Molecular design of fluorescent pH sensors based on reduced rhodol by structure-pKa relationship for imaging of lysosome

Abstract

pH-homeostasis is crucial for maintaining a living system of cells. Lysosome, a digestive system of cells, contains a number of hydrolytic enzymes which can normally function in a narrow range of acidic pH (4.5–5.5). Abnormal pH in lysosomes leads to lysosomal dysfunction, which results in various diseases such as lysosomal storage disorders, cancer, cardiovascular diseases, and neurodegenerative diseases. We previously reported a series of reduced xanthene fluorophores including fluorescein, rhodol and rhodamine, and discovered a reduced rhodol, Rh–OMe, with a methoxy and a free amino group in a xanthene ring. In this work, a series of novel pH-sensitive fluorescent sensors for monitoring acidic lysosomes was developed by structure-pKa relationship study of Rh–OMe. Rh–OMe derivatives with various substituents in an alkoxy group on xanthene ring were designed and evaluated by the analysis of pH-titration curves and photochemical properties. Reduced rhodol probes with a para-substituted benzyloxy group showed excellent fluorescence transition triggered by acidic pH, and they were successfully applied to in vitro imaging of lysosomes in HepG2 cells.