Understanding complex supramolecular landscapes: non-covalent macrocyclization equilibria examined by fluorescence resonance energy transferElectronic supplementary information (ESI) available: Synthetic details, compound characterization, DOSY experiments, emission and CD measurements and analysis of the titration and competition experiments. See DOI: 10.1039/c8sc03229g

As molecular self-assembled systems increase in complexity, due to a large number of participating entities and/or the establishment of multiple competing equilibria, their full understanding becomes likewise more complicated, and the use of diverse analytical techniques that can afford complementary information is required. We demonstrate in this work that resonance excitation energy transfer phenomena, measured by fluorescence spectroscopy in combination with other optical spectroscopies, can be a valuable tool to obtain supplementary thermodynamic data about complex supramolecular landscapes that other methods fail to provide. In particular, noncovalent macrocyclization processes of lipophilic dinucleosides are studied here by setting up a competition between intra- and intermolecular association processes of Watson-Crick H-bonding pairs. Multiwavelength analysis of the monomer emission changes allowed us to determine cyclotetramerization constants and to quantify chelate cooperativity, which was confirmed to be substantially larger for the G-C than for the A-U pair. Furthermore, when bithiophene-BODIPY donor-acceptor energy transfer probes are employed in these competition experiments, fluorescence and circular dichroism spectroscopy measurements in different regions of the visible spectrum additionally reveal intermolecular interactions occurring simultaneously at both sides of the macrocyclization reaction: the cyclic product, acting as a host for the competitor, and the monomer reactant, ultimately leading to macrocycle denaturation. Energy transfer between FRET pairs is employed herein to reveal novel intermolecular interactions between self-assembled macrocycles, built via Watson-Crick pairing between complementary dinucleosides, and mononucleoside competitors.