Characterization of the low pH sensing dye, lysosensor yellow/blue DND-160, under high hydrostatic pressures.

摘要

DND-160, a low-pH probe dye, was characterized under high hydrostatic pressures in pH buffers acetate (pKa = 4.8), MOPS (pKa = 7.2) and BES (pKa = 7.1). The effect of pressurization on the fluorescence of DND-160 shows differences compared to previously studied near-neutral pH seminapthorhodafluor (SNARF) and seminaphthofluorescein (SNAFL) dyes, including a pressure sensitivity of the deprotonated emission and a buffer solvent dependent DeltaV of the acid-base reaction for DND-160. The DeltaV was determined from the pressure dependence of the equilibrium constant pKa, calculated from the fluorescence of the dye at different pH. The DeltaV of DND-160 in acetate was about half of the value in MOPS and BES. We hypothesize that the differences in DND-160 pressure response compared to previously studied dyes are due to the differences in their fluorescence mechanism. C.SNAFL and C.SNARF emission is solely dependent on the superposition of the fluorescence of its two dye forms, i.e., the protonated and deprotonated forms, while DND-160 emission depends on two factors, the concentration of the two chemical forms and on the interaction of its locally-excited and charge transfer states.