Porphyrin-Based Honeycomb Films and Their Antibacterial Activity

May 20, 2014 - porphine chloride (an acid form, {MnTPPS}) and dimethyldioctadecylammonium bromide. (DODMABr). The morphology of the microporous thin ...
0 downloads 0 Views 8MB Size
Download PDF
Article pubs.acs.org/Langmuir

Porphyrin-Based Honeycomb Films and Their Antibacterial Activity Yanran Wang, Yan Liu, Guihua Li, and Jingcheng Hao* Key Laboratory of Colloid and Interface Chemistry and Key Laboratory of Special Aggregated Materials, Shandong University, Ministry of Education, Jinan 250100, China S Supporting Information *

ABSTRACT: Micrometer-sized porous honeycomb-patterned thin films based on hybrid complexes formed via electrostatic interaction between Mn(III) meso-tetra(4-sulfonatophenyl) porphine chloride (an acid form, {MnTPPS}) and dimethyldioctadecylammonium bromide (DODMABr). The morphology of the microporous thin films can be well regulated by controlling the concentration of MnTPPS-DODMA complexes, DODMABr, and polystyrene (PS), respectively. The formation of the microporous thin films was largely influenced by different solvents. The well-ordered microporous films of MnTPPS-DODMA complexes exhibit a more efficient antibacterial activity under visible light than those of hybrid complexes of nanoparticles modified with DODMABr, implying that well-ordered microporous films containing porphyrin composition can improve photochemical activity and more dominance in applications in biological medicine fields.

1. INTRODUCTION The surface properties of functional materials are related to their chemical compositions and topologies. Microporous films with ordered pore-structures are of significant interest because of their great potential as functional materials such as photonic band gap materials,1 cell culture scaffolds,2 superhydrophobic surfaces,3 surface-enhanced Raman spectroscopy substrates,4 size-selective separation,5 etc. Micropatterned or honeycomb film materials can be formed by various methods. One of the simplest and the most promising methods is to utilize breath figures, which the microporous patterns can be fabricated through self-assembly using water droplets as templates. The exact physical mechanism of self-assembly of breath figures is still under investigation.6,7 Although different mechanisms for the formation of highly ordered microporous structures have been proposed,6 basically rapid evaporation of the water immiscible, volatile solvent such as chloroform, carbon disulfide, and toluene decreases the temperature at the solution/humid air interface below its dew point, resulting in intensive condensation of water droplets on the surface of the solution. The droplets then sink into the solution. Because of the presence of the water-resistance polymer, the water droplets do not coalesce; instead, they rearrange into ordered hexagonal arrays driven by thermocapillary forces through Rayleigh or Marangoni convection.6,7 Upon further evaporation, the condensed water droplets are turned into regular air pores, and the honeycomb-pattern films can be formed consequently. Since François et al.8 first prepared honeycomb films of starpolymer polystyrene from CS2 solution in 1994, a variety of materials such as polymers,9,10 amphiphilic polyion complexes,11 organic/inorganic hybrids,12 surface modified nanoparticles,13 and surfactant-encapsulated polyoxometalates14,15 have been successfully employed for fabricating ordered honeycomb films. Of particular interest is the incorporation © 2014 American Chemical Society

of biomaterials, which may have potential applications in medical and nonmedical uses, such as DNA,16 peptides,17 cellulose,18 porphyrin,19,20 etc. Porphyrins are attractive functional biomaterials, which widely exist in nature. Porphyrins are commonly used as photosensitizers to produce reactive oxygen species, in particular, singlet oxygen, which causes photodynamic inactivation (PDI) of microorganisms upon exposing to visual light in the presence of oxygen.21 Many studies have reported the immobilization of porphyrins on polymeric supports22−26 to allow the reutilization of the porphyrins and avoid the effect of residual photosensitizer. To the best of our knowledge, fewer reports have studied the photodynamics of porphyrin films with ordered-pore structures. Due to the larger surface area and unique microporous patterns, the honeycomb films can provide better photoelectric properties1 and photochemical activities.27 In this work, a hydrophilic porphyrin, MnTPPS, is facilely used to construct into microporous honeycomb films (MHFs) on solid substrates via casting organic solution of MnTPPSDODMA complexes at high relative humidity. The morphology of MHFs was characterized by optical microscope (OM), scanning electron microscope (SEM), and atomic force microscope (AFM). The pore size can be achieved by controlling the concentration of the MnTPPS-DODMA complexes, polystyrene and surfactant. The solvents were selected to construct MHFs which can control the formation of the micropatterns. Photodynamics of the porphyrin-based honeycomb films was preliminarily examined in cellular suspensions of microorganisms, showing high inhibitory activities. The exciting results of photoinduced killing microReceived: December 7, 2013 Revised: May 19, 2014 Published: May 20, 2014 6419

dx.doi.org/10.1021/la501244s | Langmuir 2014, 30, 6419−6426

Langmuir

Article

Scheme 1. Chemical Structures of MnTPPS (Left) and DODMABr (Right)

organisms of ordered porous porphyrin films provide potential applications in the treatment of wasterwater containing bacterials and a new avenue unexplored for ordered porous films.

probe, 9,10-anthracenediyl-bis(methylene)dimalonic acid (ABDA). ABDA is a water-soluble derivative of anthracene that can be photobleached by singlet oxygen to its corresponding endoperoxide. 150 μmol·L−1 ABDA in PBS solution (pH 7.2) containing the films was irradiated in a 1 cm path length spectrofluorimetric cuvette (3 mL). The photobleaching of ABDA was carried out using a CHFXM500 M mercury lamp as light source, in combination with a 420 nm cutoff filter. The reaction was monitored spectrophotometrically by recording the decrease in optical density at 401 nm (λmax of ABDA) in different irradiation periods. 2.5. Antibacterial Activities of Honeycomb-Patterned Films. Escherichia coli strains (E. coli, BL21 (DE3)) were obtained from School of Life Science, Shandong University. Monocolony of E. coli on the solid Luria−Bertani (LB) agar plate was transferred to 5 mL of liquid LB culture medium and grown at 37 °C for 12 h to reach the log phage. Bacteria were harvested by centrifuging (10 000 rpm for 1 min) at 4 °C and washed by phosphate-buffered saline (PBS pH 7.2) four times, and then resuspended in 4 mL of PBS. The E. coli cells were appropriately diluted to obtain ∼105 colony forming units (cfu)/mL in PBS. A total of 100 μL of bacterial solution was dropped onto the porphyrin-based honeycomb thin films, and the samples were covered with a cover glass to prevent media evaporation. The samples were incubated for 60 min in the dark at 37 °C or illuminated with a 100 W halogen bulb for 60 min. In all cases, the controlling experiments were carried out in the presence of MnTPPS-DODMA nonporous thin films or in the absence of MnTPPS-DODMA films with cellular suspensions irradiated and in the dark. The treated films were washed with PBS to collect the bacterial cells on the films and the bacterial suspension was diluted to ∼103 cfu/mL in PBS. A 100 μL portion of the diluted bacterial E. coli was spread on the solid LB agar plate and the colonies formed after 12−16 h incubating at 37 °C were counted. The cell reduction was determined by dividing the number of cfu of the samples incubated with MnTPPS-DODMA honeycomb films by the number of cfu of the controlling that was carried out in the absence of MnTPPS-DODMA films in the dark. All data were presented as the mean standard deviation of each group. Variation among groups was evaluated using the Student’s ttest, with a confidence level of 95% (p < 0.05) considered statistically significant.

2. EXPERIMENTAL SECTION 2.1. Chemicals and Materials. Mn(III) meso-tetra(4-sulfonatophenyl) porphine chloride (an acid form, MnTPPS) was purchased from Frontier Scientific, Inc. Dimethyldioctadecyl-ammonium bromide (DODMABr) was purchased from J&K Chemical. Polystyrene (PS, Mw ≈ 250 k) was purchased from ACROS. 9,10-anthracenediylbis(methylene) dimalonic acid (ABDA) was purchased from SigmaAldrich. Organic solvents including chloroform, carbon disulfide, and tetrahydrofuran are analytical reagents and purchased from China Sinopharm Chemical Reagent co., Ltd. Water used in all experiments was treated in a three-stage Millipore Milli-Q plus 185 purification system and has a resistivity of 18.2 MΩ cm. All the compounds were used as received without further purification. 2.2. Fabrication of Honeycomb-Patterned Films. MnTPPS can dissolve in water to dissociate 4 H+ and 1 Cl− in aqueous solution. A total of 2.0 mL of 0.5 mg·mL−1 MnTPPS aqueous solution was mixed with a 2.0 mL of 1.5 mg·mL−1 chloroform solution of DODMABr, which ensures that their molar ratio between MnTPPS and DODMABr is 1:5. The hydrophobic MnTPPS(DODMA)4 complexes can form according to chemical formula, as shown in Scheme 1, via electrostatic interaction. The phase transition of MnTPPS from aqueous solution to the CHCl3 can facilely occur. Typically, the microporous thin films were prepared by directly casting 5−10 μL chloroform solution of MnTPPS(DODMA)4 complexes onto the glass substrates under a moist airflow. The glass surfaces were cleaned by immersing in ultrasonically agitated solvents (acetone, alcohol, and H2O) for 30 min at 50 °C and then rinsed with deionized water. The glass surfaces were then dried with N2 for the further preparation of porous films of MnTPPS(DODMA)4 complexes. The high humid conditions were obtained by perpendicularly blowing nitrogen gas (flow rate of 1 L·min−1) saturated with water vapor (T = 20 °C) to the upward side of the solution MnTPPS(DODMA)4 complexes. The relative humidity of the moist airflow is 85% which was confirmed by a hygrometer. After complete evaporation within 10 s, the thin films with strong bright iridescent colors under reflected light were obtained. The controlling experiments in the absence of humid airflow were conducted under ambient atmosphere (relative humidity: 20−30%), leaving only unpatterned nonporous thin films. 2.3. Characterizations. The morphology of the honeycomb films was characterized by optical microscope (Zeiss, Axioskop 40/40 FL), field-emission scanning electron microscopy (FE-SEM, JEOL JSM6700F), and atomic forcemicroscopy (AFM, Digital Instruments, NanoScope IIIa, tapping mode). Fourier transform infrared (FT-IR) spectra were measured on a VERTEX-70 (Bruker) FT-IR spectrometer. UV−vis spectra were obtained on a Hitachi U4100 Spectrometer. 2.4. Detection of Singlet Oxygen Generation. Detection of singlet oxygen was determined by photobleaching of the chemical

3. RESULTS AND DISCUSSION Typically, MnTPPS-DODMA complexes can be prepared by mixing 1.5 mg·mL−1 DODMABr chloroform solution and 0.5 mg·mL−1 MnTPPS aqueous solution at room temperature. Due to the electrostatic interaction, a phase transition of the hydrophilic MnTPPS from aqueous solution to chloroform phase occurs. MnTPPS-DODMA complexes are no longer soluble in water, implying that the hydrophobic surfaces of MnTPPS encapsulated by hydrophobic cationic double DODMA+ form. The formation of MnTPPS-DODMA complexes can also be proved from the FT-IR spectroscopy data of MnTPPS, DODMABr, and MnTPPS-DODMA complexes (Figure 1), the IR spectrum of MnTPPS-DODMA 6420

dx.doi.org/10.1021/la501244s | Langmuir 2014, 30, 6419−6426

Langmuir

Article

of DODMAB does not affect the integraty of the porphyrin ring and its electronic transition. The bands of microporous honeycomb films of MnTPPS-DODMA complexes and nonporous thin films are broader. Both absorption bands are red-shifted relative to those of the corresponding monomolecular porphyrin in methanol solution, and the red-shifts of the nonporous thin films are larger than that of microporous honeycomb films. According to Kasha’s exciton theory,29−32 a stacked face-to-face π-aggregation (sandwich-type H-aggregate) leads to a spectral blue shift relative to the monomer excitedstate level, whereas the corresponding spectral feature from a tilted “deck of cards” (J-aggregate) aggregation leads to a red shift. The red-shifted absorption peaks of the honeycomb films of MnTPPS-DODMA complexes signify that the assembly consists of the “deck of cards” aggregates (J-aggregates). During the formation of microporous honeycomb films, the MnTPPSDODMA complexes can deposit on the walls, and the repulsion force of the cooled water droplet can make the walls squeezed.33 We speculate that the different arrangements of MnTPPS-DODMA complexes on different honeycomb films can lead to slightly different red shift. The microporous honeycomb films can be produced by casting chloroform solution of MnTPPS-DODMA complexes onto glass substrates under a moist airflow, relative humidity is above 80% as determined by a hygrometer, applied to the solution surface at 20 °C. After complete evaporation of the solvent and the condensed water droplets, porous thin films with strong bright iridescent colors were obtained when viewed with reflected light, indicating a periodic refractive index of variation throughout the thickness of the films (Supporting Information (SI), Figure S1). Figure 3 displays representative

Figure 1. FT-IR spectra of MnTPPS (a), DODMAB (b), and MnTPPS-DODMA complexes (c) in solid state (KBr).

complexes shows the characteristic vibration absorptions of MnTPPS and DODMAB. Furthermore, the frequencies of the CH2 antisymmetric [νas (CH2)] and the symmetric stretching [νs (CH2)] bands are strong indicators of the presence of alkyl chains. The CH2 antisymmetric stretching band was observed between 2915 and 2918 cm−1 and the symmetric stretch band was observed between 2846 and 2850 cm−1. These data indicate that the hydrocarbon chains are highly ordered arranged.28 The absorption bands at 2918 cm−1 [νas (CH2)] and 2850 cm−1 [νs (CH2)] of the MnTPPS-DODMA complexes suggest that the hydrophobic cationic double DODMA+ are uniformly oriented on the surface of MnTPPS. These factors are important for organizing honeycomb films. The UV−vis spectra of the honeycomb films and porphyrins in solution are shown in Figure 2. The maximum absorption

Figure 2. UV−vis absorption spectra of MnTPPS in methanol (a), MnTPPS-DODMA complexes in methanol (b), honeycomb films of MnTPPS-DODMA complexes (c), and nonporous thin films of MnTPPS-DODMA complexes (d).

bands were summarized in Table 1. The MnTPPS-DODMA complexes in methanol show similar Soret band to the monomeric porphyrin in methanol, meaning that the addition Figure 3. Images of self-assembled honeycomb films of MnTPPSDODMA complexes. OM (a), SEM (b), and AFM (c) of the depth curve of well along the line and a 3-D image (d).

Table 1. Maximum Absorption Peaks of Porphyrins, MnTPPS-DODMA in Solution, the Honeycomb Films, and Nonporous Thin Films Soret band, λmax(nm)

Q-band, λmax(nm)

solution methanol

466 466

562 595 562 595

honeycomb film nonporous film

477 483

573 610 578 621

system MnTPPS methanol MnTPPS-DODMA solution MnTPPS-DODMA MnTPPS-DODMA

OM, SEM and AFM images of the microporous films from 2 mg·mL−1 MnTPPS-DODMA complexes in chloroform on surface of glass substrate. Optical micrograph (Figure 3a) reveals a relatively wide area of regular honeycomb films. A FESEM image, as shown in Figure 3b, confirms that the ordered morphologies of the self-patterned honeycomb films having regular hexagonal arrays with minimum energy state have been 6421

dx.doi.org/10.1021/la501244s | Langmuir 2014, 30, 6419−6426

Langmuir

Article

Figure 4. SEM images of honeycomb-patterned films of MnTPPS-DODMA complexes prepared at different concentrations: 1.0 (a), 1.3 (b), 1.8 (c), 2.4 (d), 3.2 (e), and 4.2 mg·mL−1 (f). The inset curve in panel f is the relationship between the pore diameter and the concentration of MnTPPSDODMA complexes.

Figure 5. SEM images of honeycomb-patterned films of MnTPPS-DODMA complexes prepared by casting 0.5 mg·mL−1 MnTPPS with different DODMAB concentrations. The ratio between MnTPPS and DODMA is 1:2 (a), 1:5 (b), and 1:10 (c).

concentration was between 1.8 and 2.4 mg·mL−1. Disordered honeycomb films were obtained when the concentration was below 1.0 mg·mL−1 or above 4.2 mg·mL−1. The concentration of free DODMAB in chloroform is also an important factor on the morphology of honeycomb films (Figure 5). MnTPPS has four sulfuric acid groups which can have four negative charges, while DODMA+ has a positive charge. When the molar ratio (r) of MnTPPS: DODMA+ ≈ 1:2, the pore sizes are not uniform and almost no ordered hexagonal arranged pores were formed (Figure 5a). It is likely that MnTPPS ions were not completely encapsulated by DODMA+ and the water droplets as template of pore fromation were not able to be well stabilized. When appropriate concentration of DODMAB (cDODMA = 1.5 mg·mL−1) and a molar ratio (r) of MnTPPS:DODMA+ ≈ 1:5 with a little extra DODMA+ were used, MnTPPS-DODMA complexes were able to self-assemble into ordered microporous honeycomb films (Figure 5b). When the concentration of DODMAB was higher and the molar ratio (r) of MnTPPS:DODMA ≈ 1:10, the walls of pores became cracked. Too many free surfactants are present, i.e., at much higher DODMAB concentration in chloroform, which can affect the stability of water droplets and lead to the destruction of the films (Figure 5c). The choice of the solvent is also a decisive factor on the formation of microporous honeycomb films. There are some requirements for selecting an appropriate solvent,35 e.g., high vapor pressure, low boiling point, higher density than water and

reported by Shimomura at al.34 The spherical pores have an average diameter of about 800 nm and the internal thickness of the microporous wall is about 250 nm. The hexagonally arranged pores can also be seen from the atomic force microscope (AFM) section analysis (Figure 3c) and the depth curve of the AFM measurements shows the average hole depth is about 325 nm. Furthermore, the 3-D morphology of the ordered porous films obviously indicates that pores are monolayered (Figure 3d). Basically, water droplets act as a template for the formation of honeycomb films. The local capillary forces and Marangoni convection drives these water droplets into a minimum energy state and different factors can affect the energy state of water droplets, which in turn determine the morphology of the microporous films.6,7 We explored the effect of the concentration of MnTPPSDODMA complexes, DODMAB, and solvents on the formation of honeycomb films, respectively. The structure of the films can be modulated by slightly adjusting the concentration of the MnTPPS-DODMA complexes, which were investigated with different concentrations on solid surface under 80% humidity (Figure 4). The relationship between the pore size and the concentration was included in Figure 4f. As shown in Figure 4, the average pore diameter of the films slightly changes from 0.84 to 0.72 μm with the increase of the concentration of the MnTPPS-DODMA complexes from 1.0 to 4.2 mg·mL−1. A classic hexagonal array was formed when the 6422

dx.doi.org/10.1021/la501244s | Langmuir 2014, 30, 6419−6426

Langmuir

Article

low solubility in water. Based on these factors, chloroform (CHCl3), carbon disulfide (CS2), and tetrahydrofuran (THF) have been reported to be the best solvents for the formation of highly ordered honeycomb films of MnTPPS-DODMA complexes. Table 2 shows the data of three solvents. In the Table 2. Effect of Different Solvents on HoneycombPatterned Films

vapor pressure (KPa) boiling point (°C) density (g·cm−3) solubility in water (g·L−1)

chloroform

carbon disulfide

tetrahydrofuran

21.3 61 1.48 8.2

39.7 46 1.26 2.9

21.6 65 0.89 miscible

Figure 7. SEM images of MnTPPS-DODMA complexes/PS honeycomb-patterned films. The concentration of PS is changed from 0.5 (a), 1.0 (b), 2.0 (c), and (d) 2.5 mg·mL−1, respectively.

optical micrographs (Figure 6), a continuous area of microporous honeycomb films can be formed from CHCl3 solution (Figure 6a). No ordered microporous structures but net-like structures can be seen from CS2 solution as a result of the low solubility of MnTPPS-DODMA complexes in CS2 (Figure 6b). Because of the miscibility of THF and water, water droplets can not orderly arrange on the THF solution surface, one is not able to obtain mircroporous films for THF as solvent (Figure 6c). In our study, CHCl3 was chosen to be the solvent for developing honeycomb-patterned films. To investigate the effect of polystyrene (PS) concentration on the morphology of microporous honeycomb films, rMnTPPS:DODMA = 1:5 was chosen as the studied mole ratio and the concentration of MnTPPS-DODMA complexes was fixed at 2.0 mg·mL−1. As shown in Figure 7, one can find that only disordered films were formed by PS. At cPS = 0.5 mg·mL−1 with cMnTPPS‑DODMA = 2.0 mg·mL−1 in chloroform, uniform microporous honeycomb films can be produced with bigger pore sizes (Figure 7a). The pore sizes become bigger with the increase of PS concentration. At cPS = 2.0 mg·mL−1 PS, the order degree of the pores decreases and the walls of the pores become cracked. We can conclude that the addition of PS not only increases the strength of the film, but also modulates the pore sizes. It is believed that porphyrin molecule can be photoexcited to its triplet state from its ground state after absorption of visible light. The energy can be transferred to biological substances, leading to the formation of oxygenated product such as superoxide ion and peroxide (Type I), or directly transfer to triplet oxygen O2 and form singlet oxygen O21, which is a key agent of cellular damage in photodynamic inactivation of microorganisms. To assess the capability of singlet oxygen (O2 1) generation of microporous honeycomb films of

MnTPPS-DODMA complexes, ABDA was employed as a probe molecule to monitor the singlet oxygen generation.36,37 In this method, the microporous honeycomb films of MnTPPSDODMA complexes were placed in spectrofluorimetric cuvettes separately, irradiated with visible light above 420 nm and the absorption intensity of ABDA at 401 nm was monitored concurrently. The controlling experiments were performed with the nonporous films of MnTPPS-DODMA complexes or the films without MnTPPS-DODMA complexes. As shown in Figure 8c, the results indicate that the absorption intensity of ABDA gradually decreased over the course of illumination in the presence of microporous honeycomb films, suggesting an increase amount of singlet oxygen produced by microporous honeycomb films. In contrast, the change of the absorption intensity of ABDA was small without MnTPPSDODMA complexes films under the same experiment condition (Figure 8a), confirming that the decrease in absorption of ABDA was a result of a combined effect of microporous honeycomb films and but not the illumination itself. The capability of the nonporous films of MnTPPSDODMA complexes which was also monitored under the same condition (Figure 8b) generates singlet oxygen. The change of the absorption intensity of ABDA was bigger than the case of the control experiment without any films but smaller than those of the microporous honeycomb films. The time-dependent changes of ABDA absorption of different films were directly compared as shown in Figure 8d, clearly demonstrating that the microporous honeycomb films can generate much more singlet

Figure 6. Optical micrographs of the honeycomb-patterned films prepared by casting 2.0 mg·mL−1 of MnTPPS−DODMA complexes in different solvents on glass substrate, chloroform (a), tetrahydrofuran (b), and carbon disulfide (c). 6423

dx.doi.org/10.1021/la501244s | Langmuir 2014, 30, 6419−6426

Langmuir

Article

Figure 8. Absorption spectra of ABDA (150 μmol·L−1) in PBS solution, photosensitized by (a) glass as control, (b) nonporous thin films, and (c) honeycomb films. (d) Time-dependent decrease in ABDA absorption as a function of illumination time, corresponding to panels a−c.

oxygen than nonporous films having irradiated with visible light above 420 nm. The antibacterial activity photoinduced by the honeycomb films of MnTPPS-DODMA complexes were demonstrated with the BL21 (DE3) E. coli strain. Bacteria survival experiments upon exposure to visible light were performed by a traditional spread plate method (Figure 9). Bacterial counting (Figure 10) shows that the rate of bacteria reduction upon irradiation of E. coli suspension incubated with honeycomb films was 83%, whereas the rate of bacteria reduction of honeycomb films in the dark was only 5%. For the nonporous thin films, the reduction rate is 43% when incubated under light and in the dark the bacteria reduction rate is 7%. In the control

Figure 10. Biocidal activity toward E. coli in the dark and under light illumination for 60 min. Dark and light control experiments were done with the cell suspensions irradiated or in the dark in the absence of photosensitizers. The light was filtered through a 15 cm glass cuvette filled with water to absorb heat. The halogen lamp was placed 30 cm above and 30 cm away from the plates. Values represent mean standard deviation of five separate experiments. Error bars represent standard deviations of data from five separate measurements.

experiments, the rate of reduction rate is 19% and 0% for ITO glass upon exposure to visible light and in the dark, respectively. These results suggest that the microporous honeycomb films with the presence of porphyrins have more efficient antibacterial activity when irradiated with visual light than stored in the dark. This is due to the fact that singlet oxygen (O21) photoinduced by porphyrins under visible light can inactivate the bacteria. The microporous honeycomb films of MnTPPS-DODMA complexes have more efficient antibacterial activity. For the bacteria size (Figure S2) we used on average is more than 1 μm, which is much bigger than the pore

Figure 9. Number of colony forming units (cfu) for E. coli on an LB agar plate. cfu of E. coli suspension incubated with honeycomb film and irradiated with visible light (a). cfu of E. coli suspension incubated with nonporous thin film and irradiated with visible light (b). cfu control with ITO irradiated with visible light (c), cfu of E. coli suspension incubated with honeycomb film in the dark (d), cfu of E. coli suspension incubated with nonporous thin film in the dark (e), and cfu control with ITO in the dark (f). Diameter of the solid LB agar plates was 100 mm. 6424

dx.doi.org/10.1021/la501244s | Langmuir 2014, 30, 6419−6426

Langmuir

Article

size of microporous honeycomb films. We deduce that the efficient antibacterial activity of honeycomb films can increase the surface area to make more porphyrin molecules interact with protons, consequently more O21 are produced to have much more significant damage for killing the bacteria. The morphology of the microporous honeycomb films after 1 h illumination in PBS solution was tested by SEM (Figure S3). The enhanced antibacterial properties of the porphyrin-based honeycomb films suggest that they can be used as antibacterial surface, which promises their potentials in broad applications in medical and nonmedical areas.

(5) Wan, L.; Li, J.; Ke, B.; Xu, Z. Ordered microporous membranes templated by breath figures for size-selective separation. J. Am. Chem. Soc. 2012, 134, 95−98. (6) Ma, H.; Dong, R.; Van Horn, J. D.; Hao, J. Spontaneous formation of radially aligned microchannels. Chem. Commun. 2011, 47, 2047−2049. (7) Ma, H.; Hao, J. Ordered patterns and structures via interfacial self-assembly: superlattices, honeycomb structures and coffee rings. Chem. Soc. Rev. 2011, 40, 5457−5471. (8) Widawski, G.; Rawiso, M.; Francois, B. Self-organized honeycomb morphology of star-polymer polystyrene films. Nature 1994, 369, 387−389. (9) Liu, C.; Gao, C.; Yan, D. Honeycomb-patterned photoluminescent films fabricated by self-assembly of hyperbranched polymers. Angew. Chem., Int. Ed. 2007, 46, 4128−4131. (10) Cheng, C. X.; Tian, Y.; Shi, Y. Q.; Tang, R. P.; Xi, F. Porous polymer films and honeycomb structures based on amphiphilic dendronized block copolymers. Langmuir 2005, 21, 6576−6581. (11) Maruyama, N.; Koito, T.; Nishida, J.; Sawadaishi, T.; Cieren, X.; Ijiro, K.; Karthaus, O.; Shimomura, M. Mesoscopic patterns of molecular aggregates on solid substrates. Thin Solid Films 1998, 327− 329, 854−856. (12) Zhao, H.; Shen, Y.; Zhang, S.; Zhang, H. A vapor phase hydrothermal modification method converting a honeycomb structured hybrid film into photoactive TiO2 film. Langmuir 2009, 25, 11032−11037. (13) Ma, H.; Hao, J. Evaporation-induced ordered honeycomb structures of gold nanoparticles at the air/water interface. Chem.Eur. J. 2010, 16, 655−660. (14) Tang, P.; Hao, J. Formation mechanism and morphology modulation of honeycomb hybrid films made of polyoxometalates/ surfactants at the air/water interface. J. Colloid Interface Sci. 2009, 333, 1−5. (15) Fan, D.; Jia, X.; Tang, P.; Hao, J.; Liu, T. Self-patterning of hydrophobic materials into highly ordered honeycomb nanostructures at the air/water interface. Angew. Chem., Int. Ed. 2007, 46, 3342−3345. (16) Sun, H.; Li, W.; Wu, L. Honeycomb-patterned films fabricated by self-organization of DNA-surfactant complexes. Langmuir 2009, 25, 10466−10472. (17) Du, M.; Zhu, P.; Yan, X.; Su, Y.; Song, W.; Li, J. Honeycomb self-assembled peptide scaffolds by the breath figure method. Chem. Eur. J. 2011, 17, 4238−4245. (18) Xu, W. Z.; Zhang, X.; Kadla, J. F. Design of functionalized cellulosic honeycomb films: site-specific biomolecule modification via “click chemistry. Biomacromolecules 2012, 13, 350−357. (19) Fan, D.; Xia, X.; Ma, H.; Du, B.; Wei, Q. Honeycomb-patterned fluorescent films fabricated by self-assembly of surfactant-assisted porphyrin/polymer composites. J. Colloid Interface Sci. 2013, 402, 146−50. (20) Zhu, L.; Wan, L.; Jin, J.; Xu, Z. Honeycomb porous films prepared from porphyrin-cored star polymers: submicrometer pores induced by transition of monolayer into multilayer structures. J. Phys. Chem. C 2013, 117, 6185−6194. (21) DeRosa, M. C.; Crutchley, R. J. Photosensitized singlet oxygen and its applications. Coordin. Chem. Rev. 2002, 233−234, 351−371. (22) Krouit, M.; Granet, R.; Krausz, P. Photobactericidal plastic films based on cellulose esterified by chloroacetate and a cationic porphyrin. Bioorg. Med. Chem. 2008, 16, 10091−10097. (23) Bonnetta, R.; Krystevab, M. A.; Lalovb, I. G.; Artarskyb, S. V. Water disinfection using photosensitizers immobilized on chitosan. Water Res. 2006, 40, 1269−1275. (24) Jesenská, S.; Plíštil, L.; Kubát, P.; Lang, K.; Brožová, L.; Popelka, S.; Szatmáry, L.; Mosinger, J. Antibacterial nanofiber materials activated by light. J. Biomed. Mater. Res. A 2011, 99, 676−683. (25) Funes, M. D.; Caminos, D. A.; Alvarez, M. G.; Fungo, F.; Otero, L. A.; Durantini, E. N. Photodynamic properties and photoantimicrobial action of electrochemically generated porphyrin polymeric films. Environ. Sci. Technol. 2009, 43, 902−908.

4. CONCLUSIONS In conclusion, through the encapsulation of a cationic doublechain surfactant (DODMA+), porphyrins can be successfully introduced into the establishment of honeycomb-patterned films. The porphyrin−surfactant complexes play important roles in stabilizing the template water droplets during the formation progress. The morphology of these films can be easily controlled by varying the complex concentration, the extra surfactant concentration, PS concentration, and the selective solvent. Antibacterial experiments indicate that these films composed of porphyrins have efficient antibacterial activities under visual light, indicating that porphyrin-based honeycomb films can serve as an interesting and promising photodynamic surface to inactivate microorganisms. Such materials have great potentials in medical industrial and household environmental applications.

■

ASSOCIATED CONTENT

S Supporting Information *

Photographs of honeycomb films with strong bright iridescent colors, SEM observations of E. coli, and SEM images of microporous honeycomb films before and after irradiated with visible light. This material is available free of charge via the Internet at http://pubs.acs.org.

■

AUTHOR INFORMATION

Corresponding Author

*E-mail: [email protected]. Tel: +86-531-88363532. Fax: +86531-88564750. Notes

The authors declare no competing financial interest.

■

ACKNOWLEDGMENTS This work was financially supported by the NSFC (Grant Nos. 21033005 and 21273134) and GIFSDU (yyx10099).

■

REFERENCES

(1) Ma, H.; Cui, J.; Chen, J.; Hao, J. Self-organized polymer nanocomposite inverse opal films with combined optical properties. Chem.Eur. J. 2011, 17, 655−660. (2) Yamamoto, S.; Tanaka, M.; Sunami, H.; Ito, E.; Yamashita, S.; Morita, Y.; Shimomura, M. Effect of honeycomb-patterned surface topography on the adhesion and signal transduction of porcine aortic endothelial cells. Langmuir 2007, 23, 8114−8120. (3) Brown, P. S.; Talbot, E. L.; Wood, T. J.; Bain, C. D.; Badyal, J. P. S. Superhydrophobic hierarchical honeycomb surfaces. Langmuir 2012, 28, 13712−13719. (4) Kong, L.; Dong, R.; Ma, H.; Hao, J. Au NP honeycomb-patterned films with controllable pore size and their surface-enhanced raman scattering. Langmuir 2013, 29, 4235−4241. 6425

dx.doi.org/10.1021/la501244s | Langmuir 2014, 30, 6419−6426

Langmuir

Article

(26) Huang, C.; Xing, C.; Wang, S.; Li, Y.; Liu, H.; Lai, S.; Che, C. Fabrication of a well ordered microspheres film for efficient antibacterial activity. Chem. Commun. 2011, 47, 7644−7646. (27) Pessoni, L.; Lacombe, S.; Billon, L.; Brown, R.; Save, M. Photoactive, porous honeycomb films prepared from rose bengalgrafted polystyrene. Langmuir 2013, 29, 10264−10271. (28) MacPhail, R. A.; Strauss, H. L.; Snyder, R. G.; Elliger, C. A. C-H stretching modes and the structure of normal-alkyl chains. 2. long, alltrans chains. J. Phys. Chem. B 1984, 88, 334−341. (29) Kasha, M. Energy transfer mechanisms and the molecular exciton model for molecular aggregates. Radiat. Res. 1963, 20, 55−70. (30) Zhao, L.; Ma, R.; Li, J.; Li, Y.; An, Y.; Shi, L. J- and H-aggregates of 5,10,15,20-tetrakis- (4-sulfonatophenyl)-porphyrin and interconversion in PEG-b-P4VP micelles. Biomacromolecules 2008, 9, 2601−2608. (31) Reddy, M. H. V.; Al-Shammari, R. M.; Al-Attar, N.; Rogers, L.; Lopez, S.; Forster, R. J.; Senge, M. O.; Keyes, T. E.; Rice, J. H. Fractal structures in n-phenyl-porphyrin J-aggregate films. Mater. Chem. Phys. 2014, 143, 963−968. (32) Khairutdinov, R. F.; Serpone, N. Photoluminescence and transient spectroscopy of free base porphyrin aggregates. J. Phys. Chem. B 1999, 103, 761−769. (33) Tang, P.; Hao, J. Photoluminescent honeycomb films templated by microwater droplets. Langmuir 2010, 26, 3843−3847. (34) Karthaus, O.; Maruyama, N.; Cieren, X.; Shimomura, M.; Hasegawa, H.; Hashimoto, T. Water-assisted formation of micrometersize honeycomb patterns of polymers. Langmuir 2000, 16, 6071−6076. (35) Escalé, P.; Rubatat, L.; Billon, L.; Save, M. Recent advances in honeycomb-structured porous polymer films prepared via breath figures. Eur. Polym. J. 2012, 48, 1001−1025. (36) Wang, T.; Zhang, L.; Su, Z.; Wang, C.; Liao, Y.; Fu, Q. Multifunctional hollow mesoporous silica nanocages for cancer cell detection and the combined chemotherapy and photodynamic therapy. ACS Appl. Mater. Interfaces 2011, 3, 2479−86. (37) Zhao, T.; Wu, H.; Yao, S. Q.; Xu, Q. H.; Xu, G. Q. Nanocomposites containing gold nanorods and porphyrin-doped mesoporous silica with dual capability of two-photon imaging and photosensitization. Langmuir 2010, 26, 14937−14942.

6426

dx.doi.org/10.1021/la501244s | Langmuir 2014, 30, 6419−6426