Subscriber access provided by ONDOKUZ MAYIS UNIVERSITESI
Article
Anomalous diffusion in thermoresponsive polymer-clay composite hydrogels probed by widefield fluorescence microscopy Beate Stempfle, Anna Große, Bernhard Ferse, Karl-Friedrich Arndt, and Dominik Wöll Langmuir, Just Accepted Manuscript • DOI: 10.1021/la503571j • Publication Date (Web): 30 Oct 2014 Downloaded from http://pubs.acs.org on November 9, 2014
Just Accepted “Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted online prior to technical editing, formatting for publication and author proofing. The American Chemical Society provides “Just Accepted” as a free service to the research community to expedite the dissemination of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully peer reviewed, but should not be considered the official version of record. They are accessible to all readers and citable by the Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore, the “Just Accepted” Web site may not include all articles that will be published in the journal. After a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web site and published as an ASAP article. Note that technical editing may introduce minor changes to the manuscript text and/or graphics which could affect content, and all legal disclaimers and ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or consequences arising from the use of information contained in these “Just Accepted” manuscripts.
Langmuir is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.
Page 1 of 13
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60
Langmuir
Anomalous diffusion in thermoresponsive polymer-clay composite hydrogels probed by widefield fluorescence microscopy Beate Stempfle,1 Anna Große,2 Bernhard Ferse,2 Karl-Friedrich Arndt,2 Dominik Wöll*1,3,4 1
Faculty of Chemistry, University of Konstanz, Universitätsstr.10, 78464 Konstanz, Germany. Physical Chemistry of Polymers, Department of Chemistry, Technical University Dresden, Mommsenstraße 13, D-01062 Dresden, Germany 3 Zukunftskolleg, University of Konstanz, Universitätsstr. 10, 78464 Konstanz, Germany. 4 Institute for Physical Chemistry, RWTH Aachen University, Landoltweg 2, 52074 Aachen, Germany. 2
Abstract Thermoresponsive materials bear an enormous potential for tissue engineering, separation systems and drug delivery. We investigated the diffusion of laponite clay nanoparticles, which serve as physical crosslinker to achieve improved material properties in Poly(Nisopropylacrylamide) (PNIPAM)-clay composite hydrogels close to the gel point. The networks are formed through physical interactions between PNIPAM chains and clay nanoparticles after mixing these two components. In contrast to previous studies, a covalent labeling strategy was chosen in order to minimize the amount of free dyes in solution. Single particle tracking of the labeled clay nanoparticles showed that their diffusion is anomalous at all temperatures used in this study reflecting the viscoelastic behavior as a crosslinker. Stepwise heating from 25 °C to 38 °C resulted in a slight rise of the diffusion coefficient and the anomality parameter up to the volume phase transition temperature of ca. 31 °C which was followed by a significant drop of both parameters reflecting strongly hindered motion of the collapsed nanoparticle aggregates.
Introduction Smart hydrogels experience a rising interest due to their multiple applications such as drug delivery systems, artificial muscles and micro-machines.1 Their “smartness” originates from the fact that these hydrogels react to an external stimulus such as temperature, pH or ionic strength. The most prominent example is crosslinked PNIPAM which, in water, exhibits a reversible volume phase transition at the lower critical solution temperature (LCST) of the non-crosslinked polymer in water (32 °C),2, 3 which is very interesting for biological and medical applications. Below this temperature the PNIPAM-hydrogel is swollen, at higher temperature it collapses expelling water. The reason for this phenomenon is a good balance between hydrophilic and hydrophobic interactions in the polymer chain. During the phase transition the water bound to the polymer chain is released in an endothermic process.4 The LCST depends on the conditions of the aqueous environment, such as presence of salts and surfactants,5, 6, 7 and can be varied by functionalization of the polymer structure. For PNIPAM copolymers the LCST is strongly influenced by the nature of the comonomer. Hydrophobic 1 ACS Paragon Plus Environment
Langmuir
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60
compounds lower the LCST and hydrophilic compounds raise it.8 Furthermore, it depends on polymer architecture (see Picos-Corrales et al.9 and reference herein). Conventional chemically crosslinked PNIPAM hydrogels bear several disadvantages limiting their application possibilities.10 They are rather brittle and thus readily broken even at low deformations in elongation, bending or compression. Their swelling and deswelling kinetics are rather slow. The reason for these disadvantages is the heterogeneous distribution of crosslinks (junction points). The hydrogels look opaque at temperatures below LCST of PNIPAM in water due to permanent structural inhomogeneities on a scale of optical wavelengths.11 To circumvent these problems, Haraguchi et al. developed a nanocomposite hydrogel consisting of a responsive organic polymer and inorganic clay nanoparticles acting as threedimensional crosslinking agents.10, 12 This opened the possibility to vary the crosslinking density and the molecular weight of the polymer chains independently.10, 13, 14 It allows for a better fine-tuning of the material properties of the hydrogel in order to synthesize transparent hydrogels with extraordinary mechanical toughness and rapid swelling and deswelling kinetics. In our case, the networks do not result from a chemical reaction forming covalent bonds between clay nanoparticles and polymer chains, but through physical interactions. It is well known that aqueous mixtures of clay and polymers can form so-called shake-gels simply by mixing the two components, e.g. mixtures of clay and polyethyleneoxide (PEO).15, 16 In contrast to crosslinking with chemical reactions, the advantage of this method for the formation of the network is that very mild conditions can be used and no unwanted low molecular weight components or initiator residues remain in the network. The properties related to the structure and dynamics within the clay-crosslinked hydrogel can strongly depend on temperature. Even though molecular pictures were developed,17 a detailed understanding of this relationship is still lacking. The strong bonding between the polymer chains and the clay surface can be attributed to physical interactions such as hydrogen bonds.11, 18 The influence of the temperature on these physical interactions and on the formation of these networks is of particular interest because the temperaturesensitive polymer PNIPAM can change its conformation between coil and globule. This conformational change is expected to affect the network formation of the shake gel. At low temperatures the polymer chains are associated to the clay particles. Rising the temperature above the volume phase transition temperature (VPTT) causes the polymer chains to collapse and to get physically much closer to the layered silicate surface. This should also influence the diffusion coefficient of the clay particles. For a deeper insight, the real-time in situ observation of the dynamics of single clay particles on the nano- to microscale is essential. Fluorescence microscopy is an ideal tool to study the diffusion of single molecules and particles including diffusional heterogeneities in polymer systems. Different techniques have been used to access motion on different time scales. Fluorescence correlation spectroscopy (FCS) has contributed significantly to this field,19, 20 becomes however not practical for slow moving molecules and particles with diffusion coefficients lower than ca. 10-14 m2 s-1 when motion through the confocal volume in average takes several minutes to hours. Such lower diffusion coefficients can be obtained by fluorescence microscopy with single molecule/particle tracking.21, 22 The advantage of this method is that the trajectories of single fluorescent objects can be followed and statistically analyzed using typically their time 2 ACS Paragon Plus Environment
Page 2 of 13
Page 3 of 13
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60
Langmuir
averaged mean square displacement (TAMSD)23. This yields diffusion coefficients and also reveals the diffusion mode, i.e. whether diffusion is normal or anomalous.24 However, it has to be carefully distinguished between different models resulting in anomalous diffusion.25, 26, 27, 28 In particular, three different types of random walk models have been considered: obstructed diffusion describes diffusion through immobile obstacles, continuous time random walk consider power-law distributed rests between periods of free diffusion, and fractional Brownian motion accounts for the motion of particles in viscoelastic media.27 The latter case applies to the PNIPAM-clay composite investigated here.
Experimental Polymer The PNIPAM homopolymer was obtained by a free radical polymerization. For this, 5 g of Nisopropylacrylamide (NIPAM, 0.0442 mol, 99% purity, Acros Organics) and 6 mg of azobisisobutyronitrile (AIBN, 0.036 mmol, 99% purity, Acros Organics) were dissolved in 85 ml of 1,4-dioxane (for synthesis, VWR). The mixture was purged with argon for 15 min and allowed to react in an oil-bath at 70 °C for 7 h under argon atmosphere. The polymer was precipitated in ice cold diethyl ether (-70 °C, for synthesis, VWR) and purified by 3 × reprecipitation from THF (for analysis, Acros Organics) into diethyl ether. Finally, the polymer was freeze-dried. The molecular weight and the molecular weight distribution of the polymer were determined by gel permeation chromatography with a JASCO instrument equipped with a triple detection, that includes a WYATT Technology DAWN DSP multi angle laser light scattering (MALLS)-detector, a RI-detector (RI-930) and a Viscotek viscosity detector (model 250). The samples were measured at 25 °C using dimethylacetamide as the mobile phase. The flow rate was 1 ml/min. This yielded a polymer with a weight-averaged molecular weight Mw of 116 kg/mol, a number-averaged weight Mn of 60 kg/mol, and a polydispersity Đ of 1.95.
Clay nanoparticles For our studies, we used nanoparticles consisting of Laponite S482 (Rockwook Inc.), a synthetic hectorite-type clay with the formal composition Na0.7+[(Si8Mg5.5Li0.3)O20(OH)4]0.7(see Fig.1) in which negatively charged counterions (mainly pyrophosphate-groups) are attracted to the positively charged edges of the disc shape particles thus preventing the formation of aggregates. In the dry state, the clay nanoparticles used in this study exhibit a height of ca. 1 nm and a diameter of ca. 30 nm. When suspended in water, they swell to approximately 40 nm in diameter and 3.5 nm thickness.29 For the clay nanoparticles, a polydispersity index of 0.564 was determined using dynamic light scattering (DLS) assuming spherical particles. DLS was carried out on commercial laser light scattering spectrometer (Malvern Nano ZS 3600) and analyzed with Malvern-Software. 3 ACS Paragon Plus Environment
Langmuir
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60
Page 4 of 13
®
Fig.1 Schematic drawing of the investigated Laponite clay nanoparticles including their charge distribution: the upper and lower side of the nanoparticles are formed by negatively charged silicate groups and their positive counter ions which can be readily exchanged in aqueous solution. At the edges, the counter ions are coordinated thus yielding a net positive charge.18, 30
Fluorescence-labeling of clay nanoparticles Clay nanoparticles have previous been labeled with rhodamine B by non-covalent interactions.11 However, for single particle tracking using widefield fluorescence microscopy (WFM), a covalent labeling strategy was used in the current work. A perylenediimide derivative was chosen as fluorophore since this class of dyes has been successfully used in previous studies of us31 and other groups.32
Fig.2 Perylenediimide derivative 1 (for synthesis see Supporting Information) used in this work to label the clay
nanoparticles. The N-hydroxysuccinimide group was reacted with the amino groups on the modified clay edges.
HO-Si Si Si-OH OH
PDI-NHS toluene 7 h reflux
CH2Cl2, 5 d, soxhlet extraction Fig.3 Fluorescence labeling of clay nanoparticles.
First, the edges of the laponites were silanized using 3-aminopropyldimethylethoxysilan (APES) bearing an amino group to attach the functionalized dyes in analogy to Wheeler et al.33 This reaction occurs preferentially at the SiOH groups at the edges of the flat clay nanoparticles. The clay particles were dried over night at 80 °C in vacuum and transferred to a 4 ACS Paragon Plus Environment
Page 5 of 13
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60
Langmuir
round bottom flask heated to 100°C in a drying oven prior to addition to remove traces of water. 8 ml of toluene and 0.93 ml (1.47 equivalents) of APES were added under inert gas atmosphere, and the mixture stirred and refluxed for 10 h. The cooled mixture was filtered through a Büchner-filter and washed with toluene. Remaining silane was removed by several times washing with toluene and centrifugation of the clay particles. As a final step, the clay particles were dried under vacuum. For the labeling step, 5 mg of perylenediimide N-hydroxysuccinimide (NHS) ester 1 (structure see Fig.2) were dissolved in 60 ml of dichloromethane, and 100 mg of the APESmodified clay particles were added. The mixture was carefully mixed on a biological shaker for 3 days at room temperature. The pinkish particles were filtered off through a Büchner funnel and washed with dichloromethane. The remaining free dye 1 was removed using a soxhlet extraction with dichloromethane for 12 h, and the particles were dried under vacuum. Using DLS, it was verified that the size distribution of the particles before and after fluorescence labeling was very similar. The labeled particles showed mainly a one- or twostep bleaching behavior indicating a very low labeling density with one or few dye molecules per particle. Due to the low labeling density the properties of the nanoparticles are not significantly affected by the fluorophore, even though the isooctylphenoxy substituted perylenediimide derivative is rather hydrophobic. Hydrophilic derivatives of perylenediimide also exist,34 but have not been tested within the framework of our studies.
Preparation of polymer/clay-mixtures and microscopy measurements Before the tracking experiments, a stem solution of clay was freshly prepared from a mixture of labeled and unlabeled Laponite clay nanoparticles (ratio ca. 1:8600). This concentration resulted in a density of labeled particles suitable for WFM experiments. For preparation of the hydrogel, 0.175 mL of clay stem solution (120 g/L) were pipetted into a glass vial and 0.0905 mL of deionised water (MilliQ) was added. Then, 0.485 mL of the polymer stem solution (65 g/L) was added dropwise while stirring. Thus, the final concentration was 28 mg/mL for the clay particles and 42 mg/mL for the polymer (weight-ratio 1.0/1.5). While adding the polymer stem solution, the mixture got more viscous and became turbid. The mixture was stirred for two more hours. After standing for 1.5 days, the hydrogel was sufficiently clear for WFM measurements. The hydrogel was filled into a microfluidic chamber (ibidi, µ slide I Luer) for microscopy, and the ends were sealed with Parafilm. Prior to measurements, the viscous hydrogel was allowed to equilibrate in the microfluidic chamber in order to minimize drifts. The temperature of the sample between 22 °C and 38 °C was adjusted with a heating stage (Pecon Erbach). The objective was also heated with the same heating stage using an appropriate adaptor. The temperature of slide and sample was monitored with an IR thermometer. The optical WFM and the FCS setup are described elsewhere.35
5 ACS Paragon Plus Environment
Langmuir
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60
Page 6 of 13
Single molecule Tracking Single molecule tracking was performed according to a procedure described previously:22 (1) The positions of molecules were determined with a custom-made recognition algorithm which determined the center of the fluorescence signal using a 2D Gaussian fit. (2) The positions were connected with an automatic algorithm using appropriate cost functions to minimize the spatiotemporal distance between points. All trajectories were checked afterwards, short and bad trajectories, i.e. trajectories with less than 20 positionings or more than 20% of missing positioning during the observation time due to blinking, respectively, were removed and obvious errors, i.e. false positives due to positioning of background noise and molecules with wrong positions close to the edge of the CCD-Chip, corrected manually.
Results Characterization of the labeled clay nanoparticles using FCS In order to check the fluorescence labeling of the clay nanoparticles, they were dispersed in water, larger aggregates filtered off, and diffusion of the clay nanoparticles measured by fluorescence correlation spectroscopy (FCS).36 The autocorrelation function shown in Fig. 4 (below) was fitted with two diffusion times according to
(1) with the number N of diffusing species, their amplitudes f1 and f2, and the ratio of the axial and lateral direction of the ellipsoid confocal volume, thus obtaining diffusion times D of 3.2 ms and 0.0083 ms, respectively. The former corresponds to a diffusion coefficient of 5.8 × 10-12 m2 s-1, which was determined using an aqueous solution of rhodamine 6G with a diffusion coefficient of D = 4.14 × 10-10 m2 s-1 as a reference37 yielding a diffusion time of 0.0445 ms with our setup. From the diffusion coefficient, the hydrodynamic radius R can be obtained using the Stokes-Einstein equation
(2) with the Boltzmann constant kB, the temperature T which was 293 K, and the viscosity of 0.001 Pas of water. The resulting value of 37 nm corresponds well with the lateral size of the clay nanoparticles. No drop in correlation corresponding to free dye was found. The correlation decay at ca. 8 s cannot be connected to a translational diffusion process, but rather to a rotation of the clay nanoparticles or photokinetics of the fluorophores.19
6 ACS Paragon Plus Environment
Page 7 of 13
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60
Langmuir
Fig. 4 Autocorrelation curve of a dispersion of labeled Laponite clay nanoparticles in water, its fit according to equation (1) and corresponding residuals.
Single particle diffusion measurements The motion of single fluorescence labeled clay nanoparticles in the nanocomposite was followed using WFM. The time evolution of their mean square displacements for different temperatures between 25 and 38 °C is presented in Fig. 5. All curves clearly deviate from the linear behavior expected for normal diffusion. Therefore, we used the following equation describing 2D anomalous diffusion to fit the data: (3) Herein, is the mean square displacement at a time interval , K is the generalized diffusion coefficient and the anomaly parameter, which is 1 for normal diffusion and below 1 for anomalous sub-diffusion. The application of a 2D diffusion model for our studies is reasonable since we could only track the lateral motion of the nanoparticles which, in order to avoid surface effects, was measured in a distance of ca. 10 m from the coverslip surface in an approximately 400 m thick gel where no significant difference in the motion in axial and lateral diffusion is expected. The influence of localization accuracy on the determination of these parameters has been discussed by Kepten et al.23 Our typical localization accuracy of ca. 10 nm, however, is sufficiently high for their unbiased determination. The curves between 25 and 29 °C are very similar, before they become steeper and less curved around the VPTT. At higher temperatures the mean square distribution (MSD) curves become more and more shallow and curved, the latter indicating stronger anomaly. The values of the fitting parameters, K and , obtained for different temperatures are presented in Fig. 6. The diffusion coefficient K of ca. 3.7 × 10-15 m2 s- obtained at r.t. gradually increases with raising temperature to ca. 4.7 × 10-15 m2 s- at 31 °C. At temperatures above this VPTT, K drops significantly to ca. 0.4 × 10-15 m2 s- at 38 °C. The anomaly parameter shows a similar 7 ACS Paragon Plus Environment
Langmuir
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60
behavior increasing from 0.60 to 0.65 between 25 and 31 °C, followed by a rather strong decrease to 0.41 at 38 °C.
Fig. 5 Dependency of the mean square displacements of the clay particles on the lag times between particle positionings for temperatures between 25 °C and 38 °C. For a clear representation, the curves of some intermediate temperatures are skipped. The mean square displacements of these temperatures and all fits according to the anomalous diffusion model of Equation 3 can be found in the Supporting Information. Each point is an average over at least 130 trajectories measured at different positions.
Fig. 6 Temperature-dependence of the parameters Kα (blue filled circles, left axis) und α (open red squares, right axis) between 25 and 38 °C. The error bars originate from the fits of the mean square displacements as presented in the Supporting Information. The dashed line corresponds to the relative change in diffusion coefficient expected from the temperature dependent decrease in water viscosity as calculated by the Stokes-Einstein equation.
8 ACS Paragon Plus Environment
Page 8 of 13
Page 9 of 13
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60
Langmuir
Discussion We observed the diffusion of dye-labeled clay nanoparticles in PNIPAM solutions and found that the evolution of their mean-square displacements (MSD) can be fitted using the anomalous subdiffusion model presented in equation 3. In the following, we will discuss how this model can be related to the structure and dynamics of clay nanoparticles in hydrogels. Anomalous subdiffusion can be observed in systems with molecular crowding,38 in particular in combination with elastic elements,39 in nanochannels,40 and in network structures.41 In Factin networks for example, Wong et al. could tune the anomaly parameter between 0 < 1 by changing the size ratio between diffusing particles and the mesh sizes of the network.41 The subdiffusive behavior was found to be caused by the particles to be temporarily trapped in the actin filaments with infrequent jumps between the filaments. In the hydrogel studied here, the clay nanoparticles serve as crosslinkers for the linear PNIPAM chains, thus forming the crosslinks of the polymer/clay-network structure. This introduces elasticity in the system which results in anomalous sub-diffusion with < 1,42 in contrast to a purely viscous material where normal diffusion with a direct proportionality between mean square displacement and time would be observed. Since the time-averaged MSDs shown in Fig. 5 exhibit significant anomaly and trajectories do not exhibit power-law distributed waiting times, the anomalous sub-diffusion observed in our studies can be described by a fractional Brownian motion model.27 The structure of the clay-PNIPAM nanocomposites depends strongly on the concentration of the clay nanoparticles and the polymer, and, due to the thermoresponsive behavior of the PNIPAM chains, also on temperature. The polydispersity (1.95) of the polymer is not expected to have a significant influence on the motion of the clay nanoparticles. At temperatures below the VPTT, we observe heterogeneous motion of the clay nanoparticles (see Supporting Movies). Approximately half of the nanoparticles move with a diffusion coefficient suitable for single particle tracking, the other particles are too fast, i.e. their diffusion coefficients are larger than 10-12 m2 s-1, which causes their trajectories to smear over the screen. It is reasonable to assume that the former are trapped in the hydrogel, acting as crosslinkers between different PNIPAM chains, whereas the fast particles move around freely without significant hindrance by the gel. For approximately 2% of the nanoparticles, we could observe that their mobility changes significantly, i.e. free particles get trapped in the network or trapped particles get released. Single particle tracking is only possible for the slower fraction. Our diffusion analysis is therefore restricted to these slow, crosslinking particles. At temperatures below the VPTT, the PNIPAM chains between the particles are swollen with water and therefore exhibit a rather low density. Therefore the network is still rather loose and the clay particles can move significantly. However, already at this point, diffusion is not totally free but viscoelastic behavior is observed (< 1). Heating towards the VPTT results in a slight increase in the mobility of these particles which is however significantly more pronounced than expected by the decrease in solvent viscosity (see Fig. 6). Additionally, the diffusion of the crosslinking particles becomes more normal, i.e. their viscoelasticity decreases. The reason for this behavior could be a slight shrinkage of Laponite-PNIPAM9 ACS Paragon Plus Environment
Langmuir
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60
Page 10 of 13
aggregates when approaching the VPTT as previously shown by DLS.18 Above the VPTT, previous NMR investigations showed that, prior to the deswelling process, the net chains in the swollen state tend to collapse rapidly forming a macro-network.43 Thus, bundles of aggregated and stretched polymer chains appear resulting in a reduced mobility of the net chains and also significantly increasing the elastic modulus of the gel.44 Our measurements confirm these NMR results. At the VPTT, formation of the macro-network causes the mobility of the cross-linking clay nanoparticles to decrease drastically, and the decreasing anomaly parameter indicates the increase in elastic modulus. Within a few degrees centigrade above the VPTT, the mobility of the particles drops significantly and at ca. 36 °C their motion is basically restricted to wobbling around the center position which results in a low anomaly parameter .
Conclusion In summary, we covalently labeled laponite clay nanoparticles with a suitable perylene diimide derivative and investigated the translational motion of these nanoparticles in thermoresponsive PNIPAM hydrogels using FCS and WFM with single particle tracking. We found anomalous diffusion of the clay nanoparticles reflecting viscoelastic behavior due to their function as crosslinkers of PNIPAM chains. The mobility changes significantly around the VPTT of the PNIPAM hydrogel. At temperatures clearly below their VPTT, the show motion with a generalized diffusion coefficient K of ca. 4 × 10-15 m2 s-, and an anomaly parameter of ca. 0.6. These values both increase with increasing temperature and, above VPTT, drop significantly showing the collapse of the hydrogel and the concomitant hindrance of motion within the collapsed hydrogel and a higher viscoelasticity. The present approach to visualize the motion of nanoobjects in hydrogels will, in future work, allow for a better understanding of the dynamics within these systems and allows for a better fine-tuning of their properties.
Acknowledgements D.W. thanks the Center for Applied Photonics (CAP) and the Zukunftskolleg of the University of Konstanz for financial and administrative support. We thank Dr. Maren Dill and Jonathan Liebmann for synthesizing the PDI-dye used for labeling.
10 ACS Paragon Plus Environment
Page 11 of 13
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60
Langmuir
Literature 1. 2. 3. 4. 5. 6. 7. 8. 9.
10. 11. 12. 13. 14. 15. 16. 17.
Gerlach, G.; Arndt, K. F. Hydrogel Sensors and Actuators - Engineering and Technology. Spinger: Heidelberg, 2010. Saeed, A.; Georget, D. M. R.; Mayes, A. G. Synthesis, characterisation and solution thermal behaviour of a family of poly (N-isopropyl acrylamide-co-N-hydroxymethyl acrylamide) copolymers. React. Funct. Polym. 2010, 70 (4), 230-237. Smith, A. E.; Xu, X.; McCormick, C. L. Stimuli-responsive amphiphilic (co)polymers via RAFT polymerization. Prog. Polym. Sci. 2010, 35 (1–2), 45-93. Schild, H. G. Poly(N-isopropylacrylamide): experiment, theory and application. Prog. Polym. Sci. 1992, 17 (2), 163-249. Garret-Flaudy, F.; Freitag, R. Unusual Thermoprecipitation Behavior of Poly(N,Ndiethylacrylamide) from Aqueous Solution in the Presence of Anionic Surfactants. Langmuir 2001, 17 (16), 4711-4716. Freitag, R.; Garret-Flaudy, F. Salt Effects on the Thermoprecipitation of Poly-(Nisopropylacrylamide) Oligomers from Aqueous Solution. Langmuir 2002, 18 (9), 34343440. Zhang, Y.; Furyk, S.; Bergbreiter, D. E.; Cremer, P. S. Specific Ion Effects on the Water Solubility of Macromolecules: PNIPAM and the Hofmeister Series. J. Am. Chem. Soc. 2005, 127 (41), 14505-14510. Yoshioka, H.; Mikami, M.; Mori, Y.; Tsuchida, E. Preparation of Poly(NMsopropylacrylamide)-B-Poly(Ethylene Glycol) and Calorimetric Analysis of Its Aqueous Solution. J. Macromol. Sci., Pure Appl. Chem. 1994, 31 (1), 109-112. Picos-Corrales, L. A.; Licea-Claverie, A.; Cornejo-Bravo, J. M.; Schwarz, S.; Arndt, K.F. Well-defined N-Isopropylacrylamide Dual-Sensitive Copolymers with LCST similar to 38 degrees C in Different Architectures: Linear, Block and Star Polymers. Macromol. Chem. Phys. 2012, 213 (3), 301-314. Haraguchi, K.; Takehisa, T. Nanocomposite Hydrogels: A Unique Organic–Inorganic Network Structure with Extraordinary Mechanical, Optical, and Swelling/De-swelling Properties. Adv. Mater. 2002, 14 (16), 1120-1124. Ferse, B.; Graf, M.; Krahl, F.; Arndt, K.-F. Photopolymerization as Alternative Concept for Synthesis of Poly(N-isopropylacrylamide)-Clay Nanocomposite Hydrogels. Macromol. Symp. 2011, 306-307 (1), 59-66. Nie, J. J.; Du, B. Y.; Oppermann, W. Swelling, elasticity, and spatial inhomogeneity of poly(N-isopropylacrylamide)/clay nanocomposite hydrogels. Macromolecules 2005, 38 (13), 5729-5736. Haraguchi, K.; Farnworth, R.; Ohbayashi, A.; Takehisa, T. Compositional Effects on Mechanical Properties of Nanocomposite Hydrogels Composed of Poly(N,Ndimethylacrylamide) and Clay. Macromolecules 2003, 36 (15), 5732-5741. Haraguchi, K.; Li, H. J. Mechanical properties and structure of polymer-clay nanocomposite gels with high clay content. Macromolecules 2006, 39 (5), 1898-1905. Zebrowski, J.; Prasad, V.; Zhang, W.; Walker, L. M.; Weitz, D. A. Shake-gels: shearinduced gelation of laponite-PEO mixtures. Colloid Surf. A-Physicochem. Eng. Asp. 2003, 213 (2-3), 189-197. Can, V.; Okay, O. Shake gels based on Laponite-PEO mixtures: effect of polymer molecular weight. Des. Monomers Polym. 2005, 8 (5), 453-462. Haraguchi, K.; Takehisa, T.; Fan, S. Effects of Clay Content on the Properties of Nanocomposite Hydrogels Composed of Poly(N-isopropylacrylamide) and Clay. Macromolecules 2002, 35 (27), 10162-10171.
11 ACS Paragon Plus Environment
Langmuir
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60
Page 12 of 13
18. Ferse, B.; Richter, S.; Eckert, F.; Kulkarni, A.; Papadakis, C. M.; Arndt, K. F. Gelation Mechanism of Poly(N-isopropylacrylamide)-Clay Nanocomposite Hydrogels Synthesized by Photopolymerization. Langmuir 2008, 24 (21), 12627-12635. 19. Wöll, D. Fluorescence Correlation Spectroscopy in Polymer Science. RSC Advances 2014, 4, 2447-2465. 20. Koynov, K.; Butt, H.-J. Fluorescence correlation spectroscopy in colloid and interface science. Curr. Opin. Colloid Interface Sci. 2012, 17 (6), 377-387. 21. Shuang, B.; Chen, J.; Kisley, L.; Landes, C. F. Troika of single particle tracking programing: SNR enhancement, particle identification, and mapping. Phys. Chem. Chem. Phys. 2014, 16 (2), 624-634. 22. Wöll, D.; Kölbl, C.; Stempfle, B.; Karrenbauer, A. Novel method for automatic single molecule tracking of blinking molecules at low intensities. Phys. Chem. Chem. Phys. 2013, 15, 6196-6205. 23. Kepten, E.; Bronshtein, I.; Garini, Y. Improved estimation of anomalous diffusion exponents in single-particle tracking experiments. Phys. Rev. E 2013, 87 (5), 052713. 24. Konopka, M. C.; Weisshaar, J. C. Heterogeneous motion of secretory vesicles in the actin cortex of live cells: 3D tracking to 5-nm accuracy. J. Phys. Chem. A 2004, 108 (45), 9814-9826. 25. Magdziarz, M.; Weron, A.; Burnecki, K.; Klafter, J. Fractional Brownian Motion Versus the Continuous-Time Random Walk: A Simple Test for Subdiffusive Dynamics. Phys. Rev. Lett. 2009, 103 (18), 180602. 26. Ernst, D.; Hellmann, M.; Köhler, J.; Weiss, M. Fractional Brownian motion in crowded fluids. Soft Matter 2012, 8 (18), 4886-4889. 27. Ernst, D.; Köhler, J.; Weiss, M. Probing the type of anomalous diffusion with singleparticle tracking. Phys. Chem. Chem. Phys. 2014, 16, 7686-7691. 28. Felix, H.; Thomas, F. Anomalous transport in the crowded world of biological cells. Rep. Prog. Phys. 2013, 76 (4), 046602. 29. Saunders, J. M.; Goodwin, J. W.; Richardson, R. M.; Vincent, B. A Small-Angle X-ray Scattering Study of the Structure of Aqueous Laponite Dispersions. J. Phys. Chem. B 1999, 103 (43), 9211-9218. 30. Bruce, D. W.; O'Hare, D.; Walton, R. I. Inorganic Materials. Wiley: 2011. 31. Flier, B. M. I.; Baier, M. C.; Huber, J.; Müllen, K.; Mecking, S.; Zumbusch, A.; Wöll, D. Heterogeneous Diffusion in Thin Polymer Films As Observed by High-Temperature Single-Molecule Fluorescence Microscopy. J. Am. Chem. Soc. 2012, 134 (1), 480-488. 32. Weil, T.; Vosch, T.; Hofkens, J.; Peneva, K.; Müllen, K. The Rylene Colorant FamilyTailored Nanoemitters for Photonics Research and Applications. Angew. Chem. Int. Ed. 2010, 49 (48), 9068-9093. 33. Wheeler, P. A.; Wang, J.; Baker, J.; Mathias, L. J. Synthesis and Characterization of Covalently Functionalized Laponite Clay. Chem. Mater. 2005, 17 (11), 3012-3018. 34. Peneva, K.; Mihov, G.; Nolde, F.; Rocha, S.; Hotta, J.-i.; Braeckmans, K.; Hofkens, J.; Uji-i, H.; Herrmann, A.; Müllen, K. Water-Soluble Monofunctional Perylene and Terrylene Dyes: Powerful Labels for Single-Enzyme Tracking. Angew. Chem. Int. Ed. 2008, 47 (18), 3372-3375. 35. Stempfle, B.; Dill, M.; Winterhalder, M.; Müllen, K.; Wöll, D. Single Molecule Diffusion and its Heterogeneity during the Bulk Radical Polymerization of Styrene and Methyl Methacrylate. Polymer Chem. 2012, 3, 2456-2463. 36. Papadakis, C. M.; Košovan, P.; Richtering, W.; Wöll, D. Polymers in focus: fluorescence correlation spectroscopy. Colloid Polym. Sci. 2014, 292, 2399-2411. 37. Müller, C. B.; Loman, A.; Pacheco, V.; Koberling, F.; Willbold, D.; Richtering, W.; Enderlein, J. Precise measurement of diffusion by multi-color dual-focus fluorescence correlation spectroscopy. EPL 2008, 83 (4), 46001. 12 ACS Paragon Plus Environment
Page 13 of 13
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60
Langmuir
38. Weiss, M.; Elsner, M.; Kartberg, F.; Nilsson, T. Anomalous Subdiffusion Is a Measure for Cytoplasmic Crowding in Living Cells. Biophys. J. 2004, 87 (5), 3518-3524. 39. Weber, S. C.; Spakowitz, A. J.; Theriot, J. A. Bacterial Chromosomal Loci Move Subdiffusively through a Viscoelastic Cytoplasm. Phys. Rev. Lett. 2010, 104 (23), 238102. 40. De Santo, I.; Causa, F.; Netti, P. A. Subdiffusive Molecular Motion in Nanochannels Observed by Fluorescence Correlation Spectroscopy. Anal. Chem. 2010, 82 (3), 9971005. 41. Wong, I. Y.; Gardel, M. L.; Reichman, D. R.; Weeks, E. R.; Valentine, M. T.; Bausch, A. R.; Weitz, D. A. Anomalous diffusion probes microstructure dynamics of entangled Factin networks. Phys. Rev. Lett. 2004, 92 (17), 178101-1-178101-4. 42. Waigh, T. A. Microrheology of complex fluids. Rep. Prog. Phys. 2005, 68 (3), 685-742. 43. Arndt, K.-F.; Knörgen, M.; Richter, S.; Schmidt, T. NMR Imaging: Monitoring of Swelling of Environmental Sensitive Hydrogels. In Modern Magnetic Resonance, Webb, G., Ed.; Springer Netherlands, 2006, pp 187-193. 44. Knörgen, M.; Arndt, K.-F.; Richter, S.; Kuckling, D.; Schneider, H. Investigation of swelling and diffusion in polymers by 1H NMR imaging: LCP networks and hydrogels. J. Mol. Struct. 2000, 554, 69-79.
Graphical abstract:
13 ACS Paragon Plus Environment
