The light-promoted
conversion of extensively used cyanine dyes
to blue-shifted emissive products has been observed in various contexts.
However, both the underlying mechanism and the species involved in
this photoconversion reaction have remained elusive. Here we report
that irradiation of heptamethine cyanines provides pentamethine cyanines,
which, in turn, are photoconverted to trimethine cyanines. We detail
an examination of the mechanism and substrate scope of this remarkable
two-carbon phototruncation reaction. Supported by computational analysis,
we propose that this reaction involves a singlet oxygen-initiated
multistep sequence involving a key hydroperoxycyclobutanol intermediate.
Building on this mechanistic framework, we identify conditions to
improve the yield of photoconversion by over an order of magnitude.
We then demonstrate that cyanine phototruncation can be applied to
super-resolution single-molecule localization microscopy, leading
to improved spatial resolution with shorter imaging times. We anticipate
these insights will help transform a common, but previously mechanistically
ill-defined, chemical transformation into a valuable optical tool.