Size-Selective Permeation of Water-Soluble Polymers through the

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Size-Selective Permeation of Water-Soluble Polymers through the Bilayer Membrane of Cyclodextrin Vesicles Investigated by PFG-NMR Sabine Himmelein,† Nora Sporenberg,‡ Monika Schönhoff,*,‡ and Bart Jan Ravoo*,† †

Organic Chemistry Institute and Graduate School of Chemistry, Westfälische Wilhelms-Universität Münster, Corrensstraße 40, 48149 Münster, Germany ‡ Institute of Physical Chemistry, Westfälische Wilhelms-Universität Münster, Corrensstraße 28/30, 48149 Münster, Germany S Supporting Information *

ABSTRACT: Cyclodextrin vesicles (CDVs) consist of a bilayer of amphiphilic cyclodextrins (CDs). CDVs exhibit CD cavities at their surface that are able to recognize and bind hydrophobic guest molecules via size-selective inclusion. In this study, the permeability of α- and β-CDVs is investigated by pulsed field gradient-stimulated echo (PFG-STE) nuclear magnetic resonance. Diffusion experiments with water and two types of water-soluble polymers, polyethylene glycol (PEG) and polypropylene glycol (PPG), revealed three main factors that influence the exchange rate and permeability of CDVs. First, the length of the hydrophobic chain of the CD amphiphile plays a crucial role. Reasonably, vesicles consisting of amphiphiles with a longer aliphatic chain are less permeable since both membrane thickness and melting temperature Tm increase. Second, the exchange rate through the bilayer membrane depends on the molecular weight of the polymer and decreases with increasing weight of the polymer. Most interestingly, a size-selective distinction of permeation due to the embedded CDs in the bilayer membrane was found. The mechanism of permeation is shown to occur through the CD cavity, such that depending on the size of the cavity, permeation of polymers with different cross-sectional diameters takes place. Whereas PPG permeates through the membrane of β-CD vesicles, it does not permeate α-CD vesicles.

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INTRODUCTION A variety of amphiphilic molecules, either natural phospholipids or synthetic amphiphiles, self-assemble in water to form vesicles.1−8 Since vesicles create a compartment composed of a bilayer of amphiphiles enclosing an aqueous interior, they can be considered as the simplest artificial cell model. Thus, the membrane surface is able to function as a scaffold for mimicking essential biological processes such as signal transduction, molecular recognition, as well as adhesion and fusion of cells.9−18 Furthermore, bilayer vesicles possess the capability of encapsulating chemical agents or drugs and can be used as nanoreactors or effective drug delivery systems.19−24 In this respect, the permeability of the vesicle membrane is of great importance. Controlled release or the transfer of a signal through the vesicle barrier requires a detailed understanding of the transport resistance of the bilayer. Cyclodextrin vesicles (CDVs) composed of amphiphilic cyclodextrins (CDs) have been extensively studied over the © 2014 American Chemical Society

past few years. The structure of CDVs is illustrated in Scheme 1. Amphiphilic CDs can be synthesized from native α-, β-, and γ-CDs by introducing one hydrophobic alkyl chain per glucose at the primary side as well as one hydrophilic oligo(ethylene glycol) group per glucose on the secondary side of the macrocycle.25 Just like liposomes, these vesicles are able to encapsulate hydrophilic molecules in their aqueous interior or hydrophobic molecules or particles in their bilayer membrane. Additionally, the surface of the vesicles exhibits multiple CD hosts that are available for specific host−guest inclusion complexation with corresponding guest molecules.26 Previous work revealed that via guest moieties, various active molecules such as carbohydrates,27 peptides,28 zwitterions,29 DNA binding moieties,30 and polymers31 can be attached to the Received: January 17, 2014 Revised: March 20, 2014 Published: March 20, 2014 3988

dx.doi.org/10.1021/la500226z | Langmuir 2014, 30, 3988−3995

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Scheme 1. Polymer Size-Selective Diffusion and Molecular Structure of Amphipilic CDsa

a

(a) Schematic representation of the size-selective diffusion of polymers with a different cross-sectional area through the membrane of CDVs. The vesicles consist of a bilayer of amphiphilic CDs. (b) Molecular structure of the amphiphilic CDs used in this study.

labeled dextran could be encapsulated into CD vesicles formed from amphiphiles with both n-dodecyl and n-hexadecyl chains. Whereas CDVs consisting of amphiphiles with the longer nhexadecyl chains are able to retain the dye molecules quite substantially as more than 75% of CF remained inside the vesicles after 3 days, the content leakage from CDVs composed of 1b chains is fast (complete leakage within hours) and the encapsulation less efficient.25,26 Recently, it could be shown that the mixing of 1b with natural lipids results in the formation of vesicles that are much less leaky (comparable to vesicles from 2) and still possess the ability to bind guest molecules into the CD cavities on their surface.45 Altogether, these experiments showed no dependence of the vesicle permeability on the size of the CD cavity but only on the thickness of the membrane. In order to obtain a more detailed picture of the transport processes through the bilayer membrane of CDVs PFG-NMR diffusion experiments were performed. Besides water, the hydrophilic polymers polyethylene glycol (PEG) and polypropylene glycol (PPG) were used as tracer molecules. PEG is known to form size-selective inclusion complexes with α-CD and PPG with β-CD.46 First of all, the permeability of CDVs composed of 1b and 2 was compared to verify the results of the dye encapsulation experiments. In a second set of experiments, the exchange rate through the bilayer membrane with regard to the molecular weight of the polymer was studied. Furthermore, PEG and PPG were used to identify if a size-selective distinction of permeation due to the embedded CDs in the bilayer membrane can be observed.

surface of the CDVs. Due to their specific functionality, these can act further as a recognition group to bind and release DNA and proteins,27,30 to change the shape of the vesicles into rodlike structures,28 or to mediate the adhesion and aggregation of the vesicles.29 For applications as an encapsulation system, the permeability of the bilayer is a crucial parameter. An interesting question is thus whether the CD cavities play any role in the permeation mechanism and whether permeation takes place through the cavity or if it is limited only by the hydrophobic layer. In order to investigate the permeation of a water-soluble molecule from the outside of a vesicle to the inside and vice versa, pulsed-field gradient nuclear magnetic resonance (PFGNMR) can be applied.32−34 In a specialized approach termed diffusion-exchange NMR, echo decays yielding information about the molecular mean square displacement are determined under variation of the observation time. In this way, the experiment can not only distinguish molecules in a site with fast diffusion versus a site with slow diffusion, but it can furthermore quantify the equilibrium exchange dynamics between these two sites. For this purpose, a two-site exchange model developed by Kärger35 for exchange in diffusion experiments has to be applied to the diffusion data sets, yielding permeation rates through vesicle walls or, more generally, exchange rates between the sites. This technique has been applied to study the permeation of water-soluble polymers or small molecules through the walls of hollow colloidal particles in various systems, such as block-co-polymer vesicles,36−38 polyelectrolyte multilayer capsules,39−41 or lipid vesicles42,43 (see also a recent review article).44 With PFGNMR, it is possible to determine the hydrodynamic vesicle radius, the equilibrium distribution of encapsulated species, and the permeability of the vesicle wall without labeling the observed molecules. Instead, the hindered diffusion of a molecule in a spherical compartment is detected by a decrease in their lateral diffusivity determined by the mean-square displacements in a given observation time (Δ). This makes PFG-NMR perfectly suitable for the investigation of the permeability of vesicle membranes without perturbing the system. In this study, we examined the permeability of CDVs formed from amphiphilic α- and β-CDs with n-dodecyl chains (1a: αC12−CD and 1b: β-C12−CD), as well as β-CD with n-hexadecyl chains (2: β-C16−CD) toward water and water-soluble polymers with PFG-NMR. So far, the permeability of CDVs has only been investigated with dye encapsulation experiments. Carboxyfluorescein (CF), sulforhodamine, and fluorescein-

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MATERIALS AND METHODS Materials. Polyethylene glycol, PEG (Mw = 200, 1020, 10000, and 25800 g/mol) and polypropylene glycol, PPG (Mw = 1000 g/mol) were purchased from PSS Polymer Standards Service. Deuterium oxide (99.9% isotopic purity) was purchased from Aldrich. Amphiphilic α- and β-CDs were synthesized, as described previously.26,47 Their chemical structure is shown in Scheme 1b. Preparation of Vesicles. Unilamellar vesicles of amphiphiles 1a, 1b, or 2, respectively (see Scheme 1b), with an average diameter of 100 nm were prepared by extrusion. Amphiphilic CD was dissolved in chloroform, dried while rotating to obtain a thin film in the flask, and left under high vacuum for 1 h. Deuterium oxide was added, and the mixture was sonicated for 1 min using a Sonorex bath sonicator (Branson) in order to suspend the amphiphiles in solution. The 3989


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in a MATLAB (The Mathworks, Inc.) program. Details about the procedure and the relevant equations have been described in previous publications.39,43,48 Here, a global set of fixed parameters consisting of the corresponding spin relaxation times (T1e,i, T2e,i) is employed, where the values for internal and external chains are identical, as shown for PEG chains in polymer capsules previously.39 Effective relaxation times Te,i of encapsulated and free chains are thus assumed to be identical. A set of free parameters (τe,i. De,i, Pe,i) is optimized. From the resulting residence times, the exchange time is calculated as

dispersion was stirred overnight and afterward extruded with a LiposoFast extruder (Avestin) through a polycarbonate membrane with a 0.1 μm pore size. If necessary, the dispersion was heated to 50 °C during the extrusion process. 1−2 wt % of the respective polymer standard were added either before or after extrusion. Dynamic Light Scattering. Dynamic light scattering analysis of the CDVs was performed after extrusion to ensure that the average size of the vesicles is 100 nm. Measurements were conducted on a Zetasizer Nano ZS device (Malvern Instruments, Ltd.). NMR Measurements. 1H diffusion experiments were performed on a Bruker 400 MHz Avance NMR spectrometer. The measurements were done with a liquid state probe head (DIFF 30, Bruker) providing pulsed field gradients of up to 1180 G/cm. The gradient coils were cooled by a water circulation unit (Haake), which is also used to control the sample temperature, T = 22 °C. Examples of 1H spectra are shown in Figure S1 of the Supporting Information. The 1H resonances of the polymer are narrow, resulting from free chains, while no broadened signal of the ethyleneoxide chains of the vesicles was observed. The diffusion coefficients were measured with a stimulated echo sequence [π/2-τ-π/2-T-π/2-τecho] in combination with two gradient pulses of duration δ = 2 ms and variable gradient strength (g) applied during each delay (τ).33 The spacing between the two gradient pulses is the observation time (Δ). This time is varied between 25 and 150 ms. Data in the results part are displayed as logarithmic, normalized intensities in dependence of the parameter k, which is k = γ2g2δ2(Δ−δ/3). Data Analysis in Two-Site Model. Diffusion data were quantitatively analyzed, employing a two-site model for diffusion experiments with the equations first derived by Kärger.35 The two sites are given by free chains in the continuous phase (exterior, site e) and encapsulated chains in the vesicle interior (site i). Both sites are described by the respective diffusion coefficients, De,i, (where Di is the diffusion coefficient of the vesicle and De that of the free chain), the fraction of polymer in either site, Pe,i, and the average residence time in the site, τe,i. The effect of exchange is then described by fitting the diffusion echo decays by a biexponential decay with apparent fractions PA,B and apparent decay constants DA,B:35

1 1 1 = + τex τe τi

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RESULTS AND DISCUSSION Permeability of β-CDVs with Different Membrane Thickness. In a first set of experiments the influence of the membrane thickness on the permeation of PEG 1020 and water in β-CDV dispersions was investigated. Here, vesicles were prepared on the one hand from β-CD amphiphiles 1b and on the other hand from amphiphiles 2. The melting temperature for the transition from the liquid-crystalline (Lα) phase to the lamellar gel (Lβ) is lower for β-C12−CDVs (Tm ≈ 0 °C) than for β-C16−CDVs (Tm ≈ 48 °C). Therefore, the membrane of βC12−CDVs is in a fluid state, whereas the membrane of β-C16− CDVs is densely packed in the PFG-NMR measurements performed at ambient temperature. Furthermore the thickness of the bilayer membrane for β-C12−CDVs is dC12 = 41.9 Å and for β-C16−CDVs, dC16 = 45.7 Å.25,26 PEG 1020 in β-C12−CDV and β-C16−CDV Dispersion. The diffusion measurements for PEG 1020 in β-C12−CDV and βC16−CDV dispersion with varying observation time Δ are shown in Figure 1. In both cases, biexponential decay curves are found, where the first region can be attributed to the rapid decay of the signal of free PEG 1020 in the continuous phase and the second region to the slow decay of the signal of encapsulated PEG 1020 in the interior of the CDVs. For βC12−CDVs (see Figure 1a), the permeation of PEG 1020 through the bilayer membrane during the observation time causes a dependence of the echo decays on Δ, as the molecular exchange between the fast and the slow diffusing site is in an intermediate regime as compared to Δ. Employing the two-site model fit, an exchange time of τex ≈ 50 ms is obtained. On the contrary, the echo decays for PEG 1020 in β-C16− CDVs show no dependence on the observation time Δ. Hence, the exchange process is slow compared to Δ (i.e., τex ≫ 100 ms). It can be concluded that the thicker and more rigid membrane of β-C16−CDVs is less permeable toward the polymer than the membrane of β-C12−CDVs. This is also in good agreement with previous results from content release experiments of CDVs.26 Water in β-C12−CDV and β-C16−CDV Dispersion. The water diffusion coefficient was measured employing the 1H signal of the residual protons in D2O, thus it represents the HDO diffusion. The sets of Δ-dependent water echo decays differ clearly between β-C12−CDV and β-C16−CDV dispersions (see Figure 2). For β-C12−CDVs, an exponential echo decay is observed. This decay is furthermore independent of the observation time (data not shown), and no signal for the encapsulated species can be detected. The diffusion coefficient is D = 1.6 × 10−9 m2/s, close to the literature value for HDO in D2O. To prove the absence of any encapsulated water, the same

PB = 1 − PA =

⎧ ⎛ ⎛ D ⎫ 1 Δ⎞ Δ⎞ ⎨Pe⎜De + ⎟ + Pi ⎜Di + ⎟ − A⎬ (DB − DA ) ⎩ ⎝ Tk Tk k ⎭ ⎝ e ⎠ i ⎠

DA,B =

∓

⎪

⎪

⎪

⎪

⎧ Δ⎛ 1 1 1 1⎞ 1⎪ ⎨(De + Di) + ⎜ + + + ⎟ τe τi ⎠ 2⎪ k ⎝ Te Ti ⎩ 2 ⎡ Δ⎛ 1 1 1 1 ⎞⎤ 4Δ2 ⎢(Di − De) + ⎜ − + − ⎟⎥ + ⎢⎣ τi τe ⎠⎥⎦ k ⎝ Ti Te τeτik 2

⎫ ⎪ ⎬ ⎪ ⎭

The effective relaxation times Te,i are given in the case of the stimulated echo by 1 T 2τ = + Te , i T1e , i T2e , i

Sets of echo decays obtained with different observation times were analyzed by a least-squares fitting procedure implemented 3990


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Figure 1. Echo decays of PEG 1020 in (a) CDV composed of 1b and (b) CDV composed of 2 for different observation time Δ. Symbols represent the experimental data points, and solid lines are results obtained by the Kärger model fit.

Figure 2. Echo decays of water in (a) CDV composed of 1b and (b) CDV composed of 2 for different observation times Δ. Symbols represent the experimental data points, and lines are results obtained by either (a) an exponential fit or (b) a Kärger model fit, respectively.

data are shown over a wider range of k in Figure S2 of the Supporting Information. Since no encapsulated water is detected, water is in fast exchange across the CDV membrane, such that the measured diffusion coefficient is a weighted average of the interior and exterior water diffusion. This implies a fast permeation (τex ≪ 25 ms) of water molecules through the vesicle membrane. In contrast, for water in β-C16−CDV dispersion, biexponential decay curves are found, which depend on the observation time Δ (Figure 2b). The Kärger model fit yields an exchange time of τex ≈ 200 ms. These results show that even for water, the β-C16−CDVs are significantly less permeable than β-C12−CDVs. In comparison, for liposomes made of phospholipids, it is well-known that water permeates their membrane with a permeation rate that is higher for liquid crystalline lipids than for lipids in the gel state.49−54 Permeability of β-C12−CDVs for PEGs with Varying Molecular Weights. Diffusion-exchange experiments of further polymers with higher molecular weights (PEG 10000 and PEG 25800) and lower molecular weights (PEG 200 and PPG 192) were performed in β-C12−CDV dispersions. Here, the influence of the different chain length of the probe molecule on the exchange through the bilayer membrane was investigated. The corresponding echo decays are shown in Figure 3 and Figure S3 of the Supporting Information. While for PEG 1020 in β-C12−CDV dispersion (Figure 1a), an intermediate exchange time of τex ≈ 50 ms was found; in contrast, for PEG 200 and PPG 192, an exponential decay is observed. Again, to prove the absence of any encapsulated PEG 200, the data of Figure 3a are shown over a wider range of k in Figure S2 of the Supporting Information. Thus, the observed

single exponential echo decay reveals a fast exchange process (τex ≪ Δ). By contrast for PEG 10000 and PEG 25800, slow exchange can be monitored as the biexponential decay curves are independent of the observation time Δ. On the whole, the exchange time increases for probe molecules with high molecular weights and the permeation through the bilayer membrane of CDVs is retarded. A qualitatively similar molecular weight dependency was also observed for PEG permeation through the walls of block copolymer vesicles55 or the walls of polyelectrolyte capsules.39 Permeability of α- and β-CDVs for PEG and PPG. In a third series of experiments, the diffusion of PEG 1020 and PPG 1000 through the bilayer membranes of α-C12− and β-C12− CDVs were examined by PFG-NMR. It is well-known that αand β-CD form size-selective inclusion complexes with PEG and PPG, respectively. The smaller α-CD forms complexes with PEG, which has a small cross-sectional diameter, whereas β-CD can complex PPG, which has methyl as the side groups, resulting in a larger cross-sectional diameter (cf. Scheme 2).46 With this experimental assay, a selectivity of permeation as function of size of the CD cavity with regard to the crosssectional area of the polymer is investigated. PEG 1020 in α-C12− and β-C12−CDV Dispersions. First, the exchange of PEG 1020 through α- and β-CD vesicles was studied, again by performing diffusion experiments with varying observation time Δ. The resulting echo decays are shown in Figure 1a and Figure 4, respectively. In both cases, biexponential decay curves are found. The slope of the decay curve in the first region yields a diffusion coefficient of D1 = 1.7 × 10−10 m2 s−1 for both the α- as well as the β-CDV dispersion. 3991


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Figure 4. Echo decays of PEG 1020 in CDV composed of 1a for different observation time Δ. Symbols represent the experimental data points and the solid line is the result obtained of a biexponential fit for Δ = 25 ms.

10−12 m2 s−1 for the β-CDV dispersion. By means of the Stokes−Einstein equation, a diameter of dα ≈ 80 nm and dβ ≈ 120 nm for α- and β-CDVs is estimated. In the case of the diffusion of PEG 1020 in α-C12−CDV (Figure 4), only for the smallest observation time Δ = 25 ms data points over the whole k-region are obtained. For longer observation times (Δ = 50− 100 ms), the signal intensity of the slow diffusing species is rather low (magnitude on the order of 10−4). Hence, with increasing gradient strength g, the signal is no longer detectable. For PEG 1020 in β-C12−CDV dispersion data points over the whole k-region are obtained for all observation times (Δ), and the two-site model fit yielded an exchange time of τex ≈ 50 ms (Figure 1a). Qualitatively, a similar or somewhat larger exchange time can be estimated for α-C12−CDVs. Thus, the experiments show that PEG 1020 permeates both α- and βCDVs within tens of milliseconds. PPG 1000 in α-C12− and β-C12−CDV Dispersions. As outlined above for PEG 1020, analog experiments were performed for PPG 1000. The resulting echo decay curves for PPG 1000 in α- and β-CDV dispersions are shown in Figure

Figure 3. Echo decays of (a) PEG 200 and (b) PEG 10000 in CDV composed of 1b for different observation times Δ. Symbols represent the experimental data points, and lines are results obtained by the Kärger model fit.

This is in good agreement with the diffusion coefficient for free PEG 1020 in D2O (D = 1.85 × 10−10 m2 s−1). The diffusion coefficient of the encapsulated species is D2,α = 4.8 × 10−12 m2 s−1 for the α-CDV dispersion and D2,β = 3.0 × 10−12 m2 s−1 for the β-CDV dispersion. They agree with the diffusion coefficients obtained by DLS measurements, which are D2,α = 4.7 × 10−12 m2 s−1 for the α-CDV dispersion and D2,β = 3.9 ×

Scheme 2. Scheme of the Dimensions of the α-CD and β-CD Cavities and the Cross-Sectional Areas of PEG and PPG

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s−1 for the β-CDV dispersion. It has to be noted that for this system a phase separation occurred within 3 days. There, the top phase still was fluid, while the bottom phase was found to be a gel. This observation can be explained by the formation of host−guest complexes of PPG with the β-CD cavities on the surface of the vesicles resulting in a cross-linked network of βCDVs that decants to the bottom of the NMR tube. Though the NMR experiment is faster than the gel formation kinetics, it might be influenced by the onset of the gelation, thus the vesicle concentration within the measurement zone decreases with time and, furthermore, the encapsulated volume of the vesicles decreases. Consequently, the encapsulated fraction of PPG is only 0.06%, while for PEG 1020 in β-CDVs, a value of 0.4% was found. In a comparison of Figure 5 (panels a and c), the conclusion is that PPG seems to be unable to cross the bilayer membrane of α-CDVs, whereas for β-CDVs encapsulated PPG can be found. However, since the α-CDV system also might suffer from gelation onset, thus reducing the encapsulated fraction, a control experiment was performed to confirm the impermeability of the α-CDVs for PPG 1000; there, PPG 1000 was added to a dispersion of α-CDVs before extrusion of the vesicles. Thereby a portion of the polymer is enclosed in the interior of α-CDV during the extrusion process and a signal of the encapsulated species should be observable. Figure 5b shows the resulting diffusion measurements with varying observation time (Δ). Now biexponential echo decays are obtained, which show almost no dependence on Δ. The slope of the decay curve in the first region yields D1 = 1.7 × 10−10 m2 s−1, whereas the diffusion coefficient of the encapsulated species is D2,α = 1.5 × 10−12 m2 s−1. The results approve the existence of an encapsulated PPG component; furthermore, there is no exchange through the vesicle membrane. The permeation of PPG through α-CDVs seems to be hindered by its larger crosssectional area due to the methyl side groups. However β-CDVs are permeable by PPG, as β-CDs possess a larger cavity that is able to form inclusion complexes with the polymer. It can be concluded that the permeation of CDVs occurs by the threading of the elongated polymer chains through the CD cavities, similar to the formation of pseudorotaxanes. The surface of the CDVs is covered with CD pores, which allow the size-selective permeation of polymers according to their crosssectional area. This is in good agreement with reports on liposomes, showing that molecules prefer to penetrate along the bilayer normal with their long principal axis. Xiang and Anderson53 demonstrated that the penetration rate through liposomal membranes has a large correlation coefficient (r = 0.94) with the minimum cross-sectional area of a molecule, but a low correlation coefficient (r = 0.59) with the molecular volume. A transbilayer displacement requires an available crosssectional area equal to or larger than the minimum crosssectional area of the diffusing molecule. For CDVs, the CD cavities serve as precast openings for the penetration of molecules. PPG with its larger cross-sectional diameter (d = 5.8 Å) compared to PEG (d = 3.1 Å) fits through the cavity of βCD but is too large for α-CD. Harada et al. previously revealed that PPG with its methyl side groups forms inclusion complexes with β-CD but not with α-CD and PEG is complexed exclusively by α-CD but not by β-CD.46 The results of the diffusion experiments of PEG 1020 and PPG 1000 in α- and βCDV dispersions lead to the conclusion that the exchange of molecules through their bilayer membrane takes place through

5. For the α-CDV dispersion, it is interesting that there is no signal of the encapsulated species. The inset in Figure 5a shows

Figure 5. Echo decays of PPG 1000 in CDV composed of 1a added (a) after and (b) before extrusion and (c) echo decays of PPG 1000 in β-C12−CDV dispersion for different observation times (Δ). Symbols represent the experimental data points, and the solid lines are fit results: (a) exponential fit, (b) biexponential fit for Δ = 25 ms, and (c) Kärger model fit.

an exemplary spectrum in the range of k values between 1 and 5 × 10−11 sm−2. Despite the same number of scans used as in the experiments shown above, no signal is detectable in this region. In contrast, for PPG 1000 in β-CDV dispersion, biexponential echo decay curves are obtained (see Figure 5c). By varying the observation time Δ and applying the Kärger model fit, an exchange time of τex ≈ 70 ms is found. The diffusion coefficient for the free PPG 1000 site is determined as De = 1.6 × 10−10 m2 s−1 and is in fair agreement with the diffusion coefficient for free PPG 1000 in D2O (D = 1.9 × 10−10 m2 s−1). The diffusion coefficient of the encapsulated species is Di,β = 1.2 × 10−12 m2 3993


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the CD cavities. Thus, a size-selectivity of permeation is resulting.

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CONCLUSIONS In the present paper, PFG-NMR was successfully applied for the investigation of exchange processes and the permeability of CD vesicles toward water and the hydrophilic polymers PEG and PPG with varying molecular weights. In cases of intermediate permeation processes [i.e., the tracer molecules cross the vesicle membrane within a time on the order of available observation times (Δ)], exchange rates of the tracer molecules could be obtained by using a two-site exchange model. In other cases, fast exchange (τex ≪ Δ) and slow exchange (τex ≫ Δ) permeation processes could be identified. On the basis of the diffusion experiments, three main factors that influence the permeability of CDVs were identified. On the one hand, CDVs consisting of amphiphiles with n-hexadecyl chains are less permeable toward water and PEG than CD vesicles formed from amphiphiles with n-dodecyl chains. This can be attributed to the fact that both the membrane thickness and the melting temperature Tm increase with the aliphatic chain length. On the other hand, the exchange rate of PEG through the membrane of CD vesicles decreased with increasing molecular weight of the polymer. These two factors are also known for liposomes and polymeric capsules. Furthermore, and most interestingly, the size-selective permeabilities of α-CD and β-CD vesicles by polymers with different cross-sectional areas were identified. PEG is able to permeate through both, α-CD and β-CD vesicles, while PPG is too large to pass through the cavity of α-CD vesicles and only penetrates through the larger cavity of β-CD vesicles. This is in good agreement with literature on complex formation of α- and β-CD with those polymers and moreover indicates that the diffusion of polymer takes place mainly through the CD cavities. Hence, the CD head groups act as pores for the penetration of the linear polymers, forming a permeation model system mimicking the DNA translocation through the defined pores of the cell membrane.

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ASSOCIATED CONTENT

* Supporting Information S

Tables of fast, intermediate, and slow exchange, 1H NMR, and echo decays. 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: schonhoff@uni-muenster.de. *E-mail: [email protected]. Notes

The authors declare no competing financial interest.

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ACKNOWLEDGMENTS This work was partially funded by the German Science Foundation (DFG) within the collaborative research center TR 61.

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REFERENCES

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Size-Selective Permeation of Water-Soluble Polymers through the

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