¹⁵N-Labelling and structure determination of adamantylated azolo-azines in solution

Determining the accurate chemical structures of synthesized compounds is essential for biomedical studies and computer-assisted drug design. The unequivocal determination of N-adamantylation or N-arylation site(s) in nitrogen-rich heterocycles, characterized by a low density of hydrogen atoms, using NMR methods at natural isotopic abundance is difficult. In these compounds, the heterocyclic moiety is covalently attached to the carbon atom of the substituent group that has no bound hydrogen atoms, and the connection between the two moieties of the compound cannot always be established via conventional ¹H-¹H and ¹H-¹³C NMR correlation experiments (COSY and HMBC, respectively) or nuclear Overhauser effect spectroscopy (NOESY or ROESY). The selective incorporation of ¹⁵N-labelled atoms in different positions of the heterocyclic core allowed for the use of ¹H-¹⁵N (J_HN) and ¹³C-¹⁵N (J_CN) coupling constants for the structure determinations of N-alkylated nitrogen-containing heterocycles in solution. This method was tested on the N-adamantylated products in a series of azolo-1,2,4-triazines and 1,2,4-triazolo[1,5-a]pyrimidine. The syntheses of adamantylated azolo-azines were based on the interactions of azolo-azines and 1-adamatanol in TFA solution. For azolo-1,2,4-triazinones, the formation of mixtures of N-adamantyl derivatives was observed. The J_HN and J_CN values were measured using amplitude-modulated 1D ¹H spin-echo experiments with the selective inversion of the ¹⁵N nuclei and line-shape analysis in the 1D ¹³С spectra acquired with selective ¹⁵N decoupling, respectively. Additional spin-spin interactions were detected in the ¹⁵N-HMBC spectra. NMR data and DFT (density functional theory) calculations permitted to suggest a possible mechanism of isomerization for the adamantylated products of the azolo-1,2,4-triazines. The combined analysis of the J_HN and J_CN couplings in ¹⁵N-labelled compounds provides an efficient method for the structure determination of N-alkylated azolo-azines even in the case of isomer formation. The isomerization of adamantylated tetrazolo[1,5-b][1,2,4]triazin-7-ones in acidic conditions occurs through the formation of the adamantyl cation.