Spatiotemporal Mapping of the Evolution of Silver Nanoparticles in Living Cells

Bioaccumulated silver nanoparticles (AgNPs) can undergo transformation and release toxic Ag+, which can be further reduced and form secondary AgNPs (Ag(0)NPs). However, the intricate interconversions among AgNPs, Ag+, and Ag(0)NPs remain speculative. Herein, we developed a bioimaging method by coupling the aggregation-induced emission method with the label-free confocal scattering and hyperspectral imaging techniques to quantitatively visualize the biodistribution and biotransformation of AgNPs, Ag(0)NPs, and Ag+ in living cells. We demonstrated that AgNPs were first dissolved in the medium, and the released Ag+ was converted into Ag(0)NPs with the presence of algal extracellular polymeric substances and light. Under these conditions, AgNPs alone accounted for 12.4% of the total AgNP toxicity, a percentage comparable to that of Ag(0)NPs (15.6%). However, Ag+ remained the primary contributor to overall AgNP toxicity. Additionally, we found that about 9.00% of the accumulated AgNPs within the algal cells were transformed after 24 h exposure. Of these transformed AgNPs, 4.70% remained as Ag+ forms (located in the apical region, nucleus, and pyrenoid), while 4.30% persisted as (AgNP)-N-0 forms (located in the cytosol) that were only detectable after a 4 h exposure. We further showed that AgNP exposure upregulated algal glutathione production with a 38.3-fold increase in glutathione reductase activity, which potentially resulted in (AgNP)-N-0 formation at the active site. Overall, this study differentiated the toxicity of AgNPs, Ag+, and Ag(0)NPs and directly visualized the ongoing transformation and translocation of AgNPs, Ag+, and Ag(0)NPs within living cells, which are critical in unveiling the toxicity mechanisms of AgNPs.