FR3061033A1 - PROCESS FOR THE PREPARATION OF SOLUTE NANOCAPSULES AND USE THEREOF - Google Patents

Abstract

The present invention relates to a process for preparing nanocapsules of solute S from a first solvent S1, a second solvent S2, and a crosslinkable polymer PCL, the solvent S1 being water, said process comprising the steps following: a / a phase diagram P1 of the solute S in the mixture of solvents [S1 + S2] is established, the solute S being miscible in the solvent S2 and immiscible in the solvent S1, the diagram P1 comprising two curves defining three domains D1 to D3, b / we establish a phase diagram P2 of the crosslinkable polymer PCL in the mixture of solvents [S1 + S2], PCL being soluble in S1 and insoluble in S2, the diagram P2 comprising a cloud point curve defining two domains D4 and D5, c / a domain corresponding to the overlap of domains D1 and D5 is defined, d / the following are prepared separately: - an E phase comprising a crosslinkable polymer PCL and a solvent S1, and - an H phase comprising: * a solvent S2, * a crosslinking agent, and * a solute S, e / a mixture M is formed by combining phase E and phase H, - the mixture M being in the overlap of domains D1 and D5, f / on forms nanocapsules of solute S in the crosslinkable polymer PCL, by precipitation and crosslinking of PCL.

Description

® Agent (s): BECKER & ASSOCIES.

® PROCESS FOR THE PREPARATION OF NANOCAPSULES OF SOLUTE AND THEIR USE.

The present invention relates to a process for preparing nanocapsules of solute S from a first solvent S1, a second solvent S2, and a crosslinkable polymer P ^CL , the solvent S1 being water, said process

FR 3,061,033 - A1 comprising the following steps:

a / a phase diagram P1 of the solute S in the mixture of solvents [S1 + S2] is established, the solute S being miscible in the solvent S2 and immiscible in the solvent S1, the diagram P1 comprising two curves defining three domains D1 to D3, b / a phase diagram P2 of the crosslinkable polymer P ^{CL is established} in the mixture of solvents [S1 + S2], P ^CL being soluble in S1 and insoluble in S2, the diagram P2 comprising a cloud point curve defining two domains D4 and D5, c / we define a domain corresponding to the overlap of domains D1 and D5, d / we prepare separately:

a phase E comprising a crosslinkable polymer P ^CL and a solvent S1, and

- a phase H comprising:

* a solvent S2, * a crosslinking agent, and * a solute S, e / a mixture M is formed by combining phase E and phase H,

- the mixture M being in the overlap of the domains D1 and D5, f / nanocapsules of solute S are formed in the crosslinkable polymer

P ^CL , by precipitation and crosslinking of P ^CL .

Mass fraction Si '[Sl + S2]

PROCESS FOR THE PREPARATION OF SOLUTE NANOCAPSULES AND THEIR USE

FIELD OF THE INVENTION

The present invention relates to a process for the preparation of nanocapsules of the heart / shell type, the heart and the shell being respectively a solute and a polymer.

These nanocapsules of solute can be used in particular in cosmetics, in vectorization applications, for example pharmaceuticals and therapeutics.

PRIOR STATE OF THE ART

The encapsulation of chemical compounds is of particular interest in applications requiring controlled release. For example, the encapsulation of a catalyst or a drug can allow its release at the appropriate time.

In general, encapsulation consists in trapping a chemical compound, the solute, within a bark such as a polymer matrix.

The solute is released after decomposition or rupture of, or by diffusion through, the polymer matrix. Thus, by adapting the nature of the latter, it is possible to control the conditions of release of the solute. It can be a porous matrix or a waterproof matrix, respectively.

The nanoprecipitation process makes it possible to encapsulate a solute by the Ouzo effect (Yan et al., Angew. Chem. Int. Ed., 2014, 53, 6910-6913).

This process combines the following phenomena:

the formation of a solute emulsion when a solvent is added to a solution of solute in a co-solvent;

precipitation of a polymer as a function of the mass fraction of solvent and co-solvent.

To obtain the Ouzo effect, the solvent and the co-solvent are miscible while the solute is insoluble in the solvent. In addition, to achieve the encapsulation of the droplets of solute thus formed, the polymer is soluble in the solvent and insoluble in the co-solvent.

Thus, when the conditions are met, the solute can be encapsulated by the polymer.

However, the Ouzo process does not allow capsules of easily controllable sizes to be obtained. In addition, the range of sizes available is limited, and mainly greater than 100 nm.

There is therefore a need to overcome, at least partially, the aforementioned drawbacks.

STATEMENT OF THE INVENTION

The Applicants have developed a process which makes it possible not only to prepare nanometric size capsules but also to predefine the size of these capsules. In addition, the method according to the invention proves to be relatively reliable and easy to implement on an industrial scale.

The present invention makes it possible to prepare nanocapsules of solute. This is a particularly interesting process in view of its ease of implementation and its versatility.

This process allows in particular to predefine the size of the capsules according to the nature of the solute and of the solvents used.

The process according to the invention also makes it possible to work in a larger area than the Ouzo area and therefore, with a wider range of hydrophilic polymers.

More specifically, the present invention relates to a process for preparing nanocapsules of solute S from a first solvent SI, from a second solvent S2, and from a crosslinkable polymer P, the solvent SI being water, said process including the following steps:

a / a phase diagram PI of the solute S in the mixture of solvents [S1 + S2] is established, the solute S being miscible in the solvent S2 and immiscible in the solvent SI,

- the PI diagram being established for the mass fraction S2 / [S 1 + S2] as a function of the mass fraction S / [S1 + S2],

- the PI diagram comprising two curves defining three domains DI to D3:

* a homogeneous domain D1, * a heterogeneous domain D2 in which the solute S is immiscible in the mixture [S1 + S2], * a heterogeneous domain D3 in which the solute S is in the form of an oil-in-water emulsion in the mixture [S1 + S2], the two curves of the PI diagram being respectively a binodal curve separating the homogeneous domain DI from the heterogeneous domains D2 and D3; and a curve separating the D2 domain from the D3 domain; these two curves having an intersection point N corresponding to the intersection of the domains Dl, D2 and D3,

CT b / a phase diagram P2 of the crosslinkable polymer P is established in the mixture of solvents [S1 + S2], P being soluble in SI and insoluble in S2,

- the diagram P2 being established for the mass fraction S2 / [S 1 + S2] as a function of the mass fraction P ^CL / [S 1 + S2],

- the diagram P2 comprising a cloud point curve defining two domains D4 and D5:

CT * a D4 domain in which P is soluble in the mixture [S 1 + S2], and * a D5 domain in which P precipitates in the mixture [S1 + S2], c / we define a domain corresponding to the overlap of the Dl domains and D5, d / we prepare separately:

a phase E comprising a crosslinkable polymer P and a solvent SI, and

- a phase H comprising:

* a solvent S2, * a crosslinking agent, and * a solute S, e / a mixture M is formed by combining phase E and phase H,

the mixture M being in the overlap of the domains Dl and D5,

the mass fraction P / [S1 + S2] of the mixture M being in the domain D5,

the mass fraction S / [S 1 + S2] of the mixture M being in the domain Dl,

- the mass fraction S2 / [S1 + S2] of the mixture M being in the domains DI and

D5,

CT f / nanocapsules of solute S are formed in the crosslinkable polymer P, by

CT precipitation and crosslinking of P

The point N corresponds to a mass fraction N _x of solute S / [S1 + S2] and to a mass fraction N _y of solvent S2 / [S 1 + S2].

The mixture M is located in the overlap of the domains DI and D5. It is therefore outside the D6 domain.

In the absence of crosslinkable polymer P and of crosslinking agent, the mixture M corresponds to a thermodynamically stable microemulsion, which is not the case for emulsions from the Ouzo domain, D3.

According to a particular embodiment, the mass fraction S / [S1 + S2] of the mixture M is in the domain DI and less than N _x while the mass fraction S2 / [S 1 + S2] of the mixture M is in the domains DI and D5 and less than N _y . This embodiment makes it possible to obtain nanocapsules whose average size is less than 100 nanometers.

In addition, the size of the nanocapsules is easily controlled by varying the ratio S2 / [S1 + S2] so as to reach diameters which, advantageously, range from 40 to 200 nm. This size corresponds to the size measured by TEM (transmission electron microscopy), that is to say on dried nanocapsules. Unless otherwise indicated, the same applies to the rest of the description.

The D3 domain of the PI phase diagram corresponds to the Ouzo domain in which the solute droplets formed have a size that is difficult to control. The emulsion formed in this area is not thermodynamically stable if no surfactant is used.

Solute S is advantageously an active principle. It can in particular be chosen from the group comprising: triglycerides, for example Miglyol®; drugs such as doxorubicin; essential oils ; the proteins ; photoluminescent markers; dyes; the perfumes ; fertilizers ; pesticides ; food supplements; the aromas ; and cosmetics.

The solvent SI is water. According to a particular embodiment, it can also be any aqueous solution, as well as water / solvent mixtures in which the hydrophilic polymer is soluble.

The solvents SI and S2 are miscible.

The solvent S2 is advantageously chosen from the group comprising: ketones, for example acetone; ethers, for example tetrahydrofuran; dimethyl sulfoxide (DMSO); alcohols, for example ethanol, methanol, isopropanol; N-methyl-2-pyrrolidone (NMP); and Ν, Ν-dimethylformamide (DMF). In general, any solvent perfectly miscible with water, under certain temperature and pressure conditions (conditions for implementing the process), can be used.

According to a preferred embodiment, the solvent S1 is water while the solvent S2 is acetone.

The crosslinkable polymer P is not miscible in the solvent S2. In other words, it precipitates when a polymer solution in the solvent SI is brought into contact with a certain amount of solvent S2. The P2 diagram makes it possible to determine this

CT quantity as a function of the concentration of crosslinkable polymer P

The crosslinkable polymer P is soluble in the solvent SI. Thus, it is water-soluble (SI = water) under normal conditions of use according to the invention.

The crosslinkable polymer P is advantageously a water-soluble synthetic polymer, a polysaccharide or a hydrophilic polypeptide.

By way of nonlimiting example, it can in particular be chosen from the group comprising: poly (N- [2- (oc-D-mannopyranosyloxy) ethyl] methacrylamide) (PEMM); poly (N- [2- (2-ethoxy) ethyl (α-D-glucopyranoside)] methacrylamide) (PEEGM); dextran (branched dextrose polymer); poly (N- (2-hydroxypropyl) methacrylamide) (PHPMA); polyvinyl alcohol (PVA); hyaluronic acid ; glycogen; pullulan (polysaccharide); and dextran derivatives, for example dextran sulfate.

In general, the crosslinkable polymer P comprises OH groups which allow crosslinking in the presence of an adequate crosslinking agent.

CT

In addition, according to a particular embodiment, the crosslinkable polymer P can comprise at least one functionalizing group. This group can in particular be chosen from the group comprising: biotin; phosphors, for example a fluorophore; metals ; nanotracer; biomarker; magnetic nanoparticles; and their mixtures.

The formation of nanocapsules is ensured by the precipitation of P on the solute, and the

CT crosslinking of P in the presence of a crosslinking agent.

The crosslinking agent comprises at least two functions capable of reacting with the polymer P. It can in particular be a compound comprising at least two isocyanate functions -N = C = O. It can also be a diepoxide, a dialdehyde or any agent capable of reacting with alcohol functions.

By way of nonlimiting example, this crosslinking agent can be advantageously chosen from the group comprising: diisocyanates, for example isophorone diisocyanate (IPDI); diepoxides, for example Bisphenol A diglycidyl ether; dialdehydes, for example glutaraldehyde; and their mixtures.

In order to avoid crosslinking of P before it is brought into contact with the solute S and when it precipitates, the crosslinking agent is present in phase E. It is therefore advantageously soluble in the solvent S2. It can also be soluble in the solvent SI. Phase H is therefore advantageously devoid of crosslinking agent of the pCL polymer

As already indicated, the method according to the invention implements the following steps: a / the PI phase diagram is established;

b / the phase diagram P2 is established;

c / the domain corresponding to the overlap of the precipitation domain of P (D5) and the homogeneous domain of S (DI) is defined in [S1 + S2];

d / a phase E and a phase H are prepared separately;

e / a mixture M is formed by combining phase E and phase H;

CT f / nanocapsules of solute S are formed in the crosslinkable polymer P

The PI phase diagram of the solute S in the solvent mixture [S1 + S2] is established in a conventional manner. More precisely, this type of diagram can be obtained according to the following two steps:

1 / the mass fraction S2 / [S 1 + S2] is varied by adding solvent SI to a solution of solute S in solvent S2, at constant S concentration; the presence of a homogeneous phase (Dl), an emulsion (D3) or a heterogeneous phase (D2) is noted;

2 / we repeat step 1 / at a different solute concentration S.

Thus, we obtain the PI diagram which includes two curves defining the three domains D1, D2 and D3 as illustrated by FIG. 1.

The domain Dl corresponds to a solution of solute S in [S1 + S2], This is a domain having a homogeneous phase corresponding to a microemulsion of solute S in the mixture [S1 + S2], The domain Dl is said "Transparent" because the microemulsion formed is invisible to the naked eye. This is the reason why this domain has generally been considered to correspond to a single-phase domain in which the solute S was completely solubilized. However, the Applicants have shown for oily solute / S2 solvent / SI solvent (water) systems that this specific domain corresponds to a microemulsion, the solute S not being miscible (solubilized) in the mixture [S1 + S2] of the domain Dl. Unlike the D3 domain emulsion, this microemulsion is thermodynamically stable even in the absence of surfactant. This microemulsion is stable over time and has a monodisperse size. The Applicants have moreover discovered that, in the domain D1, the microemulsion formed has an identical droplet size for a given S1 / S2 ratio, regardless of the concentration of solute S.

The domain D2 corresponds to a heterogeneous mixture of solute S in the solvents [S1 + S2]. This domain is said to be "opaque" or "cloudy" because the solute S is neither soluble nor emulsifiable in this domain.

The heterogeneous domain D3 corresponds to an oil in water emulsion of solute S in the mixture of solvents [S1 + S2], As already indicated, it is the Ouzo domain.

In this domain, the size of the droplets of solute S forming the emulsion varies greatly as a function of a small variation in the concentration of solute S in the mixture [S1 + S2]. The emulsion formed does not correspond to the microemulsion of the domain Dl. Those skilled in the art have always clearly distinguished these two areas, in particular because of the transparency of the Dl domain.

Step b / of the process which is the subject of the invention consists in establishing the phase diagram P2 of the crosslinkable polymer P in the mixture of solvents [S1 + S2], P being soluble in SI and insoluble in S2. For this, it is possible to follow the following steps: 1 / the mass fraction S2 / [S 1 + S2] is varied by adding solvent S2 in a solution of crosslinkable polymer P in the solvent SI, at constant P concentration ; the mass fraction S2 / [S1 + S2] corresponding to the precipitation of the crosslinkable polymer P is noted;

CT

2 / step 1 / is repeated at a different concentration of crosslinkable polymer P.

Thus, we obtain the diagram P2 which includes a cloud point curve of the crosslinkable polymer P defining the two domains D4 and D5 as illustrated in FIG. 2.

The cloud point curve separating the domains D4 and D5 corresponds to the appearance of a cloudiness when the solvent S2 is added in a solution of polymer P in water. This curve represents the solubility / precipitation limit of the polymer P ^CL in [S1 + S2],

The domain D4 corresponds to a solution of P in [S1 + S2],

CT

The domain D5 corresponds to a suspension of P in [S1 + S2],

The superimposition of the PI and P2 phase diagrams makes it possible to define the domain D6 corresponding to the overlap of the precipitation domains of P (D5) and of emulsion of S (D3) in [S1 + S2] as illustrated in FIG. 3.

The D6 domain makes it possible to obtain capsules which have a size generally greater than 100 nm and which is difficult to control.

More precisely, the superposition of the phase diagrams PI and P2 makes it possible to define the domain of interest corresponding to the superposition of the domains DI and D5.

The area of interest (D1 + D5) also has the advantage of being more important than the area D6 (D3 + D5) of the prior art. It therefore offers a more substantial work area.

Once the conditions for obtaining the area of interest defined (D1 + D5), the CT phases

E (Sl + P) and H (S2 + S + crosslinker) are prepared independently of each other.

These two phases are then mixed by introducing phase E into phase H or by introducing phase H into phase E to form the mixture M.

CT

The mixture M comprises droplets of solute S covered with P

According to a particular embodiment, phase H can also include a portion of solvent S1.

In this particular case, the quantities of the solvents SI and S2 are adjusted later

CT by addition of polymer P and / or of the crosslinking agent.

The mixture M resulting from the addition of phases E and H is advantageously produced in the absence of surfactant. Thus, the process according to the invention is preferably carried out in the absence of a surfactant.

In phase E, the concentration of P is advantageously between 10 'and 10' ³ g / g, more advantageously between 10 ' ⁴ and 5/10' ⁴ g / g, this concentration being expressed in grams of P per gram of phase E.

In phase H, the concentration of solute S is advantageously between 10 ' ⁸ and 10' ² g / g, more advantageously between 10 ' ⁶ and 10' ³ g / g, this concentration being expressed in grams of solute S per gram of phase H.

The quantity of crosslinking agent used is advantageously between 1 and 128 eq / mol polymer P, more advantageously between 24 and 48 eq / mol polymer P, this quantity being expressed in molar equivalent of crosslinking agent per mole of polymer pCL

The mixture M has a mass fraction S2 / [S 1 + S2] advantageously between 0.2 and 0.8, more advantageously between 0.4 and 0.6, depending on the size of the objects concerned.

(~ Ί

Still in the mixture M, the mass ratio P / solute S is advantageously between 0.2 / 1 and 100/1, more advantageously between 1/1 and 10/1, depending on the thickness of the membrane targeted, that is to say -display depending on the thickness of crosslinked polymer surrounding the solute.

(~ Ί

As already indicated, the nanocapsules of solute S in the crosslinkable polymer P (~ Ί are formed by precipitation and by crosslinking of P

The crosslinking of P can be carried out within a period advantageously between 10 minutes and 24 hours, more advantageously between 2 hours and 12 hours.

The crosslinking of P is advantageously carried out between 10 ° C and 50 ° C, more advantageously at room temperature.

Advantageously, the whole process (steps a / to f /) is carried out at room temperature.

The present invention also relates to the nanocapsules obtained by this process.

The size of these nanocapsules can in particular be between 10 nanometers and 300 nanometers, more advantageously between 40 nanometers and 200 nanometers.

The nanocapsules being advantageously spherical, the size advantageously corresponds to their diameter.

As already indicated, these are nanocapsules of nanocapsules of the heart / shell (or heart / shell) type, the heart and the shell being respectively a solute and a polymer. The thickness of the bark is advantageously between 5 and 30 nanometers, more advantageously between 10 and 20 nanometers.

The method according to the invention makes it possible to predefine the size of the nanocapsules as a function of the S2 / S1 ratio and the amount of solute S. It also makes it possible to obtain a dispersion of relatively narrow size.

To predetermine the size of the nanocapsules, the size of the droplets of solute S in the mixture [S1 + S2] is measured as a function of the mass fraction S2 / [S1 + S2] and of the concentration of solute S, and this in the field DI of diagram Pl.

As already indicated, it is possible to obtain nanocapsules whose size is less than 100 nanometers when the mass fraction S / [S 1 + S2] of the mixture M is in the DI domain and less than N _x and when the mass fraction S2 / [S 1 + S2] of the mixture M is in the domains DI and D5 and less than N _y .

On the other hand, the method according to the invention is particularly versatile since it allows to encapsulate any kind of solute. By adapting the nature of P, it is possible to obtain nanocapsules whose bark is porous or not. The presence of a porous bark can make it possible, if necessary, to remove any trace of solvent SI or S2 by drying the nanocapsules at a temperature which does not allow the solute to evaporate.

The field of use of these nanocapsules depends on the encapsulated solute. Thus, they can in particular be used in the treatment of cancer.

Other possible fields of application of these nanocapsules include in particular cosmetics, vectorization applications, for example pharmaceuticals and therapeutics. They also include the food industry, the marking of objects, agriculture and the dissemination of aromas or perfumes.

Thanks to the present invention, a person skilled in the art will be able to prepare nanocapsules of predefined size. Thus, it can adapt the size to the intended area of use.

The present invention and the advantages which result therefrom will emerge more clearly from the following figures and examples given in order to illustrate the invention and not in a limiting manner.

DESCRIPTION OF THE FIGURES

Figure 1 illustrates the PI phase diagram of a system consisting of a solute S in a mixture of two solvents [S1 + S2] miscible with each other.

FIG. 2 illustrates the phase diagram P2 of a system consisting of a crosslinkable polymer P in a mixture of two solvents [S1 + S2] miscible with each other.

Figure 3 corresponds to the superposition of the PI phase diagram of a system consisting of a solute S in a mixture of two solvents [S1 + S2] miscible with each other

CT and phase diagram P2 of a system consisting of a crosslinkable polymer P in the same mixture of two solvents [S1 + S2] miscible with each other.

EXAMPLES OF EMBODIMENT OF THE INVENTION

The phase diagram of FIG. 3 is obtained by superimposing the phase diagram S / [S 1 + S2] (FIG. 1) and the phase diagram P ^CL / [S 1 + S2] (FIG. 2).

The solutes S were encapsulated with the polymers P under the conditions described in Table 1 below.

The encapsulation was carried out from an aqueous solution of crosslinkable polymer CT

P and a solution of solute S and crosslinker in acetone.

CT

The encapsulation conditions are defined by the S / P / S1 / S2 phase diagram with:

S = miglyol, hexadecane;

P = PEMM (poly (N- [2- (oc-D-mannopyranosyloxy) ethyl] methacrylamide), PHPMA (poly (N- (2-hydroxypropyl) methacrylamide)), PEEGM (poly (N- [2- (2-ethoxy ) ethyl (α-D-glucopyranoside)] methacrylamide), glycogen, hyaluronic acid, pullulan or dextran;

= water;

= acetone.

This phase diagram (Figure 3) is obtained by superimposing the phase diagram S / [S 1 + S2] (Figure 1) and the phase diagram P ^CL / [S 1 + S2] (Figure 2).

1 / Determination of the size of the droplets of solute S in the acetone / water mixture

Prior to the preparation of solute S capsules, the size of the solute S droplets (miglyol 812, CAS number 37332-31-3) in the acetone / water mixture was determined as a function of the acetone / water mass fraction (S2 / [S 1 + S 2] = 0.2 to 0.8) and the concentration of solute S (S / [S 1 + S 2] = 10 ^{'11-10' 3).}

The size of the droplets is determined by dynamic light scattering (DLS) or by TEM (transmission electron microscopy).

In particular, droplets of size less than 100 nm are obtained when the acetone / water mass fraction (S2 / [S1 + S2]) is between 0.4 and 0.6:

with an acetone / water mass fraction (S2 / [S1 + S2]) of 0.6, the droplets have a monodisperse size distribution of 75-90 nm (dy / dN <1.25);

with an acetone / water mass fraction (S2 / [S1 + S2]) of 0.5, the droplets have a monodisperse size distribution of 60-75 nm (dy / dN <1.2);

with an acetone / water mass fraction (S2 / [S1 + S2]) of 0.4, the droplets have a monodisperse size distribution of 50 nm (dy / dN <1.15).

This experiment makes it possible to determine the conditions suitable for carrying out the process according to the invention, namely the acetone / water mass fraction (S2 / [S 1 + S2]).

2 / Preparation of miglyol nanocapsules according to the invention, examples were carried out (at room temperature and in the presence of oxygen) according to the following protocol corresponding to Example 1:

preparing a phase E containing 0.5 mg of crosslinkable polymer P and 600 mg of water (SI);

preparing a phase H containing 10 ' ³ mg of solute S, 10' ³ mg of crosslinking agent (Ret) and 400 mg of acetone (S2);

phase E is poured into phase H all at once.

The E + H mixture thus obtained is transparent or slightly milky, reflecting the presence of nanocapsules.

After crosslinking of the crosslinkable polymer P for 12 hours, the mixture E + H is cloudy. The size of the nanocapsules thus obtained is then measured by TEM or by DLS (measurement after isolation of the nanocapsules by filtration).

Table 1: Experimental conditions and properties of nanocapsules according to the invention.

Example Polymer (XP ^CL , g) Crosslinking (XRet, g) Solute (S / [S1 + S2], g) Mass reaction acetone / water (S2 / [S1 + S2]) Nanocapsule size (nm) 1 PEMM (5.10 ' ⁴ ) IPDI (10- ⁶⁾ MIGLYOL (10- ⁶⁾ 0.4 37 (TEM) 47 (DLS) 2 PEMM (5.10 ' ⁴ ) IPDI (10- ⁶⁾ MIGLYOL (10- ⁶⁾ 0.45 41 (TEM) 52 (DLS) 3 PEMM (5.10 ' ⁴ ) IPDI (10- ⁶⁾ MIGLYOL (10- ⁶⁾ 0.5 45 (TEM) 58 (DLS) 4 PEMM (5.10 ' ⁴ ) IPDI (10- ⁶⁾ MIGLYOL (10- ⁵⁾ 0.5 47 (TEM) 63 (DLS) 5 PEMM (5.10 ' ⁴ ) IPDI (10- ⁶⁾ MIGLYOL (10- ⁴ ) 0.6 71 (TEM) 85 (DLS) 6 PEMM (5.10 ' ⁴ ) IPDI (10- ⁶⁾ MIGLYOL (10- ⁴ ) 0.7 113 (TEM) 130 (DLS) 7 PEMM (5.10 ' ⁴ ) IPDI (10- ⁶⁾ MIGLYOL (10- ⁴ ) 0.8 187 (TEM) 190 (DLS) 8 dextran (5.10 ' ⁴ ) IPDI (10- ⁶⁾ MIGLYOL (10- ⁵⁾ 0.5 60 (TEM) 82 (DLS) 9 dextran (5.10 ' ⁴ ) IPDI (10- ⁶⁾ MIGLYOL (10- ⁴ ) 0.6 60 (TEM) 136 (DLS) 10 PHPMA (1.10 ' ⁴ ) IPDI (10- ⁶⁾ Hexadecane (W ⁴ ) 0.65 85 (TEM) 91 (DLS) 11 PHPMA (1.10 ' ⁴ ) IPDI (10- ⁶⁾ Hexadecane (W ⁴ ) 0.75 150 (TEM) 155 (DLS) 12 PHPMA (1.10 ' ⁴ ) IPDI (10- ⁶⁾ Hexadecane (W ⁴ ) 0.8 187 (TEM) 196 (DLS) 13 PEMM (1.10 ' ⁴ ) IPDI (10- ⁶⁾ Hexadecane (W ⁴ ) 0.5 50 (TEM) 67 (DLS) 14 PEMM (1.10 ' ⁴ ) IPDI (10- ⁶⁾ Hexadecane (W ⁴ ) 0.6 72 (TEM) 85 (DLS) 15 PEMM (1.10 ' ⁴ ) IPDI (10- ⁶⁾ Hexadecane (W ⁴ ) 0.7 125 (TEM) 139 (DLS)

16 PEEGM (1.10 ^-5 ) IPDI (ΙΟ ⁶ ) MIGLYOL (10- ⁵⁾ 0.5 43 (TEM) 60 (DLS) 17 PEEGM (1.10 ' ⁴ ) IPDI (ΙΟ ⁶ ) MIGLYOL (10- ⁴ ) 0.6 70 (TEM) 78 (DLS) 18 PEEGM (1.10 ' ⁴ ) IPDI (ΙΟ ⁶ ) MIGLYOL (10- ⁴ ) 0.7 103 (TEM) 138 (DLS) 19 Glycogen (1.10 ' ⁴ ) IPDI (ΙΟ ⁶ ) MIGLYOL (10- ⁵⁾ 0.5 62 (DLS) 20 Hyaluronic acid (1.10 ' ⁴ ) IPDI (ΙΟ ⁶ ) MIGLYOL (10- ⁴ ) 0.6 67 (DLS) 21 Pullulan (1.10 ' ⁴ ) IPDI (ΙΟ ⁶ ) MIGLYOL (10- ⁴ ) 0.6 84 (DLS)

XP ^CL = crosslinkable polymer mass fraction P ^CL / [S 1 + S2]

X Ret = mass fraction crosslinking agent / [Sl + S2]

The size of the nanocapsules varies depending on the measurement method used (DLS or TEM). Indeed, the nanocapsules studied by TEM are dried before the analysis. In DLS, a measurement carried out directly in emulsion, the polymer can swell with water, thus accentuating the difference in size with the measurement of TEM.

This difference in size is nothing unusual or abnormal, and is part of the knowledge of a person skilled in the art. Furthermore, the size of the polymer membrane swollen with water can be modulated according to the crosslinking density.

Examples 1 to 21 show that when leaving the conventional so-called "Ouzo" zone corresponding to the formation of solute / polymer capsules, it is possible to obtain capsules of size less than 100 nm. The Applicants have thus developed a process making it possible to control the size of the nanocapsules by predefining the appropriate mass fractions and the thickness and the crosslinking density of the membrane.

In Examples 1 to 21, the mass fraction S / [S1 + S2] is in the domain DI and less than N _x while the mass fraction S2 / [S 1 + S2] of the mixture M is in the domains DI and D5 and less than N _y . As already specified, the point N corresponds to the intersection of the domains Dl, D2 and D3. It corresponds to a mass fraction N _x of solute S / [S 1 + S2] and to a mass fraction N _y of solvent S2 / [S 1 + S2].

Claims (10)

1. A process for preparing nanocapsules of solute S from a first solvent SI, a second solvent S2, and a crosslinkable polymer P, the solvent SI being water, said process comprising the following steps: a / a phase diagram PI of the solute S in the mixture of solvents [S1 + S2] is established, the solute S being miscible in the solvent S2 and immiscible in the solvent SI, - the PI diagram being established for the mass fraction S2 / [S 1 + S2] as a function of the mass fraction S / [S1 + S2], - the PI diagram comprising two curves defining three domains DI to D3: * a homogeneous domain D1, * a heterogeneous domain D2 in which the solute S is immiscible in the mixture [S1 + S2], * a heterogeneous domain D3 in which the solute S is in the form of an oil-in-water emulsion in the mixture [S1 + S2], the two curves of the PI diagram being respectively a binodal curve separating the homogeneous domain DI from the heterogeneous domains D2 and D3; and a curve separating the D2 domain from the D3 domain; these two curves having an intersection point N corresponding to the intersection of the domains Dl, D2 and D3, CT b / a phase diagram P2 of the crosslinkable polymer P is established in the mixture of solvents [S1 + S2], P being soluble in SI and insoluble in S2, - the diagram P2 being established for the mass fraction S2 / [S 1 + S2] as a function of the mass fraction P ^CL / [S 1 + S2], - the diagram P2 comprising a cloud point curve defining two domains D4 and D5: CT * a D4 domain in which P is soluble in the mixture [S 1 + S2], and * a D5 domain in which P precipitates in the mixture [S1 + S2], c / we define a domain corresponding to the overlap of the Dl domains and D5, d / we prepare separately: - A phase E comprising a crosslinkable polymer P and a solvent SI; and - a phase H comprising: * a solvent S2, * a crosslinking agent, and * a solute S, e / a mixture M is formed by combining phase E and phase H, - the mixture M being in the overlap of the domains DI and D5, the mass fraction P / [S1 + S2] of the mixture M being in the domain D5, the mass fraction S / [S 1 + S2] of the mixture M being in the domain Dl, the mass fraction S2 / [S1 + S2] of the mixture M being in the domains Dl and D5, CT f / nanocapsules of solute S are formed in the crosslinkable polymer P, by CT precipitation and crosslinking of P 2. Method according to claim 1, characterized in that the method is carried out in the absence of surfactant. 3. Method according to claim 1 or 2, characterized in that the solvent S2 is chosen from the group comprising: acetone; tetrahydrofuran; dimethyl sulfoxide; ethanol; methanol; isopropanol; N-methyl-2-pyrrolidone; and N, N-dimethylformamide. 4. Method according to one of claims 1 to 3, characterized in that the polymer Crosslinkable CT P is a water-soluble synthetic polymer, a polysaccharide or a hydrophilic polypeptide. 5. Method according to one of claims 1 to 4, characterized in that the crosslinkable polymer P comprises at least one functionalizing group chosen from the group comprising: biotin; phosphors; metals ; nanotracer; biomarker; magnetic nanoparticles; and their mixtures. 6. Method according to one of claims 1 to 5, characterized in that the crosslinking agent is chosen from the group comprising diisocyanates; diepoxides; dialdehydes; and their mixtures. 7. Method according to one of claims 1 to 6, characterized in that the concentration of solute S in phase H is between 10 ' ⁸ and 10' ² g / g, this concentration being expressed in grams of solute S by gram of phase H. 8. Method according to one of claims 1 to 7, characterized in that the mixture CT M has a mass ratio P / solute S of between 0.2 / 1 and 100/1. 9. Method according to one of claims 1 to 8, characterized in that the mixture M has a mass fraction S2 / [S 1 + S2] between 0.2 and 0.8. 5 10. Nanocapsules obtained by the process according to one of claims 1 to 9. 1/2 Mass fraction S / [S1 + S2]