Plasmonic Nanorods Provide Reversible Control over Nanostructure

pubs.acs.org/Langmuir © 2010 American Chemical Society

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Plasmonic Nanorods Provide Reversible Control over Nanostructure of Self-Assembled Drug Delivery Materials Wye-Khay Fong,† Tracey L. Hanley,‡ Benjamin Thierry,§ Nigel Kirby, and Ben J. Boyd*,† †

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Drug Delivery, Disposition and Dynamics, Monash Institute of Pharmaceutical Sciences, Monash University (Parkville Campus), 381 Royal Parade, Parkville, Victoria 3052, Australia, ‡Bragg Institute, Australian Nuclear Science and Technology Organisation, Menai, NSW 2234, Australia, §Ian Wark Research Institute, University of South Australia, Mawson Lakes, SA 5095, Australia, and SAXS/WAXS beamline, Australian Synchrotron, Clayton, Victoria, Australia Received February 11, 2010. Revised Manuscript Received March 11, 2010 The nanostructure of mesophase liquid crystals prepared from amphiphilic lipids controls the rate of release of incorporated agents from the material, such as drug molecules, and reversible transition between different nanostructures essentially provides an “on-off” switch for release (Fong, W.-K.; Hanley, T.; Boyd, B. J. J. Controlled Release 2009, 135, 218-226). In this study, the incorporation of plasmonic hydrophobized gold nanorods (GNRs) permits reversible manipulation of nanostructure on-demand, by irradiation of the matrix using a near-infrared laser. Synchrotron small-angle X-ray scattering was used to probe the kinetics of the response of nanostructure to laser irradiation, and the specificity of the approach is shown by the lack of response in the absence of nanorods, or for GNR whose dimensions are not matched to the specific wavelength of the incident light.

Light sensitive systems are of increasing interest to researchers in applications such as drug delivery due to the noninvasive and remote nature by which the stimulus can be applied. Numerous light sensitive self-assembled systems including polymeric micelles, gels, liposomes, and nanocomposites have been reported and reviewed.2,3 However, their translation into products, particularly in the drug delivery field, has thus far been limited in part by the need to incorporate dyes or other light sensitive components of unknown toxicity. Lyotropic liquid crystals are receiving increasing interest as stimuli responsive materials for drug delivery. The self-assembled nanostructures are often thermodynamically stable in excess water4 and comprise discrete lipidic domains and aqueous channels which allow the incorporation of molecules of varying physicochemical properties.5-11 The rate of release of a drug is dictated by its size relative to that of the aqueous channels which in turn is dictated by the overall geometry of the liquid crystalline structure, and the specific local packing of the amphiphilic lipids comprising the matrix. The inverse bicontinuous cubic and hexagonal phases (denoted v2 and H2, respectively), and inverse micellar phase (L2), illustrated in Figure 1, are of particular interest in drug delivery. *To whom correspondence should be addressed. Telephone: þ61 3 99039112. Fax: þ61 3 99039583. E-mail: [email protected].

(1) Fong, W.-K.; Hanley, T.; Boyd, B. J. J. Controlled Release 2009, 135, 218–226. (2) Alvarez-Lorenzo, C.; Bromberg, L.; Concheiro, A. Photochem. Photobiol. 2009, 85, 848–860. (3) Christie, J. G.; Kompella, U. B. Drug Discovery Today 2008, 13, 124–134. (4) Kaasgaard, T.; Drummond, C. J. Phys. Chem. Chem. Phys. 2006, 8, 4957–4975. (5) Drummond, C. J.; Fong, C. Curr. Opin. Colloid Interface Sci. 1999, 4, 449–456. (6) Lee, K. W. Y.; Nguyen, T.-H.; Hanley, T.; Boyd, B. J. Int. J. Pharm. 2009, 365, 190–199. (7) Shah, J. C.; Sadhale, Y.; Chilukuri, D. M. Adv. Drug Delivery Rev. 2001, 47, 229–250. (8) Amar-Yuli, I.; Libster, D.; Aserin, A.; Garti, N. Curr. Opin. Colloid Interface Sci. 2009, 14, 21–32. (9) Boyd, B. J.; Khoo, S.-M.; Whittaker, D. V.; Davey, G.; Porter, C. J. H. Int. J. Pharm. 2007, 340, 52–60. (10) Boyd, B. J.; Whittaker, D. V.; Khoo, S.-M.; Davey, G. Int. J. Pharm. 2006, 318, 154–162. (11) Cervin, C.; Vandoolaeghe, P.; Nistor, C.; Tiberg, F.; Johnsson, M. Eur. J. Pharm. Sci. 2009, 36, 377–385.

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Transitions between these structures can be induced through changes in lipid packing. Temperature has been used as a stimulus to change liquid crystalline structure between the v2 and H2 structures in vitro and in vivo, demonstrating the potential of these systems as stimulus responsive delivery systems. Reversible switching between the v2 and H2 phase structures formed by the amphiphilic lipid 3,7,11,15-tetramethyl hexadecyl-1,2,3-triol (phytantriol, see Figure 1) using temperature provided a means to manipulate drug release.1 However, the matrix required the inclusion of a modifier (vitamin E acetate) to reduce the transition temperature to close to physiological temperature for in vivo application, thereby enabling control over structure by application of, for example, a heat pack to the skin surface after subcutaneous administration. This is a major limitation, as there is no specificity in heat source; hence, exposure to extremes of temperature may unintentionally induce drug release. Consequently, an alternative means to induce the phase transition was necessary that did not require direct heating, and did not require a reduction in the temperature at which the transition occurs, ideally occurring at the inherent transition temperature without additive (approximately 55 °C for phytantriol), removing the potential for accidental activation. The incorporation of azocontaining surfactants was considered, but this has only been reported for modifying the ordering in lamellar phases rather than transitions between other liquid crystalline structures.12,13 Toward the design of an advanced drug delivery system based on light-triggered phase transition of liquid crystalline phases, we report here the design of novel liquid crystalline matrix-gold nanorod hybrid materials. Hydrophobized gold nanorods (GNRs) have been incorporated within the liquid crystalline matrix to provide remote heating, and trigger the phase transitions on irradiation at close to their resonant wavelength. The surface of plasmonic metal nanoparticles delivers heat into surrounding material on (12) Eastoe, J.; Vesperinas, A. Soft Matter 2005, 1, 338–347. (13) Zou, A.; Eastoe, J.; Mutch, K.; Wyatt, P.; Scherf, G.; Glatter, O.; Grillo, I. J. Colloid Interface Sci. 2008, 322, 611–616.

Published on Web 04/01/2010

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Figure 1. Ensemble of the self-assembled structures commonly encountered with amphiphilic lipid systems in excess water and the structure of phytantriol, the lipid used in the current report. Hydrophilic regions incorporating water are between the headgroups, while the hydrophobic regions comprise the branched alkyl tails. For phytantriol, heating induces the v2 f H2 f L2 transition.

exposure to an appropriate light source at the plasmon resonance. The application of near-infrared sensitive nanorods has significance in designing systems for ophthalmic, subcutaneous, or deeper tissue applications, as near-infrared radiation has been reported to penetrate tissues up to 50 mm deep.14,15 For GNRs, the wavelength of light that induces the plasmon resonance can be tuned by varying the size and aspect ratio of the nanoparticle.16,17 Of most use for drug delivery are nanoparticles which absorb and respond in the near-infrared light therapeutic window, that is, wavelengths where light has its maximum depth of penetration in tissues. The efficient and localized generation of heat resulting from the strong resonant absorption of light by colloidal plasmonic nanostructures is under intense investigation toward photothermal ablation of solid tumors.18 Plasmonic nanoparticles have also been incorporated into a range of materials with a view to drug delivery, including polymer microgels,19,20 liposomes,21 and microcapsules,22 with application of a laser light source allowing remote release of encapsulated materials. Externally triggered reversible phase transitions have not been achieved in lipid-based systems; hence, the use of plasmonic nanoparticles to achieve this important breakthrough offers a novel practical solution. Gold nanorods with an average aspect ratio of 3.3 (length, 55.5 ( 9.5 nm; width, 16 ( 2.5; see the Supporting Information) were synthesized through control of experimental conditions as described previously.23 The as-synthesized cetyl trimethyl ammonium bromide (CTAB)-capped nanorods displayed a longitudinal surface plasmon resonance (LSPR) at 780 nm in water. Dodecanethiol coated nanorods were then prepared using a two-step hydrophobization procedure,23 which resulted in a red-shift of the LSPR to 820 nm when resuspended in chloroform (Figure SI 1, Supporting Information). The hydrophobized nanorods were incorporated into the phytantriol matrix by combining solutions of the lipid with GNR dispersions, both in tetrahydrofuran (THF), at different concentrations, and then removing the THF under (14) Stolik, S.; Delgado, J. A.; Perez, A.; Anasagasti, L. J. Photochem. Photobiol., B 2000, 57, 90–93. (15) Barun, V.; Ivanov, A.; Volotovskaya, A.; Ulashchik, V. J. Appl. Spectrosc. 2007, 70, 430–439. (16) Huang, C.-p.; Yin, X.-g.; Huang, H.; Zhu, Y.-y. Opt. Express 2009, 17, 6407–6413. (17) El-Brolossy, T. A.; Abdallah, T.; Mohamed, M. B.; Abdallah, S.; Easawi, K.; Negm, S.; Talaat, H. Eur. Phys. J. Spec. Top. 2008, 153, 361–364. (18) Schwartz, J. A.; Shetty, A. M.; Price, R. E.; Stafford, R. J.; Wang, J. C.; Uthamanthil, R. K.; Pham, K.; McNichols, R. J.; Coleman, C. L.; Payne, J. D. Cancer Res. 2009, 69, 1659–1667. (19) Das, M.; Mordoukhovski, L.; Kumacheva, E. Adv. Mater. 2008, 20, 2371– 2375. (20) Das, M.; Sanson, N.; Fava, D.; Kumacheva, E. Langmuir 2007, 23, 196– 201. (21) Paasonen, L.; Laaksonen, T.; Johans, C.; Yliperttula, M.; Kontturi, K.; Urtti, A. J. Controlled Release 2007, 122, 86–93. (22) Angelatos, A. S.; Radt, B.; Caruso, F. J. Phys. Chem. B 2005, 109, 3071– 3076. (23) Thierry, B.; Ng, J.; Krieg, T.; Griesser, H. J. Chem. Commun. 2009, 1724–1726.

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vacuum. Addition of water produced the bicontinuous cubic phase matrix at ambient temperature. Synchrotron small-angle X-ray scattering (SSAXS) allows the determination of structural evolution in these systems; in particular, the position of peaks in the intensity versus scattering vector (q) profile at each temperature allows the calculation of the lattice parameter for the structural unit at that temperature. Determination of the lattice parameter versus temperature behavior of the matrix under controlled temperature conditions provides a “calibration” plot, from which the apparent matrix temperature can be obtained after irradiation in the presence or absence of GNRs. Figure 2A shows the calibration plot of lattice parameter versus temperature determined by heating the matrices under thermostat control. The transition from v2 to H2 phase occurs from approximately 50 °C and from H2 to L2 at approximately 65 °C, consistent with previous studies in the absence of GNR.24 The first notable feature is that the lattice parameter in the presence of nanorods was independent of GNR concentration. This implies that, at least at the concentrations of GNRs that were employed in these studies, the presence of the GNRs does not substantially change the phase structure of the liquid crystal and that the calibration curves can be applied to any GNR concentration up to 3 nM. Although the presence of nanorods did not change the lattice parameter of a particular phase structure at specific temperatures, Figure 2B shows that there is a subtle effect on the temperature at which the phase transitions occur on heating. There is a slight reduction in the transition temperature from the v2 to H2 phase with increasing concentration. It has been proposed that an overall increase in packing frustration in the transition from v2 to H2 phase, due to space filling requirements between lipid chains in the H2 phase, renders this an unfavorable process.25 Hydrophobic additives can relieve this packing frustration, thereby lowering the overall temperature required for the transition to occur. Thus, it is reasonable to suggest that the hydrophobic nanorods are interacting with the hydrophobic chain regions of the nanostructure, leading to the observed reduced transition temperature. It is nevertheless a subtle effect, and broadly speaking the GNRs have not disrupted or substantially modified the behavior of the liquid crystalline nanostructure in the absence of laser irradiation. In order to investigate the effect of irradiation of the GNR on the temperature and nanostructure of the liquid crystalline systems, an 808 nm NIR diode laser was mounted 20 cm from the sample at a tangential angle to the X-ray beam and was remotely operated through a computer control system. Scattering patterns were accumulated and recorded every 100 ms. A series of (24) Dong, Y.-D.; Larson, I.; Hanley, T.; Boyd, B. J. Langmuir 2006, 22, 9512– 9518. (25) Seddon, J. M. Biochim. Biophys. Acta, Rev. Biomembr. 1990, 1031, 1–69. Perutkova, S.; Daniel, M.; Dolinar, G.; Rappolt, M.; Kralj-Iglic, V.; Iglic, A. Advances in Planar Lipid Bilayers and Liposomes, Liu, A. L., Ed.; Elsevier: 2009; Vol. 9, pp 237-278.

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Figure 2. Effect of gold nanorods (GNRs) on equilibrium structure of the phytantriol þ water system with controlled temperature from SAXS experiments. (A) Lattice parameter of the phytantriol þ water system in the presence of 0 mM (b), 1.5 nM (9), or 3 nM (1) GNRs. (B) Influence of GNR concentration on the transition temperatures of the phytantriol þ water system (blue region = v2 phase, green = H2, yellow = L2, hashed regions are mixed phase).

experiments were performed with varying pulse duration, and repeat pulse experiments allowed investigation of the reversibility of the system. Representative scattering profiles over time, obtained using a 5 s on/5 s off sequence, are illustrated in the Supporting Information. In the absence of nanorods the structure observed is the v2 phase (Figure SI 2A), with “double diamond” or Pn3m space group. This phase structure has been reported for the phytantriol þ water system at temperatures below 55 °C. The “q” value of the main peak (1.29 nm-1) is invariant with irradiation indicating that no heating of the matrix was occurring, and is consistent with previous studies.24 Irradiation of the system, on inclusion of a low concentration of GNRs (0.3 nM) in the matrix, did not induce a change in phase structure away from the Pn3m bicontinuous cubic phase (Figure SI 2B). However, the shift in peak position to higher q values indicates a shift in lattice dimension to a smaller value, which in turn indicates, from Figure 2, that heating of the matrix has occurred. The structure relaxed back to the original position when the laser was off. On repeated application of the 5 s laser pulse, the system again displayed the heating effect and relaxed back to the starting position when irradiation was complete. This contraction and expansion of the lattice on heating and cooling has been termed the “ breathing mode” by de Campo, as it is accompanied by concurrent expulsion and uptake of water from the matrix to satisfy the changes in lattice dimension.26 Thus, the heating effect is both reversible and reproducible, but at 0.3 nM GNR concentration it was not sufficient to convert the phase nanostructure to the H2 or L2 phases that exist at higher temperatures. In the case of higher concentrations of GNR, the 5 s on pulse for both the 1.5 and 3 nM systems did induce a phase change to the H2 and L2 phase structures. At 3 nM, complete transformation to the L2 phase occurred by the end of the 5 s irradiation, while the lower 1.5 nM concentration resulted in a mixed H2 þ L2 phase, indicating a reduced heating effect. Again, when the laser was switched off, the system ultimately returned to the initial v2 (Pn3m) phase structure. Interestingly, on conversion from the L2 or L2 þ H2 state back to the v2 structure, the “gyroid” bicontinuous cubic phase with Ia3d space group was encountered. The gyroid phase coexisted with the H2 phase initially and then with the v2 (Pn3m) phase for approximately 5-6 s after the laser was switched off. This is highly unusual, as the samples are present in excess water and the gyroid phase only exists at equilibrium in dehydrated liquid (26) de Campo, L.; Yaghmur, A.; Sagalowicz, L.; Leser, M. E.; Watzke, H.; Glatter, O. Langmuir 2004, 20, 5254–5261.

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Figure 3. Effect of laser irradiation on apparent temperature of the phytantriol þ water matrix (Tapp) with change in GNR concentration. Tapp was derived from lattice dimensions in Figure SI 2 in the Supporting Information, using the calibration data from Figure 2. [GNR] = 0 nM are white symbols, 0.3 nM blue symbols, 1.5 nM yellow symbols, and 3 nM black symbols. Circles indicate v2 phase, triangles indicate v2 þ H2, squares indicate H2 þ L2, and diamonds indicate L2. The horizontal lines indicate the typical equilibrium phase boundaries derived for the data in Figure 2. The cartoon on the right indicates the type of phase structure present with increasing temperature (v2, H2, and L2, adapted from ref 30).

crystalline structures for the phytantriol þ water system.24 No gyroid phase was observed during the heating phase of the cycle. The existence of the gyroid phase on rapid cooling of the phytantriol þ water system has been recently reported in studies using T-drop experiments.27 It was hypothesized that the contraction of the lattice structure traps the system in a temporarily dehydrated state, which requires water flux back into the matrix to re-establish the equilibrium double diamond cubic phase structure. The q values were converted to apparent sample temperature (Tapp) using the calibration curves in Figure 2, and the resulting plot of Tapp versus time is illustrated in Figure 3. In the case where no GNRs were added to the matrix, there was no significant change in Tapp upon laser irradiation. The reversibility of the heating effect in the presence of the GNR is evident from the Tapp profiles for the three samples containing the nanorods. The sample containing 0.3 nM GNR heated to approximately 50 °C, just below the transition above which coexisting H2 phase occurs. (27) Dong, Y.-D.; Tilley, A. J.; Larson, I.; Lawrence, M. J.; Amenitsch, H.; Rappolt, M.; Hanley, T.; Boyd, B. J. Langmuir 2010, accepted; DOI: 10.1021/ la904803c.

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Increasing the nanorod concentration to 1.5 nM induces heating to approx 70 °C, while 3 nM GNR heated the matrix to an apparent temperature of 75 °C. The repeat irradiation provided the same peak temperature within 1-2 °C and reproducible kinetics of heating and cooling. The phase structures observed at the calculated Tapp values closely agreed with the equilibrium structures apparent from the calibration plots at those temperatures, although H2 þ L2 coexisting phases were observed for a number of samples in Figure 3 (squares) at 65-75 °C which was not apparent in the equilibrium behavior in Figure 2. The coexisting L2 phase in the H2 þ L2 system on cooling at the higher concentration of GNR was evident at temperatures below the equilibrium heating transition temperature, indicating a kinetic supercooling effect. This effect has also been observed in the T-drop experiments with the phytantriol þ water system,27 and was also attributed to the generally unfavorable conversion to the H2 phase. The heating effect observed in these experiments is clearly a function of nanorod concentration, although the relationship between concentration and maximum temperature at differing irradiation conditions requires further investigation and is likely complicated by concurrent cooling. The “breaking wave” shape of the profiles, indicating nonlinear heating/cooling gradients in the material, is consistent with previous studies on heat transfer from gold nanoparticles into and out of a bulk system.28,29 The dissipation of resonant energy is dependent on the particle dimensions, laser power, and environmental conditions, such as heat capacity of the material, in a process which occurs in pico- to nanoseconds across the particle-matrix interface. The heat transfer from the GNR to the matrix appears to plateau, and then, on removal of the heat source, cooling of the system is determined by bulk heat

Acknowledgment. This research was undertaken on the SAXS/WAXS beamline at the Australian Synchrotron, Victoria, Australia, and we acknowledge the Australian Institute of Nuclear Science and Engineering for funding under AINGRA09120.

(28) Hu, M.; Hartland, G. V. J. Phys. Chem. B 2002, 106, 7029–7033. (29) Roper, D. K.; Ahn, W.; Hoepfner, M. J. Phys. Chem. C 2007, 111, 3636– 3641. (30) Holmberg, K.; J€onsson, B.; Kronberg, B.; Lindman, B. Surfactants and Polymers in Aqueous Solution, 2nd ed.; John Wiley & Sons Ltd.: Chichester, 2002; 562 p

Supporting Information Available: Details of gold nanorod synthesis, hybrid material preparation, and small-angle X-ray scattering. This material is available free of charge via the Internet at http://pubs.acs.org.

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diffusion in the matrix and as a function of the difference between sample temperature and ambient temperature.28 Interestingly, the inclusion of a thermocouple in the sample showed an increase of only 1-2 °C in the macroscopic temperature of the matrix, even after 10 repeated on/off cycles at high nanorod concentrations, indicating that the heating effect giving rise to structural changes is a nanoscopic rather than macroscopic heating effect. This finding has important repercussions in the ultimate use of these materials in drug delivery, as the localized heating of adjacent tissue structures will not occur if the effect translates into the in vivo behavior of the material. To test the specificity of the prepared hybrid materials, GNRs of different aspect ratios were also investigated. Irradiation at 808 nm of the liquid crystalline matrix containing 3.0 nM GNRs with a resonant wavelength of 660 nm (aspect ratio = 2.6) did not induce a heating effect, demonstrating the specificity of the plasmonic heating effect. In conclusion, GNRs embedded in liquid crystalline matrices produce localized plasmonic heating of the hybrid matrix, enabling fine control over nanostructure. The phase transitions resulting from photothermal heating were fully reversible and specific to the GNR/laser wavelength combination. Localized plasmonic heating of the liquid crystal did not compromise the integrity of the lipid molecules in any of the mesophases. These findings represent a significant advance toward effective, light-activated drug delivery systems with potential to solve unmet medical needs.

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