Transient 19F photo-CIDNP: A practical tool to distinguish intermediate radical species and determine isotropic hyperfine coupling constants of 19F nuclei

Fluorine-containing flavin derivatives can be used as probes in flavin-binding proteins forming radical pairs to exploit the photo-chemically induced dynamic nuclear polarization (photo-CIDNP) effect. Knowledge of the hyperfine structure is crucial for studying the mechanism of intramolecular radical-pair formation in proteins. Transient 19F photo-CIDNP NMR has so far not been used to determine the isotropic hyperfine coupling constants of 19F nuclei. Here, we show that this method provides reliable results by studying three monofluorinated flavin mononucleotide (FMN) derivatives in conjunction with 6-fluoro-tryptophan. Combining this method with transient 1H photo-CIDNP spectroscopy leads to a more accurate interpretation of the intermediate radical species forming a radical pair. The gathered information can be used to identify the most promising FMN derivative for usage as a probe for formation of radical pairs in proteins.