Orthogonal binding and displacement of different guest types using a coordination cage host with cavity-based and surface-based binding sites

The octanuclear Co(ii) cubic coordination cage system H (or HW if it bears external water-solubilising substituents) has two types of binding site for guests. These are (i) the partially-enclosed central cavity where neutral hydrophobic organic species can bind, and (ii) the six ''portals'' in the centres of each of the faces of the cubic cage where anions bind via formation of a network of CH⋯X hydrogen bonds between the anion and CH units on the positively-charged cage surface, as demonstrated by a set of crystal structures. The near-orthogonality of these guest binding modes provides the basis for an unusual dual-probe fluorescence displacement assay in which either a cavity-bound fluorophore (4-methyl-7-amino-coumarin, MAC; λ_em = 440 nm), or a surface-bound anionic fluorophore (fluorescein, FLU; λ_em = 515 nm), is displaced and has its emission ‘switched on’ according to whether the analyte under investigation is cavity-binding, surface binding, or a combination of both. A completely orthogonal system is demonstrated based using a Hw/MAC/FLU combination: addition of the anionic analyte ascorbate displaced solely FLU from the cage surface, increasing the 515 nm (green) emission component, whereas addition of a neutral hydrophobic guest such as cyclooctanone displaced solely MAC from the cage central cavity, increasing the 440 nm (blue) emission component. Addition of chloride results in some release of both components, and an intermediate colour change, as chloride is a rare example of a guest that shows both surface-binding and cavity-binding behaviour. Thus we have a colourimetric response based on differing contributions from blue and green emission components in which the specific colour change signals the binding mode of the analyte. Addition of a fixed red emission component from the complex Ru(bipy)₃]²⁺ (Ru) provides a baseline colour shift of the overall colour of the luminescence closer to neutral, meaning that different types of guest binding result in different colour changes which are easily distinguishable by eye.

Orthogonal binding of neutral or anionic fluorophores to the cavity or surface, respectively, of a coordination cage host allows a dual-probe displacement assay which gives a different fluorescence colorimetric response according to where analyte species bind.