Langmuir 2005, 21, 4705-4709
4705
Encapsulation of Carbon Nanotubes by Self-Assembling Peptide Amphiphiles Michael S. Arnold,† Mustafa O. Guler,‡ Mark C. Hersam,† and Samuel I. Stupp*,†,‡,§ Department of Materials Science and Engineering, Northwestern University, Evanston, Illinois 60208, Department of Chemistry, Northwestern University, Evanston, Illinois 60208, and Feinberg School of Medicine, Northwestern University, Chicago, Illinois Received December 13, 2004. In Final Form: February 11, 2005 We demonstrate the dispersion and noncovalent functionalization of carbon nanotubes in water using peptide amphiphiles each consisting of a short hydrophobic alkyl tail coupled to a more hydrophilic peptide sequence. The assembly of peptide amphiphile molecules on the surfaces of carbon nanotubes adds biofunctionality to these one-dimensional conductors and simultaneously eliminates the hydrophobic nanotube-water interface, thus dispersing them in the aqueous medium. This should occur without the degradation of their structural, electronic, and optical properties caused by covalent functionalization and without the need for specific peptide sequences designed to bind with nanotube surfaces. The encapsulation by peptide amphiphiles is confirmed using transmission electron microscopy and optical absorbance spectroscopy and may have significant future applications in biosensing or medicine.
Introduction Carbon nanotubes are intriguing nanomaterials for a wide range of mechanical,1,2 electronic,3-8 optical,9,10 sensing,11,12 and biological applications.13-15 These nanostructures are appealing for use in molecular electronics because of their nanoscale, wirelike geometry6,16,17 and * To whom correspondence should be addressed. E-mail:
[email protected]. † Department of Materials Science and Engineering. ‡ Department of Chemistry. § Weinberg School of Medicine. (1) Dalton, A. B.; Collins, S.; Munoz, E.; Razal, J. M.; Ebron, V. H.; Ferraris, J. P.; Coleman, J. N.; Kim, B. G.; Baughman, R. H. Nature 2003, 423, 703-703. (2) Yu, M. F.; Files, B. S.; Arepalli, S.; Ruoff, R. S. Phys. Rev. Lett. 2000, 84, 5552-5555. (3) Wei, B. Q.; Vajtai, R.; Ajayan, P. M. Appl. Phys. Lett. 2001, 79, 1172-1174. (4) Charlier, J. C.; Issi, J. P. Appl. Phys. A: Mater. Sci. Process. 1998, 67, 79-87. (5) Saito, R.; Fujita, M.; Dresselhaus, G.; Dresselhaus, M. S. Phys. Rev. B: Condens. Matter Mater. Phys. 1992, 46, 1804-1811. (6) Derycke, V.; Martel, R.; Appenzeller, J.; Avouris, P. Nano Lett. 2001, 1, 453-456. (7) Javey, A.; Guo, J.; Wang, Q.; Lundstrom, M.; Dai, H. J. Nature 2003, 424, 654-657. (8) Odom, T. W.; Huang, J. L.; Kim, P.; Lieber, C. M. J. Phys. Chem. B 2000, 104, 2794-2809. (9) O’Connell, M. J.; Bachilo, S. M.; Huffman, C. B.; Moore, V. C.; Strano, M. S.; Haroz, E. H.; Rialon, K. L.; Boul, P. J.; Noon, W. H.; Kittrell, C.; Ma, J. P.; Hauge, R. H.; Weisman, R. B.; Smalley, R. E. Science 2002, 297, 593-596. (10) Arnold, M. S.; Sharping, J. E.; Stupp, S. I.; Kumar, P.; Hersam, M. C. Nano Lett. 2003, 3, 1549-1554. (11) Chen, R. J.; Bangsaruntip, S.; Drouvalakis, K. A.; Kam, N. W. S.; Shim, M.; Li, Y. M.; Kim, W.; Utz, P. J.; Dai, H. J. Proc. Natl. Acad. Sci. U.S.A. 2003, 100, 4984-4989. (12) Shim, M.; Kam, N. W. S.; Chen, R. J.; Li, Y. M.; Dai, H. J. Nano Lett. 2002, 2, 285-288. (13) Pantarotto, D.; Partidos, C. D.; Hoebeke, J.; Brown, F.; Kramer, E.; Briand, J. P.; Muller, S.; Prato, M.; Bianco, A. Chem. Biol. 2003, 10, 961-966. (14) Pantarotto, D.; Partidos, C. D.; Graff, R.; Hoebeke, J.; Briand, J. P.; Prato, M.; Bianco, A. J. Am. Chem. Soc. 2003, 125, 6160-6164. (15) Kam, N. W. S.; Jessop, T. C.; Wender, P. A.; Dai, H. J. J. Am. Chem. Soc. 2004, 126, 6850-6851. (16) Wind, S. J.; Appenzeller, J.; Martel, R.; Derycke, V.; Avouris, P. Appl. Phys. Lett. 2002, 81, 1359-1359.
the possibility of ballistic electronic conduction.7 Furthermore, carbon nanotubes are promising materials for sensing due to their robust mechanical strength,1,2 exceptionally large surface-to-volume ratio,4,5 high electrical conductivity,3-8 and electrical and optical properties that depend on their external environment.18,19 To take advantage of the excellent properties of carbon nanotubes, especially for sensing and biological applications, an external functionalization is often necessary.9,11-15,20-29 Ideally, this functionalization should be self-assembling and stable, and it should not degrade the outstanding physical characteristics of the carbon nanotubes. In addition, because carbon nanotubes without modification aggregate in most solvents due to strong inter-nanotube van der Waals interactions, it should also create a physical (17) Martel, R.; Schmidt, T.; Shea, H. R.; Hertel, T.; Avouris, P. Appl. Phys. Lett. 1998, 73, 2447-2449. (18) Arnold, M. S.; Lan, S.; Cruz, S. C.; Sharping, J. E.; Stupp, S. I.; Kumar, P.; Hersam, M. C. Proc. SPIE-Int. Soc. Opt. Eng. 2004, 5359, 376. (19) Spataru, C. D.; Ismail-Beigi, S.; Benedict, L. X.; Louie, S. G. Phys. Rev. Lett. 2004, 92. (20) Chen, Y.; Haddon, R. C.; Fang, S.; Rao, A. M.; Lee, W. H.; Dickey, E. C.; Grulke, E. A.; Pendergrass, J. C.; Chavan, A.; Haley, B. E.; Smalley, R. E. J. Mater. Res. 1998, 13, 2423-2431. (21) Star, A.; Liu, Y.; Grant, K.; Ridvan, L.; Stoddart, J. F.; Steuerman, D. W.; Diehl, M. R.; Boukai, A.; Heath, J. R. Macromolecules 2003, 36, 553-560. (22) Dieckmann, G. R.; Dalton, A. B.; Johnson, P. A.; Razal, J.; Chen, J.; Giordano, G. M.; Munoz, E.; Musselman, I. H.; Baughman, R. H.; Draper, R. K. J. Am. Chem. Soc. 2003, 125, 1770-1777. (23) Tang, B. Z.; Xu, H. Y. Macromolecules 1999, 32, 2569-2576. (24) Zheng, M.; Jagota, A.; Semke, E. D.; Diner, B. A.; McLean, R. S.; Lustig, S. R.; Richardson, R. E.; Tassi, N. G. Nat. Mater. 2003, 2, 338-342. (25) Holzinger, M.; Vostrowsky, O.; Hirsch, A.; Hennrich, F.; Kappes, M.; Weiss, R.; Jellen, F. Angew. Chem., Int. Ed. 2001, 40, 4002. (26) Richard, C.; Balavoine, F.; Schultz, P.; Ebbesen, T. W.; Mioskowski, C. Science 2003, 300, 775-778. (27) Wang, S. Q.; Humphreys, E. S.; Chung, S. Y.; Delduco, D. F.; Lustig, S. R.; Wang, H.; Parker, K. N.; Rizzo, N. W.; Subramoney, S.; Chiang, Y. M.; Jagota, A. Nat. Mater. 2003, 2, 196-200. (28) Zorbas, V.; Ortiz-Acevedo, A.; Dalton, A. B.; Yoshida, M. M.; Dieckmann, G. R.; Draper, R. K.; Baughman, R. H.; Jose-Yacaman, M.; Musselman, I. H. J. Am. Chem. Soc. 2004, 126, 7222-7227. (29) Kang, Y. J.; Taton, T. A. J. Am. Chem. Soc. 2003, 125, 56505651.
10.1021/la0469452 CCC: $30.25 © 2005 American Chemical Society Published on Web 03/25/2005
4706
Langmuir, Vol. 21, No. 10, 2005
or electrostatic barrier to aggregation and improve solubility.30 In this work, we focus on functionalizing carbon nanotubes with peptide-based molecules. The association of carbon nanotubes with peptides is expected to be useful in biosensor applications and in the development of new bioactive nanomaterials. The biological potential of carbon nanotubes in immunology has been explored by Pantarotto et al.,13,14 who have demonstrated enhanced in vivo antibody response from covalently linked nanotubepeptide conjugates, and by Kam, who observed the nontoxic uptake of carbon nanotubes into cells.15 Toward this end, Wang has identified peptide sequences with specific affinities for carbon nanotubes using phage display,27 and Dieckmann and Zorbas have designed an amphiphilic peptide sequence which folds into an R-helix on nanotube sidewalls.22,28 For sensing, Shim and Chen have functionalized SWNTs grown on substrates with ethylene oxide-based surfactants.11,12 Subsequently, these nonionic surfactants were covalently associated with proteins to electrically detect the binding of antibodies.11 Our approach to the functionalization of carbon nanotubes is to utilize peptide amphiphile (PA) molecules in which an amino acid sequence is covalently coupled to a hydrophobic alkyl tail.31,32 Since the surface of carbon nanotubes is nonpolar and hydrophobic, peptide amphiphiles are expected to self-assemble on this surface from aqueous solution, minimizing the interfacial energy of the nanotube-water interface.9,26,29 This approach offers several potential benefits. First, using the noncovalent assembly of peptide amphiphiles, the nanotube sidewalls should not be chemically modified, thus maintaining the outstanding electrical, mechanical, and optical properties of unmodified carbon nanotubes. Second, the hydrophobic tail of the peptide amphiphile is expected to interact with the hydrophobic nanotube surface, leaving the peptide sequence exposed on the exterior for sensing or other biological applications. Furthermore, the water solubility of nanotubes biofunctionalized with peptide amphiphiles should be controllable by adjusting pH since the net charge of peptide segments and the resulting ionic repulsion among nanotubes in solution will vary with pH. Finally, this approach is expected to be generally useful for either positively or negatively charged peptide sequences, without the need to incorporate sequences that specifically bind to nanotube surfaces. Synthesis of Peptide Amphiphiles In this work, branched anionic and cationic peptide amphiphiles (Chart 1) were utilized for the biofunctionalization of carbon nanotubes. Branched peptide structures have been proposed to enhance the accessibility of bioactive sequences and to enable the incorporation of multiple sequences on a single molecule.33 For the functionalization of multiwalled carbon nanotubes (MWNTs), PA 1 and PA 2 were used, and for the functionalization of single-walled carbon nanotubes (SWNTs), PA 3 was used. The branched PAs33 were prepared by Fmoc solid-phase peptide synthesis (SPPS). Peptides were constructed on MBHA Rink amide resin. Amino acid couplings were accomplished with 4 equiv of Fmoc-protected amino acid, 3.95 equiv of HBTU, and 6 equiv of DIEA for 4 h. Fmoc deprotections were performed with 30% piperidine/DMF solution for 10 min. Mtt removal was (30) Ausman, K. D.; Piner, R.; Lourie, O.; Ruoff, R. S.; Korobov, M. J. Phys. Chem. B 2000, 104, 8911-8915. (31) Hartgerink, J. D.; Beniash, E.; Stupp, S. I. Proc. Natl. Acad. Sci. U.S.A. 2002, 99, 5133-5138. (32) Hartgerink, J. D.; Beniash, E.; Stupp, S. I. Science 2001, 294, 1684-1688. (33) Guler, M. O.; Soukasene, S.; Stupp, S. I. Nano Lett. 2005, 5, 249-252.
Arnold et al. Chart 1. Structure of Peptide Amphiphilesa
a Simple, branched anionic (PA1) and cationic (PA2) peptide amphiphiles and a branched anionic peptide amphiphile with a biologically active sequence (Arg-Gly-Asp-Ser) (PA3) were used for the encapsulation of carbon nanotubes.
accomplished with 4% TFA/dichloromethane solution in the presence of TIS for 5 min. For PA 1 and PA 2, Fmoc-Lys(Mtt)-OH (Mtt: 4-methyltrityl) was coupled to MBHA Rink amide resin, followed by cleavage of the Mtt protecting group on the -amine to couple palmitic acid without affecting Fmoc protection. This was followed by Fmoc removal on the -amine to grow the peptide segment of the PA. Four consecutive cysteine residues (Fmoc-Cys(Trt)-OH) followed by two leucine residues (Fmoc-Leu-OH) were coupled to promote β-sheet formation between the peptide segments, favoring cylindrical assembly around the nanotubes. The branching point in the PAs was introduced at a lysine dendron using Fmoc-Lys(Fmoc)-OH. For PA 1 two L-aspartic acid di-tert-butyl ester hemisuccinate residues were included in the sequence to create negatively charged, water-soluble PAs at neutral pH and basic conditions, as well. For PA 2 two lysine residues (BocLys(Boc)-OH) were coupled to obtain molecules with net positive charge and water solubility at neutral and acidic conditions. The branching of the peptide headgroup of PA 3 was achieved using orthogonal protecting group chemistry for the amines at the R and positions of the lysine residue. First, Fmoc-Lys(Mtt)OH was coupled to MBHA Rink amide resin, followed by cleavage of the Mtt protecting group on the amine to couple palmitic acid without affecting Fmoc protection. This was followed by Fmoc removal on the R amine to grow the Leu-Leu-Leu-Ala-Ala-Ala peptide segment of the PA 3. The branching point was introduced at a lysine dendron using Fmoc-Lys(Mtt)-OH. To grow the first arm of the PA 3, Fmoc on the R amine was removed before Mtt. L-Aspartic acid di-tert-butyl ester hemisuccinate residue was coupled to the first branch. Later, Mtt on the amine of the lysine residue was removed and the bioactive peptide sequence Arg-Gly-Asp-Ser was added. Cleavage of the PA 1 and PA 2 from the resin was carried out with a mixture of TFA/EDT/H2O/TIS in a ratio of 94:2.5:2.5:1 for 3 h, and PA 3 was cleaved with a mixture of TFA/H2O/TIS in a ratio of 95:2.5:2.5. Excess TFA was removed by rotary evaporation. The remaining viscous peptide solution was triturated with cold ether, and the resulting white product was dried under vacuum. PAs were characterized by matrix-assisted laser desorption ionization-time-of-flight mass spectrometry (MALDI-TOF MS) and electrospray ionization mass spectrometry (ESI-MS). MBHA Rink amide resin, HBTU, and peptides were purchased from Novabiochem. All other chemicals were purchased from Aldrich and used without further purification.
Encapsulation of Carbon Nanotubes
Langmuir, Vol. 21, No. 10, 2005 4707
Figure 1. Photographs of solutions of PA and MWNTs, diluted 56-fold unless otherwise noted. (A) Control solution of MWNTs in water without the PA molecule. Except for the absence of PA, this solution was processed identically with that of solution B and solution C, which contain PA 1 and PA 2, respectively. The MWNTs without PA are water-insoluble from pH 1 to 13. (B) Homogeneous aqueous solution of MWNTs functionalized by PA 1 at a pH of 7. (C) Homogeneous aqueous solution of MWNTs functionalized by PA 2 at a pH of 7, diluted 84-fold for the photograph. (D) Aqueous solution of MWNTs functionalized by PA 1 at a pH of 1. (E) Aqueous solution of MWNTs functionalized by PA 2 at a pH of 9. (F) Aqueous gel of PA 2 with embedded, peptide-functionalized MWNTs at a pH of 9 not diluted. At this concentration of 1 wt % PA 2, a selfsupporting aqueous gel is formed. (PA 1 does not form a gel under analogous conditions.)
Encapsulation of MWNTs 34
MWNTs were dispersed in aqueous solutions of 1 wt % PA 1 or PA 2 (at a ratio of 10:1 by weight of PA to raw MWNT material) via bath ultrasonication (Bransonics 3510) for 1 h. Additionally, 12 mol of dithiothreitol (DTT)/mol of peptide amphiphile was added to prevent cross-linking of cysteine residues, and potassium hydroxide and hydrochloric acid were utilized to adjust pH. Control solutions of MWNTs with DTT and acid or base but without amphiphile were also prepared by following identical procedures. Dispersions of PA functionalized MWNTs and control solutions of MWNTs without PA were characterized by transmission electron microscopy (TEM). To analyze a solution by TEM, 5 µL was placed on a nickel TEM grid coated with a Quantifoil Multi-A holey carbon film (SPI Supplies, Inc., West Chester, PA). After 30 s, the applied solution was wicked through the grid using filter paper, and the grid was rinsed for 10 s in deionized water to remove excess material. The TEM grids were examined using a Hitachi 8100 transmission electron microscope with an energy of 200 keV and magnifications of 10-300 thousand times. The nanotubes were examined both over holes in the holey carbon grid and on the carbon thin film, as well. A clear difference is apparent between the solutions of MWNTs with and without peptide amphiphile (Figure 1A-C). In water without the peptide amphiphile, MWNTs were insoluble even after ultrasonication (Figure 1A), but in aqueous solutions of either 1 wt % PA 1 or PA 2 at their respective soluble pH, the MWNTs were homogeneously dispersed after ultrasonication (Figure 1B,C, respectively). This suggests that peptide amphiphiles act as surfactants, resulting in the noncovalent functionalization of the MWNTs and enhancing MWNT solubility. The dispersions of MWNTs with PA 1 or PA 2 were stable for months after ultrasonication. Since the net ionic charge of both PA 1 and PA 2 can be controlled by pH, the solubility of the peptide functionalized MWNTs can be tuned, as well. By reduction of the net charge of the peptide sequence, the ionic repulsion among free peptide amphiphile molecules and among (34) MWNTs were purchased from Nanocyl SA (Namur, Belgium); product name “very thin purified MWNTs without functionalization”.
Figure 2. TEM micrographs of MWNTs without peptide amphiphile (A) at low magnification (bar ) 50 nm) and (B) at high magnification (bar ) 10 nm). False color is used is used to highlight the MWNTs using a red-scale. The exterior of the nonfunctionalized MWNTs appears clean of adsorbed species. The scale bar is 10 nm.
peptide amphiphile functionalized MWNTs can be reduced, leading to aggregation (Figure 1D-F). For instance, at neutral and basic pH, the peptide sequence of PA 1 molecules is negatively charged. In this state, PA 1 molecules homogeneously disperse and functionalize the MWNTs (Figure 1B). After addition of acid to lower the pH and decrease the net charge of each PA molecule, the PA 1 functionalized MWNTs phase separate from solution, forming aggregates (Figure 1D). On the other hand, for the positively charged PA 2, aggregation of the MWNTs occurs at basic pH. Furthermore, for concentrations greater than 0.5 wt % PA 2 will form a self-supporting aqueous gel at basic pH. Under such conditions, after addition of base, the MWNTs do not phase separate into visible aggregates but remain embedded in a gel matrix (Figure 1F). Given the large thickness of MWNTs, a clear difference was observed by TEM between nonfunctionalized and PAfunctionalized tubes (Figures 2-4). At both low and high magnifications, MWNTs from a control solution without peptide amphiphile appeared bare on their exterior (Figure 2). Fresnel fringes due to the concentric graphitic layers of MWNTs were easily visible, separated by 3.5 ( 0.2 Å (Figure 2B). The outer diameter for the MWNTs used in this study was expected to range from 3 to 15 nm, while the inner diameter was expected to range from 2 to 7 nm.34 The TEM images confirm this expectation as the MWNTs appear appreciably hollow in their interiors. In contrast with nonfunctionalized MWNTs, an organic coating was observed by TEM for peptide amphiphile functionalized MWNTs. PA 1 functionalized MWNTs at a pH of 1 (Figure 1D) and PA 2 functionalized MWNTs
4708
Langmuir, Vol. 21, No. 10, 2005
Figure 3. TEM micrographs of MWNTs with PA 1 (A) at low magnification (bar ) 50 nm) and (B) at high magnification (bar ) 10 nm). False color is used to highlight the MWNTs using a red-scale and the surrounding organic coating using a bluescale. To false color TEM images, a distinction between the organic coating and the background was determined by a sharp difference in contrast, and a distinction between the MWNT and the organic coating was made by using the Fresnel fringes to identify the MWNT.
at a pH of 9 (Figure 1E) were examined for comparison with control solutions of nonfunctionalized MWNTs. In Figure 3A, the MWNTs are discernible by a reduction in contrast through their hollow interior and by their long aspect ratio. At high magnification, the Fresnel fringes of the MWNTs are visible, but an exterior organic coating which was not present on MWNTs from control solutions is now also apparent (Figure 3B). A similar organic coating was observed on the exterior of MWNTs functionalized by PA 2 (Figure 4). At low magnification (Figure 4A), MWNTs are visible (identified by a reduction in contrast through their hollow interior), and at high magnification (Figure 4B), an organic film is also visible on the exterior of the MWNTs. The measured thickness of the PA 1 coating varied from less than 0.5 nm to as thick as 9 nm, while the thickness of the PA 2 coating varied from 0.5 to 4 nm. For both molecules, the thickness was observed to vary along the length of the nanotubes. This variability in thickness suggests that the PA molecules are not wellordered on the surface of the MWNTs but rather may be entangled, forming multiple layers with varying orientation on the nanotube surface.
Arnold et al.
Figure 4. TEM micrographs of MWNTs with PA 2 at low magnification (bar ) 50 nm) and (B) at high magnification (bar ) 10 nm).
semiconducting SWNTs can be modulated by the external environment, these are the nanostructures of interest for sensing applications. However, direct observation by TEM is more challenging given the similarity in thicknesses of the expected PA coating and the diameter of a SWNT. Therefore, we used the optical properties of semiconducting SWNTs to verify the ability of PAs to encapsulate and isolate individual tubes. Additionally, in this case, we used a PA, which contains a bioactive peptide sequence (ArgGly-Asp-Ser) well-known to bind to cell membrane proteins known as integrins (see PA 3, Chart 1).35,36 The peptide in this PA also has a branched structure to enhance the biological recognition of this sequence.37 CoMoCAT (Co-Mo catalyst) SWNTs (Southwest Nanotechnologies, Inc.) containing enhanced concentrations of (6, 5) chirality tubes were dispersed in aqueous solutions of 0.94 wt % PA 3 (100:1 w/w PA to raw SWNT material, adjusted to pH 11 with KOH after PA addition) via horn ultrasonication for 30 min using a tapered microtip extension, which was immersed into the solution (Fisher Scientific, Sonic Dismembrator 550). Immediately following the horn ultrasonication, the PA 3-SWNT solution appeared homogeneously black and opaque to the naked eye. The isolated and dispersed SWNTs were then separated from the aggregated SWNTs and the insoluble material by ultracentrifugation (24 krpm, 75 min, 22 °C, Beckman-Coulter TLA100.3 rotor). The upper 60% of the supernatant, containing the slower sedimenting, presum-
Encapsulation of SWNTs Having demonstrated the encapsulation of MWNTs with PAs by direct observation using TEM, we explored the encapsulation of SWNTs using the same strategy. Because the electronic conductivity and optical properties of
(35) Schaffner, P.; Dard, M. M. Cell. Mol. Life Sci. 2003, 60, 119132. (36) Ziong, J.-P.; Stehle, T.; Zhang, R.; Joachimiak, A.; Frech, M.; Goodman, S.; Arnaout, M. A. Science 2002, 296, 151-5. (37) Storrie, H.; Guler, M. O.; Stupp, S. I. Manuscript under preparation.
Encapsulation of Carbon Nanotubes
Langmuir, Vol. 21, No. 10, 2005 4709
have attributed to the increased accessibility of the nanotube to the aqueous environment.18 A comparable red-shift can also be induced by lowering the concentration of SDS in aqueous solution, which disturbs the organization of SDS on the nanotube surface.38 The spectral broadening of the measured van Hove transitions for the PA 3-isolated SWNTs may arise from a variability in spectral shift due to a variability in the packing of the peptide amphiphile around the nanotube surface, as observed for the MWNTs (Figures 2-4). The observation of the van Hove transitions in the absorbance spectra of PA 3-functionalized SWNTs also suggests that the SWNTs are not appreciably covalently modified by the peptide amphiphile molecules. The covalent modification of nanotube sidewalls is expected to disrupt the delocalization of the free electronic carriers in SWNTs, causing the interband transitions between van Hove singularities to disappear.39 Figure 5. Optical abosrbance spectra of SDS isolated SWNTs (dotted line), PA 3 isolated SWNTs from the supernatant (solid line), and aggregated SWNTs (dashed line) in the visible and infrared regions of the electromagnetic spectrum. The # symbol indicates the second van Hove transition for the (6,5) chirality SWNT, and the * marks the corresponding first van Hove transition.
ably smaller and more isolated nanotubes, was decanted and saved for analysis. The pellet, containing the faster sedimenting, aggregated nanotubes, was redispersed in water by brief bath ultrasonication (15 min), diluted 30-fold, and adjusted to pH 11 with KOH. For comparison, sodium dodecyl sulfate (SDS) isolated SWNTs were also prepared by horn ultrasonication and centrifugation using similar procedures, as previously described elsewhere.9,10,18 The encapsulation and isolation of SWNTs was confirmed by measuring the optical absorbance spectra of the SWNT solutions in the visible and near-infrared regions of the optical spectrum (Cary 500, Varian, Inc.). Solutions of isolated SWNTs should exhibit sharp peaks in optical absorbance for energies corresponding to direct, electronic interband transitions between van Hove singularities,9 while for aggregated SWNTs these transitions should be broadened and significantly less intense. The optical absorbance spectra of the PA 3-functionalized SWNTs, collected from the supernatant and pellet, along with that of SDS isolated SWNTs are depicted in Figure 5. Both the PA 3-functionalized SWNTs from the supernatant and the SDS-isolated SWNTs exhibit sharp peaks in absorbance in the visible and infrared regions due to optical interband transitions between van Hove singularities, as expected for solutions rich in isolated tubes. On the other hand, this absorbance is significantly decreased and broadened for solutions of SWNTs from the pellet, as expected for aggregated SWNTs. This suggests that, during ultrasonication, the peptide amphiphile molecules are successfully solubilizing a fraction of the SWNTs and that to a large extent these isolated, PA-functionalized SWNTs can be separated from aggregates of SWNTs by ultracentrifugation. The optical absorbance spectra were invariant over several months. The first- and second-order van Hove transitions of the PA 3-isolated SWNTs were red-shifted with respect to the van Hove transitions of SDS-isolated SWNTs (Figure 5). A red-shift of 16.5 and 19.1 meV was observed for the second- and first-order van Hove transitions of the (6, 5) chirality nanotubes, respectively. This shift is similar to that observed in aqueous solutions of single-stranded DNA-wrapped SWNTs (17.8 and 19.4 meV), which we
Conclusions We have demonstrated the dispersion and noncovalent functionalization of carbon nanotubes with peptide amphiphile molecules. While carbon nanotubes without functionalization are insoluble in water, we have shown that these nonpolar, one-dimensional nanostructures can be dispersed by peptide amphiphiles in aqueous solution. Furthermore, by adjustment of pH, the solubility of functionalized carbon nanotubes can be controlled. For PA concentrations of 1 wt %, it is even possible to trap functionalized nanotubes in a peptide amphiphile aqueous gel. Inspection of the PA-functionalized MWNTs by TEM confirms the existence of an organic coating on the exterior sidewall of the outermost nanotube shell. Additionally, from the analysis of optical absorbance spectra, we can conclude that aqueous solutions of peptide amphiphile functionalized SWNTs are rich in isolated tubes that are not appreciably covalently modified. Future studies requiring the absorption of peptides onto carbon nanotubes may benefit from this strategy of coupling a hydrophobic alkyl tail to a more hydrophilic peptide sequence. Also, this approach to functionalizing nanotubes with peptide amphiphiles can likely be translated to other carbon nanostructures such as carbon onions and carbon scrolls, as well as to other nonpolar, water-insoluble nanoscale materials. Furthermore, by using derivatives of these peptide amphiphiles containing bioactive sequences for the specific binding of proteins and other analytes (such as PA 3), it may be possible to create encapsulated carbon nanotubes for applications in biosensing and medicine. Acknowledgment. This work was supported by the National Science Foundation under NSF Award Nos. DMR-0134706 and EEC-0118025 and the Department of Energy under Award No. DE-FG02-00ER45810/A001. Support from a Beckman Young Investigator Award (M.C.H.) and a National Defense Science and Engineering Graduate Fellowship (M.S.A.) is gratefully acknowledged. We graciously thank the EPIC facility for experimental assistance and for use of their transmission electron microscopes. We also thank Randal C. Claussen for preparing L-aspartic acid di-tert-butyl ester and Samantha Cruz for preparation of SWNT samples. LA0469452 (38) Strano, M. S.; Moore, V. C.; Miller, M. K.; Allen, M. J.; Haroz, E. H.; Kittrell, C.; Hauge, R. H.; Smalley, R. E. J. Nanosci. Nanotechnol. 2003, 3, 81-86. (39) Hu, H.; Zhao, B.; Hamon, M. A.; Kamaras, K.; Itkis, M. E.; Haddon, R. C. J. Am. Chem. Soc. 2003, 125, 14893-14900.
