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Simultaneous ¹⁹F hyperpolarization of aromatic molecules in aqueous solution

Frederike Euchner¹, Christian Bruns¹, Johannes Bernarding¹, and Markus Plaumann¹
¹Institute of Biometrics and Medical Informatics, Otto-von-Guericke University Magdeburg, Magdeburg, Germany

Synopsis

Fluorinated aromatic substrates are of high interest in biomedical and pharmaceutical applications. To increase the low ¹⁹F MR signal intensity in aqueous solutions, the hyperpolarization technique photo-Chemical Induced Dynamic Nuclear Polarization (photo-CIDNP) can be used. Compounds such as 2-fluoro-tyrosine, 3-fluoro-tyrosine or 3-fluoro-4-hydroxyphenylacetic acid were hyperpolarized repeatable inside the detection field (7T). Strong ¹⁹F signal enhancements could be detected for aromatic systems, which were fluorinated in meta-position. The simultaneous hyperpolarization of different substrates allows e.g. the study of molecule-molecule interactions and their binding behavior.

Introduction

MR signal enhancements can be achieved by using e. g. the hyperpolarization technique photo-Chemical Induced Dynamic Nuclear Polarization (photo-CIDNP).^1,2 Previous studies have shown that ¹⁹F MR hyperpolarization in aqueous solutions is a challenge.^3,4 Metabolism examination of fluorinated drugs as well as their intermolecular interactions are of high interest.^5,6 To date, only photo-chemically induced dynamic nuclear polarization (photo-CIDNP) allows the ¹⁹F hyperpolarization in pure water.^3,7 Here, we used a low-cost LED-based set-up.^7-9

Theory

In photo-CIDNP experiments, the polarization forms out of the reactions between radical pairs. The signal formation is determined by the radical and radical pair parameters such as hyperfine couplings and g-factors in the detection field. photo-CIDNP is strongly related to the singlet and triplet states, which are formed by the interaction of two electrons with spin-1/2. These radical pairs occur because of their limited motion in the solvent cage. Formation of radicals can be achieved e. g. by photolytic excitation of a precursor molecule (here: riboflavin 5’-monophosphate sodium salt hydrate). During the lifetime of the solvent cage the correlated radical pair can recombine (recombination/geminate products) or diffuse apart and pass to side reactions (escape/transfer products). The achieved nuclear hyperpolarization can also be transferred to heteronuclei e.g. ¹³C, ¹⁵N or ¹⁹F, among coupled spins.³

Methods

In the current study, different fluorinated aromatic substrates were dissolved in aqueous solution. For example: The amino acids 2-fluoro-DL-tyrosine (2.4 mg) and 3-fluoro-DL-tyrosine (2.1 mg), as well as 2.2 mg of 3-fluoro-4-hydroxyphenylacetic acid (a compound the can be found in olive oil in the non-fluorinated form) were hyperpolarized in D₂O, respectively physiologic salt solution. As photosensitizer riboflavin 5’-monophosphate sodium salt hydrate (1.2 mg) were dissolved in 6 ml physiologic salt solution. 600 µl of this stock solution were used in MR measurements (5 mm NMR tube). An optical fiber connected to a Cree XP E high power LED (455 nm, 3.4 V, ~400 mA) was centrally positioned in the solution. The irradiation was controlled via a self-written pulse program of a 7 T NMR spectrometer (Bruker WB-300 Ultrashield). Irradiation times were between 0 s and 15 s. ¹⁹F NMR spectra were measured using a 90° pulse (P1 = 32.5 µs, PL1 = 17 W).

Results

Figure 1 shows the ¹⁹F NMR spectrum in thermal equilibrium of a mixture of 2-fluoro-DL-tyrosine, 3-fluoro-DL-tyrosine and 3-fluoro-4-hydroxyphenylacetic acid. Without light irradiation only small signals can be detected.
When switch on the LED, the signals of 3-fluoro-DL-tyrosine and 3-fluoro-4-hydroxyphenylacetic acid will be decreased. The signal amplification depends, among other things, on the concentration of the respective substrate and the photosensitizer used. It was found that with lower concentrations the signal amplification increases.⁷
Figure 2 shows, for example, the signal enhancements in dependence of the irradiation time in the case of a 2 mM solution of 3-fluoro-DL-tyrosine in aqueous solution.
In the case of 2-fluorotyrosine no signal enhancement can be detected at a field strength of 7 T. To observe here is a sign change with increasing irradiation time. Thus, a polarization transfer to ¹⁹F is measured.
Further substrates that have been studied in our lab are 3-fluoro-4-hydroxybenzoic acid and 4-hydroxy-3-(trifluoromethyl)benzoic acid. 3-Fluoro-4-hydroxybenzoic acid shows comparable signal enhancements as 3-fluoro-DL-tyrosine and 3-fluoro-4-hydroxyphenylacetic acid.

Discussion

The results show that the hyperpolarization of ¹⁹F nuclei in aqueous solution is possible. With photo-CIDNP this is repeatable without adding a substrate again. The type or phase of the hyperpolarized signals is dependent on the substitution of the aromatic system and the magnetic field in which is measured. In addition, photo-CIDNP allows the simultaneous hyperpolarization of various substrates directly in the detection field. No further transfer or transport is necessary.

Conclusion

Photo-CIDNP was shown to significantly enhance ¹⁹F NMR signals of different fluorinated aromatic systems. High signal enhancements were achieved for molecules such as 3-fluoro-DL-tyrosine or 3-fluoro-4-hydroxyphenylacetic acid. Low or no increase of ¹⁹F signals is observable for ortho-substituted aromatic rings (e. g. 2-fluoro-DL-tyrosine). The simultaneous hyperpolarization of different substrates directly inside the detection field allows a fast examination of molecule interactions as well as investigations of metabolisms.

Acknowledgements

No acknowledgement found.

References

1. Bargon J, et al. Kernresonanz-Emissionslinien während rascher Radikalreaktionen. Zeitschrift Naturforschung Teil A, 1967;22:1551-1555.

2. Goez M, Photo-CIDNP spectroscopy. Annual reports on NMR Spectroscopy, 2009;66:77-147.

3. Flögel U, Ahrens E, 1. Ed. , Fluorine Magnetic Resonance Imaging, Pan Stanford Publishing, Singapore, 2017.

4. Hövener JB, Pravdivtsev AN, Kidd B, et al. Parahydrogen-Based Hyperpolarization for Biomedicine. Angew. Chem. Int. Ed., 2018;57:11140 – 11162.

5. Khan F, Kuprov I, Craggs TD, et al. ¹⁹F NMR Studies of the Native and Denatured States of Green Fluorescent Protein JACS, 2006;128:10729–10737.

6. Ojima I, Fluorine in Medical Chemistry and Chemical Biology, 1.Ed., Wiley-Blackwell: Chichester, 2009.

7. Bernarding J, Euchner F, Bruns C, et al. Low-cost LED-based Photo-CIDNP Enables Biocompatible Hyperpolarization of ¹⁹F for NMR and MRI at 7 T and 4.7 T. ChemPhysChem., 2018;19:2453-2456.

8. Feldmeier C, Bartling H, Riedle E, Gschwind R.M. LED based NMR illumination device for mechanistic studies on photochemical reactions – Versatile and simple, yet surprisingly powerful. J. Magn. Res., 2013;232:39-44.

9. Feldmeier C, Bartling H, Magerl K, Gschwind R.M. LED-Illuminated NMR Studies of Flavin-Catalyzed Photooxidations Reveal Solvent Control of the Electron-Transfer Mechanism. Angew. Chem., 2015;127:1363-1367.

Figures

Figure 1: ¹⁹F MR spectrum (7 T, number of scans: 8) measured without hyperpolarization. The MR signals arise from 2-fluoro-DL-tyrosine (-115.9 ppm), 3-fluoro-DL-tyrosine (-136.8 ppm) and 3-fluoro-4-hydroxyphenylacetic acid (-137.4 ppm).

Figure 2: The ¹⁹F MR signal enhancement of 3-fluoro-DL-tyrosine depends on the duration of irradiation. As many different measurements in our lab have shown, the maximum signal amplification is already reached after about 8 seconds of irradiation time. Longer irradiation times only show a slight effect with regard to the signal amplification.

Figure 3: The irradiation of blue light (455 nm) leads in the case of 2-fluorotyrosine initially to a signal reduction, and then a phase reversal is detectable.

Proc. Intl. Soc. Mag. Reson. Med. 28 (2020)

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