In contrast, a set of new signals appeared in the 1 H-NMR spectra of macrocycle 2 upon addition of H2PO4 -or H2P2O7 2-, characteristic of slow exchange on the NMR chemical shift timescale, which allowed a more complete analysis of the binding events (Figure ). The addition of H2PO4 -to 2 led to a gradual decrease of the original Hb signal (8.37 ppm), with a new signal appearing at 8.5 ppm (Hb') during the first part of the titration (up to 1.3 eq. of H2PO4 -), which disappeared upon further addition of guest with another signal appearing at 8.9 ppm (Hb''). This behavior is indicative of two binding events, the binding of a first guest anion, followed by formation of a 1:2 complex with H2PO4 -. Such 1:2 complexes have been reported previously and their existence has been attributed to the formation of phosphate dimers involving Anti-Electrostatic Hydrogen Bonds (AEHBs). During the titration with H2PO4 -(Figure ), a gradual decrease in the intensity of the Hd azomethine signal at 9.0 ppm was observed concomitant with the appearance of only one new signal Hd' at 9.4 ppm. Signal Hc, only showed some broadening when less than 1 eq. of H2PO4 -was added and it is only upon further addition of the guest that a new signal appeared at 14.1 ppm (Hc'), which is clearly visible in the spectrum with 2.1 eq. of H2PO4 -. The variation of the integration of the Hc and Hd signals upon titration are shown in Figure . The integrations suggest that signal Hd' is a superposition of this proton's signals in both the 1:1 complex and the 1:2 complex, whereas signal Hc would be the superposition of the signals arising from the 1:1 complex and the free receptor. The first H2PO4 -binding event can thus be observed from the azomethine Hd signals and the second event is reflected by the behavior of the NHc signals.