An Efficient Method for Covalent Surface Functionalization of Ultrasmall Metallic Nanoparticles by Surface Azidation Followed by Copper‐Catalyzed Azide‐Alkyne Cycloaddition (Click Chemistry)

Abstract

The azidation of glutathione (GSH)‐functionalized ultrasmall gold nanoparticles (2 nm) by the azide transfer reagent imidazole‐1‐sulfonyl azide hydrogen sulfate leads to azide‐terminated nanoparticles with high yield. A subsequent copper‐catalyzed azide‐alkyne cycloaddition (CuAAC), i. e. a click reaction, leads to covalently functionalized nanoparticles. This was demonstrated with two alkyne‐functionalized dyes, i. e. FAM‐alkyne and AlexaFluor‐647‐alkyne, that were covalently coupled to the nanoparticles. The integrity of the glutathione ligand and the successful surface azidation were demonstrated by one‐dimensional and two‐dimensional NMR spectroscopy. The surface composition of the nanoparticles was determined by quantitative NMR spectroscopy and UV/vis spectroscopy. Each nanoparticle carries 125 glutathione molecules of which 118 were substituted by an azide group. After dye conjugation, either 6 FAM molecules or 11 AlexaFluor‐647 molecules were present on each nanoparticle, respectively. The dye‐clicked nanoparticles were highly fluorescent due to the absence of surface plasmon resonance. The post‐functionalization of GSH avoids a chemical reaction of a functional ligand during the reduction reaction, gives a high yield (up to 50 mg nanoparticles per batch), is based on water as solvent, and is applicable for metallic nanoparticles in general.

Covalent Attachment of Aggregation-Induced Emission Molecules to the Surface of Ultrasmall Gold Nanoparticles to Enhance Cell Penetration

“…Analytical data of AIE luminophores, clicked to gold nanoparticles at c(NP) = 617 nM (30 mg L −1 Au). To compute the number of AIE molecules on each particle, the diameter of the solid core was set to 2 nm, as known from HRTEM data [39]. The clicking efficiency was defined as the ratio of the number of clicked AIE ligands to the number of available azide groups (118) on each particle [39].…”

“…To compute the number of AIE molecules on each particle, the diameter of the solid core was set to 2 nm, as known from HRTEM data [39]. The clicking efficiency was defined as the ratio of the number of clicked AIE ligands to the number of available azide groups (118) on each particle [39]. Note that the hydrodynamic diameter by DCS also includes the ligand shell.…”

“…The AIE luminophores 1 to 3 were then clicked to the surface of azide-terminated glutathione-stabilized ultrasmall gold nanoparticles (2 nm) [ 39 ] to enhance their dispersibility in water and their ability to enter cells. Disc centrifugal sedimentation (DCS) gave the hydrodynamic particle diameter ( Figure 2 ).…”

“…The Au-AIE-3 nanoparticles were bigger (3.2 nm) than the two other particles (1.4 and 1.7 nm) due to the hydrated (PEG) 4 group that contributed to the hydrodynamic diameter. The gold nanoparticles before copper-catalyzed azide–alkyne click reaction (CuAAc) had a hydrodynamic diameter by DCS of 1.5 nm [ 39 ]. Note that DCS systematically underestimates the particle diameter due to the hydration shell [ 42 ].…”

“…Here, we describe the covalent attachment of AIE emitters by Cu(I)-catalyzed azide–alkyne cycloaddition to the surface of ultrasmall gold nanoparticles and demonstrate their potential as imaging agents. Unlike larger gold nanoparticles (10 nm or more) [ 37 , 38 ], ultrasmall gold nanoparticles do not quench the emission of attached fluorophores [ 39 ], making them excellent carriers for AIE molecules.…”