In-Situ Encapsulation of Quantum Dots into Polymer Microspheres

Huan Meng , Helmuth Möhwald , Paul Mulvaney , Andre E. Nel , Shuming Nie , Peter Nordlander , Teruo Okano , Jose Oliveira ..... Brent Fisher , Jo...
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Langmuir 2006, 22, 3782-3790

In-Situ Encapsulation of Quantum Dots into Polymer Microspheres Wenchao Sheng, Sungjee Kim, Jinwook Lee, Sang-Wook Kim, Klavs Jensen, and Moungi G. Bawendi* Department of Chemistry, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139-4307 ReceiVed July 21, 2005. In Final Form: December 22, 2005 We have incorporated fluorescent quantum dots (QDs) into polystyrene microspheres using functionalized oligomeric phosphine (OP) ligands. We find that a uniform distribution of quantum dots is loaded inside each polymer bead. Some local close-packing of quantum dots in the beads is attributed to the self-polymerization of the functionalized ligands. The presence of quantum dots disturbs the nucleation and growth processes during the formation of polymer microspheres and results in a wider size distribution of the quantum dot-embedded polystyrene beads than for the control without dots. The change in quantum efficiency of the quantum dots before (∼20%) and after (12%) loading into the beads substantiates the protection of oligomeric phosphine ligands yet indicates that the properties of these quantum dots are still affected during processing.

1. Introduction Semiconductor nanocrystals, also known as quantum dots, have drawn intense research interest in the past decade because of their unique electronic and optical properties, strong stability against photobleaching, high quantum efficiency, and narrow excitonic emission.1-3 The most intriguing feature about quantum dots is that their absorption and emission are strongly sizedependent. Quantum dots, despite being different sizes, are passivated by the same ligands, which determines that their surface chemistry is the same. Therefore, we can incorporate differentsized QDs into a matrix without changing the processing route. In addition, because of continuous absorption at wavelengths shorter than the band edge absorption peak, a single wavelength can be used to excite different-sized dots. This makes multiplexing possible.4 All of these features give quantum dots advantages over organic dyes and make quantum dots a promising chromophore for many applications. Incorporating quantum dots in microspheres is of interest for both fundamental studies on light-matter interaction5 and practical applications, such as semiconductor microlasing6 and biological tags.7 This dot-in-a-dot structure confines electrons and photons in all three dimensions. Radiative dynamics of QDs embedded in microcavities was studied in ref 8, where enhanced spontaneous emission from the QDs was demonstrated. In practical applications, quantum dots can be encapsulated into polystyrene or bioconjugated microspheres and used for genotyping of single nucleotide polymorphisms.9 This approach is not practical using traditional organic dyes because of the difficulty of exciting multiple dyes with a single wavelength. (1) Alivisatos, A. P. Science 1996, 271, 933. (2) Li, J. J.; Wang, A.; Guo, W.; Keay, J. C.; Mishima, T. D.; Johnson, M. B.; Peng, X. J. Am Chem. Soc. 2003, 125, 12567. (3) Murry, C. B.; Norris, D. J.; Bawendi, M. G. J. Am. Chem. Soc. 1993, 115, 8706. (4) Rosenthal, S. J. Nat. Biotech. 2001, 19, 621. (5) Klimov, V. V.; Ducloy, M.; Letokhov, V. S. Phys. ReV. A 1999, 59, 2996. (6) Cha, J. N.; Bartl, M. H.; Wong, M. S.; Popitsch, A.; Deming, T. J.; Stucky, G. D. Nano Lett. 2003, 3, 907. (7) Bruchez, M.; Moronne, M.; Gin, P.; Weiss, S.; Alivisatos, A. P. Science 1998, 281, 2013. (8) Fan, X.; Lonergan, M. C.; Zhang, Y.; Wang, H. Phys. ReV. B 2001, 64, 115310. (9) Xu, H.; Sha, M. Y.; Wong, E. Y.; Uphoff, J.; Xu, Y.; Treadway, J. A.; Truong, A.; O’Brien, E.; Asquith, S.; Stubbins, M.; Spurr, N. K.; Lai, E. H.; Mahoney, W. Nucleic Acids Res. 2003, 31, 43.

Methods to incorporate quantum dots into microbeads include the following: (1) The dispersion of commercially available or synthesized microbeads and quantum dots in a solvent/nonsolvent mixture that is chosen to result in the partial swelling of microbeads. Quantum dots from the solution penetrate into the microbeads and, as the solvent is removed, are captured in the subsurface region of the microbeads.10 (2) The use of a layerby-layer strategy to adsorb quantum dots consecutively onto oppositely charged microbeads. In this scheme, quantum dots are electrostatically bound to the surface of the microspheres.11 (3) The growth of a silica/QD shell around preformed silica spheres.12 (4) Loading QDs into polymer microbeads through emulsion13 and suspension polymerization.14,15 The first three methods are reproducible, and the synthesized quantum dotembedded beads are uniform. Although such beads have been employed in biological applications, for cases 1 and 2 the incorporated dots may leave the beads when the chemical environment is changed because the dots are physically captured inside or on the beads and are not chemically bound to the polymer matrix. In addition, because QD/microsphere constructs made using all three methods have dots only in the surface region or in a shell, the volume fraction of embedded QDs can be limited. There is therefore a desire to demonstrate the encapsulation of dots throughout the volume of a microsphere. For case 4, recent reports have shown the encapsulation of quantum dots into polystyrene microspheres by emulsion polymerization13 and suspension polymerization,14,15 but important characterization of the dot/microsphere construct after encapsulation is generally lacking. Although it has been shown that the spatial distribution of quantum dots within polymer spheres is uniform,14 information such as the loading fraction and the quantum efficiency of the dot/sphere construct has not been established. In this work, we develop a strategy to incorporate quantum dots chemically into polystyrene microspheres by using functionalized oligomeric phosphine (OP) ligands. We characterize (10) Han, M.; Gao, X.; Su, J.; Nie, S. Nat. Biotechnol. 2001, 19, 631. (11) Wang, D.; Rogach, A. L.; Caruso, F. Nano Lett. 2002, 2, 857. (12) Chan, Y.; Zimmer, J. P.; Stroh, M.; Steckel, J. S.; Jain, R. K.; Bawendi, M. G. AdV. Mater. 2004, 16, 2092. (13) Yang, X.; Zhang, Y. Langmuir 2004, 20, 6071. (14) Li, Y.; Liu, E. C. Y.; Pickett, N.; Skabara, P. J.; Cummins, S. S.; Ryley, S.; Sutherland, A. J.; O’Brien, P. J. Mater. Chem. 2005, 15, 1238. (15) O’Brien, P.; Cummins, S. S.; Darcy, D.; Dearden, A.; Masala, O.; Pickett, N. L.; Ryley, S.; Sutherland, A. J. Chem. Commun. 2003, 20, 2532.

10.1021/la051973l CCC: $33.50 © 2006 American Chemical Society Published on Web 03/17/2006

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Scheme 1. Schematic Illustration of the Formation of Pure Polystyrene Microspheres (PSMS)

the resulting dot/microsphere constructs and analyze the pros and cons of such an approach. We find that the quantum efficiency is reasonably maintained during processing. We also find that the quantum dots are uniformly distributed in the polymer matrix of the spheres, with some localized areas of higher concentrations due to self-polymerization of the quantum dots before incorporation into the growing microspheres. We find that at higher quantum dot concentrations the formation of self-polymerized QD aggregates limits the loading fractions possible. Moreover, and more importantly, the addition of QDs and OP ligands perturbs the nucleation and growth processes of polymer beads and results in a widening of the size distributions. The strategy of chemically incorporating quantum dots into a polymer matrix by using polymerizable ligands hinders the formation of uniform microspheres. We suggest that this finding is not restricted to our specific example and generally limits the incorporation of quantum dots into growing polymer spheres. This suggests a need for new strategies for the uniform loading of quantum dots into microspheres at high loading fractions. 2. Experimental Section 2.1. Synthesis of CdSe/ZnCdS/ZnS Core/Shell Structured Quantum Dots. Dimethylcadium (Cd(Me)2) and diethylzinc (Zn(Et)2) were obtained from both Strem and Organometallics and were purified by filtration (0.2 µm) before use. Hexamethyldisilathiane ((TMS)2S), trioctylphosphine (TOP 97%), and trioctylphosphine oxide (TOPO 90%) were purchased from Fluka, Strem, and Alfa Aesar, respectively, and used without further purification. Cadmium selenide (CdSe) quantum dots were synthesized through the pyrolysis of organometallic precursors of dimethylcadium and trioctylphosphine selenide (1.0 M selenium shots dissolved in TOP). TOPO (30 g) was degassed at 160 °C for at least 2 h. A well-mixed solution consisting of 200 µL of Cd(Me)2, 4 mL of 1.0 M TOPSe, and 15 mL of TOP was injected at 350 °C. The CdSe QDs formed were allowed to grow at 160 °C until the desired size was obtained. The QDs were then annealed at 80 °C overnight. Overcoating CdSe dots was performed via the following two-step procedure: (i) 30 g of TOPO was degassed at 160 °C for at least

2 h. A predetermined number of CdSe dots were twice precipitated from growth solution using methanol and redispersed in hexane and then transferred into the degassed TOPO at 70 °C. After the complete removal of hexane under reduced pressure, a solution of Zn(Et)2, Cd(Me)2, (TMS)2S, and TOP was slowly dripped at 150 °C into the reaction flask at a rate of about 1 drop/s. The quantities of these materials were determined to obtain a 12.5 nm deposition of Zn80Cd20S around CdSe dots. The CdSe/ZnCdS core/shell dots then were stirred at 150 °C for half an hour. (ii) After the deposition of the alloyed shell, a second overcoating solution containing Zn(Et)2, (TMS)2S, and TOP was slowly dripped at 170 °C into the flask for another 6.2 nm shell of ZnS. After addition, the dots were stirred for another half an hour and annealed at 80 °C overnight. The final CdSe/ZnCdS/ZnS quantum dots were kept in the growth solution. 2.2. Encapsulation of Quantum Dots into Polystyrene Microspheres (QD/PSMS). 2,2′-Azobisisobutyronitrile (AIBN, 98%), styrene (99%), poly(vinylpyrrolidone) (PVP, MW ) 10 000), and ethyl alcohol (99.5%) were obtained from Aldrich. Styrene was purified using an inhibitor-removing column (Aldrich), but all other materials were used as purchased without further purification. A typical procedure for preparing QD/PSMS is described as follows:16 a predetermined number of CdSe/ZnCdS/ZnS QDs were precipitated and redispersed twice using methanol and hexane and finally redispersed in an oligomeric phosphine (OP) ligand/DMF solution (20 wt %) developed by Kim et al.17 Excess OP ligands were used to guarantee that QDs remain fully passivated during the cap-exchange process. After vigorous stirring, the solution became clear, indicating that the QD surfaces were successfully passivated by OP ligands. After removing DMF under reduced pressure, QD/ OP ligands were dissolved in 0.5 mL of ethanol and held at 60 °C for 3 h for the purpose of prepolymerization where the majority of MMA groups are cross linked, forming a polymer shell that protects the QDs from the environment. The time for this process can be adjusted by monitoring unreacted MMA using NMR until it is barely detectible. These remaining unreacted MMA groups then participate in the following polymerization reaction with styrene. Styrene (0.1 (16) Lim, J. W. Ph.D. Thesis, Massachusetts Institute of Technology, Cambridge, MA, 2002. (17) Kim, S.; Bawendi, M. G. J. Am. Chem. Soc. 2003, 125, 14652.

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Figure 1. (a) SEM image of QD/PSMS at an acceleration voltage of 7 kV. The inset is the pure PSMS at 20 kV. (b) Histograms of the size distribution of QD/PSMS and pure PSMS (inset). mL), AIBN (1 mg), PVP (20 mg), and ethanol (0.5 mL) were then added to a vial containing pretreated QDs. Slow N2 flow was introduced into the mixture through a needle for half an hour. The solution was then placed in an oil bath at 60 °C with a slow stirring rate of about 60 rpm for 24 h. The quantum dot-embedded polystyrene microspheres (QD/PSMS) formed were collected by centrifugation and then washed with ethanol. Centrifugation and washing were repeated twice. QD/PSMS was kept in ethanol. 2.3. Characterization. 2.3.1. Fluorescence Microscopy. The samples were prepared by drop casting the QD/PSMS ethanol solution onto a glass slide. Optical and fluorescent images of QD/PSMS were obtained from a Nikon eclipse ME600 microscope using a 100× objective with a working distance of 0.3 mm. 2.3.2. Scanning Electron Microscopy. Scanning electron microscopy (SEM) was used to illustrate the surface morphology and size distribution of QD/PSMS. Samples were prepared by drop casting an ethanol dispersion solution of QD/PSMS onto an ∼1 cm2 silicon wafer. SEM measurements were performed on a JEOL JSM 6060 SEM at an acceleration voltage of 7 kV. 2.3.3. Microtomy and Scanning Transmission Electron Microscopy. Spatial and elemental information on QDs inside polymer beads can be obtained by using scanning transmission electron microscopy (STEM). The samples were prepared via the following procedure: dried QD/PSMS samples were fully mixed with a silicone elastomer base and a curing agent (Wbase/Wcuring agent ) 10:1) and cured overnight under house vacuum. The formed microsphere/ PDMS block was trimmed and microtomed into ∼2 mm × 2 mm × 40 nm slices at -170 °C. The slices were placed on a TEM grid

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Figure 2. SEM images of OP ligand/styrene copolymer microspheres. Samples a to f were prepared from 90 mg of styrene, 1 mg of AIBN, 20 mg of PVP, and 1.0 mL of ethanol with (a) 0, (b) 4, (c) 8, (d) 15, (e) 30, and (f) 60 mg of OP ligands, respectively. Sample g contains 90 mg of styrene, 1 mg of AIBN, 1.0 mL of ethanol, and 60 mg of OP ligands. Table 1. Reaction Conditions for OP Ligands/Polystyrene Microspheres a-g

a b c d e f g

styrene (mg)

AIBN (mg)

PVP (mg)

ethanol (mL)

OP ligands (mg)

90 90 90 90 90 90 90

1 1 1 1 1 1 1

20 20 20 20 20 20 0

1.0 1.0 1.0 1.0 1.0 1.0 1.0

0 4 8 15 30 60 60

for STEM analysis. STEM measurements of QD/PSMS slices were carried out on an HB603STEM. 2.3.4. Quantum Efficiency Measurements. Quantum yield measurements of dots in microspheres were carried out using an integrating sphere in order to eliminate the problem of light scattering. Relative quantum yields of QD/PSMS were obtained relative to rhodamine 590. The absorbance at the excitation wavelength (485 nm) was determined using the dissolution of QD/PSMS in toluene. The dissolution of the sphere eliminated scattering issues. The absorbances of rhodamine 590 and QDs were matched accordingly. The samples of QD/PSMS for QY measurements were prepared by dispersing the same amount of QD/PSMS in ethanol. Some pure polystyrene beads were intentionally added to another solution of rhodamine 590 with the same absorbance at 485 nm as a control on the effect of light scattering on the QY measurement. All of the

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Scheme 2. Schematic Illustration of the Formation of OP Ligand/Styrene Copolymer Microspheresa

a

The concentration of OP ligands is low.

emission intensities of the four samples were measured using the integrating sphere. The basic setup can be found in ref 18.

3. Results and Discussion The challenge of in-situ encapsulation of QDs into a polymer matrix lies in the method of providing chemical compatibility between the QDs and the polymer matrix while preserving the optical properties of the QDs. The surfaces of QDs are usually passivated by hydrophobic ligands such as TOP/TOPO. However, QDs with such ligands have limited solubility in a variety of solvents and have low sustainability in harsh environments. For example, the fluorescence of TOP/TOPO/QDs is quickly quenched by free radicals in radical-initiated polymerizations.16 The oligomeric phosphine (OP) ligands of ref 17 represent a useful ligand platform for further chemistry. OP ligands functionalized with methyl methacrylate (MMA) groups can serve the following purposes: (1) They bind more strongly to quantum dot surfaces than monodentate ligands and can more efficiently prevent ligand loss during processing. (2) The MMA functional groups render the QDs soluble in ethanol, which is the dispersive solvent in our microsphere system. (3) After prepolymerization for several hours, the majority of the MMA groups are cross linked, forming a polymer shell that protects the QDs from quenching by AIBN. (4) The remaining MMA groups react with vinyl groups on styrene and chemically incorporate the dots into the interior of the polystyrene beads. Lack of MMA groups results in the phase separation of QDs and polystyrene due to their different solubilities. Pure polystyrene beads can be synthesized by dispersion polymerization, which is driven by the solubility difference of the monomer and the polymer.19 The styrene monomer is soluble in ethanol whereas polystyrene microspheres (PSMS) are not. Scheme 1 sketches the process of dispersion polymerization. (18) Ware, W. R.; Rothman, W. Chem. Phys. Lett. 1976, 39, 449. (19) Tseng, C. M.; Lu, Y. Y.; El-aasser, M. S.; Vanderhoff, J. W. J. Polymer Sci., Part A 1986, 24, 2995.

The starting mixture is a homogeneous ethanol solution containing a monomer (styrene), an initiator (AIBN), and a stabilizer (PVP) (I). The polymerization reaction begins as AIBN decomposes to the free radical [NC-C(CH3)2•], which reacts with the styrene monomer, initiating the growth of the polymer chains (II). When the chains reach a critical length, their decreased solubility in ethanol makes them unstable in solution, and they begin to aggregate into small particles. These small particles become coated with the PVP stabilizer, which keeps them suspended in solution (III). These stabilized particles serve as nucleation sites. Styrene monomers and short polystyrene oligomers diffuse into these particles, driven by the higher solubility of styrene inside the particle than in ethanol. The polymerization reaction then takes place inside the particles and proceeds until all of the polymerizable units are used up (IV). Typical polystyrene beads prepared through this polymerization route are about 1.2 µm in diameter with a polydispersity of 2% (defined as the ratio between the standard deviation and the mean size) (insets of Figure 1a and b, respectively). When modified QDs are added, however, QD/PSMS lose the monodispersity of the pure polystyrene beads. The SEM image of Figure 1a shows that as-synthesized QD/ PSMS can have a very smooth surface morphology similar to that of pure polystyrene microspheres using the same dispersion polymerization route, but these QD/PSMS have a smaller mean diameter of about 813 nm and a significantly broader size distribution (Figure 1 b). These results imply that the polymerization process has been perturbed significantly because of the introduction of OP ligand-treated QDs. Panels a-g of Figure 2 are a series of SEM images of styrene/ OP ligand copolymer microspheres. They display the effect of OP ligand concentration on the formation of polystyrene beads without the presence of QDs. The reaction conditions for each sample are summarized in Table 1. Figure 2a shows polystyrene microspheres (PSMS) with nearly perfect spherical shapes. The average diameter is 1.24 µm with a polydispersity of 2%. With a small sample of OP ligands (4 mg), panel b also shows an

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Scheme 3. Schematic Illustration of the Formation of OP Ligand/Styrene Copolymer Microspheresa

a

The concentration of OP ligands is high.

Figure 3. SEM images of QD/PSMS microspheres. Samples a to e were prepared from 90 mg of styrene, 1 mg of AIBN, 20 mg of PVP, and 1.0 mL of ethanol. Sample f contains 90 mg of styrene, 1 mg of AIBN, and 1.0 mL of ethanol. The quantities of QDs and OP ligands for each sample are (a) 0.14 nmol/4 mg, (b) 0.28 nmol/8 mg, (c) 0.56 nmol/15 mg, (d) 1.13 nmol/30 mg, (e) 2.25 nmol/60 mg, and (f) 2.25 nmol/60 mg. All six samples have the same ratio between QDs and OP ligands.

average diameter of 1.24 µm, but with an increased polydispersity of 7%. As the concentration of the OP ligand increases, the size of styrene/OP ligand copolymer microspheres decreases from c

to f. Sample f has the smallest particle diameter of about 250 nm. The copolymer microspheres lose their smooth surface morphology and spherical shape when more OP ligands are added. Without the PVP stabilizer, the spheres in sample g have sizes and shapes that are similar to those of samples e and f. We find that, as the concentration of OP ligands increases, the spheres get smaller while the size distribution worsens. In addition, starting at sample c, the spheres begin to lose their surface morphology as well as their spherical shape. Scheme 2 gives a potential mechanistic explanation for the formation of OP ligand/ styrene copolymer microspheres where the OP ligands are at a low concentration as in sample b. OP ligands were prepolymerized for several hours before being introduced into the styrene/ethanol solution. Most of the OP ligands have formed short linear or branched chains. These small oligomers are quite soluble in ethanol (I). When the mixture is heated, AIBN decomposes and initiates styrene monomers or the MMA group on the OP ligands. Polymer chains begin to grow (II). The nucleation step (III) is quite similar to that for pure PSMS: Chains of styrene and OP ligand copolymer that have reached a critical length form small particles that are stabilized by PVP. After this nucleation process, monomeric styrene, OP ligands, and short chains of styrene or OP ligands diffuse into those particles and polymerize until they are used up (IV). In this case, short OP ligand oligomers behave similarly to styrene monomers and participate in both nucleation and growth processes. When the concentration of OP ligands is increased, the role that the OP ligands play changes, as shown in Scheme 3. The OP ligands are now highly cross linked during prepolymerization, and there are now many long chains of OP ligands in solution (I). Because of their phosphine groups, these long OP ligand chains are likely to form micelles in ethanol, with the phosphines facing out with the hydrocarbon groups inside the micelle (II). The reaction begins with the initiation of styrene monomers, OP ligands, or OP ligand chains. Monomers and short oligomers of

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Table 2. Reaction Conditions for QDs/PSMS a-f

a b c d e f

styrene (mg)

AIBN (mg)

PVP (mg)

ethanol (mL)

OP ligandsa (mg)

QDsa (nmol)

90 90 90 90 90 90

1 1 1 1 1 1

20 20 20 20 20 0

1.0 1.0 1.0 1.0 1.0 1.0

4 8 15 30 60 60

0.14 0.28 0.56 1.13 2.25 2.25

a The QDs and OP ligands in the six samples came from the same batch solutions. The concentration of QDs in the batch solution was determined by UV-vis measurement through the QD absorption cross section. After dispersing the QDs into OP ligand solutions, concentrations were determined from the total volume of the final solution.

styrene and OP ligands can diffuse into the OP ligand micelles, initiating polymer growth inside the micelles (II). As in the previous schemes, aggregates of polystyrene can separately form particles stabilized by PVP (III). There are now two types of nucleation sites in solution: PVP-stabilized particles of styrene oligomers and OP ligand micelles. Styrene monomers and OP ligands continue diffusing and polymerizing into both types of nuclei (IV). Because there are now many more nucleation sites that are inherently inhomogeneous in character, the average size of the final microspheres decreases as shown in Figure 2e and f, and the polydispersity further increases. This argument is consistent with sample g, which shows microspheres prepared as in Figure 2f but without PVP, indicating that prepolymerized OP ligand long chains do act as stabilizers. Figure 3 shows the effect of adding more ligands and quantum dots, at a fixed ratio, on the formation of QD/PSMS. The reaction conditions for each sample are summarized in Table 2. All of the samples have the same ratio of OP ligands and QDs. The trends are similar to those of the styrene-OP ligand copolymer microspheres without QDs (Figure 2). When the concentrations of OP ligand and QD increase, the sizes of QD/PSMS gradually decrease to as small as about 200 nm, as shown in Figure 3e.

Figure 4. SEM images of QD/PSMS prepared with different quantities of QDs. All four samples contain 4 mg of OP ligands. The QD quantities used in the experiments are (a) 0.15 nmol, (b) 1.5 nmol, (c) 3.0 nmol, and (d) 4.5 nmol.

The surface morphology is similarly roughened. Without using PVP, sample f has a size and shape that are similar to those of sample e. A comparison of Figures 2 and 3 shows that for the same concentration of OP ligands the presence of the QDs causes a further decrease in the size of the microspheres (e.g., Figure 2c and 3b). Scheme 4 provides a possible explanation for this observation. After the QDs are cap-exchanged with OP ligands, a fraction of the OP ligands remain free in the solution. After prepolymerization for a few hours, most of the MMA functional groups on the QDs are cross linked together. These OP ligand cap-exchanged QDs are quite soluble in ethanol. At the same time, free OP ligands in ethanol also polymerize through their MMA groups and form long or short OP ligand chains (I). Upon heating, the AIBN now initiates the polymerization of the styrene,

Scheme 4. Schematic Illustration of the Formation of QD-Embedded Polystyrene Microspheres (QD/PSMS)

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Figure 5. (a) STEM image of a slice of QD/PSMS. The elliptical shape results from the diamond knife cut during microtoming. (b) STEM image of the left part (in the white square) of the same slice at higher magnification. The brightest spots at the center and on the top edge are due to charging. (c) Energy-dispersive X-ray (EDX) analysis spectrum of the white dots in the STEM image.

the MMA groups on free OP ligands, and the small number of unreacted MMA groups on the QDs (denoted as MMA(QD)). Therefore, QDs can now copolymerize with the styrene and become functionalized with some solvate polystyrene oligomers. These QD/MMA/polystyrene oligomer constructs become unstable in ethanol. They likely arrange themselves as in a micelle, with the polystyrene oligomers pointing in and the MMA(QD) surface pointing out. Styrene monomers and MMA(QD)s can also diffuse into the micelle like OP ligand oligomers that have formed as in Scheme 3. In the growth step, monomeric styrene and QDs now diffuse into (1) PVP-stabilized particles, (2) OP ligand chain-stabilized particles, and (3) MMA(QD)/polystyrene micelles. As the concentrations of QDs and OP ligands increase in Figure 3 from a to e, there are increasing numbers of OP ligand oligomer-stabilized particles and MMA(QD)/polystyrene micelles. As a result, the number of nucleation sites increases, and the size of the microspheres becomes smaller. At high enough concentrations, the QDs and OP ligands effectively act as

stabilizers, and PVP is no longer needed, as shown in Figure 3f, supporting our hypothesis that QDs and OP ligand oligomers form micellar structures. The solutions with smallest concentration of QDs and OP ligands (Figure 3a and b) had nearly all of the QDs incorporated into the polymer beads. Free QDs could not be detected in the supernatant after centrifugation. The loading efficiency was therefore ∼100%. However, as the concentration of QDs and OP ligands increased, free QDs become clearly observable in the supernatant. At the concentration in Figure 3e, only 5-10% of the QDs are incorporated into the polymer beads, and 90-95% of the QDs remain in the supernatant. (Data were obtained by comparing the luminescence of QDs/PSMS dissolved in tetrahydrofuran to that of the starting solution of QDs/hexane.) Because of this limited loading efficiency, the volume fraction of QDs in polystyrene microspheres is no higher than 0.3% (1.6 mass %). The low loading efficiency is caused by the selfpolymerization of QDs capped with OP ligands. The following

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Table 3. Reaction Conditions for QDs/PSMS a-d

a b c d

styrene (mg)

AIBN (mg)

PVP (mg)

ethanol (mL)

OP ligands (mg)

QDa (nmol)

90 90 90 90

1 1 1 1

20 20 20 20

1.0 1.0 1.0 1.0

4 4 4 4

0.15 1.5 3.0 4.5

a The concentration of QDs in the four samples was determined by UV-vis measurement on the batch solution using the known absorption cross section of the QDs. The QDs were then dispersed in four indentical solutions of OP ligands.

Figure 6. Radial distribution function of QDs inside a microsphere obtained using Figure 5b. A square patch (220 nm × 220 nm) at the center of Figure 5b was chosen. For each QD in the patch, the number of QDs within a given radius up to 45 nm was counted. The number of QDs, normalized by the bulk density, is plotted as a function of distance.

four reactions summarize the polymerization process in the QDOP ligand-styrene mixture k11

MMA(QD)* + MMA(QD) 98 MMA(QD) - MMA(QD)* k12

MMA(QD)* + styrene 98 MMA(QD) - styrene* k22

styrene* + styrene 98 styrene - styrene* k21

styrene* + MMA(QD) 98 styrene - MMA(QD)* where * represents the active centers and the k’s are the reaction rate constants. From a classical analysis of monomer reactivity ratios in radical copolymerization,20 we expect that k11/k12 = 0.46 and k22/k21 = 0.52. These two numbers are less than 1, meaning that the copolymerization of MMA(QD)s with styrene is preferable over the polymerization of MMA(QD)s among themselves. However, these ratios are large enough that there is a reasonable probability of self-polymerization. The propensity for self-polymerization increases with higher concentrations of MMA(QD)s, becoming competitive with copolymerization with styrene and producing QD aggregates. These aggregates, which are much larger than styrene, free QDs, or small oligomers of styrene or OP ligands, do not easily diffuse into the polystyrene spheres. They either remain free in solution or adhere to the surface of the microspheres. (20) Odian, G. Principles of Polymerization, 4th ed.; Wiley-Interscience: Hoboken, NJ, 2004; p 492.

Figure 7. (a) Optical and (b) fluorescent microscope images of QD/PSMS using a 100× objective.

Figure 4 displays four samples prepared by increasing only the concentration of QDs at a fixed concentration of OP ligands. The reaction conditions for each sample are summarized in Table 3. The size of as-synthesized QD/PSMS decreases with a widening size distribution as the relative number of QDs increases. In addition, some agglomerates with elongated shape begin to appear in b to d. QD/polymer small aggregates are also observed in the supernatant. Increasing the concentration of QDs further resulted in the formation of a bulk solid. The trend is very similar to that shown in Figure 3. This implies that the small concentration of OP ligands used is sufficient to passivate as many as 4.5 nmol of QDs. The loading efficiency here also decreases as the concentration of QDs increases. As the ratio of QDs to OP ligand increases, it is possible that the surfaces of the QDs are not fully passivated by the OP ligands, leading to QD aggregation and precipitation from solution. If the QD quantity is above 4.5 nmol, then a bulk cross-linked solid is formed. The distribution of QDs in the microspheres was shown in Figure 5. Figure 5a is an STEM image of a 40-nm-thick slice of QD/PSMS. The spherical slice was elongated in one direction during microtoming. The longer radius is about ∼510 nm, and the shorter radius is ∼340 nm. The brightest spots in the middle and at the top edge are the result of damage by the electron beam. Figure 5b is the left part of the same slice (surrounded by white lines) at a higher magnification. Figure 5c is the energy-dispersive

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(C) represent the QD/hexane solution; and the dark triangles (D) represent the QD/PSMS ethanol solution. The integrated areas below each curve from 500 to 650 nm are 8.08 × 105 (A), 7.77 × 105 (B), 1.69 × 105 (C), and 9.64 × 104 (D). The peak position of QD/hexane emission is at 550 nm, and the peak position of QD/PSMS/ethanol emission is at 555 nm. These results give a relative quantum efficiency of 20% before and 12% after encapsulation, compared with the results for rhodamine 590. We also observed a red shift of ∼5 nm in the photoluminescence of QDs after encapsulation. Considering the QD distribution inside polymer beads discussed above, we believe that this red shift is due to energy transfer from smaller dots to larger dots. This energy transfer occurs when QDs are close enough together that their emission and absorption spectra overlap.21

4. Conclusions

Figure 8. Photoluminescence of rhodamine 590/methanol (O), rhodamine 590/polystyrene bead/methanol (b), QD/hexane (4), and QD/PSMS/ethanol (2) solutions. The excitation wavelength was 485 nm.

X-ray (EDX) analysis of the small white spots in Figure 5b. The presence of elemental cadmium, zinc, sulfur, and phosphorus confirms that the small white spots are CdSe/CdZnS/ZnS quantum dots. The copper signal comes from the TEM grid. Figure 6 displays the radial distribution of QDs obtained from a ∼220 nm × 220 nm square patch at the center of Figure 5b. The x axis is the center-to-center distance between pairs of QDs. The y axis is the ratio between the QD distribution density at a given separation and the bulk QD density. The distribution is flat with a weak peak at 9.0 nm, indicating that the QDs are largely uniformly distributed in the microsphere. The weak peak corresponds to dots separated by ∼5 nm surface-to-surface. The even distribution of QDs in the bead is the result of copolymerization between MMA(QD)s and styrene. The 9.0 nm peak likely indicates local close packing of QDs due to some selfpolymerization prior to incorporation into the microspheres. Panels a and b of Figure 7 are optical and fluorescent microscope images of QD/PSMS. Most of the QD/PSMS are less than 1 µm. All of the QD/PSMS are well separated from each other and fluoresce under UV excitation, proving that QDs have been well protected by OP ligands and successfully embedded into the polymer matrix. The photoluminescent quantum efficiency of QDs inside polystyrene beads was determined using an integrating sphere. Figure 8 shows the emission profiles of dyes and QD/PSMS using an integrating sphere to measure the quantum yield. The empty circles (A) represent the dye/methanol solution; the dark circles (B) represent the dye/methanol and pure PS beads mixture; the empty triangles

By using specially designed oligomer phosphine (OP) ligands, we have successfully encapsulated quantum dots in polystyrene microspheres. The chemical incorporation of the QDs into the polymer matrix through chemical bonds makes for a particularly robust system. QDs do not leak out of the beads even after highfrequency sonication, solvent exchange, and long shelf times, which makes these beads prime candidates for a variety of applications in harsh environments and thus potentially preferable for the work described in refs 10 and 11. Embedded QDs maintained their optical properties as a result of the protection by the OP ligands. When the incorporation reaction is carried out at a low concentration of QDs, as-synthesized QD/PSMS have a mean diameter of about 800 nm with a reasonable polydispersity and an even distribution of QDs within polymer beads and from bead to bead. However, efforts to encapsulate high concentrations of QDs result in the aggregation of QDs, preventing the formation of QD/PSMS. We suggest that this finding is not restricted to our specific example and generally limits the incorporation of quantum dots into growing polymer spheres produced by dispersion polymerization reactions. New strategies are therefore needed in order to load more QDs into microspheres. Acknowledgment. We thank Dr. Anthony Garratt-Reed for his assistance with the scanning transmission electron microscope and Dr. Preston Snee for his assistance with the calculation of the radial distribution function. This work made use of the MRSEC Shared Facilities supported by the NSF under award number DMR-0213282, as well as the Harrison Spectroscopy Laboratory (NSF-011370-CHE). This work was supported by the Institute for Collaborative Biotechnologies through grant DAAD 19-03D0004 from the Army Research Office. LA051973L (21) Kagan, C. R.; Murray, C. B.; Bawendi, M. G. Phys. ReV. B 1996, 54, 8633.