Water based hydrogels of (A) P2 at 1.9 wt% (19 mg mL -1 ) in presence of 1 equiv. of Zn(II) in 10% PBS; (B) P2 at 1.9 wt% (19 mg mL -1 ) in 10% PBS; (C) P2 in presence of 0.5 equiv. of Zn(II) in aqueous solution. [inversion test was used as an initial indication of gel formation, while gelation was verified via rheology measurements] 3.2 Structural conformation of stimuli-responsive hydrogels and microstructure P1 and P2 each contain two histidines (Fig. ), which are positively charged below pH 6 (His(imidazole) pK a = 6.0). Under these conditions, the resulting electrostatic repulsion of peptides prevents self-assembly. Conversely, above pH 6, the deprotonation of the histidine moieties removes the electrostatic repulsion and facilitates Zn(II) binding, leading to rapid self-assembly and hydrogel formation (Fig. ). 53 The secondary conformation of the peptides in the samples in presence and absence of Zn(II) was analysed by circular dichroism. In the absence of Zn(II), both P1 (Fig. ) and P2 (Fig. ) display a random coil conformation (negative peak between 190 and 200 nm) at low concentrations (0.5 mM, 4.86 mg mL -1 ). As the concentration is increased, these conformations change, with P1 transitioning to an α-helix conformation at 50 mM, evidenced by the replacement of the negative peak at 197 nm by two at 207 nm and 225 nm, whereas at 30 mM P2 gives a strong positive peak at 199 nm, with no negative peaks at longer wavelength, indicating a conformation related to that of a β-sheet. In the presence of 0.5 equivalents of Zn(OAc) 2 (Fig. and), increasing the peptide concentration had little effect on the conformations of either of the peptides, with both giving CD signals consistent with random coil, as well as displaying some other small peaks which are not straightforward to assign. This suggests that the coordination crosslinks restrict the conformational space of the peptide and take precedence over prior conformational tendencies. In a 10% MES buffer and in the presence of Zn(II) (Fig. and), both peptides gave a classical random coil CD signal which increased in intensity as concentration was raised. In general, the presence of Zn(II) leads to Zn-imidazole coordination, which stabilizes the random coil structure, and may hinder the formation of a well-defined secondary structure. Thus, both peptides appear to have a relatively disordered, but flexible structure. showing a change in their structural conformation from random coil to α-helix and to β-sheet, respectively. In graph B (P1) and graph E (P2), the scans were conducted in an aqueous solution at pH 7 with 0.5 equiv. of Zn(OAc) 2 , while in graph C (P1) and graph F (P2) additional scans were performed in a 10% MES buffer solution with 0.5 equiv. of Zn(OAc) 2 . The peaks at 218 nm and 254 nm of D, E and F can be attributed to the Fmoc group.