Heteronuclear ¹⁹F-¹H statistical total correlation spectroscopy as a tool in drug metabolism: Study of flucloxacillin biotransformation

We present a novel application of the heteronuclear statistical total correlation spectroscopy (HET-STOCSY) approach utilizing statistical correlation between one-dimensional ¹⁹F/¹H NMR spectroscopic data sets collected in parallel to study drug metabolism. Parallel one-dimensional (1D) 800 MHz ¹H and 753 MHz ¹⁹F{¹H} spectra (n = 21) were obtained on urine samples collected from volunteers (n = 6) at various intervals up to 24 h after oral dosing with 500 mg of flucloxacillin. A variety of statistical relationships between and within the spectroscopic datasets were explored without significant loss of the typically high 1D spectral resolution, generating ¹H-¹H STOCSY plots, and novel ¹⁹F-¹H HET-STOCSY, ¹⁹F-¹⁹F STOCSY, and ¹⁹F-edited ¹H-¹H STOCSY (X-STOCSY) spectroscopic maps, with a resolution of ∼0.8 Hz/pt for both nuclei. The efficient statistical editing provided by these methods readily allowed the collection of drug metabolic data and assisted structure elucidation. This approach is of general applicability for studying the metabolism of other fluorine-containing drugs, including important anticancer agents such as 5-fluorouracil and flutamide, and is extendable to any drug metabolism study where there is a spin-active X-nucleus (e.g., ¹³C, ¹⁵N, ³¹P) label present.