Fast Disinfecting Antimicrobial Surfaces
Ahmad E. Madkour,†Jeffery M. Dabkowski,†,‡Klaus Nu ¨sslein,‡and Gregory N. Tew*,†
Department of Polymer Science & Engineering and Department of Microbiology, UniVersity of
Massachusetts, Amherst, Massachusetts 01003
ReceiVed September 8, 2008. ReVised Manuscript ReceiVed October 30, 2008
Silicon wafers and glass surfaces were functionalized with facially amphiphilic antimicrobial copolymers using the
“grafting from” technique. Surface-initiated atom transfer radical polymerization (ATRP) was used to grow
became highly antimicrobial and killed S. aureus and E. coli 100% in less than 5 min. The molecular weight and
grafting density of the polymer were controlled by varying the polymerization time and initiator surface density.
Antimicrobial studies showed that the killing efficiency of these surfaces was independent of polymer layer thickness
or grafting density within the range of surfaces studied.
Hospital-acquired infections pose a major global healthcare
issue. Over 2 million cases are reported annually in the USA
alone, leading to 100 000 deaths and adding nearly 5 billion
dollars to U.S. healthcare costs.1,2Contamination of medical
skin, and the ambient atmosphere of the hospital.4In addition,
Medicare recently adopted a new policy that will not reimburse
hospitals for these acquired infections.5,6
Bacterial contamination of surfaces typically begins with the
initial adherence of only a few microorganisms to the surface
following implantation,7but then develops into a biofilm in less
and the host’s own immune system. In order to reduce or even
prevent these infections, an attractive strategy is to consider
materials that resist the initial phase of bacterial colonization,
thus preventing biofilm formation.9Strategies to make antimi-
crobial materials include the addition of leachable biocides to
the material.8,10However, these materials have disadvantages
as short durations of antimicrobial action due to rapid leaching.
Alternatively, biocides can be covalently bonded to the surface,
or a nonleaching biocide can be used.1N-Halamines are
commonly used to make biocidal materials.11-13Quaternary
ammonium compounds (QAC) are widely used as biocides and
were successfully applied as antimicrobial layers over glass14
QACs have been previously covalently attached onto different
materials such as glass,14,16-19polymers,20,21paper,19and
The killing mechanism of bacteria by those surfaces, and
especially the effect of polymer length on killing efficiency, is
still under debate. Klibanov and co-workers hypothesized that
the killing efficiency of immobilized polycationic chains would
be enhanced if the chains are sufficiently long and flexible.17
chain lengths (160 kDa and 25 kDa rather than 60 kDa and 2
kDa, respectively) had stronger bactericidal activity than the
shorter ones. However, their “grafting onto” technique allowed
the backbone; thus, the actual free chain length of the attached
polymer is not precisely known. Ober and co-workers demon-
strated that the antimicrobial properties of surfaces coated with
mers were related to the molecular composition and polymer
organization in the top 2-3 nm of the surface. Furthermore,
their results suggest that molecular weight does not seem to be
* Corresponding author. Prof. Dr. Gregory N. Tew, University of
120 Governors Drive, Amherst, MA 01003, USA. Fax: +1-413-545-0082,
†Department of Polymer Science & Engineering.
‡Department of Microbiology.
(1) Madkour, A. E.; Tew, G. N. Polym. Int. 2008, 57, 6–10.
(2) Gabriel, G. J.; Som, A.; Madkour, A. E.; Eren, T.; Tew, G. N. Mater. Sci.
Eng., R 2007, R57, 28–64.
(3) Stamm, W. E. Ann. Intern. Med. 1978, 89, 764–769.
(4) An, Y. H.; Friedman, R. J. J. Hosp. Infect. 1996, 33, 93–108.
(5) Wald, H. L.; Kramer, A. M. JAMA 2007, 298, 2782–2784.
(6) Department of Health and Human Services. Centers for Medicare &
Medicaid Services. Fed Regist.: 2008; Vol. 73, pp 23528-23938.
(7) von Eiff, C.; Jansen, B.; Kohnen, W.; Becker, K. Drugs 2005, 65, 179–
(8) Hetrick, E. M.; Schoenfisch, M. H. Chem. Soc. ReV. 2006, 35, 780–789.
(9) Williams, J. F.; Worley, S. D. J. Endourol. 2000, 14, 395–400.
(10) Danese, P. N. Chem. Biol. 2002, 9, 873–880.
(11) Kocer, H. B.; Akdag, A.; Ren, X.; Broughton, R. M.; Worley, S. D.;
Huang, T. S. Ind. Eng. Chem. Res. 2008, 47, 7558–7563.
(12) Makal, U.; Wood, L.; Ohman, D. E.; Wynne, K. J. Biomaterials 2006,
(13) Chen, Z. B.; Sun, Y. Y. Ind. Eng. Chem. Res. 2006, 45, 2634–2640.
(14) Isquith, A. J.; Abbott, E. A.; Walters, P. A. Appl. Microbiol. 1972, 24,
(15) El Ola, S. M. A.; Kotek, R.; White, W. C.; Reeve, J. A.; Hauser, P.; Kim,
J. H. Polymer 2004, 45, 3215–3225.
2007, 28, 4870–4879.
(17) Tiller, J. C.; Liao, C.-J.; Lewis, K.; Klibanov, A. M. Proc. Natl. Acad.
Sci. U.S.A. 2001, 98, 5981–5985.
(18) Ku ¨gler, R.; Bouloussa, O.; Rondelez, F. Microbiology 2005, 151, 1341–
(19) Lee, S. B.; Koepsel, R. R.; Morley, S. W.; Matyjaszewski, K.; Sun, Y.;
Russell, A. J. Biomacromolecules 2004, 5, 877–882.
(20) Tiller, J. C.; Lee, S. B.; Lewis, K.; Klibanov, A. M. Biotechnol. Bioeng.
2002, 79, 465–471.
(21) Cen, L.; Neoh, K. G.; Kang, E. T. Langmuir 2003, 19, 10295–10303.
M.; Sciannamea, V.; Lenoir, S.; Cossement, D.; Gouttebaron, R.; Je ´ro ˆme, R.;
Je ´ro ˆme, C. Langmuir 2004, 20, 10718–10726.
(23) Lin, J.; Qiu, S.; Lewis, K.; Klibanov, A. M. Biotechnol. Bioeng. 2003,
(24) Krishnan, S.; Ward, R. J.; Hexemer, A.; Sohn, K. E.; Lee, K. L.; Angert,
E. R.; Fischer, D. A.; Kramer, E. J.; Ober, C. K. Langmuir 2006, 22, 11255–
Langmuir 2009, 25, 1060-1067
10.1021/la802953v CCC: $40.75
2009 American Chemical Society
Published on Web 12/16/2008
co-workers25used both “grafting from” and “grafting onto”
techniques to prepare quaternized poly(2-dimethylamino)ethyl
methacrylate immobilized on both silicon wafers and glass
surfaces. In these studies, they prepared surfaces with various
polymer chain lengths and surface charge density. Their results
of polymer chain length but is dependent on surface charge
Facially amphiphilic, cationic polymers belong to a class of
molecules that mimic natural host defense peptides.2,26They
contain hydrophobic and hydrophilic side chains (protonated
amines), which can segregate to opposite regions, or faces, of
the molecule forming a facially amphiphilic polymer. These
polymers have been termed synthetic mimics of antimicrobial
peptides (SMAMPs).26a–cThis facially amphiphilic topology
and polycationic nature leads to bacterial membrane insertion,
from biocidal QAC is the fact that SMAMPs can be designed
to be antimicrobial yet nontoxic to mammalian cells.2
In this study, we used ATRP to graft one family of these
SMAMPs, poly(butylmethacrylate)-co-poly(Boc-aminoethyl meth-
acrylate)27(3), from silicon wafers and glass surfaces in order
to determine if their antimicrobial activity was retained. Our
results showed that these surface-bound polymers retained their
antibacterial properties and killed S. aureus and E. coli 100%
by contact in less than 5 min. We show that the antimicrobial
properties of these polymers are independent of polymer chain
length and grafting density.
Materials. 5-Hexen-1-yl 2-bromo-2-methylpropionate (1) was
prepared according to literature.28Butylmethacrylate (BMA, 99%,
Sigma-Aldrich) was vacuum-distilled and stored in an air-free flask
in the freezer. Karsted’s catalyst, platinum(0)-1,3-divinyl-1,1,3,3-
tetramethyldisiloxane complex, solution in vinyl-terminated poly-
dimethylsiloxane (Alfa Aesar), N,N-diisopropylethylamine (DIEA,
Aldrich), chlorodimethylsilane (97%, Alfa Aesar), n-butyldimeth-
99.7%, Aldrich), dichloromethane (DCM, Fisher), and hydrogen
according to literature procedure.27BBLMueller-Hinton Broth was
wafers (100 orientation, P/B doped, thickness from 450 to 575 µm
and resistivity from 7.0 to 20.0 ohm cm) were obtained from
International Wafer Service Inc. Glass slides were purchased from
Instruments.1H NMR spectra were recorded on a Bruker DPX-
with a Rudolph Research model SL-II automatic ellipsometer with
an angle of incidence of 70° from the normal. The light source was
a He-Ne laser with λ ) 632.8 nm. Measurements were performed
on 3-5 different locations on each sample. X-ray photoelectron
spectra (XPS) were recorded on a Physical Electronics Quantum
2000 spectrometer with Al KR excitation at a spot size of 100 µm
at 25 W. Spectra were obtained at 15° and 75° takeoff angles with
respect to the plane of the sample surface.
(2). To a dry Schlenk flask were added 1 (8.59 g, 34.5 mmol) and
chlorodimethylsilane (43 mL, 387 mmol) followed by the addition
of Karsted’s catalyst (715 µL). The mixture was stirred at room
temperature under N2. Excess chlorodimethylsilane was removed
under reduced pressure and the product was purified by vacuum
1.68 (quint, 2H, J ) 6.6 Hz, OCH2CH2), 1.93 (s, 6H, 2CH3), 4.17
(t, 2H, J ) 6.6 Hz, OCH2).
Silicon wafers or glass slides were cut into 1.2 × 1.2 cm2pieces,
rinsed with DCM and ethanol, and then dried under a stream of
nitrogen. The samples were then treated with oxygen plasma for 10
were placed in a custom-designed glass holder and immersed in dry
toluene (10 mL) containing DIEA (275 µL, 1.58 mmol) and ATRP
mixed with chlorodimethylbutylsilane at various molar ratios (1%,
10%, 40%, 70%, and 100% initiator, 1.46 mmol total). Samples
with hexanes, ethanol, ethanol-water (1:1), ethanol, then water,
followed by drying under a stream of N2.
CuBr (114.7 mg, 0.8 mmol), CuBr2(17.84 mg, 0.08 mmol), dnbpy
(0.72 g, 1.76 mmol), Boc-AEMA (8.0 g, 34.9 mmol), BMA (3 mL,
18.9 mmol), and anisole (11 mL). The mixture was purged with N2
for 10 min at 80 °C (to dissolve the monomers). The surfaces with
immobilized initiator were then added, and the flask was sealed and
were extracted with DCM and ethanol before drying under a stream
glass surfaces were placed in a flask and covered with hydrogen
chloride solution (4.0 M in dioxane) and left at RT for 12 h. They
were then rinsed with ethanol and dried under a stream of N2.
Bacterial Strains and Microbiological Media. Staphylococcus
in 30% (v/v) glycerol solution at -80 °C) were grown overnight in
sterile BBL Mueller-Hinton Broth at 37 °C with rotary agitation at
Antimicrobial Activity Assay. The following studies were
conducted using a modified version of the Japanese Industrial
Standard JIS Z 2801:2000 Antibacterial ProductssTest for Anti-
bacterial ActiVity and Efficacy.1An overnight culture of S. aureus
(∼109cells/mL) was diluted to approximately 106cells/mL. Silicon
wafer test samples were placed in individual Petri plates, and the
in a fume hood using standard chromatography sprayer (VWR
Scientific). Petri plates were immediately covered and transferred
3 min, 50 µL of phosphate buffered saline was added by pipet onto
phosphate buffered saline on the test surface was mixed by pipet to
ensure removal of bacteria from the test surface. An aliquot of this
phosphate buffer was removed, diluted, and spread using sterile
technique on Mueller-Hinton agar plates. Plates were incubated
overnight at 37 °C without agitation, and then, viable bacterial
colonies were counted. Bacterial growth inhibition was determined
as a percentage of the colonies on the control sample. Each test
sample was dried after each spraying to avoid prolonged contact
between bacteria, liquid, and the test surface.
studies were carried out using the antimicrobial surface studies
described above. After the initial spraying, test samples were dried
and recovered in their respective Petri plates and placed on the
benchtop at room temperature. The same samples were tested the
following day using the identical procedure. This was carried out
for 3 or 5 consecutive days.
(25) Huang, J.; Koepsel, R. R.; Murata, H.; Wu, W.; Lee, S. B.; Kowalewski,
T.; Russell, A. J.; Matyjaszewski, K. Langmuir 2008, 24, 6785–6795.
(26) (a) Lienkamp, K.; Madkour, A. E.; Musante, A.; Nelson, C. F.; Nu ¨sslein,
S.; Ivanov, I.; Tew, G. N. Biopolymers 2008, 90, 83–90. (c) Gabriel, G. J.; Tew,
G. N. Org. Biomol. Chem. 2008, 6, 417–423.
(27) Kuroda, K.; DeGrado, W. F. J. Am. Chem. Soc. 2005, 127, 4128–4129.
(28) Theato, P.; Kim, K. J.; Yoon, D. Y. Phys. Chem. Chem. Phys. 2004, 6,
Fast Disinfecting Antimicrobial SurfacesLangmuir, Vol. 25, No. 2, 2009 1061
Fluorescence Microscopy. An Olympus BX51 reflected fluo-
rescence microscope (Optical Analysis Corp. Nashua, NH) with a
100 W mercury lamp (Chin Technical Corp.) was used for
fluorescence studies. A live/dead BacLight kit L-7007 (Invitrogen,
temperature for 15 min before applying to a modified glass surface.
Bacteria were viewed under a green filter (excitation/emission,
420-480 nm/520-800 nm) or a red filter (480-550 nm/590-800
nm). Viable bacterial cells appeared green, whereas cells with
compromised membrane appeared red.
Results and Discussion
Polymer Synthesis and Characterization. With our goal of
from” technique to attach the antimicrobial polymer 3 to silicon
2 that contains a reactive chlorodimethylsilane, able to self-
condense with silica surfaces, and an R-bromoester that can be
used as an ATRP initiator. The chlorodimethylsilane group was
chosen for the formation of a well-defined monolayer of the
ATRP initiator via condensation with the free hydroxyl groups
were not used as they polymerize in the presence of a trace
The initiator 2 was synthesized by reacting 5-hexene-1-ol with
2-bromoisobutyl bromide to form the ester 1, which was
hydrosilylated with chlorodimethylsilane using platinum(0)
as a catalyst. The chlorodimethylsilyl group of the initiator was
reacted with the surface silanol groups of cleaned silicon wafers
or glass in the presence of DIEA in dry toluene under N2. This
resulted in the formation of a well-defined covelantly linked
monolayer containing ATRP surface-bound initiators. Addition
of the methacrylate monomers followed by heating at 80 °C in
the presence of anisole and CuBr/dnbpy generated the desired
polymers. CuBr2 was used to ensure a sufficiently high
concentration of deactivator, enabling better control over the
polymerization by reducing the concentration of active radical
end groups. The surface-bound polymer was treated with 4.0 M
the active, cationically charged surfaces.
with different polymer thicknesses (5-106 nm) by varying the
polymerization time. Figure 1 shows the evolution of the elli-
psometric brush thickness as a function of reaction time. The
time, which is consistent with a loss of active chain ends during
ATRP.31Removing the Boc-protecting group with HCl led to
the formation of the active polymer 3 with an expected decrease
in layer thickness due to the loss of the bulky group. This effect
is not compensated by electrostatic repulsion of the repeat units,
mostly because the ellipsometry measurments were performed
on dry samples. Thus, active surfaces with polymer layer
thicknesses between 3 and 70 nm were obtained (Figure 1).
In order to vary the initiator density on silicon wafer surfaces,
butyldimethylchlorosilane was used as a blocking agent. It was
premixed with 2 in different molar ratios and allowed to react
with the silicon surfaces. All of the surfaces shown in Figure 2
were polymerized in the same reaction vessel to expose them all
to the identical reaction conditions. Figure 2 shows a nonlinear
relationship between film thickness and initiator concentration,
which may be due to a facilitated radical recombination near the
surface early in the polymerization for the samples with high
initiator density.32Nevertheless, films with thicknesses from 3
to 57 nm were obtained.
Removal of the Boc-group and formation of the amine
hydrochloride salt were monitored by XPS. The N 1s XPS
spectrum (Figure 3a) showed a peak at 399.3 eV, which
(29) Fadeev, A. Y.; McCarthy, T. J. Langmuir 1999, 15, 3759–3766.
(30) Fadeev, A. Y.; McCarthy, T. J. Langmuir 2000, 16, 7268–7274.
(31) Ramakrishnan,A.;Dhamodharan,R.;Ru ¨he,J.Macromol.RapidCommun.
2002, 23, 612–616.
(32) Bao, Z.; Bruening, M. L.; Baker, G. L. Macromolecules 2006, 39, 5251–
Scheme 1. Reaction Scheme for the Synthesis of 3 on Surfaces
1062 Langmuir, Vol. 25, No. 2, 2009Madkour et al.
corresponds to the binding energy of Boc-protected amines.33
Figure 3b shows the N 1s XPS spectra for the surface-bound
polymer after deprotection using HCl. The peak has shifted to
401.1 eV corresponding to the protonated amine group. A small
peak remains at 399.0 eV, which may be due to residual Boc
groups or deprotonated amines. It was observed that, after
extensive washing of the deprotected surface with EtOH
at 401.1 eV corresponding to the protonated amine disappeared
and was replaced with a peak at 399.0 eV, most likely the free
amine (see Figure 3c).
Effect of Polymer Chain Length and Initiator Density on
Antimicrobial Activity. As described above, active surfaces
previous literature reports, it was expected that a trend would be
increasing as film thickness was increased. Surprisingly, our
results shows that all surfaces killed S. aureus 100% in less than
5 min, even those with thickness as low as 3 nm (Figure 4).
Similarly, Figure 5 shows that surfaces with initiator densities
nm, were able to fully kill S. aureus regardless of the initiator
density. We expected that a polymer layer thickness of 2 or 3
is estimated to be about ∼30-37 nm thick.34,35However, our
results are in agreement with the findings of Russell and co-
grafted from surfaces with thicknesses as low as 10 nm killed
bacteria cells.16Similarly, Isquith and co-workers showed that
a thin layer of quaternary ammonium compounds on glass had
good antimicrobial properties.14
In order to ensure that the killing ability of the surfaces was
not due to leaching of the active polymer into the solution, the
(33) Strother, T.; Hamers, R. J.; Smith, L. M. Nucleic Acids Res. 2000, 28,
(34) Friedrich, C. L.; Moyles, D.; Beveridge, T. J.; Hancock, R. E. W.
Antimicrob. Agents Chemother. 2000, 44, 2086–2092.
(35) Matias, V. R. F.; Beveridge, T. J. J. Bacteriol. 2006, 188, 1011–1021.
Figure 1. Evolution of the ellipsometric dry brush thickness with time
for the formation of the polymer on silicon wafers before (9) and after
(O) deprotection with HCl.
of the polymer as a function of initiator density.
Figure 3. N 1s XPS spectra of surface-grafted Boc-protected polymer
Fast Disinfecting Antimicrobial SurfacesLangmuir, Vol. 25, No. 2, 2009 1063
surfaces were evaluated using the Kirby-Bauer test.36Treated
and untreated silicon wafers were placed active side down onto
an agar plate previously inoculated with 1 × 109cells/mL of
S. aureus. After 24 h incubation, treated surfaces did not exhibit
a zone of inhibition around the wafer, which indicated the
nonleaching nature of these surfaces (Figure 6).
Serial Subsequent Exposure Studies. All surfaces with
varying polymer thicknesses and initiator densities exhibited
excellent antimicrobial activities, even those with a polymer
layer thickness of 3 nm or an initiator density of 1% (2 nm
thickness). These surfaces were re-examined with successive
dried, replaced in their respective Petri plates, and stored on the
benchtop at room temperature. The same samples were tested
the following day using the identical test procedure with no
cleaning steps between successive sprays in order to examine
days. As shown in Figures 4 and 5, surfaces still exhibited near-
quantitative bactericidal activity upon a second exposure;
however, it was also observed that the third and subsequent
exposures were not able to kill as many bacteria. Gorham and
co-workers observed complete loss of the antimicrobial activity
of polyethylene surfaces with covalently linked monolayers of
quaternary ammonium compounds after the second exposure
with S. aureus.37They concluded that this loss in activity might
be due to the adsorption of organic materials, such as proteins,
on the positively charged membrane surface. If that was the
case, removal of this material would regenerate the surface
activity. We therefore washed the surface thoroughly; however,
or TEA overnight, followed by sonication. The integrity of the
This leads to the assumption that biological components were
not inhibiting the activity, as such strong acids and/or bases
would have been able to remove any cells, lipids, or cellular
debris from the surface, thus allowing viable cells to once again
contact the antimicrobial polymer.38
completely when heated at 80 °C in sealed ampoules for 4 days,
whereas they retained their activity for long times if stored at
low temperature (-20 °C). This suggested that the loss of
antimicrobial activity might be due to chemical modification or
rearrangement of the polymer. Armes and co-workers39showed
that a similar polymer system, poly(2-aminoethyl methacrylate)
hydrochloride, underwent chemical reactions under alkaline
conditions in solution. They proposed several pathways: Under
alkaline conditions, the ammonium salt is deprotonated to give
the free amine which may attack its own ester group intramo-
lecularly to form an amide via a rearrangement. Alternatively,
an interchain or intrachain condensation can occur between the
primary amine and a neighboring carbonyl (see Scheme 2).
According to Ikada and co-workers, poly N-[3(N,N-dimethy-
lamino)propylacrylamide and poly 2-(dimethylamino)ethyl meth-
and 7.9, respectively.40This shows that the pKavalues of their
values of the corresponding monomers. By analogy, we assume
that the pKavalue of our grafted polymer will be in the range
(37) McCubbin, P. J.; Forbes, E.; Gow, M. M.; Gorham, S. D. J. Appl. Polym.
Sci. 2006, 100, 381–389.
(38) Nylander, T.; Samoshina, Y.; Lindman, B. AdV. Colloid Interface Sci.
2006, 123-126, 105–123.
(39) He, L.; Read, E. S.; Armes, S. P.; Adams, D. J. Macromolecules 2007,
(40) Uchida, E.; Uyama, Y.; Ikada, Y. Langmuir 1993, 9, 1121–1124.
(41) Geurts, J. M.; Gottgens, C. M.; Van Graefschepe, M. A. I.; Welland,
R. W. A.; Van Es, J. J. G. S.; German, A. L. J. Appl. Polym. Sci. 2001, 80,
1966, 45, 493–496.
layer thickness compared to untreated silicon wafers. The modified
efficacy was seen on subsequent exposures.
density compared to untreated silicon wafers. The modified surfaces
killed 100% of bacteria on the first exposure, while decreasing efficacy
was seen on subsequent exposures.
1064 Langmuir, Vol. 25, No. 2, 2009Madkour et al.
Thus, when surfaces are sprayed with bacterial suspension in
distilled water, part of the polymer will be in the free amine
form, allowing polymer rearrangement to occur in pathways
This rearrangement is presumably faster on the surface than in
solution because of the close proximity of the polymer chains
on the surface. To further test this assumption, silicon wafer
surfaces with active polymer in the ammonium chloride form
were covered with PBS buffer (pH 7.4) for 3 days, washed with
If the surface was just deprotonated in the buffer, treatment with
7) showed that the peak at 401.1 eV, corresponding to the
protonated amine nitrogen was not present, but a new peak at
399.5 eV was. This broad peak is consistent with the presence
of amide nitrogens. Antimicrobial assays showed that samples
treated the same way only retained 60% of their antimicrobial
activity, not 100%. These results support the hypothesis that
chemical rearrangement rather than contamination was respon-
sible for the loss of the surface activity.
polymer kills bacteria by contact on the surface, we used the
live/dead two-color fluorescence method. In this viability assay,
a mixture of SYTO9 green fluorescent nucleic acid stain and
propidium iodide, a red fluorescent nucleic acid stain, was used.
The SYTO9 stain labels bacteria with both intact and compro-
bacteria with compromised membranes. When both dyes are
present, propidium iodide competes with the SYTO9 stain for
nucleic acid binding sites.
The dye mixture was incubated with the bacteria (108cell/
mL) for 15 min before applying to plain glass and polymer-
that all bacteria had intact cell membranes. On the other hand,
Figure 8e,f shows that almost all bacterial cells turned red after
2 min of exposure to the modified surface. It is seen that the
iodide (Figure 8b,c). Similar results were observed when E. coli
was used instead of S. aureus (Figure 9). The very fast killing
effect observed using fluorescent microscopy corroborates the
results in Figures 4 and 5, which showed that these surfaces kill
100% S. aureus within only 5 min exposure.
Silicon wafers and glass surfaces were functionalized with
polymer 3 using “grafting from” techniques. ATRP allowed for
control of the polymer layer thickness. Surfaces with varying
grafting density and polymer thicknesses from 2 to 70 nm were
prepared. This polymer is known to be antimicrobial in solution
with molecular weights as low as 1.3 kDa.27The mechanism of
Figure 7. N 1s XPS spectra of surface-grafted polymer 3 (a), after
treatment with PBS buffer (pH 7.4), followed by 4.0 M HCl/dioxane
Scheme 2. Possible Rearrangement Mechanisms for Polymer 3 in PBS Buffer (pH 7.4)
Fast Disinfecting Antimicrobial SurfacesLangmuir, Vol. 25, No. 2, 2009 1065
of the polymer into the bacterial phospholipid membrane. Our
These surfaces were able to kill S. aureus 100% in 5 min. A
high-density polymer layer thickness of 3 nm would not have
are independent. Alternatively, even though the cell envelope is
disruption occurs within 2 min but do not indicate how this
membrane and the electrostatic compensation of the negative
Figure 8. Fluorescence microscopy images of S. aureus: (a,d) on unmodified glass surfaces after 10 min exposure using green and red filters,
respectively, and (b,e) on polymer-modified glass surfaces after 2 min exposure using green and red filters, respectively.
and (b,d) on polymer-modified glass surfaces after 2 min exposure using green and red filters, respectively.
1066 Langmuir, Vol. 25, No. 2, 2009Madkour et al.
charge of the phospholipid membrane by the surface cationic Download full-text
charge.18Regardless of mechanism, surfaces modified with
SMAMPs led to a rapid cell death and, importantly, to interact
cells as opposed to significant lysis. The lack of killing in serial
bacterial loading experiments is related to the chemical rear-
rangement of 3, and alternative polymers that eliminate these
side reactions are easily envisaged. Surfaces that do not support
bacterial growth will be more important as “super-bugs” like
MRSA become more common. This paper, and the other work
in this area, will be critical for meeting this challenge.
Acknowledgment. We thank Jacob Hirsch for his help with
the ONR (N00014-07-1-0520 P00002), and PolyMedix Inc. are
greatly acknowledged for financial support.
Fast Disinfecting Antimicrobial SurfacesLangmuir, Vol. 25, No. 2, 2009 1067