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May 3, 2011 - In this work, we have employed a suite of complementary analytical techniques to shed light on the nanocomposite structures formed durin...
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New Insights into the Structure of PAMAM Dendrimer/Gold Nanoparticle Nanocomposites Lee W. Hoffman, Gunther G. Andersson,* Anirudh Sharma, Stephen R. Clarke, and Nicolas H. Voelcker* Centre for Nanoscale Science and Technology, School of Chemical and Physical Sciences, Flinders University, Adelaide, South Australia, 5001 ABSTRACT: In this work, we have employed a suite of complementary analytical techniques to shed light on the nanocomposite structures formed during gold nanoparticles (AuNPs) synthesis in the presence of poly(amidoamine) (PAMAM) dendrimers. Nanocomposites of AuNPs and either fourth or eighth generation amine-terminated PAMAM dendrimers (G4 or G8) were prepared. The size distributions of AuNPs and the nanocomposites were determined by transmission electron microscopy. Atomic force microscopy phase imaging and neutral impact collision ion scattering spectroscopy (NICISS) were utilized for the first time to investigate and compare nanocomposite structures formed from G4 and G8. Our results suggest that G4 stabilized the AuNP by capping the AuNP particle surface but that a certain fraction of the gold surface was still barely covered. In contrast, the metal nanoparticle surface was completely covered by G8. In addition, NICISS results provided evidence that nanocomposites deformed when being deposited directly onto a substrate.

’ INTRODUCTION Interest in poly (amidoamine) (PAMAM) dendrimers and metal nanoparticles formed in the presence of PAMAM dendrimers has resulted in recent years in more extensive characterization of these materials, which in turn has led to a better understanding of nanoparticle formation.1 14 As nanotechnology has developed over the past several decades, interest in the properties and capabilities of the zerovalent metal nanoparticles formed by single metal ions or various derivatives obtained through combination of different metals has been steadily increasing. While the formation of gold nanoparticles (AuNPs) in the presence of PAMAM dendrimers has been the subject of several studies over the past few decades (e.g., refs 15 18), most of the work has concerned larger generation dendrimers (i.e., > sixth generation). In addition, there has been much effort devoted to the modification of the periphery of PAMAM dendrimers to help facilitate nanoparticle encapsulation.19 22 A significant dependence of the nanocomposite structure on the dendrimer generation has been observed. While AuNPs are known to form inside the dendrimer for PAMAM dendrimers of sixth generation or higher, those of smaller generations (4th or less) create complexes where AuNPs are formed and their surfaces are subsequently capped by these lower generation PAMAM dendrimers (Figure 1).23,24 These observations have sparked interest in a better understanding of the mechanism of nanocomposite formation and in suitable models to describe and predict the experimental results.6 Much of the previously reported evidence has been provided through transmission electron microscopy (TEM) and small angle neutron and X-ray scattering (SANS and SAXS, respectively)15,16,25 or work involving, UV vis, Fourier Transform Infrared Spectroscopy (FT-IR), ζ potential measurements, nuclear magnetic resonance (NMR), and matrix-assisted laser desorption/ionization time-of-flight spectroscopy (MALDITOF).24,26,27 Of the above methods, SANS and SAXS provides the r 2011 American Chemical Society

most comprehensive picture of the structure of the nanocomposites. However, for analysis of the SANS and SAXS data, the measurements have to be fitted to models, where the models have to be selected before the fit can be made. As a result, only the location of where the AuNP is situated with respect to the dendrimer has been reported but not the thickness of the layer covering the AuNPs. These investigations have led to conclusions where AuNPs are formed in different dendrimer generations. In any case, it is presumed that the AuNP is completely covered by the dendrimer. To our knowledge, studies on the resulting surfaces or outer shell of these materials (i.e., dendrimer/ AuNP nanocomposite) and the extent to which the dendrimer encapsulates the AuNP have not yet been reported. For this work, our interest is in the agglomeration of the nanocomposites as gold nanoparticles are formed, the size distribution of these nanoparticles and nanocomposites, and how the structure of the nanocomposites is changing when placed on a flat surface. In order to gain an advanced understanding of nanocomposite formation, we have performed microscopy experiments using atomic force microscopy (AFM) including phase imaging and transmission electron microscopy (TEM) in combination with the depth profiling capabilities provided by neutral impact collision ion scattering spectroscopy (NICISS). AFM phase imaging is a wellestablished surface-sensitive technique that has been used for imaging changes in heterogeneous surfaces,28 30 and has recently been used to study the surface morphology of Langmuir Blodgett monolayers of amphiphilic polybutadiene-poly(ethylene glycol).31 More recently, NICISS has successfully been used to determine the structure of the shell of polymer stabilized platinum nanoparticles in an ionic Received: December 25, 2010 Revised: March 8, 2011 Published: May 03, 2011 6759

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Figure 1. Illustration of the complexation of gold ions with 4th generation amine-terminated (G4) PAMAM dendrimer in aqueous solution followed by addition of reducing agent, leading to the formation of dendrimer-capped, zerovalent AuNP. The AuNP are not formed inside the G4 PAMAM dendrimer.

liquid.32 The combined use of AFM and TEM allows for the determination of the size distribution as well as elucidation of the resulting agglomeration of these nanoparticles and nanocomposites. The combination of phase imaging AFM and NICISS allows the detailed analysis of the distribution of PAMAM and the AuNPs within the nanocomposites when adsorbed on a surface. The aim of this work is to study nanocomposites formed when AuNPs are synthesized in the presence of PAMAM dendrimers (dendrimer/AuNP nanocomposites). The formation of the nanocomposites depends on the size of the dendrimer and we investigate nanocomposites formed by the fourth generation and the eighth generation, amine terminated PAMAM dendrimers (G4 and G8, respectively). We describe how the nanoparticles are stabilized and to what extent the gold nanoparticle surface is covered with dendrimer after nanoparticle formation. In addition, this work provides information on how the size of the dendrimer determines whether the AuNPs are formed inside or outside the dendrimers. Employing AFM and NICISS on both G4 and G8 nanocomposites has resulted in direct information about the positioning of the AuNPs within the nanocomposites. Finally, we investigate how the nanocomposites are deformed when placed on silicon surfaces.

’ EXPERIMENTAL SECTION Materials. G4 and G8 amine-terminated PAMAM dendrimers were purchased from Dendritech, Inc., Midland, MI, USA, (10.0% w/w and 4.95% w/w in MeOH, respectively). Tetrachloroauric (III) acid trihydrate (HAuCl4 3 3H2O, g99.9%) and sodium borohydride (NaBH4, 98%) were purchased from Sigma-Aldrich Chemical Co. Pty Ltd., Castle Hill, NSW, Australia and used without further purification. Sodium hydroxide (NaOH, 99.0%) was purchased from Hem-Supply Pty Ltd., Port Adelaide, SA, Australia, and used without further purification. Milli-Q water (18.2 MΩ 3 cm) was obtained via a Labconco WaterPro PS and used in all experiments. Any reagents used were of analytical grade or better. Gold Nanoparticle Formation in the Presence of PAMAM Dendrimer. Molar ratios of gold ions:dendrimer of 55:1 and 480:1 were used for G4 and G8, respectively. The molar ratio was set according to the number of peripheral amine groups on the dendrimers. Synthesis of Dendrimer/AuNP Nanocomposites. G4/AuNP and G8/AuNP were prepared in similar manner. A 12.5-mL portion of Milli-Q water was placed in a 25 mL round-bottom flask. To this solution, approximately 150 mg of G4 or 174.5 mg of G8 solution was added and stirred for 15 min to ensure complete distribution of dendrimer throughout the solution. A dilute solution of Au(III) ion was then added to the dendrimer solution following a procedure developed by Kim et al.19 This solution was stirred for 1 h to ensure proper complexation of Au ion with dendrimer. Finally, a dilute solution of sodium borohydride in sodium hydroxide was quickly added to reduce the Au ion to Au NPs. Through this process, the solution changed from bright yellow to dark yellow upon complexation with PAMAM dendrimers and to red upon addition of the

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reducing agent. Dendrimer/AuNP nanocomposites were removed from the salt solution by centrifugal ultrafiltration. A Sigma 3 18K laboratory centrifuge was operated at 5000 RCF, using Satorius Stedim Biotech Vivaspin 2 (10,000 MW cutoff) centrifuge tubes. The nanocomposites were taken up into Milli-Q water. Synthesis of AuNPs. For AFM analysis, AuNPs were prepared similar to what has been described above. However, no dendrimer or any other stabilizer was used. A 40-mL portion of Milli-Q (18.2 MΩcm) water was placed in a 50 mL round-bottom flask. A dilute solution of Au(III) ion was added. Finally, a dilute solution of sodium borohydride in sodium hydroxide was quickly added and the Au ion was immediately reduced to Au NPs. An aliquot of AuNP was removed, further diluted with Milli Q water, followed by 4 μL placed on a freshly cleaved mica surface. Atomic Force Microscopy. Tapping-mode AFM (TM-AFM) were performed on a Multimode Nanoscope IV controller (Veeco Instruments) utilizing the “E” scanner. TM experiments were performed under ambient conditions, and both height and phase data were simultaneously recorded. Commercially available cantilevers (Veeco Probes, FESP) were employed, with a nominal tip radius on the order of 8 10 nm. Images were typically recorded with scan rates averaging 2 Hz, an amplitude set point approximately 80% of the free cantilever amplitude. Initially, a quick scan of 5  5 μm area was collected, followed by smaller scan sizes in the desired regions of interest. Computer analysis of raw data was performed using NanoScope (Digital Instruments) software, version 5.31. Transmission Electron Microscopy. TEM data were collected on a Jeol 200EX instrument with a Megaview 3 Digital Camera. Carboncoated copper grids, 100-mesh, were used as received. A dilute solution of PAMAM/AuNP nanocomposites was placed on the grid and allowed to evaporate under ambient conditions. The substrate was then inverted and the sample was placed in contact with an aqueous solution of phosphotungstic acid solution (2% w/w), The grid was then blotted with filter paper and allowed to dry. TEM images were obtained at 80 kV first using a low resolution to find an area of interest. Final TEM images were obtained using magnifications of 100 000 and 300 000.

Neutral Impact Collision Ion Scattering Spectroscopy. Solutions of dendrimer/AuNP nanocomposite were applied to clean silicon wafers, resulting in a variety of sample thicknesses ranging from less than a monolayer to multiple layers. The measurements were averaged over a sample size of about a few square millimeters to keep the damage to the sample due to the impact of the ions low. The helium ion dose was kept below 5  1013 ions/cm2. Equipment used in this investigation is similar to that described in ref 33 using 3 keV helium ions. A Leybold-Heraeus ion source with a Wien mass filter from SPECS for masses separation was used to generate ion beams with energies from 1 to 5 keV inert gas ions. Two pairs of deflection plates in the ion source and mass filter were used to pulse the ion beam. Applying pulsed voltages with a repetition frequency of about 50 kHz to the deflection plates, the ion beam was swept over an aperture generating short ion pulses of about 50 ns. The target was positioned on a mechanical manipulator unit that allows for in situ vertical and horizontal alignment of the target with an operational range of several centimeters. The time-of-flight (TOF) of backscattered projectiles from the target to the detector was recorded by a multichannel analyzer (MCA). The detector consists of a set of channel plates. The angle between the ion beam and TOF path was 168°. The TOF detector formed a cone with a dihedral angle of 0.5°.

’ RESULTS AND DISCUSSION Size and Location of AuNPs. Figure 2(A) shows a representative TEM micrograph of G4/AuNP nanocomposites stained with phosphotungstic acid to help visualize the PAMAM component. Figure 2(C) shows a histogram of nanoparticle size compiled from at least 50 measurements each from three different TEM micrographs. As can be observed in Figure 2(C), 6760

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Figure 2. TEM image of G4 PAMAM/AuNP nanocomposites (A) and corresponding gold nanoparticle size analysis (C) and TEM image of G8 PAMAM/AuNP nanocomposites (B) with corresponding gold nanoparticle size analysis (D).

AuNPs averaging 2.7 nm, with a range of 1.5 8.0 nm can be observed when formed in the presence of G4. The “tail” in the histogram was produced in large part by a second size distribution centered at 5.5 nm, or roughly double that of the major size distribution. This phenomenon is attributed to formation of agglomerates. Agglomeration is attributed to hydrogen bonding between the peripheral NH2 groups of the PAMAM dendrimers, or interaction of the dendrimer branches as previously reported in ref 34. G4 dendrimers do not allow much freedom in the conformation of the dendrimer branches due to the short length of the branches which results in dendrimers with a large number of NH2 groups at the outside of the dendrimer. As the NH2 groups show strong interaction and the ability for dendrimer arms to interdigitate, agglomerates are formed for G4 deposited on surfaces. The size of the PAMAM dendrimer surrounding the AuNP (the gray region of 3.0 4.0 nm diameter surrounding the darker AuNPs in Figure 2(A)) is slightly smaller than the reported diameter of 4.5 nm for a G4. The reduced size is attributed to the deformation of the dendrimer upon binding to the AuNP (i.e., the spherical conformation of the dendrimer in solution “flattens” as the NH2 groups bind to the Au surface). The overall size of a single G4/AuNP nanocomposite particle ranges from 7.0 to 15.0 nm. These measurements strongly suggest that dendrimers surrounded the AuNP, where the size of the AuNP ranges from 1.7 to 7.9 nm, as shown in Figure 2(C), rather than the AuNP forming within the dendrimer interior, consistent with results reported in earlier studies.35,36 We also investigated nanocomposites formed in the presence of G8. Samples were prepared similar to those described above for the G4, except that the dendrimers were not stained. As can be seen in Figure 2(B), the gold nanoparticles appear to be much more evenly dispersed throughout the TEM grid, with little (if any) agglomeration. Figure 2(D) shows a histogram of AuNP size (50 AuNPs were measured from four different TEM micrographs). This is in agreement with the fact that G8 branches are

more densely packed, causing the dendritic arms and the peripheral NH2 groups to fold to the inside of the dendrimers resulting in less interaction between dendrimer arms and thus less agglomeration as shown in ref 37. While a majority of particles formed are around 2.8 nm in diameter, there are peaks in the histogram at larger particle sizes around 5.3, 7.3, and 8.8 nm. These larger distributions are attributed to the formation of multiple AuNPs within the G8 that have fused together, thus forming larger particles within the G8. The spherical nature of these larger AuNPs suggests that they are still single particles rather than artifacts from nanocomposite agglomeration. More important to our work is the distribution of the main peak (bulk of AuNPs). The differences in the histograms presented in Figure 2(C),(D) are consistent with different formation mechanisms for G4 and G8. In the case of G4, the AuNPs are capped by the dendrimers after formation38 leading to a monomodal distribution, whereas in the case of eighth generation dendrimers AuNP formation occurs inside the dendrimer.39 The formation inside the dendrimer may permit different sizes of particles to form or fusion of AuNPs within the G8. AFM Phase Image Analysis of G4/AuNP Nanocomposites. TM-AFM is an established technique for the nondestructive study of the morphology of materials with high resolution.40,41 Phase imaging in TM-AFM measures the phase difference between the external excitation of the vibrating probe and its response as a result of tip surface interactions.42 The magnitude of this shift is related to the local energy dissipation on the surface during the tip surface interaction.43 Heterogeneous materials of similar height show contrasts in phase images due to different energy dissipation of the material’s constituents.28,29,44,45 AFM phase imaging was applied to AuNPs formed in the presence of G4 to elucidate the size and location of the gold nanoparticles in nanocomposites adsorbed from a diluted solution (∼5 μg/mL) on a freshly cleaved mica surface. We also imaged G4 and AuNPs alone. In the first instance, obtaining a 6761

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Figure 3. AFM height (left column) and phase (right column) images of G4 (top row), G8 (middle row) PAMAM dendrimers along with individually synthesized gold nanoparticles (bottom row) all adsorbed on freshly cleaved mica. Materials for A D were prepared using identical conditions (i.e., dendrimer concentration in solution was the same).

monolayer of G4 particles on mica turned out quite difficult, as these dendrimers have a well-known tendency to aggregate upon adsorption to a surface.37,46,47 As shown in Figure 3(A), rounded features of different lateral and vertical sizes were observed. These were attributed to various states of aggregation of the dendrimer. Features in Figure 3(A) range are labeled in order of size from A to I. Z-scale measurements (heights) were obtained from raw data using Digital Instruments Nanoscope software, and ranged from 0.6 to 4.4 nm. On the basis of literature evidence, we may assume that the smallest feature observed (height of 0.6 nm) corresponds to a single G4 and the larger features are aggregates of multiple G4. The height of the smaller features was well below the accepted 4.5 nm diameter for this G4, but PAMAM dendrimers are known to flatten out upon adsorption to a surface.37,48 50 The diameters of the features in Figure 3(A) range from 25 to 100 nm. In regard to the smaller features, we believe that they are affected by tip convolution (the AFM tip radius of curvature is 10 nm), whereas the larger features are too large to be affected by tip convolution and thus the result of dendrimer aggregation. The phase signal observed for G4 (Figure 3(B)) is close to that of the underlying mica surface, although there is some phase contrast on the periphery of the feature, which is attributed to a topography effect. For G8 (Figure 3(C,D)) the measured heights ranged from 2.0 to 2.5 nm. These heights are quite consistent and close to the expected height for individual G8 on a surface. In contrast to G4, aggregation appears to be absent in G8. Without aggregation, the

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measured diameters of the G8 are consistently near 20 nm, which again is the smallest lateral feature that can be measure by an AFM tip with a radius of curvature of 10 nm. Thus, the measured values are affected by tip convolution. Previously reported research suggests that, in the case of the higher generation PAMAM dendrimers, steric hindrance results in the lowest energy conformation of the peripheral amines such that some of the amines are directed toward the dendrimer interior.51,52 The difference in aggregation behavior may be a result of this difference in surface charge. The phase signal for G8 is similar to that described above for the G4. Namely, the phase response is similar to that observed for the substrate. The results for neat AuNPs are presented in Figure 3(E,F). While these particles were formed through reduction of gold ions using the same chemistry as those formed in the presence of dendrimer, these particles were not stabilized by the primary amines on the periphery of the dendrimer. The size of AuNPs stabilized by the presence of G4 as measured by TEM was typically in the range of 1.5 8.0 nm. The AuNPs shown in Figure 3(E) have a measured height of 1.1 nm on average and are thus of comparable size to those formed in the presence of dendrimer. Since the size of these objects is small compared to the AFM tip’s radius of curvature, changes in phase response correspond to changes in material composition. When the phase response of the AFM tip to the AuNPs was measured, all parameters affecting phase response (i.e., amplitude set point and proportional and integral gains) were held constant. G4 show a small positive phase response between the AFM tip and the dendrimer, whereas AuNPs exhibit a larger negative phase response (darker region) as compared to the substrate. We considered that this contrasting phase response between the dendrimer and AuNP might allow the detection of the location of AuNPs within the nanocomposites. Analysis of the G4/AuNP nanocomposites was performed, and a typical result is shown in Figure 4(A ,B). The height of the nanocomposite [Figure 4(A)] ranged from 2.0 to 11.6 nm. This range is in agreement with the notion that various G4 form a stabilizing coating around the AuNP, thus the total height includes at least one layer of dendrimer underneath and one on top of the AuNP. This architecture is consistent with our TEM results. It is, of course, also conceivable that some dendrimers were present that were not associated with AuNPs. While TEM images identified only the AuNP core of the nanocomposite, AFM analysis detected dendrimer in addition to the AuNP core. Thus, the results reported by AFM analysis have extended to a lower range due to detection of dendrimer not associated with the AuNP core. The AFM phase image of the dendrimer/AuNP nanocomposite [Figure 4(B)] shows that there are features in the height image, which produce a negative (appearing darker than the mica substrate) response in the phase image. Black arrows highlight exemplar features. Others show a positive (appearing lighter than mica) phase response, which are denoted by white arrows. The majority of the features seen in the height images give a mixed phase response. A few of the features show only a positive phase shift (white arrowhead) or only a negative phase shift (black arrowhead). This behavior is consistent with the proposed composite structure where one constituent, the AuNP, gives a negative phase response (see Figure 3(F)) and the other, the PAMAM dendrimer, a positive one (see Figure 3(B)). Whether a positive, negative, or mixed phase response is observed would arguably depend on various factors. First, it depends on the 6762

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Figure 4. AFM height (A) and phase (B) images of G4 PAMAM/ AuNP nanocomposites. Panels (C) and (D) were collected from G8/ AuNP nanocomposites with similar instrument settings to those for data collected in (A) and (B), yet no phase response is observed. For (E) and (F), the same sample as (C) and (D) was analyzed, however the amplitude set point was reduced, resulting in the tip striking the surface with more force and allowing for the elucidation of gold nanoparticles in the G8 dendrimer via the phase image. While dendrimer weight concentration in solution was held constant for all samples, panels A/B show a larger number of nanocomposites due to the larger number of dendrimer being present (i.e., higher molar concentration for G4 than G8 due to difference in molar mass between the two dendrimers).

orientation of the nanocomposite particle. Second, the position of the AuNP within the nanocomposite influences the signal. Third, the signal is influenced by the size of the individual AuNPs, which, as the TEM images presented earlier showed (Figure 2), were dispersed in size. Any of these factors increasing the area of gold exposed to the AFM tip resulted in an increase in the ratio of negative to positive phase response within an individual composite particle. Where the AuNP was effectively shielded, only a positive phase response from the PAMAM is observed. Alternatively, we cannot exclude the possibility of PAMAM dendrimer (aggregates) persisting without central AuNP. Features that produced negative phase responses could be attributed to large and exposed AuNPs.53 55 Previous literature data and our TEM results show that the location of the AuNP within an amine-terminated PAMAM dendrimer complex varies depending on the generation of the selected dendrimer. AFM phase imaging offers an opportunity to study differences in the localization of the AuNP between G4 and G8. Figure 4 (C,D) show height and phase images of nanocomposites formed from G8. The relative tapping force (tapping amplitude set at 90% of the free vibrational amplitude) and the

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gains were held constant, allowing for the phase response to be similar between the samples shown in Figure 4. There was a marked difference in the height and phase response between G4/ AuNP and G8/AuNP. First, the height image from the G8/ AuNP sample [Figure 4 (C)] shows regular features on the order of 1.6 6.9 nm in height, with a few particles smaller than that. This range is narrower than the range of feature heights observed for G4/AuNP, and is consistent with our interpretation of the nanocomposite structure. According to our proposed structure, in G4/AuNP, dendrimers coat the AuNP and there were many possible configurations, due to the flexible nature of the G4, leading to dispersion in feature height. In contrast, for G8/AuNP, the range of different configurations on the core shell nanocomposite is limited due to the lack of potential agglomeration for the G8 as a result of the dendrimer’s having a more restricted conformation. This results in a dense packing of G8 branches as mentioned earlier. While the G4/AuNP nanocomoosites have a propensity to aggregate as shown in Figure 4 (A), this aggregation is absent in the G8/AuNP particles [see Figure 4 (C)]. Flattening effects during adsorption on the mica surface confound the dispersion in particle height, which can be applied to both G4 and G8. Therefore, the measured height range is lower than the average diameter of a G8 dendrimer in aqueous solution of 9.7 nm, which is calculated from the hydrodynamic radius.56 The measured height range of the G8/AuNP particles was similar to that reported earlier on G8 dendrimers.57 Incorporation of AuNP hence does not significantly increase the height as measured by AFM, which is again consistent with our proposed structure where the AuNP grows within the dendrimer molecule. In regard to the phase data, the absence of a prominent negative phase signal for the individual G8/AuNP particles is notable. It appears that the G8 PAMAM dendrimer effectively shielded the gold surface, so that only a small positive phase shift (corresponding to the dendrimer itself) was observed. Reducing the amplitude set point in the experiment from 90% to 50% of the free vibrational amplitude of the AFM tip resulted in the appearance of significant negative phase shifts. Lowering the amplitude set point restricts the cantilever vibration resulting in the tip striking the surface with more force. It appears that at higher applied force, the AFM tip penetrates through the dendrimer layer and interacts with the AuNP of the G8/AuNP nanocomposites. A control experiment was performed whereby the nascent G8 was analyzed using a higher applied force. No difference in phase response was observed.

’ CONCENTRATION DEPTH PROFILES DERIVED FROM NICIS SPECTRA AFM and TEM are techniques with a high lateral resolution and give insight into the topography of surfaces and shapes of particles. In the present work, information about the shape and size of the dendrimer/AuNP nanocomposites is gained. However, both methods provide only very limited information about the structure parallel to the surface normal. In order to gather information on the inner structure of the dendrimer/AuNP nanocomposites we applied the depth profiling technique NICISS. With NICISS, we investigated samples with dendrimer/ AuNP nanocomposites deposited on the surface of the substrate from solutions with various dendrimer/AuNP nanocomposites concentration: 0.56, 5.6, and 56 μg/mL. The medium concentration is the same as that used in the AFM experiments. 6763

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Figure 5. NICISS TOF spectrum of G8/AuNP nanocomposite solutions deposited from a low concentration onto a silicon substrate.

In Figure 5, a NICISS TOF spectrum of G8/AuNP nanocomposites deposited from a low concentration solution (0.56 μg/mL) onto a silicon substrate is shown. The nature of NICIS spectra are explained in detail in ref 58. Here, we briefly explain a few details. In a NICIS spectrum, helium projectiles backscattered from a sample are recorded with contributions from the elements constituting the target. The shape of the contribution of each element is given by its concentration depth profile. In Figure 5, at 3.8 μs, the onset of the signal due to helium projectiles backscattered from gold can be seen, at 4.9 and 6.0 μs, those due to silicon and oxygen and at 6.6 and 7.3 μs those due to nitrogen and carbon. In Figure 5, the signals due to the presence of carbon and gold show a peak-like structure as the AuNP/ dendrimer nanocomposites form a thin layer. The signal due to the presence of silicon and oxygen appear as a step as these elements form the substrate. The nitrogen signal is small as the cross section for backscattering from nitrogen is small and the concentration of nitrogen in these samples is considerably lower than that of carbon. For the NICISS experiments, silicon was used instead of mica as NICISS requires using a conducting or at least semiconducting substrate to avoid charging of the samples. This change in substrate did not have any significance with regard to the results we have reported here. The interaction between the dendrimers and the substrate will be not very different for mica and silicon and thus not strongly influence the results. In Figure 6, the NICISS depth profiles of gold for the G4/ AuNP and G8/AuNP nanocomposites adsorbed on silicon are shown. The procedure how to derive the concentration depth profiles for each element from an NICIS-TOF spectrum is described in ref 59. The zero mark of the concentration depth profiles is determined via the equations of energy and momentum conservation.58 The accuracy determining the zero mark via this procedure is a few Å. The more accurate procedure by measuring gas phase spectra of the respective element is not required here. We varied the concentration of the nanocomposites in solution and therefore the surface coverage in the adsorption experiment. The concentration depth profiles have not been deconvoluted as in the present case the deconvoluted profiles would be almost the same as the measured profiles. The reason is that the gradient of the concentration depth profiles is small compared to the gradient of the function that has to be used for the deconvolution. The gold concentration depth profiles are shown in absolute concentration. The profiles of the samples using low and medium

Figure 6. NICISS results of G4/AuNP and G8/AuNP nanocomposites. Top data set collected from G4/AuNP nanocomposite solutions at 0.56 μg/mL (A), 5.6 μg/mL (B), and 56 μg/mL (C) deposited on a clean silicon surface. Bottom data set collected from G8/AuNP nanocomposite solutions at 0.56 μg/mL (D), 5.6 μg/mL (E), and 56 μg/mL (F) deposited on a clean silicon surface. Units on the vertical axis are in absolute concentration. The profiles of the samples using concentrations of 0.56 μg/mL and 5.6 μg/mL are multiplied with a factor given in the legend.

concentrations (0.56 μg/mL and 5.6 μg/mL) are multiplied with a factor given in the legend as we want to compare the shape of the profiles. Comparing the concentration depth profiles of the different surface coverages, four observations can be made which inform about the structure of the AuNP/dendrimer nanocomposites adsorbed on a solid surface. 1. The shape of the gold profiles for both G4 and G8/gold nanocomplexes are almost the same for samples obtained from solutions with low and medium concentration, i.e., low and medium surface coverage. Figure 6 also shows that that the lower the concentration is, the lower the count rate of projectiles backscattered from gold thus the lower the height of the respective concentration depth profile becomes. This provides evidence that the AuNP/dendrimers form an incomplete layer on the surface as from a certain depth close to the surface the concentration of gold is decreasing (from about 60 Å for the low concentration and about 80 Å for the medium concentration). If a closed layer had formed, then varying the amount of deposited substances would only increase the range of the concentration depth profile in which gold is found and not the height. A nonclosed layer means that the silicon substrate is only partially covered with AuNP/dendrimers. Further, at low coverage of the surface with AuNP/dendrimers, the profiles show a clear maximum with the gold concentration dropping close to zero for larger depth. This concentration 6764

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agglomeration occurs or whether the AuNP/dendrimer nanocomposites are in direct contact with the surface of the silicon wafer. AuNP/dendrimers directly sitting on a silicon surface will experience a strong attraction from the substrate and deform by flattening of the nanocomposite. This results in concentration depth profiles with gold closer to the surface as observed for the low surface coverage. Previous investigations have shown that a strong interaction between peripheral NH2 groups and surfaces is responsible for dendrimer deformation at surfaces.60 This interaction results from van der Waals forces, ionic binding, chemical bonding, elastic/plastic compression, long-range electrostatic interactions, and capillary forces. AuNP/dendrimer nanocomposites on top of other AuNP/dendrimer nanocomposites or aggregates of AuNP/dendrimer nanocomposites, however, will not experience such strong attraction and will be much less deformed. Thus, the concentration depth profiles of the 5.6 and 56 μg/mL concentrations will then not show a maximum as for the isolated dendrimers of the low surface coverage at 0.56 μg/ mL but an almost constant gold concentration after the initial increase. Also the onset of the gold concentration will be shifted toward larger depth when increasing the surface coverage. Another observation is that the shape of the concentration depth profiles of G4/AuNP nanocomposites and G8/AuNP nanocomposites are very similar for low and medium surface coverage, i.e., 0.56 and 5.6 μg/mL, (curves A and B compared with curves D and E in Figure 6) even though the properties of the G4 are different: the size of G4 is smaller than that of G8 and the G8 is more rigid than a G437,48 as outlined above. Thus it may be assumed that the interaction between the silicon and the G4/AuNP nanocomposites and G8/AuNP nanocomposites dominates over the differences in properties of the G4 and G8. The discussion about the deformation AuNP/dendrimer nanocomposites must be distinguished from the question whether or not all AuNPs are covered with dendrimer material as discussed earlier. AuNPs not fully covered with dendrimer material will still not be fully covered when the AuNP/ dendrimer nanocomposites deform, while fully covered AuNPs will remain fully covered even if the AuNP/dendrimer nanocomposites deforms. In the latter case, the AuNPs will only move closer to the surface. 4. The slope for the onset of the gold concentration depth profile is almost the same for medium and high (5.6 and 56 μg/mL) concentrations of the G8 nanocomposite (curve E and F in Figure 6, center of the rising edge both at 30 Å) while in the case of the G4 nanocomposite, the onset of the gold profile of the highest surface coverage is at a larger depth (curve C in Figure 6, center of the rising edge at 60 Å) compared to the onset of the profile of the medium coverage (curve B in Figure 6, center of the rising edge at 30 Å). This finding can be understood when considering the dendrimer/AuNP nanocomposite structure: in the case of G4, the AuNPs are surrounded by the dendrimers and do not form inside the dendrimer, while in the case of G8, the AuNPs form inside the dendrimer. Thus, when the AuNP/ dendrimer nanocomposite is not deformed, in the case of G4 a thicker layer of dendrimer material than in the case of a G8 dendrimer covers the AuNP. As a consequence, when dendrimer/AuNP nanocomposites stack and are not strongly

depth profile is consistent with a single, incomplete layer of AuNP/dendrimers on the silicon surface. It also can be seen in Figure 6 that the coverage with AuNPs is larger in the case of G4 compared to G8. This corresponds to the finding in Figure 4 whereby the nanocomposite surface coverage is greater in the case of G4 versus G8. Both observances are attributed to the fact that equal solution concentrations of the nanocomposites measured in mass per volume have been used, resulting in lower molar concentrations for G8 as opposed to G4 by a factor of nearly 16. 2. In all concentration depth profiles, we find that there is a range of coverages of the AuNPs with material forming the dendrimer from barely covered (i.e., thin layer of dendrimer material) to densely covered (i.e., thicker layer of dendrimer material). AuNPs barely covered with dendrimer material result in a gold signal close to the zero mark of the depth scale while strongly covered AuNPs result in a gold signal setting on at a larger depth. As each AuNP contributes gold signal over some range of the depth scale due to the finite size of the AuNP, the gold profile appears with a finite slope at the onset of the gold profile over a range of 30 to 50 Å. Such a gradient of the slope is significantly lower compared to the case that there would be a homogeneous coverage of the AuNPs, i.e., coverage of the same thickness with dendrimer material. In the case of a homogeneous coverage of the AuNPs, only the energy resolution would contribute to the slope of the onset.The onset of the gold profile at zero penetration depth for the more dilute solutions (curves A, B, and D of Figure 6) indicates that at least some gold appears to be barely covered or uncovered by the dendrimer as the onset of the gold concentration depth profile is at zero Å. In the case where all gold would be covered with dendrimer material, the onset of the gold concentration depth profile would be shifted toward positive values of the depth scale and at a depth close to zero, no signal would be found. 3. When comparing the concentration depth profiles of samples with increasing coverage of AuNP/dendrimers (i.e., comparing the sequence A, B, C and the sequence D, E, F in Figure 6) the onset of the gold signal shifts to larger depth and the maximum becomes less pronounced. As an example, the rising edge of curve B in Figure 6 has shifted about 10 Å to larger depth compared to the rising edge of curve A. Also the maximum of the profile has shifted from 40 Å (curve A) to 70 Å (curve B). The maximum of curve B is only about 15% higher than the bulk value while the maximum of curve A is about three times larger than that of the bulk. In a similar way, we can observe for the G8 dendrimers a shift of about 15 Å of the rising edge from curve D to E, a shift of the maximum from 50 Å for curve D to 75 Å for curve E. The maximum of curve E is only about 25% higher than the bulk value, while the maximum of curve F is about two times larger than that of the bulk. These observations mean that the AuNPs shift to a larger depth in the sequence A, B, C and in the sequence D, E, F and thus are covered with a thicker layer of organic material provided by the dendrimer. The formation of agglomerates by the AuNP/dendrimers and stacking on top of each other can explain this finding. The interaction of the AuNP/dendrimer nanocomposites will depend on whether stacking or 6765

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Figure 7. Schematic proposing a simplified version of proposed agglomerate formation obtained from (A) dilute solution of G4/AuNP, (B) dilute solution of G8/AuNP, (C) concentrated solution of G4/ AuNP, and (D) concentrated solution of G8/AuNP solutions deposited on a clean silicon surface.

deformed by the interaction with the silicon substrate, the AuNP will be found at a larger depth in a G4/AuNP nanocomposite than in a G8/AuNP nanocomposite.

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’ CONCLUSIONS AFM and TEM results show that G4/AuNP tend to form agglomerates more when compared to G8/AuNP. NICISS results provide evidence that nanocomposites deposited on a surface undergo conformational changes due to the interaction between the dendrimer and the substrate. We have successfully used AFM phase imaging to gain insights into the structure of dendrimer AuNP nanocomposites. G4/ AuNP showed strong (negative) phase responses when imaged at 90% of free vibrational amplitude, which were attributed to gold surfaces exposed to the AFM tip. This strong phase response was absent in the case of G8/AuNP. Together with the TEM data and the AFM height images, our results suggest that G4 stabilizes the AuNP upon capping the AuNP exterior. In contrast, the surface of AuNPs was completely covered by dendrimer when AuNPs were formed in the presence of G8. In that case, the G8 seems to effectively shield the AuNP. The NICISS results support results obtained through AFM and TEM. This information will be critical when pursuing applications of dendrimer/ metal nanoparticle systems, such as catalysis, drug delivery, and sensing applications where the gold surface exposure will be an important factor in the performance of the system. ’ AUTHOR INFORMATION Corresponding Author

’ DISCUSSION OF THE AFM, TEM, AND NICISS RESULTS The interpretation of the AFM, TEM, and NICISS results are schematically illustrated in Figure 7. Only one or two of the methods gives evidence for some of the conclusions while other conclusions are based on the results of all methods. The TEM and AFM results show that the G4 dendrimer/ AuNP nanocomposite have a strong tendency to agglomerate, while the agglomeration of the G8 dendrimer/AuNP nanocomposite is limited. The agglomeration can be attributed to the interaction between the NH2 groups. The agglomeration cannot be seen directly in the NICIS spectra, as the current NICISS setup does not provide lateral resolution. NICISS gives evidence as to the deformation of the dendrimer/AuNP nanocomposite when deposited on surfaces. The further the dendrimer/AuNP nanocomposite moves away from the surface, i.e., by forming stacks of dendrimer/AuNP nanocomposites, the thicker the coverage of the AuNP with dendrimer material. G4/AuNP nanocomposites are formed by dendrimers surrounding the AuNPs. In G8/AuNP nanocomposites, however, the AuNP is located inside the dendrimer. When G4 are surrounding AuNPs, in some cases, structures are formed leaving the AuNP partially uncovered. This is not possible when AuNPs are formed inside the dendrimers, as in the case of the G8. AFM phase imaging gives evidence for this as only when mapping the G4 dendrimer/AuNP nanocomposites with the AFM tip interaction between the AFM tip and the AuNPs can be detected. It must be emphasized that the interaction between the AFM tip and the AuNP is possible not only when the top of the AuNP is not covered but also when the AuNP is laterally not completely surrounded by dendrimers. The evidence from NICISS for this conclusion comes from the observation that G4 on average forms a thicker layer around the AuNPs than the G8.

*E-mail: gunther.andersson@flinders.edu.au (G.G.A.), nico. voelcker@flinders.edu.au (N.H.V.).

’ ACKNOWLEDGMENT This work was made possible through an Australian EIPRS scholarship. The authors would also like to thank Kerry Gascoigne for expertise provided with the TEM analysis at Flinders Microscopy and Dr. Chris Gibson for his assistance in interpreting some of the AFM data. ’ REFERENCES (1) Zhou, L.; Russell, D. H.; Zhao, M.; Crooks, R. M. Macromolecules 2001, 34, 3567. (2) Zhang, Z.; Yu, X.; Fong, L. K.; Margerum, L. D. Inorg. Chim. Acta 2001, 317, 72. (3) Xu, Y.; Zhao, D. Environ. Sci. Technol. 2005, 39, 2369. (4) Wang, D.; Tran, H.; Margerum, L. D. In 219th ACS National Meeting; American Chemical Society: Washington, D. C: San Francisco, CA, 2000. (5) Venditto, V. J.; Regino, C. A. S.; Brechbiel, M. W. Mol. Pharm. 2005, 2, 302. (6) Tran, M. L.; Gahan, L. R.; Gentle, I. R. J. Phys. Chem. B 2004, 108, 20130. (7) Shi, X.; Ganser, T. R.; Sun, K.; Balogh, L. P.; B., J. R., Jr. Nanotechnology 2006, 17, 1072. (8) Perignon, N.; Marty, J.-D.; Mingotaud, A.-F.; Dumont, M.; RicoLattes, I.; Mingotaud, C. Macromolecules 2007, 40, 3034. (9) Mark, S. S.; Bergkvist, M.; Yang, X.; Angert, E. R.; Batt, C. A. Biomacromolecules 2006, 7, 1884. (10) Li, Y.; El-Sayed, M. A. J. Phys. Chem. B 2001, 105, 8938. (11) Li, G.; Luo, Y. Inorg. Chem. 2008, 47, 360. (12) Krot, K. A.; de Namor, A. F. D.; Aguilar-Cornejo, A.; Nolan, K. B. Inorg. Chim. Acta 2005, 358, 3497. (13) Hendricks, T. R.; Dams, E. E.; Wensing, S. T.; Lee, I. Langmuir 2007, 23, 7404. (14) Goodson, T.; Varnavski, O.; Wang, Y. Int. Rev. Phys. Chem. 2004, 23, 109. 6766

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