Synthesis of a Photo-Protected Fluorophore to Investigate the Viability of EANBP to Deprotect Amines with Rapid Efficiency

Abstract

In the past decade, photo-removable protecting groups (PPGs) have served as valuable tools in probing biochemical pathways with high spatial and temporal control. When attached to a substrate of choice, they render it chemically inert – allowing it to be introduced into biological environments without affecting cells. Researchers can then use lasers to stimulate the PPG in desired areas to release the substrate, allowing its interaction with receptors in tight localizations. Modern-day PPGs have developed the compatibility with two-photon excitation (2PE) and the ability to release substrates at incredibly quick rates. However, a universal deficiency that these PPGs contain is the inability to deprotect amines with the latter property. This has limited their application with many types of important amine containing molecules such as monamines. The novel PPG, 2-(4’-((di(tris-ethoxy (methyl))amino)-4-nitro-[1,1’-biphenyl]-3-yl)propyl (EANBP), was developed by Donato et al. in 2012 and has demonstrated unprecedented 2PE ability in deprotecting various substrates on microsecond timescales; however, it has not been researched in its ability to do the same with amines. We attempted to synthesize EANBP attached the fluorophore 7-amino-4-methyl-3- coumarinylacetic acid (AMCA-H) to visualize the rates in which EANBP deprotects these functional groups. EANBP-AMCA-H was never achieved; however, in the process we discovered reductive amination was a facile way to directly link amines to the nitrophenylpropyl moiety of EANBP. In addition, we addressed optimization issues concerning the synthesis of the reactive aldehyde precursor needed for the reaction. We are confident that this developed pathway will aide future endeavors to investigate if EANBP/nitrophenylpropyl caging moieties can support the rapid deprotection of amines.