A versatile synthetic route to the preparation of 15 N heterocycles

Abstract

A robust medium-scale (approximately 3 g) synthetic method for ¹⁵ N labeling of pyridine (¹⁵ N-Py) is reported based on the Zincke reaction. ¹⁵ N enrichment in excess of 81% was achieved with approximately 33% yield. ¹⁵ N-Py serves as a standard substrate in a wide range of studies employing a hyperpolarization technique for efficient polarization transfer from parahydrogen to heteronuclei; this technique, called SABRE (signal amplification by reversible exchange), employs a simultaneous chemical exchange of parahydrogen and a to-be-hyperpolarized substrate (e.g., pyridine) on metal centers. In studies aimed at the development of hyperpolarized contrast agents for in vivo molecular imaging, pyridine is often employed either as a model substrate (for hyperpolarization technique development, quality assurance, and phantom imaging studies) or as a co-substrate to facilitate more efficient hyperpolarization of a wide range of emerging contrast agents (e.g., nicotinamide). Here, the produced ¹⁵ N-Py was used for the feasibility study of spontaneous ¹⁵ N hyperpolarization at high magnetic (HF) fields (7 T and 9.4 T) of an NMR spectrometer and an MRI scanner. SABRE hyperpolarization enabled acquisition of 2D MRI imaging of catalyst-bound ¹⁵ N-pyridine with 75 × 75 mm² field of view (FOV), 32 × 32 matrix size, demonstrating the feasibility of ¹⁵ N HF-SABRE molecular imaging with 2.4 × 2.4 mm² spatial resolution.

Keywords: 15N; MRI; PHIP; contrast agent; hyperpolarization; parahydrogen; pyridine.

Figure 1.

A schematic diagram of Signal Amplification by Reversible Exchange (SABRE)^{[48, 49, 73, 74]} for the case of efficient hyperpolarization of the ¹⁵N site of ¹⁵N-pyridine using the SABRE-SHEATH approach. Note the simultaneous chemical exchange of pH₂ and the to-be-hyperpolarized substrate (here, ¹⁵N-pyridine) in the equatorial positions of this IrIMes hexacoordinate complex^[75] and the spontaneous nature of polarization transfer via two-bond spin-spin couplings when the process is performed at the matching magnetic field of approximately 1 micro-Tesla (μT).^{[56, 57, 76]}

Figure 2.

The overall scheme of ¹⁵N-Py enrichment with ¹⁵N (from ¹⁵NH₄Cl source) via Zincke salt formation.

Figure 3.

Reaction scheme from the previous report^[83] of ¹⁵N-pyridine synthesis.

Figure 4.

Reaction scheme and mechanism for the preparation of ¹⁵N-nicotinamide via formation of Zincke salt.

Figure 5.

High-resolution mass spectrometry of the final product (¹⁵N-Py) performed by direct liquid infusion using an Orbitrap mass spectrometer (Thermo-Finnigan, San Jose, CA) equipped with an Ion-Max source housing and an atmospheric pressure chemical ionization (APCI) probe in positive ion mode at a resolving power of 60,000 (at m/z 400). Note the presence of the peak at ca. 82.05 due to contribution from ¹³C natural abundance of ~1.1% (therefore, the probability of having one ¹³C in any of the five carbon positions is greater than 5%).

Figure 6.

¹⁵N NMR spectra obtained for (a) ¹⁵N signal reference of ¹⁵NH₄Cl (37%) aqueous solution, (b) HF-SABRE, and (c) SABRE-SHEATH of HP ¹⁵N resonances of ¹⁵N-Py.

Figure 7.

¹⁵N MRI image of high-field (HF) ¹⁵N SABRE of HP ¹⁵N-pyridine obtained using an EPI pulse sequence with 11 ms echo time (TE), 32×32 matrix size (2.4×2.4 mm² pixel size) over 75×75 mm² field of view (FOV), and 4 acquisitions (total experimental time was approximately 4 seconds).