WO2019030661A1

WO2019030661A1 - Fiber and electrospun matrix based on silk proteins, process for its preparation, manufactured products comprising it and their use in therapy and cosmetics

Info

Publication number: WO2019030661A1
Application number: PCT/IB2018/055926
Authority: WO
Inventors: Stefano LORENZONI; Chiara MARCOLIN
Original assignee: Leonardino S.R.L.
Priority date: 2017-08-07
Filing date: 2018-08-07
Publication date: 2019-02-14
Also published as: IT201700091328A1

Abstract

Object of the present invention is an electrospun fiber and matrix based on silk proteins, in particular comprising mixed fibroin and sericin. Further object of the invention is a process for the preparation of said matrix as well as the manufactured products comprising it and their use in therapy and cosmetics.

Description

"FIBER AND ELECTROSPUN MATRIX BASED ON SILK PROTEINS, PROCESS FOR ITS PREPARATION, MANUFACTURED PRODUCTS COMPRISING IT AND THEIR USE IN THERAPY AND

COSMETICS"

***

Technical field

Silk produced by silkworms of Bombyx mori species is in the form of double filament, named "bave". Such bave is made of two inner "flosses" made of a protein called fibroin, which represents about 70-80% of the filament weight, which remain coupled thanks to the adhesive effect of a second protein named sericin, which represents the remaining 20-30% of the filament.

The use of derivatives of silk proteins, fibroin and sericin is known, for the preparation of products suitable for the skin treatment, e.g. for the wound healing or the cosmetic use.

As a matter of fact silk fibroin is a protein widely known for its applications as biomaterial and has interesting properties. The protein is made of two chains, one heavy and one light, of about 350 and 25 kDa respectively, which are bonded by disulfide bridges. Fibroin is studied, in laboratory, for research purposes in the biomedical field.

The adhesive protein, sericin, is on the other hand made of three water-soluble glycoprotein fractions, with molecular weight of about 150 KDa, 250 KDa and 400 KDa.

It is known in the art to solubilize fibroin and obtain a fiber from such solution, e.g. by electrospinning; the obtained fibers are recovered to form a nonwoven fabric, as described e.g. in Kim et al. Polymer Journal, 35, 2, (2003), 185-190. The nonwoven fabric of fibroin fibers is known also as fiber "matrix" and, in the present application, such term "matrix" will be used.

WO2016/059611, in the name of the present Applicant, describes the production of medical devices for the wound treatment; the device comprises a structure, i.e. a matrix, of electrospun fibroin. The electrospun fibroin described in WO2016/059611 is defined as being "pure" (page 11 , row 7 and claim 20), and forms therefore an electrospun matrix consisting essentially of fibroin at 100%, thus meaning that it is free from other components, in particular, but not limited to, free from PEO (page 4, rows 21-23 of WO2016/059611).

The two proteins can be separated and isolated by a known way, starting from the silkworm cocoon by means of a process called "degumming". Sericin, as mentioned, is water-soluble and is therefore extracted by subjecting the cocoons to cycles of high temperature and pressure; the aqueous solution of sericin obtained from the degumming is brought to dryness in order to remove the solvent, to provide a sericin powder. On the other hand, fibroin is difficult to process, and is required to subject it to chemical treatments in order to solubilize and thereafter process it, e.g. by electrospinning.

Most of medical or cosmetic compositions claiming the use of silk proteins actually contain derivatives containing of decomposition products of such proteins. It is therefore clear that such compositions can not have the characteristics of the native silk proteins, being hydrolyzed poly- or oligo-peptides.

It is also known the use in cosmetics of hydrolyzed products deriving from the silk proteins. For example, WO2016/176633 describes a hydrating composition based on fibroin protein fragments free from sericin, which are emulsified in water with an oil or butter and with hyaluronic acid. Still by way of example, Hang et al., Int. J. Biol. Macromol., 51(2012), 980-986, describes an electrospinning process of two separate solutions of fibroin and sericin and explicitly points out that said proteins are inevitably degraded with breakdown of the peptide chains (page 981, left column). However the known products are poor performing, at least in the cosmetic field. Moreover, the electrospinning process of Hang et al. requires a coaxial electrospinning of fibroin and sericin from two different aqueous solutions; the fibroin is extruded as "core" and the sericin as "shell", mimicking the structure of the yarn of the cocoon. However this electrospinning method of fibroin and sericin is complex and expensive to be realized and hardly usable in an industrial process. There is therefore the need of providing new products comprising the silk proteins in an improved form with respect to what is known in the art, to be used in the medical and cosmetic field. Objects of the Invention

It is an object of the present invention to solve the above mentioned problems and provide a fiber and a matrix of improved electrospun fibers. Such object is achieved by means of the present invention providing a fiber containing fibroin and sericin, preferably both in the native state, or said fiber being able to contain further components, in particular compounds useful for the medical and/or cosmetic use and having an advantageous action in such sense when applied topically. It is another object of the invention a fiber matrix as indicated above; a further object of the invention is a process for the preparation of said fiber and said matrix.

It is a further object of the invention to provide manufactured products comprising said matrix and the use of said matrix and said manufactured products in the therapeutic and/or cosmetic field.

Such objects are achieved by means of the present invention, which concerns a fiber containing a mixture of fibroin and sericin optionally combined with other components, according to claim 1. The invention also refers to a fiber matrix containing a mixture of fibroin and sericin according to claim 6. As discussed above, such matrix can be in the form of a nonwoven fabric (nwf) made of the electrospun fibers. The invention also refers to a process for the electrospinning production of said fiber and the relative matrix, according to claim 8. Objects of the invention are also the uses in cosmetics and therapy of the fiber and matrix, according to claims 13 and 14.

Preferably the proteins used in the invention are in the native form. Sericin and fibroin are preferably in a non-hydrolyzed or a substantially non-hydrolyzed form and have a MW (molecular weight) identical, similar or analogous to that of the native proteins. In an embodiment, suitable fibroin and sericin have the following characteristics: the heavy chain (H chain) of fibroin used in the invention has a molecular weight greater than 200, preferably greater than 300 and more preferably greater than 320 kDa. The water-soluble sericin fractions have molecular weight greater than 80 kDa, preferably greater than 100 and more preferably greater than 130 kDa. As discussed above, in a preferred embodiment fibroin and sericin are in the substantially native state or have been slightly hydrolyzed, as stated above with the reported molecular weights. Suitable methods to determine the molecular weight of sericin and fibroin are, for example, those described at page 981, section 2.1 of the publication by Hang et al., cited above; in particular, the MW can be determined by gel electrophoresis on sodium dodecyl sulfate-polyacrylamide (SDS-PAGE), in particular for the fibroin, and by MALDI-MS 4800 proteomics Analyzer (Applied Biosystems Co.), in particular for the powdered sericin.

Brief Description of the Drawings

Figure 1 represents an image of the matrix of the invention, collected by scanning electron microscopy (SEM) with a magnification of 15000x.

Figure 2 represents an image of the matrix of the invention, collected by scanning electron microscopy (SEM) with a magnification of 15000x wherein the size of some fibers is reported.

Figure 3 represents an image of the matrix of the invention, collected by scanning electron microscopy (SEM) with a magnification of lOOx.

Figure 4 represents an image of the matrix of the invention collected by optical microscopy, after Coomassie Brilliant Blue staining which highlights the presence of sericin.

Figure 5 shows a section of the "bave" wherein it is possible to note the fibroin flosses wrapped by sericin (5A) and a filament of the matrix according to the invention (5B).

Figure 6 shows the negative result of the attempt of solubilizing the cocoons (or silk cortices) without prior treatment, as described in Example 9.1.

Description of the Invention

Object of the present invention, according to one the aspects thereof, is a fiber containing fibroin and sericin and a matrix of such fibers. Differently from known fibers, where sericin and fibroin are in the form of structures different from each other, even though in contact (e.g. as coaxial and/or co-extruded structures), in the fibers of the present invention fibroin and sericin are present in mixture. In other words, the two proteins are, in the fiber of the invention, mixed together in a random way in the final fiber, e.g. as it can be obtained by electrospinning of a single solution containing a mixture of the peptide polymers forming the two proteins. The fibers of the invention are preferably part of a matrix, e.g. in the form of nonwoven fabric, which is used to produce manufactured products and products used in the fields of application of interest. Examples of the products containing a matrix according to the invention are compresses, masks for the skin of the face, neck, hands and other parts of the body; dressings; patches; gauzes; bandages and bandaging; diapers, sanitary pads and incontinence pads.

In an embodiment, the fibers consist of fibroin and sericin which are present together in the fiber; sericin being water-soluble and releasable from the fiber or a fiber matrix. In embodiments, the fiber can comprise also further components useful in the cosmetic and/or medical field; such components of the fiber are compounds or active ingredients performing an advantageous action in the medical and/or cosmetic field, in particular when applied topically.

In an aspect, the invention refers to a matrix essentially consisting of regenerated silk fibers, where "regenerated" means a recreated silk (by bringing back together its two protein components) downstream of a solubilization step of the raw fiber (cocoon) and of the separation of the two proteins or, in general, a silk recreated after one or more steps to which the raw fiber (of the cocoon) is subjected. It should be noted that the regenerated silk of the invention, differently from the prior art, has not separated and differentiated structures (a fibroin structure and a sericin structure) mimicking the natural, untreated silk. In the fibers of the invention the structure is only one and comprises mixed fibroin and sericin.

In the present case the two protein components, i.e. sericin and fibroin, are brought together in specific weight ratios in a solution that is electrospun to give mixed fibers containing both fibroin and sericin, intimately mixed together.

According to an aspect of the present invention, the fibers of the matrix comprise mixed fibroin and sericin and are obtained from a solution containing both sericin and fibroin. The used sericin and fibroin are preferably native or substantially in native form. In the following description with the expressions "native fibroin and sericin", "fibroin and sericin in the native state" and the like, it is meant that the two proteins are not in hydrolyzed form, i.e. that the primary structure (i.e. the amino acid chain) of the proteins has not been substantially or significantly altered. This is made possible by the mild nature of the treatments performed in the process for its preparation.

By "essentially" it is meant herein that the fibroin and sericin represent at least the 90 wt%, preferably at least the 95 wt%, more preferably at least the 98 wt% of the fiber matrix of the invention.

In an embodiment, the proteins used in the invention are in the native form. Sericin and fibroin have preferably a MW (molecular weight) identical, similar or analogous to that of the native proteins. In an embodiment, partially hydrolyzed fibroin and sericin can be used. However the hydrolyzation must not be excessive, in order to avoid the loss of the "filming" effect typical of sericin. The ideal hydrolyzation level is the one with which the sericin retains its filming effect having at the same time an increased solubility.

In an embodiment, the heavy chain of the fibroin used in the invention has a molecular weight greater than 200, preferably greater than 300 and more preferably greater than 320 kDa. The water-soluble sericin fractions have molecular weight greater than 80 kDa, preferably greater than 100 and more preferably greater than 130 kDa.

Suitable methods to determine the molecular weight of sericin and fibroin are for example those described at page 981, section 2.1 of the publication by Hang et al., cited above; in particular, the MW can be determined by gel electrophoresis on sodium dodecyl sulfate-polyacrylamide (SDS-PAGE), in particular for the fibroin, and by MALDI-MS 4800 proteomics Analyzer (Applied Biosystems Co.), in particular for the powdered sericin.

The matrix of the invention is the product of an electrospinning process, providing a product consisting of fibers with average diameter lower than one micron. The fiber matrix of the invention (hereinafter also only matrix) is in the form of a nonwoven fabric and the fibers of the matrix have preferably a mean diameter from 200 to 800 nm, preferably from 300 to 700 nm (measured by a scanning electron microscope SEM MIRA3, TESCAN).

In the matrix of the invention, at least 85% of the fibers have a mean diameter between 200 and 800 nm, preferably at least 55% of the fibers have a mean diameter between 300 and 700 nm. According to a preferred embodiment, the fibroin/sericin weight ratio in the matrix is between 60/40 and 95/5, preferably between 70/30 and 95/5, more preferably between 75/25 and 92/8, advantageously between 85/15 and 90/10, respectively.

Object of the present invention, according to another of the aspects thereof, is a process for the preparation of a fiber matrix essentially consisting of fibroin and sericin, both in the native state, which comprises electrospinning a mixture of fibroin and sericin in a suitable solvent, preferably a weak acid, characterized in that the starting fibroin is pre-treated in an acidic or basic environment with binary salts. According to a preferred embodiment said weak acid is formic acid, generally, as solvents also other organic solvent, such as hexafluoroisopropanol (HFIP) or trifluoroacetic acid can be used, in particular solvents showing Lewis acid properties. By "binary salts" it is meant herein salts consisting of a metal and a non-metal. Preferred binary salts are calcium chloride (CaCl₂), lithium bromide (LiBr) and magnesium chloride (MgCl₂), calcium chloride being particularly preferred.

According to a preferred embodiment, the starting fibroin, before being electrospun with sericin, is pre-treated with CaCl₂ and formic acid as described in WO2016/059611, in the name of the present Applicant; this treatment allows to keep intact the primary structure of fibroin.

According to a preferred embodiment, the fibroin/sericin weight ratio in the mixture used in the process of the invention is in the range from 60/40 to 95/5, preferably from 70/30 to 95/5, more preferably from 75/25 to 92/8, advantageously for example from 85/15 to 90/10,

The electrospinning process can be carried out under different conditions of temperature and humidity. According to a preferred embodiment, the electrospinning is carried out at temperatures between 15 and 40°C, advantageously at 20-25°C. According to a preferred embodiment, the electrospinning is carried out in an environment with a relative humidity from 25 to 55%, preferably from 25 to 45% advantageously from 30 to 40%. According to a particularly preferred embodiment, the electrospinning is carried out at 20 and 25°C and in an environment with a relative humidity of 30-40%. In order to carry out the electrospinning process, the two proteins, fibroin and sericin, can be separated from the cocoon by means of conventional degumming processes known in the art. Before being subjected to the electrospinning according to the invention, fibroin is pre-treated as stated above, i.e. with binary salts under an acidic or basic environment, advantageously with calcium chloride and formic acid as described in WO20016/059611, herein incorporated by reference. According to such art, for example, fibroin is dissolved in CaCl₂ and formic acid under mild stirring at room temperature; then the so-obtained viscous solution is poured into a container suitable to favor the evaporation of the formic acid (or the solvent); the fibroin is purified from calcium chloride thus obtaining a fibroin film. Electrospinning can be carried out by means of needles or by the "needleless" technique.

The sericin used as starting material for the electrospinning with the fibroin can be, for example, isolated by bringing to dryness the waters used in the degumming process according to the known techniques.

Object of the invention, according to another of the aspects thereof, is the matrix that can be obtained and/or is obtained by means of the process described above. Exemplary examples of such process are reported in the following Experimental Section; the parameters of the solution, the electrospinning process and the environment are reported in the table according to examples 2-5.

Surprisingly, the process of the invention allows a fiber to be obtained and, therefore, a matrix consisting of a mixture of sericin and fibroin in the native state, where by "native state" is meant the state as defined above and as reported in claim 2. Sericin and fibroin are in the fibers with a "random" distribution, i.e. as can be obtained by electrospinning a single solution containing the two substances, without using coaxial electrospinning of two different solutions. In other words, with reference to Figure 5, the fibers of the matrix according to the invention (5) are recognized from the fibers of natural silk (1) and from the fibers of the known art (Hang et al.) since they do not have one or more "cores" of fibroin (3) and an external "shell" of sericin (4) as in the natural bave, but the intimate mixture of the two proteins, fibroin (3) and sericin (4), in the electrospun fiber (5) as shown in Figure 5B.

Moreover, at least part of the sericin present in the fibers of the matrix of the invention is water-soluble, therefore it can be released from the matrix in presence of water; this property is particularly useful in the applications of the matrix of the present invention. It is clear that the sericin release in water will be significant thanks to the high exchange surface due to the reduced size of the fibers (lower than one micron).

As a matter of fact, due the natural function of sericin, which tends to "stick" and keep connected the fibroin molecules, it was not expected to be able to obtain the matrix of the invention. Moreover, sericin could have deteriorate in the solvent used for the electrospinning and finally, but not less important, it was not possible a priori to expect obtaining the fibers. Even only for this series of reasons, the obtained matrix represents an original and unexpected result.

Once electrospun, the matrix of the invention can be also coupled to other materials, e.g. with one or more layers, preferably one layer, of a material selected from viscose, polyester, cellulose, cotton fibers, foams, gel forming materials, gauze, polyurethanes and the mixtures thereof, when possible. The matrix, possibly coupled to other materials as stated above, can be easily sized in different ways, by simply cutting it to the desired form, thus obtaining a ready-to-use manufactured product. According to another embodiment, object of the invention is a fiber and a matrix of electrospun fibers; which comprises fibroin and sericin, both pre-treated as stated above, mixed each other and together with other components. Such components are in general components with an advantageous activity when applied topically, in particular useful to hair and skin wellness. Such components are soluble or dispersible in the solvent used for the electrospinning step. Such components include, e.g., hyaluronic acid or a salt thereof which is acceptable for the topical application, collagen, Vitamins e.g. Vitamin C, vegetable extracts and the like.

According to a preferred embodiment, the matrix of electrospun fibers comprises fibroin and sericin, both pre-treated as stated above, and hyaluronic acid or a salt thereof acceptable for the topical application, advantageously sodium hyaluronate. For the preparation of the matrix comprising fibroin and sericin together with other components to the solution to be electrospun, which contains fibroin and sericin, preferably in the ratios specified above, it is added the desired quantity of the additional component or components and the electrospinning is then carried out. In this manner matrices, and fibers, consisting of the components present in the solution, are obtained. By way of example, aqueous solutions of fibroin at 5-10% and sericin at 0.1-5%, where "%" means the gram ratio of component per 1 ml of the solution to be electrospun, can be electrospun.

The other possible components, preferably hyaluronic acid and/or Vitamin C and/or collagen, can be added in an amount of 0.1-5% to the above mentioned solutions. The matrix described above, in its different forms, i.e. only consisting of fibroin and sericin or consisting of fibroin, sericin and other components, and the manufactured products comprising it, can be used in the therapy and/or cosmetic field. The term "manufactured product" means herein any product derived from handwork or a machine work, comprising the matrix of the invention.

By way of example, in the cosmetic field, the matrix and its manufactured products can be used topically, e.g. as ready-to-use mask, for the skin of the face, neck, hands and, in general, for the body and the hair; thanks to the presence of silk proteins, such compress provides a hydrating, calming and anti- wrinkle effect.

The release of sericin from the matrix, in particular if it is native, on the skin and hair produces indeed a protective film preventing dehydration and keeping therefore well hydrated the part on which it is deposited. It is also expected that sericin could work as a carrier for the delivery of the hyaluronic acid or another active component, from the matrix to the part of the user and that it could, thanks to its filming properties, favor the application of the active (i.e. advantageous) component to said part of the user.

Therefore object of the invention, according to another of the aspects thereof, is the use of the fiber and matrix of the invention and the manufactured products containing it, in the cosmetic field.

In order to carry out its action, the matrix or the manufactured product containing it, can be placed on the part of the body of interest; the matrix or, more preferably, the part of the body of interest are moistened with water or any other water-based product compatible with the use, such as for example, a toner or thermal water, to solubilize the sericin in the fibers and bring about its release. If present, also the additional component is released. The sericin release occurs in a quite fast way, since the protein is immediately soluble in water and in water-based liquids and, moreover, the nanomatrix has a high surface/volume ratio. The expected application time can be 1-90 minutes, preferably 5-75 minutes, e.g. 10-20 minutes. However other application times can be used, without any limitation.

With particular reference to the masks for the skin of the face, neck, hands and, in general, of the body, an important advantage of the invention with respect to known products, is represented in that the matrix is in the form of a solid and dry thin layer. Once placed in contact with a part of the body moisten with water or any other water-based product compatible with the use, such as for example a toner or thermal water, the matrix adheres thereto and releases sericin, thus producing the beneficial effects on the skin, either intact or damaged. The presence of fibroin as well produces advantageous effects, in particular when applied on damaged skin. Moreover, an advantage linked to the structure/morphology of the matrix is that the latter has a high surface/volume ratio, thus allowing a very effective sericin release. Therefore, contrary to the conventional masks, which are marketed already imbibed with liquid or semi-solid components which must therefore be packaged in a suitable way in order to avoid drying (e.g. in "Terra pak"^®), the mask obtained with the matrix of the invention can be packaged in less complex and cheaper containers, an aspect representing a significant industrial advantage. As a matter of fact, being a solid and dry layer, it does not require packaging preserving its humidity. Moreover, contrary to the conventional masks, handling the mask of the invention produces a pleasant feeling, since it is not sticky or gluey.

At the time of use, as mentioned, it is enough to moisten the part of the body where the matrix has to be applied in order to obtain a perfect adhesion of the mask and the following sericin release.

Alternatively, the matrix and the manufactured products containing it can be used in the medical field, e.g. for the wound treatment. It is clear therefore that the matrix of the invention can be used indifferently on intact or damaged skin.

The manufactured products containing the matrix, such as for example masks for the skin of the face, neck, hands and, in general, of the body, dressings, patches, bandages, bandaging, diapers and sanitary pads, incontinence pads and the like, are a further object of the invention.

Object of the invention, according to another of the aspects thereof, is a method to hydrate and protect the skin, the nails and the hair, which comprises applying the matrix or a manufactured product containing it, on the part of interest for said hydration and protection. The term "protect" includes the formation of a film, mainly due to the sericin release from the matrix, on the skin thus avoiding its dehydration. The matrix of the invention can also be used as a carrier to topically deliver substances, such as for example hyaluronic acid, collagen, other advantageous substances for the skin and, when possible, combinations thereof. In an embodiment of the invention it is provided the presence of microencapsulated substances added to the above described matrix of sericin and fibroin fibers according to the invention. It should be noted that sericin is well suited to encapsulation, being a globular protein. The possibility of adding a posteriori substances to the pre-formed matrix, still of fibroin and sericin, not only of fibroin, could be useful in the case of solubilization problems of some active ingredients in the solvents used in the electrospinning.

The following examples illustrate the invention without any limitation.

Experimental Section

Example 1

General procedure

From the cocoon of the silk worm, by means of the conventional degumming process, the two proteins constituting the silk, fibroin and sericin, have been separated. Sericin has been recovered from the degumming waters which have been freeze-dried to provide a pure sericin powder. The solid residue of degumming, i.e. fibroin, is contacted with CaCl₂ and formic acid (with a concentration of 1-10% p/v, preferably of about 2% of CaCl₂ and a concentration of fibroin in the range 1-15% p/v, preferably around 8% p/v) under mild stirring and at room temperature. The so- obtained viscous solution is poured into a suitable container and the formic acid is left to evaporate. Thereafter fibroin is purified from CaCl₂, by means of washing with water, in order to obtain pure fibroin.

The pre-treatment process with calcium chloride and formic acid is described in detail in the Patent Application WO2016/059611, in the name of the same Applicant. In order to prepare the electrospun matrix of the present invention, the two proteins are separately weighted and dissolved in formic acid. Once the solution is homogeneous by eye (about 20-30 minutes), the electrospinning is carried out.

Examples 2-5

Preparation of the electrospun matrix, by means of conventional electrospinning (with needles)

It is carried out as described in the general procedure, by using the following conditions:

Figures 1 -3 show images of the matrix of example 2 collected by scanning electron microscopy (SEM) with two magnifications. The nonwoven fabric structure can be noted, which consists of fibers and low presence of defects (Figure 1). The images at higher magnification (Figures 2 and 3) highlight the cylindrical morphology of the fibers, the absence of microscopic defects and a good uniformity of the fiber size, generally between 400 and 800 nm.

Figures 4 and 5 show two images at the optical microscope of the matrix after the treatment with Coomassie Brilliant Blue; such dye can selectively bind to the sericin (and not to the fibroin). The Figures show blue fibers, confirming the presence of sericin in the fibers and its homogeneous distribution within the matrix.

Example 6 Sericin release assay

In order to verify the sericin release from the matrix in presence of water, the following assay has been performed.

100 mg of electrospun samples of fibroin/sericin according to the invention and, separately, 100 mg of matrices of fibroin only as controls, are soaked in 10 ml of distilled water. The samples, immersed into the aqueous medium, are placed in an oven at 37°C for 30 minutes or, according to an analogous procedure, for 1 hour. Thereafter, the fibroin/sericin or fibroin only matrices are extracted, and the aqueous medium, in which the release happened, is brought to dryness. In this way it has been possible to observe and quantify, e.g. by weighing, the presence of residual sericin in one case (electrospun blend), and the absence of any protein release in the control matrix of fibroin only.

Example 7

Evaluation of the relative amounts of the two proteins in the matrix

The amount of the two proteins in the matrix can be easily verified by carrying out the following steps:

a) carrying out the release assay of sericin of example 6,

b) controlling the sericin release in the matrix by subjecting it to a treatment with the Coomassie Brilliant Blue dye,

c) controlling the absence or a decrease of blue color in the matrix where sericin has been removed, and let it dry,

d) bringing to dryness the aqueous solution containing sericin or, alternatively, freeze-drying it,

e) weighing the matrix where sericin has been removed obtained from step c) and the sericin obtained from step d).

Example 8

Preparation of electrospun matrices comprising fibroin, sericin and sodium hyaluronate

It is carried out as described in Example 2, by adding 2% (g 100 ml) of sodium hyaluronate to the solution to be electrospun. A matrix of fibroin, sericin and sodium hyaluronate is thus obtained. Example 9

Solubilization assays of the native cocoons and comparison assays on the sericin release by the native cocoons and the matrix of the invention.

Example 9.1

An attempt to solubilize the cocoons as such (i.e. in the native state, i.e. not previously subjected to any treatment) in a solution of formic acid and calcium chloride (as stated in Example 1) has been done: even after 24 hours under stirring the cocoons are not completely dissolved, as it can be seen from the image shown in Figure 6.

Example 9.2

The sericin release has been evaluated by introducing the same amount of cocoons as such and of matrix according to the invention, which consists of fibroin and sericin only, in water at 37°C for two hours: it has been verified that the sericin released from the matrix was 500% higher than that released by the cocoons.

Claims

1. A fiber comprising fibroin and sericin, said fibroin and sericin being present in the form of mixture; said fiber optionally comprising at least one further active component in the cosmetic or medical field when applied topically.

2. The fiber according to claim 1, wherein the heavy chain of fibroin has a molecular weight greater than 200, preferably greater than 300 and more preferably greater than 320 kDa and the water-soluble sericin fractions have molecular weight greater than 80 kDa, preferably greater than 100 and more preferably greater than 130 kDa.

3. The fiber according to claim 1 or 2, wherein the fibroin/sericin weight ratio in the matrix is in the range from 60/40 to 95/5, from 70/30 to 95/5, preferably from 75/25 to 92/8, more preferably from 85/15 to 90/10.

4. The fiber according to any one of claims 1 to 3, wherein said further components are selected from hyaluronic acid or a salt thereof which is acceptable for topical application, collagen, Vitamins, e.g. Vitamin C and vegetable extracts.

5. The fiber according to any one of claims 1 to 3, wherein said fiber is a mixture essentially consisting of fibroin and sericin.

6. A fiber matrix containing a mixture of fibroin and sericin according to one of claims 1 to 5.

7. The fiber matrix according to claim 6, wherein said matrix is coupled to one or more layers of support materials, said materials being preferably selected from viscose, polyester, cellulose, cotton fibers, foams, gel forming materials, gauze, polyurethanes and the possible mixtures thereof.

8. The matrix according to claim 6 or 7, wherein said matrix comprises microcapsules containing at least one of said further components.

9. A process for the preparation of a fiber as defined in any one of claims 1 to 5, or of a fiber matrix according to one of claims 6 to 8, characterized in that it comprises the steps of: treating the starting fibroin under acidic or basic environment with binary salts; mixing said treated fibroin with sericin in an acidic solution, to form a fibroin and sericin solution, adding to said solution the possible further components and electrospinning said solution to give fibers comprising a mixture of sericin and fibroin.

10. The process according to claim 9, wherein said binary salt is calcium chloride.

11. The process according to claim 9 or 10, wherein the fibroin/sericin weight ratio in the matrix is in the range from 60/40 to 95/5.

12. The process according to claim 11, wherein said weight ratio is in the range from 70/30 to 95/5, preferably from 75/25 to 92/8, more preferably from 85/15 to 90/10.

13. The process according to any one of claims 9 to 12, characterized in that said electrospinning step is carried out at a temperature between 20 and 30°C and under environment with relative humidity of 30-40%.

14. The fiber matrix containing fibroin and sericin, characterized in that it is obtained by the process according to any one of claims 9 to 13.

15. Use of the fiber according to one of claims 1 to 6, and/or of the matrix according to any one of claims 7-8 or 14, in the cosmetic field.

16. The fiber according to one of claims 1 to 5 and/or matrix according to any one of claims 6-8 and 14, for its use in therapy.

17. The fiber according to one of claims 1 to 5 and/or matrix according to any one of claims 6-8 and 14, for its use in therapy.

18. A manufactured product comprising a fiber according to one of claims 1 to 5 and/or a matrix according to any one of claims 6-8 and 14.

19. The manufactured product according to claim 18, which is selected among compresses, masks for the skin of the face, neck, hands and other parts of the body; dressings; patches; gauzes; bandages and bandaging; diapers, sanitary pads and incontinence pads.