High density orthogonal surface immobilization via photoactivated copper-free click chemistry

Article abstract of DOI:10.1021/ja105066t

Surfaces containing reactive ester polymer brushes were functionalized with cyclopropenone-masked dibenzocyclooctynes for the light activated immobilization of azides using catalyst-free click chemistry. The photodecarbonylation reaction in the amorphous brush layer is first order for the first 45 s with a rate constant of 0.022 s^-1. The catalyst-free cycloaddition of surface bound dibeznocyclooctynes proceeds rapidly in the presence of azides under ambient conditions. Photolithography using a shadow mask was used to demonstrate patterning with multiple azide containing molecules. This surface immobilization strategy provides a general and facile platform for the generation of multicomponent surfaces with spatially resolved chemical functionality.

Full text of DOI:10.1021/ja105066t

Published on Web 07/22/2010
High Density Orthogonal Surface Immobilization via Photoactivated
Copper-Free Click Chemistry
Sara V. Orski, Andrei A. Poloukhtine, Selvanathan Arumugam, Leidong Mao, Vladimir V. Popik,* and
Jason Locklin*
Department of Chemistry, Faculty of Engineering, and the Center for Nanoscale Science and Engineering,
UniVersity of Georgia, Athens, Georgia 30602
Received June 10, 2010; E-mail: jlocklin@chem.uga.edu; vpopik@chem.uga.edu
Scheme 1a
Abstract: Surfaces containing reactive ester polymer brushes
were functionalized with cyclopropenone-masked dibenzocy-
clooctynes for the light activated immobilization of azides using
catalyst-free click chemistry. The photodecarbonylation reaction
in the amorphous brush layer is first order for the first 45 s with
a rate constant of 0.022 s-1. The catalyst-free cycloaddition of
surface bound dibeznocyclooctynes proceeds rapidly in the
presence of azides under ambient conditions. Photolithography
using a shadow mask was used to demonstrate patterning with
multiple azide containing molecules. This surface immobilization
strategy provides a general and facile platform for the generation
of multicomponent surfaces with spatially resolved chemical
functionality.
a (a) Attachment of cyclopropenone (1) to poly(NHS4VB) brushes. (b)
Subsequent photoactivation (2) and functionalization (3) of the polymer
brush pendant groups with azide-derived fluorescent dyes azido-FL and
azido-RB (see Figure SI.1 for structures).
The advancement of engineered particles and surfaces with
spatially resolved chemical functionality is of interest to many areas
of science and technology including the fabrication of biochips,
microfluidic devices, targeted drug delivery, and microelectronic
devices. Among several immobilization strategies developed,1,2 the
alkyne-azide Huisgen 1,3-dipolar cycloaddition is emerging as an
ideal coupling approach.2-4 These “click” reactions are especially
appealing for biological attachment due to their quantitative
reactivity, small size, and the ability to incorporate azides in
biomolecules through native cell machinery.5 While conventional
copper(I)-catalyzed click chemistry is ideal for many applications,6-8
the cytotoxicity of the Cu catalyst can limit bioorthogonal conjuga-
tion. Recently, catalyst-free click reactions have emerged that utilize
a reactive cycloalkyne to promote the [3+2] cycloaddition with
comparable reaction rates.9,10
brush matrix under aminolysis conditions with quantitative conver-
sion (Scheme 1a). When irradiated with UV light (350 nm, 3.5
mW/cm2), 1 undergoes rapid decarbonylation to yield the reactive
dibenzocyclooctyne 2. Cyclooctyne 2 can then undergo catalyst-
free cycloadditions with azides to yield the triazole-linked conjugate
in quantitative yield under ambient conditions (Scheme 1b).
Unexposed cyclopropenone 1 does not react with azides and is
thermally stable at 60 °C for 12 h without decomposition.12,13
Unreacted 1 can be subsequently decarbonylated by further irradia-
tion with UV light.
The consecutive functionalization steps of the brush coatings
were characterized using ellipsometry, contact angle, and grazing-
incidence attenuated total reflectance Fourier transform infrared
spectroscopy (GATR-FTIR). Upon functionalization, polymer brush
thickness increases with the additional molecular weight of the
pendant group and static contact angle measurements confirm
anticipated surface wettability changes (Table SI.1). Figure 1 shows
the progression from the poly(NHS4VB) brush to the covalent
attachment of a fluorescein-azide conjugate (azido-FL) via pho-
toactivated click chemistry. Upon functionalization with 1, the
disappearance of the NHS CdO stretch at 1738, 1769, and 1801
cm-1 is observed along with the appearance of the cyclopropenone
CdO stretch at 1846 cm-1 and conjugation of the CdCsCdO
stretch at 1608 cm-1 (Figure 1a and b). Upon irradiation, the
cyclopropenone CdO stretch disappears yielding cyclooctyne 2
(Figure 1c). After the click reaction, the appearance of carboxylic
acid stretches at 1757 and 1447 cm-1 indicate the attachment of
the azido-FL (see Table SI.2 for complete peak assignments).
The photodecarbonylation of cyclopropenones to alkynes in
solution proceeds quantitatively on the order of picoseconds with
high quantum efficiency (Φ ) 0.2-1.0).13 In the solid crystalline
Herein, we report the functionalization of activated ester polymer
brushes with a cyclopropenone masked dibenzocyclooctyne com-
pound that allows selective immobilization of azido-containing
substrates only upon activation with light. A poly(n-hydroxysuc-
cidimide 4-vinyl benzoate) (poly(NHS4VB)) brush coating was
chosen as a versatile surface platform because it is densely packed
and provides a facile template for postfunctionalization.11 The
electrophilic n-hydroxysuccinimide (NHS) ester pendant group
allows coupling of a wide variety of functional groups, and the
controlled nature of surface initiated polymerization allows for
homopolymer, copolymer, and block copolymer coatings with
precise control of functionality and microenvironment. These
surface bound polymer coatings also allow one to decouple sensitive
chemistry or multistep monomer synthesis from the polymer brush
geometry.
Scheme 1 outlines the surface immobilization strategy. First
poly(NHS4VB) coatings (125 nm) were prepared using surface-
initiated ATRP.11 Cyclopropenone 1 was then immobilized to the
9
11024 J. AM. CHEM. SOC. 2010, 132, 11024–11026
10.1021/ja105066t  2010 American Chemical Society

C O M M U N I C A T I O N S
state, it has also been observed that diarylcyclopropenones can
undergo decarbonylation with a remarkable quantum efficiency (Φ
> 1).14 In order to examine the kinetics of photodecarbonylation
in the amorphous brush layers, time dependent UV-vis spectros-
copy on quartz substrates was performed. Absorption spectra were
recorded at light exposure (350 nm, 3.5 mW/cm2) intervals of 1 s
up to 150 s and are shown in Figure 2. The bands at 335 and 353
nm of cyclopropenone 1 rapidly decrease upon initial exposure to
350 nm light along with the simultaneous emergence of bands at
308 and 326 nm for dibenzocyclooctyne 2. The spectral observations
in the brush layer are consistent with the photodecarbonylation of
1 in solution.12
Figure 2. UV-vis absorption spectroscopy of the simultaneous decar-
bonlyation of cyclopropenone and formation of cyclooctyne on the polymer
brush irradiated with 350 nm UV light. Spectra shown are every 5 s until
complete disappearance of cyclopropenone after 150 s.
of the protected regions for further click reactions. Azido-FL was
then immobilized to generate a multifunctional substrate. Figure 3
shows a fluorescence microscopy image of the photopatterned
substrates. There is negligible cross-contamination between the two
dyes, with excellent segregation between the selectively activated
regions.
Figure 1. Grazing incidence attenuated total reflection Fourier transform
infrared spectroscopy (GATR-FTIR) of the original poly(NHS4VB) brush
functionalized (a), postfunctionalization with cyclopropenone 1 (b), conver-
sion of cyclopropenone to dibenzocyclooctyne 2 (c), and functionalization
with azido-FL 3 (d).
Figure 3. Fluorescence microscope images of a photopatterned surface
fabricated by sequential photoactivation of dibenzocycloocytnes. (a) Click
functionalized azido-RB excited at 550 nm, (b) azido-FL excited at 477
nm, and (c) both dyes imaged under wide UV excitation (350 nm).
The decay of the cyclopropenone absorbance at 353 nm was
found to be first order for the first 45 s of exposure with a rate
constant of 0.022 s-1 (Figure SI.3). Decarbonylation after 45 s
deviates from first-order behavior even though the photoconversion
is unimolecular. The alteration of decarbonylation kinetics is likely
due to nonequivalent sites in the polymer matrix that influence the
absorption coefficient and/or quantum yield such as differing states
of aggregation or a heterogeneous free volume distribution in the
amorphous film.15,16 Overall, 95% of the brush decarbonylation is
complete within 90 s of irradiation with a hand-held UV lamp and
quantitative conversion occurs within 150 s. No photodegradation
of the polymer substrate or cyclooctyne moieties was observed in
the UV-vis spectra after several minutes of exposure to 350 nm
light.
To further demonstrate the versatility of the photoclick substrates,
cyclopropenone functionalized substrates were irradiated through
a shadow mask to form multicomponent surfaces with spatially
resolved chemical functionality. A square patterned transmission
electron microscope grid (12 µm pitch) was used to mask 1 during
irradiation. Substrates were irradiated through the shadow mask
and then immersed in a solution of Lissamine Rhodamine B-azide
conjugate (azido-RB). The cycloaddition is complete within 20 min
and occurred only in the exposed areas, where cyclopropenone
groups underwent decarbonylation to generate dibenzocyclooctyne.
A subsequent flood of irradiation of the substrate liberates the
remaining cyclooctyne groups and allows for the functionalization
In conclusion, we have demonstrated the surface immobilization
of cyclopropenones that undergo photoinduced decarbonylation to
yield dibenzocyclooctynes for catalyst-free cycloaddition with
azides. The decarbonylation reaction occurs quickly and quantita-
tively with low power UV irradiation. This photoactivated surface
platform allows the creation of multifunctional surfaces with
spatially resolved chemical functionality and can be extended to
biological, nanoparticle, and materials science applications.
Acknowledgment. The authors gratefully acknowledge financial
support from the National Science Foundation (ECCS 0901141).
Supporting Information Available: Detailed experimental proce-
dures, ellipsometry and contact angle measurements, decarbonylation
kinetics, and FTIR peak assignments. This material is available free of
charge via the Internet at http://pubs.acs.org.
References
(1) Hensarling, R. M.; Doughty, V. A.; Chan, J. W.; Patton, D. L. J. Am. Chem.
Soc. 2009, 131 (41), 14673–14675.
(2) Im, S. G.; Bong, K. W.; Kim, B.-S.; Baxamusa, S. H.; Hammond, P. T.;
Doyle, P. S.; Gleason, K. K. J. Am. Chem. Soc. 2008, 130 (44), 14424–
14425.
(3) White, M. A.; Johnson, J. A.; Koberstein, J. T.; Turro, N. J. J. Am. Chem.
Soc. 2006, 128 (35), 11356–11357.
(4) Iha, R. K.; Wooley, K. L.; Nystro¨m, A. M.; Burke, D. J.; Kade, M. J.;
Hawker, C. J. Chem. ReV. 2009, 109 (11), 5620–5686.
(5) Prescher, J. A.; Bertozzi, C. R. Nat. Chem. Biol. 2005, 1 (1), 13–21.
(6) Sumerlin, B. S.; Vogt, A. P. Macromolecules 2009, 43 (1), 1–13.
9
J. AM. CHEM. SOC. VOL. 132, NO. 32, 2010 11025

C O M M U N I C A T I O N S
(7) Canalle, L. A.; Berkel, S. S. v.; Haan, L. T. d.; Hest, J. C. M. v. AdV.
Funct. Mater. 2009, 19 (21), 3464–3470.
(8) Bernardin, A.; Cazet, A. l.; Guyon, L.; Delannoy, P.; Vinet, F.; Bonnaffe´,
D.; Texier, I. Bioconjugate Chem. 2010, 21 (4), 583–588.
(9) Baskin, J. M.; Prescher, J. A.; Laughlin, S. T.; Agard, N. J.; Chang, P. V.;
Miller, I. A.; Lo, A.; Codelli, J. A.; Bertozzi, C. R. Proc. Natl. Acad. Sci.
U.S.A. 2007, 104 (43), 16793–16797.
(10) Jewett, J. C.; Bertozzi, C. R. Chem. Soc. ReV. 2010, 39 (4), 1272–1279.
(11) Orski, S. V.; Fries, K. H.; Sheppard, G. R.; Locklin, J. Langmuir 2010, 26
(3), 2136–2143.
(12) Poloukhtine, A. A.; Mbua, N. E.; Wolfert, M. A.; Boons, G.-J.; Popik,
V. V. J. Am. Chem. Soc. 2009, 131 (43), 15769–15776.
(13) Poloukhtine, A.; Popik, V. V. J. Org. Chem. 2003, 68 (20), 7833–7840.
(14) Kuzmanich, G.; Gard, M. N.; Garcia-Garibay, M. A. J. Am. Chem. Soc.
2009, 131 (32), 11606–11614.
(15) Mita, I.; Horie, K.; Hirao, K. Macromolecules 1989, 22 (2), 558–563.
(16) Sekkat, Z.; Knoll, W. PhotoreactiVe Organic Thin Films; Academic Press:
2002.
JA105066T
9
11026 J. AM. CHEM. SOC. VOL. 132, NO. 32, 2010