Studies on the Formation of Organosilica Nanoparticles and Their

J. Phys. Chem. C 2008, 112, 17063–17070

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Studies on the Formation of Organosilica Nanoparticles and Their Ability To Host Hydrophobic Substances Maria Stjerndahl,† Patrik Jarvoll,‡ Martin Andersson,†,‡ Ryan Kohout,† and Randolph S. Duran*,† Department of Chemistry, UniVersity of Florida, GainesVille, Florida 32601, and Department of Chemical and Biological Engineering, Chalmers UniVersity of Technology, SE-412 96, Go¨teborg, Sweden ReceiVed: March 2, 2008; ReVised Manuscript ReceiVed: August 14, 2008

Water-dispersible organosilica nanoparticles were synthesized using microemulsions and micellar solutions. Octadecyltrimethoxysilane (OTMS) was used as the silica source resulting in particles having a hydrophobic interior with the ability to host oil. The diameters of the formed particles could be varied between 60 and 200 nm, depending on the amount of added oil and OTMS. The size of the particles was determined using dynamic light scattering and transmission electron microscopy. Exchange-coupled diffusion nuclear magnetic resonance experiments were performed to study the exchange rates between the particles and the surrounding media. Triethylamine and tributylamine were used as probe molecules, and it was shown that they had longer mean residence times in the particles compared with in the bulk. Moreover, it was found that the mean residence time of the probe molecules increased significantly when the particles contained oil. The results also showed that the mean residence time of tributylamine was longer than that of triethylamine. Furthermore, by the use of UV-vis spectrophotometry, it was shown that the particles were able to take up benzophenone from water solutions. Introduction Biocompatible nanosized particles have been shown to have great potential in medical applications such as biodetoxification and drug delivery.1-6 The small size of the particles opens up the possibility of new routes for delivery and uptake because they are able to penetrate small biological passages such as the mucosal membrane7 and the blood-brain barrier.8 Moreover, the nanosized particles can easily be presented to the body through direct injection into the blood stream followed by transportation as part of the systemic circulation system.9 Several kinds of nanoparticles with the ability to host specific molecules have been presented in the literature. As examples, particles having core-shell architectures such as hollow spheres,10 bioresorbable particles,11 and particles with continuous porous structures12 can be mentioned. Within the context of being used for medical applications, organic particles,13,14 silica particles,6,10,15 solid lipid particles,4 and metallic particles16,17 have been suggested. One often desired property of the nanoparticles is that they should be dispersible in aqueous media but that their interior should constitute a hydrophobic, that is, oil-like, environment. This would give the particles the ability to host hydrophobic compounds, which would be of interest due to the lack of approved oils for direct injection purposes available for human applications.18 Silica, SiO2, has proven to be a promising material for in ViVo applications owing to its high stability, good biocompatibility and excellent possibilities for anchoring derivatizing molecules.19,20 Methods for the preparation of nanosized particles of SiO2 are numerous and include reactions performed under Sto¨ber conditions21 and template-assisted syntheses using * To whom correspondence should be addressed. E-mail: duran@ chem.ufl.edu. † University of Florida. ‡ Chalmers University of Technology.

surfactants and polymers.22,23 One of the most promising formation techniques is the use of oil-in-water microemulsions, which are thermodynamically stable systems consisting of nanosized oil droplets stabilized by a monolayer of surfactants in a continuous aqueous phase.24,25 The Duran group has earlier reported on the use of microemulsions for the formation of core-shell particles, containing a hydrophobic liquid interior and a hydrophilic pure silica shell.6,26 These core-shell particles have proven to have the ability to sequester hydrophobic compounds in their liquid like interior. Upon further investigation of these particles using transmission electron microscopy and atomic force microscopy, it has been suggested that the core of these core-shell particles has a more robust character than the earlier presented liquid-like core.27 One of the reasons for drawing this conclusion was that the core part of the particles retained its structure after being subject to high vacuum. Hence, an outer silica shell to increase the robustness of the core portion of the particle might not be needed. Herein, we report on the use of a micellar and an oil-in-water (O/W) microemulsion system for the preparation of waterdispersible organosilica nanoparticles having a hydrophobic interior. Syntheses were performed according to procedures presented before, both with and without pharmaceutically acceptable oil (ethylbutyrate) present.28 However, in contrast to the earlier works, no additional pure silica shell was grown onto the particles. Octadecyltrimethoxysilane (OTMS) was used a silica source resulting in a hydrophobically modified silica matrix with the ability to host oil. Various sized particles were formed and characterized using dynamic light scattering (DLS) and transmission electron microscopy (TEM). To show that the particles can sequester hydrophobic substances, diffusion nuclear magnetic resonance (NMR) was used, and the dynamic exchange rates of triethylamine and tributylamine as probe molecules were monitored.29-31 The diffusion was followed using stimulated echo pulse field gradient experiments (STE-

10.1021/jp803867s CCC: $40.75  2008 American Chemical Society Published on Web 10/09/2008

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PFG) with varied diffusion times.32-34 Furthermore, UV-vis spectrophotometry was used to study the uptake of benzophenone from water solutions. Materials and Methods Materials. All chemicals were obtained from commercial sources and were used without further purification. Tween 80 (poly(oxyethylene)20-sorbitan monooleate), sodium octanoate, ethylbutyrate (EB), octadecanol, triethylamine, tributylamine, and sodium chloride were obtained from Aldrich. Benzophenone came from Fisher Scientific. Octadecyltrimethoxysilane (OTMS) was purchased from Gelest Inc. All water used was obtained from a milli-Q water purification system. Synthesis of the Nanoparticles. A surfactant solution was prepared by dissolving Tween 80 (8 g) and sodium octanoate (8 g) in 220 mL of saline (9 g/L NaCl). To 5 g of the surfactant solution, the appropriate amount of EB was added, and the mixture was vortexed and placed in an orbital shaker at 75 °C. After 2 h, OTMS was added, and the sample was vortexed before being put back in the 75 °C shaker overnight. After reaching room temperature, the sample was sonicated for 10 min, and the pH was then adjusted to 10 using sodium hydroxide. After 30 min of shaking at room temperature, the pH was adjusted to neutral using hydrochloric acid, and the sample was sonicated for 10 min before being filtered through 450 nm pore size filters (Fisherbrand). The formed nanoparticles were purified by dialysis (Spectra/Por regenerated cellulose MWCO 6-8 kDa) against water containing Bio-beads, normally used to sequester surfactants used during protein purification, for four days changing the water and the beads at least twice daily. All syntheses were repeated at least three times. Before NMR analysis, additional purification was performed. Most of the solvent was removed from the suspension, concentrating the nanoparticles, and the solvent was changed from H2O to D2O, by forcing the solvent through a 25 nm pore size filter (Millipore). Samples were sonicated for at least 15 min before NMR analysis. Before performing the uptake studies using UV-vis spectrophotometry, the particle suspensions (4 mL) were dialyzed in the presence of Bio-beads for at least 1 month, changing the water and the beads every third day. Determination of the Phase Boundaries of the Microemulsions. To 5 g of the surfactant solution, different amounts of EB were added, and the mixtures were vortexed and shaken at 75 °C for 2 h. The samples were then inspected by the naked eye to determine the existence of a microemulsion. Clear, transparent solutions were judged as being microemulsions. Similar analyses were performed to establish the phase boundary between the microemulsion phase and the emulsion phase in the presence of OTMS. However, since OTMS would hydrolyze under these conditions, forming particles, octadecanol was used as a stable analogue. Trials both with and without EB were made. Transmission Electron Microscopy (TEM). TEM studies were performed using a Hitachi H-7000 microscope operated at 75 kV. Specimens were prepared by placing a drop of the purified particles suspended in water onto a Formvar carboncoated copper grid followed by drying at room temperature. Size determinations were performed manually, by measuring the diameter of at least 100 particles. Dynamic Light Scattering (DLS). DLS measurements were performed using a Precision Detectors PDDLS/CoolBatch+90T instrument. The Precision Deconvolve32 program was used to analyze the data. The measurements were performed at 20 °C at a 90° scattering angle using a 683 nm laser source. Final

Figure 1. An illustration describing the pulse sequence used in the STE-PFG experiments.

sizes were obtained from the average of at least three reproducible results. Generally 10 µL of the particle dispersion was diluted into approximately 10 mL. Additional dilutions were performed until the count rate was between the recommended 200 000 and 400 000 counts per second. Furthermore, dilution series were performed to ensure that no multiple scattering was effecting the measurement. NMR Diffusion. The exchange-coupled diffusion stimulated echo pulse field gradient (STE-PFG) experiments were performed on a Bruker 750 MHz equipped with a diff60 gradient capable of providing 30 T/m in field strength along the z-axis. For the experiments, a maximum field strength of 22 T/m was used since the gradient profile was not linear above 24 T/m. A Bruker 10 mm diffusion probe was used but with 5 mm NMR tubes. The pulse sequence used is described in Figure 1. The particles used were prepared from 40 mg of OTMS containing no oil and 40 mg of EB, respectively. All NMR experiments were performed at 25 °C. As probe molecules, triethylamine and tributylamine were chosen at a concentration of 0.5 µM. The intensity of the peaks at approximately 2.45 ppm for triethylamine and 2.29 ppm for tributylamine was determined as a function of the gradient strength (g), which was varied linearly in 32 steps. The duration time of the gradient (δ) was kept constant at 1.1 ms, and the diffusion time (∆) was constant within each experiment. In a set of six experiments, ∆ was varied: 10, 25, 50, 100, 250, and 500 ms. Each experiment had 64 scans and a delay time of 6 s between the scans. Sixteen dummy scans were performed before starting the sampling of the signal. The intensity signal of the echo decay, I/I0, for free diffusion of a STE-PFG experiment for one component can be described as

(

)

ta 2tb I 1 (k) ) exp - exp(-kD) I0 2 T1 T2

(1)

where k ) γ2δ2g2(∆ - δ/3), γ is the gyromagnetic ratio, ta is the time between the gradient pulses when T1 relaxation occurs, and tb is the time when T2 relaxation occurs, that is, between the first and second 90° pulse and after the third 90° pulse until sampling of the signal. The data from the exchange-coupled diffusion using STEPFG experiments were fitted to a model presented by Scho¨nhoff and described by Ka¨rger.35,36 The model describes the diffusion for a two site exchange situation (A and B), and the mean residence times (τA and τB) can be extracted from it. The echo decays, φ, from all ∆ were fitted simultaneously to the expression

φ(k) ) pA exp(-RA) + pB exp(-RB) where R is

(2)

Organosilica Nanoparticles

RA(+),B(-) )

)

{

J. Phys. Chem. C, Vol. 112, No. 44, 2008 17065

[(

)(

ta/∆ 2tb/∆ ta/∆ 1 k(DA + DB) + ∆ + + + 2 T1A T2A T1B

] [(

[( ])

) ]}

ta/∆ 2tb/∆ 2tb/∆ 1 1 + + ( k(DA - DB) + ∆ + T2B τA τB T1A T2A

and p is

pB )

(

[

)

ta/∆ 2tb/∆ 1 1 + + T1B T2B τA τB

(

)

2

+

(

4∆2 τAτB

1⁄2

(3)

)

τB τA 1 ∆ ∆ kDA + + kDB + RB - RA τA + τB TA τA + τB TB

]

RA (4) pA ) 1 - pB

(5)

In this system, the two sites are the bulk phase, A, and the organosilica nanoparticles, B. The fitting procedure was performed in several steps, where DA, T1A, and T2A were fixed in the final fit. Fixed starting values for the parameters in the bulk phase were taken from experiments performed separately when no particles were present. The starting values for the initial fit of the diffusion of probe molecules within the organosilica particles were taken from the lower part of the echo decay for the shortest ∆ (10 ms). Initial starting values were T1B ) T1A and T2B ) T2A, for the probe molecules in the particles. The mean residence times were set to 100 ms (τA ) τB) and the weighting parameters were allowed to match in each echo decay. The average values from 500 minimizations, using Monte Carlo simulated noise, were obtained to validate the precision of the initial parameters.37 This first fit was performed in order to ensure that all parameters were within reasonable ranges. These parameters were then used as starting values and DA, T1A and T2A were fixed to the values obtained from the measurements in bulk solution in the absence of particles. The procedure was iterated until all fitted parameters were within 5% of their starting values. These optimized starting values were then used in a final fit using 1000 Monte Carlo simulations. For the diffusion experiments performed to determine the possible presence of micelles in the particle dispersion, a 500 MHz Varian Inova equipped with a DOTY diffusion probe capable of gradients of maximum 4.8 T/m was utilized. For these experiments, sinus-shaped gradient pulses (δ ) 2 ms) were used, which slightly changed the diffusion expression to k ) γ2δ2g2(4∆ - δ/π2). Diffusion times of ∆ ) 25, 100, and 500 ms were employed. NMR Relaxation. Relaxation experiments were performed with an inversion recovery sequence for T1 and a Carr-PurcellMeiboom-Gill (CPMG) sequence for T2.38 In the CPMG sequence, the total delay between the 180° pulses was 2.5 ms, which is comparable to how T2 is affected in most of the diffusion experiments. Uptake Studies Using UV-Vis Spectrophotometry. Uptake studies were performed by keeping the nanoparticle solution trapped within a dialysis tube submerged in a water solution containing the UV-active hydrophobic molecule benzophenone. Before use, the dialysis tube (Fisherbrand regenerated cellulose, 12-14 kDa MWCO) was soaked in water overnight and rinsed. The uptake experiment was started by placing the dialysis tube in the benzophenone solution, 30 mL, for 45 min. A sample, which corresponded to t ) 0, was then withdrawn. The sonicated nanoparticle suspension, 1 or 2 mL, was placed inside the dialysis tube, which was then sealed. To compensate for the

Figure 2. Photograph of particle dispersions prepared from 40 mg of OTMS without (left) and with 80 mg of ethylbutyrate (right).

dilution of the benzophenone solution due to the addition of the nanoparticle solution, saturated benzophenone solution was added. The uptake system was placed on a shaker, and aliquots of 0.5 mL were withdrawn from the solution outside the dialysis tube at certain times. The absorption at 258 nm was monitored using a PharmaSpec UV-1700 (Shimadzu). Particles made from 40 mg of OTMS containing no oil and particles made from 40 mg of OTMS with 40 mg of EB were investigated. In order to ensure that the dialysis tube itself did not absorb benzophenone, the above-described experiment was performed in the absence of nanoparticles. Those experiments did not result in any change in the intensity of the absorbance with time. Results Synthesis of the Nanoparticles. The particle syntheses resulted in the formation of suspensions that ranged from being translucent bluish to opaque white, see Figure 2. Syntheses performed without or at low concentrations of EB, in combination with low to moderate amounts of OTMS, were generally bluish and totally homogeneous. At higher EB concentrations (40 mg and above), white suspensions were formed, which sometimes displayed white flakes at the surface. The same type of flakes could be seen in samples with high amounts of OTMS (80 mg and above). The relation between the initial composition of the reaction solution and the resulting particle size, as determined by DLS, can be seen in Figure 3. The diameters of the formed particles as a function of the ethylbutyrate (EB) concentration are shown in Figure 3a. The two series presented contain different amounts of OTMS (20 and 40 mg). From 0 to 50 mg of EB, the particle diameter increased linearly from about 60 to 110 nm. At EB concentrations above 50 mg, a deviation from linearity toward larger particles was observed. In Figure 3b, the particle diameter as a function of OTMS concentration is presented. Two series are shown, one with no EB present and one with the addition of 10 mg of EB. For both series, the diameter of the particles increased with increasing amounts of OTMS. However, the overall diameters were larger when EB was present. In both series, the diameters seemed to first increase, then level off, and then increase again. When Figure 3, panels a and b, are compared, it is obvious that the changes in the diameters were not as pronounced when OTMS was varied as compared with when EB was varied. All syntheses were repeated at least three times and the diameters presented in Figure 3 are the mean values. Also included in the figures are the standard deviations. When the OTMS concentrations were varied, the reproducibility was excellent. When EB concentration was varied, the reproducibility was very good for amounts lower than 60 mg. At higher EB concentrations, samples with a somewhat larger size variation were obtained.

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Figure 3. DLS results showing the diameters of the formed particles (a) as a function of the amount of ethylbutyrate (EB) and (b) as a function of the amount of octadecyltrimethoxysilane (OTMS). The presented diameters are the mean values from three separate syntheses, and the error bars represent the standard deviation. When no error bars are visible, the errors are so minor that they are covered by the symbols.

The particles were analyzed using TEM, which gave information not only about the size but also about the morphology. In Figure 4, TEM micrographs of particles synthesized with different amounts of EB (0, 30, 60, and 80 mg) are presented. In the case of no EB, the particles are spherical in shape having a quite narrow size distribution and a diameter of 68 ( 6 nm (arithmetic mean value and standard deviation). With increasing oil content, the particles grew larger and the size distributions became wider. The micrographs b-d illustrate how the particles seem to become more and more deformable as the oil content increases. Especially in micrograph d, the particles look much squeezed. Moreover, for some of the oil-containing particles the center appears lighter than the rest of the particle, which makes them appear hollow. Determination of the Phase Boundaries of the Microemulsions. The determination of the phase boundaries of the microemulsion system at 75 °C showed that up to about 70 mg of EB can be added to a 5 g surfactant solution before the microemulsion breaks and an ordinary emulsion is formed. Since OTMS starts hydrolyzing as soon as it gets in contact with water, a stable analogue had to be used to explore its phase boundaries. Octadecanol was chosen since it has the same length of the hydrophobic tail and a polar but not ionic headgroup. It was found that to a microemulsion system consisting of 5 g of surfactant solution and 20 mg of octadecanol, about 60 mg of EB can be added before the microemulsion turns into an emulsion.

Stjerndahl et al. NMR. The organosilica nanoparticles themselves are not detectible by NMR, and the oil within them, ethylbutyrate, gave weak NMR signals due to fast relaxation.39,40 Consequently, probe molecules had to be used in order to monitor the diffusion of the particles. By choice of hydrophobic probes, the ability of the organosilica particles to host hydrophobic substances could be determined using relaxation and diffusion NMR. Because the sensitivity of NMR is relatively low, probe molecules with fairly high water solubility had to be chosen. Two amines, triethylamine and tributylamine, were suitable probe molecules since the presence of nitrogen atoms make the NMR signals easy to differentiate from hydrocarbon signals. Triethylamine has a moderately high water solubility, 17 g/100 mL, and an octanol/water partition coefficient of log Pow ) 1.45. The corresponding values for the more hydrophobic tributylamine are 0.52 g/100 mL water and log Pow ) 1.52. At neutral pH, both probe molecules are positively charged. It is reasonable to assume that the continuous aqueous phase of the particle suspensions contains some surfactants even after dialysis. As a consequence, initial experiments were performed to determine the diffusion rate and relaxation times of triethylamine in surfactant systems of different concentrations, see Table 1. The surfactant system consisted of a 1:1 mixture, by weight, of Tween 80 and sodium octanoate. The concentrations of Tween 80 are reported in Table 1. Both the T1 and the T2 relaxation obtained from the solutions at or above 0.2 mM Tween 80 solution corresponded well with the slow relaxation times obtained from measurements on the particle dispersions, which will be presented later. The critical micelle concentration (CMC) is 0.01 mM for Tween 80, and hence the so-called bulk solution was a micellar solution. From the exchange-coupled diffusion experiments performed on the nanoparticle suspensions, two sets of T1 and T2 were extracted for both the probes, triethylamine and tributylamine. The slower relaxation rates can be assumed to originate from the probes when present in the bulk, T1bulk and T2bulk, and the faster when present in the particles, T1SiP, T1SiP+oil and T2SiP, T2SiP+oil, where SiP and SiP+oil denotes organosilica particles without and with oil, respectively. The exchange coupled diffusion fits are shown in Figure 5 as intensity, I, vs k for both probes and for particles with and without oil. The fits correspond well with the data; however, higher noise levels were observed in the case of tributylamine. For further validation, Monte Carlo simulations (1000) were performed, and the results are shown in Figure 6 as histogram plots of the distributions. The diffusion coefficients of the probes within the particles DSiP and DSiP+oil are 5.22 × 10-12 and 3.36 × 10-12 m2 s-1, respectively, in the case of triethylamine and 5.24 × 10-12 and 3.29 × 10-12 m2 s-1, respectively, in the case of tributylamine. These values correspond to a particle size of approximately 100 nm for the organosilica particles and 150 nm for the EB-containing organosilica particles according to the Stokes-Einstein relation.41 Lower relaxation values were generally obtained for the probes present in the particles compared with those in the bulk, except in the case of T2SiP+oil using the tributylamine probe (Figure 6d). This discrepancy was probably a fitting error caused by a higher noise level. The shorter relaxation in the particles compared with bulk was an expected result since the probe dynamics were significantly more restricted in the particles. Furthermore, the relaxation times, particularly the T2 relaxation, became shorter for particles containing oil compared with the ones without oil. The mean residence times, τ, were generally larger in the particles, especially for those containing oil,

Organosilica Nanoparticles

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Figure 4. TEM micrographs showing organosilica particles prepared from 40 mg of OTMS without and with oil. Syntheses without EB gave particles with a mean diameter of 68 ( 6 nm (a), 30 mg of EB yielded diameters of 105 ( 16 nm (b), 60 mg of EB resulted in diameters of 160 ( 32 nm (c), and 80 mg of EB yielded diameters of 183 ( 30 nm (d). The values reported are the arithmetic mean value followed by the standard deviation (().

TABLE 1: The Diffusion Rates, D, and Relaxation Times, T1 and T2, of Triethylamine in Surfactant Solutions of Different Concentrations CTween80 (mM)

D (m2 s-1)

T1 (s)

T2 (s)

0.05 0.1 0.2 0.5 2.5

7.9 × 7.9 × 10-10 7.9 × 10-10 7.9 × 10-10 7.9 × 10-10

1.82 1.79 1.80 1.78 1.75

1.49 1.32 0.44 0.44 0.42

10-10

compared with those in the bulk. This relation was considerably more apparent in the case of tributylamine. To conclude, these results show that the probes have a higher probability of existing in the particles as compared with in the bulk. Uptake Studies Using UV-Vis Spectrophotometry. The ability of the nanoparticles to sequester hydrophobic substances was also investigated using UV-vis spectrophotometry. The highly UV-active substance benzophenone was used as hydrophobic probe molecule. In Figure 7, the intensity of the UV absorbance is presented as a function of the time that the particles have been in contact with an aqueous benzophenone solution. Particles with and without oil were used, and different volumes of the particle suspensions, 1 and 2 mL, were investigated. As can be seen, most of the benzophenone was sequestered during the first hour, after which the uptake slowed considerably. When the volume of the particle suspension was increased, that is when the number of particles able to host the probe was doubled, the uptake increased. Unfortunately, it is impossible to compare the particles with and without oil and draw any quantitative conclusions since the exact concentrations of the particles were unknown because they were altered during the excessive filtering and dialyzing steps. Discussion The synthesis procedure for preparation of the nanoparticles was based on the hydrolysis and subsequent condensation of the octadecyltrimethoxysilane, OTMS, in the environment of a micellar or microemulsion solution. The surfactant solution used to prepare the nanoparticles consisted of two surfactants, the

nonionic Tween 80 and the anionic sodium octanoate. The inclusion of a charged species makes the formed micelles and microemulsion droplets negatively charged and, hence, electrostatically stabilized. The nanoparticles could be formed both with and without the addition of oil and hence both in a micellar solution and in a microemulsion. By varying the concentration of OTMS and oil, the size of the obtained particles could be altered. In this study, the oil utilized was EB, although other oils, such as toluene and mineral oil, have been successfully employed. At room temperature, OTMS is close to its melting point. When added to the surfactant solution or EB-containing microemulsion, it will thus have a tendency to form a separate solid layer on top of the surfactant solution prohibiting an efficient inclusion into microemulsion droplets. At 75 °C, the OTMS is liquid, and it is easily dispersed as microemulsion droplets. As soon as the OTMS is added to the heated surfactant solution, the hydrolysis of the silane groups will start, resulting in the formation of reactive silanol groups. These silanol groups can react with other silanol groups forming siloxane linkages. Because each OTMS carries three silane groups, it can be linked covalently to up to three other hydrolyzed OTMS molecules. We believe that this results in the formation of a threedimensional hydrophobic matrix, eventually forming spherical nanoparticles. In the case of oil present during the reaction, it will be included as part of the particles and hence the particle size will increase upon its addition. The dispersion was heated overnight, and to ensure that all silane groups were hydrolyzed and condensed, the dispersion was then made alkaline for 30 min to further catalyze the reaction. It is known that when OTMS is hydrolyzed and condensated, the alkyl chains still remain covalently bounded to the Si, resulting in an organic/ inorganic organosilane.42 The alkyl chains can be removed, however, for example by a calcination step. When this extra heating step was performed on the particles (450 °C), with the aim of forming porous pure silica particles, the structure collapsed resulting in a powder of scattered silica, as seen by electron microscopy (results not shown here). Hence, the structure is not strong enough to remain as particles of pure silica alone. Considering the size of the organic part of the

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Figure 5. Exchange-coupled diffusion fits obtained for particles without oil (a, c) and with oil (b, d). Triethylamine was used as probe in the experiments of panels a and b, and tributylamine was used in the experiments of panels c and d. Each graph shows the echo decays obtained for a series of diffusion times: 10 (b), 25 (2), 50 (9), 100 (O), 250 (4), and 500 ms (0).

Figure 6. Evaluation distributions of Monte Carlo simulations obtained from the fits shown in Figure 5. The distributions, the mean values, and the standard deviations, in parentheses, are presented. The diffusion coefficients, D, are presented in m2 s-1; the relaxations times, T1 and T2, are presented in s; and the mean residence times, τ, are presented in s. Triethylamine was used as probe in the experiments of panels a and b. Tributylamine was used in the experiments of panels c and d. In the experiments of panels b and d, oil-containing particles were used and in those of panels a and c particles without oil.

OTMS compared with the silica network forming part, this is not surprising. The dominating fraction of the structure of the nanoparticle will in fact consist of the organic octadecyl chains. The size of the obtained particles was dependent upon the composition of the microemulsion. As seen from the phase studies, the microemulsion can only withstand oil contents up to 60 mg of EB. At oil concentrations below this limit, the particle size increases linearly with increasing amounts of oil. The reproducibility of the syntheses was also very good as seen from the error bars. At higher oil concentrations, the reaction

was no longer performed in a microemulsion but an emulsion system, which resulted in particles of larger size and also larger differences between repeated syntheses. The particle size seemed to be less sensitive to the amount of OTMS than to oil. Increased amount of OTMS also resulted in an increased particle size, but the increase was less pronounced. At low OTMS concentrations, the reproducibility of the syntheses was at a similar level as when EB was varied. At high OTMS concentrations, the reproducibility was still surprisingly good, even though the syntheses were not performed within a microemulsion. For

Organosilica Nanoparticles

Figure 7. The decrease in UV absorbance of aqueous benzophenone in the presence of nanoparticles with and without EB. The uptake for two different volumes of particle suspensions, 1 and 2 mL, was investigated. The absorbance was measured at 258 nm.

reactions performed at high concentrations of oil or OTMS, flakes or larger chunks of polymerized OTMS were formed along with the nanoparticles. These had to be removed by filtration before any analyses could be performed. In the present study, three independent methods were used to determine the size of the formed particles. The obtained diameters correlate well, especially between the DLS and TEM. When the results from the diffusion NMR were translated into diameters using Stokes-Einstein, the particle size generally became somewhat larger compared with DLS and TEM. However, the Stokes-Einstein relation assumes that each single particle executes movement that is independent of the movement of all other particles. In the DLS measurements, the samples were highly diluted to reduce the effects of multiple scattering, but in the case of the NMR experiments, the dispersion was much more concentrated. The high concentration of particles in the NMR experiment was necessary in order to ensure a reliable signal-to-noise ratio for the uptake of the probe molecules. As a consequence, particle-particle interaction will cause obstruction effects resulting in slower diffusion, and hence, overestimated particle sizes will be obtained. The particle suspensions were dialyzed and concentrated before the NMR analysis to remove excessive surfactants; however, as mentioned in the results for the NMR experiments, the bulk solution corresponded to a micellar solution. The micellar solutions could, however, still be considered as onephase systems since the exchange rates for the probes between micelles and pure water were significantly faster than the time scales used in the diffusion experiments. In the experiments performed on the surfactant solutions containing no particles, only one diffusion coefficient, one T1 and one T2 could be extracted with the experimental parameters used, see Table 1. Due to the fast exchange rate, the diffusion coefficients will not significantly change with increasing surfactant concentration. However, the relaxation rate will be affected by the presence of micelles. The NMR relaxation studies performed on the particle systems showed that the probes’ relaxation rates were lower in the particles that did not contain oil compared with those in the oil-containing particles. Hence, the probes experienced a higher restriction in the particles without oil, indicating that the hydrophobic domains within the particles were larger when oil was present. This result supports the observation that the particles synthesized in the presence of oil are larger due to the inclusion of oil in the organosilica matrix. The NMR diffusion

J. Phys. Chem. C, Vol. 112, No. 44, 2008 17069 studies showed that the mean residence times were longer for the oil-containing particles indicating that the oil increased the ability to sequester hydrophobic compounds. When the effect of the different probes was compared, it could be seen from the mean residence times that the tributylamine had a higher probability to be absorbed into the particles compared with the triethylamine. This was expected since tributylamine is less water-soluble than triethylamine. To investigate whether the oil would leak from the particles and be taken up by the micelles in the bulk, the relaxation and diffusion of the surfactants were monitored for three days. The obtained relaxation and diffusion values did not change with time, and hence, the composition of the micelles was not altered, that is, they were not swollen by the oil. As a consequence, it can be concluded that the EB remains inside the organosilica particles. To ensure that the uptake of the amine probes was indeed due to hydrophobic and not electrostatic interaction, control experiments were performed by measuring the diffusion of the amines in the presence of pure solid silica particles formed by TEOS hydrolysis in water under Sto¨ber conditions. The results obtained by NMR measurements showed that the positively charged amines did not adsorb onto the negatively charged pure silica particles, ruling out the possibility of electrostatic interaction. Further support for the uptake being governed by hydrophobic interaction came from the uptake studies performed using benzophenone. It was shown that both the oil-containing particles and the particles without oil have the ability to sequester uncharged hydrophobic substances. However, since the concentration of the particle suspensions was unknown it was difficult to perform quantitative analyses of the differences between the two types. Conclusion Organosilica nanoparticles were prepared using microemulsions and micellar solutions. Octadecyltrimethoxysilane (OTMS) was used as the silica precursor forming particles with a hydrophobic interior. The particle size could be varied between 60 and 200 nm in diameter dependent on the concentrations of oil and OTMS used in the synthesis. The particles were robust enough to withstand degassing under vacuum as observed by TEM. Both types of particles, with and without oil, had the ability to sequester benzophenone from water solutions. The pulse field gradient NMR technique was successfully used to monitor the exchange rates of triethylamine and tributylamine, which were used as probes, between the organosilica nanoparticles and the surrounding bulk solution. The probes had a longer mean residence time in the particles than in the bulk, that is, a higher probability to be found in the particles. The probability of finding tributylamine in the particles, when compared with the surrounding bulk solution, was higher than that of triethylamine. Furthermore, particles synthesized in the presence of oil increased the probability of finding the probes in the particles. Acknowledgment. S. Blackband at the Advanced Magnetic Resonance Imaging and Spectroscopy (AMRIS) facility at the University of Florida is acknowledged for providing NMR access and for general assistance regarding the NMR analysis. K. Kelley at the Electron Microscopy Core Laboratory, Biotechnology Program, at the University of Florida is a thanked for technical assistance. The Swedish NMR-Centrum is acknowledged for granting NMR spectrometer time. Financial support was from the Knut and Alice Wallenberg Foundation (M.S. and M.A.), and the NHMFL internal research program (R.D.).

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