water interfaces covered by nonionic amphiphilic polysaccharides. Correlation with stability of oil-in-water emulsions

Colloids and Surfaces A: Physicochem. Eng. Aspects 441 (2014) 312–318

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Dilational rheology of air/water interfaces covered by nonionic amphiphilic polysaccharides. Correlation with stability of oil-in-water emulsions Rudy Covis a,b , Jacques Desbrieres c , Emmanuelle Marie a,b,1 , Alain Durand a,b,∗ a

CNRS, LCPM, FRE 3564, Nancy F-54001, France Université de Lorraine, LCPM, FRE 3564, Nancy F-54001, France c Université de Pau et des Pays de l’Adour, IPREM, UMR 5254, 2 Avenue Président Angot, F-64053 Pau, France b

h i g h l i g h t s

g r a p h i c a l

a b s t r a c t

• Amphiphilic polysaccharides were obtained by modifying dextran with 1,2-epoxydodecane. • Dilational rheology of air/water interface was studied according to polymer structure. • Oil-in-water submicronic emulsions were prepared with various dextran derivatives. • Optimal dextran modification was similar for emulsifying and interfacial properties.

a r t i c l e

i n f o

Article history: Received 30 June 2013 Received in revised form 27 August 2013 Accepted 19 September 2013 Available online 27 September 2013 Keywords: Dextran Amphiphilic polysaccharide Oil-in-water emulsion Dilational rheology

a b s t r a c t Biodegradable polymeric emulsifiers were prepared by covalent attachment of aliphatic hydrocarbon groups onto dextran macromolecules (a bacterial nonionic polysaccharide), varying the number of attached hydrocarbon tails. Submicronic oil-in-water emulsions were prepared by sonication using previous polymeric stabilizers with oil volume fractions between 10 and 50% and two oils (hexadecane and nujol). An optimal range of dextran hydrophobic modification was evidenced conciliating a strong anchoring of polymer to oil/water interface and sufficient steric repulsions between droplets. These results were correlated to dilational rheology of air/water interfaces covered by amphiphilic dextrans. The storage stability of these emulsions was of several weeks. © 2013 Elsevier B.V. All rights reserved.

1. Introduction Emulsions are commonly encountered in formulated products for which it is necessary to combine several immiscible liquid

∗ Corresponding author at: Universite´ de Lorraine, LCPM. Tel.: +33 03 83 17 52 92; fax: +33 03 83 37 99 77. E-mail address: [email protected] (A. Durand). 1 Current address: ENS – Département de Chimie, UMR 8640 CNRS – ENS – UPMC, 24 Rue Lhomond, F-75005 Paris, France. 0927-7757/$ – see front matter © 2013 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.colsurfa.2013.09.027

phases in order to obtain targeted end use properties. The main characteristics of emulsions are: droplet size distribution, rheological behavior, nature and volume fraction of liquid phases. In many applications, submicronic droplets diameters are desired in order to reach high surface-to-volume ratios. Thus performances of stabilizers in relation to interface properties become key formulation parameters. Surfactants and stabilizers obtained from renewable resources are attracting an increasing interest from both industry and academia. Especially, biopolymers are more and more considered as alternative formulation ingredients and among them proteins

R. Covis et al. / Colloids and Surfaces A: Physicochem. Eng. Aspects 441 (2014) 312–318

dextran sample, T40® from Amersham Pharmacia was used. Its weight-average and number-average molar masses were characterized by size exclusion chromatography and found equal to 40,000 g/mol and 33,000 g/mol, respectively.

( O OH O )x OH

HO

313

( O

2.2. Synthesis of dextran derivatives

O )y O CH2 CH

The experimental procedure for polymer synthesis was that discussed previously [8]. Briefly, commercial dextran (5 g) was dissolved in 100 mL of milliQ water. The required amount of tetrabutylammonium hydroxide (40% wt solution in water) was added (1.5 mol per repeat unit). After 1 h stirring, the mixture was freeze-dried. The resulting solid was dissolved in 100 mL dimethylsulfoxide (DMSO) and, if necessary, heated to 50 ◦ C. Then, the required amount of 1,2-epoxydodecane was added. The reaction was left proceed during 96 h under magnetic stirring. The crude reaction medium was transferred to a dialysis bag (molar mass cut off equal to 6–8000 g/mol) and dialyzed against water/ethanol mixture (50/50 v/v) and finally water. The final aqueous solution was freeze-dried.

OH HO

( CH2 )

9

CH3

OH Fig. 1. Repeat units of dextran derivatives.

and polysaccharides [1–4]. Various strategies have been reported for using biopolymers as stabilizers in dispersed systems like the use of adsorbing or non-adsorbing biopolymers, mixing proteins and polysaccharides, mixing biopolymers and molecular surfactants, etc. One particular way is to chemically modify native biopolymers in order to obtain functional macromolecules which combine biopolymer characteristics to new properties brought by modification procedure. Apart from the ability to specifically adjust physico-chemical properties to targeted applications, this strategy can also contribute to fundamental investigations. Indeed, series of chemically modified biopolymers with regularly varying structural characteristics can be useful tools for experimental investigation and acquisition of deeper knowledge about structure–property relationships. For many years we have been studying the synthesis and physico-chemical properties of amphiphilic polymers derived from polysaccharides. Such compounds combine strong adsorption capacity at liquid/liquid interfaces together with biocompatibility and biodegradability. These properties are relevant for certain applications involving specific interfacial interactions with environmental or biological media [5,6]. Previously, we studied the synthesis of amphiphilic polysaccharides by reacting dextran (Fig. 1), a nonionic bacterial polysaccharide, with aliphatic 1,2-epoxyalkanes [7]. The amount of grafted alkyl tails was varied by modifying the feed composition. Degrees of substitution (DS) up to 164% were obtained, DS being defined as the molar ratio of attached hydrocarbon groups to glucose repeat units (Eq. (1)) [8]: DS = 100 ×

number of attached alkyl tails number of glucose repeat units

(1)

In the case of 1,2-epoxydodecane, as long as DS was lower than 30%, dextran derivatives remained water soluble (up to 50 g/L at 25 ◦ C) and were shown to be efficient as stabilizers of oil-in-water submicronic emulsions with oil volume fractions lower than 20%. Within that range of oil volume fractions, we showed that emulsion aging was dominated by molecular diffusion [9]. The aging rate was strongly influenced by the solubility of oil in water, as predicted by the theory of Lishitz, Slyozov and Wagner [9]. This work was focused on the interfacial adsorption and emulsifying properties of water-soluble dextran derivatives modified by 1,2-epoxydodecane with DS values up to 30%. These amphiphilic polymers were used for preparing oil-in-water submicronic emulsions with oil volume fractions up to 50%. The droplet size distributions were correlated to the structural characteristics of amphiphilic dextrans. 2. Experimental 2.1. Materials All chemicals were from Aldrich with the highest purity available. MilliQ water was used in all experiments. A commercial

2.3. Polymer characterization Size exclusion chromatography analyses were performed with a system comprising a Merck L6200A pump (0.7 mL/min), a Degazys DG 1310, Uniflow unit, a 200 ␮L injection loop, a PL aquagel-OH Guard pre-column followed by 2 PL aquagel-OH 40 and PL aquagelOH 50 columns (Polymer Laboratories). The detection system was a miniDawn (Wyatt Technology Corporation) photodiffusiometer (wavelength 690 nm and detection at 41.6◦ , 90◦ and 138.4◦ ) followed by a Merck RI-71 differential refractometer. The eluent was 0.1 mol/L NaNO3 with 0.2 g/L NaN3 . The refractive index increment value (dn/dc) used for calculations was 0.145 mL/g. 2.4. Rheological measurements with aqueous solutions Aqueous solutions of polymers were prepared by gentle stirring in milliQ water for 24 h. Steady shear and dynamic measurements were performed at 13 ◦ C on a AR 2000 rheometer (TA instruments® ) using a parallel plate geometry (diameter 25 mm). The temperature of 13 ◦ C was chosen so as to limit the risks of drying of the samples during the measurements. Oscillatory experiments were performed within the linear viscoelasticity region, where storage (G ) and loss (G ) moduli were independent of the stress magnitude. 2.5. Dynamic surface tension and interfacial tension measurements A drop tensiometer (Tracker, IT Concept, France) was used to measure the surface tension  by analyzing the axial symmetric shape (Laplace profile) of the rising bubble in aqueous polymer solution [10,11]. All the measurements were performed at a controlled temperature (24 ◦ C). The two-dimensional complex modulus E of the adsorption layers was found from the measurement of the surface tension variations as the response to a sinusoidal variation of the surface area [11,12]. All the measurements were made during a sufficiently long time to follow the effect of the aging on the surface tension and on the dilatational rheological properties of the adsorption layers. Technically, the study of the structure formation kinetics inside the adsorption layers of surface-active compounds was performed by applying a small dilational perturbation A(t) to the bubble area (noted A) and simultaneously recording the variation of the surface tension (t) [or the surface pressure (t) =  0 − (t), where  0 is the surface tension of the solvent]. Periodic variation of the drop area A according to the sinusoidal law ε(t) = εa exp(iωt) (where

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ε = A/A is the relative dilational deformation of the layer, and εa , is the amplitude of this relative deformation) produces a variation of the surface pressure (t) = a exp[i(ωt + ˚)], where ˚ is the phase angle. In the linear deformation domain, the frequency-dependent complex modulus may be expressed by Eq. (2). E(ω) =

d dε

=

−d dε

= E  (ω) + iE  (ω)

(2)

In Eq. (2), E (ω) and E (ω) are the real and the imaginary parts, which depend on the applied frequency ω (i.e. elastic and viscous moduli respectively). For all experiments, the period of sinusoidal variations of the drop area was 30 s. The amplitude of the relative deformation of the layer, εa , was kept constant to 0.1 and it was checked that it was within the linear regime at the used frequency. To do so, several experiments were carried out, keeping the same period and changing εa . The linear domain corresponded to the range of εa values within which the modulus E kept unchanged. The reproducibility of the dilational rheological measurements had been checked. With DexC1010 and DexC1017 , two independent series of experiments carried out with two weeks in between led to superimposed curves of surface tension vs. time. As for dilational modulus, the found values obtain for the two series of experiments differed by about 1 mN/m. 2.6. Preparation of oil-in-water emulsions Oil-in-water emulsions were prepared by sonication using Vibracell (600 W), Sonic & Material Inc with 50% active cycle and an applied power equal to 5 W. The followed procedure was the same for all samples. An aqueous solution of dextran derivative was prepared by mixing dry polymer and milliQ water containing 10−3 g/L NaN3 .The mixture was stirred overnight. The required amount of oil (n-hexadecane or nujol) was added such that the weight ratio of dextran derivative to oil was 0.1. The mixture was vortexed during 30 s and sonicated during 30 s. This sequence was done four times successively. After this emulsification procedure no excess oil could be observed. Emulsion was sampled to determined droplet size distribution after preparation and stored at 4 ◦ C. Control experiments were performed to check that dextran derivatives were not degraded during emulsification procedure. For those experiments, the same procedure was followed without the addition of oil. 2.7. Determination of droplet size distribution and average droplet diameter of emulsions The diameter distributions of emulsion droplets were determined by light scattering measurements using a Master Sizer® apparatus from Malvern. The reported diameters D(0.1), D(0.5) and D(0.9) were diameters at 10, 50 and 90% of the volume distribution of droplets. In order to get an idea of the broadness of diameter distributions, we calculated the Span, defined by Eq. (3). Span =

D(0.9) − D(0.1) D(0.5)

(3)

Surface-average and volume-average droplet diameter calculated from light scattering measurements we denoted D[3,2] and D[4,3], respectively. 3. Results and discussion

1

Viscosity (Pa.s)

314

0.1

0.01

0.001 0

10

20

30

40

50

Polymer concentration (g/L)

Fig. 2. Viscosity of DexC107 aqueous solutions at 13 ◦ C measured with a plate–plate geometry at a stress equal to 0.1 Pa.

degree of substitution (DS, Eq. (1)). The label “C10” was used by reference to the number of carbon atoms in the alkyl tail following the carbon carrying the hydroxyl group from by the opening of epoxide ring (Fig. 1). 3.2. Aqueous solutions of amphiphilic dextrans The solution behavior of amphiphilic dextran derivatives had been discussed previously on the basis of capillary viscometry and light scattering experiments [7,13]. Dextran derivatives obtained by modification with phenoxy groups or aliphatic tails with 6 or 10 carbon atoms remained readily soluble in water and DMSO (up to 50 g/L) as long as DS was lower than 30%. This can be related to the fact that water and DMSO are good solvents for native dextran [14,15]. In aqueous solutions, it was showed that in the dilute domain, DexC10DS with DS ≥ 10% were under the form of compact aggregates gathering several macromolecules. Above a critical association concentration, Cass , associations between aggregates started to form and thus build a physical network. This range of concentration was considered as the semi-dilute domain. For DS values between 11 and 31%, the value of Cass in pure water was between 33 and 38 g/L according to capillary viscometry experiments. Using flow and oscillatory experiments, we investigated the rheological behavior of aqueous solutions of DexC107 with polymer concentrations varying between 10 and 50 g/L (Fig. 2). Over the explored range of strain, dynamic viscosity exhibited no significant variation. A significant raise of dynamic viscosity was detected between 30 and 40 g/L. This was attributed to the transition between dilute and semi-dilute domains. Oscillatory measurements carried out between 0.1 and 10 Hz showed that over that range of concentration, aqueous solutions behaved as viscous fluids, with loss modulus, G , higher than elastic modulus, G . The latter reached measurable values only for aqueous solutions with concentrations above 40 g/L. These results were consistent with previous data obtained by viscometry and showed that, for concentrations above 40 g/L in pure water, amphiphilic dextrans formed physical networks. Our previous work showed that native dextran T40® formed dilute aqueous solutions up to 60 g/L [7]. Thus, the observed variation of rheological behavior between 30 and 40 g/L should be attributed to additional polymer–polymer interactions brought by hydrocarbon tails. 3.3. Dilational rheology of air–water interfaces covered by amphiphilic dextrans

3.1. Polymeric emulsifiers synthesized from dextran Dextran derivatives were synthesized as reported previously [8]. Native dextran had a weight-average molar mass equal to 40,000 g/mol and a number-average molar mass equal to 33,000 g/mol. Polymers were named DexC10DS according to their

Modified dextrans adsorb at air–water and oil–water interfaces via covalently attached hydrocarbon groups. By considering the number-average degree of polymerization of dextran backbone which can be approximated to 204 and the range of DS, between 5 and 17%, the average number of hydrocarbon tails per

R. Covis et al. / Colloids and Surfaces A: Physicochem. Eng. Aspects 441 (2014) 312–318

315

60

Surface Tension (mN/m)

50

40

Surface Tension Dilational Module

30

20

10

0

4

6

8

10

12

14

16

18

20

Substitution Degree (%) Fig. 3. Surface tension and dilational modulus of 0.2 g/L DexC1010 aqueous solution as a function of time at 25 ◦ C.

macromolecule should vary between 10 and 35. Thus, in their adsorbed state, modified dextrans should have several hydrophobic units simultaneously in close contact with interface. This fact has several consequences like the kinetic irreversibility of adsorption as well as viscoelastic behavior of interfacial layer. Apart from the number of hydrocarbon tails, their chemical nature and their distribution within dextran backbone should have also significant effects on interfacial properties as it has been observed for aqueous solution properties of dextran or cellulose ether derivatives [16–18]. Since kinetic and equilibrium lowering of air–water and oil–water interfacial tension by water-soluble dextran derivatives had already been reported, this work was focused essentially on dilational rheology of fluid interfaces covered by a series of amphiphilic dextrans [19]. When following surface tension and dilational modulus as a function of time, a continuous variation was observed over more than 104 s in the case of 0.2 g/L DexC1010 aqueous solution at 25 ◦ C (Fig. 3). Surface tension decreased upon adsorption of an increasing number of hydrocarbon tails. According to our previous work, surface tension variations over that time interval and with those concentrations could not be attributed to diffusion limitations between bulk and interface. These variations were attributed to interfacial phenomena, either diffusion within the interfacial sublayer or conformational rearrangements, which could not be identified on the basis of these experiments. Over the same time interval, a continuous increase of dilational modulus was observed. This increase was caused by progressive structuration within the whole adsorbed layer (including both adsorbed units and polymer loops protruding into aqueous phase) which accompanied previously mentioned interfacial phenomena. At a given time, the surface tension almost did not vary strongly with DS value between 5 and 17% (Fig. 4). This indicated that, whatever the substitution degree, after 5000 s, the number of hydrocarbon chains per unit of area was approximately unchanged. On the contrary, after 5000 s, the dilational modulus varied significantly, reaching significantly higher value for DS = 10% (the reproducibility of the experiments had been checked, see Section 2). More precisely, the elastic modulus, E , was much higher for DS = 10% while the viscous modulus, E , was almost independent of DS (Table 1). Taking into account the limited number of DS values, we concluded that there was a range of DS values, between 7 and 17%, for which elastic modulus was significantly higher. In order to relate these observations to macromolecular parameters, the average distance between two hydrocarbon tails was calculated by estimating the length of one

Fig. 4. Surface tension and dilational modulus of 0.2 g/L DexC10DS aqueous solutions as a function of degree of substitution (DS) after 5000 s and at 25 ◦ C.

Table 1 Dilational moduli of air/water interfaces for 0.2 g/L aqueous of dextran derivatives at 25 ◦ C after 5000 s (the period of oscillations was 30 s). Polymer

lAU (Å)a

E (mN/m)

E (mN/m)

E” (mN/m)

DexC105 DexC107 DexC1010 DexC1017

90 66 45 24

11.1 12.4 40.7 8.0

8.7 10.7 40.2 7.0

6.9 6.2 6.3 4.0

a Estimated mean distance between hydrocarbon tails taking the length of anhydroglucose repeat units equal to 5 A˚ [14].

anhydroglucose unit to 5 A˚ [20]. The found values ranged between 90 and 24 A˚ when DS increased from 5 to 17% (Table 1). Additionally the persistence length of dextran T40® in aqueous solution has been determined to be 25 A˚ [21]. Thus, the elastic modulus E was significantly higher when the average distance between two successive hydrocarbon tails was comparable to the persistence length of polysaccharide. We must notice that the calculated values were only average distances between hydrocarbon tails. Nevertheless, since these polymers have been synthesized in dimethyl sulfoxide (see Section 2) where both dextran and 1,2-epoxydodecane were solubilized, we admitted that the distribution of alkyl substituents along dextran backbone was statistical and could be represented by a single average value [22,23]. The concentration used for those experiments (0.2 g/L) was lower than that corresponding to the minimum of surface tension (1 g/L or above) [24]. Consequently, macromolecules were expected to adopt a conformation at the interface such that the maximal number of hydrophobic tails were immersed into the air phase [25]. Increasing the degree of substitution allowed increasing the number of hydrophobic tails at the air-solution interface provided that the average distance between two successive hydrocarbon tails remained larger than the persistence length of the polymer. This structuration of the adsorbed layer gave rise to an increase of the elastic modulus since surface delation or compression affected an increased number of sticking hydrocarbon tails. When the average distance between two successive hydrocarbon tails became comparable or even lower than the persistence length of the polymer, some hydrocarbon tails were no longer in direct contact with the surface, by an increasing number of loops. Thus,

316

R. Covis et al. / Colloids and Surfaces A: Physicochem. Eng. Aspects 441 (2014) 312–318

upon dilation or compression of the surface, some could be transferred from the surface into hydrophobic aggregates within the adsorbed layer and vice versa, thus reducing the elasticity of the surface and consequently lowering E . A similar effect has been reported in the case of alkylated chitosans [26].

Table 2 Droplet diameter distribution of hexadecane-in-water emulsions stabilized with dextran derivatives. Measurements were carried out within 1 h following emulsion preparation. Stabilizer

Hexadecane (v/v)

D[3,2] (␮m)

D[4,3] (␮m)

Spana

DexC107

0.1 0.2 0.3 0.4 0.5

0.17 0.15 0.17 0.14 0.32

0.20 0.32 0.48 0.69 0.92

0.8 1.0 6.0 3.8 12.0

DexC1010

0.1 0.2 0.3 0.4 0.5

0.17 0.16 0.21 0.21 0.29

0.20 0.19 0.26 0.26 0.44

1.6 1.6 0.8 0.8 0.7

DexC1030

0.1 0.2 0.3

0.21 0.20 0.22

0.26 0.24 0.27

0.7 0.7 0.7

3.4. Oil-in-water submicronic emulsions 3.4.1. Design of experimental procedure for emulsion preparation Water-soluble amphiphilic dextrans have already been reported as efficient stabilizers for oil-in-water submicronic emulsions prepared with hydrocarbon oils. Our previous work was focused on emulsions with oil volume fractions lower than 20% and mainly around 10%. In that domain, with hydrocarbon oils like dodecane, we established that the main aging process was Ostwald ripening, at least within the first week following preparation. This aging process could be strongly limited by using oils with very low water solubilities like hexadecane [9,27,28]. In the present work, we examined the preparation and stability of submicronic oil-in-water emulsions having oil volume fractions between 10 and 50% using ultrasound emulsification procedure [29]. For all emulsions reported in that section, the weight ratio of amphiphilic dextran to hydrocarbon oil was set to 0.1. On the basis of previous results, this stabilizer-to-oil weight ratio was known as being convenient for obtaining submicronic oil droplets with fully covered surfaces [30]. We used ultra hydrophobic hydrocarbon oils, hexadecane and nujol, so as to suppress Ostwald ripening and thus assess the ability of amphiphilic dextrans to prevent coalescence or flocculation. In order to prevent any bacterial development during emulsion aging, the aqueous phase contained 10−3 g/L NaN3 and emulsions were stored at 4 ◦ C. We first designed the experimental procedure for preparing submicronic emulsions (see Section 2). We applied sonication steps to biphasic mixtures containing the required amounts of both liquid phases. Because of important viscosity increase (especially for oil volume fractions higher than 20%), four successive sonication steps (30 s each) were separated by vortexing steps (30 s each). Following that procedure, no excess oil was observed after the last sonication step. Having defined the procedure for emulsion preparation, we checked that polymeric stabilizers were not degraded by application of ultrasounds. Indeed, it had been reported that dextran in aqueous solution may be degraded when exposed to ultrasounds [31–34]. Aqueous solutions (20 g/L) of native dextran T40© and dextran derivatives, DexC105 and DexC107 , were submitted to ultrasounds during 120 s. Molar mass distributions of native dextran T40© and DexC105 were found unchanged, within experimental uncertainty, after ultrasound application. DexC107 was characterized by 1 H NMR in DMSOd6 before and after sonication. DS values of both samples were identical within experimental uncertainty. As a conclusion, dextran derivatives were not significantly degraded during emulsion preparation.

a The Span was calculated by (D(0.9) − D(0.1))/D(0.5) and was used as an indication of the broadness of droplet diameter distribution.

3.4.2. Distribution of droplet diameters and aging of oil-in-water emulsions Distributions of droplet diameters were determined by multi-angle light scattering for hexadecane-in-water emulsions stabilized with various dextran derivatives (Table 2). DexC1015 polymer led to experimental results very similar to DexC1010 (data not shown). Up to 20% hexadecane volume fraction, all dextran derivatives with DS between 7 and 30% led to similar results regarding average droplet diameters and broadness of size distribution. Submicronic emulsions were obtained. Emulsions prepared with DexC107 and oil volume fractions between 30 and 50%, had much wider size distributions, with droplets having diameters higher than 1 ␮m. This was attributed to a lack of anchoring points within polymer chains limiting their efficiency to prevent coalescence during emulsification. As for surface activity, within the explored range of DS no significant difference could be observed from one polymer to another (Fig. 4). Using the polymer with the highest DS, DexC1030 , it was not possible to prepare submicronic emulsions with oil volume fractions equal to 40 or 50%. Indeed, for those emulsions, the required polymer concentration in the aqueous phase was such that DexC1030 could not be dissolved completely. As a consequence very large droplet diameters distributions were obtained with diameters higher than 100 ␮m. These emulsions were not further considered in that work. These results evidenced that there was an “optimal” range of DS, roughly between 10 and 20%, for which submicronic hexadecanein-water emulsions were obtained. The limits of that “optimal” DS range corresponded to the requirement of having enough physical anchoring points with dextran backbone to provide steric repulsions preventing droplet coalescence (for the low DS limit) and to the necessity of maintaining high enough water solubility (for the high DS limit). Furthermore, we can notice that, within that DS

Table 3 Droplet diameter distribution of nujol-in-water emulsions stabilized with dextran derivatives after sonication and after freeze-drying and reconstitution (for details see Section 3.4.3). Stabilizer

DexC107

a

Nujol (v/v)

0.1 0.2 0.3 0.4 0.5

After sonication

After freeze-drying and reconstitution

D[3,2] (␮m)

D[4,3] (␮m)

Spana

D[3,2] (␮m)

D[4,3] (␮m)

Spana

0.79 0.81 0.77 0.68 0.39

1.66 1.70 1.60 1.36 0.70

2.3 2.3 2.3 2.2 2.1

1.25 1.29 0.98 1.60 0.29

12.88 13.87 7.88 8.84 4.09

11.1 6.9 7.4 3.6 3.6

The Span was calculated by (D(0.9) − D(0.1))/D(0.5) and was used as an indication of the broadness of droplet diameter distribution.

R. Covis et al. / Colloids and Surfaces A: Physicochem. Eng. Aspects 441 (2014) 312–318

displaced toward higher values after reconstitution. Nevertheless, dried samples were fully re-dispersed by simple stirring. Reconstituted emulsions contained no particles with diameters higher than 100 ␮m. It seemed that amphiphilic polysaccharide prevented droplet coalescence to a large extent. Another contribution came from non-adsorbed polymer chains which provided supplementary physical barriers between oil droplets in the dried state. Better results could be reasonably expected by selecting dextran derivatives with convenient DS, on the basis of results reported in Section 3.4.2. Further investigations are ongoing in the case of polymeric nanoparticles.

0.8 0.7 0.6

D[4,3] (µm)

317

0.5 0.4 0.3 0.2

4. Conclusion 0.1 0

10

20

30

40

50

60

Time (Days) Fig. 5. Volume-average droplet diameter of hexadecane-in-water emulsions as a function of storage time. The stabilizers were DexC107 with 50% v/v oil () and DexC1015 with 10% (䊉) and 50% () v/v oil.

range, dilational modulus of air/water interface was shown to reach a maximal value (see Section 3.3). In order to examine the effect of oil viscosity, hexadecane was replaced by nujol, following the same emulsification procedure (Table 3). When DexC107 was used as the stabilizer, emulsions prepared with nujol exhibited higher droplet diameters than those prepared with hexadecane. This result was explained by the higher viscosity of disperse phase [29]. The storage stability of emulsions was studied over several weeks by monitoring droplet diameter distributions versus storage time at 4 ◦ C (Fig. 5). For all emulsions, it was observed that droplet diameter distributions remained almost unchanged for both oils, i.e. hexadecane and nujol. Thus, oil droplets were conveniently covered by amphiphilic polysaccharides so as to prevent coalescence or flocculation during storage. Over the period of time where emulsion shelf life was checked, estimations of droplet size variation induced by Ostwald ripening showed that this contribution was negligible both at room temperature and at the storage temperature (4 ◦ C). As for coalescence phenomena, since the storage temperature was much below the melting point of hexadecane (18 ◦ C), crystals were formed in the storage conditions which may be a source of colloidal instability [35–37]. Nevertheless, our results demonstrated that the surface coverage of hexadecane droplets by dextran derivatives was efficient for preventing coalescence events. As for nujol oil, no crystallization was observed (with a 1 mL oil sample) in the storage conditions. 3.4.3. Lyophilization and re-dispersion of oil-in-water emulsions Freeze-drying of oil-in-water emulsions has been considered for many years as a very convenient method for limiting sample volume and possible degradations during storage or transport. Nevertheless one important limitation is the ability of stabilizers to ensure easy re-dispersion of previously dried emulsion samples as well as to limit droplet size variation. One usual way to prevent droplet coalescence during emulsion drying is to add a highly water soluble compound (often called “solid carrier”) into the continuous aqueous phase (carbohydrates like sucrose or glucose have been reported) [38,39]. Nujol-in-water emulsions stabilized by DexC107 were freeze dried and reconstituted by addition of water and magnetic stirring (Table 3). The case of hexadecane-in-water emulsions was not investigated. The solid content of emulsion samples recovered after lyophilization was, within experimental uncertainty, equal to the one calculated taking into account oil and polymeric stabilizer (data not shown). Droplet diameter distribution was

Hydrophobically modified dextran samples carrying hydrocarbon tails with 10 carbon atoms were studied as stabilizers for oil-in-water emulsions. Dilational rheology of air/water interfaces covered by these polymers was characterized using the pendant drop technique. These experiments revealed that elastic modulus reached a maximum for degrees of substitution between 10 and 17% which was interpreted by considering the conformation of adsorbed dextran chains. Submicronic oil-in-water emulsions of hexadecane were prepared with oil volume fractions between 10 and 50% and exhibited shelf lives longer than one month. Better results in terms narrowness of droplet diameter distributions were obtained with polymers having degrees of substitution between 10 and 20%. Replacing hexadecane by nujol induced a significant displacement of droplet diameter distribution toward higher sizes, which was attributed to the higher viscosity of oil. Nujol emulsions with oil contents between 10 and 50% v/v were freeze-dried and reconstituted by water addition and simple stirring. Although droplet diameter distributions were displaced toward higher values, they remained well below 100 ␮m. This work showed that nonionic dextran derivatives modified by C10 hydrocarbon chains behaved as efficient stabilizers of oil-inwater emulsions. Ongoing work deals with the use of amphiphilic dextrans for preparing micronic emulsions with mechanical processes and comparison with molecular surfactants. References [1] E. Dickinson, Food Hydrocoll. 17 (2003) 25–39. [2] E. Bouyer, G. Mekhloufi, V. Rosilio, J.-L. Grossiord, F. Agnely, Int. J. Pharm. 436 (2012) 359–378. [3] D. Dalgleish, Food Hydrocoll. 20 (2006) 415–422. [4] O.G. Jones, D.J. McClements, Compr. Rev. Food Sci. Food Saf. 9 (2010) 374–397. [5] C. Rouzes, R. Gref, M. Léonard, A. De Sousa Delgado, E. Dellacherie, J. Biomed. Mater. Res. 50 (2000) 557. [6] C. Fournier, M. Leonard, I. Le Coq-Leonard, E. Dellacherie, Langmuir 11 (1995) 2344. [7] E. Rotureau, C. Chassenieux, E. Dellacherie, A. Durand, Macromol. Chem. Phys. 206 (2005) 2038–2046. [8] R. Covis, C. Ladavière, J. Desbrières, E. Marie, A. Durand, Carbohydr. Polym. 95 (2013) 360–365. [9] E. Rotureau, M. Léonard, E. Marie, E. Dellacherie, T. Camesano, A. Durand, Colloids Surf., A 288 (2006) 62–70. [10] R. Nagarajan, D.T. Wasan, J. Colloid Interface Sci. 159 (1993) 164–173. [11] P. Saulnier, F. Boury, A. Malzert, B. Heurtault, T. Ivanova, A. Cagna, I. Panaïotov, J.E. Proust, Langmuir 17 (2001) 8104–8111. [12] R. Wustneck, P. Enders, T. Ebisch, R. Miller, Thin Solid Films 298 (1997) 39–46. [13] M. Léonard, E. Marie, M. Wu, E. Dellacherie, T. Camesano, A. Durand, ACS Symp. Ser. 996 (2008) 322–340. [14] A. Güner, J. Appl. Polym. Sci. 72 (1999) 871. [15] E. Catiker, A. Güner, Polym. Bull. 41 (1998) 223–230. [16] M. Vigouret, M. Rinaudo, J. Desbrières, J. Chim. Phys. 93 (1996) 858–869. [17] J. Desbrières, M. Hirrien, M. Rinaudo, ACS Symp. Ser. 688 (1998) 332–348. [18] R. Covis, C. Ladaviere, J. Desbrières, E. Marie, A. Durand, Colloids Surf., A 436 (2013) 744. [19] E. Rotureau, M. Léonard, E. Dellacherie, A. Durand, J. Colloid Interface Sci. 279 (2004) 68. [20] P.E. Marszalek, A.F. Oberhauser, Y.-P. Pang, J.M. Fernandez, Nature 396 (1998) 661–664. [21] S.K. Garg, S.S. Stivala, J. Polym. Sci. Polym. Phys. Ed. 16 (1978) 1419–1434. [22] B. Magny, F. Lafuma, I. Iliopoulos, Polymer 33 (1992) 3151–3154.

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R. Covis et al. / Colloids and Surfaces A: Physicochem. Eng. Aspects 441 (2014) 312–318

[23] M. Hirrien, C. Chevillard, J. Desbrières, M. Axelos, M. Rinaudo, Polymer 39 (1998) 6251–6259. [24] C. Rouzes, M. Léonard, A. Durand, E. Dellacherie, Colloids Surf., B 32 (2003) 125–135. [25] V.G. Babak, J. Desbrières, V.E. Tikhonov, Colloids Surf., A 255 (2005) 119–130. [26] V.G. Babak, J. Desbrières, Colloid Polym. Sci. 284 (2006) 745–754. [27] A. Durand, E. Marie, E. Rotureau, M. Léonard, E. Dellacherie, Langmuir 20 (2004) 6956–6963. [28] V. Sadtler, P. Imbert, E. Dellacherie, J. Colloid Interface Sci. 254 (2002) 355. [29] J.P. Canselier, H. Delmas, A.M. Wilhelm, B. Abismaïl, J. Disp. Sci. Technol. 23 (2002) 333–349. [30] E. Rotureau, M. Léonard, E. Marie, E. Dellacherie, T.A. Camesano, A. Durand, Colloids Surf., A 288 (2006) 131–137. [31] J.P. Lorimer, T.J. Mason, T.C. Cuthbert, E.A. Brookfield, Ultrason. Sonochem. 2 (1995) S55–S57.

[32] G.L. Cote, J.L. Willet, Carbohydr. Polym. 39 (1999) 119–126. [33] G. Portenlänger, H. Heusinger, Ultrason. Sonochem. 4 (1997) 127–130. [34] Q. Zou, Y. Pu, Z. Han, N. Fu, S. Li, M. Liu, L. Huang, A. Lu, J. Mo, S. Chen, Carbohydr. Polym. 90 (2012) 447–451. [35] E. Fredrick, P. Walstra, K. Dewettinck, Adv. Colloid Interface Sci. 153 (2010) 30–42. [36] F. Leal-Calderon, F. Thivilliers, V. Schmitt, Curr. Opin. Colloid Interface Sci. 12 (2007) 206–212. [37] F. Thivilliers-Arvis, E. Laurichesse, V. Schmitt, F. Leal-Calderon, Langmuir 26 (2010) 16782–16790. [38] J. Beirowski, S. Inghelbrecht, A. Arien, H. Gieseler, J. Pharm. Sci. 100 (2011) 1958–1968. [39] P. Shahgaldian, J. Gualbert, K. Aïssa, A.W. Coleman, Eur. J. Pharm. Biopharm. 55 (2003) 181–184.