Unravelling the correlation between metal induced aggregation and cellular uptake/subcellular localization of Znsalen: An overlooked rule for design of luminescent metal probes

Unravelling the unique effects of metal coordination on biological behaviours is of importance to design metal based therapeutic and diagnostic agents. In this work, we chose luminescent Znsalen (ZnL₁) as a case study to demonstrate that metal induced aggregation arising from the intermolecular Zn⋯O interaction influences its cellular uptake and subcellular localization. Comparative studies with the free bases (L₁ and L₂) show that ZnL₁ undergoes cellular uptake through caveolae-mediated endocytosis and internalizes in endosomal/lysosomal compartments, in contrast to the localization of L₁ and L₂ in the mitochondria. Further studies of photophysical properties, TEM imaging and DLS analysis suggest that ZnL₁ tends to form large sized fibrous structures in aqueous media. To investigate the relationship between ZnL₁ aggregation and the biological behaviour, we used pyridine to tune the "aggregation-to-deaggregation" transition and found that, in the presence of pyridine, ZnL₁ could localize in the mitochondria and internalize into cells through the passive diffusion pathway. Such distinctive biological behaviours resulting from the different Znsalen species clearly point out the importance of metal induced aggregation or metal speciation analysis in designing metal complexes as biological probes.