A novel photoreactive amphiphile of nitrophenylazide for immobilization of bioactive proteins

Monolayers of an amphiphilic nitrophenylazide (ANPA) derivative were prepared and their ability to immobilize a protein molecule by a photochemical binding reaction was demonstrated as a novel means to constitute a biofunctional membrane. Molecular orientations and photoreactions in the ANPA monolayers were investigated by Fourier transform IR spectroscopy. Rapid photolysis of ANPA was observed which reflects decomposition of the azide group to a reactive nitrene radical intermediate. As an enzyme, glucose oxidase (GOD) was immobilized on a monomolecular film of ANPA, coated on a substrate, by means of enzyme adsorption at the solid-solution interface and subsequent photolysis of ANPA. Radioisotope labelling experiments revealed that the enzyme was immobilized at a high surface concentration which corresponds to a closely packed monolayer of GOD. The specific activity of the immobilized enzyme was estimated to be high compared with those of other systems involving the adsorption of this enzyme. The usefulness of the present technique for fabricating biofunctional monolayer assemblies is discussed.