 The percentages are 0%~35%, 20%~40%, 20%~45%, 20%~50%, 25%~30%, 25%~35%, 25%~40%, 25%~45%, 25%~50%, 30%~35%, 30%~40%, 30%~45%, 30%~50%, 35%~40%, 35%~45%, 35%~50%, 40%~45%, 40%~50%, or 45%~50%. In some embodiments, the collective difference is between spin states containing parahydrogenated protons and spin states containing other nuclear spins, e.g., additional protons on the compound. In some embodiments, the parahydrogenated precursor contains side chains, and the parahydrogenated spins may be located on the side chains.

[0088] In some embodiments, the concentration of the hydrogenation catalyst during hydrogenation is at least about 0.1 mM, 0.2 mM, 0.3 mM, 0.4 mM, 0.5 mM, 0.6 mM, 0.7 mM, 0.8 mM, 0.9 mM, 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 6 mM, 7 mM, 8 mM, 9 mM, 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, 100 mM, or more, up to The values ​​are approximately 100mM, 90mM, 80mM, 70mM, 60mM, 50mM, 40mM, 30mM, 20mM, 10mM, 9mM, 8mM, 7mM, 6mM, 5mM, 4mM, 3mM, 2mM, 1mM, 0.9mM, 0.8mM, 0.7mM, 0.6mM, 0.5mM, 0.4mM, 0.3mM, 0.2mM, 0.1mM, or less, or within the range defined by any two of the aforementioned values.

[0089] The disclosed embodiments may include methods implemented by the disclosed system for generating hyperpolarized bio-relevant contrast agents. The methods of the disclosed invention may include mixing a solution (e.g., by a mixing mechanism) comprising a precursor and a hydrogenation catalyst to a bio-relevant contrast agent. The mixing mechanism may be a device for introducing, holding, and facilitating a blend, mixture, or solution of two or more materials. In some embodiments, the mixing mechanism is located within a chamber, and the mixing occurs within the chamber. In some embodiments, the solutions are mixed at a location away from the chamber. The solution may be at least about 1 milliliter (ml), 2 ml, 3 ml, 4 ml, 5 ml, 6 ml, 7 ml, 8 ml, 9 ml, 10 ml, 20 ml, 30 ml, 40 ml, 50 ml, 60 ml, 70 ml, 80 ml, 90 ml, 100 ml, or more, or at most 100 ml, 90 ml, 80 ml, 70 ml, 60 ml, 50 ml, 40 ml, 30 ml, 20 ml, 10 ml, 9 ml, 8 ml, 7 ml, 6 ml, 5 ml, 4 ml, 3 ml, 2 ml, 1 ml, or less, or within the volume range defined by any two of the aforementioned values.

[0090] In some embodiments, the mixing mechanism is a gas-liquid exchange mechanism. For example, the gas-liquid exchange mechanism may be a bubbler or a diffusion system. In some embodiments, the mixing mechanism includes a membrane adapted to allow the diffusion of molecular hydrogen. In some embodiments, mixing may be carried out using a spray chamber, in which the solution is sprayed into a chamber filled with pressurized parahydrogen.

[0091] In some embodiments, the catalyst is a molecule, complex, or particle system that catalyzes hydrogenation. In some embodiments, the catalyst includes a homogeneous metal catalyst such as a rhodium complex or a ruthenium complex. Rhodium complexes can be used to prepare and activate precursor molecules and parahydrogens. In some embodiments, heterogeneous metal catalysts are attached to nanoparticles.

[0092] Various embodiments of this disclosure describe introducing a solution comprising a precursor of a bio-related contrast agent and a hydrogenation catalyst into a chamber configured to hold the solution during polarization transfer. In some embodiments, the solution is mixed within the chamber. In some embodiments, the solution is hydrogenated within the chamber. In some embodiments, the chamber is located within a magnetic shield (e.g., a mu-metal shield). The magnetic shield can reduce the effects of the Earth's magnetic field (or other external magnetic fields) and allows modulation of the amplitude of the low-level magnetic field applied to the solution. Therefore, placing the solution within a chamber may include placing the solution within a magnetic shield.

[0093] As described herein, in some embodiments, parahydrogenation occurs before polarization transfer (e.g., before amplitude modulation, such as in a magnetic field applied to a solution). In some embodiments, parahydrogenation occurs during polarization transfer. For example, parahydrogen can be combined with the solution during modulation of the magnetic field amplitude (e.g., flowed through the solution or bubbled).

[0094] In some embodiments, parahydrogen gas is combined with a solution in a hydrogenation chamber under pressure. The pressure may be at least about 10 bar, 15 bar, 20 bar, 30 bar, 50 bar, or higher, and may be at most about 50 bar, 30 bar, 20 bar, 15 bar, 10 bar, or within the range defined by any two of the aforementioned values. In some embodiments, parahydrogen is combined with a solution in a metal chamber capable of withstanding the pressure. Parahydrogen may be combined with the solution over a time interval (or the dissolution of parahydrogen may occur in less than a time interval). The time interval can be up to approximately 90 seconds, 60 seconds, 30 seconds, 20 seconds, 10 seconds, 9 seconds, 8 seconds, 7 seconds, 6 seconds, 5 seconds, 4 seconds, 3 seconds, 2 seconds, 1 second, or less, at least approximately 1 second, 2 seconds, 3 seconds, 4 seconds, 5 seconds, 6 seconds, 7 seconds, 8 seconds, 9 seconds, 10 seconds, 20 seconds, 30 seconds, 60 seconds, 90 seconds, or more, or within the range defined by any two of the aforementioned values. In some embodiments, hydrogenation is carried out or occurs within the time interval.

[0095] Polarization shift using high-frequency waveforms In some embodiments, the concentration of the precursor in the solution before polarization transfer is at least about 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, 100 mM, 200 mM, 300 mM, 400 mM, 500 mM, 600 mM, 700 mM, 800 mM, 900 mM, 1,000 mM, or more. The maximum values ​​are approximately 1,000 mM, 900 mM, 800 mM, 700 mM, 600 mM, 500 mM, 400 mM, 300 mM, 200 mM, 100 mM, 90 mM, 80 mM, 70 mM, 60 mM, 50 mM, 40 mM, 30 mM, 20 mM, 10 mM, or less, or within the range defined by any two of the aforementioned values. The volume of the solution is at least approximately 1 ml, 2 ml, 3 ml, 4 ml, 5 ml, 6 ml, 7 ml, 8 ml, 9 ml, 10 ml, 20 ml, 30 ml, 40 ml, 50 ml, 60 ml, 70 ml, 80 ml, 90 ml, 100 ml, 200 ml, 300 ml, 400 ml, 500 ml, 600 ml, 700 ml, 800 ml, 900 ml, 1000 ml, 2000 ml, or more, with a maximum of approximately 2000 ml. It may be 000ml, 900ml, 800ml, 700ml, 600ml, 500ml, 400ml, 300ml, 200ml, 100ml, 90ml, 80ml, 70ml, 60ml, 50ml, 40ml, 30ml, 20ml, 10ml, 9ml, 8ml, 7ml, 6ml, 5ml, 4ml, 3ml, 2ml, 1ml, or less, or within the range defined by any two of the aforementioned values.

[0096] Various embodiments of this disclosure describe the application of polarization-transferring magnetic perturbations aimed at generating a magnetic field around a solution (for example, around a solution containing Formula II as described herein). In some embodiments, the magnetic field is at least about 0.1 Gauss (G), 0.2G, 0.3G, 0.4G, 0.5G, 0.6G, 0.7G, 0.8G, 0.9G, 1G, 2G, 3G, 4G, 5G, 6G, 7G, 8G, 9G, 10G, 20G, 30G, 40G, 50G, 60G, 70G, 80G, 90G, 100G, 200G, 300G, 400G, 500G, 600G, 700G, 800G, 900G, 1,000G, 2,000G, 3,000G, 4,000G, 5,000G, 6,000G, 7,000G, 8,000G, 9,000G, 10,000G, 20,000G, 30,000G, 40,000G, 50,000G, 60,000G, 70,000G, 80,000G, 90,000G, 100,000G, 200,000G, or more, up to approximately 200 ,000G, 100,000G, 90,000G, 80,000G, 70,000G, 60,000G, 50,000G, 40,000G, 30,000G, 20,000G, 10,000G, 9,000G, 8,000G, 7,000G, 6,000G, 5,000G, 4,000G, 3,000G, 2,000G, 1,000G, 900G, 800G, 700G, 600G, 500G The magnetic field has an intensity of 400G, 300G, 200G, 100G, 90G, 80G, 70G, 60G, 50G, 40G, 30G, 20G, 10G, 9G, 8G, 7G, 6G, 5G, 4G, 3G, 2G, 1G, 0.9G, 0.8G, 0.7G, 0.6G, 0.5G, 0.4G, 0.3G, 0.2G, 0.1G, or less, or within the range defined by any two of the aforementioned values. In some embodiments, the magnetic field has an intensity of 0.1G to 200,000G around the solution. Magnetic perturbations can be generated by electromagnets or permanent magnets. The magnetic field can be applied to the sample in pulse or continuous wave (CW) form. Magnetic perturbations may be static or vary over time.

[0097] The signal generator may be configured to generate one or more radio frequency (RF) waveforms that can be applied to a sample to shift its polarization. The signal generator may include another computing unit, processor, controller, associated memory, PC, computer service, or any device that can perform computing operations using inputs and generate outputs. In some embodiments, the RF coil may emit or "apply" a pulse sequence containing a first RF waveform. In some embodiments, the RF coil may have one or more channels. The channels may be paths for the RF signal. At least one channel may be provided for each different type of NMR spectroscopy. In some embodiments, 1 H has at least one channel, 2 H, 13 C, 15 N, 19 F, and 31 P has at least one channel. For example, the first RF waveform is from one or more high-frequency coils (RF coils) arranged around the sample. 1 This can be applied to H channels. In some embodiments, the second RF waveform is the RF coil 13 Applies to C channels. In some embodiments, 1 H channel and 13 The RF waveform on the C channel is configured to apply a polarization shift sequence such as PH-INEPT, Goldman's sequence, S2M, S2hM, SLIC, ADAPT, or ESOTERIC.

[0098] In some embodiments, the RF waveform is configured to support polarization movement even in the presence of a large proton full width half-maximum (FWHM). Such RF waveforms may include a pulse sequence that may contain tens to hundreds of RF pulses. The sequence may be configured so that the pulses protect against the detrimental effects of magnetic field inhomogeneity on polarization movement.

[0099] In some embodiments, the polarization pulse sequence is, for example, when the chemical shift difference is greater than the J-coupling between them, two non-equivalents 1It is configured to transfer spin order from hydrogenated spins. ESOTHERIC may be, for example, a pulse sequence suitable for polarization transfer in this regime.

[0100] In some embodiments, the pulse sequence is, for example, equivalent when the chemical shift differences are smaller than the J-coupling between them. 1These pulse sequences are configured to transfer spin order from the H hydrogen spin. T, 9mT, 10mT, 20mT, 30mT, 40mT, 50mT, 60mT, 70mT, 80mT, 90mT, 100mT, 200mT, 300mT, 400mT, 500mT, 600mT, 700mT, 800mT, 900mT, 1,000mT, 2,000mT, 3,000mT, 4,000mT, 5,000mT, 6,000mT, or more, up to approximately 6,000mT. 5,000mT, 4,000mT, 3,000mT, 2,000mT, 1,000mT, 900mT, 800mT, 700mT, 600mT, 500mT, 400mT, 300mT, 200mT , 100mT, 90mT, 80mT, 70mT, 60mT, 50mT, 40mT, 30mT, 20mT, 10mT, 9mT, 8mT, 7mT, 6mT, 5mT, 4mT, 3mT, 2mT, 1mT These can be used in magnetic fields having strengths of 0.9mT, 0.8mT, 0.7mT, 0.6mT, 0.5mT, 0.4mT, 0.3mT, 0.2mT, 0.1mT, 0.09mT, 0.08mT, 0.07mT, 0.06mT, 0.05mT, 0.04mT, 0.03mT, 0.02mT, 0.01mT, or less, or within the range defined by any two of the aforementioned values. Examples of such sequences may be Goldman sequences (M. Goldman, H. Johannesson, CRPhys. 2005, 6, 575-581, this reference is incorporated herein by reference in relation to pulse sequence configurations for transferring spin order), singlet-to-heteronuclear magnetization (S2hM) sequences, or other sequences used in singlet NMR (e.g., ADAPT, SLIC, etc.).

[0101] In some embodiments, the magnetic shield is applied to solutions with magnetic fields of at least about 0mG, 0.1mG, 0.2mG, 0.3mG, 0.4mG, 0.5mG, 0.6mG, 0.7mG, 0.8mG, 0.9mG, 1mG, 2mG, 3mG, 4mG, 5mG, 6mG, 7mG, 8mG, 9mG, 10mG, 20mG, 30mG, 40mG, 50mG, 60mG, 70mG, 80mG, 90mG, 100mG or higher, up to about 100m The magnetic shield is configured to maintain a magnetic field that is within the range defined by any two of the aforementioned values ​​when applied to a solution with a magnetic field of G, 90mG, 80mG, 70mG, 60mG, 50mG, 40mG, 30mG, 20mG, 10mG, 9mG, 8mG, 7mG, 6mG, 5mG, 4mG, 3mG, 2mG, 1mG, 0.9mG, 0.8mG, 0.7mG, 0.6mG, 0.5mG, 0.4mG, 0.3mG, 0.2mG, or 0.1mG or less, or when applied to such a solution. The magnetic shield can maintain the magnetic field strength within the polarization chamber even at such amplitudes when a polarization waveform is applied to one or more high-frequency coils.

[0102] In accordance with the disclosed embodiments, the RF waveform may be applied to a solution containing a parahydrogenated precursor.

[0103] Polarization shift using magnetic field modulation In some embodiments, polarization-transferring magnetic perturbations are carried out within a magnetic shield (e.g., a mu-shield) to achieve a homogeneous low magnetic field. The magnetic shield operates in a microtesla (μT) magnetic field, which is below the Earth's magnetic field. 13This enables polarization transfer to the C nuclear spin. Low magnetic fields range from approximately 0mG, 0.1mG, 0.2mG, 0.3mG, 0.4mG, 0.5mG, 0.6mG, 0.7mG, 0.8mG, 0.9mG, 1mG, 2mG, 3mG, 4mG, 5mG, 6mG, 7mG, 8mG, 9mG, 10mG, 20mG, 30mG, 40mG, 50mG, 60mG, 70mG, 80mG, 90mG, 100mG, or higher, with a maximum of approximately 100mG and 90mG. , 80mG, 70mG, 60mG, 50mG, 40mG, 30mG, 20mG, 10mG, 9mG, 8mG, 7mG, 6mG, 5mG, 4mG, 3mG, 2mG, 1mG, 0.9mG, 0.8mG, 0.7mG, 0.6mG, 0.5mG, 0.4mG, 0.3mG, 0.2mG, 0.1mG, or less, or within the range defined by any two of the aforementioned values.

[0104] In such a magnetic field, polarization is related to proton spin, 2 H, 13 C, 15 N, 19 F, and 31 The polarization is moved by utilizing level pseudocrossing (LAC) between P and other spin species of interest. In some embodiments, the magnetic field can be adjusted to a specific magnetic field strength relative to the LAC. In various embodiments, the magnetic field strength can be adjusted over time to enable robust polarization movement in larger volume samples. For example, the magnetic field strength can be swept through the LAC conditions. Alternatively or additionally, the sample may be physically moved within the magnetic field. Such modulation can relax constraints on magnetic field homogeneity and magnetic field offset. Thus, robust polarization movement can be achieved in larger volumes with greater efficiency. Furthermore, relaxing constraints on magnetic field homogeneity and magnetic field offset can enable the use of less complex, more precise, or more expensive polarization systems.

[0105] The lower limit of magnetic field modulation is at least approximately -10μT, -9μT, -8μT, -7μT, -6μT, -5μT, -4μT, -3μT, -2μT, -1μT, -0.9μT, -0.8μT, -0.7μT, -0.6μT, -0.5μT, -0.4μT, -0.3μT, -0.2μT, -0.1μT, or higher, with a maximum of approximately -0.1μT. The values ​​may be -0.2μT, -0.3μT, -0.4μT, -0.5μT, -0.6μT, -0.7μT, -0.8μT, -0.9μT, -1μT, -2μT, -3μT, -4μT, -5μT, -6μT, -7μT, -8μT, -9μT, -10μT, or less, or within the range defined by any two of the aforementioned values. The upper limit of the modulation may be at least approximately 0.1μT, 0.2μT, 0.3μT, 0.4μT, 0.5μT, 0.6μT, 0.7μT, 0.8μT, 0.9μT, 1μT, 2μT, 3μT, 4μT, 5μT, 6μT, 7μT, 8μT, 9μT, 10μT, or more, and at most approximately 10μT, 9μT, 8μT, 7μT, 6μT, 5μT, 4μT, 3μT, 2μT, 1μT, 0.9μT, 0.8μT, 0.7μT, 0.6μT, 0.5μT, 0.4μT, 0.3μT, 0.2μT, 0.1μT, or less, or within the range defined by any two of the aforementioned values.

[0106] The magnetic field is applied to volumes of at least approximately 1 ml, 2 ml, 3 ml, 4 ml, 5 ml, 6 ml, 7 ml, 8 ml, 9 ml, 10 ml, 20 ml, 30 ml, 40 ml, 50 ml, 60 ml, 70 ml, 80 ml, 90 ml, 100 ml, 200 ml, 300 ml, 400 ml, 500 ml, 600 ml, 700 ml, 800 ml, 900 ml, 1,000 ml, 2,000 ml, or more, with a maximum of approximately 2,000 ml, 1,000 ml, 90 ml. Such amplitudes may be present over volumes of 0 ml, 800 ml, 700 ml, 600 ml, 500 ml, 400 ml, 300 ml, 200 ml, 100 ml, 90 ml, 80 ml, 70 ml, 60 ml, 50 ml, 40 ml, 30 ml, 20 ml, 10 ml, 9 ml, 8 ml, 7 ml, 6 ml, 5 ml, 4 ml, 3 ml, 2 ml, 1 ml, or less, or within the range defined by any two of the aforementioned values. Modulation may be performed over a certain duration. Duration: at least approximately 100 milliseconds (ms), 200 milliseconds, 300 milliseconds, 400 milliseconds, 500 milliseconds, 600 milliseconds, 700 milliseconds, 800 milliseconds, 900 milliseconds, 1,000 milliseconds, 2,000 milliseconds, 3,000 milliseconds, 4,000 milliseconds, 5,000 milliseconds, 6,000 milliseconds, 7,000 milliseconds, 8,000 milliseconds, 9,000 milliseconds, 10,000 milliseconds, 20,000 milliseconds, 30,000 milliseconds, 40,000 milliseconds, 50,000 milliseconds, or more, up to approximately 50,000 milliseconds. This may be milliseconds, 40,000 milliseconds, 30,000 milliseconds, 20,000 milliseconds, 10,000 milliseconds, 9,000 milliseconds, 8,000 milliseconds, 7,000 milliseconds, 6,000 milliseconds, 5,000 milliseconds, 4,000 milliseconds, 3,000 milliseconds, 2,000 milliseconds, 1,000 milliseconds, 900 milliseconds, 800 milliseconds, 700 milliseconds, 600 milliseconds, 500 milliseconds, 400 milliseconds, 300 milliseconds, 200 milliseconds, 100 milliseconds, or less, or within the range defined by any two of the aforementioned values.

[0107] Therefore, the rate of change of the magnetic field amplitude is at least approximately 0.01 μT / sec, 0.02 μT / sec, 0.03 μT / sec, 0.04 μT / sec, 0.05 μT / sec, 0.06 μT / sec, 0.07 μT / sec, 0.08 μT / sec, 0.09 μT / sec, 0.1 μT / sec, 0.2 μT / sec, 0.3 μT / sec, 0.4 μT / sec, 0.5 μT / sec, 0.6 μT / sec, 0.7 μT / sec The rate of change of the magnetic field amplitude may be 0.8 μT / s, 0.9 μT / s, 1 μT / s, or more, with a maximum of approximately 1 μT / s, 0.9 μT / s, 0.8 μT / s, 0.7 μT / s, 0.6 μT / s, 0.5 μT / s, 0.4 μT / s, 0.3 μT / s, 0.2 μT / s, 0.1 μT / s, or less, or within the range defined by any two of the aforementioned values. The upper limit of the rate of change of the magnetic field amplitude may be determined by the capabilities of the equipment used to perform the sweep.

[0108] In some embodiments, when the magnetic field is within the upper and lower limits disclosed above, the spatial deviation of the magnetic field across the modulated volume is less than about half (or a quarter, or an eighth, or a tenth) of the amplitude of the magnetic field. For example, when the magnetic field strength is less than 2 μT (or greater than -2 μT), the spatial deviation of the magnetic field across the modulated volume may be less than 1 μT. As an additional example, when the magnetic field strength is less than 10 μT (or greater than -10 μT), the spatial deviation of the magnetic field across the modulated volume may be less than 5 μT. The spatial deviation can be measured, for example, by taking at least about 10, 20, 30, 40, 50, 60, 70, 80, 90, 100, 200, 300, 400, 500 or more spatially randomly sampled or spatially equally distributed measurements of the magnetic field in the volume and calculating the standard deviation of the sampled magnetic field measurements. Such homogeneity can be achieved within a large homogeneous magnetic shield, for example, by having a large puncture solenoid through the magnetic shield, or by using a large Helmholtz coil with a large homogeneous region for generating magnetic field amplitude modulation. In some embodiments, the modulation is a sweep of the magnetic field. In some embodiments, the magnetic field amplitude modulation includes non-adiabatic jumps, monotonic amplitude changes, or a combination thereof.

[0109] In some embodiments, following the polarization transfer step, a bio-relevant contrast agent (for example, a bio-relevant contrast agent) 13 C or 15 The non-hydrogen nuclear spins of N) have a nuclear spin polarization of at least about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, or more, and a maximum of about 50%, 45%, 40%, 35%, 30%, 25%, 20%, 15%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or within the range defined by any two of the aforementioned values. For example, in some embodiments, following the polarization transfer step, the non-hydrogen nuclear spins of the bio-related contrast agent are 10%~15%, 10%~20%, 10%~25%, 10%~30%, 10%~35%, 10%~40%, 10%~45%, 10%~50%, 15%~20%, 15%~25%, 15%~30%, 15%~35%, 15%~40%, 15%~45%, 15%~50%, 20%~25%, 2 It has nuclear spin polarization of 0%~30%, 20%~35%, 20%~40%, 20%~45%, 20%~50%, 25%~30%, 25%~35%, 25%~40%, 25%~45%, 25%~50%, 30%~35%, 30%~40%, 30%~45%, 30%~50%, 35%~40%, 35%~45%, 35%~50%, 40%~45%, 40%~50%, or 45%~50%.

[0110] In some embodiments, this polarization is achieved for solution volumes of approximately 1 ml, 2 ml, 3 ml, 4 ml, 5 ml, 6 ml, 7 ml, 8 ml, 9 ml, 10 ml, 20 ml, 30 ml, 40 ml, 50 ml, 60 ml, 70 ml, 80 ml, 90 ml, 100 ml, 200 ml, 300 ml, 400 ml, 500 ml, or more, up to approximately 500 ml, 400 ml, 300 ml, 200 ml, 100 ml, 90 ml, 80 ml, 70 ml, 60 ml, 50 ml, 40 ml, 30 ml, 20 ml, 10 ml, 9 ml, 8 ml, 7 ml, 6 ml, 5 ml, 4 ml, 3 ml, 2 ml, 1 ml, or less, or within the range defined by any two of the aforementioned values.

[0111] In some embodiments, after polarization transfer, a portion of the collective difference in the parahydrogenated proton spin state is the target of the bio-related contrast agent (e.g., 13 C or 15 N) Transferred to the polarization of nuclear spin. This portion may be at least about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, or more, and at most about 50%, 45%, 40%, 35%, 30%, 25%, 20%, 15%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or within the range defined by any two of the aforementioned values. For example, in some embodiments, this portion is 10%~15%, 10%~20%, 10%~25%, 10%~30%, 10%~35%, 10%~40%, 10%~45%, 10%~50%, 15%~20%, 15%~25%, 15%~30%, 15%~35%, 15%~40%, 15%~45%, 15%~50%, 20%~25%, 20%~30%, The percentages are 20%~35%, 20%~40%, 20%~45%, 20%~50%, 25%~30%, 25%~35%, 25%~40%, 25%~45%, 25%~50%, 30%~35%, 30%~40%, 30%~45%, 30%~50%, 35%~40%, 35%~45%, 35%~50%, 40%~45%, 40%~50%, or 45%~50%.

[0112] In some embodiments, magnetic field modulation includes a non-adiabatic jump in the magnetic field. The non-adiabatic jump may be performed for magnetic fields where level pseudocrossing occurs, including proton spins and aproton spins. Considering J-coupling between nuclear spins in the system, this value may be analytically calculated or identified by plotting the energy levels of the Hamiltonian for different magnetic fields and identifying the LAC. In some embodiments, the duration for which the magnetic field amplitude is in the LAC condition is up to about 5 seconds, 4 seconds, 3 seconds, 2 seconds, 1 second, 0.9 seconds, 0.8 seconds, 0.7 seconds, 0.6 seconds, 0.5 seconds, 0.4 seconds, 0.3 seconds, 0.2 seconds, 0.1 seconds or less, at least about 0.1 seconds, 0.2 seconds, 0.3 seconds, 0.4 seconds, 0.5 seconds, 0.6 seconds, 0.7 seconds, 0.8 seconds, 0.9 seconds, 1 second, 2 seconds, 3 seconds, 4 seconds, 5 seconds or more, or within the range defined by any two of the aforementioned values.

[0113] In some embodiments, modulating the magnetic field amplitude involves monotonically changing the magnetic field amplitude (or monotonically over each of a limited number of intervals, such as increasing intervals of 1 to 10 and / or decreasing intervals of 1 to 10). In some embodiments, modulating the magnetic field amplitude involves linearly changing the magnetic field amplitude. The initial and final magnetic field amplitudes of the sweep, and the total duration of the sweep, can be optimized for the target molecule. In some embodiments, the magnetic field amplitude during the sweep is within lower and upper limits. The lower limit may be at least approximately -2μT, -1μT, -0.9μT, -0.8μT, -0.7μT, -0.6μT, -0.5μT, -0.4μT, -0.3μT, -0.2μT, -0.1μT or higher, and at most approximately -0.1μT, -0.2μT, -0.3μT, -0.4μT, -0.5μT, -0.6μT, -0.7μT, -0.8μT, -0.9μT, -1μT, -2μT or lower, or within the range defined by any two of the aforementioned values. The upper limit may be at least approximately 0.1 μT, 0.2 μT, 0.3 μT, 0.4 μT, 0.5 μT, 0.6 μT, 0.7 μT, 0.8 μT, 0.9 μT, 1 μT, 2 μT, or more, and at most approximately 2 μT, 1 μT, 0.9 μT, 0.8 μT, 0.7 μT, 0.6 μT, 0.5 μT, 0.4 μT, 0.3 μT, 0.2 μT, 0.1 μT, or less, or within the range defined by any two of the aforementioned values. For some MOSFETs, the modulation duration is at least approximately 100ms, 200ms, 300ms, 400ms, 500ms, 600ms, 700ms, 800ms, 900ms, 1,000ms, 2,000ms, 3,000ms, 4,000ms, 5,000ms, 6,000ms, 7,000ms, 8,000ms, 9,000ms, 10,000ms, or longer, up to approximately 10 The amplitude may be 000ms, 9,000ms, 8,000ms, 7,000ms, 6,000ms, 5,000ms, 4,000ms, 3,000ms, 2,000ms, 1,000ms, 900ms, 800ms, 700ms, 600ms, 500ms, 400ms, 300ms, 200ms, 100ms, or less, or within the range defined by any two of the aforementioned values. In some embodiments, the rate of amplitude change varies along the amplitude profile.In some embodiments, constant adiabatic sweep is calculated by selecting a specific subgroup of level pseudocrossings in the spin system. In some embodiments, magnetic amplitude modulation includes a combination of non-adiabatic jumps, monotonic amplitude modulation, and rate of change sign inversion.

[0114] Purification and separation In some embodiments, the precursor may be selected or designed such that, following hydrogenation and other potential chemical reactions, one of the products is a bio-relevant contrast agent usable in hyperpolarized NMR or MRI applications. In some embodiments, the bio-relevant contrast agent is produced through additional chemical reactions following hydrogenation. Such additional chemical reactions may include, for example, cleaving the side arms of the molecule (e.g., cleaving the compound of formula II as described herein) by hydrolysis to form the bio-relevant contrast agent and the side arms (e.g., the compound of formula III as described herein). After hydrogenation and polarization transfer, the para-hydrogenated precursor (e.g., the compound of formula II as described herein) may be cleaved to produce a hyperpolarized bio-relevant contrast agent.

[0115] The volume of the solution containing the bio-related contrast agent after cutting (and the concentration of the bio-related contrast agent produced) may depend on the volume of the solution used for polarization transfer and the concentration of the precursor in that solution. Exemplary ranges of solution volume and precursor concentration are described herein. As further specific examples, solutions of at least about 1 ml, 2 ml, 3 ml, 4 ml, 5 ml, 6 ml, 7 ml, 8 ml, 90 ml, 100 ml, 200 ml, 300 ml, 400 ml, 500 ml, or more can be produced, containing at least about 10 mM, 20 mM, 30 mM, 400 mM, 500 mM, or more of the bio-related contrast agent.

[0116] In accordance with the disclosed embodiments, after polarization transfer and cleavage of the precursor to generate a hyperpolarized bio-relevant contrast agent, the properties of the solution containing the hyperpolarized bio-relevant contrast agent can be modified to induce precipitation of the hyperpolarized bio-relevant contrast agent.

[0117] Such precipitates can allow for the separation of hyperpolarized bio-related contrast agents from other substances in the solution (e.g., sidearm fragments). The precipitates may form crystals, amorphous solid particles, polycrystalline materials, etc. After precipitation, a fraction of at least solid hyperpolarized bio-related contrast agent (or hyperpolarized precursor) can be separated from the solution and other substances in the solution. For example, a mixture of precipitate and solution may be filtered to remove particles. The filtered precipitate may be washed with a second solvent. The second solvent may be selected to remove the original solvent and other substance residues in the solution without completely dissolving the filtered precipitate. In some embodiments, the cleaning process takes place for a maximum of about 300 seconds, 200 seconds, 100 seconds, 90 seconds, 80 seconds, 70 seconds, 60 seconds, 50 seconds, 30 seconds, 20 seconds, or 10 seconds, and at least about 10 seconds, 20 seconds, 30 seconds, 50 seconds, 60 seconds, 70 seconds, 80 seconds, 90 seconds, 100 seconds, 200 seconds, 300 seconds, or longer, or within the range defined by any two of the aforementioned values.

[0118] The precipitate consists of acetic acid, acetone, acetonitrile, benzene, 1-butanol, 2-butanol, 2-butanone, t-butyl alcohol, carbon tetrachloride, chlorobenzene, chloroform, cyclohexane, 1,2-dichloroethane, diethylene glycol, diethyl ether, digleam (diethylene glycol dimethyl ether), 1,2-dimethoxyethane (gleam, DME), dimethylformamide (DMF), dimethyl sulfoxide (DMSO), 1,4-dioxane, ethanol, ethyl acetate, ethylene glycol, and glycerin. The process may be carried out using aqueous solutions or solutions containing organic solvents such as n, heptane, hexamethylphosphoramide (HMPA), hexamethylphosphoric triamide (HMPT), hexane, methanol, methyl t-butyl ether (MTBE), methylene chloride, N-methyl-2-pyrrolidone (NMP), nitromethane, pentane, petroleum ether (ligroin), 1-propanol, 2-propanol, pyridine, tetrahydrofuran (THF), toluene, triethylamine, water, heavy water, o-xylene, m-xylene, and p-xylene. In some embodiments, the organic solvent is used in the polarization and precipitation steps. For example, parahydrogen may be more soluble in organic solvents than in aqueous solutions. Therefore, hydrogenation may occur more efficiently in such solvents. Thus, PHIP may occur in a solution formed with an organic solvent. In some embodiments, the aqueous solution is used for hydrogenation and polarization transfer. In some embodiments, this aqueous solution may be mixed with a miscible organic solvent before precipitation of the hyperpolarized bio-related contrast agent.

[0119] In some embodiments, precipitation of bio-relevant contrast agents can be induced by changing the pH of the solvent. In organic solvents, certain bio-relevant contrast agents (e.g., carboxylic acids, pyridines, etc.) may be soluble at concentrations suitable for polarization using PHIP. However, salts of these bio-relevant contrast agents may be highly insoluble. For example, these salts have a maximum concentration of approximately 10mM, 9mM, 8mM, 7mM, 6mM, 5mM, 4mM, 3mM, 2mM, 1mM, 1mM, 0.9mM, 0.8mM, 0.7mM, 0.6mM, 0.5mM, 0.4mM, 0.3mM, 0.2mM, 0.1mM, 0.09mM, 0.08mM, 0.07mM, 0.06mM, 0.05mM, 0.04mM, 0.03mM, 0.02mM, 0.01mM, or less, at least approximately 0.01mM, 0.02 It may be insoluble at concentrations of mM, 0.03 mM, 0.04 mM, 0.05 mM, 0.06 mM, 0.07 mM, 0.08 mM, 0.09 mM, 0.1 mM, 0.2 mM, 0.3 mM, 0.04 mM, 0.05 mM, 0.6 mM, 0.7 mM, 0.8 mM, 0.9 mM, 1 mM, 2 mM, 3 mM, 4 mM, 5 mM, 6 mM, 7 mM, 8 mM, 9 mM, 10 mM, or higher, or within the range defined by any two of the aforementioned values. The organic solution may contain a bio-related contrast agent at a concentration suitable for hyperpolarization using PHIP. After polarization, the pH of the organic solution may be changed to induce precipitation of salts of the bio-related contrast agent. In aqueous solutions, salts of certain biocompatible contrast agents (e.g., fumarates, glutamates, etc.) are more soluble than their acidic forms. Therefore, aqueous solutions can contain biocompatible contrast agents at concentrations suitable for hyperpolarization using PHIP. After polarization, the pH of the aqueous solution can be lowered to induce precipitation of the acidic form of the biocompatible contrast agent.

[0120] In accordance with the disclosed embodiments, a pH change can be induced by the addition of an acidic or basic molecule to a solution such as sodium chloride or sodium hydroxide. In some embodiments, a pH change can be induced by mixing the solution with another solution having a substantially different pH. In some embodiments, precipitation due to a pH change occurs for a maximum of about 100 seconds, 90 seconds, 80 seconds, 70 seconds, 60 seconds, 50 seconds, 40 seconds, 30 seconds, 20 seconds, 10 seconds, 9 seconds, 8 seconds, 7 seconds, 6 seconds, 5 seconds, 4 seconds, 3 seconds, 2 seconds, 1 second or less, and for at least about 1 second, 2 seconds, 3 seconds, 4 seconds, 5 seconds, 6 seconds, 7 seconds, 8 seconds, 9 seconds, 10 seconds, 20 seconds, 30 seconds, 40 seconds, 50 seconds, 60 seconds, 70 seconds, 80 seconds, 90 seconds, 100 seconds or more, or within the range defined by any two of the aforementioned values.

[0121] In some embodiments, precipitation of a bio-relevant contrast agent can be induced without changing the pH of the solution, which induces the hyperpolarized bio-relevant contrast agent to change between salt and acid forms.

[0122] In some embodiments, hydrogenation and polarization transfer occur in a first solution having a first solvent. The bio-relevant contrast agent may have high solubility in the first solvent. Precipitation can be induced by mixing the first solution with a second solvent to form a second solution. The second solvent may be selected such that the solubility of the bio-relevant contrast agent in the second solution is low enough to initiate precipitation.

[0123] In some embodiments, the temperature of the solution changes, thereby reducing the solubility of hyperpolarized molecules and initiating precipitation. In most solvents, solubility decreases as the temperature decreases. The change in temperature to a desired temperature with lower solubility may be carried out for a maximum of about 100 seconds, 90 seconds, 80 seconds, 70 seconds, 60 seconds, 50 seconds, 40 seconds, 30 seconds, 20 seconds, 10 seconds, 9 seconds, 8 seconds, 7 seconds, 6 seconds, 5 seconds, 4 seconds, 3 seconds, 2 seconds, 1 second, or less, and at least within about 1 second, 2 seconds, 3 seconds, 4 seconds, 5 seconds, 6 seconds, 7 seconds, 8 seconds, 9 seconds, 10 seconds, 20 seconds, 30 seconds, 40 seconds, 50 seconds, 60 seconds, 70 seconds, 80 seconds, 90 seconds, 100 seconds, or more, or within the range defined by any two of the aforementioned values. For example, the maximum molar fraction of acetic acid in n-heptane is 0.935 at 14.8°C, only 0.02 at -29.2°C, and even lower at lower temperatures. The temperature chosen for precipitation can be selected as a temperature above the freezing point of the solvent.

[0124] In some embodiments, the surface area of ​​the solution is increased to induce nucleation of the precipitate. This can be achieved, for example, by spraying the solution through a nozzle to create very small droplets with a high surface area. In some embodiments, microcrystalline seeds (which may be biocompatible contrast agents or other biocompatible compounds) are added to the solution to induce precipitation.

[0125] In some embodiments, the pressure of the solution changes, thereby reducing the solubility of the hyperpolarized compound and initiating precipitation.

[0126] In some embodiments, the concentration of a hyperpolarized bio-relevant contrast agent rises above its solubility limit, thereby inducing precipitation without reducing the solubility level of the compound in the solvent. This can be achieved, for example, by adding unpolarized molecules of the bio-relevant contrast agent or by evaporating a specific volume of the solvent, thereby increasing the concentration of the bio-relevant contrast agent above its solubility limit.

[0127] In some embodiments, the polarization parameters of PHIP are optimized to exceed the solubility limit of its acid or salt form of the biorelevant contrast agent, for example, at the expense of achieving lower polarization, for high concentrations. This can be achieved, for example, by starting with a high concentration of precursor for hydrogenation and selecting a long hydrogenation time that allows polarization to decrease due to relaxation, in order to achieve high concentrations of the biorelevant contrast agent.

[0128] In some embodiments, precipitation is accelerated by the addition of mechanical energy or by improved mixing of the mixture over a certain period of time. This can be done, for example, by applying ultrasound to the mixture through an ultrasonic solicitor, or by mechanical or magnetic mixing of the sample. In some embodiments, the additional mixing or introduction of mechanical energy is performed for a period of at least about 0.1 seconds, 0.2 seconds, 0.3 seconds, 0.4 seconds, 0.5 seconds, 0.6 seconds, 0.7 seconds, 0.8 seconds, 0.9 seconds, 1 second, 2 seconds, 3 seconds, 4 seconds, 5 seconds, 6 seconds, 7 seconds, 8 seconds, 9 seconds, 10 seconds or more, and at most about 10 seconds, 9 seconds, 8 seconds, 7 seconds, 6 seconds, 5 seconds, 4 seconds, 3 seconds, 2 seconds, 1 second, 0.9 seconds, 0.8 seconds, 0.7 seconds, 0.6 seconds, 0.5 seconds, 0.4 seconds, 0.3 seconds, 0.2 seconds, 0.1 seconds or less, or within the range defined by any two of the aforementioned values.

[0129] In some embodiments, precipitation is induced by a chemical reaction involving a hyperpolarized bio-relevant contrast agent. The hyperpolarized bio-relevant contrast agent may react with another compound or in response to an external stimulus such as electromagnetic radiation (e.g., ultraviolet irradiation). The product of this reaction has reduced solubility compared to the hyperpolarized bio-relevant contrast agent, thereby inducing precipitation. For example, the external stimulus may modify the structure of the hyperpolarized bio-relevant contrast agent, thereby reducing its solubility. Following the redissolution of the precipitate, additional reactions may be carried out to produce the desired final product (e.g., a bio-relevant contrast agent, an NMR material, etc.).

[0130] In some embodiments, hydrolysis of the precursor induces precipitation. In some embodiments, the solvent is selected such that the precursor is more soluble than the bio-relevant contrast agent. The concentration of the precursor in the solution may be selected so that the bio-relevant contrast agent precipitates from the solution after cleavage of the side chains and generation of the bio-relevant contrast agent. In some such embodiments, cleavage is initiated by changing the pH of the solution. For example, cleavage may be initiated by adding a base (e.g., sodium hydroxide or another suitable base). In some embodiments, the solution may be formed using an organic solvent, and cleavage is carried out under basic conditions. After cleavage, the less soluble bio-relevant contrast agent undergoes rapid precipitation while retaining its polarization. The same solution may be used for hydrogenation, polarization transfer, and precipitation, or a different solvent may be mixed with the solution used for hydrogenation and polarization transfer.

[0131] In some embodiments, precursor precipitation occurs before cleavage. Such embodiments may be preferred when the precursor is more stable than the bio-relevant contrast agent. Precipitation of the precursor can be induced by modifying the pH of the solution so that the solubility of the precursor is reduced, or by mixing the precursor in a solution or compound that reduces its solubility. In such embodiments, cleavage of the precursor can be carried out after the precursor is redissolved in the solvent. The precursor can be filtered and washed as described herein to remove other substances present in the original solution, such as a hydrogenation catalyst. The precursor can then be reacted to form a bio-relevant contrast agent (e.g., by cleaving side chains). The bio-relevant contrast agent can be separated from other reaction products by liquid-liquid extraction or by an additional precipitation step, consistent with the precipitation methods described herein. In some embodiments, precipitation may occur after the formation of the bio-relevant contrast agent from the precursor. Such precipitation can be carried out according to the methods described herein.

[0132] In some embodiments, the conversion of spin order to polarization occurs before the compound solidifies and precipitates. In other embodiments, the conversion occurs after the redissolution of the crystal with a biocompatible contrast agent.

[0133] In some embodiments, several steps among precipitation, washing, and redissolution are performed. This may be advantageous in further purifying the bio-related contrast agent, increasing the relaxation time of the precipitate, or further separating the polarization and cleavage steps. For example, the first precipitate may be used to wash the catalyst, while the second precipitate is a crystalline form with a longer relaxation time and may be used for transport. In some embodiments, the first precipitate is an ester of the compound of formula II after hydrogenation and polarization transfer, and the second precipitate is performed after cleavage.

[0134] In accordance with the disclosed embodiments, the precipitation (and optionally washing) step can separate the hyperpolarized bio-related contrast agent from other substances in the original solution (e.g., catalyst, original solvent, reaction product, etc.). For example, a large portion of the hydrogenation catalyst present in the original solution may be retained in the original solution after precipitation of the bio-related contrast agent. In some embodiments, the precipitate (after optionally washing) is up to about 1%, 0.9%, 0.8%, 0.7%, 0.6%, 0.5%, 0.4%, 0.3%, 0.2%, 0.1%, 0.09%, 0.08%, 0.07%, 0.06%, 0.05%, 0.04%, 0.03%, 0.02%, 0.01%, 0.009%, 0.008%, 0.007%, 0.006%, 0.005%, 0.004%, 0.003%, 0.002%, 0.001%, or less of the hydrogenation catalyst, at least about 0.001%. It can hold hydrogenation catalyst in amounts of 0.002%, 0.003%, 0.004%, 0.005%, 0.006%, 0.007%, 0.008%, 0.009%, 0.01%, 0.02%, 0.03%, 0.04%, 0.05%, 0.06%, 0.07%, 0.08%, 0.09%, 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1%, or more, or an amount of hydrogenation catalyst within the range defined by any two of the aforementioned values.Similarly, precipitates may consist of up to approximately 1%, 0.9%, 0.8%, 0.7%, 0.6%, 0.5%, 0.4%, 0.3%, 0.2%, 0.1%, 0.09%, 0.08%, 0.07%, 0.06%, 0.05%, 0.04%, 0.03%, 0.02%, 0.01%, 0.009%, 0.008%, 0.007%, 0.006%, 0.005%, 0.004%, 0.003%, 0.002%, 0.001%, or less, cleavage by-products (e.g., side arms or other residues from the cleavage, at least approximately 0.001%). It may retain cleavage by-products in the following proportions: %, 0.002%, 0.003%, 0.004%, 0.005%, 0.006%, 0.007%, 0.008%, 0.009%, 0.01%, 0.02%, 0.03%, 0.04%, 0.05%, 0.06%, 0.07%, 0.08%, 0.09%, 0.1%, 0.2%, 0.3%, 0.4%, 0.5%, 0.6%, 0.7%, 0.8%, 0.9%, 1%, or more, or an amount of cleavage by-products within the range defined by any two of the aforementioned values.

[0135] In some embodiments, after precipitation and separation from the original solvent (and potential transport of the hyperpolarized precipitate), the precipitate is redissolved in the solvent (for example, for use as a drug in hyperpolarized NMR or MRI). The solvent may be a biocompatible solvent such as an aqueous solution. In some embodiments, redissolution may be carried out for a period of at least about 1 second, 2 seconds, 3 seconds, 4 seconds, 5 seconds, 60 seconds, 60 seconds, or longer, or within the range defined by any two of the aforementioned values, with a maximum of about 60 seconds, 50 seconds, 40 seconds, 30 seconds, 20 seconds, 1 second, or less.

[0136] In some embodiments, after redissolution, the bio-related contrast agent may be present at a higher concentration than that used during PHIP polarization. In some embodiments, the concentration of the hyperpolarized bio-related contrast agent after redissolution is at least about 100 mM, 150 mM, 200 mM, 250 mM, 300 mM, 350 mM, 400 mM, 450 mM, 500 mM or higher, and at most about 500 mM, 450 mM, 400 mM, 350 mM, 300 mM, 250 mM, 200 mM, 150 mM, 100 mM or lower, or within the range defined by any two of the aforementioned values. Therefore, the concentration of the precursor or bio-related contrast agent during polarization may be independent of the concentration of molecules in the injection solution. The concentration of the precursor or bio-related contrast agent during polarization may be selected for efficient polarization transfer. In some embodiments, this polarization concentration is less than the concentration of the hyperpolarized bio-related contrast agent after redissolution. For example, the polarization concentration may be at least about 300 mM, 200 mM, 150 mM, 140 mM, 130 mM, 120 mM, 110 mM, 100 mM, 90 mM, 80 mM, 70 mM, 60 mM, 50 mM, 40 mM, 30 mM, 20 mM, 10 mM, or less, and at most about 10 mM, 20 mM, 30 mM, 40 mM, 50 mM, 60 mM, 70 mM, 80 mM, 90 mM, 100 mM, 110 mM, 120 mM, 130 mM, 140 mM, 150 mM, 200 mM, 300 mM, or more, or within the range defined by any two of the aforementioned values.

[0137] In some embodiments, the precipitate sample exhibits polarization of at least about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 40%, 45%, 50%, or more, and at most about 50%, 45%, 40%, 35%, 30%, 25%, 20%, 15%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less, or within the range defined by any two of the aforementioned values. For example, in some embodiments, the precipitate sample is 10%~15%, 10%~20%, 10%~25%, 10%~30%, 10%~35%, 10%~40%, 10%~45%, 10%~50%, 15%~20%, 15%~25%, 15%~30%, 15%~35%, 15%~40%, 15%~45%, 15%~50%, 20%~25%, 20%~30%, 2 It exhibits polarization in the following ranges: 0%~35%, 20%~40%, 20%~45%, 20%~50%, 25%~30%, 25%~35%, 25%~40%, 25%~45%, 25%~50%, 30%~35%, 30%~40%, 30%~45%, 30%~50%, 35%~40%, 35%~45%, 35%~50%, 40%~45%, 40%~50%, or 45%~50%.

[0138] After the redissolution of the particles, the concentrations of the catalyst, precursor, or cleavage byproducts are, respectively, up to approximately 1 μM, 900 nanomoles (nM), 800 nM, 700 nM, 600 nM, 500 nM, 400 nM, 300 nM, 200 nM, 100 nM, 90 nM, 80 nM, 70 nM, 60 nM, 50 nM, 40 nM, 30 nM, 20 nM, 10 nM, 9 nM, 8 nM, 7 nM, 6 nM, 5 nM, 4 nM, 3 nM, 2 nM, 1 nM, or less. It may be at least about 1 nM, 2 nM, 3 nM, 4 nM, 5 nM, 6 nM, 7 nM, 8 nM, 9 nM, 10 nM, 20 nM, 30 nM, 40 nM, 50 nM, 60 nM, 70 nM, 80 nM, 90 nM, 100 nM, 200 nM, 300 nM, 400 nM, 500 nM, 600 nM, 700 nM, 800 nM, 900 nM, 1 μM, or more, or within the range defined by any two of the aforementioned values. In some embodiments, the purity of the hyperpolarized bio-related contrast agent after redissolution is at least about 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or higher, and at most about 99%, 98%, 97%, 96%, 95%, 94%, 93%, 92%, 91%, 90%, or lower, or within the range defined by any two of the aforementioned values. In some embodiments, at least the hyperpolarized compound fraction is separated from cleaved side chains or, if present, other reaction by-products.

[0139] transportation In line with the disclosed embodiments, the polarization transfer and use of bio-related contrast agents may occur at different locations. In some embodiments, the precursor (in the solution used for polarization transfer, or as a precipitate) is transported to another location after precipitation. In some embodiments, the bio-related contrast agent (in the solution used for polarization transfer, as a precipitate, or redissolved) is transported to another location. The disclosed embodiments are not necessarily limited to any particular transport distance or duration. Instead, the maximum distance or duration may be determined based on the target molecule, its initial degree or polarization, the desired final degree of polarization, and the transport conditions. In some embodiments, the precipitate is transported at least 1 meter in a suitable transport device.

[0140] In accordance with the disclosed embodiments, the transport device may be configured to transport a sample of a precursor or a bio-related contrast agent. The transport device may be arranged and configured to transport one or more samples simultaneously. The transport device may include a transport chamber configured to receive one or more samples. The transport device may be configured to maintain the transport chamber within a predetermined temperature range and a predetermined magnetic field strength. The transport device may be configured to maintain one or more samples in a magnetic field of at least approximately 10G, 20G, 30G, 40G, 50G, 60G, 70G, 80G, 90G, 100G, 200G, 300G, 400G, 500G, 600G, 700G, 800G, 900G, 1,000G or higher, and at most approximately 1,000G, 900G, 800G, 700G, 600G, 500G, 400G, 300G, 200G, 100G, 90G, 80G, 70G, 60G, 50G, 40G, 30G, 20G, 10G or lower, or within the range defined by any two of the aforementioned values.

[0141] Permanent magnets or electromagnets included in the transport device can provide a magnetic field. In some embodiments, the permanent magnets or electromagnets are shielded to reduce the strength of the magnetic field outside the transport device. The transport device may also include a cooling system. The cooling system may be configured to maintain the sample at a predetermined temperature or within a predetermined temperature range during transport. For example, the cooling system may be configured to maintain the sample at a temperature below 270K, below 80K, or below 4K. In some embodiments, the transport device is configured to maintain the sample at approximately liquid nitrogen temperature. The transport device may include insulation between the cooling system and the outside of the transport device to minimize heat exchange with the external environment. In some embodiments, the cooling system is configured to maintain the sample temperature using a cold gas flow. In some embodiments, the cooling system is configured to maintain the sample temperature using a coolant. In some embodiments, the transport device includes a dewar to provide cooling for the sample. To distribute hyperpolarized samples over long distances, containers may be transported by standard transport vehicles such as airplanes, trains, trucks, cars, and ships.

[0142] In some embodiments, the hyperpolarized precipitate particles are transported within a transport device. In some embodiments, the relaxation time of the hyperpolarized precipitate particles within the transport device is at least about 1 minute, 2 minutes, 3 minutes, 4 minutes, 5 minutes, 6 minutes, 7 minutes, 8 minutes, 9 minutes, 10 minutes, 20 minutes, 30 minutes, 40 minutes, 50 minutes, 1 hour, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, 8 hours, 9 hours, 10 hours or more, and at most about 10 hours, 9 hours, 8 hours, 7 hours, 6 hours, 5 hours, 4 hours, 3 hours, 2 hours, 1 hour, 50 minutes, 40 minutes, 30 minutes, 20 minutes, 10 minutes, 9 minutes, 8 minutes, 7 minutes, 6 minutes, 5 minutes, 4 minutes, 3 minutes, 2 minutes, 1 minute or less, or within the range defined by any two of the aforementioned values.

[0143] Generation of polarized bio-associated contrast agents Figure 1 shows a first exemplary process 100 for generating a polarized bio-relevant contrast agent according to various embodiments. In the embodiments shown, process 100 includes providing a composition comprising a compound of formula I in step 110. In some embodiments, the compound of formula I is (i) 1 H (proton), 2 The R1 group comprises a PHIP transfer moiety as described herein, the R2 group comprises an optionally substituted hydrocarbon, alkoxy group, primary amine, secondary amine, or tertiary amine, or a solubilizing moiety as described herein, and the R3 group comprises a bio-related contrast agent as described herein.

[0144] In step 120, the double or triple bond in the compound of formula I is hydrogenated with parahydrogen to form a parahydrogenated derivative of the compound of formula I, and the parahydrogenated derivative is a compound having the structure of formula II. In some embodiments, the compound of formula II is (i) 1 H (proton), 2 (ii) Parahydrogenated carbon-carbon single bonds (-CH*-CH*-) substituted to include H (deuterium) or a combination thereof 1 H (proton), 2 The compound comprises Z', which is a parahydrogenated carbon-carbon double bond (-CH*=CH*-) substituted to include H (deuterium) or a combination thereof, where H* is a hydrogen having spin order derived from parahydrogen, the R1 group comprises a PHIP transfer moiety as described herein, the R2 group comprises an optionally substituted hydrocarbon, alkoxy group, primary amine, secondary amine, or tertiary amine, or solubilizing moiety as described herein, and the R3 group comprises a bio-related contrast agent as described herein. In some embodiments, the compound of formula I is hydrogenated with parahydrogen using the hydrogenation process described herein.

[0145] In step 130, a polarization transfer waveform is applied to transfer nuclear spin order from at least one H* in the side arm of the compound of formula II to any non-hydrogen nuclear spin in the bio-relevant contrast agent of the compound of formula II, as described herein, thereby forming a derivative of the compound of formula II having a hyperpolarized bio-relevant contrast agent. In some embodiments, the nuclear spin order is transferred using any polarization transfer process described herein.

[0146] Figure 2 shows a second exemplary process 200 for generating a polarized bio-relevant contrast agent according to various embodiments of the present disclosure. In the embodiments shown, process 200 includes providing a composition comprising a compound of formula II in step 210. In some embodiments, formula II is (i) 1 H (proton), 2 Parahydrogenated carbon-carbon single bonds (-CH*-CH*-) substituted to include H (deuterium) or a combination thereof, or (ii) as described herein, 1 H (proton), 2 The R1 group comprises a Z' group which is a parahydrogenated carbon-carbon double bond (-CH*=CH*-) substituted to include H (deuterium) or a combination thereof, where H* is a spin-ordered hydrogen derived from parahydrogen, the R1 group comprises a PHIP transfer moiety as described herein, the R2 group comprises an optionally substituted hydrocarbon, alkoxy group, primary amine, secondary amine, or tertiary amine, or solubilizing moiety as described herein, and the R3 group comprises a bio-related contrast agent as described herein.

[0147] In step 220, a polarization transfer waveform is applied to transfer nuclear spin order from at least one H* in the side arm of the compound of formula II to any non-hydrogen nuclear spin in the bio-relevant contrast agent of the compound of formula II, as described herein, thereby forming a derivative of the compound of formula II having a hyperpolarized bio-relevant contrast agent.

[0148] In step 230, the derivative compound of formula II is hydrolyzed to form a composition comprising a hyperpolarized bio-related contrast agent and a separate side-arm compound of formula III. In some embodiments, the compound of formula III is (i) 1 H (proton), 2 Parahydrogenated carbon-carbon single bonds (-CH*-CH*-) substituted to include H (deuterium) or a combination thereof, or (ii) as described herein, 1 H (proton), 2 The R1' group comprises a parahydrogenated carbon-carbon double bond (-CH*=CH*-) Z'' which is substituted to include H (deuterium) or a combination thereof, the R1' group comprises a parahydrogen-induced polarization (PHIP) transfer moiety as described herein, and the R2 group comprises an optionally substituted hydrocarbon, alkoxy group, primary amine, secondary amine, or tertiary amine, or solubilizing moiety as described herein.

[0149] In step 240, the hyperpolarized bio-related contrast agent is washed once or more times with an organic solvent. In some embodiments, the non-hydrogen nuclear spins in the bio-related contrast agent have non-hydrogen spin polarizations within the range defined by at least about 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, or more after the washing step, and at most about 50%, 45%, 40%, 35%, 30%, 25%, 20%, 15%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, or less after the washing step, or within the range defined by any two of the aforementioned values.

[0150] In some embodiments, process 100 or process 200 includes one or more additional steps or operations. In some embodiments, process 100 or process 200 omits one or more steps or operations. In some embodiments, one or more steps or operations of process 100 are combined with one or more steps or operations of process 200. In some embodiments, all steps or operations of process 100 and process 200 are combined to obtain a complete process for producing a hyperpolarized contrast agent from a precursor having the structure of formula I.

[0151] Enumerated embodiments The aforementioned non-limiting embodiments disclosed herein include:

[0152] Embodiment 1. A composition comprising the compound of formula (I), [ka] In the formula, Z is (i) 1 H (proton), 2 A composition comprising (ii) a carbon-carbon double bond (-C=C-) or a carbon-carbon triple bond (-C≡C-) substituted to include H (deuterium) or a combination thereof, wherein R1 comprises a parahydrogen-induced polarization (PHIP) transfer moiety, R2 comprises an optionally substituted hydrocarbon, alkoxy group, primary amine, secondary amine, or tertiary amine, and R3 comprises a bio-related contrast agent containing a non-hydrogen nuclear spin.

[0153] Embodiment 2. A composition comprising the compound of formula (II), [ka] In the formula, Z' is (i) 1 H (proton), 2 (ii) Parahydrogenated carbon-carbon single bonds (-CH*-CH*-) substituted to include H (deuterium) or a combination thereof, or 1 H (proton), 2A composition comprising a parahydrogenated carbon-carbon double bond (-CH*=CH*-) substituted to include H (deuterium) or a combination thereof, wherein H* is a hydrogen having a spin order derived from parahydrogen, R1 comprises a parahydrogen-induced polarization (PHIP) transfer portion, R2 comprises an optionally substituted hydrocarbon, alkoxy group, primary amine, secondary amine, or tertiary amine, and R3 comprises a bio-related contrast agent containing a non-hydrogen nuclear spin.

[0154] Embodiment 3. A composition comprising (i) a bio-related contrast agent containing non-hydrogen nuclear spins and (ii) a compound of formula (III), [ka] In the formula, Z'' is (i) 1 H (proton), 2 (ii) Parahydrogenated carbon-carbon single bonds (-CH*-CH*-) substituted to include H (deuterium) or a combination thereof, or 1 H (proton), 2 A composition comprising a parahydrogenated carbon-carbon double bond (-CH*=CH*-) substituted to include H (deuterium) or a combination thereof, wherein R1' contains a parahydrogen-induced polarization (PHIP) transfer moiety, and R2 contains an optionally substituted hydrocarbon, alkoxy group, primary amine, secondary amine, or tertiary amine.

[0155] Embodiment 4. (i) A composition comprising a hyperpolarized bio-related contrast agent containing non-hydrogen nuclear spins, and (ii) a compound of formula (IV), [ka] In the formula, Z is (i) 1 H (proton), 2A composition comprising (ii) a carbon-carbon double bond (-C=C-) substituted to include H (deuterium) or a combination thereof, or (ii) a carbon-carbon triple bond (-C≡C-), wherein R1' comprises a para-hydrogen-induced polarization (PHIP) transfer moiety, and R2 comprises an optionally substituted hydrocarbon, alkoxy group, primary amine, secondary amine, or tertiary amine.

[0156] Embodiment 5. The composition according to any one of Embodiments 1 to 4, wherein the PHIP transfer portion comprises an optionally substituted C1 hydrocarbon or an optionally substituted C2 hydrocarbon.

[0157] Embodiment 6. The PHIP moving portion includes *CR4R5, *CR4Y, *C=Y, or any deuterated version thereof, where *C is 12 C or 13 It is a carbon isotope of C, and R4 and R5 are each independent of each other. 1 H, 2 H, 3 The composition according to any one of Embodiments 1 to 5, wherein H is selected from linear, branched, or cyclic C1-C10 alkyl hydrocarbons, C6 aryl, benzyl, phenyl, heteroaryl, and haloalkyl groups, and Y is selected from a spin 1 / 2 atom and a spin 1 / 2 atom covalently bonded to one or more chemical parts selected from heteroatoms such as N, O, and S that are optionally substituted with linear, branched, or cyclic C1-C10 alkyl hydrocarbons, C6 aryl, benzyl, phenyl, heteroaryl, halogen, or haloalkyl groups, or linear, branched, or cyclic C1-C10 alkyl hydrocarbons, C6 aryl, benzyl, phenyl, heteroaryl, halogen, or haloalkyl groups.

[0158] Embodiment 7. The PHIP moving portion includes *CR6R7-*CR8R9, or any deuterated version thereof, where *C is 12 C or 13 The carbon isotopes R6, R7, R8, and R9 are each independently... 1 H, 2 H, 3A composition according to any one of Embodiments 1 to 5, selected from H, linear, branched, or cyclic C1-C10 alkyl hydrocarbons, C6 aryl, benzyl, phenyl, heteroaryl, and haloalkyl groups.

[0159] Embodiment 8. The PHIP moving portion includes *CH2, *CH2-*CH2, *CHY, *C=Y, or any deuterated version thereof, where *C is 12 C or 13 The composition according to any one of Embodiments 1 to 5, wherein Y is a carbon isotope of C, and Y is selected from a spin 1 / 2 atom and a spin 1 / 2 atom covalently bonded to one or more chemical parts selected from heteroatoms such as N, O, and S that are optionally substituted with a linear, branched, or cyclic C1-C10 alkyl hydrocarbon, C6 aryl, benzyl, phenyl, heteroaryl, halogen, or haloalkyl group, or a linear, branched, or cyclic C1-C10 alkyl hydrocarbon, C6 aryl, benzyl, phenyl, heteroaryl, halogen, or haloalkyl group.

[0160] Embodiment 9. Spin 1 / 2 atoms, 1 H, 13 C 15 N, 19 F, or 31 A composition according to embodiment 6 or 8, selected from P.

[0161] Embodiment 10. The composition according to any one of Embodiments 1 to 9, wherein the PHIP moving portion includes at least one atom that is J-coupled with a non-hydrogen nucleus spin of at least 0.1 Hertz (Hz).

[0162] Embodiment 11. The composition according to any one of Embodiments 1 to 10, wherein Z comprises at least one atom that is J-coupled with a non-hydrogen nuclear spin of at least 0.1 Hertz (Hz).

[0163] Embodiment 12.R2 is the composition according to any one of Embodiments 1 to 11, wherein the composition includes a solubilized portion.

[0164] Embodiment 13. The composition according to any one of Embodiments 1 to 12, wherein R2 comprises a hydrophobic and / or organic affinity moiety.

[0165] Embodiment 14. The composition according to Embodiment 13, wherein R2 includes an organically solubilized portion.

[0166] Embodiment 15. The composition according to any one of Embodiments 1 to 12, wherein R2 includes a hydrophilic and / or organic affinity moiety.

[0167] Embodiment 16. A composition according to any one of Embodiments 1 to 15, wherein R2 comprises or is selected from a methyl group, an ethyl group, a propyl group, an isopropyl group, an n-butyl group, an s-butyl group, a t-butyl group, an isobutyl group, a hydroxyl group, a methyl alcohol group, an ethyl alcohol group, an n-propanol group, an isopropyl alcohol group, an alcohol propionic acid group, an n-butyl alcohol group, an s-butyl alcohol group, a t-butyl alcohol group, an isobutyl alcohol group, a methoxy group, an ethoxy group, an ethoxy group, an isopropoxy group, an alcohol propionic acid group, an ethoxy group, a t-butoxy group, an s-butoxy group, an ester group, a phenyl group, a substituted phenyl group, a primary amine group, a secondary amine group, a tertiary amine group, a primary amide group, a secondary amide group, and a tertiary amide group.

[0168] Embodiment 17. A bio-related contrast agent is, Formula R 10 It contains a compound C(=O)X-, in which R 10 However, selected from linear, branched, or cyclic C1-C10 alkyl groups, one or more C atoms are optionally substituted with C=C, CO, COH, CNH2, COOH, CH2COOH, CONH2, or OC(=O), and X is NR 11 Selected from , S, and O, in the formula, R 11 but, 1 H, 2 H, 3A composition according to any one of Embodiments 1 to 16, wherein H and an amino protecting group optionally selected from trifluoroacetyl, acetyl, benzoyl, carbobenzoxy, tert-butyl carbonate, and benzyl.

[0169] Embodiment 18. The composition according to any one of Embodiments 1 to 17, wherein the bio-related contrast agent is selected from pyruvate, glutamate, glutamine, lactic acid, acetic acid, acetoacetic acid, zymonate, alanine, fructose, fumaric acid, bicarbonate, urea, dehydroascorbic acid, α-ketoglutaric acid, dihydroxyacetone, glucose, ascorbic acid, and their conjugate acids.

[0170] Embodiment 19. The composition according to any one of Embodiments 1 to 18, wherein the composition has a solubility in water of less than 50 millimoles (mM).

[0171] Embodiment 19a. The composition according to any one of Embodiments 1 to 18, wherein the composition has a solubility in an organic solvent (e.g., acetone, ethanol, chloroform, toluene) of less than 50 mmol (mM).

[0172] Embodiment 20. The composition according to any one of Embodiments 1 to 19a, wherein reacting the composition with parahydrogen yields a chemical yield of at least 30% of the parahydrogenated product.

[0173] Embodiment 21. A composition according to any one of Embodiments 1 to 20, for use in a para-hydrogen-induced polarization (PHIP) process.

[0174] Embodiment 22. A method for preparing a hyperpolarized bio-relevant contrast agent or a pharmaceutically acceptable salt thereof, wherein the method provides a composition comprising a compound of formula (I), [ka] In the formula, Z is (i) 1 H (proton),2 (ii) a carbon-carbon double bond (-C=C-) or a carbon-carbon triple bond (-C≡C-) substituted to include H (deuterium) or a combination thereof, wherein R1 comprises a parahydrogen-induced polarization (PHIP) transfer moiety, R2 comprises an optionally substituted hydrocarbon, alkoxy group, primary amine, secondary amine, or tertiary amine, and R3 comprises a bio-related contrast agent containing a non-hydrogen nuclear spin, and (b) a double bond or triple bond in a compound of formula I is hydrogenated with parahydrogen to form a parahydrogenated derivative of the compound of formula I, wherein the parahydrogenated derivative has the structure of formula (II), [ka] In the formula, Z' is (i) 1 H (proton), 2 (ii) Parahydrogenated carbon-carbon single bonds (-CH*-CH*-) substituted to include H (deuterium) or a combination thereof, or 1 H (proton), 2 A method comprising (c) forming a parahydrogenated carbon-carbon double bond (-CH*=CH*-) which is substituted to include H (deuterium) or a combination thereof, wherein H* is a hydrogen having a spin order derived from parahydrogen, R1 includes a parahydrogen-induced polarization (PHIP) transfer portion, R2 includes an optionally substituted hydrocarbon, alkoxy group, primary amine, secondary amine, or tertiary amine, and R3 includes a bio-relevant contrast agent containing a non-hydrogen nuclear spin, and (c) applying a polarization transfer waveform to the non-hydrogen nuclear spin to transfer the nuclear spin order from at least one H* in the compound of formula II, thereby forming a derivative of formula II having a hyperpolarized bio-relevant contrast agent.

[0175] Embodiment 23. A method for preparing a hyperpolarized bio-relevant contrast agent or a pharmaceutically acceptable salt thereof, wherein the method provides (a) a composition comprising formula (II) of a compound, [ka] In the formula, Z' is (i) 1 H (proton), 2 (ii) Parahydrogenated carbon-carbon single bonds (-CH*-CH*-) substituted to include H (deuterium) or a combination thereof, or 1 H (proton), 2 (b) A parahydrogenated carbon-carbon double bond (-CH*=CH*-) substituted to include H (deuterium) or a combination thereof, wherein H* is a hydrogen having a spin order derived from parahydrogen, R1 comprises a parahydrogen-induced polarization (PHIP) transfer portion, R2 comprises an optionally substituted hydrocarbon, alkoxy group, primary amine, secondary amine, or tertiary amine, and R3 comprises a bio-relevant contrast agent containing a non-hydrogen nuclear spin; and a method comprising (b) applying a polarization transfer waveform to a non-hydrogen nuclear spin to transfer nuclear spin order from at least one H* in the compound of formula II, thereby forming a derivative of formula II having a hyperpolarized bio-relevant contrast agent.

[0176] Embodiment 24. Hydrolysis of a derivative of formula II yields (i) a hyperpolarized bio-related contrast agent containing non-hydrogen nuclear spins, and (ii) a compound of formula (III), [ka] In the formula, Z'' is (i) 1 H (proton), 2 (ii) Parahydrogenated carbon-carbon single bonds (-CH*-CH*-) substituted to include H (deuterium) or a combination thereof, or 1 H (proton), 2 The method according to Embodiment 22 or 23, further comprising providing a composition comprising a compound having a parahydrogenated carbon-carbon double bond (-CH*=CH*-) substituted to include H (deuterium) or a combination thereof, wherein R1' includes a parahydrogen-induced polarization (PHIP) transfer moiety, and R2 comprises an optionally substituted hydrocarbon, alkoxy group, primary amine, secondary amine, or tertiary amine.

[0177] Embodiment 25. The method according to Embodiment 24, further comprising washing the hyperpolarized bio-related contrast agent with an organic solvent once or more times.

[0178] Embodiment 26. The method according to Embodiment 25, wherein the non-hydrogen nucleus spins have more than 10% non-hydrogen nucleus spin polarization after the washing step.

[0179] Embodiment 27. The method according to any one of Embodiments 22 to 26, wherein the PHIP transfer portion comprises an optionally substituted C1 hydrocarbon or an optionally substituted C2 hydrocarbon.

[0180] Embodiment 28. The PHIP moving portion includes *CR4R5, *CR4Y, *C=Y, or any deuterated version thereof, where *C is 12 C or 13 It is a carbon isotope of C, and R4 and R5 are each independent of each other. 1 H, 2 H, 3 The method according to any one of Embodiments 22 to 26, wherein H is selected from linear, branched, or cyclic C1-C10 alkyl hydrocarbons, C6 aryl, benzyl, phenyl, heteroaryl, and haloalkyl groups, and Y is selected from a spin 1 / 2 atom and a spin 1 / 2 atom covalently bonded to one or more chemical parts selected from heteroatoms such as N, O, and S, which are optionally substituted with linear, branched, or cyclic C1-C10 alkyl hydrocarbons, C6 aryl, benzyl, phenyl, heteroaryl, halogen, or haloalkyl groups.

[0181] Embodiment 29. The PHIP moving portion includes *CR6R7-*CR8R9, or any deuterated version thereof, where *C is 12 C or 13 The carbon isotopes R6, R7, R8, and R9 are each independently... 1 H, 2 H, 3The method according to any one of embodiments 22 to 26, selected from H, linear, branched, or cyclic C1-C10 alkyl hydrocarbons, C6 aryl, benzyl, phenyl, heteroaryl, and haloalkyl groups.

[0182] Embodiment 30. The PHIP moving portion includes *CH2, *CH2-*CH2, *CHY, *C=Y, or any deuterated version thereof, where *C is 12 C or 13 The method according to any one of embodiments 22 to 26, wherein Y is a carbon isotope of C, and Y is selected from a spin 1 / 2 atom and a spin 1 / 2 atom covalently bonded to one or more chemical parts selected from heteroatoms such as N, O, and S, which are optionally substituted with a linear, branched, or cyclic C1-C10 alkyl hydrocarbon, C6 aryl, benzyl, phenyl, heteroaryl, halogen, or haloalkyl group.

[0183] Embodiment 31. Spin 1 / 2 atoms, 1 H, 13 C, 15 N, 19 F, or 31 The method according to embodiment 28 or 30, selected from P.

[0184] Embodiment 32. The method according to any one of Embodiments 22 to 31, wherein the PHIP moving portion includes at least one atom that is J-coupled with a non-hydrogen nucleus spin of at least 0.1 Hz.

[0185] Embodiment 33. The method according to any one of Embodiments 22 to 32, wherein Z or Z' comprises at least one atom that is J-coupled with a non-hydrogen nuclear spin of at least 0.1 Hz.

[0186] Embodiment 34.R2 is the method according to any one of Embodiments 22 to 33, wherein the solubilized portion is included.

[0187] Embodiment 35. The method according to any one of Embodiments 22 to 34, wherein R2 includes a hydrophobic and / or organic affinity moiety.

[0188] Embodiment 36.R2 is the method according to Embodiment 35, wherein the organic solubilization portion is included.

[0189] Embodiment 37. The method according to any one of Embodiments 22 to 33, wherein R2 includes a hydrophilic and / or organic affinity moiety.

[0190] Embodiment 38. The method according to any one of Embodiments 22 to 37, wherein R2 comprises or is selected from a methyl group, an ethyl group, a propyl group, an isopropyl group, an n-butyl group, an s-butyl group, a t-butyl group, an isobutyl group, a hydroxyl group, a methyl alcohol group, an ethyl alcohol group, an n-propanol group, an isopropyl alcohol group, an alcohol propionic acid group, an n-butyl alcohol group, an s-butyl alcohol group, a t-butyl alcohol group, an isobutyl alcohol group, a methoxy group, an ethoxy group, an ethoxy group, an isopropoxy group, an alcohol propionic acid group, an ethoxy group, a t-butoxy group, an s-butoxy group, an ester group, a phenyl group, a substituted phenyl group, a primary amine group, a secondary amine group, a tertiary amine group, a primary amide group, a secondary amide group, and a tertiary amide group.

[0191] Embodiment 39. A bio-related contrast agent is, Formula R 10 It contains a compound C(=O)X-, in which R 10 However, selected from linear, branched, or cyclic C1-C10 alkyl groups, one or more C atoms are optionally substituted with C=C, CO, COH, CNH2, COOH, CH2COOH, CONH2, or OC(=O), and X is NR 11 Selected from , S, and O, where R 11 but, 1 H, 2 H, 3The method according to any one of Embodiments 22 to 38, wherein H and an amino protecting group optionally selected from trifluoroacetyl, acetyl, benzoyl, carbobenzoxy, tert-butyl carbonate, and benzyl.

[0192] Embodiment 40. The method according to any one of Embodiments 22 to 39, wherein the bio-related contrast agent is selected from pyruvate, glutamate, glutamine, lactic acid, acetic acid, acetoacetic acid, zymonate, alanine, fructose, fumaric acid, bicarbonate, urea, dehydroascorbic acid, α-ketoglutaric acid, dihydroxyacetone, glucose, ascorbic acid, and their conjugate acids.

[0193] Embodiment 41. A hyperpolarized bio-associated contrast agent or a pharmaceutically acceptable salt thereof, produced by the method described in any one of Embodiments 22 to 40. [Examples]

[0194] Example 1 - NMR Procedure The NMR spectrum, 1 H is 400.13MHz, and 13 The data was recorded using a Bruker Avance Neo 400MHz spectrometer operating at 100.61MHz in C. Dichloromethane (CHCl3) 1 It is used as a solvent for 1H NMR experiments, while deuterated dichloromethane (CDCl3) is used. 13 It was used as a solvent for 13C NMR experiments. The NMR chemical shift (δ) is reported in ppm. 1 H and 13 For the C spectrum, the solvent signal is internally calibrated. 1 ¹H NMR: δ(CHCl3)7.26; 13 [C NMR: δ(CDCl3)77.16] was used. 1313C NMR spectra were recorded in proton separation mode. Flash chromatography was performed using a Biotage® Sfar HC Duo column as the solid phase and a Biotage® Selekt Flash Purification System with a mixture of cyclohexane and ethyl acetate as the mobile phase. Column chromatography was performed on silica gel (Silica 60, 63-200 μm, Macherey-Nagel) with a mixture of cyclohexane and ethyl acetate as the mobile phase.

[0195] Example 2 - General Synthesis Procedure Synthesis of esters in the a,b-unsaturated sidearm [ka] Combination of side arms 1a-g Alcohols 1a, 1c, 1d, and 1g were prepared using the published procedures (1a: T. Yoshinao, K. Masanari, T. Shuji, K. Sigeru, Y. Zenichi, Bull. Chem. Soc. Jpn. 1994, 67, 2838-2849; 1c: BRBlank, I. Andrews, O. Kwon, ChemCatChem 2020, 12, 4352-4372; 1d: GCTsui, K. Villeneuve, E. Carlson, W. Tam, Organometallics 1g was prepared according to the procedure used to synthesize 1a, starting with tetrahydro-2-(2-propynyloxy)-2H-pyran and isopropyl chloroformate, followed by deprotection of the THP group with pyridinium p-toluenesulfonate. The synthesis of 1c-d9 with selective deuteration of the tert-butyl group was performed using di-tert-butyl bicarbonate instead of di-tert-butyl bicarbonate-d18 The synthesis of 1c was carried out according to the procedure using [the specified method]. Selective monodeuteration at 1c-d1 was performed by the following reaction scheme: [ka]

[0196] Alcohol 1e was prepared according to a published one-step synthesis from tert-butyl propiophosphate and benzaldehyde (A. Kondoh, R. Ozawa, M. Terada, Chem. Lett. 2019, 48, 1164-1167, which is incorporated herein by reference in connection with a method for synthesizing alcohol 1e). Alcohol 1f was prepared by esterification with benzylol under Mitsunobu conditions, starting from tetrahydro-2-(2-propynyloxy)-2H-pyran via carboxylation using CO2, followed by a known procedure in the literature (O. Mitsunobu, Y. Yamada, Bulletin Chem. Soc. Japan 1967, 40(1), 2380-2382, which is incorporated herein by reference in connection with a method for synthesizing alcohol 1f) and THP cleavage with 1c. Alcohol 1g-d5 was prepared according to 1g using benzoyl chloride-d5 instead of benzoyl chloride.

[0197] Synthesis of esters 2-4 in a,b-unsaturated side arms General procedure A: The target carboxylic acid (1.0-2.0 equivalents) was dissolved in anhydrous tetrahydrofuran (THF, 200-350 mmol), and 1 a-g of the corresponding alcohol was added. The solution was cooled to 0°C in an ice / water bath. At this temperature, pyridine (4.0-5.0 equivalents) was added dropwise over 5 minutes, followed by methanesulfonyl chloride (1.2-2.0 equivalents) (or p-toluenesulfonyl chloride dissolved in THF) dropwise over 10 minutes. The mixture was stirred until all of the corresponding alcohols (1 a-g) were consumed (if methanesulfonyl chloride and pyruvate are used in high excess, 2-5 hours at 0°C; if only slightly excess methanesulfonyl chloride or p-toluenesulfonyl chloride is used, up to 18 hours at ambient temperature). 1 (Monitored via 1H NMR). A colorless precipitate formed in all cases during the reaction. After the reaction was complete, the reaction mixture was poured into a stirred mixture of 0.5N HCl aqueous solution (same volume as the reaction solvent) and diethyl ether (same volume as the reaction solvent). The phases were separated, the organic phase was collected, and the aqueous phase was washed with diethyl ether (2 × 200 mL). The combined organic phase was washed with saturated NaHCO3 aqueous solution (100 mL), followed by washing with brine (100 mL). The organic phase was dried over Na2SO4, and volatile components were removed under reduced pressure (800 → 1 mbar, 40°C water bath). The crude product was purified by flash chromatography or column chromatography.

[0198] General procedure B: 1 a-g (1.0 equivalent) of the corresponding alcohol was dissolved in anhydrous THF (200 mmol) and cooled to -78°C in an acetone / dry ice bath. Anhydrous triethylamine (NEt3, 1.0 equivalent) was added all at once to the clear mixture, followed by the dropwise addition of the target carboxylic acid chloride (1.0 equivalent) over 3 minutes. After the complete addition of the acid chloride, the cooling bath was removed and the mixture was allowed to warm to ambient temperature. The suspension was quenched by pouring it into a stirred mixture of 0.5 N HCl aqueous solution (same volume as the reaction solvent) and diethyl ether (same volume as the reaction solvent). The phases were separated, the organic phase was collected, and the aqueous phase was washed with diethyl ether (2 × 200 mL). The combined organic phase was washed with saturated NaHCO3 aqueous solution (100 mL), followed by washing with brine (100 mL). The organic phase was dried over Na2SO4, and volatile components were removed under reduced pressure (800 → 1 mbar, 40°C water bath). The crude product was purified by flash chromatography as needed.

[0199] Example 3 - Methyl 4-((2-oxopropanoyl)oxy)buta-2-inoate (2a) [ka] Structure 2a was prepared from 300 mL of THF containing alcohol 1a (7.08 g, 62.1 mmol), pyruvate (10.9 g, 124 mmol, 2.0 equivalents), pyridine (25 mL, 310 mmol, 5.0 equivalents), and methanesulfonyl chloride (9.6 mL, 124 mmol, 2.0 equivalents) at 0°C for 4 hours (h) according to general procedure A. Purification was carried out by column chromatography yielding 2a as a colorless, slightly viscous liquid (7.1 g, 38 mmol, 61% yield).

[0200] Figure 3A is an example corresponding to structure 2a. 1 The 1H NMR (400 MHz, CDCl3) spectrum is shown. The spectrum showed peaks at d=2.52 (single (s), combined signal: corresponding to 3H, CH3), 3.80 (s, 3H, OCH3), and 4.96 (s, 2H, OCH2).

[0201] Figure 3B is an example corresponding to structure 2a. 13 The 13C NMR (101 MHz, CDCl3) spectrum is shown. The spectrum showed peaks at d=26.78 (CH3), 52.84 (OCH3), 53.05 (OCH2), 78.61 (C≡C), 79.52 (C≡C), 153.00 (COOCH3), 159.36 (COO), and 190.23 (C=O).

[0202] Example 4-Isopropyl 4-((2-oxopropanoyl)oxy)buta-2-inoate (2b) [ka] Structure 2b was prepared from alcohol 1b (6.36 g, 40.7 mmol), pyruvate (7.16 g, 81.4 mmol, 2.0 equivalents), pyridine (16.4 mL, 203 mmol, 5.0 equivalents), and p-toluenesulfonyl chloride (15.5 g, 81.4 mmol, 2.0 equivalents) in THF (200 mL) according to general procedure A, at 0°C for 2 hours and at ambient temperature for 15 hours. Purification was performed via flash chromatography to obtain 2b as a yellowish, slightly viscous liquid (5.7 g, 25 mmol, 62% yield).

[0203] Figure 4A is an exemplary model corresponding to structure 2b. 1 The 1H NMR (400 MHz, CDCl3) spectrum is shown. In the NMR spectrum, d = 1.28 (double line (d)), 3 J H,H =6.29Hz, 6H, 2×CH3), 2.50(s, 3H, CH3), 4.94(s, 2H, OCH2), and 5.09(hept, 3 J H,H The peak was observed at 6.26Hz, 1H, OCH.

[0204] Figure 4B is an exemplary representation of structure 2b. 13The ¹³C NMR (101 MHz, CDCl3) spectrum is shown. The NMR spectrum is d=21.61(2×CH3), 26.79(CH3), 52.93(OCH2), 70.64(OCH), 78.62(C≡C), 79.33(C≡C), 152.15(COO t Peaks were observed at Bu, 159.43 (COO), and 190.25 (C=O).

[0205] Example 5: Tert-butyl 4-((2-oxopropanoyl)oxy)buta-2-inoate(2c) [ka] Structure 2c was prepared from THF (500 mL) containing alcohol 1c (30.0 g, 192 mmol), pyruvate (33.8 g, 384 mmol, 2.0 equivalents), pyridine (77.0 mL, 954 mmol, 5.0 equivalents), and methanesulfonyl chloride (30.0 mL, 388 mmol, 2.0 equivalents) at 0°C for 3 hours according to general procedure A. Purification was performed via flash chromatography to produce 2c as a yellowish, slightly viscous liquid (33 g, 146 mmol, 76% yield).

[0206] Figure 5A is an illustrative example corresponding to structure 2c. 1 The 1H NMR (400MHz, CDCl3) spectrum is shown. The NMR spectrum is d=1.49(s, 9H, t Peaks were observed at Bu, 2.50 (s, 3H, CH3), and 4.93 (s, 2H, OCH2).

[0207] Figure 5B is an example corresponding to structure 2c. 13 The 13C NMR (101 MHz, CDCl3) spectrum is shown. The NMR spectrum is d=26.81(CH3), 27.97(3×CH3), 53.02(OCH2), 76.78(C≡C), 80.21(C≡C), 84.36(OC- t Bu), 151.59 (COO t Peaks were observed at Bu), 159.47 (COO), and 190.32 (C=O).

[0208] Example 6 -Tert-butyl 4-((2-oxopropanoyl-1- 13 C) Oxy) Buta-2-inoate (2c- 13 C) [ka] Structure 2c- 13 C( 13 1C-labeled 2c) is mixed with alcohol 1c (4.40g, 28.1mmol), pyruvate-1- 13 A solution was prepared from 80 mL of THF containing 14C (5.00 g, 56.1 mmol, 2.0 equivalents), pyridine (11.3 mL, 140 mmol, 5.0 equivalents), and methanesulfonyl chloride (4.34 mL, 56.1 mmol, 2.0 equivalents) at 0°C for 4 hours according to general procedure A. Purification was performed by flash chromatography to obtain 2c- as a colorless, slightly viscous liquid (4.50 g, 19.8 mmol, 70% yield). 13 C was produced.

[0209] Figure 6A shows structure 2c- 13 Example corresponding to C 1 The 1H NMR (400MHz, CDCl3) spectrum is shown. The NMR spectrum is d=1.49(s, 9H, t Bu), 2.50(d, 3 J C、H =1.58Hz, CH3), and 4.93(d, 3 J C、H A peak was observed at 3.54Hz, 2H, and OCH2.

[0210] Figure 6B shows structure 2c- 13 Example corresponding to C 13 The ¹³C NMR (10¹ MHz, CDCl3) spectrum is shown. The NMR spectrum is d = 26.91 (d, 2 J C,C =17.08Hz, CH3), 28.07(3×CH3), 53.12(d, 2 J C、C =2.58Hz, OCH2), 76.76(d, 3 J C,C=2.51Hz, C≡C), 80.35(C≡C), 84.49(OC- t Bu), 151.69 (COO t Bu), 159.56 (COO), and 190.39 (d, 1 J C、C The peak was observed at =67.07 (C=0).

[0211] Example 7-2-(methyl-d3)propan-2-yl-1,1,1,3,3,3-d64-((2-oxopropanoyl-1- 13 C)Oxy)buta-2-inoate(2c-d9- 13 C) [ka] Structure 2c-d 9- 13 C(9 × deuterated, 13 The 1C-labeled structure 2c) was treated with alcohol 1c-d9 (5.50g, 33.2 mmol), pyruvate-1- 13 A solution was prepared from 130 mL of THF containing 140C (5.00 g, 56.1 mmol, 1.7 equivalents), pyridine (11.3 mL, 140 mmol, 4.2 equivalents), and methanesulfonyl chloride (4.3 mL, 56.1 mmol, 1.7 equivalents) at 0°C for 5 hours according to general procedure A. Purification was performed by flash chromatography to obtain a colorless, slightly viscous liquid (5.08 g, 21.6 mmol, 65% yield) in 2c-d. 9- 13 C was produced.

[0212] Figure 7A shows structure 2c-d 9- 13 Example corresponding to C 1 The H NMR (400 MHz, CDCl3) spectrum is shown. The NMR spectrum is d=2.48(d, 3 J C,H =1.56Hz, CH3), and 4.91(d, 3 J C,H A peak was observed at 3.55Hz, 2H, and OCH2.

[0213] Figure 7B shows 2c-d9- 13 Example corresponding to C 13 The ¹³C NMR (10¹ MHz, CDCl3) spectrum is shown. The NMR spectrum is d=26.62~27.41 (multiline (m), 3×CD3), 26.84 (d, 2 J C,C =17.06Hz, CH3), 53.06(d, 2 J C,C =2.68Hz, OCH2), 76.76(d, 3 J C,C =2.45Hz, C≡C), 80.27(C≡C), 83.99(OC- t Bu), 151.64 (COO t Bu), 159.50 (COO), and 190.35 (d, 1 J C,C The peak was observed at =67.13 (C=0).

[0214] Example 8: Tert-butyl 4-((2-oxopropanoyl)oxy)buta-2-inoate-4-d(2c-d1) [ka] Structure 2c-d1 (1×deuterated structure 2c) was prepared from THF (400 mL) of alcohol 1c-d1 (14.0 g, 89.0 mmol), pyruvate (15.8 g, 178 mmol, 2.0 equivalents), pyridine (36.0 mL, 445 mmol, 5.0 equivalents), and methanesulfonyl chloride (13.8 mL, 178 mmol, 2.0 equivalents) at 0°C for 1 hour and then at ambient temperature for 3 hours, according to general procedure A. Purification was performed by flash chromatography to obtain 2c-d1 as a yellowish, slightly viscous liquid (13.2 g, 58.1 mmol, 65% yield).

[0215] Figure 8A is an exemplary representation corresponding to structure 2c-d1. 1 The 1H NMR (400MHz, CDCl3) spectrum is shown. The NMR spectrum is d=1.49(s, 9H, t Bu), 2.51 (s, 3H, CH3), and 4.92 (triple line (t), 2 JH,D A peak was observed at 2.34Hz, 2H, and OCH2.

[0216] Figure 8B is an example corresponding to structure 2c-d1. 13 The ¹³C NMR (10¹ MHz, CDCl3) spectrum is shown. The NMR spectrum is d = 26.66 (CH3), 27.84 (3×CH3), 52.66 (t, 1 J C,D =23.56Hz, OCHD), 76.78(C≡C), 80.00(C≡C), 84.19(OC- t Bu), 151.46 (COO t Peaks were observed at Bu, 159.36 (COO), and 190.24 (C=O).

[0217] Example 9: Tert-butyl 4-((2-oxopropanoyl)oxy)penta-2-inoate(2d) [ka] Structure 2d was prepared from THF (80 mL) containing alcohol 1d (4.83 g, 28.4 mmol), pyruvate (5.00 g, 56.8 mmol, 2.0 equivalents), pyridine (11.5 mL, 142 mmol, 5.0 equivalents), and methanesulfonyl chloride (4.40 mL, 56.8 mmol, 2.0 equivalents) at 0°C for 3 hours and then at ambient temperature for 1 hour, according to general procedure A. Purification was performed via flash chromatography to produce 2d as a yellowish, slightly viscous liquid (4.85 g, 20.2 mmol, 71% yield).

[0218] Figure 9A is an exemplary model corresponding to structure 2d. 1 The H NMR (400 MHz, CDCl3) spectrum is shown. The NMR spectrum is d = 1.48 (s, 9H, t Bu), 1.64(d, 3 J H,H =6.80Hz, OCHCH3), 2.49(s, 3H, CH3), and 5.58(quadruple (q), 3 J H、H A peak was observed at 6.79Hz, 1H, and OCHCH3.

[0219] Figure 9B is an example corresponding to structure 2d. 13 The ¹³C NMR (101 MHz, CDCl3) spectrum is shown. The NMR spectrum is d=20.35(OCHCH3), 26.77(CH3), 27.97(3×CH3), 61.87(OCHCH3), 78.58(C≡C), 80.62(C≡C), 84.21(OC- t Bu), 151.82 (COO t Peaks were observed at Bu, 159.26 (COO), and 190.75 (C=O).

[0220] Example 9: Tert-butyl 4-((2-oxopropanoyl)oxy)-4-phenylbuta-2-inoate(2e) [ka] Structure 2e was prepared from 100 mL of THF containing alcohol 1e (5.81 g, 25.0 mmol), pyruvate (4.40 g, 50.0 mmol, 2.0 equivalents), pyridine (10.7 mL, 125 mmol, 5.0 equivalents), and methanesulfonyl chloride (3.87 mL, 50.0 mmol, 2.0 equivalents), following general procedure A, at 0°C for 1.5 hours and at ambient temperature for 2.5 hours. Purification was performed via flash chromatography to produce 2e as a yellowish, viscous liquid (5.00 g, 16.5 mmol, 66% yield).

[0221] Figure 10A is an example corresponding to structure 2e. 1 The 1H NMR (400 MHz, CDCl3) spectrum is shown. The NMR spectrum is d = 1.50 (s, 9H, t Bu), 2.48(s, 3H, CH3), 6.57(s, 1H, OCHPh), 7.40-7.43(m, 3H, H Ph ), and 7.53-7.56 (m, 2H, H Ph It showed a peak at ).

[0222] Figure 10B is an example corresponding to structure 2e. 13The 13C NMR (101 MHz, CDCl3) spectrum is shown. The NMR spectrum is d=26.98(CH3), 28.09(3×CH3), 67.23(OCHPh), 78.88(C≡C), 80.69(C≡C), 84.51(OC- t Bu), 128.26(C Ph ), 129.18(C Ph ), 130.09(C Ph ), 134.23(C Ph ), 151.86 (COO t Peaks were observed at Bu, 159.16 (COO), and 190.56 (C=O). Example 10 - Benzylhydryl 4-((2-oxopropanoyl)oxy)buta-2-inoate(2f) [ka]

[0223] Structure 2f was prepared from 1f (1.60 g, 6.00 mmol) according to general procedure B, yielding 2f as a colorless, viscous oil (1.92 g, 5.70 mmol, 95% yield).

[0224] Figure 11 is an example corresponding to structure 2f. 1 The 1H NMR (400 MHz, CDCl3) spectrum is shown. The NMR spectrum is d=2.51(s, 3H, CH3), 4.97(s, 2H, OCH2), 6.94(s, 1H, OCH), and 7.28~7.38(m, 10H, H Ph It showed a peak at ).

[0225] Example 11-4-oxo-4-phenylbuta-2-in-1-yl 2-oxopropanoate (2g) [ka] 2 g of the structure was purified by flash chromatography, and a preparation was made from 1 g (2.49 g, 15.5 mmol) according to general procedure B, yielding 2 g-d5 as a slightly orange solid (1.86 g, 8.06 mmol, yield 52%).

[0226] Figure 12A is an example corresponding to structure 2g. 1 The 1H NMR (400 MHz, CDCl3) spectrum is shown. The NMR spectrum is d=2.54(s, 3H, CH3), 5.11(s, 2H, OCH2), 7.48-7.52(m, 2H, H Ph ), 7.61-7.66(m, 1H, H Ph ), and 8.09-8.11(m, 2H, H Ph It showed a peak at ).

[0227] Figure 12B is an example corresponding to structure 2g. 13 The 13C NMR (101 MHz, CDCl3) spectrum is shown. The NMR spectrum is d=26.81(CH3), 53.24(OCH2), 84.65(C≡C), 85.56(C≡C), 128.78(C Ph ), 129.65(C Ph ), 134.65(C Ph ), 136.03(C Ph Peaks were observed at 159.59 (COO), 176.91 (C(O)Ph), and 190.34 (C=O).

[0228] Example 12-4-oxo-4-(phenyl-d5)buta-2-in-1-yl 2-oxopropanoate (2g-d5) [ka] Structure 2g-d5 was prepared from 1g-d5 (4.66g, 28.2 mmol) according to general procedure B, and 2g-d5 was obtained as a brownish solid (6.1g, 25.9 mmol, 92%).

[0229] Figure 13A is an example corresponding to structure 2g-d5. 1 The 1H NMR (400 MHz, CDCl3) spectrum is shown. The NMR spectrum showed peaks at d=2.49 (s, 3H, CH3) and 5.09 (s, 2H, OCH2).

[0230] Figure 13B is an example corresponding to structure 2g-d5. 13The ¹³C NMR (101 MHz, CDCl3) spectrum is shown. The NMR spectrum is d=26.79(CH3), 53.23(OCH2), 84.62(C≡C), 85.55(C≡C), 128.24(t, 1 J C,D =24.58Hz, C Ph ), 129.23(t, 1 J C,D =24.58Hz, C Ph ), 134.14(t, 1 J C,D =22.45Hz, C Ph ), 135.83(C Ph Peaks were observed at 159.54 (COO), 176.91 (C(O)Ph), and 190.35 (C=O).

[0231] Example 13-1-(4-(Tert-butoxy)-4-oxobuta-2-in-1-yl)5-ethyl 2-oxopentanedioate (3c) [ka] Structure 3c was purified from THF (40 mL) containing alcohol 1c (0.92 g, 5.9 mmol), mono-ethyl α-ketoglutaric acid (1.02 g, 5.85 mmol, 1.0 equivalent), pyridine (1.88 mL, 23.3 mmol, 4.0 equivalents), and methanesulfonyl chloride (543 μL, 7.02 mmol, 1.2 equivalents) at 0°C for 5 hours and 7°C for 40 hours, according to general procedure A. Purification was performed via flash chromatography, and since esterification was not quantitative and 3c and 1c had similar retention times, 3c was produced as a colorless viscous liquid containing 20% ​​of 1c (1.40 g, 4.48 mmol, yield 77%).

[0232] Figure 14A is an exemplary representation of structure 3c. 1 The 1H NMR (400 MHz, CDCl3) spectrum is shown. The NMR spectrum is d=1.25(t, 3 J H,H =7.14Hz,3H, OCH2CH3), 1.49(s, 9H, t Bu), 2.68(t,3 J H,H =6.49Hz, 2H, CH2CH2), 3.17(t, 3 J H,H =6.36Hz, 2H, CH2CH2), 4.14(q, 3 J H,H Peaks were observed at 7.14Hz (2H, OCH2CH3) and 4.94(s, 2H, OCH2).

[0233] Figure 14B is an example corresponding to structure 3c. 13 The ¹³C NMR (101 MHz, CDCl3) spectrum is shown. The NMR spectrum is d=14.26(OCH2CH3), 27.87(CH2CH2), 28.06(3×CH3), 28.10(CH2CH2), 53.16(OCH2), 61.15(OCH2CH3), 76.74(C≡C), 80.37(C≡C), 84.46(OC- t Bu), 151.69 (COO t Peaks were observed at Bu), 159.33 (COO), 171.95 (COOEt), and 191.27 (C=O).

[0234] Example 14-Methyl 4-(2,2-dichloroacetoxy)buta-2-inoate (4a) [ka] Structure 4a was prepared from 1a (4.00 g, 35.0 mmol) according to general procedure B, and 4a was produced as a colorless liquid (7.72 g, 34.3 mmol, 98% yield).

[0235] Figure 15A is an example corresponding to structure 4a. 1 The 1H NMR (400 MHz, CDCl3) spectrum is shown. The NMR spectrum showed peaks at d=3.80 (s, 3H, OCH3), 4.97 (s, 2H, OCH2), and 5.99 (s, 1H, CHCl2).

[0236] Figure 15B is an example corresponding to structure 4a. 13The 13C NMR (101 MHz, CDCl3) spectrum is shown. The NMR spectrum showed peaks at d=53.19 (OCH2), 54.07 (OCH3), 63.60 (CHCl2), 79.05 (C≡C), 79.11 (C≡C), 153.08 (COOCH3), and 163.75 (COO).

[0237] Example 15-Tert-butyl 4-((acetoxy)buta-2-inoate(5c) [ka] Structure 5c was prepared from 1c (5.50 g, 35.2 mmol) according to general procedure B, and purified by flash chromatography to produce 5c as a colorless liquid (5.72 g, 28.8 mmol, 82% yield).

[0238] Figure 16A shows an exemplary 1H NMR (400 MHz, CDCl3) spectrum corresponding to structure 5c. The NMR spectrum is δ = 1.49 (s, 9H, t Peaks were observed at Bu), 2.10(s, 3H, CH3), and 4.76(s, 2H, OCH2).

[0239] Figure 16B shows an exemplary 13C NMR (101 MHz, CDCl3) spectrum corresponding to structure 5c. The NMR spectrum is δ = 20.68 (CH3), 28.08 (3×CH3), 51.50 (OCH2), 78.59 (C≡C), 79.32 (C≡C), 84.16 (OC- t Bu), 151.96 (COO t The peaks were observed at Bu) and 170.03 (COO).

[0240] Example 16 - Improved Hydrogenation Efficiency Figure 17A shows an exemplary 1H NMR (400 MHz, CDCl3) spectrum corresponding to para-hydrogenated 3-phenylpropa-2-in-1-yl-2-oxopropanoate (i.e., 3-phenylallyl-2-oxopropanoate with two H*). The para-hydrogenation reaction between 3-phenylpropa-2-in-1-yl-2-oxopropanoate and para-hydrogen was carried out at 60°C in the presence of 1 mol% [Rh(dppb)(COD)]BF4 for 10 seconds at a para-hydrogen pressure of 10 bar. The reaction formed the previously known 3-phenylallyl-2-oxopropanoate with two H*, where the * symbol indicates a molecule containing a proton derived from para-hydrogen. In the examples shown, the NMR signals marked with the * symbol correspond to the para-hydrogen molecule added across the carbon-carbon triple bond of 3-phenylpropa-2-in-1-yl-2-oxopropanoate. 1 Based on the 1H NMR spectrum, the yield of the parahydrogenation reaction was determined to be approximately 29%.

[0241] Figure 17B is an exemplary representation of para-hydrogenated tert-butyl 4-((2-oxopropanoyl)oxy)buta-2-inoate (i.e., tert-butyl 4-((2-oxopropanoyl)oxy)buta-2-enoate having two H*). 1 The 1H NMR (400 MHz, CDCl3) spectrum is shown. The parahydrogenation reaction between this novel molecule and parahydrogen was carried out at 60°C in the presence of 1 mol% [Rh(dppb)(COD)]BF4 for 10 seconds at a parahydrogen pressure of 10 bar. The reaction formed tert-butyl 4-((2-oxopropanoyl)oxy)buta-2-enoate with two H*, where * indicates a molecule containing a proton derived from parahydrogen. In the shown examples, the NMR signals marked with * correspond to the parahydrogen molecules added across the carbon-carbon triple bond of tert-butyl 4-((2-oxopropanoyl)oxy)buta-2-inoate. 1 Based on the 1H NMR spectrum, the yield of the parahydrogenation reaction was determined to be approximately 85%.

[0242] As shown in Figures 17A and 17B, the parahydrogenation reaction between tert-butyl 4-((2-oxopropanoyl)oxy)buta-2-inoate and parahydrogen yielded a significantly higher yield than the parahydrogenation reaction between 3-phenylpropa-2-in-1-yl 2-oxopropanoate and parahydrogen.

[0243] Figure 18A shows an exemplary 1H NMR (400 MHz, CDCl3) spectrum corresponding to para-hydrogenated 3-phenylpropa-2-in-1-yl-2-oxopropanoate (i.e., 3-phenylallyl-2-oxopropanoate with two H*). The para-hydrogenation reaction between 3-phenylpropa-2-in-1-yl-2-oxopropanoate and para-hydrogen was carried out at 45°C in the presence of 1 mol% [Rh(dppb)(COD)]BF4 for 5 seconds at a para-hydrogen pressure of 10 bar. The reaction formed the previously known 3-phenylallyl-2-oxopropanoate with two H*, where the * symbol indicates a molecule containing a proton derived from para-hydrogen. In the examples shown, the NMR signals marked with the * symbol correspond to para-hydrogen molecules added across the carbon-carbon triple bond of 3-phenylpropargylpyruvic acid. 1 Based on the 1H NMR spectrum, the yield of the parahydrogenation reaction was determined to be approximately 21%.

[0244] Figure 18B shows an exemplary corresponding to para-hydrogenated 4-oxo-4-phenylbuta-2-in-1-yl 2-oxopropanoate (i.e., 4-oxo-4-phenylbuta-2-en-1-yl 2-oxopropanoate having two H*). 1The 1H NMR (400 MHz, CDCl3) spectrum is shown. The parahydrogenation reaction between 4-oxo-4-phenylbuta-2-in-1-yl-2-oxopropanoate and parahydrogen was carried out at 45°C in the presence of 1 mol% [Rh(dppb)(COD)]BF4 for 5 seconds at a parahydrogen pressure of 10 bar. The reaction formed 4-oxo-4-phenylbuta-2-in-1-yl-2-oxopropanoate with two H*, where * indicates a molecule containing a proton derived from parahydrogen. In the shown examples, the NMR signals marked with * correspond to parahydrogen molecules added across the carbon-carbon triple bond of 4-oxo-4-phenylbuta-2-in-1-yl-2-oxopropanoate. 1 Based on the 1H NMR spectrum, the yield of the parahydrogenation reaction was determined to be approximately 44%.

[0245] As shown in Figures 18A and 18B, the parahydrogenation reaction between 4-oxo-4-phenylbuta-2-in-1-yl 2-oxopropanoate and parahydrogen yielded significantly higher yields than the parahydrogenation reactions between known pyruvate derivatives and between phenylallyl 2-oxopropanoate and parahydrogen.

[0246] Example 17 - Improved 13 C polarization Figure 19A is an exemplary representation of 200 mM parahydrogenated 3-phenylpropane-2-in-1-yl 2-oxopropane (i.e., 3-phenylallyl 2-oxopropane with two H*) in acetone-d6. 13 The 13C NMR (101 MHz, CDCl3) spectrum is shown. Para-hydrogenated 3-phenylpropa-2-in-1-yl 2-oxopropanoate was formed using the procedure described herein with respect to Figure 17A. In the shown examples, the para-hydrogenated 3-phenylpropa-2-in-1-yl 2-oxopropanoate was approximately 9.8% 13 It was determined that it has C polarization.

[0247] Figure 19B shows an exemplary representation of 200 mM para-hydrogenated tert-butyl 4-((2-oxopropanoyl)oxy)buta-2-inoate (i.e., tert-butyl 4-((2-oxopropanoyl)oxy)buta-2-enoate having two H* atoms) in acetone-d6. 13 The 13C NMR (101 MHz, CDCl3) spectrum is shown. Para-hydrogenated tert-butyl 4-((2-oxopropanoyl)oxy)buta-2-inoate was formed using the procedure described herein with respect to Figure 17B. In the shown examples, the para-hydrogenated tert-butyl 4-((2-oxopropanoyl)oxy)buta-2-inoate was approximately 17.5% 13 It was determined that it has C polarization.

[0248] As shown in Figures 19A and 19B, para-hydrogenated tert-butyl 4-((2-oxopropanoyl)oxy)buta-2-inoate yielded significantly higher polarization than para-hydrogenated 3-phenylpropa-2-in-1-yl 2-oxopropanoate. It should be noted that para-hydrogenated 3-phenylpropargylpyruvic acid represents one of the current gold standard precursors for producing hyperpolarized pyruvate via PHIP-SAH. Therefore, the compositions described herein may enable the production of pyruvate and other bio-relevant contrast agents with significantly enhanced polarization compared to known precursors.

[0249] Figure 20A is an exemplary representation of 200 mM parahydrogenated 3-phenylpropane-2-in-1-yl 2-oxopropanoate (i.e., 3-phenylallyl 2-oxopropanoate with two H*) in acetone-d6. 13 The 13C NMR (101 MHz, CDCl3) spectrum is shown. Para-hydrogenated 3-phenylpropa-2-in-1-yl 2-oxopropanoate was formed using the procedure described herein with respect to Figure 18A. In the shown examples, the para-hydrogenated 3-phenylpropa-2-in-1-yl 2-oxopropanoate was approximately 13.9% 13 It was determined that it has C polarization.

[0250] Figure 20B shows an exemplary solution corresponding to 200 mM parahydrogenated 4-oxo-4-phenylbuta-2-in-1-yl 2-oxopropanoate (i.e., 4-oxo-4-phenylbuta-2-en-1-yl 2-oxopropanoate having 2 H*). 13 The 13C NMR (101 MHz, CDCl3) spectrum is shown. Para-hydrogenated 4-oxo-4-phenylbuta-2-in-1-yl 2-oxopropanoate was formed using the procedure described herein with respect to Figure 18B. In the shown examples, the para-hydrogenated 4-oxo-4-phenylbuta-2-in-1-yl 2-oxopropanoate was approximately 10.3% 13 It was determined that it has C polarization.

[0251] Figure 21 shows an exemplary corresponding to 133 mM para-hydrogenated methyl 4-((2-oxopropanoyl)oxy)buta-2-inoate (i.e., methyl 4-((2-oxopropanoyl)oxy)buta-2-enoate having two H* atoms). 13 The 13C NMR (101 MHz, CDCl3) spectrum is shown. The parahydrogenation reaction between this novel molecule and parahydrogen was carried out at 60°C in the presence of 1 mol% [Rh(dppb)(COD)]BF4 for 10 seconds at a parahydrogen pressure of 10 bar. The reaction formed methyl 4-((2-oxopropanoyl)oxy)buta-2-enoate with two H*, where * indicates a molecule containing a proton derived from parahydrogen. In the shown example, the parahydrogenated methyl 4-((2-oxopropanoyl)oxy)buta-2-inoate was approximately 17.2% 13 It was determined that it has C polarization.

[0252] Example 18 - Hyperpolarized α-ketoglutarate Figure 22A is an exemplary corresponding to para-hydrogenated 1-(4-(tert-butoxy)-4-oxobuta-2-in-1-yl)5-ethyl 2-oxopentanedioate (i.e., 1-(4-(tert-butoxy)-4-oxobuta-2-in-1-yl)5-ethyl 2-oxopentanedioate having 2 H*). 1 The 1H NMR (400 MHz, CDCl3) spectrum is shown. The parahydrogenation reaction between 1-(4-(tert-butoxy)-4-oxobuta-2-in-1-yl)5-ethyl 2-oxopentanedioate and parahydrogen was carried out at 60°C in the presence of 1 mol% [Rh(dppb)(COD)]BF4 for 10 seconds at a parahydrogen pressure of 10 bar. The reaction formed 1-(4-(tert-butoxy)-4-oxobuta-2-in-1-yl)5-ethyl 2-oxopentanedioate with two H*, where * indicates a molecule containing a proton derived from parahydrogen. In the shown examples, the NMR signals marked with * correspond to parahydrogen molecules added across the carbon-carbon triple bond of 1-(4-(tert-butoxy)-4-oxobuta-2-in-1-yl)5-ethyl 2-oxopentanedioate.

[0253] Figure 22B is an exemplary sample corresponding to 70 mM parahydrogenated 1-(4-(tert-butoxy)-4-oxobuta-2-in-1-yl)5-ethyl 2-oxopentanedioate (i.e., 1-(4-(tert-butoxy)-4-oxobuta-2-in-1-yl)5-ethyl 2-oxopentanedioate having 2 H*) in acetone-d6. 13 The 13C NMR (101 MHz, CDCl3) spectrum is shown. Para-hydrogenated 1-(4-(tert-butoxy)-4-oxobuta-2-in-1-yl)5-ethyl 2-oxopentanedioate was formed using the procedure described herein with respect to Figure 22A. In the shown examples, para-hydrogenated 1-(4-(tert-butoxy)-4-oxobuta-2-in-1-yl)5-ethyl 2-oxopentanedioate was approximately 12.8% 13 It was determined that it has C polarization.

[0254] The foregoing description is provided for illustrative purposes only. It is not exhaustive and is not limited to the exact forms or embodiments disclosed. Modifications and adaptations of embodiments will be apparent from the specification and practice of the disclosed embodiments. For example, the described implementations include hardware, but systems and methods consistent with this disclosure can be implemented using hardware and software. Furthermore, while certain components are described as being combined with one another, such components may be integrated with one another or distributed in any preferred manner.

[0255] Furthermore, while exemplary embodiments are described herein, the scope includes any and all embodiments having equivalent elements, modifications, omissions, combinations (e.g., aspects across various embodiments), adaptations, or alterations based on this disclosure. The elements of the claims should be interpreted broadly in accordance with the language used in the claims, and not limited to the examples described herein or in the application of the application, and such examples should be interpreted as non-exclusive. Furthermore, the steps of the methods of this disclosure can be modified in any way, including reordering steps, or inserting or deleting steps.

[0256] The features and advantages of this disclosure are evident from the detailed specification, and therefore the appended claims are intended to cover all systems and methods that fall within the true spirit and scope of this disclosure. Where used herein, the indefinite articles "a" and "an" mean "one or more." Similarly, the use of plural words does not necessarily mean plural unless ambiguous in a given context. Furthermore, since numerous modifications and variations readily arise from the study of this disclosure, it is undesirable to limit the exact structure and operation of this disclosure to examples and descriptions only. Therefore, all suitable modifications and equivalents may be utilized so as to fall within the scope of this disclosure.

[0257] As used herein, unless otherwise specifically stated, the term “or” encompasses all possible combinations, except in cases where it is not feasible. For example, if it is stated that a component may include A or B, then unless otherwise specifically stated or unless it is not feasible, the component may include A or B, or A and B. In a second embodiment, if it is stated that a component may include A, B, or C, then unless otherwise specifically stated, the component may include A or B or C, or A and B, or A and C, or B and C, or A and B and C.

[0258] Embodiments may be further described using the following clauses.

[0259] Other embodiments will be apparent from the description and implementation considerations of the embodiments disclosed herein. The description and examples are intended to be considered solely as examples having the true scope and spirit of the embodiments disclosed as set forth by the following claims. Examples of embodiments of the present invention are listed in the following sections [Aspect 1] to [Aspect 42]. [Aspect 1] A composition comprising a compound of formula (I), [ka] In the formula, Z is (i) 1 H (proton), 2 (ii) A carbon-carbon double bond (-C=C-) that is substituted to include H (deuterium) or a combination thereof, or (ii) a carbon-carbon triple bond (-C≡C-), R 1 However, it includes a para-hydrogen-induced polarization (PHIP) transfer portion, R 2 However, it contains optionally substituted hydrocarbons, alkoxy groups, primary amines, secondary amines, or tertiary amines. R 3 A composition comprising a bio-related contrast agent containing non-hydrogen nuclear spins. [Aspect 2] A composition comprising the compound of formula (II), [ka] In the formula, Z' is (i) 1 H (proton), 2 (ii) Parahydrogenated carbon-carbon single bonds (-CH*-CH*-) substituted to include H (deuterium) or a combination thereof, or1 H (proton), 2 A para-hydrogenated carbon-carbon double bond (-CH*=CH*-) which is substituted to include H (deuterium) or a combination thereof, In the formula, H* is hydrogen having a spin order derived from parahydrogen, R 1 However, it includes a para-hydrogen-induced polarization (PHIP) transfer portion, R 2 However, it contains optionally substituted hydrocarbons, alkoxy groups, primary amines, secondary amines, or tertiary amines. R 3 A composition comprising a bio-related contrast agent containing non-hydrogen nuclear spins. [Aspect 3] (i) A bio-relevant contrast agent containing non-hydrogen nuclear spins, and (ii) a compound of formula (III),

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Claims

1. A composition comprising a compound of formula (I), 【Chemistry 1】 In the formula, Z is (i) １ H (proton), ２ (ii) A carbon-carbon double bond (-C=C-) or a carbon-carbon triple bond (-C≡C-) that is substituted to include H (deuterium) or a combination thereof, R １ However, it includes a para-hydrogen-induced polarization (PHIP) transfer portion, R ２ However, it contains optionally substituted hydrocarbons, alkoxy groups, primary amines, secondary amines, or tertiary amines. R ３ However, it includes a bio-related contrast agent that contains non-hydrogen nuclear spins and is at least partially isotope-labeled with said non-hydrogen nuclear spins, The PHIP moving part is *CH ２ , *CH ２ -*CH ２ , *CHY, *C=Y, *CR ４ R ５ , *CR ４ Y, *C=Y, *CR ６ R ７ -*CR ８ R ９ or any deuterated version thereof, *C is １２ C or １３ It is a carbon isotope of C, R ４ and R ５ However, each independently, hydrogen, １ H, ２ H, ３ Selected from H, linear, branched, or cyclic C1-C10 alkyl hydrocarbons, C6 aryls, benzyls, phenyls, heteroaryls, and haloalkyl groups, Y is selected from a spin 1 / 2 atom and a spin 1 / 2 atom covalently bonded to one or more chemical parts selected from the group consisting of N, O, and S, which are optionally substituted with a linear, branched, or cyclic C1-C10 alkyl hydrocarbon, C6 aryl, benzyl, phenyl, heteroaryl, halogen, or haloalkyl group, and the spin 1 / 2 atom is 1H, 13C, 15N, 19F, or 31P. R ６ , R ７ , R ８ , and R ９ However, each operates independently. １ H, ２ H, ３ A composition selected from H, linear, branched, or cyclic C1-C10 alkyl hydrocarbons, C6 aryls, benzyls, phenyls, heteroaryls, and haloalkyl groups.

2. The composition according to claim 1, wherein the PHIP moving portion includes at least one atom that is J-coupled with the non-hydrogen nucleus spin at least 0.1 Hertz (Hz), or Z includes at least one atom that is J-coupled with the non-hydrogen nucleus spin at least 0.1 Hertz (Hz).

3. R ２ The composition according to claim 1, wherein the group is selected from a methyl group, an ethyl group, a propyl group, an isopropyl group, an n-butyl group, an s-butyl group, a t-butyl group, an isobutyl group, a hydroxyl group, a methyl alcohol group, an ethyl alcohol group, an n-propanol group, an isopropyl alcohol group, an alcohol propionic acid group, an n-butyl alcohol group, an s-butyl alcohol group, a t-butyl alcohol group, an isobutyl alcohol group, a methoxy group, an ethoxy group, an ethoxy group, an isopropoxy group, an alcohol propionic acid group, an ester group, an ester group, a phenyl group, a substituted phenyl group, a primary amine group, a secondary amine group, a tertiary amine group, a primary amide group, a secondary amide group, and a tertiary amide group.

4. The aforementioned bio-related contrast agent is formula R １０ It contains a compound of C(=O)X-, where R １０ The alkyl group is selected from a linear, branched, or cyclic C1-C10 alkyl group, and one or more C atoms are C=C, CO, COH, CNH ２ COOH, CH ２ COOH, CONH ２ , is optionally substituted with OC (=O), and X is NR １１ Selected from , S, and O, in the formula, R １１ The composition according to claim 1, wherein is selected from hydrogen and an amino protecting group optionally selected from trifluoroacetyl, acetyl, benzoyl, carbobenzoxy, tert-butyl carbonate, and benzyl.

5. The composition according to claim 1, wherein the bio-related contrast agent is selected from pyruvate, glutamate, glutamine, lactic acid, acetic acid, acetoacetic acid, zymonate, alanine, fructose, fumaric acid, bicarbonate, urea, dehydroascorbic acid, α-ketoglutaric acid, dihydroxyacetone, glucose, ascorbic acid, and their conjugate acids.

6. The composition according to claim 1, wherein the composition has a solubility in water of less than 50 mmol (mM).

7. The composition according to claim 1, wherein the composition has a solubility in an organic solvent of less than 50 mmol (mM), and the organic solvent is acetone, ethanol, chloroform, or toluene.

8. The composition according to claim 1, wherein reacting the composition with parahydrogen yields a chemical yield of at least 30% of the parahydrogenated product.

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