Adhesive coating intended for tissue engineering and bone regeneration

Abstract

The present invention relates to an adhesive coating intended for tissue engineering and bone regeneration that comprises hydroxyapatite (HA) synthesised from Carbocal from the sugar industry and having strong binding mediated by a commercial medical-grade epoxy adhesive. The adhesive coating is an osteoinductive composite material with biocompatible characteristics as of the first 24 hours after use on implantable surfaces or as independent bone filler. In the latter case, it is a material that solidifies in less than three hours and, given that it exhibits mesopores throughout the volume thereof, accepts blood infiltration from the patient who is being surgically operated on.

Description

[0001] DESCRIPTION

[0002] ADHESIVE COATING ORIENTED TO TISSUE ENGINEERING AND BONE REGENERATION.

[0003] TECHNICAL SECTOR

[0004] The present invention belongs to the field of adhesive technology, chemical engineering and biomedical engineering.

[0005] The technical object consists of a biomaterial for the priming process of metallic and non-metallic implantable surfaces, formed by a mixture of an epoxy-type adhesive and hydroxyapatite (HA) obtained from a by-product of sugar beet processing, as well as for application as an adhesive bone filler, which allows providing osteoinductive mass and a protective barrier for metallic implantable prostheses in the biomedical, traumatological, orthopedic and oral dental industries.

[0006] BACKGROUND OF THE INVENTION

[0007] Any foreign body in the body promotes the development of infections and, consequently, rejection (Wood et al., 2015). This phenomenon is particularly common with dental implants, which are covered by saliva, whose glycoproteins promote colonization by microorganisms (especially streptococci), which proliferate in the oral cavity (Siqueira et al., 2012); (Subramani et al., 2009).

[0008] A similar situation occurs with prostheses in traumatology (for example, in total knee arthroplasty (TKA) or hip THA), whose materials used have been based on stainless steels, titanium alloys, Cr-Co-Ni alloys, Vitalium® and other metals hazardous to humans (Keegan et al., 2007). In this case, the problem arises from a lack of demonstrated medium- or long-term biocompatibility. The goal of these pieces is to increase their useful life and facilitate their integration into the bone by incorporating a cemented material into the implantable metallic surfaces, which serves as a true osteoinductive barrier and protects against infection. Despite achieving a high success rate in these interventions, 19% of patients still express dissatisfaction with the interventions they have undergone, leading to a revision (Bourne et al., 2010).

[0009] The literature has described coating methods based on the application of zirconium or aluminum nanoparticles to improve the osseointegration of implants (Salou et al., 2015), including antimicrobial chlorhexidine-hexametaphosphate (CHX-HMP) nanoparticles. These nanoparticles promote good biological fixation in the implants and enhance the antibacterial effect on the bone (Wood et al., 2015). In fact, most failures in this type of implants are due to poor stability of the piece caused by bone loss that causes it to become unstable. It is possible to find references that use bioactive glass nanoparticles using calcium precursors using Sol-Gel techniques (Firzok et al., 2019).In addition, abrasive treatment processes have been described on the metallic surfaces of implants to make them rougher (1-2 pm) and, thus, more osteointegrable, but the results are debatable and these abrasives leave particles deposited on the implants that behave as contaminants (Schupbach et al., 2019).

[0010] Similarly, adhesives have been used to improve screw-implant and screw-abutment (interface) threaded joints (Bosqué, C.; Azevedo, AD; Arenas, VA; Ávila-Campos, 2019) (Suffo, Vilches-Prez, & Salido-Peracaula, 2020). So that, on the one hand, the threaded tightening is improved in the area of ​​the thread threads where they may remain without contact and, in the upper part where the screw head sits with the prosthetic abutment, in this way the adhesive acts as a retainer of the threaded joint. In this way, the entry of bacteria that induce pathologies is prevented (Caldas et al., 2019).

[0011] Furthermore, the sugar industry based on beet, regardless of its variety (Mahmoud et al., 2018), generates high volumes of by-products / waste, reaching 80% of the original raw material (Nanou & Fuentes, 2002). Approximately 140 g of sugar can be obtained from each ton of beet; the rest is valuable by-products / waste (Canales et al., 2004). These by-products, which are suitable for valorization as part of other value-added products, include "Carboca!®", "dried pulp", and "stem leaves" (plant matter). Currently, these by-products / waste are used as animal feed, including in the production of cement for the construction sector, but a large fraction of these usually end up in landfills.

[0012] The scientific literature provides different studies that validate the use of a good number of agricultural residues as additives or reinforcements of synthetic thermoplastics for different industrial sectors (Ramesh et al., 2017), (Váisánen et al., 2016), (Saba et al., 2016). Among the most used additives in the updated bibliography, it is worth mentioning: calcium carbonate (CaCOs), glass fibers, talc, kaolin, mica, wollastonite, silica, graphite, synthetic fillers (e.g. PET or PVA based fibers), high performance fibers (carbon, aramid, etc.) (Ozen & §¡m§ek, 2015), and reinforcing fibers (hemp, jute, sisal, kenaf, bamboo, etc.) (Gurunathan et al., 2015), (Váisánen et al., 2016).Additionally, patent records have been found that mix dried pulp in polymeric matrices for the manufacture of packaging compounds in the food industry [W02011114101A1], [EP0506650A2], [US2017015820A1], [W02014074086A1], [AU2017200475A1], However, no bibliographic references or registered patents have been found referring to the use of Carbocal in order to guarantee a homogeneous mixture together with an adhesive resin in order to obtain a biomatehal for priming implantable surfaces, as a protective barrier for these surfaces. On the other hand, the reference (Suffo, Mata, et al., 2020) describes the manufacture of an agrocomposite® from Carbocal® and a synthetic thermoplastic for the food industry.

[0013] A previous study (Boyetey et al., 2023) describes the production of a hydroxyapatite derived from sugarcane filter cake. This cake has similar properties to Carbocal, however, the latter has a higher percentage of CaCOs. Furthermore, the process described here uses sodium dihydrogen phosphate monohydrate as a phosphorus source, while the previous reference uses orthophosphoric acid. Finally, the hydroxyapatite obtained in that reference is produced by a precipitation process, while the one presented here is obtained by a phosphatization process.

[0014] The patent Keller & Bohacek GmhH, 1988, ES2003166 (A6), describes a process for obtaining enzymes from beet peelings, but there is no further scientific contribution to the valorization of by-products / waste, nor as part of the aforementioned composite mixtures. In addition, there are numerous studies in the scientific literature on CaCOs (main basic constituent of Carbocal), as an additive in thermoplastics such as (RN Rothon, 2003), (Ozen & §¡m§ek, 2015). Finally, the reference (Kashyap & Datta, 2017) carries out a feasibility study of CaCOs-rich waste from the paper industry as a filler for thermoplastics, justifying the improvement of the original properties of the polymeric matrix.

[0015] Since the first studies carried out by (Roy et al., 1974), it was proven how nature has provided biomaterials that through physical-chemical processing, structured biocompatible compounds such as HA have been generated (Tampieri et al., 2009) (Parfen'eva et al., 2006), in a temperature range between 100-1000°C, between 14-140 MPa of pressure, and a pH between 5 and 12. The reference (Ruffini et al., 2013), transforms rattan wood into HA in a 96-hour process, with a basic pH and at low temperatures (20-60°C). Through hydrothermal synthesis, the well-known chemical reaction of phosphatization of CaCO3 gives rise to bioceramic structures of calcium phosphate (HA) and beta-tricalcium phosphate (P-TCP) and biphasic calcium phosphate (BCP) within a specific range of temperature, pressure and pH; also known in the literature (Roy & Linnehan, 1974), (Roy et al., 1974). According to reference (Yoshimura et al., 2004), the best mechanism for HA formation from CaCOs is precipitation / dissolution. These compounds are marketed at a high market price, approaching €400 / 100g, and depending on the manufacturing stages, this will add even more economic cost to the final compound, as well as the energy consumed throughout the process. Furthermore, clinical practice currently requires a large amount of bone filler, with approximately 6,000 such procedures performed annually in Andalusia alone, which represents a high cost.However, Carbocal®, as described above, being a by-product from the processing of beet sugar, has a low market price and contains more than 80% CaCOs, apart from some trace elements that make it more attractive for use in biomedical applications and no patent records have been found referring to such applications, nor as part of an adhesive mixture for priming implantable surfaces.

[0016] Similarly, adhesive products for medicinal use based on cyanoacrylates have been registered [US2293969A], [US2467927A], [US2768109A]; [US5684042]; [US3483870]; [US5580565]; [US4764377]; [US4892736]; [US4940579]; [US5,254, 132]; [WO 01 / 32319 A2], with a longer or shorter polymerization period and forming compounds together with therapeutic agents or antibiotics. There are also those from the acrylic family and, finally, epoxy resins [WO2019181721 (A1)]. The aforementioned reference (Suffo, Vilches-Pérez, et al., 2020) compares the adhesion strength of some commercial medical adhesives in dental applications. This reference concluded that the adhesive known as LOCTITE M31CL exhibited the best adhesion strength performance on materials used in additive manufacturing (3D printing) such as ULTEM1010™, a biocompatible rigid thermoplastic from the PEI (Polyetheremide) family.[US6455064B1] describes a method for applying a polymerizable adhesive monomer to a biologically active medicament that acts as an adhesive polymerization initiator. The result is an adhesive compound that is applied to wounds as a replacement for sutures. Since the 1960s, acrylic cements and those based on polymethyl methacrylate (PMMA) mixtures have been used. These cements polymerize rapidly and are applied directly to the implant itself. Unfortunately, bone loss is common in areas where prostheses made of these materials have been implanted, mainly due to corrosion.

[0017] The implant surface is considered to be the surface that will be in direct contact with oral tissues, either soft tissue (gum and connective tissue) or hard tissue (bone). The biology of both is completely different, so the part of the implant that will come into contact with them has been modified to allow for greater interconnection between them. Thus, implant surfaces can be classified as smooth or treated. The smooth surface is one that has not undergone any processing other than that required for manufacturing, from the turning of the bar. It is almost flat, with only the roughness caused by the milling process, and has a lower impact on bone formation. It is used to be the surface in contact with the soft tissues, in order to achieve a possible union with them and an area that is easier to clean. The treated surface, on the other hand, is irregular and seeks to connect with the bone tissue.The pattern obtained depends on how the smooth surface is modified to achieve it. This can be achieved by additive methods (plasma, material coating, anodizing, etc.), subtractive methods (shot peening, acid etching, laser, etc.), oxidation, or a combination of vapors. Implants obtained through synthesis (metal 3D printing) always have a rough surface and must be refinished to smooth it. Coatings are usually porous, based on hydroxyapatite (HA), and produced by diffusion and plasma spraying (Heimann, 2016). Among other technologies developed to modify surfaces and coat metallic implants, chemical and electrochemical treatments stand out, such as electrophoretic deposition (Balamurugan et al., 2009), thermal spray techniques (Cañas et al., 2016) and the sol-gel method (Sergi et al., 2020).Unfortunately, some of these technologies present disadvantages, such as a weak bond between implants and coatings, modification of the properties of the metallic implant or the coating and the presence of impurities. For example, reference [RU2684617C1] describes a method for applying a bioactive coating on titanium implants; the coating is an electrolyte composed of orthophosphoric acid, hydroxyapatite (HA, Cas(PO4)3(OH)), germanium and distilled water. Reference (Jun et al., 2010) refers to a hybrid coating material consisting of a silicon and chitosan xerogel for coating titanium implant surfaces. Also, (Kim et al., 2015) disclose a method to obtain material based on a silica aerogel with an epoxy resin, while, (Guzel Kaya et al., 2018) describe the synthesis of a material based on a silica xerogel with an epoxy resin.

[0018] More recently, (Palla-Rubio et al., 2019) performed a sol-gel coating on titanium implants based on silica-chitosan using a dipcoating process. However, to achieve a homogeneous solution, small amounts of chitosan are required, since increasing the concentration of the biopolymer makes the solution too viscous and difficult to adhere.

[0019] Finally, (Reyes-Peces et al., 2020) refer to a material for bone regeneration, consisting of a TEOS aerogel, chitosan (4-20% w / w chitosan) and glycidopropoxymethylthethoxysilane (GPTMS), obtained by sol-gel synthesis. Additionally, the reference (Perez-Moreno et al., 2020) discloses the synthesis and method of a hybrid aerogel of silicon (TEOS) and 8% chitosan for use in tissue engineering. As a result of these last two works, the recent patent P202130841 was requested (Suffo, M.; Piñero, M.; Salido M.; Fernández-Montesinos R.; Reyes-Peces M.V.;De la Rosa, 2021), which describes a hybrid adhesive / sol-gel coating material characterized in that it is composed of a sol-gel material (aerogel or xerogel), consisting of tetraethylorthosilicate (TEOS) and chitosan (4-20% by weight), with solid bonding mediated by a commercial medical-grade epoxy adhesive, mixed in certain proportions for application on metallic and non-metallic implantable surfaces, in tissue and biomechanical engineering. In the explanation of the invention of said patent, bioactivity is demonstrated by introducing the samples into simulated biological fluid and verifying that HA grows on its surface after 7 days. Said patent evidences a manifest invention but the manufacturing process of the described sol-gel is laborious and expensive, in addition to not providing more inventiveness than hybridization with the adhesive in a solid bond, since the manufacture of aerogels is well known in the literature.However, lattice shear tests or trials according to the LINE-EN ISO 2409 standard, which confirm correct fixation of the coating on the implantable surface, are not indicated. Nor are the sizes and morphologies of the HA crystalline structures formed specified, considering that micro / nanocrystalline structures with a Ca / P ratio of 1.67 are the most suitable for manufacturing biomimetic biomaterials similar to human bone (Lin et al., 2014).

[0020] Additionally, the literature reviewed does not report cases of obtaining a biomaterial for priming metallic or non-metallic implantable surfaces, consisting of a mixture of HA and an epoxy resin-based adhesive, such as the one described in this invention. This biomaterial potentially plays a bioactive role and has the capacity to induce cell nucleation and growth on the surface of the biomaterial, regardless of whether it is metallic or non-metallic. Furthermore, no references have been found that mention HA synthesized from byproducts of sugar beet processing.

[0021] The hybrid adhesive biomaterials and biocomposites resulting from the processing described in this report avoid the main problems that arise with implants based on metallic materials such as stainless steel, titanium alloys or chromium-cobalt-nickel, which do not guarantee bone formation and cause problems in the vicinity of surgery and, above all, corrosion phenomena.

[0022] Current prostheses used in traumatology and dentistry, in general, are not designed to receive a coating like the one described in this report. At most, they use bone cement as a protective barrier against the metallic material. Tests conducted on metallic and non-metallic surfaces designed to test the effectiveness of the coating showed similar results.

[0023] The invention demonstrates the possibility of coating non-bioactive and non-biodegradable metallic surfaces on implantable elements with these characteristics, but with the guarantee of non-detachment. References used

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[0066] EXPLICACIÓN DE LA INVENCIÓN

[0067] The invention develops a biomaterial composed of hydroxyapatite (HA) synthesized from a byproduct of sugar beet processing known as Carbocal, with solid bonding mediated by a commercial medical-grade epoxy adhesive, for use in primers that prepare implantable surfaces to make them osteoinductive.

[0068] The bioactivity of the biomaterial is demonstrated by a cytotoxicity assay, which shows cell growth at almost the same level as the positive control on its surface within the first 24 hours. The correct adhesion of the biomaterial for proper priming is demonstrated by the lattice shear test according to the UNE-EN ISO 2409 standard. This results in a biomaterial for priming implantable surfaces that meets the three basic requirements for osteoinduction: biocompatible, bioactive, and biodegradable. This biomaterial is used in tissue engineering as a scaffold and also as an adhesive filler that promotes bone regeneration.

[0069] The proposed surface-priming biomaterial, which is the result of a mixture of HA synthesized from Carbocal and a commercial medical-grade epoxy resin, begins with the synthesis of HA in a hydrothermal reactor. The sample prepared from Carbocal residue exhibits a biphasic character, which is important within the framework of the concept of bioactivity proposed by Daculsi in France and Lynch, Nery, and LeGero in the USA in the 1980s (Daculsi et al., 1989). As is well known, this concept was based on the unique properties of biphasic calcium phosphate ceramics (BCPs). BCP bioceramics consist of a mixture of hydroxyapatite (HA) (more stable) and beta-tricalcium phosphate (P-TCP) (more soluble), with a variable HA / b-TCP ratio. This material, structurally similar to the HAcc prepared in this work, gradually dissolves in the body, generating new bone formation as it releases Ca2+ and PO43- ions into the biological medium.Furthermore, both phases are biocompatible, an essential property to avoid adverse reactions and promote integration with the surrounding tissue. BCP is commercially available in Europe, Brazil, Japan, the USA, and Australia as a bone graft or bone substitute material for orthopedic and dental applications under several brand names (BCP®, MBCP®, Triosite®, Hatric®, Eurocer®, Biceram®, Bicalfoss®).

[0070] The hydrothermal technique was selected for the synthesis of the material due to its simplicity, speed, and, above all, low cost. In this particular case, Carbocal powder from the owner company (AB Azucarera) will be used, as it comes from the sugar refining process. Applying the well-known recipe for hydrothermal phosphatization of CaCO3 (Tampieh et al., 2009) (Yoshimura et al., 2004), but using a sodium salt instead of potassium and respecting the molar ratio (CO3 / PO4 = 5 / 3), the process will be carried out under identical conditions.

[0071] BRIEF DESCRIPTION OF THE DRAWINGS

[0072] Figure 1. Results obtained in the cell viability assay.

[0073] PREFERRED EMBODIMENT OF THE INVENTION

[0074] Two comparative tests with the same characteristics are carried out, that is, 9 grams of Carbocal (80% w / w) and 6 grams of sodium hydroxyphosphate are mixed in 100 ml of water in a 200 ml beaker until both reagents are completely dissolved. At this point, a constant bubbling of CO2 is observed as a result of the reaction taking place. The mixture is left stirring for 2 hours to complete the reaction and then the resulting pH is measured (approx. 7).

[0075] Using separate portions of water, the product is transferred to a 250 ml Teflon-coated vessel, reaching a volume of 80% of its nominal capacity (200 ml in total). It is sealed in the steel hydrothermal reactor and transferred to the oven at 200 ° C for 24 hours. Once cooled to room temperature, the contents of the vessels are extracted, washed, centrifuged (5000 rpm for 5 minutes), the liquid residue is removed and dried in an oven at 120 ° C for 24 hours. Once the material is dry, it is ground in an agate mortar and sieved through a 75-micron mesh size to obtain a homogeneous powder. The results of both tests are identical, and a total of 14.15 grams of reaction product are obtained.

[0076] The HA obtained has a biphasic character: 75% w / w HA and 24.83% w / w p-TCP, which is beneficial for its biocompatibility, as previously explained. In the FTIR analysis of HA, a peak corresponding to the presence of CO3 groups is observed, which are not present in commercial HA analysis. Therefore, the synthesis of HA from Carbocal has another benefit, since the natural HA present in bones also has these CO3 groups.

[0077] Table 1. Density and specific surface area results

[0078] The synthesized HA powder is then manually mixed with the biocompatible epoxy adhesive until a greenish-brown biomaterial with a homogeneous texture is obtained.

[0079] The adhesive used is a medical grade epoxy called LOCTITE® M31 CLTM from the manufacturer HENKEL IBÉRICA, SA. It was selected from among 5 products based on the results obtained from the adhesion test carried out in the aforementioned reference (Suffo, Vilches-Pérez, et al., 2020). It is a two-component adhesive based on a resin and a hardener (2:1 ratio, resin:hardener), the mixture of which is almost transparent in color, has low viscosity and a specific gravity of 1.07 kg / cm3 at 25°C. The mixture with the adhesive was made following different proportions for HA as shown in Table 2, which affects its behavior. These mixtures cure in 90 minutes at 30°C and have withstood the autoclave sterilization process up to 80°C without deformation or phase changes.

[0080] Table 2. Proportions used for the biomaterial

[0081] *Use of CHA doped with beet pulp

[0082] Finally, cell cultures were performed with human fetal osteoblastic cell lines (hFOB 1.19) in the presence of the generated biomaterials to determine whether the materials affect cell viability. The hFoB (Human Fetal Osteoblasts) cell line is an established cell line widely used in research related to bone biology and osteogenesis, as it is derived from human fetal bone tissue. Being osteoblasts, i.e., cells specialized in the formation and mineralization of bone tissue, allows them to be used in research related to bone regeneration.

[0083] For this cell viability study, hFOB were cultured in osteogenic media, and viability assays were performed at 24, 48, 72 hours, and 7 days. In this assay, cells were incubated with MTS, and absorbance was subsequently quantified. As a negative control, hFOB cultures were incubated with 70% methanol for 30 minutes before the procedure. Untreated hFOB cultures were used as a positive control.

[0084] Normal human fetal osteoblasts were purchased from ATTC (hFOB cell line

[0085] 1.19, CRL-11372TM). hFOB cultures were grown in osteogenic media containing Dulbeccos' modified Eagle's biotin (DMEM - Gibco), 10% fetal bovine serum (FBS - Corning) and antibiotic (G418 30 mg / uL - PAN Biotech). Cells were expanded by incubation at 35°C in 75 cm2 flasks with 5% CO2.

[0086] Viability assays were performed for 7 days. As a negative control, hFOB cultures were incubated with 70% methanol for 30 minutes before the labeling process. Untreated hFOB cultures were used as a positive control.

[0087] Before testing, the materials were sterilized using an autoclave at 120°C for 20 minutes. For the viability assay, cells were cultured with complete medium in 96-well plates at a concentration of 2 x 10 4 cells / well. These plates were incubated at 35°C with 5% CO2 for 24 hours. In the case of positive control (PC) and negative control (NC), the wells were allowed to grow without adding anything, while in the wells corresponding to the tests, the different biomaterials were added.

[0088] To measure viability, the MTS assay was used. Before labeling, as a negative control, cells were incubated with 70% methanol for 30 minutes. Twenty ul of MTS was added to all wells and incubated at 34°C for 1 hour. After this time, absorbance at 490 nm was measured using the Varioskan LUX plate reader. This process was performed at 24, 48, 72 hours, and 7 days of incubation with the different HA sample solutions. The test was carried out in n = 3 replicates, repeating the test on 3 different samples.

[0089] In addition, the LIVE / DEAD cytotoxicity assay was performed to determine whether cells exhibited growth adherent to the surface of the tested biomaterials. To do this, cells were grown on the surface of the materials under the same conditions described above for 24 hours. After this time, the reagents Calcein (3.5 uM) and EthD-1 (7 uM) were added, and the cells were incubated for 1 hour at 37°C. For the positive control, cells were grown without the presence of any biomaterial. For the negative control, the same materials were used, treated with the reagents, without cell culture. The biomaterials, as well as the controls, were observed under a fluorescence microscope to determine whether there was growth of adherent cells on the surface.The results obtained in the viability test can be seen in Figure 1 , where the cell viability of each material is expressed as a percentage, compared to the positive control, considered as 100%. The negative control never exceeded 30% viability. It is observed that all the proportions used are biocompatible, with mixtures VI and VII showing the highest viability at 7 days. It is observed how, when beet pulp is used as an additive to HA, the required proportion of this, with respect to the epoxy, is lower to achieve the same viability (0.83:1). Therefore, the HA:epoxy ratio range that maintains viability is > 0.10:1 and < 1.22:1.

[0090] The LIVE / DEAD assay revealed that cells grow adherent only on the VI and VII ratio, thus being the most optimal for the established purpose.

[0091] How the invention is susceptible to industrial application

[0092] Currently available bone coatings are dispensed independently of the prosthesis. Commercially available medical-grade epoxy adhesives, such as LOCTITE M31CL, are dispensed in double-fluted cans for application with a gun. The application originating from the present invention in the biomedical sector must be dispensed together with the prosthesis itself, with the appropriate coating dose, or separately in sterilized, plastic-coated sachets, ready for direct application to an implantable surface or as a bone filler to be applied to the area, admitting a few drops of blood serum prior to application.

Claims

CLAIMS 1. Adhesive coating oriented to tissue engineering and bone regeneration, characterized by the fact that it is composed of a hydroxyapatite (HA) synthesized from the byproduct of the Carbocal sugar industry, biphasic with an HA phase and a TCP phase, with solid bonding mediated by a commercial medical-grade epoxy adhesive.

2. Adhesive coating, according to claim 1, characterized in that the HA has a biphasic character: 75% w / w HA and 24.83% w / w p-TCP.

3. Adhesive coating, according to claims 1 and 2, characterized by presenting cell viability, according to the HA:epoxy adhesive ratio, in the following range: > 0.10:1 and < 1.22:1 (w / w).

4. Adhesive coating, according to claim 3, characterized by presenting the maximum cell viability, according to the HA:epoxy adhesive ratio, at 1.22:1 (w / w).

5. Adhesive coating, according to claim 3, characterized by presenting the maximum cell viability, according to the HA:epoxy adhesive ratio, at 0.83:1 (w / w), when using a HA doped with beet pulp.

6. Use of the adhesive coating, according to claims 1 to 5, for its application in tissue engineering and biomechanics.

7. Use of the adhesive coating, according to claims 1 to 5, for application on surfaces of implantable elements.

8. Use of the adhesive coating, according to claims 1 to 5, for application in metal prostheses.

9. Use of the adhesive coating, according to claims 1 to 5, for application in non-metallic prostheses.

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