Photophysical and Cellular Studies of pH-Sensitive Boron-Functionalised Far-Red Fluorophores

Two novel chromenylium-based probes were developed by functionalisation of the 5-position of the xanthene core with either a para-N-methyldiaminoacetic acid (MIDA) boronate ester (CRpMD) or closo-1,7-carborane (CRmCB). CRpMD revealed near-infrared emission at 655 nm, demonstrating a bathochromic shift compared to the analogous phenyl derivative (CRPh). Both boron-containing probes were found to be sensitive to changes in acidic pH, with CRpMD also displaying a dual fluorescence response at alkaline pH (pKa1 = 2.40 and pKa2 = 9.16). CRmCB showed a ca. 3-fold increase in fluorescence intensity at 638 nm from pH 7.01 to 2.00, with a pKa = 2.08. Moreover, CRpMD and CRmCB were successfully taken up by A549 lung cancer cells to assess their intracellular biodistributions, thereby showcasing the future potential of these probes for time-resolved pH measurements using confocal microscopy.