Temperature Controlled Fluorescence on Au@ Ag@ PNIPAM-PTEBS

Article pubs.acs.org/Langmuir

Temperature Controlled Fluorescence on Au@Ag@PNIPAM-PTEBS Microgels: Effect of the Metal Core Size on the MEF Extension Rafael Contreras-Caceres,*,† Paulino Alonso-Cristobal,‡ Diego Mendez-Gonzalez,‡ Marco Laurenti,‡ Ana Maldonado-Valdivia,§ Francisco Garcia-Blanco,‡ Enrique López Cabarcos,‡ Antonio Fernandez-Barbero,§ José Manuel Lopez-Romero,† and Jorge Rubio-Retama*,‡ †

Department of Organic Chemistry, Science Faculty, Málaga University, Málaga 29071, Spain Department of Physical-Chemistry II, Faculty of Pharmacy, Complutense University of Madrid, 28040 Madrid, Spain § Department of Physical-Chemistry, Almeria University, 04120 Almeria, Spain ‡

S Supporting Information *

ABSTRACT: In this work, we present a novel method to produce thermoresponsive, monodisperse microgels which display temperature-dependent photoluminescence. The system is based on bimetallic cores of Au@Ag encapsulated within thermoresponsive poly(N-isopropylacrylamide) microgels and coated with a photoluminescent polymer (poly[2-(3thienyl)ethoxy-4-butylsulfonate] (PTEBS) using the Layer-by-Layer technique. The electromagnetic radiation used to excite the PTEBS induces a local electromagnetic field on the surface of the bimetallic cores that enhances the excitation and emission rates of the PTEBS, yielding a metal enhanced fluorescence (MEF). This effect was studied as a function of the bimetallic core size and the separation distance between the PTEBS and the bimetallic cores. Our results permit evaluation of the effect that the metallic core size of colloidal particles exerts on the MEF for the first time, and prove the relevance of the metallic cores to extend the effect far away from the metallic surface. dielectric nanoparticles on different characteristics of fluorophores, such as absorption, peak shapes, fluorescence lifetime, and quantum yields for emission and for nonradiative energy transfer. They provided a theoretical understanding for the observed optical properties of the fluorophores when they were adsorbed on silver island films or silver nanoparticles. These preliminary works showed an increment in the PL intensity of certain fluorophores when they were close to a metallic surface. This effect was attributed to the local electromagnetic field generated by the collective oscillation of the conduction band electrons of the metallic surfaces,13,14 which is similar to the effect observed in surface enhanced Raman spectroscopy (SERS).15,16

1. INTRODUCTION Photoluminescent polymers (PLPs) constitute an interesting group of materials with a wide range of applications in biosensing, optoelectronics, electroactive polymers, solar cells, and light-emitting diodes.1−5 The photoluminescence of these polymers can be easily modified by changing the chemical structure or the environmental conditions. This responsiveness has prompted scientists to use them as sensors because their photoluminescent (PL) properties are superb compared with other organic dyes.6 Due to this environmental sensitivity, tiny variations in the medium can induce a dramatic change in the PL emission, making the PLPs sensitive to external stimuli such as pH, ionic strength, polarity, or polymer conformation.7−10 The presence of a local electromagnetic field increases tremendously the photoluminescent emission of the fluorophores.11 That has an enormous importance to increase the sensitivity of fluorescence analytical techniques. Gersten and Nitzan12 performed pioneering studies about the effect of © 2014 American Chemical Society

Received: September 29, 2014 Revised: November 26, 2014 Published: December 1, 2014 15560

dx.doi.org/10.1021/la503864f | Langmuir 2014, 30, 15560−15567

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Lakowicz17−20 and Cotton21 described the effects that the separation distance between silver particles and fluorophores produce on its fluorescence emission. These studies were later extrapolated to colloidal systems rendering fluorescent beacons that respond upon variations of this separation distance.22,23 However, to the best of our knowledge, there is no experimental information that assesses the influence of the metallic particles size on the distance range at which the MEF is extended. These points could help to clarify the roll of the local electromagnetic field on the MEF. For this reason, we have produced three types of monodisperse fluorescent microgels with a core@shell structure. The bimetallic core size was varied from 57 to 101 nm for each sample, yielding microgels with different metal enhanced fluorescence properties. The synthesized Au@Ag cores exhibited a broad surface plasmon resonance (SPR) band with two maximum peaks at 405 and 535 nm that overlapped the absorption band of the complexed PTEBS. In addition, the PNIPAM shell permitted us to control the separation distance by changing the temperature providing an easy way to study the distance dependency of MEF versus the metallic core size. The analysis of the steady-state fluorescence intensity and the radiative decay rate permitted us to determine the influence of the bimetallic core size on the enhancement of the absorption and the emission rate of the PTEBS. The results demonstrate the importance of the metallic core size to extend the effect far away from the metallic surface.

UV−Vis spectrophotometer. Transmission electron microscopy (TEM) studies were carried out using a JEOL JEM 2100 operated at 200 kV and a JEOL JEM 1010 operated at 80 kV. TEM samples were prepared by placing a drop of diluted solutions of each synthesized Au@Ag@PNIPAM hybrid microgels on a formvar coated copper grid. Dynamic light scattering (DLS) and Z-potential experiments were carried out using a Malvern Nano-ZS system equipped with a He−Ne laser working at 632.8 nm. The suspensions of microgels were diluted to a concentration of 0.02% (w/w) to avoid multiple scattering and to diminish colloidal interactions. The time correlation function of the scattered intensity, g(t) = ⟨I(0)I(t)⟩, was measured, and the mean hydrodynamic diameter was obtained as a function of the temperature. The UV-Vis and the PL spectra were taken after pouring 50 μL of the microgel dispersion in a quartz cuvette containing 3 mL of Milli-Q water at pH 7. The same sample dispersions were used to study the decay time measurements. 2.3. Synthesis of the Thermoresponsive Fluorescent Microgels. 2.3.1. Synthesis of PNIPAM Microgels with a Polystyrene Core (PS@PNIPAM). In this work, we have used 68 ± 8 nm polystyrene particles coated with PNIPAM (PS@PNIPAM) for control experiments. The synthesis of fluorescent PS@PNIPAM microgels was carried out in two steps. Initially, polystyrene nanoparticles with a mean diameter of 68 ± 8 nm were produced by radical emulsion polymerization and used as seeds for growing a PNIPAM shell. Briefly, 300 mg of sodium dodecyl sulfate, 2.5 g of styrene, and 25 mg of NIsopropylacrylamide were dissolved in 30 mL of water. The mixture was purged with nitrogen and mechanically stirred at room temperature (200 rpm, 20 min). The nitrogen inlet and outlet were removed, and the flask was placed into a preheated oil bath at 70 °C. The polymerization was initiated after 20 min by injecting 25 mg of potassium persulfate dissolved in 1 mL of water. After 12 h, the reaction was stopped and the particles were centrifuged three times at 13 000 rpm for 1 h. The hydrodynamic diameter of the polystyrene particles was measured by DLS (68 ± 8 nm). Afterward, 250 mg of NIsopropylacrylamide, 25 mg of N,N-methylenebis(acrylamide) (BIS) and 3.75 mL of the previously synthesized polystyrene dispersion were mixed in 6.25 mL of milli-Q water. The flask was sealed with a septum and heated up to 70 °C under nitrogen atmosphere. The polymerization was started after adding 25 mg of potassium persulfate dissolved in 1 mL of water. The reaction was stopped after 4 h and the particles centrifuged three times at 13 000 rpm for 1 h. The precipitate containing the PS@NIPAM particles was dispersed in 10 mL of water and stored. 2.3.2. Synthesis of Au@PNIPAM Microgels. Initially, 45 nm of spherical Au nanoparticles were synthesized by seed-mediated growth.24 Briefly, 35 mL of ∼15 nm spherical gold nanoparticles (prepared by citrate reduction)25 were diluted with 15 mL of a 0.03 M CTAB aqueous solution. Then, 50 mL growth solution containing ascorbic acid (1 × 10−3 M), HAuCl4 (0.5 × 10−4 M) and CTAB (0.015 M) at 35 °C were prepared. Finally, 2.76 mL of the previously prepared 15 nm gold nanoparticles were added under middle magnetic stirring and after 30 min the excess of surfactant was removed by centrifugation at 4500 rpm for 30 min. The supernatant was discarded and the precipitate was redispersed in 50 mL of Milli-Q water. The Au core size measured by TEM was 45.31 ± 2.49 nm. The Au@PNIPAM particles were synthesized using these Au nanoparticles as seeds. The colloidal dispersion containing spherical 45 nm Au nanoparticles was heated at 70 °C. Then, 50 μL of 3-butenoic acid were added, vigorously stirred for 30 seconds, and kept at 70 °C for 1 h. After this time, the mixture was cooled to room temperature and the excess of 3butenoic acid was removed by centrifugation at 4500 rpm for 30 min. The precipitate was redispersed with 50 mL of 4 mM CTAB and centrifuged again. The precipitate containing the 3-butenoic acid functionalized Au nanoparticles was redispersed in 10 mL of Milli-Q water. This solution was heated at 70 °C under nitrogen atmosphere and N-Isopropylacrylamide (0.1698 g) and N,N′ methylenebis(acrylamide) (0.0234 g) were added under magnetic stirring. After 15 min, the nitrogen gas flow was removed, and 2,2′-Azobis(2methylpropionamidine) dihydrochloride (100 μL 0.1 M) were added.

2. EXPERIMENTAL SECTION 2.1. Materials. Ascorbic acid (reagent grade), N,N′-Methylenebis(acrylamide) (99%), potassium persulfate (≥99%), sodium dodecyl sulfate (≥98.5%), styrene (≥99%), silver nitrate (99%), trisodium citrate (≥99%), were purchased from Sigma. N-Isopropylacrylamide (NIPAM, 97%), butenoic acid (≥97%a) sodium borohydride (NaBH4, ≥98%), Chloroauric acid trihydrate (HAuCl4·3H2O, 99.999%), polyethylene imine (Mw, 10 000 aqueous solution 10% w/v) where supplied by Aldrich. Hexadecyltrimethylammonium bromide (CTAB, ≥99%) was supplied by Fluka and 2,2′-Azobis(2-methylpropionamidine) dihydrochloride (V50, 99%) was supplied by Acros Organics. Sodium poly[2-(3-thienyl)ethoxy-4-butylsulfonate] (PTEBS) with average molecular weight of 269 000 g/mol was purchased from American Dye Source. 2.2. Characterization. The fluorescence intensity decays were analyzed in terms of the multiexponential model as the sum of individual single-exponential decays:

I(t ) =

∑ αi·e−(t / τi) i

In this expression, τi represents the decay times, αi represents the amplitudes, and Σ·αi = 1. The fractional contribution of each component to the steady-state intensity is described by the following: αi·τi fi = ∑i αi·τi The average lifetime is represented by the following: τ ̅ = fi ·τi The amplitude-weighted lifetime is given by the following: ⟨τ ⟩ =

∑ αi·τi i

The values of αi and τi were recorded using a QM3PH instrument from Photon Technology International. The goodness of the fit was analyzed by DW and χ2. Photoluminescence (PL) spectra were collected using a JASCO spectrofluorometer equipped with a thermostatic bath for temperature control. Absorption spectra were taken with a Varian Cary 300 Bio 15561


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metric analysis). The as-synthesized PS@PNIPAM, Au@Ag1@ PNIPAM, Au@Ag2@PNIPAM, and Au@Ag3@PNIPAM microgels were negatively charged, as it was demonstrated by the Z-potential measurements. For this reason, it was necessary to cover the microgels first with polyethylenimine (PEI), a positively charged polymer. After that, the surface of the microgels became positively charged and suitable to complex PTEBS, which is a negatively charged polyelectrolyte. This procedure was performed as follows; 5 mL of an aqueous dispersion of microgels at pH 5 were poured onto 5 mL of a PEI solution (40 mg/mL) under vigorous stirring and left for 48 h. After this time, the microgels were purified by centrifugation at 13 000 rpm for 1 h. This purification step was repeated three times. The supernatant was discarded, and the precipitate was dispersed in 5 mL of water at pH 5. After the PEI deposition, the surface charge of the microgels became positive. Then, the microgel dispersion was dropped onto 10 mL of a PTEBS solution (2 mg/mL) under vigorous stirring and left to react for 48 h. After this time, the microgels were centrifuged at 13 000 rpm for 1 h, and the precipitate was dispersed with 5 mL of water at pH 5. This purification was repeated three times. Finally, another layer of PEI was deposited using the same procedure used above. As a result, the PTEBS fluorescent polymer was deposited onto the microgels in a sandwich-like structure (PEI@PTEBS@PEI).

After 2 h, the reddish turbid dispersion was allowed to cool down to room temperature, diluted up to 50 mL with Milli-Q water and centrifuged at 4000 rpm for 30 min. The centrifugation step was repeated 5 times for the complete purification of Au@PNIPAM microgels and the removal of any unreacted monomers/oligomers and free PNIPAM microgels. The general synthetic process is depicted in Scheme 1A.

Scheme 1. (A) Schematic Representation of the Synthesis of Au@PNIPAM Particles and for the Growth of a Silver Shell Using Different R* Values. (B) Representation of the Layer by Layer (LBL) Deposition Performed for the Complexation of PTEBS on Au@Ag@PNIPAM Particles

3. RESULTS AND DISCUSSION The synthetic route used to obtain the photoluminescent material began with the growth of PNIPAM shell around the 45 nm spherical Au nanoparticles. Then, the Au cores were coated with Ag using different quantities of AgNO3 and reducing reagents, yielding different shell thickness. To produce the Au@Ag cores, we used a methodology that employs Au nanoparticles as seeds for the silver growth. This technique permitted us to ensure the synthesis of highly monodisperse bimetallic cores, with controlled sizes and with superb plasmonic properties, which would render an intense electromagnetic field. The Au@Ag core size measured by TEM was 57.4 ± 5.4 nm for Au@Ag1, 78.6 ± 5.7 nm for Au@Ag2 and 101.6 ± 12.4 nm for Au@Ag3. Table 1 summarizes the silver Table 1. Amount of Silver Nitrate Used in Each Case and Dimensions of the Bimetallic Core for Au@Ag1PNIPAM, Au@Ag2@PNIPAM, and Au@Ag3@PNIPAM Microgels

2.3.3. Silver Growth and Bimetallic Au@Ag@PNIPAM Production. The Au@Ag@PNIPAM particles were prepared using the assynthesized Au@PNIPAM microgels as seeds. Three different Au@ Ag@PNIPAM particles with different silver shell thickness were grown and named as Au@Ag1, Au@Ag2, and Au@Ag3. The silver shell growth was performed by a procedure previously published,26 using R* ratios of 1.2, 6, and 12, where R* is the molar ration between Ag+ and the Au atoms in the Au@PNIPAM solution. First, 10 mL of a growth solution containing 0.4 M glycine buffer solution (adjusted at pH 9.5), Au@PNIPAM particles ([Au] = 0.25 mM) and 50 mM CTAB was prepared. For each growth solution different amount of AgNO3 (300 mM) and ascorbic acid (100 mM) solutions were added under gentle magnetic stirring: 10 μL AgNO3 and 30 μL ascorbic acid for Au@Ag1; 50 μL AgNO3 and 150 μL ascorbic acid for Au@Ag2; and 100 μL AgNO3 and 300 μL ascorbic acid for Au@Ag3. A representation of the process is depicted in Scheme 1A. 2.3.4. Deposition of PTEBS on the Surface of Au@Ag@PNIPAM Microgels. The deposition of the photoluminescent polymer (PTEBS) was carried out by using the layer by layer (LBL) technique,27 see Scheme 1B. Prior to the PTEBS deposition, the PS@PNIPAM and the Au@Ag@PNIPAM microgels were diluted to obtain a particle concentration of 1 × 1011 particles/mL (determined by thermogravi-

sample

R* = [Ag+]/[Au]

Aucore (nm)

AuAg (nm)

Agshell (nm)

AuAg1 AuAg2 AuAg3

1.2 6 12

45.3 45.3 45.3

57.4 78.7 101.6

6.05 16.7 28.2

concentration, gold core size, bimetallic Au@Ag size, and silver shell of the as-synthesized systems. The whole procedure permitted to obtain monodisperse microgels with bimetallic cores with a controllable diameter. In this hierarchical structure, the Ag shell provides an intense surface plasmon resonance band absorption due to the high interband transitions (∼3.2 eV), which minimizes the dumping of the plasmon.28 In addition, we also produced PS@PNIPAM microgels that were used as controls in the photoluminescent experiments. The synthesized Au@PNIPAM and PS@PNIPAM microgels were monodisperse with a mean diameter of 130 ± 10 nm as measured by TEM (see Figure S1B in the Supporting Information, SI). The deposition of the silver layer onto the Au core of the Au@PNIPAM microgels produced Au@Ag@ PNIPAM microgels, which are depicted in Figure 1. Figure 1A−C show that under the TEM chamber conditions, the mean diameter of the hybrid microgels remained constant around 180 nm independently on the shell thickness of Ag 15562


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Figure 1. Representative TEM micrographs of the Au@Ag@PNIPAM microgels with different Ag shell thickness, (A) Au@Ag1@PNIPAM, (B) Au@Ag2@PNIPAM, and (C) Au@Ag3@PNIPAM microgels. A1, B1, and C1 are detailed HR-TEM images of each sample while the insets A2, B2, and C2 represent the electron energy loss of Au (green) and Ag (red) chemical mapping of each sample.

deposited onto the Au core. A detailed analysis using highresolution TEM (insets A1, B1, and C1 of Figure 1) revealed that as result of the reduction of Ag+ the size of the bimetallic core increased through the formation of a shell, which is lighter (less electrodense) than the Au nucleus. The electron energy loss spectroscopy (EELS) (insets A2, B2, and C2 of Figure 1) corroborated that this shell is made of silver. The thickness of the Ag outer shell was controlled from 6 to 28 nm by changing the concentration of Ag+ and reducing agent used in the reaction. As result, it was possible to obtain bimetallic cores with different sizes. Table 1 summarizes R* = [Ag+]/[Au], the gold core size, the final bimetallic core size and the thickness of the silver shell for Au@Ag1PNIPAM, Au@Ag2@PNIPAM, and Au@Ag3@PNIPAM samples. In aqueous dispersion, the mean hydrodynamic diameter of each hybrid microgels was 300 nm (measured by DLS). The lack of variation in the mean hydrodynamic diameters of these microgels upon depositing the Ag shell indicated that the polymer matrix did not interact with the ions during the growth step, which occurs from the surface of the metallic core toward the outer part, without collapsing or modifying the swelling state of the polymer. This fact is tremendously important, since it ensured us to have hybrid microgels with the same outer polymer shell in terms of specific surface and chemical behavior, but with different bimetallic core sizes. As a result of the presence of a bimetallic core within the microgels, these systems exhibited optical properties due to the localized surface plasmon resonance (LSPR). This phenomenon was studied by UV−Vis spectroscopy, see Figure 2A. The absorption band with the maximum at 530 nm (black line in Figure 2A) was attributed to the collective oscillation of the electrons of the Au surface. In addition, we observed a small band located at 650 nm due to the presence of Au cylinders in the sample. The UV−Vis absorption spectra of the microgels after the Ag deposition changed due to the different optical properties between Ag and Au. In all cases, the UV−Vis absorption spectra revealed the existence of a LSPR band with a

Figure 2. (A) UV−Vis spectra of the different hybrid microgels; Au@ PNIPAM (black line), Au@Ag1@PNIPAM (blue line), Au@Ag2@ PNIPAM (red line), and Au@Ag3@PNIPAM (green line). (B) Absorption spectra of the hybrid microgels after the PTEBS deposition. The dotted black line corresponds to the fluorescent PS@PNIPAM-PTEBS microgels particles that were used as references.

maximum at 420 nm attributed to the silver layer.26 The intensity of LSPR bands increased when the thickness of the silver shell grew. This is due to the different contribution of the dielectric function of silver for each core-shell nanoparticle.29,30 Interestingly, when the PTEBS was deposited onto the surface of the hybrid microgels a new absorption band appeared in the spectra at 435 nm, which completely overlapped the SPR bands of the bimetallic cores, see Figure 2B. In these hierarchical hybrid microgels, the PNIPAM shell acts as a separation layer, maintaining the PTEBS apart from the surface of the bimetallic core. However, the thermal responsiveness of PNIPAM was able to control the swelling degree as a function of the temperature and, therefore, to 15563


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modify the separation distance between the PTEBS and the bimetallic cores. In fact, when the temperature was above 32 °C, the polymer matrix collapsed, and consequently, the distance between the bimetallic cores and the PTEBS was reduced. The distance shortening induced after increasing the temperature was studied by dynamic light scattering (DLS), see Figure 3.

Figure 4. Fluorescence intensity of the synthesized microgels; PS@ PNIPAM-PTEBS (black line), Au@Ag1@PNIPAM-PTEBS (green line), Au@Ag2@PNIPAM-PTEBS (blue line), and Au@Ag3@ PNIPAM-PTEBS (red line) at (A) 25 °C and (B) 50 °C, respectively. Excitation wavelength 405 nm. Figure 3. Temperature dependence of the hydrodynamic diameter for the different fluorescent hybrid microgeles versus the temperature; Au@Ag1@PNIPAM-PTEBS (blue line), Au@Ag2@PNIPAM-PTEBS (red line), and Au@Ag3@PNIPAM-PTEBS (green line).

As one can observe in Figure 3, these microgels had a lower critical solution temperature (LCST) close to 35 °C, which was slightly higher than the LCST observed for pure PNIPAM microgels (32 °C).31 This could be related to two effects: First, the electrostatic repulsion that the outer layer of PEI could impose, and second, the steric hindrance that the PEI@PTEBS inner layers could introduce in the thermal-responsive matrix. Both effects would hamper the aggregation of the PNIPAM, and consequently, the hybrid microgels would require a higher temperature to overcome these barriers and collapse its structure.32,33 In all the samples, the mean hydrodynamic diameter of the microgels was reduced from 300 nm at 20 °C to a minimum of 190 nm at 50 °C. This reduction brought the PTEBS closer to the surface of the bimetallic core, which produced an enhancement of the fluorescence intensity of the complexed PTEBS, as shown in Figure 4. Figure 4 shows that for all the hybrid microgels having a bimetallic core, the temperature rising produced an enhancement of the fluorescence intensity. This increment was more intense for those microgels with larger cores. This fluorescence enhancement could be affected by two factors: the distance shortening between both components and the bimetallic core size. In order to distinguish between the contribution of both factors, the fluorescence enhancement was studied for each sample as a function of the temperature in the range between 20 °C and 50 °C, see Figure 5. Figure 5A shows that the Normalized Fluorescence Enhancement increased steeply with the temperature, reaching a plateau at temperatures above 45 °C. At this temperature, the microgels were completely collapsed and therefore the separation distance between the PTEBS and the bimetallic surface was at a minimum. These data were related with dynamic light scattering results to infer the fluorescent enhancement as a function of the separation distance for

Figure 5. (A) Normalized Fluorescence Enhancement as a function of the temperature observed for PS@PNIPAM-PTEBS (black), Au@ Ag1@PNIPAM-PTEBS (green), Au@Ag2@PNIPAM-PTEBS (blue), and Au@Ag3@PNIPAM-PTEBS (red). The normalization was done by dividing the fluorescence intensity of each sample by the mean fluorescence intensity obtained from the PS@PNIPAM-PTEBS microgels obtained in the experimental temperature range. (B) Normalized Fluorescence Enhancement as a function of the separation distance between the bimetallic core and the PTEBS. Excitation wavelength 405 nm.

each sample, see Figure 5B. This reveals two results; first that the fluorescence enhancement increased as the separation distance diminished. This result may be due to the modification of the electric field near the metal surface, which is more intense for larger metallic cores. This modification would change both the field applied to the PTEBS and the field radiated by it. As a consequence, the local electromagnetic field could increase the number excited fluorophores (rate of absorption) by a factor of Nabs as well as the radiative decay rate by a factor of Ne.34,35 Second, we observe that at larger distances, all the samples displayed a fluorescence enhancement, which was also strongly affected by the bimetallic core size. The existence of this fluorescence enhancement could be 15564


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exponential decay functions were necessary to describe the experimental data fit (χ2 between 1.1 and 1.3, residuals 1.7) in all the experiments. The fluorescence intensity decay time was analyzed at 25 and 50 °C, which corresponded to the maximum and the minimum separation distance between the PTEBS and the bimetallic cores, respectively. The average decay time of the PS@ PNIPAM microgels remained constant at both temperatures, as shown in Figure 6B. On the contrary, the average decay time of the Au@Ag@PNIPAM-PTEBS microgels showed a progressive reduction that was the highest for the Au@Ag3@PNIPAMPTEBS microgels at 50 °C. This was attributed to the distance between the fluorophores and the bimetallic surface, which was the lowest in the Au@Ag3@PNIPAM-PTEBS microgels. Our experimental data of fluorescence intensity and timeresolved fluorescence were fitted to eq 5, considering that Nabs(d=0), Ne(d=0), Rabs, and Re are free parameters for each sample as it was previously published by Lackowiz et al.19 The results are summarized in Table 2.

related with the increment of the light scattering provoked by larger metal cores. As result of the increment of the light scattering, the possibility to excite a higher number of fluorophores should increase, that would result in an augmentation of the excitation rate of the fluorophores. Under this scenario, the total fluorescence intensity in the presence of a metallic surface (Im) can be described in terms of a phenomenological model that accounts for the expected interactions. These interactions consist in an increment of the radiative decay by a factor Ne and an increase in the rate of excitation by a factor of Nabs, both factors being (Ne, Nabs) distance dependent. φ Imet = Io· met [Nabs(d)] φo (1) Imet = Io·[Ne(d)]·

τ0 ·[Nabs(d)] τmet

(2)

For each sample, the augmentation of the emission (Ne) and excitation (Nabs) rates depends on the distance (d) between the metal and the fluorophore. Therefore, the fluorescence enhancement by metallic surface can be described in an easy way as a distance depending factor: Ne(d) =

Ned = 0·e(−d / R e)

+1

d = 0 (−d / R abs) Nabs(d) = Nabs ·e +1

Table 2. Nabsd=0, Ned=0, Rabs, and Re Obtained for Each Sample after Being Fitted with Eq 5

(3) (4)

where Re and Rabs are the characteristic distances over which these effects decrease to 1/e exponentially. Nabs(d=0) and Ne(d=0) are the absorption and emission rates when the distance between polymer and the bimetallic core is 0, and they should be dependent on the size of the bimetallic cores. For that, the fluorescent intensity (eq 1) can be rewritten in terms of τm and τ0, and fluorescent enhancement (eq 5): F (d ) = τ0 τmet

sample

Nabs(d=0)

Ne(d=0)

Rabs(nm)

Re(nm)

Au@Ag1 Au@Ag2 Au@Ag3

1.49 1.98 2.35

2.35 3.27 5.12

21 18 13

115 69 51

These results demonstrate that the core size affects the Nabs(d=0), Ne(d=0), in such a way that the absorbance and emission rates were substantially enhanced when the Ag shell was thicker. Such an effect could be related with the intensity of the electromagnetic field generated by the bimetallic core, which is supposed to be much more intense in the case of Au@ Ag3@PNIPAM-PTEBS. This factor, together with the minimum separation distance between the photoluminescent polymer and the metallic surface in the samples Au@Ag3@ PNIPAM-PTEBS, provoked and intensified the observed fluorescence intensity.

Im d = 0 (−d / R abs) = [Nabs ·e + 1]·[Ned = 0·e(−d /R e) + 1]· I0 (5)

4. CONCLUSIONS We have developed hybrid microgels with photoluminescent and thermoresponsive properties. These microgels were composed by a bimetallic Au@Ag core with tunable silver shell thickness, coated with a PNIPAM shell, and wrapped with a PTEBS layer by using the Layer by Layer technique. With these systems, we studied the Metal-Enhanced Fluorescence of the PTEBS by the Au@Ag cores. The thermoresponsive PNIPAM shell permitted to control the distance between the PTEBS layer and the bimetallic core surface by varying the temperature. When the temperature was above the lower critical solution temperature of the PNIPAM, the fluorophoreto metal distance was shortened, and the fluorescence intensity increased. Our control experiments demonstrated that this fluorescent enhancement was due to the bimetallic core. In addition, we observed that at large separation distance, the enhancement effect was more intense for the particles with larger bimetallic cores. This result could be attributed to the increment of the light scattering provoked by the bigger cores, which could increase the number of excited fluorophores. The time-resolved fluorescence spectroscopy analysis revealed that the presence of the bimetallic core shortened the average decay time. These data permitted us to infer that the metal enhanced

In order to solve eq 5, we studied the fluorescence enhancement with time-resolved fluorescence measurements, see Figure 6A. For fitting the decay−time profiles, two

Figure 6. (A) Intensity decay−time profile of the microgels at 25 and 50 °C, λexc = 460 nm. (B) Representation of the average decay time for all the samples at 25 °C (blue) and 50 °C (red) λexc = 460 nm. 15565


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fluorescence occurred by increasing both the absorption (Na) and the emission (Ne) rates, which were directly related with the increment of the core size.

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ASSOCIATED CONTENT

S Supporting Information *

(Table 1S) Z-potential of the micro gels along the LbL deposition step; (Figure S1) representative TEM micrographs for (A) Au@PNIPAM microgels used as template in the synthesis of Au@Ag@PNIPAm microgels and (B) PS@ PNIPAm microgels used for the preparation of the reference fluorescent particles; additional equations; and an additional reference. This material is available free of charge via the Internet at http://pubs.acs.org.

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AUTHOR INFORMATION

Corresponding Authors

*E-mail: [email protected]. *E-mail: [email protected]. Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS The Marie Curie COFUND program “U-mobility” cofinanced by University of Malaga and the European Communitýs Seventh Framework Programme under Grant Agreement No. 246550 and the Spanish Ministry of Science for the projects MAT2010-15349 and CTQ-48418P are gratefully acknowledged for the financial support of this project. P.A. acknowledges the Spanish Ministry of Education for the FPU grant AP2010-1163.

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Temperature Controlled Fluorescence on Au@ Ag@ PNIPAM-PTEBS

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