Fig. 1. (A) Equilibrium between the two pH-dependent forms of RHG. (B) Fluorescence microscopy observations of (a) DOPC/DOPE/TOCL (2 : 1 : 1) and (b) DOPC/DOPG (1 : 2) GUVs. GUVs were prepared in 1 mM HEPES buffer, pH 6.5, containing 200 mM sucrose. The total concentration of the lipids was 0.5 mM. RHG was added to GUVs for 0.06% of the total lipid concentration of GUVs. The red colour represents RHG fluorescence. White bars represent 5 μm. (C) ¹H NMR (500 MHz) spectra of RHG (1.5 mM) in D₂O medium (a) in the absence of lipids at pH 4.5, (b) in the presence of DOPC/DOPE/TOCL (2 : 1 : 1) LUVs (total lipid, 15 mM) at pH 6.5 and (c) in the presence of DOPG LUVs (total lipid, 15 mM) at pH 5.5. The pD value of the solution was adjusted by addition of 0.01 M CF₃COOH. The protons labeled in (C) correspond to the protons labeled in (A).