Tuning nuclear permeability of naphthalimide Zn²⁺ probes through amine modification: From molecular design to...

Zinc ions (Zn²⁺) function as vital second messengers, and their nuclear localization critically influences gene expression and cell fate. However, research progress has been hindered by the lack of probes capable of efficiently penetrating the nucleus. In this work, seven naphthalimide Zn²⁺ fluorescent probes with different amine modifications (monoamine, diamine, triamine) and BOC lipophilic groups were synthesized. The effect of amine modification on the nuclear permeability of naphthalimide based zinc ion probe was discussed. These probes exhibit nanomolar sensitivity (LOD = 9.3-13.1 nM), strong Zn²⁺-triggered fluorescence enhancement in PBS (pH 7.2), high selectivity, and stability across pH 6-14. Live-cell imaging in MC3T3-E1, SiHa, and C33A cells revealed that amine modifications are essential for nuclear entry. Specifically, probes with fewer nitrogen atoms (monoamines 1a, diamines 2a) demonstrated significantly enhanced nuclear permeability compared to triamine probes. Shorter alkyl chains and moderate hydrophobicity further improved nuclear translocation. Probes 1a and 2a maintained good cell viability while enabling effective subcellular Zn²⁺ tracking. Notably, probe 2a achieved consistent whole-cell localization, whereas 1a exhibited reduced permeability in C33A cells. These rationally designed probes enable spatiotemporally resolved tracking of Zn²⁺ dynamics across subcellular compartments, establishing a molecular toolbox that bridges synthetic chemistry with live-cell biology applications.