ES2371898A1 - Cyclodextrin nanogels - Google Patents

Abstract

The invention relates to cyclodextrin nanogels and to a method for obtaining nanogels by cross-linking/emulsifying/evaporating the organic phase of cyclodextrins or the derivatives thereof, or cyclodextrins or the derivatives thereof and hydrosoluble polymers or the derivatives thereof, using molecules containing at least two glycidyl ether groups in the structure thereof as a cross-linking agent. The invention also relates to the compositions thus obtained, which can include active substances and pharmaceuticals and which form inclusion complexes with cyclodextrins. The invention further relates to the use of same as components of controlled-release devices, such as transdermal pharmaceutical forms, buccal, oral, rectal, ocular, otic or vaginal transmucosal forms and parenteral implants, which are designed to administer active substances or pharmaceuticals to humans, animals or plants, or as components of cosmetic formulations. In addition, the invention relates to the use of said compositions as chelating agents in the extraction of biological or toxic molecules from living organisms or contaminating substances from water.

Description

Nanogels of cyclodextrins.

Technical sector

The present invention relates to the development of hydrogels, more specifically refers to the development of Hydrogels of nanometric size.

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State of the art

Cyclodextrins are cyclic oligomers consisting of units of α-D-glucose in a variable number, in general 6 (α-), 7 (β-), or 8 (γ-cyclodextrin). Cyclodextrins have a toroidal structure with a hydrophobic internal surface and a hydrophilic external face (Düchene and Wouessidjewe, Pharm. Technol . 14: 22-30, 1990). This conformation gives them the ability to form complexes with substances of diverse nature (Uekama, Chem. Pharm. Bull . 52: 900-915, 2004). Complex formation has been used for decades to modify the solubility, stability and volatility of active drugs and molecules. More recently, the ability of cyclodextrins to form complexes has been used in the preparation of hydrogels with loading capacity and control of the transfer of improved active molecules (Frank van de Manakker, Tina Vermonden, Cornelus F. van Nostrum, and Wim E. Hennink, Biomacromolecules 10: 3157-31 75, 2009 ).

This ability to incorporate active molecules and control its assignment, makes cyclodextrins a material of interest in the preparation of drug delivery systems. Within these systems are the hydrogels that are characterized by its ability to incorporate water.

In the state of the art hydrogels are known constituted by cyclodextrins for whose preparation they have developed procedures that involve a) the previous formation of an acrylic or vinyl derivative of cyclodextrin susceptible to react with other acrylic or vinyl monomers (Siemoneit, U., Schmitt, C., Alvarez-Lorenzo, C., Luzardo, A., Otero-Espinar, F., Concheiro, A., Blanco-Mendez, J. Acrylic / cyclodextrin hydrogels with enhanced drug loading and sustained release capability. Int. J. Pharm 312, 66-74, 2006); or b) crosslinking Direct cyclodextrins using molecules capable of react simultaneously with several hydroxyl groups cyclodextrins (Rodriguez-Tenreiro, C., Alvarez-Lorenzo, C., Rodriguez-Perez, A., Concheiro, A., Torres-Labandeira, J.J. Estradiol sustained release from high affinity cyclodextrin hydrogels. Eur. J. Pharm. Biopharm 66, 55-62, 2007).

In order to implement procedures simple and compatible with the environment, the direct cross-linking of cyclodextrins, in aqueous medium and mild conditions, using molecules as crosslinking agents which contain two or more glycidyl ether groups in their structure. In the WO2006 / 089993 A2 patent application describes a procedure for obtaining hydrogels of cyclodextrins or their derivatives and water-soluble cellulose ethers or their water-soluble derivatives, or cyclodextrins or their derivatives and guar gums or their derivatives, using molecules that contain two or more as the crosslinking agent glycidyl ether groups in its structure. In the procedure according to WO2006 / 089993 A2 patent application, the dissolution of cyclodextrin with the crosslinking agent "is transferred to a mold adequate "and after crosslinking," the hydrogels are divided into portions of appropriate shape and size and are used as found when removed from the washing liquid or after subject them to desiccation. "

For certain pharmaceutical applications and in particular when a vectorization to specific tissues or cells is required, the use of hydrogels as drug carriers is only possible if they are presented as multiparticulate systems consisting of nanogels smaller than 200 nm in size ( Raemdonck, K; Demeester, J; De Smedt, S. Advanced nanogel engineering for drug delivery, SOFT MATTER 5, 707-715, 2009; Jung Kwon Oha, Ray Drumright, Daniel J. Siegwart, Krzysztof Matyjaszewski. The development of microgels / nanogels for drug delivery applications Prog. Polym. Sci. 33 (2008) 448-477 ). The nanogels could be obtained from monolithic structures, by spraying or crushing, but this procedure ("top-bottom" approach) is difficult to implement in practice and the resulting particles have irregular shapes and their size dispersion is wide. The procedures that allow nanogels to form directly from their bottom-up approach are more suitable for nanogels of controlled size and mainly spherical shape ( Jung Kwon Oha, Ray Drumright, Daniel J. Siegwart, Krzysztof Matyjaszewski. development of microgels / nanogels for drug delivery applications. Prog. Polym. Sci. 33 (2008) 448-477 ).

A recently published procedure is refers to the synthesis of nanogels by polymerization-precipitation of vinyl monomers cyclodextrin together with other acrylic or vinyl comonomers in water at 70 ° C and ultrafiltration (Markus J. Kettel, Fiete Dierkes, Karola Schaefer, Martin Moeller, Andrij Pich. Aqueous nanogels modified with cyclodextrin. Polymer 52 (2011) 1917-1924). This procedure requires in a first stage the prior preparation of vinyl derivatives of cyclodextrin, followed, in a second stage, by copolymerization with other acrylic or vinyl monomers.

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Description of the invention

The authors of the invention have developed nano-sized gels, which solve the limitations of corresponding monolithic hydrogels, as they are useful for applications for which monoliths cannot be used. From Thus, the invention provides nanometric gels that possess well differentiated properties of the corresponding hydrogels monolithic The size of the gels of the invention provides a high specific surface which facilitates exchanges of matter with the environment in which they are and makes it possible for cross cell membranes by endocytosis and elude the recognition by the mononuclear phagocytic system.

Thus, in one aspect the invention is directed to a hydrogel characterized by a lower mean hydrodynamic diameter at 1 micrometer, which comprises a matrix of cyclodextrins, where the cyclodextrins are linked together through a spacer to which they are joined by an ether or amino group.

A particular embodiment of the invention is refers to a hydrogel as defined above that additionally it comprises a water-soluble polymer.

The cyclodextrins that make up the hydrogel of the invention give them a high capacity to incorporate drugs, active substances and biological or toxic molecules with structures and physicochemical properties very diverse. Thus, the hydrogel as defined above can further comprise an active ingredient, a biological molecule, or A toxic molecule

In another aspect the invention is directed to a pharmaceutical composition comprising the hydrogel as has defined above and at least one pharmaceutically excipient acceptable.

In another aspect the invention is directed to a cosmetic composition comprising the hydrogel as defined previously.

In another aspect the invention is directed to a phytosanitary composition comprising the hydrogel as it has been defined above.

The invention further provides a method. suitable for the preparation of the nanogels of the invention which is based on an approximation from the materials that they constitute, and not in a crushing of the monolithic hydrogels corresponding. This procedure has the advantage of preparing the nanogels in a single step, integrating the cross-linking phase and emulsification, and also allows the control of the structure and Morphology of the nanogels.

So in another aspect, the invention is refers to a process for the preparation of the hydrogel as You have previously defined that you understand:

a) preparing an aqueous solution comprising one or more cyclodextrins, a crosslinking agent that possess two or more functional groups that are able to react with the groups hydroxyl of cyclodextrins to form ether groups or with amino groups of cyclodextrins to form amino groups, and a substance of acidic or basic character, and optionally a polymer water soluble,

b) prepare an organic solution that comprises an organic solvent, and optionally an agent surfactant,

c) mix the solutions under stirring prepared in stages a) and b),

d) mix the emulsion obtained in step c) with water.

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In a particular embodiment, the procedure further comprises adding an active ingredient or a molecule biological

In another aspect, the invention relates to the use of the hydrogel as defined above to prepare a medicine.

In another aspect, the invention relates to the use of the hydrogel as defined above, in capable systems of sequestering toxic substances, molecules produced by living organisms, pollutants or liquid waste.

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Description of the figures

Figure 1. Microscopy photomicrograph of electronic transmission of nanogels of γ-cyclodextrin before undergoing the process Freeze drying.

Figure 2. Microscopy photomicrograph of electronic transmission of nanogels of γ-cyclodextrin and hydroxypropylmethylcellulose before undergoing the lyophilization process.

Figure 3. Infrared spectra of γCD (1), HP? CD (2), HPMC (3), agar-agar (4), γCD_ {0,0. 5} (5), γCD-HPMC_2,1 (6), HP? 0.0. 5} (7) and HP? -CD-agar_ 1.0. 5} (8) in the region 1600-800 cm -1.

Figure 4. Broadcast profiles of 3-MBA from a solution without nanogels or from γ-cyclodextrin nanogel dispersions or of γ-cyclodextrin e hydroxypropyl methylcellulose.

Figure 5. Dexamethasone Assignment Profiles obtained from a dexamethasone solution (Dex code), of a dexamethasone solution to which it was incorporated γ-free cyclodextrin (code γCD), and of γ-cyclodextrin and HPMC nanogels prepared with different proportions of Span 80 in the organic phase (0%, code \ gammaCD-HPMC_ {2; 0}; 0.5%, code γCD-HPMC_ {2,0. 5}; 1.0% code γCD-HPMC2,1; 2.0% code γCD-HPMC2,2).

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Detailed description of the invention

For the purposes of the present invention the term "hydrogel" refers to a network of chains of hydrophilic polymers, which may be crosslinked by different methods, and it contains a high proportion of water. The hydrogels can be presented macroscopically or confined in smaller dimensions It is understood by nanogel, a hydrogel of submicron size (Jung Kwon Oha, Ray Drumright, Daniel J. Siegwart, Krzysztof Matyjaszewski. The development of microgels / nanogels for drug delivery applications. Prog. Polym. Sci. 33 (2008) 448-477). As described above, the present invention is directed to a hydrogel characterized by a mean hydrodynamic diameter less than 1 micrometer, comprising a matrix of cyclodextrins, where the cyclodextrins are bound each other through a spacer to which they are joined by a group ether or amino. The nanogels of the invention are capable of incorporate high proportions of water without dissolving.

One of the objectives of the invention is provide useful nanogels in the transport and release of drugs and that are also able to target tissues or concrete cells Thus, in a particular embodiment, the hydrogel as described above it has a hydrodynamic diameter medium between 1 nm and 400 nm. In one more embodiment In particular, the nanogel of the invention has a diameter medium hydrodynamic between 1 nm and 200 nm.

For the best understanding of the invention is important to differentiate nanometer size hydrogels from others nano-sized systems such as nanoparticles. The nanoparticles are solid particles of included size between 1 and 1000 nm (Encyclopedia of Pharmaceutical Technology, J. Swarbrick and J.C. Boylan Vol. 10. Marcel Dekker Inc., New York, p. 165) which, depending on their internal structure, differ in two Groups: nanocapsules and nanospheres. The nanocapsules consist of a cavity surrounded by a layer of polymer; and the nanospheres are continuous matrix systems (F. Rocha Formiga, E. Ansorena, A. Estella-Hermoso De Mendoza, E. Imbuluzqueta, D. González, M. J. Blanco Prieto. Nanosystems based on polyesters. Monograph XXVIII of the Royal National Academy of Pharmacy, Madrid 2009, p.41).

In comparison to liposomes and nanocapsules, nanogels are physically more stable; and the nanoparticles do not have the ability to incorporate high quantities of water without disintegrating, characteristic of gels.

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Cross-linked cyclodextrins

The term "cyclodextrins" herein invention relates to natural cyclodextrins, cyclodextrins synthetic and semi-synthetic, so that this term includes example, without this being a limitation, cyclodextrins of 6, 7, and 8 members (known as alpha-beta and gamma, respectively), cyclodextrins with more than 8 members (known as large cyclodextrins) and cyclodextrin derivatives. Be understood by cyclodextrin derivatives, cyclodextrins substituted in some of its hydroxyl groups by groups functional, for example those listed in the following table:

one

Cyclodextrin derivatives also include its pharmaceutically acceptable salts.

The amino derivatives of cyclodextrins can be obtained by the procedure described in Nature Protocols , 2008, 3, 691-697.

In the nanogel of the invention cyclodextrins they are crosslinked, so that a cyclodextrin is attached to one or more spacers to which other ones are attached cyclodextrins, thus forming a matrix. The cyclodextrins and the spacer are covalently linked through an ether group or Not me.

In a particular embodiment, the spacer it comprises a carbon skeleton that is selected from alkyl, aryl, arylalkyl and polyether chains, linear or branched, optionally substituted. In one embodiment In particular, the spacer is a polyether.

"Alkyl" refers to a chain linear or branched hydrocarbon, cyclic or acyclic formed by carbon and hydrogen atoms, without unsaturations, from 1 to 12, preferably one to eight, more preferably one to four carbon atoms, optionally substituted.

"Aryl" refers to a hydrocarbon aromatic of 6 to 10 carbon atoms, such as phenyl or naphthyl, optionally substituted by an alkyl or oxyalkyl group, which They may be substituted.

"Arylalkyl" refers to one or more aryl groups attached to the rest of the molecule by a radical alkyl, for example, benzyl, 3- (feni1) -propyl, etc.

"Polyether" refers to a chain with one or more ether groups. In a particular embodiment, the polyether is a unsaturated alkyl, or an aryl alkyl, in which they are intercalated one or more ether groups in the alkyl chain, and that is optionally substituted by a functional group selected from hydroxyl, C1-C6 alkyl, hydroxyalkyl C1-C6, C1-C6 alkyloxy, and OR 3, where R 3 is a C1-C4 alkyl optionally substituted by hydroxyl, alkyl C1-C6, hydroxy C1-C6 alkyl or glycidyl ether.

In a particular embodiment, the polyether present the formula - (CHR_ {1} -CHR_ {O}) n-, where n has a value between 1 and 100, preferably between 1 and 50, more preferably between 1 and 10, R1 and R2 may be the same or different and are selected from the group consisting of hydrogen, hydroxyl, C1-C6 alkyl, hydroxyalkyl C1-C6, C1-C6 alkyloxy, and OR 3, where R 3 is a C1-C4 alkyl optionally substituted by hydroxyl, alkyl C1-C6, hydroxy C1-C6 alkyl or glycidyl ether.

The nanogel spacers of the invention, as described above, they come from crosslinking agents that have two or more functional groups that are capable of react with the hydroxyl groups of cyclodextrins to form ether groups or with amino groups of cyclodextrins to form amino groups. These functional groups are known by the person skilled in the art and some examples, not limited to they are the following: 1,2-epoxyethane group, glycidyl ether, primary alkyl halide, alkyl tosylate primary, etc. In a more particular embodiment, the group capable of reacting with the hydroxyl or amino groups of cyclodextrins is a glycidyl ether group.

In a particular embodiment, the agent crosslinker comprises an optionally substituted polyether, and two or more glycidyl ether groups.

Glycidyl ethers have the advantage that they have a very low toxicity. Their wide safety margins, together with the absence of reproductive and endocrine effects and carcinogenic effects, make them suitable as components of containers that remain in prolonged contact with food (Poole et al., Food Additives & Contaminants 21: 905 -919, 2004). Crosslinking agents with glycidyl ether groups (also known as epoxides, oxiranes or alkene oxides; Allinger et al ., Organic Chemistry, 2nd Ed. Reverté SA, Barcelona, 1988, p. 639), also allow obtaining water-soluble polysaccharide hydrogels without need of previously forming derivatives of these polymers to introduce polymerizable groups or of previously modifying their structure (Rodríguez et al., op. cited 2003).

In a more particular embodiment, the agents crosslinkers are selected from diglycidyl ether, ethylene glycol glycidyl ether, diethylene glycol glycidyl ether, polyethylene glycol glycidyl ether, polyglycerol polyglycidyl ether, propylene glycol glycidyl ether, glycerol glycidyl ether, glyceroltriglycidylether, or bisphenol A diglycidylether.

In a particular embodiment, the proportion in dry weight of cyclodextrin is between 1 and 95%, and the dry weight ratio of the crosslinking agent is comprised between 99% and 5% of the total dry hydrogel weight of the invention as described above. In one more embodiment In particular, the dry weight ratio of cyclodextrin is between 4 and 70%, and the dry weight ratio of crosslinking agent is comprised between 96% and 30% of the weight Total dry hydrogel.

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Water soluble polymer

As described above, the nanogels of the invention can incorporate water soluble polymers. By "water-soluble polymer" means any macromolecule natural, semi-synthetic or synthetic that can be dispersed in medium aqueous forming colloidal solutions or systems.

In order to modulate the properties of hydrogels of the invention the authors found that Water-soluble polysaccharides are suitable for modulating affinity of the nanogels by the drugs and to modulate the profiles of assignment. The polysaccharides are constituted by carbohydrates that they have reactive hydroxyl groups similar to those of cyclodextrins, so comparatively to other polymers, they have the advantage of being able to react with the crosslinking agent of similar to the way cyclodextrins do, which facilitates obtaining homogeneous fabrics.

In a particular embodiment, the polymer Water soluble is a water soluble polysaccharide or its derivatives. Be understood by derivatives of a water-soluble polysaccharide, its salts pharmaceutically accepted, and water-soluble polysaccharides resulting from the substitution in some of its hydroxyl groups with functional groups, such as, for example, alkyl, aryl, arylalkyl, alkylcarbonyl, alkoxycarbonyl, etc.

In a more particular embodiment, the Water-soluble polysaccharide is selected from the group consisting of dextrans, alginates, starch, glycogen, chitosan, guar gums, agar-agar, garrafina gums and ethers of water soluble cellulose, and its pharmaceutically salts acceptable.

In a particular embodiment, the ethers of Water soluble cellulose are selected from methyl cellulose (MC), hydroxyethylmethylcellulose (HEMC), hydroxypropylcellulose (HPC), hydroxypropylmethylcellulose (HPMC), hydroxyethylcellulose (HEC), ethylhydroxyethyl cellulose (EHEC), sodium carboxymethyl cellulose (CMCNa), quaternary ammonium salts of hydroxyethyl cellulose with substituent trimethylammonium (Polyquaternium 10), and hydroxyethyl copolymers cellulose and dimethyl diallyl ammonium chloride (Polyquaternium 4).

Examples of guar gum derivatives are their hydroxypropylated or carboxyhydroxypropylated ethers, their derivatives cationic (Ecopol) and products resulting from the depolymerization of guar gums.

In a particular embodiment, the polymer Water soluble is a neutral or ionizable acrylic polymer with the condition of being soluble in aqueous medium. Polymer Acrylic is interpenetrated in cyclodextrin interweaving, not is covalently bound to cyclodextrins or spacers

In a more particular embodiment, the polymer water soluble acrylic is selected from the group constituted by polyacrylic acid, poly-N-isopropylacrylamide, copolymers of methacrylic acid and ethyl acrylate, copolymers of methacrylic acid, methyl acrylate and methyl methacrylate, and copolymers of dimethylaminoethyl methacrylate, butyl methacrylate and methyl methacrylate

In a particular embodiment, the proportion in dry weight of cyclodextrin is between 1% and 95%; the dry weight ratio of water soluble polymer is comprised between 0.05% and 95%; and the dry weight ratio of the agent crosslinker is between 98.95% and 4% of the total weight of the Dry hydrogel of the invention as described above. In a more particular embodiment, the dry weight ratio of cyclodextrin is between 4 and 70%, the proportion in dry weight of water-soluble polymer is between 0.1% and 20%; and the dry weight ratio of the crosslinking agent is between 96% and 30% of the total weight of the hydrogel dry.

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Active ingredient and biological molecules

The nanogels object of the present invention They are suitable for associating active ingredients or molecules biological regardless of solubility characteristics thereof. The ability to associate will depend on the molecule correspondent.

The term "active ingredient" refers to to any substance that is used in the treatment, cure, prevention or diagnosis of a disease or that is used to improve the physical and mental well-being of humans and animals, as well as that compound that is intended to destroy, prevent action, counteract or neutralize, any harmful organism, or either any substance that is used as cosmetic or hygiene, as well as that compound that is intended to regenerate tissues or in tissue engineering

By "biological molecules" is meant any molecule synthesized in a living organism, as per example, proteins, carbohydrates, lipids, nucleic acids, amino acids, etc. In a particular embodiment, the molecule Biological is selected from peptides, proteins, compounds lipid or lipophilic, saccharide compounds, compounds of nucleic acids or nucleotides as oligonucleotides, polynucleotides or combinations of the aforementioned molecules.

In a particular embodiment of the invention, the active ingredient or the biological molecule has activity antifungal, antiseptic or anti-inflammatory, or it is a molecule of interest in tissue engineering, regenerative medicine, Cosmetic or hygiene. In a particular embodiment, the Active ingredient is an anti-inflammatory. In one more embodiment Particularly the active ingredient is an anti-inflammatory Steroid

The proportion of active ingredient or molecule Biological incorporation will depend in each case on the nature of the active ingredient or biological molecule to be incorporated, the indication for which it is used and the efficiency of administration.

According to another particular embodiment, the nanogel of The invention is in lyophilized form.

Given the characteristics of cyclodextrins, presenting ocular compatibility, the nanogels of the invention they are suitable as active ingredient release systems or Biological molecules for ophthalmic treatments.

In a particular embodiment, the nanogel of the invention is used in the preparation of a medicament for the Treatment of ophthalmic diseases.

The nanogels of the invention, with or without active ingredient or biological molecules incorporated, can be use as such or as base components of pharmaceutical forms, medicines and phytosanitary products for the treatment of pathological or physiological states in humans, animals and plants.

In a particular embodiment, the compositions Pharmaceuticals according to the invention are suitable for transdermal, oral, oral, rectal, ocular, nasal, administration otic, vaginal, or parenteral implant.

They can also be used as agents sequestrants of biological or toxic substances in organisms live, for example cholesterol, glucose or bile acids, or in the environment.

The nanogels of the invention are also very suitable to control the transfer of drugs or substances active, incorporated into the nanogels. The compositions They provide different transfer speeds depending on your qualitative and quantitative composition, and of the properties physicochemicals of the drug, especially its water solubility and of its affinity for the cyclodextrin cavity.

They can also serve to direct drugs towards specific areas in living beings, through changes associated with environmental conditions, in the degree of swelling of the nanogel or in the affinity of the drug for the nanogel components.

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Preparation Procedure

The process of the invention takes place in mild conditions, high temperatures and conditions are avoided drastic, and so that it is possible to incorporate ingredients biological assets or molecules without damaging their integrity and without danger of degradation

The process of the invention has the advantage that does not require prior modification of the structure of cyclodextrin or water-soluble polymer, or the use of a mold to form the nanogel. In addition, the procedure of the invention allows modulating the size and shape of the nanogel by means of the control of the emulsion droplet formation process of stage c) and by the proportion of system components.

As described above, the invention also refers to a procedure for the preparation of hydrogel as defined above comprising:

a) preparing an aqueous solution comprising one or more cyclodextrins, a crosslinking agent that has two or more functional groups that are able to react with the groups hydroxyl of cyclodextrins to form ether groups or with amino groups of cyclodextrins to form amino groups, and a substance of acidic or basic character, and optionally a polymer water soluble,

b) prepare an organic solution that comprises an organic solvent, and optionally an agent surfactant,

c) mix the solutions under stirring prepared in stages a) and b),

d) mix the emulsion obtained in step c) with water.

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The preparation of the aqueous solution of the stage a) it is possible to carry it out in several ways without There are variations in the result obtained. Thus it is possible to add first, cyclodextrin in water and then a substance of acidic or basic character in order to modify the pH to adjust it for the crosslinking to take place, or to dissolve the cyclodextrin directly in a medium with the appropriate pH. In cases where a water-soluble polymer is incorporated, the addition of said polymer Water can be done before or after dissolving cyclodextrin. And finally, the agent is added to the resulting solution crosslinker in solid, liquid or aqueous solution.

The acidic substance may be a organic or inorganic acid, for example triluoroacetic acid, p-toluenesulfonic acid, camforsulfonic acid, acetic acid, acidic ion exchange resin, Lewis acid, etc. The substance of basic character includes organic bases and inorganic for example potassium or sodium carbonate, cyanides, hydrides, primary, secondary amines, sodium or potassium methoxide, sodium or potassium ethoxide, etc.

In a particular embodiment, the solution aqueous prepared in step a) is incubated for a period of time between 1 and 60 minutes, more preferably between 5 and 45 minutes In a particular embodiment, incubation of the aqueous solution prepared in step a) is carried out at a controlled temperature between 5ºC and 80ºC, more preferably between 20 ° C and 60 ° C. In stage a) the crosslinking but without giving rise to an increase in viscosity that prevent subsequent emulsification with the organic phase.

In step b) a solution is prepared organic They are examples of organic phases, although the invention is not limited to these, chlorobenzene, chloroform, cyclohexane, dichloromethane, hexane, tetrahydrofuran, toluene, octanol, heptanol

Optionally it is possible to add an agent surfactant to organic solution. The term "agent surfactant "refers to a molecule composed of one part hydrophobic and a hydrophilic moiety. These molecules present, by both amphiphilic properties, which makes in a mixture water / organic solvent, these migrate to the surface between the water and organic solvent. Thus, the hydrophilic head is maintains in the aqueous phase and the hydrophobic tail interacts with the organic solvent altering the surface properties of the water / solvent interface and allowing the formation of an emulsion, as well as its stabilization.

In a particular embodiment, the agent surfactant is selected from the group consisting of derivatives long chain hydroxylics of 8 to 18 carbon atoms (alcohols fatty), ethoxylated carboxylic acids, ethoxylated amides, ethoxylated glycerides, glycol esters and derivatives, monoglycerides, polyglyceryl esters, esters and ethers of polyalcohols, sorbitan / sorbitol esters, acid triesters phosphoric, ethoxylated derivatives of fatty alcohols and ethers of polyethylene glycol In a more particular embodiment, the agent Surfactant is a sorbitan ester. In a particular embodiment, the sorbitan ester is selected from the group consisting of mono-, di-, tri- or sesqui-sorbitan oleate, mono-, di-, tri- or sesqui-sorbitan laurate, mono-, di-, tri- or sesqui-palmitate sorbitan, mono-, di-, tri- or sesqui-sorbitan stearate and mono-, di-, sorbitan tri-sesqui-isostearate, as well as, combinations of the above.

In a particular embodiment, the proportion in weight of surfactant added in step b) is between 0% and 5%, preferably between 0% and 3%.

In a particular embodiment, in step c) the mixture is homogenized for a time between 1 second and 2 minutes, preferably between 30 and 60 seconds. This homogenization can be performed by vigorous agitation, by example, using a high homogenizer stirrer performance.

In a particular embodiment, the volume of the organic phase of stage b) is equal to or greater than the phase volume aqueous resulting from step a).

In a particular embodiment, the mixture of the step c) is stirred for a period of time between 5 minutes and 5 hours, preferably between 10 minutes and 50 minutes. In a particular embodiment, said stirring is performed at a temperature controlled between 5 ° C and 80 ° C, preferably between 40 ° C and 70 ° C

In a particular embodiment, in step d) the emulsion obtained in step c) is diluted with a volume of water between an equal volume and up to 10 times greater than that of the mixture of stage a). In a particular embodiment, the mixture of the dilution is incubated for 10 minutes to 10 hours, preferably between 1 hour to 5 hours. In one embodiment In particular, said incubation is carried out under controlled temperature between 5ºC and 80ºC. Through this process the crosslinking ends and it is possible to evaporate the organic solvent.

In a particular embodiment, the procedure further comprises adding an active ingredient or a molecule biological

In a particular embodiment of the invention, the incorporation of the active ingredient or the biological molecule It can be carried out by one of the following processes: i) direct immersion of a hydrogel of the invention, as described above, in a solution or in a suspension of the ingredient active or of the biological molecule, at a temperature included between 0 and 100 ° C and at atmospheric pressure, optionally using ultrasound, ii) in autoclave at a temperature between 100 and 130 ° C, or, iii) addition of the active ingredient or molecule biological to the aqueous phase a).

In a more particular embodiment, the The method further comprises a stage e) after stage d) which comprises the dialysis of the mixture.

In a more particular embodiment, the The method further comprises a stage f) after stage e) comprising lyophilization of the mixture.

In a more particular embodiment, the The method further comprises a stage g) after stage f) which includes the rehydration of the nanogels.

In another aspect, the invention relates to a nanogel obtainable by the procedure described previously.

The structure of the nanogel obtainable through the procedure described above, is relevant in the tests performed as set out in the examples herein memory. These nanogels have a high capacity of incorporation of drugs, active substances, biological molecules or toxic with structures and properties very diverse physicochemicals that form complexes of inclusion with the cyclodextrins of the nanogels. The nanogels they form stable colloidal systems: dispersed in aqueous medium, they have high physical stability, resisting centrifugation at 5000 rpm for 10 min without precipitating.

The size and shape of the nanogels can be modulate controlling the process of formation of the droplets of the emulsion. The shape of the nanogels of the invention is spherical and the size distribution of nanogels in the colloidal system is narrow.

All these features can be modulated to through an adequate selection of the variety and / or proportion of cyclodextrin / s and of water-soluble polymers or their derivatives to accompany her / him. The low or no toxicity of cyclodextrins, water-soluble polymers and their derivatives, and agents glycidyl ether crosslinkers make the resulting compositions can be used as components of pharmaceutical forms, of cosmetic preparations or "trap" systems to capture molecules of living organisms or the environment, without posing problems of biocompatibility or environmental impact.

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Examples of the invention

Here are some examples that show the obtaining of nanogels using cyclodextrins or their derivatives, or cyclodextrins or their derivatives and polymers water soluble. Also included are examples of the preparation of compositions that incorporate active substances and control their assignment.

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Materials and methods

γ-Cyclodextrin (γCD, W8) and 2-hydroxypropyl-? -Cyclodextrin (HP? CD, W7 HP, Mw 1309.24 Da) were from Wacker (Barcelona, Spain), hydroxypropyl methylcellulose (HPMC, Methocel K4M Premium EP) of Colorcon Iberica S.L. (Barcelona, Spain), Guinama agar-agar (Valencia, Spain), Fluka ethylene glycol diglycidyl ether (EGDE, 50% w / w in water) Louis, IL, USA), dichloromethane from Normaolv, Scharlau, S.A. (Barcelona, Spain), Sorbitan monooleate (Span 80) by Fluka (Schnelldorf, Germany), 3-methyl benzoic acid (3-MBA) from Merck (Darmstadt, Germany). Water purified by reverse osmosis (MilliQ®, Millipore, Spain). The Remaining reagents were of analytical quality.

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Example 1 Procedure for obtaining nanogels based on γ-cyclodextrin or HPβCD

A solution of γ-cyclodextrin or HPβCD, 20% (w / w), in 0.2M NaOH. Then, 10 mL of solution is given added 4 mL of a solution of ethylene glycol glycidyl ether to 50% (w / w) in water, so that the final concentration of agent crosslinker was 14.28%. The solution was stirred for 25 minutes at 60 ° C to start the reaction of reticulation.

Separately the organic phase was prepared consisting of a 2% Span 80 solution (w / w) in dichloromethane at 20 ° C.

The aqueous solution of cyclodextrin- ethylene glycol glycidyl ether was added to 20 ml of the organic phase and the whole was subjected to the action of a homogenizing stirrer of high performance (8000 rev./min; Ultra-Turrax T25, Janke & Kunkel, INK-Labortechnik, Germany) for 30 seconds Then the emulsion remained in stirring (magnetic stirrer 300 rev./min) for 30 min in a thermostated bath at 60 ° C. Next, the emulsion was poured into 100 ml of distilled water and kept under stirring (stirrer magnetic 300 rev./min) for 210 min in a thermostated bath at 60 ° C, to complete the formation of the nanogels.

50 ml of the colloidal system were taken containing the nanogels and dialyzed for 72 hours using dialysis tubes of 12-14 kDa. Behind the dialysis, the colloidal system containing the nanogels was dried by lyophilization (VirTis Genesis freeze-dryer, USES). Once lyophilized, the nanogels were redispersed with ease in water giving rise to a colloidal system with the size of particle similar to that registered before lyophilization (40-500 nm).

Figure 1 shows the photomicrograph of transmission electron microscopy (Philips CM-12 TEM apparatus, FEI Company, The Netherlands) of the nanogels before undergoing the lyophilization process. The sizes of the nanogels were determined using a team of dynamic light scattering using an optical system ALV-5000 F equipped with a Nd: YAG laser (400 mW) connected to a CW diode pump (400 mW) operated at 532 nm (Coherent Inc., Santa Clara, CA, USA). The average size turned out to be of 151.36 nm.

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Example 2 Procedure for obtaining nanogels based on γ-cyclodextrin and hydroxypropylmethylcellulose (HPMC)

A solution of 1% HPMC K4M was prepared (w / w) in 0.2M NaOH. Then, 10 mL of this solution is given added 2 grams of γ-cyclodextrin or HPβCD and, after homogenization, 4 mL of a solution of 50% ethylene glycol glycidyl ether (w / w) in water. The solution is Stirred for 25 minutes at 60 ° C to start the crosslinking reaction.

Separately the organic phase was prepared consisting of a solution of 1% Span 80 (w / w) in dichloromethane at 20 ° C.

The aqueous solution of HPMA-cyclodextrin-ethylene glycol glycidyl ether 20 ml of the organic phase was added and the whole was subjected to the action of a high performance homogenizer agitator (8000 rev./min; Ultra-Turrax T25, Janke & Kunkel, INK-Labortechnik, Germany)) for 30 seconds. TO then the emulsion was kept under stirring (stirrer magnetic 300 rev./min) for 30 min in a thermostated bath at 60 ° C Then, the emulsion was poured into 100 ml of water distilled and kept under stirring (magnetic stirrer 300 rev./min) for 210 min in a thermostated bath at 60ºC, to complete the formation of the nanogels.

50 ml of the colloidal system were taken containing the nanogels and dialyzed for 72 hours using dialysis tubes of 12-14 kDa. Behind the dialysis, the colloidal system containing the nanogels was dried by lyophilization. Once lyophilized, the nanogels will easily redispersed in water resulting in a system colloidal with particle size similar to that registered before the lyophilization (40-500 nm).

Figure 2 shows the photomicrograph of transmission electron microscopy (Philips CM-12 TEM apparatus, FEI Company, The Netherlands) of the nanogels before undergoing the lyophilization process. The sizes of the nanogels were determined using a team of dynamic light scattering using an optical system ALV-5000 F equipped with a Nd: YAG laser (400 mW) connected to a CW diode pump (400 mW) operated at 532 nm (Coherent Inc., Santa Clara, CA, USA). The average size turned out to be of 93.68 nm.

Following the procedures of examples 1 and 2 and varying the proportions of surfactant and the proportion different nanogels were obtained from polysaccharide as collected in table 1.

The nanogel formulations obtained are identify using code cyclodextrin-polysaccharide_ {x, y} where the Cyclodextrin is γ-CD or HPβCD, the polysaccharide is HPMC or agar, "x" is the concentration of HPMC or agar (0%, 1% or 2%) in the aqueous phase and "y" is the concentration Span 80 in the organic phase (0%, 0.5% or 2%) as used in hydrogel preparation as described in examples 1 and 2.

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Table 1. The data of the surfactant concentration, process performance Preparation (Rto), dynamic light analysis results scattering (DLS) of the nanogels suspension, area of each peak, hydrodynamic radius, and mass distribution.

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By gas chromatography (Finnigan Trace GC ultra Thermo, USA) -mass spectrometry (Finnigan Trace DSQ Thermo, USA) the residual amount of chloride was determined methylene lyophilized nanogels, which was in all cases close to the limit of quantification: 1 ppm.

The stability of the nanogels obtained as aqueous dispersions, after centrifugation at 5000 r.p.m. for 10 minutes or 10,000 r.p.m. for 30 minutes to assess the tendency of nanogels to precipitate, simulating thus an aging process during storage. Be noted that centrifugation at 5000 r.p.m. for 10 minutes no caused precipitation of the nanogels. While centrifugation at 10,000 r.p.m. for 30 minutes led to small amounts precipitates that were more intense in the nanogels that were HPMC incorporated them in the case of constituted nanogels only by cyclodextrins. In all cases, after shaking the precipitated, it redispersed again.

Infrared spectra were taken from the γ-CD nanogels in a range of between 400 and 4000 cm -1, on a Brucker IFS 66V spectrophotometer FT-IR (using the potassium bromide technique). These spectra are shown in Figure 3. Group formation ether between the crosslinking agent and cyclodextrins is put in evidence in infrared (IR) spectra when comparing Starting species against cross-linked cyclodextrins.

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Example 3 Control of the transfer of 3-methylbenzoic acid (3-MBA) from nanogels of γ-cyclodextrin and nanogels of γ-cyclodextrin e hydroxypropyl methylcellulose

Lyophilized nano-angels dispersed in 3-MBA solutions (0.08 mg / ml) to obtain a 2% w / v nanogel dispersion, which was maintained at 20 ° C for 60 hours. The transfer of 3-MBA from nanogel dispersions were evaluated using diffusion cells vertical, using cellophane membrane (MWCO 3500, 0.785 cm2) as a separation barrier between the donor compartment and the receiving compartment The tests were carried out at 37 ° C using 2 ml of nanogel dispersion and 5.5 ml of water purified, subjected to magnetic stirring at 300 rpm, as a medium receiver. It was also tested, under the same conditions, a 3-MBA solution (0.08 mg / ml) without nanogels. TO preset time intervals 750 samples were taken µl of the receiving medium and replaced by fresh medium. The 3-MBA concentration in the receptor medium is determined by UV spectrophotometry at 281 nm. Figure 4 shows show the assignment profiles obtained.

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Example 4 Control of the transfer of dexamethasone from nanogels of γ-cyclodextrin and nanogels of γ-cyclodextrin e hydroxypropyl methylcellulose

Lyophilized nano-angels dispersed in saturated dexamethasone solutions (0.14-0.16 mg / ml) to obtain a dispersion of 2% w / v nanogels, which held at 20 ° C for 16 hours or 7 days. Dexamethasone Assignment from the dispersions of nanogels was evaluated using vertical diffusion cells, using cellophane membrane (MWCO 12-14 kDa, 1.77 cm2) as separation barrier between the donor compartment and the receiver compartment. The Assays were carried out at 37 ° C using 2 ml dispersion of nanogels and 12 ml of purified water, subjected to magnetic stirring at 300 rpm, as receiving medium. It was also tested, in them conditions, saturated dexamethasone solution without nanogels. TO preset time intervals 150 samples were taken µl of the receiving medium and replaced by fresh medium. The Dexamethasone concentration in the receptor medium was determined by HPLC Figure 5 shows the assignment profiles obtained from from the dissolution of dexamethasone (Dex code), of a dexamethasone solution to which it was incorporated γ-free cyclodextrin (code γCD), or of γ-cyclodextrin and HPMC nanogels prepared with different proportions of Span 80 in the organic phase (0%, code \ gammaCD-HPMC_ {2,0}; 0.5%, code γCD-HPMC_ {2,0. 5}; 1.0% code γCD-HPMC2,1; 2.0% code γCD-HPMC2,2).

Claims (27)

1. Hydrogel characterized by an average hydrodynamic diameter of less than 1 micrometer, comprising a matrix of cyclodextrins, where the cyclodextrins are linked together through a spacer to which they are joined by an ether or amino group. 2. Hydrogel according to claim 1, wherein the average hydrodynamic diameter is between 1 nm and
400 nm 3. Hydrogel according to claim 2, wherein the average hydrodynamic diameter is between 1 nm and
200 nm 4. Hydrogel according to claim 1, wherein the spacer comes from a crosslinking agent that has two or more functional groups that are able to react with the groups hydroxyl of cyclodextrins to form ether groups or with amino groups of cyclodextrins to form amino groups. 5. Hydrogel according to claim 4, wherein the crosslinking agent comprises an optionally substituted polyether, and two or more glycidyl ether groups. 6. Hydrogel as defined according to any of claims 1 to 5, further comprising a polymer water soluble. 7. Hydrogel according to claim 6, wherein the Water-soluble polymer is a water-soluble polysaccharide or its derivatives. 8. Hydrogel according to claim 6, wherein the Water-soluble polymer is a neutral acrylic polymer or ionizable with the condition of being soluble in aqueous medium. 9. Hydrogel as defined according to any of claims 1 to 8, further comprising an ingredient active, a biological molecule, or a toxic molecule. 10. Hydrogel according to claim 9, wherein the active ingredient or biological molecule possess activity antifungal, antiseptic or anti-inflammatory, or it is a molecule of interest in tissue engineering, regenerative medicine, Cosmetic or hygiene. 11. Hydrogel according to claim 10, wherein The active ingredient is a steroidal anti-inflammatory. 12. Hydrogel as defined as any one of claims 1 to 11, wherein the hydrogel is found in lyophilized form. 13. Pharmaceutical composition comprising the hydrogel as defined according to any of the claims 1 to 11, and at least one pharmaceutically excipient acceptable. 14. Pharmaceutical composition according to claim 13, for transdermal, oral, oral administration, rectal, ocular, nasal, otic, vaginal, or parenteral implant. 15. Cosmetic composition comprising the hydrogel as defined according to any of the claims 1 to 11. 16. Phytosanitary composition comprising the hydrogel as defined according to any of the claims 1 to 11. 17. Procedure for the preparation of hydrogel as defined according to any of the claims 1 to 11, comprising: a) preparing an aqueous solution comprising one or more cyclodextrins, a crosslinking agent that has two or more functional groups that are able to react with the groups hydroxyl of cyclodextrins to form ether groups or with amino groups of cyclodextrins to form amino groups, and a substance of acidic or basic character, and optionally a polymer water soluble, b) prepare an organic solution that comprises an organic solvent, and optionally an agent surfactant, c) mix the solutions under stirring prepared in stages a) and b), d) mix the emulsion obtained in step c) with water.

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18. Method according to claim 17, where the surfactant is selected from the group consisting of long chain hydroxylic derivatives from 8 to 18 carbon atoms (fatty alcohols), carboxylic acids ethoxylates, ethoxylated amides, ethoxylated glycerides, esters of glycol and derivatives, monoglycerides, polyglyceryl esters, esters and polyol ethers, sorbitan / sorbitol esters, tri esters of phosphoric acid, ethoxylated derivatives of fatty alcohols and polyethylene glycol ethers. 19. Method according to claim 18, where the surface active agent is a sorbitan ester. 20. Procedure according to any of the claims 17 to 19 further comprising adding an ingredient active or a biological molecule. 21. Method according to claim 20, where the incorporation of the active ingredient or molecule Biological is carried out by one of the following processes: i) direct immersion of a hydrogel of the invention, as described above, in a solution or in a suspension of the active ingredient or biological molecule, to a temperature between 0 and 100 ° C and at atmospheric pressure, optionally using ultrasound, ii) in autoclave at a temperature between 100 and 130 ° C, or, iii) addition of the active ingredient or of the biological molecule to the aqueous phase a).

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22. Procedure according to any of the claims 17 to 21, further comprising a step e) after stage d) comprising the dialysis of the mixture. 23. Method according to claim 22, which further comprises a stage f) after stage e) which It comprises lyophilization of the mixture. 24. Method according to claim 23, which further comprises a stage g) after stage f) which It comprises the rehydration of the nanogels. 25. Use of the hydrogel as defined according to any of claims 1 to 11, to prepare a medicine. 26. Use of the hydrogel according to claim 25, where the medicine is for the treatment of diseases ophthalmic 27. Use of the hydrogel as defined according to any of claims 1 to 11, in systems capable of sequester toxic substances, molecules produced by organisms live, pollutants or liquid waste.