Synthesis of 15N-Pyridines and Higher Mass Isotopologs via Zincke Imine Intermediates

Abstract

Methods to incorporate stable radioisotopes are integral to pharmaceutical and agrochemical development. However, despite the prevalence of pyridines in candidate compounds, methods to incorporate ¹⁵N atoms within their structures are limited. Here, we present a general approach to pyridine ¹⁵N-labeling that proceeds via ring-opening to NTf-Zincke imines and then ring-closure with commercially available ¹⁵NH₄Cl salts. This process functions on a range of substituted pyridines, from simple building block-type compounds to late-stage labeling of complex pharmaceuticals, and ¹⁵N-incorporation is >95% in most cases. The reactivity of the Zincke imine intermediates also enables deuteration of the pyridine C3- and C5-positions, resulting in higher mass isotopologs required for LCMS analysis of biological fluids during drug development.

Scheme 1.. Current Methods To Synthesize ¹⁵N-Pyridines and an Approach via Zincke Imine Intermediates

Scheme 2.. Development of Two General Protocols for Pyridine ¹⁵N Labeling

^aYields determined by ¹H NMR analysis using mesitylene as an internal standard. ^bIsolated yield. ^c1H NMR yield reported due to the volatility of 3b. ^dAn analytically pure sample of 3b·HCl was obtained in 30% yield.

Scheme 3.. Upgrading Percentage ¹⁵N Incorporation Using Zincke Iminium Intermediates

^a1H NMR yield reported with reference to an internal standard due to product volatility.

Scheme 4.. Deuterium and ¹⁵N Labeling Sequences for [M+2] and [M+3] Pyridines^a,b,c

^aIsolated yields are shown. Percentage deuterium incorporation measured by ¹H NMR spectra. ^cPercentage ¹⁵N incorporation values extrapolated from values from Tables 1 and 2. ^dDeuteration conducted using 200 equiv of AcOD in d₃-MeCN at 80 °C and ring closure with 8 equivalents each of ¹⁵NH₄Cl and NaOAc.