Journal of Colloid and Interface Science 344 (2010) 438–446

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Journal of Colloid and Interface Science

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Solubilisation of model adjuvants by Pluronic block copolymers

Melissa A. Sharp a,c,*, Clive Washington b, Terence Cosgrove a

a School of Chemistry, University of Bristol, Cantock’s Close, Bristol BS8 1TS, UKb AstraZeneca, Charter Way, Silk Road Business Park, Macclesfield, Cheshire SK10 2NA, UKc ESS Scandinavia, P.O. Box 117, SE-221 00 Lund, Sweden

a r t i c l e i n f o a b s t r a c t

Article history:Received 3 November 2009Accepted 6 January 2010Available online 11 January 2010

Keywords:PluronicP85P105F127MicelleAdjuvantSANSPFG-NMRBenzyl benzoateBenzyl alcohol

0021-9797/$ - see front matter Crown Copyright � 2doi:10.1016/j.jcis.2010.01.005

* Corresponding author. Address: ESS Scandinavia, PSweden.

E-mail address: melissa.sharp@esss.se (M.A. Sharp

The effect of two model adjuvants (benzyl benzoate and benzyl alcohol) on the structure and dynamics ofthree Pluronic triblock copolymers (P85, P105 and F127) was studied using small-angle neutron scatter-ing and pulsed-field gradient NMR. The two adjuvants studied have different aqueous solubilities. It wasfound that both adjuvants promoted the micellisation of the Pluronic block copolymers. In addition theylead to a swelling of the micelles, as shown by small-angle neutron scattering. From the pulsed-field gra-dient NMR results it was possible to determine the amount of adjuvant bound to the micelles.

Crown Copyright � 2010 Published by Elsevier Inc. All rights reserved.

1. Introduction

Pluronic block copolymers are amphiphilic triblock copolymersconsisting of ethylene oxide (EO) and propylene oxide (PO) in theform (EO)n(PO)m(EO)n. The polymers are water soluble and showa rich phase behaviour, which has been widely studied [1–9].The polymers are available in a variety of EO:PO ratios and molec-ular weights, which govern their behaviour in aqueous solution. Byvarying these factors one can therefore optimise the polymers foruse in a wide range of applications, and Pluronics have thus founduse in a number of industrial applications, such as cleaning prod-ucts, detergents, emulsification and fermentation [1,10–12].

It has been found that the phase behaviour of the Pluronics inaqueous solutions is very sensitive to both the polymer concentra-tion and temperature [1,3–5]. Typically the Pluronics will bepresent as unimolecular ‘‘unimers” at low concentrations and tem-peratures. As the polymer concentration and/or temperature is in-creased the polymers begin to form spherical micelles, althoughthere will still be a significant concentration of unimer present.Eventually, if the concentration and temperature are increased fur-ther, a ‘‘micellar liquid” phase will be formed, which consists ofspherical micelles and a small number of unimers, and finally a

010 Published by Elsevier Inc. All r

.O. Box 117, SE-221 00 Lund,

).

gel phase will be reached [4,1,6]. At still higher temperaturesrod-like phases and lamellar phases may be observed [4,6,7,13].

In recent years there has also been an interest in studying theability of polymeric micelles to solubilise organic compounds[1,14,15]. The focus of these studies has mainly been the solubilis-ing capacity of the polymer and the partitioning of the organiccompound between the polymer and the solvent [16–18]. Small-angle scattering techniques have, also been used to characterisethese systems. Lettow et al. [19] have for example used small-angleneutron scattering to study two aromatic compounds added to Plu-ronic P123, while Arleth et al. [20] have studied the reverse micelleformation of Pluronic L64 with xylene. Alexandridis and coworkers[21–23] have used small-angle X-ray scattering to study the liquidcrystalline phases that may be formed by Pluronic–water–xylenemixtures.

The interest in this type of system arises from their potentialuse as drug delivery agents [24–27]. In this context it is importantto gain a better understanding of the structural changes that occurupon the addition of organic compounds, that may be used in drugdelivery systems, to the micellar solutions of the Pluronics. In addi-tion it is important to understand the dynamics in this type of sys-tem. This paper describes structural investigations that werecarried out using small-angle neutron scattering (SANS) as wellas an investigation of the dynamics of the system using pulsed-field gradient nuclear magnetic resonance (PFG-NMR). The effectof adding benzyl benzoate and benzyl alcohol, two model adju-

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M.A. Sharp et al. / Journal of Colloid and Interface Science 344 (2010) 438–446 439

vants, to aqueous solutions of three Pluronic block copolymers(Pluronics P85, P105 and F127) was studied. Combining these tech-niques provides new and unique insight into this type of system.The choice of compounds was governed by their use in the phar-maceutical industry as adjuvants. From a physico-chemical per-spective it was thought to be interesting to compare these twocompounds, due to their different aqeuous solubilities. Comparedto several studies reported in the literature on ternary Pluronic sys-tems [17,28–30], the compounds studied here have such low aqeu-eous solubilities that they cannot be considered ‘‘cosolvents”. Thearticle will discuss the results obtained for Pluronic P85 in detail.This will be followed by a briefer discussion of the results obtainedfor Pluronics P105 and F127, together with a comparison of the re-sults obtained from the different Pluronics.

2. Experimental

2.1. Materials

Pluronic P85, P105 and F127 were kindly supplied by BASF.Fully hydrogenated benzyl benzoate (99.9+%) was obtained fromFisher (Acros Organics), while partly deuterated benzyl benzoate(d5-benzyl benzoate, 98%) was obtained from CDN Isotopes Inc.Fully hydrogenated benzyl alcohol (99+%) was obtained from Al-drich, as was the partly deuterated benzyl alcohol (d5-benzyl alco-hol, 98%). Deuterium oxide (D2O, 99.9%) was obtained from Goss.All materials were used as received.

The samples were prepared by weighing out the polymer, adju-vant and solvent into vials. The vials were placed on a roller mixerfor at least 24 h in order to allow the polymer to dissolve and forthe samples to equilibrate. The sample preparation was carriedout at room temperature.

2.2. Measurements

2.2.1. Small-angle neutron scatteringThe SANS measurements were carried out on the LOQ instrument

at ISIS, Didcot, UK and the SANS-1 instrument at GKSS, Hamburg,Germany. The instrument at ISIS is a fixed-geometry instrument,using a spread of wavelengths between 2.2 and 10 Å. The Q-rangeobtained was 0.009–0.29 Å�1. The instrument at GKSS is a fixed-wavelength instrument. A wavelength of 8 Å was used, combinedwith four sample-detector distances: 0.7 m, 1.8 m, 4.5 m and9.7 m. This gave a Q-range of approximately 0.005–0.25 Å�1.

Before the measurements of the samples began, backgroundmeasurements were carried out. The calibration sample used atISIS was a partially deuterated homopolymer blend. Deuteratedwater was run as a background sample. In addition an empty cellwas run. At GKSS water was used as the calibration sample. Thesolvent mixtures used to prepare the samples were measured asthe background samples.

All samples were measured in pure D2O. The samples where theadjuvants were solubilised have also been measured at two con-trast match points: firstly the solvent was contrast matched tothe adjuvant, leaving only the scattering from the polymer, sec-ondly the solvent was contrast matched to the polymer, leavingonly the scattering from the adjuvant.

The samples prepared in pure D2O were measured in 2 mmpathlength Hellma quartz rectangular cells, while the samples withcontrast matching were measured in 1 mm pathlength cells, due tothe higher hydrogen content.

2.2.2. Pulsed-field gradient NMRThe PFG-NMR measurements were carried out using a Bruker

300 MHz spectrometer, with a 1H diffusion probe. The pulse se-

quence used in the experiments was the stimulated echo sequence[31–33]. This pulse sequence employs three 90� radiofrequencypulses and two magnetic field gradient pulses, of strength g andlength d. The first and second radiofrequency pulses are separatedby time s while the second and third pulse are separated by time T.The two gradient pulses are separated by the diffusion time, D.Depending on the sample, the value of the gradient duration, d,varied between 1 and 3 ms, while the diffusion time, D, was variedbetween 30 and 1000 ms. The maximum gradient strength usedwas 10 T/m.

2.3. Analysis

Pluronic micelles are thought to consist of a spherical core con-taining the hydrophobic part of the copolymer (poly(propyleneoxide)). This core is surrounded by a shell of dissolved polymerchains consisting of the hydrophilic part of the copolymer(poly(ethylene oxide)) [6,7,34–37]. Pedersen et al. [35,38] haveproposed an analytical expression for the form factor of a sphericalmicelle consisting of a dense spherical core and polymer chains at-tached to the surface. The form factor for the micelle is composedof four different terms: the normalised self-correlation of thesphere (Ps(Q, R)), the self-correlation of the chains (Pc(Q, L, b)),the cross term between the sphere and the chains (Pcc(Q)) andthe cross term between the different chains (the poly(ethyleneoxide)):

PmicðQÞ ¼ N2aggq

2s PsðQ ;RcoreÞ þ Naggq2

c PcðQ ; L; bÞþ NaggðNagg � 1Þq2

c PccðQÞ þ 2N2aggqsqcPscðQÞ ð1Þ

where Nagg is the aggregation number of the micelle, qs and qc arethe scattering length densities of the spherical core and the chain,respectively, Rcore is the radius of the core, and L and b are the con-tour length and Kuhn segment length of the chain (PEO).

Further details of these form factors can be found in the papersby Pedersen et al. [35,38].

The model uses a Schultz distribution of the aggregation num-ber to take into account the polydispersity of the micelles, andtakes into account the presence of both unimers and micellesand also introduces a structure factor to account for the intermicel-lar interactions. The structure factor used is based on the hardsphere structure factor derived by Lekner [39]. The overall expres-sion for the scattering cross section is given by:

@R@XðQÞ ¼ UmicPmicðQÞSðQÞ þ ð1�UmicÞPuniðQÞ þ bg ð2Þ

where Umic is the fraction of polymer micellised, Pmic(Q) is the formfactor of the micelle, Puni(Q) is the form factor of the unimer, S(Q) isthe structure factor and bg is the incoherent background.

The Pedersen model was used to fit the SANS data obtained forthe pure Pluronics, as well as the data where the adjuvants werecontrast matched to the solvent.

Due to the large number of variables in the model some con-straints were initially used in the fitting procedure. These con-straints were relaxed during the latter part of the fitting as thefits began to converge. Known parameters, such as molecular vol-umes and concentrations, were, however, kept fixed throughoutthe fitting procedure.

A model with a spherical form factor and hard sphere structurefactor was used to fit the data obtained where the Pluronics werecontrast matched to the solvent. The form factor is in this case gi-ven by:

PðQÞ ¼ 3ðsinðQRÞ � QR cosðQRÞÞðQRÞ3

" #2

ð3Þ

440 M.A. Sharp et al. / Journal of Colloid and Interface Science 344 (2010) 438–446

Finally some samples were fitted using a core–shell rod model com-bined with a hard sphere structure factor. The form factor for therods is given by:

PðQÞ ¼ NZ p=2

0F2ðQÞ sinðaÞda ð4Þ

where

FðQÞ ¼ ðDqÞVsin 1

2 QL cosðaÞ� �

12 QL cosðaÞ

2J1ðQR sinðaÞÞQR sinðaÞ ð5Þ

where J1(x) is the first order Bessel function of the first kind, V is thevolume of the rod, L and R are the length and radius of the rod,respectively, and a is the angle between the axis of the rod andthe scattering vector Q.

In the simplest case of the PFG-NMR experiment a single spe-cies is present, diffusing isotropically and unrestricted. The ob-served attenuation curve will in this case fit well to a singleexponential decay. However, if the species is present at two sites,for example a block copolymer chain may be in the unimeric orthe micellar state, the shape of the observed attenuation curve willdepend on the rate of exchange of the species. A single exponentialcurve will be obtained if the exchange of the species is fast on thetimescale of the NMR experiment, i.e. compared to the diffusiontime D. The diffusion coefficient obtained is then an average ofthe two diffusion coefficients:

Deff ¼ Us � Ds þUf � Df ð6Þ

where Deff is the diffusion coefficient obtained experimentally, Us

and Uf are the fractions of the slow and fast species, respectively,and Ds and Df are the diffusion coefficients of the slow and fast spe-cies, respectively.

Alternatively, if the exchange of the species is slow on the NMRtimescale, a double exponential decay will be observed:

wðT þ 2sÞ ¼ Us expð�kDsÞ þUf expð�kDf Þ ð7Þ

where

k ¼ q2 D� d3

� �ð8Þ

and q = cgd, c is the gyromagnetic ratio.Other more complex equations arise in the case of intermediate

exchange, but these are not relevant in the cases considered here.

3. Results and discussion

In the first experiment the amount of benzyl benzoate and ben-zyl alcohol that could be solubilised by the aqueous polymer solu-tions was investigated. This was done by preparing solutions withincreasing adjuvant concentrations and observing the point atwhich the adjuvant could no longer be solubilised. The resultsare summarised in Table 1. While benzyl benzoate is insolubleand benzyl alcohol is only slightly soluble in pure water, significant

Table 1Amount of the adjuvants that can be solubilised by the different Pluronics. The error is±0.1%. The Pluronic solutions all consist of 10 w/w% solutions of the Pluronic in water.bb: benzyl benzoate, ba: benzyl alcohol. n and m are the number of EO and PO units in(EO)n(PO)m(EO)n. MW is the molecular weight of the Pluronic. HLB is the hydropho-bic–lipophilic balance of the Pluronic. cmc is the critical micellisation concentrationreported at 25 �C by Alexandridis et al. [40].

Solution bb (%) ba (%) n m MW (g/mol) HLB cmc (w/v%)

H2O 0 2 – – – – –P85 2 6 26 40 4600 16 4P105 1.5 7 37 56 6500 15 0.3F127 1 10 99 65 12600 22 0.7

amounts can be solubilised by the polymer solutions, as expected[14,15]. The amount of adjuvant that can be solubilised was notfound to correlate with the hydrophobic-lipophilic balance, butrather with the size of the hydrophobic block of the polymer. Itcan be seen that as the amount of the hydrophobic component,poly(propylene oxide), in the polymer increases, the amount ofbenzyl benzoate that can be solubilised decreases, in contrast tothe observations made by Gadelle et al. [18], where the oppositetrend was found. On the other hand, the amount of benzyl alcoholthat can be solubilised increases as the amount of poly(propyleneoxide) increases, in agreement with the observations made byGadelle et al. [18]. The discrepancy presumably arises from the factthat Gadelle measured the quantity that the polymer itself couldsolubilise, while here it was the amount that the solution as awhole could solubilise, while the difference in the behaviour ofbenzyl alcohol and benzyl benzoate arises from their differentaqueous solubilities.

3.1. Small angle neutron scattering

In the SANS measurements contrast variation was first of allused to determine where in the system the adjuvants were pres-ent. For a given concentration of polymer and adjuvant in D2Othe scattering intensity was compared for hydrogenated and deu-terated adjuvant. The results for Pluronic P85 with benzyl alcoholand benzyl benzoate are shown in Fig. 1. Comparing the curves ob-tained for the two adjuvants it can be seen that the curves for ben-

Fig. 1. Top: P85 with 2% benzyl benzoate added. Bottom: P85 with 5% benzylalcohol added. The solvent was D2O in both cases. All data was obtained at ISIS.

Fig. 2. Top: P85 with 1% and 2% benzyl benzoate added; data obtained at ISIS.Bottom: P85 with 2%, 4% and 6% benzyl alcohol added; data obtained at GKSS. Alldata obtained at 25 �C. The benzyl benzoate and benzyl alcohol are contrastmatched to the aqueous solvent. The scattering curve shown for the pure polymerhas been scaled from the fit obtained in pure D2O to how it would scatter at theD2O:H2O ratio used in contrast matching the adjuvants.

Table 3Fit parameters from the Pedersen model used to fit P85 with benzyl benzoate (bb)added at 25 �C. The benzyl benzoate has been contrast matched to the aqueoussolvent.

10% P85 +1% bb +2% bb

Nagg 25.04 ± 0.37 49.06 ± 1.71 87.89 ± 0.38Ucore 0.367 ± 0.008 0.413 ± 0.021 0.459 ± 0.027RGPEO (Å) 11.22 ± 0.20 10.23 ± 0.547 9.59 ± 0.54UHS 0.126 ± 0.001 0.190 ± 0.002 0.227 ± 0.002DR (Å) 23.98 ± 0.42 30.93 ± 0.548 32.16 ± 0.44R 0.371 ± 0.016 0.380 ± 0.022 0.351 ± 0.019Umic 0.497 ± 0.008 0.726 ± 0.025 0.760 ± 0.030RGuni (Å) 19.95 ± 0.488 19.19 ± 1.05 18.39 ± 1.31Rcore (Å) 41.79 53.83 66.31RHS (Å) 65.77 84.76 98.47

2

M.A. Sharp et al. / Journal of Colloid and Interface Science 344 (2010) 438–446 441

zyl benzoate have steeper slopes and more pronounced shoulderscompared to the curves obtained for benzyl alcohol. This signifiesmicelles with smaller polydispersity and sharper interfaces. Com-paring the hydrogenated/deuterated curves it can be seen thatthe maxima and minima occur at the same Q-values, but that thescattering intensity is greater for the hydrogenated samples. Thescattering cross section is proportional to the square of the differ-ence in scattering length density (the contrast). Since all else isequal, the difference in the observed scattering cross sections mustbe directly related to the difference in the contrast. There are threepossible scenarios for where the adjuvant is present: Either it ispresent within the micelles or it is within the solvent, or it maybe a combination of these two scenarios. By calculating the differ-ence in the contrast for these two possibilities it can be deducedthat if the adjuvant is present in the micelles the hydrogenatedversion will have a larger contrast than the deuterated version,while if the adjuvant is present in the solvent the deuteratedversion will have the larger contrast. The effect here wouldhowever be much smaller.

Quantitatively, it can be calculated from the scattering lengthdensities that the scattering cross section should be around 10%higher for the hydrogenated sample in the case of 2% benzyl benzo-ate and approximately 37% higher for 5% benzyl alcohol, if theadjuvant is situated within the core of the micelle. In contrast, ifthe adjuvant were situated in the solvent, the deuterated sampleswould have scattering cross sections approximately 0.5% and2.5% higher for the benzyl benzoate and benzyl alcohol samples,respectively. From the scattering curves shown in Fig. 1, it wasfound that the scattering cross sections were 13% and 59% higherfor the hydrogenated samples for benzyl benzoate and benzyl alco-hol, respectively. These results show that the adjuvant is in bothcases predominately present within the micelles rather than inthe solvent.

In order to reduce the complexity of the analysis of the system,contrast matching was used. This has the advantage of avoidinghaving to make assumptions about the exact location of the adju-vants in the micelles and their volume fractions in the differentparts of the micelle. The scattering length densities of the differentcomponents are given in Table 2. Fig. 2 shows the scattering curvesobtained for Pluronic P85 with deuterated benzyl benzoate (top)and deuterated benzyl alcohol (bottom), where the aqueous sol-vent has been contrast matched to the adjuvants. Only the scatter-ing from the polymer should remain. The best fits obtained usingthe Pedersen model are shown with the fit parameters given inTables 3 and 4. Qualitatively, it can be seen that there is a sharp in-crease in the scattering intensity upon the addition of either benzylbenzoate or benzyl alcohol, indicating an increase in the number ofmicelles. In addition there is a shift to lower Q of the scatteringcurves, both the maxima and the minima, showing that the struc-tures present are increasing in size. Finally it can be seen that theslope of the scattering curve in the mid-Q region becomes notica-bly steeper in the case of benzyl benzoate, indicating that theroughness of the micelles is decreasing.

For both benzyl benzoate and benzyl alcohol it was found thatthere was a significant increase in the aggregation number (Nagg)

Table 2Scattering length densities of the compounds used. ba is benzyl alcohol, bb is benzylbenzoate, PEO is poly(ethylene oxide) and PPO is poly(propylene oxide).

Compound SLD (10�6 �2) Compound SLD (10�6 �2)

D2O 6.38 H2O �0.56d5-ba 4.14 h-ba 1.31d5-bb 3.46 h-bb 1.90

PEO 0.61PPO 0.34

v 0.728 0.534 0.777

upon the addition of the adjuvants. For benzyl benzoate the frac-tion of solvent in the core of the micelles (Ucore) was found to in-crease slightly, while for benzyl alcohol an initial decrease wasobserved followed by a significant increase. This increase in Ucore

is consistent with the adjuvant being solubilised into the core ofthe micelles, since the fraction of solvent in the core of the micellereflects the combined water and adjuvant content in the micellarcore for these measurements because the adjuvants and solventare contrast matched.

Table 4Fit parameters from the Pedersen model used to fit P85 with benzyl alcohol (ba)added at 25 �C. The benzyl alcohol has been contrast matched to the aqueous solvent.

10% P85 +2% ba +4% ba +6% ba

Nagg 25.04 ± 0.37 27.47 ± 0.51 33.85 ± 0.41 33.19 ± 0.52Ucore 0.367 ± 0.008 0.354 ± 0.014 0.450 ± 0.007 0.618 ± 0.006RGPEO (Å) 11.22 ± 0.20 10.04 ± 0.52 9.86 ± 0.31 12.85 ± 0.43UHS 0.126 ± 0.001 0.235 ± 0.004 0.252 ± 0.003 0.168 ± 0.005DR (Å) 23.98 ± 0.42 27.19 ± 0.50 28.42 ± 0.42 41.46 ± 0.90R 0.371 ± 0.016 0.386 ± 0.013 0.397 ± 0.010 0.424 ± 0.013Umic 0.497 ± 0.008 0.650 ± 0.005 0.780 ± 0.005 0.894 ± 0.005RGuni (Å) 19.95 ± 0.488 27.09 ± 0.84 29.24 ± 1.06 23.28 ± 1.33Rcore (Å) 41.79 43.10 49.01 55.66RHS (Å) 65.77 70.29 77.43 97.12v2 0.728 64.84 45.82 35.39

442 M.A. Sharp et al. / Journal of Colloid and Interface Science 344 (2010) 438–446

The radius of the core of the micelles (Rcore), which is calculatedfrom these two parameters as well as the polydispersity (R), wasfound to increase significantly upon the addition of benzyl benzo-ate and benzyl alcohol. The increase in size of the micelles is inagreement with the predictions made by Nagarajan for the addi-tion of hydrophobic moieties to a number of Pluronics [15] andalso with the results obtained by Lettow et al. [19] for the additionof 1, 3, 5-trimethylbenzene and 1, 2-dichlorobenzene to PluronicP123.

From the fitting it is found that the radius of gyration of thepoly(ethylene oxide) chains in the micelles (RGPEO) decreasesslightly as benzyl benzoate is added, while for benzyl alcohol aslight decrease is observed initially, followed by an increase forthe highest concentration of benzyl alcohol added. These trendsare consistent with the changes in the size of the micelles formed,however the increase for the 6% benzyl alcohol sample indicatesthat there is a change in shape of the micelle to a more elongatedstructure, as will be discussed in more detail further on. It can alsobe seen that the radius of gyration of the unimers (RGuni) does notchange significantly as the adjuvants are added, suggesting thatthe adjuvants are only interacting with the micelles.

In addition it can be seen that the fraction of polymer micellised(Umic) increases as benzyl benzoate and benzyl alcohol are added.In effect, the critical micellisation concentration has been loweredfrom around 5% to around 2.5% at 25 �C for benzyl benzoate, whilefor benzyl alcohol it is lowered to around 1%. Such an effect hasalso been observed for Pluronics upon the addition of some saltsor the use of relatively hydrophobic cosolvents [41–43]. Addition-ally differential scanning calorimetry measurements show a shiftin the critical micellisation temperature to lower temperatures asbenzyl alcohol is added. For benzyl benzoate such a shift is lessclear, as shown in the supplementary material.

From the scattering curves it can be seen that the scatteringintensity increases as benzyl alcohol is added. This is similar towhat is observed for benzyl benzoate. In contrast to benzyl benzo-ate, however, the scattering intensity in the low-Q region starts toincrease at the highest benzyl alcohol concentration. This region ofthe scattering curves is dominated by the structure factor, which,in the model used for the fitting, uses a hard sphere potential.The fit parameters that are related to the structure factor are theinteraction radius (DR) and the volume fraction of hard spheresUHS. From the fitting it was found that the interaction radius in-creases as the benzyl alcohol concentration increases, while thefraction of hard spheres in the system was initially found to in-crease, but then shows an apparent decrease at the highest benzylalcohol concentration studied (6%). However, since the fraction ofpolymer micellised and the overall hard sphere radius (RHS) in-crease as benzyl alcohol is added both the number of micellesand the volume fraction of hard spheres must also increase. Thissuggests that there could be a change in the intermicellar interac-

tion potential from repulsive to attractive, with the micelles begin-ning to aggregate to some extent. Aggregation of polymericmicelles has been observed for other polymeric systems [44,45].The parameters describing the structure factor in the model wouldthen no longer be physically meaningful, since they relate to a hardsphere repulsive interaction. The alternative explanation is that weare observing a change in the shape of the micelle with the mi-celles becoming elongated at this high benzyl alcohol concentra-tion. A change to elongated structures has been reported for purePluronics at much higher temperatures [4,46] or upon the additionof salts and cosolvents [47–50]. However, Nagarajan, who carriedout a study of the predicted solubilisation and equilibrium shapesexpected for a number of Pluronics with different hydrophobiccompounds added, does not predict a change in shape for PluronicP85 at room temperature [15]. The argument for this is the high ra-tio of ethylene oxide to propylene oxide, which favours the spher-ical shape. There are however other factors affecting the preferredstructure, and if the presence of benzyl alcohol lowers the solubili-sate–solvent interfacial tension, this would begin to favour a cylin-drical structure [15]. The increase in the radius of gyration ofpoly(ethylene oxide) and the decrease in the fraction of hardspheres are consistent with a change to a more cylindrical shape,and it was therefore decided to attempt to fit the 6% benzyl alcoholsample to a cylindrical model. The fit obtained is also shown inFig. 2, and gives a radius of 51 Å, consistent with the radius ob-tained from the spherical model, and a length of the rods of139 Å. While both the spherical and cylindrical models give excel-lent fits to the data, the cylindrical model is more consistent withthe other parameters already discussed. It should however benoted that the cylindrical fit suggests only a slight elongation ofthe micelles. Recent shear-SANS measurements on this samplesupports the formation of elongated structures at the highest ben-zyl alcohol concentration, with lengths of at least 500 Å and ananisotropy factor [51] of around 0.4 [52].

In order to investigate the structure of the micelles further itwas decided to contrast match the solvent to the polymer, leavingonly the scattering from the adjuvant present. In fact what we didwas match the solvent to the ethylene oxide block. While thismeans that in principle propylene oxide is still visible, the differ-ence in scattering length density between ethylene and propyleneoxide (0.61 � 10�6 and 0.34 � 10�6 �2) compared to the scatter-ing length density of deuterated benzyl alcohol (4.14 � 10�6 �2)means that in practice the scattering from the propylene oxideblock is negligible. This has also been verified by carrying out ameasurement with Pluronic P85 in the absence of adjuvants andobserving the scattering at this contrast; no scattering was ob-served above the background level.

Fig. 3 shows the scattering curves obtained for Pluronic P85with deuterated benzyl benzoate (top) and deuterated benzyl alco-hol (bottom), where the polymer has been contrast matched out.For this contrast no scattering curves are shown at the lowest con-centrations of adjuvant studied for the two systems (1 w/w% ben-zyl benzoate and 2 w/w% benzyl alcohol) since the scatteringobtained from these systems was not above the background level.This is most likely a consequence of the poor contrast between theparticle and the solvent, due to a significant amount of solventwithin the core of the micelles for these samples. As described ear-lier, contrast variation shows that the adjuvant is present withinthe micelle.

The solid lines show the best fits obtained using a sphericalform factor with a hard sphere structure factor. The fit parametersused to obtain these fits are given in Tables 5 and 6. It was foundthat the contrast between the particle (in this case the adjuvant)and the solvent (the aqueous solution) was not as large asexpected. The scattering length density difference is expected tobe around 2.85 � 10�6 �2 for benzyl benzoate and around

Fig. 3. Top: P85 with 2% benzyl benzoate added; data obtained at ISIS. Bottom: P85with 4% and 6% benzyl alcohol added; data obtained at GKSS. In both cases theethylene oxide part of the polymer is contrast matched to the aqueous solvent.

Table 5Fit parameters from a simple hard spheres model used to fit P85 with 2% benzylbenzoate. The ethylene oxide component of the Pluronic has been contrast matchedto the aqueous solvent.

P85 +2% bb

SLD diff. (Å�2) 1.42 � 10�6 ± 0.13 � 10�6

Volume fraction 0.016 ± 0.002Radius (Å) 32.55 ± 0.88Polydispersity 0.146 ± 0.010Background 0.753 ± 0.004v2 0.497

Table 6Fit parameters from the hard sphere model used to fit P85 with benzyl alcohol added.The poly(ethylene oxide) has been contrast matched to the aqueous solvent.

+4% ba +6% ba

SLD dif. (Å�2) 0.85 � 10�6 ± 0.05 � 10�6 1.44 � 10�6 ± 0.03 � 10�6

Volume fraction 0.032 ± 0.003 0.031 ± 0.001Radius (Å) 30.36 ± 1.05 40.27 ± 0.76Polydispersity 0.323 ± 0.016 0.221 ± 0.011Background 0.018 ± 0.001 0.022 ± 0.001v2 stat. 2.67 3.55

Fig. 4. Attenuation plot for Pluronic P85 at 25 �C with benzyl alcohol added. Theresults are for the methyl peak of the polymer.

M.A. Sharp et al. / Journal of Colloid and Interface Science 344 (2010) 438–446 443

3.53 � 10�6 �2 for benzyl alcohol. The lower than expected scat-tering length density difference shows the presence of somepoly(propylene oxide) and/or water within the core of the micelle,

thereby reducing the contrast. This is to be expected. As the benzylalcohol concentration increases it can be seen that the contrast im-proves. This is consistent with the mechanism proposed by Gadelleet al. [18] for the solubilisation of compounds by block copolymers,which predicts the displacement of water from the core by the sol-ubilisate, in this case benzyl alcohol.

The radii obtained from these fits are consistent with the radiiobtained for the core of the micelle obtained at the other contrast,even though the radius obtained is slightly lower. Based on theseobservations it is proposed that the centre of the micelle consistsof the adjuvant with some solvent and poly(propylene oxide). Thisis surrounded by a thin shell of poly(propylene oxide) and solvent,while the outer corona of the micelle consists of highly swollenpoly(ethylene oxide).

Despite the excellent fit obtained for the 6% benzyl alcohol sam-ple it was also attempted to fit the data using a cylindrical model,due to the evidence of the micelles becoming elongated at thisadjuvant concentration, as discussed earlier. The fit obtained isshown in Fig. 3, and it was found that the rods have a radius of40 Å and a length of 110 Å. The radius is again consistent with thatobtained using the spherical model, but smaller than for the fit ob-tained with the cylindrical model at the other contrast, as is thelength obtained. The results are consistent with the adjuvant beingpresent only in the centre of the micelles.

3.2. Pulsed-field gradient NMR

Both systems were also studied using PFG-NMR, which providesinformation about the diffusional behaviour of the different com-ponents in the system. The diffusion coefficients were obtainedby fitting the attenuation curves to a single diffusion constant forboth the polymer and the adjuvants. For the Pluronics, only themethyl peak was used, while for benzyl benzoate and benzyl alco-hol both the CH2 peak and the peaks from the aromatic rings wereused in the analysis. The attenuation plot obtained for the polymermethyl peak when benzyl alcohol is added is shown in Fig. 4. It canbe seen that a single diffusion coefficient is obtained. Benzyl ben-zoate shows similar results. The diffusion coefficients obtainedhave been plotted as a function of the adjuvant concentration.The results are shown in Fig. 5 for benzyl benzoate and for benzylalcohol. For benzyl alcohol the diffusion coefficient of benzyl alco-hol in aqueous solution in the absence of polymer is also shown, upto 2%, the concentration to which it is soluble.

For Pluronic P85 with benzyl benzoate it can be seen that ben-zyl benzoate always diffuses at a rate smaller than the polymer.

Fig. 5. Diffusion coefficient against adjuvant concentration for Pluronic P85 at25 �C. Results are shown for benzyl benzoate and benzyl alcohol together. ‘‘P85(bb)” and ‘‘P85 (ba)” are the values obtained for the polymer in the presence ofbenzyl benzoate and benzyl alcohol, respectively, while ‘‘bb (P85)” and ‘‘ba (P85)”are the values obtained for the adjuvants in the presence of the polymer.

444 M.A. Sharp et al. / Journal of Colloid and Interface Science 344 (2010) 438–446

This shows that benzyl benzoate is fully solubilised by the poly-meric micelles, in agreement with the results obtained by SANS.The diffusion coefficient obtained for the polymer is an averageof the diffusion coefficients of the unimer and the micelle. Sincethe polymer is undergoing fast exchange on the timescale of theNMR measurement only one diffusion coefficient is obtained. Fromthe relative diffusion coefficients the fraction of polymer that ismicellised can be calculated, since:

Deff ¼ Ufree � Dfree þUmic � Dmic ð9Þ

where Deff is the effective polymer diffusion coefficient, which is ta-ken as the value of the polymer with benzyl benzoate added, Dfree isthe free polymer diffusion coefficient, taken as the value when nobenzyl benzoate has been added, and Dmic is the diffusion coefficientof the polymer in the micelle, taken as the value of the diffusioncoefficient of benzyl benzoate. Ufree and Umic are the fractions offree and micellised polymer, respectively. The results obtained forthe fraction of Pluronic P85 that has been micellised is shown inFig. 6, and it was found that as the concentration of benzyl benzoateincreases, the fraction of polymer micellised increases, in agree-ment with the small-angle neutron scattering results. The numeri-

Fig. 6. Fraction of Pluronic P85 micellised as a function of the concentration ofbenzyl benzoate that has been added, as obtained by PFG-NMR at 25 �C.

cal values obtained are in reasonable agreement with the valuesobtained by SANS.

With benzyl alcohol added to Pluronic P85 it is found that thebenzyl alcohol diffuses significantly faster than the polymer at allbenzyl alcohol concentrations. This means that some or all of thebenzyl alcohol is diffusing freely in the aqueous solution, whichis not surprising since benzyl alcohol is partly water soluble. Fromthe diffusion coefficients obtained it is possible to estimate whatfraction of benzyl alcohol is bound to the polymeric micelles andwhat fraction is diffusing freely in the solution, since

Deff ¼ Ufree � Dfree þUbound � Dbound ð10Þ

where Deff is the average diffusion coefficient of benzyl alcohol ob-tained in the presence of the polymer, Ufree and Ubound are the frac-tions of free and bound benzyl alcohol, respectively and Dfree andDbound are the diffusion coefficients of the free and bound benzylalcohol, as obtained in the presence of the polymer. For the calcula-tion some assumptions have to be made: First of all the value takenfor Dfree is assumed to be equal to the diffusion coefficient obtainedfor the free benzyl alcohol in aqueous solution and Dbound is taken tohave the value of the diffusion coefficient of the polymer. It shouldbe noted that the calculation will not be entirely accurate, since,first of all, the value of the free benzyl alcohol diffusion coefficientis likely to change as the concentration is increased, secondly, thediffusion coefficient obtained for the polymer in the presence ofbenzyl alcohol will be an average of the diffusion coefficients ofthe unimer and micelle. This must be the case, since SANS showsthat not all the polymer is micellised. It is therefore possible thatthe values obtained for the fraction of benzyl alcohol bound tothe micelles is slightly higher than the real value. The fraction isshown in Fig. 7, and is found to increase as the benzyl alcohol con-centration increases. From the values obtained it is also possible tocalculate the percentage out of the overall system that benzyl alco-hol makes up in free solution and what percentage is bound to themicelles. This is shown in Fig. 8, where it can be seen that both thepercentage of free and bound benzyl alcohol increase as the amountof benzyl alcohol added to the system increases. Since benzyl alco-hol is soluble to around 2 w/w% in aqueous solution it was thoughtthat the amount of free benzyl alcohol would remain constant atthis value, with any benzyl alcohol present in the system in excessof this amount situated within the micelles. This is clearly not thecase. It therefore appears that it is favourable for benzyl alcoholto be situated within the micelles, even at concentrations whereit is soluble in the aqueous solution.

Fig. 7. Fraction of benzyl alcohol bound to the Pluronic P85 micelles, plotted as afunction of the concentration of benzyl alcohol added to the polymeric solution, ascalculated from the results obtained from PFG-NMR.

Fig. 8. Percentage, of the overall system, of benzyl alcohol free and bound to thePluronic P85 micelles, plotted as a function of the concentration of benzyl alcoholadded to the polymeric solution. The calculation is based on the results obtainedfrom PFG-NMR.

M.A. Sharp et al. / Journal of Colloid and Interface Science 344 (2010) 438–446 445

One important observation that can be made from the diffusioncoefficients obtained for benzyl alcohol is that there is a significantdrop in the diffusion coefficient of the polymer above 5% benzylalcohol concentration. This drop in diffusion coefficient indicatesthe presence of much larger structures forming, consistent withthe micelles becoming elongated to form cylindrical micelles. Tak-ing the ratio of the diffusion coefficient obtained before and afterthe observed decrease, one can compare with the ratio predictedfor the diffusion coefficient of rods and spheres, by using the fol-lowing expression for the hydrodynamic radius of a rod [53–55]:

RH �L

2 lnðL=dÞ ð11Þ

where L is the length and d is the diameter of the rods. A reasonableagreement is achieved between the measured and predicted ratios,thereby supporting the evidence that rod-like micelles are formed,when using the diffusion coefficient obtained by NMR and the sizesas obtained by neutron scattering, and taking into account thechange in viscosity as benzyl alcohol is added.

3.3. Pluronics P105 and F127

Measurements were also carried out for Pluronic P105 and F127with benzyl benzoate and benzyl alcohol using both SANS andPFG-NMR. The observed trends were generally similar to those ob-tained for Pluronic P85, therefore only significant differences shallbe discussed here.

While there was a significant increase in the fraction of polymermicellised for P85, the increase observed for the other Pluronics ismuch smaller. From the values obtained it is possible to calculatethe change in the cmc as the adjuvants are added. For P105 thecmc decreased from around 1.8% to around 1.4% in the presenceof benzyl benzoate, while for F127 the decrease in the cmc wasfrom 2.2% to 1.6%. In both cases the maximum benzyl benzoateconcentration studied was 1%. In the presence of benzyl alcoholthe cmc decreased from 1.8% to 0.7% for P105 and from 2.2% to0.01% for F127. The maximum benzyl alcohol concentration was7% in the case of P105 and 10% in the case of F127. This smaller ef-fect is most likely a consequence of both P105 and F127 alreadybeing highly micellised at this polymer concentration (10 w/w%).

While benzyl benzoate diffuses at a similar rate or slower forboth P85 and P105, it was found to diffuse slightly faster thanF127 at all concentrations studied. From the results obtained by

small-angle neutron scattering it is possible to estimate the inter-micellar distances [56]. It was found that the mean surface-to-sur-face distance between F127 micelles was less than 14 Å in thepresence of benzyl benzoate. This value is significantly lower thanfor P105 (around 45 Å) and P85 (around 68 Å). Since the micellesare very close together it is therefore possible that the benzyl ben-zoate is exchanging between the F127 micelles. This is not possiblefor P85 or P105, due to the larger intermicellar distances.

For all three Pluronics benzyl alcohol diffuses significantly fas-ter than the polymer. However, while for both P85 and P105 asharp drop in the diffusion coefficient is observed at high benzylalcohol concentrations, no such observation is made for F127.

The fraction of bound and free benzyl alcohol as well as the per-centage of the overall system that benzyl alcohol makes up wascalculated for P105 and F127 in the same way as for P85, and inter-estingly it was found that, at the maximum concentration of benzylalcohol that could be solubilised, the percentage of free benzylalcohol lies around 2%. This was the case for all three polymersstudied. It therefore seems possible that this is the critical concen-tration for determining the maximum amount of benzyl alcoholthat can be solubilised.

4. Conclusions

The solubilisation of two model adjuvants, benzyl benzoate andbenzyl alcohol, by micellar solutions of Pluronics P85, P105 andF127 was studied using SANS and PFG-NMR. The work has led toa better understanding of how the structure of the micelles is af-fected by the presence of these model adjuvants, but also of thedynamics in such systems. It was found that the micelles swelledsignificantly upon the addition of the adjuvants, and led to a de-crease in the critical micelle concentration. Using contrast varia-tion techniques it was shown that the adjuvants are presentwithin the core of the micelles. Pulsed-field gradient NMR allowedfor the calculation of the amount of adjuvant bound to the micelles.It was found that the insoluble adjuvant (benzyl benzoate) wasonly present within the micelles, while the partially soluble adju-vant (benzyl alcohol) was present in both the micelles and the sur-rounding solution. It is normally assumed that any solubilisates arepresent in the solution up to the concentrations at which they aresoluble, and only above this concentration do they become solubi-lised by the micelles. Our work shows that this is in fact not thecase, and great care must be taken when calculating the partition-ing of solubilisates.

This study has shown that while for benzyl benzoate and lowconcentrations of benzyl alcohol the micelles swell significantlythey remain spherical in shape as predicted theoretically. However,when the amount of benzyl alcohol added increases significantlythere is a change in the shape of the micelles to elongated struc-tures. This is contrary to the theoretical calculations for these Plur-onics and has not previously been observed with such lowconcentrations of additives at this temperature.

Since the system studied is pharmaceutically important, it isimportant to also gain an understanding of the changes that takeplace as the temperature is increased from room temperature tobody temperature. This work will be described in a separate paper.

Acknowledgments

This research project has been supported by the European Com-mission under the 6th Framework Programme through the KeyAction: Strengthening the European Research Area, Research Infra-structures. Contract no.: RII3-CT-2003-505925. We thank GKSSand ISIS for the provision of neutron beamtime, in particular wethank Dr. Vasil Haramus (GKSS) and Dr. Stephen King (ISIS) for

446 M.A. Sharp et al. / Journal of Colloid and Interface Science 344 (2010) 438–446

their help during the measurements. We thank Dr. Youssef Espidelfor his assistance with the NMR measurements. We thank Jan SkovPedersen and Richard Heenan for their help with model fitting theSANS data. MAS acknowledges the EPSRC and AstraZeneca forfunding of her doctoral work.

Appendix A. Supplementary data

Supplementary data associated with this article can be found, inthe online version, at doi:10.1016/j.jcis.2010.01.005.

References

[1] P. Alexandridis, T. Hatton, Colloids Surf., A 96 (1995) 1.[2] W. Brown, K. Schillen, M. Almgren, S. Hvidt, P. Bahadur, J. Phys. Chem. 95

(1991) 1850.[3] W. Brown, K. Schillen, S. Hvidt, J. Phys. Chem. 96 (1992) 6038.[4] K. Mortensen, J. Phys.: Condens. Matter 8 (1996) A103.[5] I. Goldmints, F. von Gottberg, K. Smith, T. Hatton, Langmuir 13 (1997) 3659.[6] G. Wanka, H. Hoffmann, W. Ulbricht, Macromolecules 27 (1994) 4145.[7] K. Zhang, A. Khan, Macromolecules 28 (1995) 3807.[8] A. Iovino, C. La Mesa, D. Capitani, A. Segre, Colloid Polym. Sci. 281 (2003) 1136.[9] A. Caragheorgheopol, H. Caldararu, I. Dragutan, H. Joela, W. Brown, Langmuir

13 (1997) 6912.[10] <http://www.basf.com/performancechemical/bcperfapplications.html> URL

valid at time of publication.[11] G. Riess, Prog. Polym. Sci. 28 (2003) 1107.[12] I. Hamley, The Physics of Block Copolymers, first ed., Oxford University Press,

Oxford, 1998.[13] K. Mortensen, J. Pedersen, Macromolecules 26 (1993) 805–812.[14] R. Nagarajan, M. Barry, E. Ruckenstein, Langmuir 2 (1986) 210–215.[15] R. Nagarajan, Colloids Surf., B 16 (1999) 55.[16] P. Hurter, T. Hatton, Langmuir 8 (1992) 1291.[17] S. Vauthey, M. Leser, N. Garti, H. Watzke, J. Colloid Interface Sci. 225 (2000) 16.[18] F. Gadelle, W. Koros, R. Schechter, Macromolecules 28 (1995) 4883.[19] J. Lettow, T. Lancaster, C. Glinka, J. Ying, Langmuir 21 (2005) 5738.[20] L. Arleth, B. Svensson, K. Mortensen, J. Pedersen, U. Olsson, Langmuir 23 (2007)

2117.[21] P. Alexandridis, U. Olsson, B. Lindman, Macromolecules 28 (1995) 7700.

[22] R. Ivanova, B. Lindman, P. Alexandridis, Langmuir 16 (2000) 3660.[23] R. Ivanova, B. Lindman, P. Alexandridis, Langmuir 16 (2000) 9058.[24] M. Malmsten, Soft Matter 2 (2006) 760.[25] N. Rapoport, Colloids Surf., B 16 (1999) 93.[26] C. Allen, D. Maysinger, A. Eisenberg, Colloids Surf., B 16 (1999) 3.[27] A. Kabanov, E. Batrakova, V. Alakhov, J. Controlled Release 82 (2002) 189.[28] R. Ivanova, P. Alexandridis, B. Lindman, Colloids Surf. 183–185 (2001) 41.[29] P. Alexandridis, R. Spontak, Curr. Opin. Colloid Interface Sci. 4 (1999) 130.[30] P. Alexandridis, R. Ivanova, B. Lindman, Langmuir 16 (2000) 3676.[31] C. Johnson, Prog. Nucl. Magn. Reson. Spectrosc. 34 (1999) 203.[32] W. Price, Concepts Magn. Reson. 9 (1997) 299.[33] W. Price, Concepts Magn. Reson. 10 (1998) 197.[34] K. Mortensen, Collids Surf. A 183–185 (2001) 277.[35] J. Pedersen, M. Gerstenberg, Macromolecules 29 (1996) 1363.[36] I. Goldmints, G.-E. Yu, C. Booth, K. Smith, T. Hatton, Langmuir 15 (1999) 1651.[37] P. Linse, Macromolecules 26 (1993) 4437.[38] J. Pedersen, J. Chem. Phys. 114 (2001) 2839.[39] J. Percus, G. Yevick, Phys. Rev. 110 (1958) 1.[40] P. Alexandridis, J. Holzwarth, T. Hatton, Macromolecules 27 (1994) 2414.[41] P. Bahadur, P. Li, M. Almgren, W. Brown, Langmuir 8 (1992) 1903.[42] N. Jain, V. Aswal, P. Goyal, P. Bahadur, J. Phys. Chem. B 102 (1998) 8452.[43] P. Alexandridis, L. Yang, Macromolecules 33 (2000) 5574.[44] L. Lobry, N. Micali, F. Mallamace, C. Liao, S. Chen, Phys. Rev. E 60 (1999) 7076.[45] L. Shen, H. Wang, G. Guerin, C. Wu, I. Manners, M. Winnik, Macromolecules 41

(2008) 4380.[46] S. King, R. Heenan, V. Cloke, C. Washington, Macromolecules 30 (1997) 6215.[47] E. Jørgensen, S. Hvidt, W. Brown, K. Schillen, Macromolecules 30 (1997) 2355.[48] G. Mao, S. Sukumaran, G. Beacage, M.-L. Saboungi, P. Thiyagarajan,

Macromolecules 34 (2001) 552.[49] R. Ganguly, V. Aswal, P. Hassan, I. Gopalakrishnan, J. Yakhmi, J. Phys. Chem. B

109 (2005) 5653.[50] L. Guo, R. Colby, P. Thiyagarajan, Physica B 385–386 (2006) 685.[51] M.T. Truong, L. Walker, Langmuir 16 (2000) 7991.[52] M. Sharp, T. Cosgrove, Unpublished, 2009.[53] J. Riseman, J. Kirkwood, J. Phys. 18 (1950) 512.[54] V. Castelletto, P. Parras, I. Hamley, P. Bäverbäck, J. Pedersen, P. Panine,

Langmuir 23 (2007) 6896.[55] L. Bastardo, J. Iruthayaraj, M. Lundin, A. Dedinaite, A. Vareikis, R. Makuska, A.

van der Wal, I. Furo, V. Garamus, P. Claesson, J. Colloid Interface Sci. 312 (2007)21.

[56] M. Sharp, Using block copolymers to solubilise model adjuvants, Ph.D. Thesis,University of Bristol, 2007.