ARTICLE pubs.acs.org/Langmuir
Antibacterial Activity of Conjugated Polyelectrolytes with Variable Chain Lengths Eunkyung Ji,† Anand Parthasarathy,‡ Thomas S. Corbitt,† Kirk S. Schanze,*,‡ and David G. Whitten*,† †
Department of Chemical and Nuclear Engineering, Center for Biomedical Engineering, University of New Mexico, Albuquerque, New Mexico 87131-1341, United States ‡ Department of Chemistry, University of Florida, Gainesville, Florida 32611-7200, United States
bS Supporting Information ABSTRACT: Cationic poly(phenylene ethynylene)- (PPE-) based conjugated polyelectrolytes (CPEs) with six different chain lengths ranging in degree of polymerization from ∼7 to ∼49 were synthesized from organic-soluble precursor polymers. The molecular weight of the precursor polymers was controlled by the amount of a monofunctional “end-capping” agent added to the polymerization reaction. Cationic CPEs were prepared by quaternization of amine groups to tetraalkylammonium groups. Their structureproperty relationships were investigated by observing their photophysical properties and antibacterial activity. The polymers were found to exhibit a chain-length dependence in their photophysical properties. It has also been observed that the polymers exhibit effective antibacterial activity against both Gram-positive and Gram-negative bacteria under UV irradiation, whereas they show little antibacterial activity in the dark. An effect of chain length on the light-activated antibacterial activity was also found: The shortest polymer (n = 7) exhibited the most effective antibacterial activity against both Gram-positive and Gram-negative bacteria.
’ INTRODUCTION Among various classes of antibacterial polymers, cationic conjugated polyelectrolytes (CPEs) are of great interest because of their long-lasting and effective antimicrobial activity.16 The cationic pendant groups are thought to be responsible for initiating antibacterial activity through their attachment to or penetration of cell membranes.2,3,7,8 We recently reported the novel light-induced antibacterial activity of CPEs containing tetraalkylammonium pendant groups.1,9,10 In the light-induced antibacterial activity, generation of singlet oxygen is considered to play a crucial role; the singlet oxygen or subsequently produced reactive oxygen intermediates interact with the bacteria causing bacterial death.1,1115 We also found that a CPE containing a thiophene substituted for one of phenyl rings exhibits highly effective dark antibacterial activity.9 This has been attributed to the high lipophilicity of the thiophene polymer and possibly more accessible quaternary ammonium groups. Ikeda and coworkers reported that the antimicrobial activities of poly(methyl acrylate) and poly(tributyl 4-vinylbenzyl phosphonium chloride) against Staphylococcus aureus increase with increasing molecular weight lower than 5 104 Da and range from 1.6 104 to 9.4 104 Da, respectively.2,1618 From the observation of variable effects of structure and molecular weight, it is clear that there is a need to explore the structureproperty relationships between structures of CPEs and their antibacterial activities. In view of the fact that the properties of CPEs depend strongly on molecular r 2011 American Chemical Society
weight (MW) and polydispersity index (PDI), the molecular weights of antimicrobial polymers could affect their antimicrobial activities and mechanisms of activity.16,17 Recently, we synthesized a series of cationic oligo(p-phenylene ethynylene)s (OPEs) with different chain lengths (one, two, and three repeat units) and examined structurereactivity relationships in their photophysical and antibacterial activities.19,20 This study demonstrated that the shortest oligomer (n = 1) exhibited a relatively high triplet yield, where the triplet yield determines the ability of an OPE to sensitize singlet oxygen. The singlet oxygen yield decreases with increasing chain length, reflecting a decrease in the triplet yield with increasing OPE length. In the dark, longer OPEs have better antibacterial activity against both Gram-negative and Gram-positive bacteria. However, their light-induced antimicrobial activity against Gram-positive bacteria shows two different trends depending on their end groups on each side of backbone. For asymmetric OPE-n series containing hydrogen (H) and carboxylic acid ester (COOC2H5) on each side of backbone as end groups, OPE-3 has higher activity than OPE-1 and OPE-2. For symmetric S-OPE-n(H) series containing hydrogen (H) as an end group, the light-induced biocidal activity is higher than that of the OPE-n series, and S-OPE-1 kills more bacteria than S-OPE-2 Received: May 14, 2011 Revised: July 6, 2011 Published: July 08, 2011 10763
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Scheme 1. Synthetic Scheme for Polymerization
and S-OPE-3. It was also observed that UV irradiation enhances OPE antimicrobial activity. These observations suggest that (1) the formation of triplet state followed by singlet oxygen generation is vital to bacterial killing and (2) antimicrobial activity can be tuned through differences in oligomer structure, including chain length and side groups. To further explore the effects of MW on antibacterial activity, we synthesized polymers featuring longer chains than OPEs. In this article, we describe the preparation of variable-chain-length CPEs with structures similar to those of OPEs. Additionally, we report their MW-dependent photophysical properties and light-activated antibacterial activities. This series of cationic poly(phenylene ethynylene)- (PPE-) based CPEs with variable chain lengths were synthesized according to previously reported procedures.21 The degree of polymerization of the neutral precursor polymers was controlled by the amount of a monofunctional “end-capping” agent added to the polymerization reaction. The synthesis of organic-soluble precursor polymers was followed by quaternization of the amino groups to obtain CPEs substituted with tetraalkylammonium side groups. Subsequently, we investigated the photophysics of the quaternary salts of the CPEs in methanol and water, including the chain-length dependence of absorption and emission spectra and the relative triplet and singlet oxygen yields. Finally, their antimicrobial activity was tested against Gramnegative and Gram-positive bacteria such as Escherichia coli and Staphylococcus epidermidis. These results allowed for an examination of the effect of polymer chain length on antibacterial activity. Interestingly, we found that the polymer of the lowest molecular weight showed the highest antibacterial activity.
’ EXPERIMENTAL SECTION Materials and Synthesis. Monomer 1 and monomer 2 were synthesized according to previously described procedures.21,22 Pd(PPh3)4,
triethylamine, and tetrahydrofuran (THF) were used as received from Sigma-Aldrich. Detailed synthetic procedures for a series of polymers with various chain lengths are included in the Supporting Information. All sample solutions were prepared using water that was distilled and purified on a Millipore purification system. The concentrations of the polymers are provided in polymer-repeat-unit (PRU) concentrations. Photophysical Characterization. Absorption and fluorescence spectra were obtained on a Varian-Cary 100 UVvis absorption dualbeam spectrometer and a spectrofluorometer from Photon Technology International, respectively. Transient absorption spectra were collected using a laser system that is described elsewhere.23 The optical density of the solutions was adjusted to ∼0.7 at the excitation wavelength (355 nm), with the laser energy set at 67 mJ pulse1. Solutions were purged with argon for 45 min before transient absorption spectroscopy measurements were made. Singlet oxygen quantum yields were measured using a Photon Technology International Quantamaster near-IR spectrophotometer equipped with an InGaAs photodiode detector, an optical chopper, and a lock-in amplifier. Biocide Studies, Dead/Live Assays. Escherichia coli and Staphylococcus epidermidis were grown in a nutrient broth (NB, Difco) for 18 h at 37 °C. Then, the bacteria were prepared by centrifuging and washing with 0.85% NaCl three times in a previously described procedure1 and counted in a hemocytometer to normalize the bacterial concentration (∼107 cells/mL) for antibacterial tests. Live control samples containing only bacterial cells and desired concentrations of polymer/bacteria mixture samples were prepared and treated by UV light exposure (UVA lamp centered at ∼350 nm) for 30, 60, and 120 min using a photoreactor from Luzchem. Another set of samples was kept in the dark for 30, 60, and 120 min. Then, dead/live assays were conducted using propidium iodide (red fluorescence = dead) and SYTO9 (green fluorescence = live) purchased from Invitrogen. After the bacteria were incubated with the polymers under light and dark conditions, the dyes (2.4 μL) were added in a 1:1 ratio to the samples, which were kept in the dark for 15 min. Finally, the bacteria were examined using a Zeiss LSM 510 Meta confocal laser scanning microscope (40 oil objective) and an 10764
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Table 1. 1H NMR End-Group Analysis and Pulsed-Gradient Spin-Echo (PGSE) NMR Data for Neutral Precursor Polymers PGSE NMR spectroscopy end-cap content (mol %)
DP by end-group analysis
Mn
PPE-NMe2-7-COOEt
55
7
4210
9
PPE-NMe2-9-COOEt
45
9
5140
12
PPE-NMe2-11-COOEt
35
11
6880
16
PPE-NMe2-14-COOEt
25
14
10970
26
PPE-NMe2-20-COOEt
15
20
15950
39
PPE-NMe2-49-COOEt
10
49
17650
43
Accuri C6 flow cytometer, in which 5 105 cells were analyzed from a 107-cell solution. The numbers of live and dead cells, corresponding to green and red fluorescence, respectively, were counted and compared.
’ RESULTS AND DISCUSSION Synthesis and Characterization of Neutral Precursor Polymers. The organic-soluble precursor polymers (PPE-NMe2-n-
COOEt, n = degree of polymerization) with variable chain lengths were prepared using the difunctional comonomers 1,4diethynyl benzene (1) and 3,30 -[(2,5-diiodo-1,4-phenylene)-bis(oxy)]bis(N,N-dimethylpropan-1-amine) (2), and their molecular weight was controlled by the amount of a monofunctional end-capping agent, ethyl 4-iodobenzoate, in the polymerization reaction mixtures (Scheme 1). Polymerization reactions were conducted under the same conditions in a THF/Et3N (3:2, v/v) mixture with constant concentration (25 mM) of difunctional monomers to minimize the other effects on the chain length.21 All polymers were characterized by 1H NMR spectroscopy to confirm structures and purities. NMR end-group analysis was used to evaluate the degree of polymerization (DP).21 As shown in Figure S1 (Supporting Information), the 1H NMR spectrum of PPE-NMe2-n-COOEt exhibits signals at 8.03 and 7.05 ppm corresponding to the aromatic protons on the ethyl 4-iodobenzoate end group and 2,5-disubstituted-1,4-phenylene repeat units, respectively. The DP values of the series of polymers were calculated by comparing the integration at 8.03 ppm to the integration at 7.05 ppm for each sample. The number-average molecular weights (Mn) and DP values were also determined by the pulsed-gradient spin-echo (PGSE) NMR technique (using polystyrene standards).24 The estimated DP values of the polymers are listed in Table 1. The molecular weight of PPENMe2-7-COOEt was also characterized by gel permeation chromatography (using polystyrene standards), which afforded Mn = 5740 g mol1 and Mw = 7130 g mol1 (where Mw is the weightaverage molecular weight), giving PDI = 1.24. The calculated DP values obtained by NMR end-group analysis and PGSE NMR spectroscopy differ by a factor 1.32.0. These observations are consistent with previous studies demonstrating that the numberaverage molecular weights of rigid rod polymers such as PPENMe2-n-COOEt are overestimated by molecular weight analysis using polystyrene standards.21,25,26 After the characterization of the precursor polymers, water-soluble CPEs (PPE-NMe3-n-COOEt) were prepared according to a literature procedure (Scheme 1).22 The resulting polyelectrolytes were characterized by 1H NMR spectroscopy in CD3OD, confirming the quaternization by chemical shifts of the signals in the aliphatic regions (OCH2 CH2CH2N) and N(CH3)2 of the organic-soluble precursor polymers.
DP
Photophysical Characterization. Figure 1 shows the absorption and fluorescence spectra (normalized according to the fluorescence quantum yield) of six different chain lengths of neutral precursor polymers in chloroform solution. The absorption maximum red shifts from 412 nm (PPE-NMe2-7-COOEt) to 417 nm (PPE-NMe2-20-COOEt) and 420 nm (PPE-NMe249-COOEt) as a result of the increase in average conjugation length.21 Generally, the molar absorption coefficient (per polymer repeat unit) increases with DP except for PPE-NMe2-49-COOEt (Table 2). The exceptional trend for PPE-NMe2-49-COOEt is possibly explained by the onset of polymer aggregation causing a red-shifted absorption maximum band.21 As shown Figure 1b, all CPEs exhibited the same fluorescence spectra with a constant band maximum at 447 nm. The fluorescence quantum yields of the polymers are listed in Table 2. The absorption and fluorescence spectra of the set of six quaternized polyelectrolytes in methanol and water were also obtained, and the results are presented in Figure 2 and Table 3. With increasing DP, the absorption maximum red shifts by 3 nm in methanol and 8 nm in water. This small increase in absorption maximum with increasing DP can be explained by our previously reported molecular simulations. In particular, density functional theory calculations of oligomers (with n = 1, 2, and 3 repeat units) featuring the same repeat-unit structure as the quaternized polymers (PPE-NMe3-n-COOEt) indicates that the limit of conjugation length is approached for a repeat unit consisting of three phenylene ethynylene repeat units.20 Li et al. also demonstrated that the absorption maximum of PPE-based polymers mostly depends on the conjugation length of the polymers and not directly on total polymer chain length.28 For the fluorescence spectra, the λmax value is constant in both solvents; however, in water, the intensity of the shoulder band at longer wavelength (∼500 nm) increases with increasing chain length, giving rise to broad spectra likely due to the effects of polymer aggregation. Because the triplet excited state of CPEs plays a crucial role in their generation of singlet oxygen and subsequent light-activated antibacterial activity,1 we examined the transient absorption of the PPE-NMe3-n-COOEt series in methanol and water (Figure 3 and Table 3) to detect the triplet state by triplettriplet absorption. As previously shown for other CPEs,1,9,29,30 the transient absorption spectra of the PPE-NMe3-n-COOEt series exhibit a broad intense band centered around 760 nm in both methanol and water solutions (Figure 3 and Supporting Information) that is assigned to the triplettriplet absorption. The triplet lifetime ranged from 1.6 to 2.8 μs in methanol solution and from 5.8 to 16.9 μs in water solution for the PPE-NMe3-nCOOEt series. Although absolute triplet yields were not measured, we use the initial transient absorption amplitude (ΔAt=0, Table 3) as a measure of the relative triplet yield. Another 10765
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Figure 1. (a) Absorption and (b) fluorescence spectra of PPE-NMe2-n-COOEt in chloroform. Polymer concentration (polymer repeat units) = 1 μM, and the fluorescence spectra were obtained with excitation at 410 nm. Emission spectra are normalized according to the fluorescence quantum yield.
Table 2. Photophysical Properties of Neutral Precursor Polymers in Chloroform n
λabs max (nm)
εmax (104) (M1 cm1)b
λflmax (nm)
Φfla,b
7
412
2.9
447
0.32
9
412
6.0
447
0.25
11
412
6.9
447
0.27
14
412
6.4
447
0.31
20
417
12.9
447
0.27
49
420
11.4
447
0.22
Coumarin 30 in MeOH as standard, Φfl = 0.307 (ref 27). b Errors are less than 5%. a
important point to note is that all of the polymers showed lower amplitude for triplet absorption and longer triplet lifetime in water compared to methanol, which is in line with earlier reports.24,3133 The above observations in water solution are consistent with the diminished fluorescence yield of these polymers, which signals enhanced nonradiative decay by internal conversion in this medium, which, in turn, reduces the triplet yield. In methanol, the amplitude of the transient absorption decreased with increasing DP, implying that the triplet yield decreases with increasing chain length. The same trend was observed in water solution except for PPE-NMe3-49-COOEt. Taken together, the transient absorption studies reveal that photoexcitation of each member of the PPE-NMe3-n-COOEt series generates the triplet state with moderate efficiency. By comparing the initial amplitude of the transient absorption immediately following the laser pulse (ΔAt=0, Table 3), it can be concluded that the relative triplet yield was the highest for the shortest member (n = 7) of the series. The trend of decreasing triplet yield with increased DP is consistent with our earlier investigation of a series of monodisperse oligomers in which the same trend was seen.19 Having confirmed the formation of the triplet excited state of these polymers by transient absorption spectroscopy, we set out to determine their ability to sensitize the formation of singlet oxygen in CD3OD solution by using the emission of 1O2 at 1260 nm as a probe. (Deuterated methanol was selected for this study, as the emission yield of 1O2 is strongly attenuated in water or D2O as a result of solvent quenching.) All members of the polymer series are capable of producing singlet oxygen, as evidenced
by its emission at 1260 nm, and the singlet oxygen yields are listed in Table 3. In general, a slight decrease (∼20%) in the singlet oxygen yields was observed with increasing DP, which is consistent with a decrease in the relative triplet yield with increasing chain length. However, again, it is evident that, in each case, the CPEs are capable of producing singlet oxygen in moderate yield. Antibacterial Activity of CPEs. Our previous works have described light-induced antibacterial action of cationic CPEs and OPEs.1,9,10,19 Recently, it was demonstrated that the antibacterial action of OPEs depends on their molecular weight.19 To further explore the role of molecular weight in determining the antibacterial activity, we investigated the light-induced killing ability of the PPE-NMe3-n-COOEt series against the same bacteria used in our prior study: Gram-negative bacteria E. coli and Grampositive bacteria S. epidermidis. Polymer solutions were incubated with E. coli or S. epidermidis under UV light illumination or in the dark, and the antibacterial effect of the polymers was evaluated at times 0, 30, 60, and 120 min by flow cytometry analysis and confocal microscopy after the cellular suspensions had been stained with live/dead fluorescent dyes (SYTO9, green fluorescence = live; propidium iodide, red fluorescence = dead). Figure 4 shows plots of the percentage of dead bacteria cells versus light exposure time of the PPE-NMe3-n-COOEt series against E. coli at two different concentrations. These plots were derived from flow cytometry analysis of the cellular suspensions after live/dead staining (flow cytometry data are provided in the Supporting Information). Two trends were found: (1) For the 1 μg/mL solution concentration, the polymer with the shortest chain (n = 7) exhibited the strongest antibacterial activity against E. coli under light exposure, and (2) the polymers' antibacterial efficiency increased with increasing irradiation time. As indicated in Figure 4a,b, ∼99% of bacteria were killed by PPE-NMe3-nCOOEt after 120 min of irradiation at both concentrations. It has also been observed that a higher concentration of polymer solution (10 μg/mL) shows an inner filter effect, leading to suppression in the light-induced killing efficacy of the polymers.35 In particular, for the CPEs with DP = 7 and 20, 10 μg/ mL of polymer solutions exhibited less bacterial killing than 1 μg/mL of polymer solutions. This finding can be explained by protection of polymer-coated bacteria through the inner filter effect at the higher concentration of polymer. The polymer/bacteria suspensions were also examined using confocal microscopy after 10766
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Figure 2. (a) Absorption and (b) fluorescence spectra of PPE-NMe3-n-COOEt in water. Polymer concentration (polymer repeat units) = 1 μM, and the fluorescence spectra were obtained after excitation at 400 nm.
Table 3. Photophysical Properties of CPEs in Methanol and Water Solution n
solvent
λabs max (nm)
εmaxa (105) (M1 cm1)
λflmax (nm)
Φfla,b
triplet abs (ΔAt=0)
τtriplet (μs)
ΦΔc
7
MeOH
394
14.0
435
0.33
0.082
2.7
0.120 ((0.02)
9
H2O MeOH
396 397
13.5 6.3
436 435
0.04 0.33
0.022 0.069
8.7 2.8
0.101 ((0.02)
H2O
396
5.4
436
0.04
0.018
5.8
MeOH
397
5.1
435
0.29
0.068
2.1
H2O
397
4.5
436
0.04
0.017
16.9
MeOH
397
5.6
435
0.26
0.048
1.6
H2O
401
5.1
436
0.05
0.015
8.3
20
MeOH
397
10.6
435
0.23
0.046
2.8
0.098 ((0.02)
49
H2O MeOH
403 397
10.0 3.1
436 435
0.04 0.23
0.012 0.046
11.3 2.4
0.099 ((0.02)
H2O
404
2.8
436
0.05
0.019
10.5
11 14
0.122 ((0.02) 0.117 ((0.02)
a Errors are less than 5%. b Coumarin 30 in MeOH as standard, Φfl = 0.307 (ref 27). c Measured in CD3OD using 20 -acetonaphthone (ΦΔ = 0.79) as an actinometer.
Figure 3. Transient absorption difference spectra of PPE-NMe3-nCOOEt (n = 7, OD ≈ 0.7 at 355 nm, excited with a laser energy of ∼7 mJ) in methanol (initial delay =70 ns, subsequent delay increment = 1 μs).
live/dead staining, and the results were in accord with the flow cytometry analysis. A sample where polymer was not added to the bacterial suspension served as a control (bacteria only), and it was found that most cells (>90% as quantified by flow cytometry;
data not shown) were viable under UV light illumination or in the dark after 0, 30, 60, and 120 min, with most of the bacteria appearing green in the confocal fluorescence microscope images (Figure 4c). Consistent with the flow cytometry data, the fluorescence microscopy image of the polymer/bacteria suspension after 120 min of irradiation (Figure 4d) mainly showed red fluorescence indicating dead cells. Antibacterial activity against the Gram-positive bacteria S. epidermidis was investigated by the same procedure. As shown in Figure S9 (Supporting Information), S. epidermidis is more sensitive to the light-activated antibacterial action of the polymers (n = 7 and 49) than Gram-negative bacteria. After 30 min of irradiation, both polymers killed ∼99% of bacteria at two different concentrations of the polymer solution. The observation of a higher antibacterial efficiency against Gram-positive bacteria might be due to the different outer membrane structures for Gram-negative and Gram-positive bacteria. Gram-negative bacteria cell walls contain an additional outer membrane layer consisting of negatively charged lipopolysaccharides (LPS), lipoproteins, phospholipids, and proteins-like porins, allowing a higher resistance to membrane-permeable cationic antimicrobials compared to Gram-positive bacteria.36,37 10767
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Figure 4. Viability of Escherichia coli cells with polymer solution following exposure to UV light and incubation in the dark. Plots of percentages of dead cells with (a) 1 and (b) 10 μg/mL polymer solutions. Fluorescence confocal microcopy images of (c) 0-min control and (d) 120-min light-exposed polymer (n = 7, 1 μg/mL) with E. coli,. Green fluorescent (SYTO 9) cells represent live cells, whereas red fluorescent (propidium iodide) cells represent dead or compromised cells.
Dark antimicrobial activity of the polymers was not observed: The polymers (1 and 10 μg/mL) showed only ∼10% and 20% of dark killing against E. coli and S. epidermidis, respectively, after the incubation of bacteria with the polymers in the dark for 30, 60, and 120 min (Figures S8 and S9, Supporting Information). Therefore, the mechanism of antimicrobial activity of PPENMe3-n-COOEt could be mainly explained with the adsorption and/or penetration of the cationic polymers onto the negatively charged bacterial membrane. This is followed by singlet oxygen generation at the interface between the polymers and the bacteria surfaces or inside the bacterial membrane upon exposure to UV light, leading to the inactivation of bacteria through the oxidization of bacterial proteins or lipids. This mechanism is supported by results of experiments on the polymer chain-length-dependent disruption of the bacterial membrane with different-sized PPE-NMe3-n-COOEt species (n = 7, 20, and 49): (1) There are strong interactions between the polymers and bacterial membrane, leading to dramatic changes in the polymer photophysical properties after addition of E. coli total lipid extract vesicles to the polymer solutions, and (2) the bacterial membrane is physically disrupted by the polymer, giving rise to dye leakage from the vesicles.38 It was also observed that the shortest polymer (n = 7)
exhibited the most significant membrane perturbation ability.38 As mentioned above, the shortest polymer featuring relatively higher transient absorption amplitude also exhibited the most effective antibacterial activity. On the basis of these findings, we conclude that the efficient light-induced antimicrobial action of the polymers occurs through their accumulation in or at the cytoplasmic membrane, which is a crucial target for damage by the singlet oxygen generated upon exposure to light.39
’ CONCLUSIONS We have documented an investigation of the photophysical properties and antimicrobial activity of poly(phenylene ethynylene)-based cationic CPEs with variable chain lengths. The photophysical properties of the series of neutral precursor polymers and cationic polymers were studied. The neutral polymer solutions in chloroform were found to exhibit a slightly red-shifted (∼8 nm) absorption band and increased molar absorption coefficient with increasing polymer chain length. The quaternized polymer solutions in methanol and water also showed a small increase (3 and 8 nm, respectively) in the absorption maximum with increasing chain length. These small 10768
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changes in the maximum absorption support our previous finding that the limiting chromophore is a segment consisting of three phenylene ethylene repeat units.20 The molecular-weight-dependent antimicrobial activity of the cationic polymers was also investigated. The polymers were found to exhibit effective light-induced antimicrobial activity against both Gram-negative and Gram-positive bacteria at low concentrations (1 and 10 μg/mL). The polymer with the shortest chain (n = 7) killed bacteria most effectively because of its highest triplet yield, as observed for the amplitude (ΔAt=0) of the transient absorption, and most efficient membrane perturbation ability, as described in a companion article.38
’ ASSOCIATED CONTENT
bS
1
H NMR spectrum of PPENMe2-n-COOEt, fluorescence lifetime data of PPE-NMe3-nCOOEt in methanol and water, transient absorption spectra of PPE-NMe3-n-COOEt in methanol and water, flow cytometry data of antibacterial activity of PPE-NMe3-n-COOEt, and details of polymer synthesis. This material is available free of charge via the Internet at http://pubs.acs.org. Supporting Information.
’ AUTHOR INFORMATION Corresponding Author
*E-mail:
[email protected] (D.G.W.),
[email protected]fl.edu (K.S.S.). Tel.: 505-277-5736 (D.G.W.), 352-392-9133 (K.S.S.). Fax: 505-277-1292 (D.G.W.), 352-392-2395 (K.S.S.).
’ ACKNOWLEDGMENT This research was supported by the Defense Threat Reduction Agency (Contract W911NF-07-1-0079). Confocal images were obtained using the confocal laser scanning microscope housed in the W. M. Keck Confocal Microscopy Facility of the UNM Keck Nanofluidics Laboratory. ’ REFERENCES (1) Chemburu, S.; Corbitt, T. S.; Ista, L. K.; Ji, E.; Fulghum, J.; Lopez, G. P.; Ogawa, K.; Schanze, K. S.; Whitten, D. G. Langmuir 2008, 24, 11053. (2) Ikeda, T.; Yamaguchi, H.; Tazuke, S. Antimicrob. Agents Chemother. 1984, 26, 139. (3) Kugler, R.; Bouloussa, O.; Rondelez, F. Microbiology 2005, 151, 1341. (4) Thorsteinsson, T.; Masson, M.; Kristinsson, K. G.; Hjalmarsdottir, M. A.; Hilmarsson, H.; Loftsson, T. J. Med. Chem. 2003, 46, 4173. (5) Bromberg, L.; Hatton, T. A. Polymer 2007, 48, 7490. (6) Lin, J.; Tiller, J. C.; Lee, S. B.; Lewis, K.; Klibanov, A. M. Biotechnol. Lett. 2002, 24, 801. (7) Kwon, D. H.; Lu, C. D. Antimicrob. Agents Chemother. 2006, 50, 1623. (8) Johnston, M. D.; Hanlon, G. W.; Denyer, S. P.; Lambert, R. J. W. J. Appl. Microbiol. 2003, 94, 1015. (9) Corbitt, T. S.; Ding, L. P.; Ji, E. Y.; Ista, L. K.; Ogawa, K.; Lopez, G. P.; Schanze, K. S.; Whitten, D. G. Photochem. Photobiol. Sci. 2009, 8, 998. (10) Corbitt, T. S.; Sommer, J. R.; Chemburu, S.; Ogawa, K.; Ista, L. K.; Lopez, G. P.; Whitten, D. G.; Schanze, K. S. ACS Appl. Mater. Interfaces 2009, 1, 48. (11) Halliwell, B.; Gutteridge, J. M. C. Lancet 1984, 1, 1396. (12) Foote, C. S. Photochem. Photobiol. 1991, 54, 659.
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dx.doi.org/10.1021/la2018192 |Langmuir 2011, 27, 10763–10769
