Photochemical Sciences Ph.D. Dissertations


Photoaffinity Labeling Via Nitrenium Ion Chemistry: The Photochemistry of 4-aminophenylazides

Date of Award


Document Type


Degree Name

Doctor of Philosophy (Ph.D.)


Photochemical Sciences

First Advisor

R. Marshall Wilson, PhD

Second Advisor

Zhaohui Xu, PhD (Committee Member)

Third Advisor

Michael Y. Ogawa, PhD (Committee Member)

Fourth Advisor

Thomas H. Kinstle, PhD (Committee Member)


Phenyl azides with powerful electron-donating substituents are known to deviate from the usual photochemical behavior of other phenyl azides. They do not undergo ring expansion, but form basic nitrenes that protonate to form nitrenium ions. The photochemistry of the widely used photoaffinity labeling system 4-amino-3-nitrophenyl azide, has been studied by transient absorption spectroscopy from femtosecond to microsecond time domains and from a theoretical perspective. The nitrene generation from 4-amino-3-nitrophenyl azide occurs on the S2 surface, in violation of Kasha’s rule. The resulting nitrene is a powerful base and abstracts protons extremely rapidly from a variety of sources to form a nitrenium ion. In methanol, this protonation occurs in about 5 ps, which is the fastest intermolecular protonation observed to date. Suitable proton sources include alcohols, amine salts, and even acidic C-H bonds such as acetonitrile. The resulting nitrenium ion is stabilized by the electron-donating 4-amino group to afford a diiminoquinone-like species that collapses relatively slowly to form the ultimate crosslinked product. In some cases in which the anion is a good hydride donor, cross-linking is replaced by reduction of the nitrenium ion to the corresponding amine. However, the efficiency of 4-amino-3-nitrophenyl azide in generating the reactive nitrenium is impaired by associated photochemistry of the nitro group and a tendency for the initially formed nitrene to undergo reduction to the corresponding amine. We have examined possible alternative molecules that might yield reactive nitrenium ion more efficiently, and thus, provide more effective photoaffinity labeling agents. The 2-(N,N-diethylamino)-5-azidopyridine has been investigated as this regard and found to offer substantial advantages over 4-amino-3- iii nitrophenyl azide as a photoaffinity agent. Ultrafast transient spectroscopy confirms that 2-(N,Ndiethylamino)- 5-azidopyridine proceed via a reactive nitrenium ion species, and the same reactive species is formed both photochemically and thermally at relatively low temperatures. The coupling reactions of 2-(N,N-diethylamino)-5-azidopyridine proceed more rapidly than those of 4-amino-3-nitrophenyl azide in reactions with fewer side products, including reduction to the amino analogs. In addition, 2-(N,N-diethylamino)-5-azidopyridine displays fluorescence that ceases upon conversion to products, a property that might offer distinct advantages in photoaffinity labeling studies in complex biological systems.