WO2007093662A1

WO2007093662A1 - Eugenol-derived acrylic polymers and monomers, formulations and compositions containing same and biomedical uses thereof

Info

Publication number: WO2007093662A1
Application number: PCT/ES2007/070031
Authority: WO
Inventors: Luis Rojo Del Olmo; Maria Blanca Vazquez Lasa; Julio San Roman Del Barrio; Sanjukta Deb
Original assignee: Consejo Superior De Investigaciones Científicas
Priority date: 2006-02-15
Filing date: 2007-02-14
Publication date: 2007-08-23
Also published as: ES2303430B1; ES2303430A1

Abstract

The invention relates to eugenol-derived monomeric compounds, the synthesis of polymers and copolymers from the derivatives obtained and the development of self-curing acrylic formulations. Said systems have analgesic and antiseptic properties originating from the eugenol molecule that is chemically anchored to the macromolecular chains. The aforementioned self-curing formulations can be applied directly when used as dental resins or can be injected when used in the treatment of osteoporotic fractures in minimally-invasive surgery.

Description

TITLE

MONOMERS AND ACRYLIC POLYMERS DERIVED FROM EUGENOL, FORMULATIONS AND COMPOSITIONS CONTAINING THEM AND THEIR BIOMEDICAL APPLICATIONS

SECTOR OF THE TECHNIQUE.

This invention is part of the self-healing formulations that are used as controlled release systems for medications in minimally invasive surgery and dental applications.

STATE OF THE TECHNIQUE

Eugenol (4-allyl-2-methoxyphenol) is a product of natural origin that is found as the main component of essential oils, specifically it is the main constituent of clove essence. This compound has analgesic and antiseptic properties, being one of the most commonly used products for pain relief of irritated or diseased dental pulp (Sticht FD, Smith PM. "Eugenol: Some pharmacologic observations". J Dent Res 1971, 50, 1531-1535). Low concentrations of Eugenol have anti-inflammatory effects on dental pulp but high concentrations can be cytotoxic. Eugenol exhibits antimicrobial and anti-aggregating activity as demonstrated in vitro, as well as an action on platelet function in vivo (Laekeman GM, Van Hoof L, Haemers A, Vanden Berghe DA, Herman AG, Vlietinck AJ. "Eugenol a valuable compound for in vitro experimental research and worthwhile for further in vivo investigation ". Phytother Res 1990, 4, 90-96). It also has an antipyretic action when administered intravenously and can reduce fever through a central action (Feng J, Lipton JM. "Eugenol: Antipyretic activity in rabbits." Neuropharmacology 1987, 26, 1775-1778). Likewise, antianaphylactic properties have been described for Eugenol (Kim HM, Lee EH, Kim CY, Chung JG, Kim SH, Lim JP, Shim TY. "Antianaphylactic properties of Eugenol". Pharmacol Res 1997, 36, 475-480) . Eugenol, like any phenolic derivative, is very reactive against free radicals and can inhibit lipid peroxidation at the initiation stage (Okada N, Satoh K, Atsumi T, Tajima M, Ishihara M, Sugita Y, Yokoe I, Sakagami H , Fujisawa S. "Radical modulating activity and cytotoxic activity of synthesized Eugenol-related compounds". Anticancer Res 2000, 20, 2955-2960).

Eugenol has been used in combination with zinc oxide to give rise to ZOE (zinc oxide-Eugenol) cements that are used as pulp covering agents in temporary cements and as root canal filling cements (Fujisawa S, Kashiwagi Y, Atsumi T, Iwakura I, Ueha T, Hibiro Y, Yokoe I. "Application of bis- Eugenol to a zinc oxide Eugenol cement". J Dent 1999, 27, 291-295). Cement is formed by reacting zinc oxide with Eugenol to give rise to zinc eugenolate, a reaction that needs the presence of water and can be accelerated with the presence of zinc acetate. In these formulations, Eugenol acts as a sealing agent by softening the irritated pulp. However, this type of cement usually produces an inflammatory reaction in vivo, probably due to the release of Eugenol, which in certain concentrations can cause irritation in the tissues and induce an inflammatory reaction on the membrane of the oral mucosa. The use of ZOE cements as provisional cements has to be carefully considered since Eugenol, due to its antioxidant character, inhibits the radical polymerization of most resins that set or harden by radical route. Studies taken to In order to determine the mechanism of the action of Eugenol as an inhibitor, they conclude that the inhibitory effect of Eugenol is governed by the molar ratio of Eugenol to benzoyl peroxide (BPO) (Fujisawa S, Kadoma Y. "Action of Eugenol as a retarder against polymerisation of methyl methacrylate by benzoyl peroxide ". Biomaterials 1997, 18, 701-703). In relation to this phenomenon, there are numerous references that describe the harmful action of temporary cements containing Eugenol on permanent cements based on acrylic resins, leading to a worsening of the physical properties of the resins (Hotz P, Schlatter D, Lussi A. "The modification of the polymerisation of composite materials by Eugenol- containing temporary fillings". Schweiz Monatsschr Zahnmed 1992, 102, 1461-1466).

Regarding the incorporation of Eugenol to macromolecular chains, the references found in the literature are scarce. Mention should be made of the preparation of methyl methacrylate copolymers with Eugenol derivatives bearing hanging urethane groups (Tyagi AK, Choundary V, Varma IK. "Copolymerisation of methyl methacrylate and a novel allyl monomer." Eur Polym J 1992, 28, 419- 22).

DESCRIPTION

Brief description of the invention.

The present invention is related to the preparation of monomeric compounds derived from Eugenol, the synthesis of polymers and copolymers from the derivatives obtained as well as the development of self-healing acrylic formulations bearing Eugenol with acrylic and polyacrylic components. These systems have analgesic and antiseptic properties from the chemically anchored Eugenol molecule to macromolecular chains. These self-healing formulations can be applied directly when used as dental resins, or they can be injected when used in the treatment of osteoporotic fractures in minimally invasive surgery.

Detailed description of the invention.

The present invention is based on the fact that the inventors have developed a monomeric acrylic compound derived from Eugenol of general structure such as that presented in Formula (I), capable of polymerizing and copolymerizing with acrylic monomers and of being part in liquid phase compositions of self-healing formulations. On the other hand, the polymers or copolymers thus obtained can be used in the preparation of the solid phase composition of self-healing formulations alone or in conjunction with other commercial polymers. In relation to these polymers it was observed that cell viability is not affected by the presence of the extracts of any of these released polymers or copolymers (see Example 6).

By mixing both solid and liquid phases of any of the self-curable formulations, and the consequent radical polymerization of the liquid phase, a fluid paste capable of being injected, or applied directly and finally cured "in situ" is obtained. These self-healing formulations can be applied directly when used as dental resins, or they can be injected when used in the treatment of osteoporotic fractures in minimally invasive surgery. The self-healing systems thus obtained have the following advantages:

1.- Eugenol's self-healing formulations reach peak or maximum temperatures, lower than those of commercial compositions in the process of curing or polymerization. This reduction in temperature could potentially reduce the damage caused to adjacent tissues during "in situ" hardening.

2. - The self-healing formulations carrying Eugenol, chemically anchored to the macromolecular chains, offer the advantage of having a therapeutic action as analgesics and antiseptic in situ.

3.- The most outstanding feature of Eugenol's self-healing formulations is that they offer the advantage of having the Eugenol molecule chemically anchored, which is expected to reduce the toxicity caused by the release of it to adjacent tissues and its incorporation into systemic circulation

Thus, an object of the present invention is an acrylic monomer derived from Eugenol, hereinafter acrylic monomer of the invention, of general formula (I)

Where: n is 0, 2, 6 or 11

Ri is a methyl, ethyl, propyl, isopropyl, butyl, isobutyl or tert-butyl radical.

R.2 is a methyl, ethyl, propyl, isopropyl, butyl, isobutyl or tert-butyl radical.

R ₃ is a methyl, ethyl, propyl, isopropyl, butyl, isobutyl or tert-butyl radical.

On the other hand, the acrylic monomer of the invention can group several families of compounds. So, an object Particular of the invention is an acrylic monomer of the general formula Ia:

Where :

A particular embodiment of the invention is the acrylic compound of the invention, belonging to the formula Ia, eugenyl methacrylate

(EgMA) (Example 1).

Another particular object of the invention is an acrylic monomer of the general formula Ib

Where: n is 2, 6 or 11.

Another particular embodiment of the invention is the acrylic compound derived from Eugenol, belonging to the formula Ib, ethoxyieugenyl methacrylate (EEgMA) (Example 2).

Another object of the invention is a process for obtaining the acrylic monomer of the invention, hereinafter procedure for obtaining the acrylic monomer, and more preferably of the compounds belonging to the general formula Ia which is carried out under mild conditions and because it comprises the following steps (see Example 1): i) Eugenol is dissolved with triethylamine in stoichiometric amounts, using diethyl ether as a solvent at room temperature, ii) under a nitrogen atmosphere, a stoichiometric amount of methacryloyl chloride is added dropwise to the solution of i) and allowed to react for 48 hours at room temperature under stirring, and iii) the acrylic monomer is isolated and purified as a reaction product. Another particular object is a process for obtaining an acrylic monomer of the invention, preferably belonging to the general formula Ib characterized in that it is carried out under mild conditions and because it comprises the following steps: i) synthesis of 1-hydroxy, n- alkyl Eugenol by treatment of Eugenol with the corresponding OC, Gϋ-chloro-alkyl alcohol in a hydroalcoholic medium, applying a Williamson reaction [March 's advanced organic chemictry (5th Edition) BM Smith and J. March. John Wiley and Sons. New York 2001]. ii) the corresponding 1-hydroxy, n-alkyl Eugenol is dissolved with triethylamine in stoichiometric amounts, using diethyl ether as solvent, iii) under a nitrogen atmosphere, a stoichiometric amount of methacryloyl chloride is added dropwise to the solution from i) and allowed to react for 48 h at room temperature, and iv) the acrylic monomer is isolated and purified as a reaction product.

The purification of the Eugenol acrylic monomer of the invention can be carried out by different techniques, preferably by a silica chromatographic column.

On the other hand, the acrylic monomer of the invention can also be used for the preparation of a polymer or a copolymer carrying Eugenol.

Thus, another object of the invention is a polymer carrying Eugenol, hereinafter acrylic polymer of the invention, comprising an acrylic monomer derived from

Eugenol of the invention, preferably belonging to the formula Ia or Ib. Another particular embodiment of the The invention constitutes a polymer of the invention, by way of illustration and without limiting the scope of the invention, belonging to the following group: poly (eugenyl methacrylate) (PEgMA) and poly (ethoxyethylene methacrylate) (PEEgMA) polymers (see Example 3).

Another particular object of the invention is the acrylic polymer of the invention where the polymer is a copolymer comprising an acrylic monomer derived from Eugenol of general formula (I) and a second acrylic monomer different from the previous one belonging, by way of illustration and without limit the scope of the invention, to the following group: methyl methacrylate (MMA) or ethyl methacrylate (EMA). Another particular embodiment of the invention is a copolymer of the invention, by way of illustration and without limiting the scope of the invention, belonging to the following group: eugenyl methacrylate-ethyl methacrylate (EgMA / EMA) copolymer and methacrylate copolymer of ethyl ethoxynethylene methacrylate (EEgMA / EMA) (see Example 4). Another object of the invention is a process for obtaining the acrylic polymer of the invention comprising a radical polymerization step of any of the monomeric compounds of general formula (I), and is carried out by dissolving the corresponding monomer in toluene, using azobisisobutyronitrile (AIBN) as a radical initiator and at a temperature of 50-60 ° C. In the case of a copolymer, the process for obtaining comprises a copolymerization step in the presence of a radical initiator of any of the compounds of general formula (I) as the first monomer, with a different acrylic monomer as the second belonging monomer, by way of illustration and without limiting the scope of the invention, to the following group: methyl methacrylate (MMA) or ethyl methacrylate (EMA). The Eugenol-derived acrylic monomers and polymers of the invention can be used in the preparation of a self-curable formulation comprising a system of two compositions or phases: a liquid phase and a solid phase. Thus, another object of the invention is a self-curable formulation, hereinafter self-curable formulation of the invention, of acrylic systems derived from Eugenol comprising two compositions or phases: a liquid phase and a solid phase. Another particular object of the invention is a self-curable formulation of the invention in which the self-healing acrylic liquid phase composition comprises one or more of the Eugenol-derived acrylic monomers of the invention, in an amount between 20-60% - p with respect to the total weight of the liquid phase, and a second acrylic monomer in an amount between 80-40% -p with respect to the total weight of the liquid phase belonging, by way of illustration and without limiting the scope of the invention, to the following group: methyl methacrylate (MMA) or ethyl methacrylate (EMA).

In addition, the self-curing liquid phase acrylic composition of the formulation of the invention comprises an aromatic tertiary amine as an activator in an amount between 0.5-2.5% -p, preferably 2% -p, and one or more inhibitors in an amount of up to 0.01% -p. The inhibitor used may, for example, belong to the family of quinones.

Another particular object of the invention is a self-curable formulation of the invention in which the self-curing solid-phase acrylic composition comprises prepolymerized poly (methyl methacrylate) (PMMA), or prepolymerized poly (ethyl methacrylate) (PEMA) particles. copolymers of MMA or EMA with other monomers, present in an amount comprised between 20-80% -p with respect to the total weight of the solid phase. Among other polymers or copolymers that can be used in the solid phase composition are the polymers or copolymers described in the present invention. On the other hand, the self-healing acrylic solid phase composition of the formulation of the invention can also comprise zinc oxide (ZnO) in an amount between 50-80% -p with respect to the total weight of the solid phase (see Example 8). Likewise, the self-curing acrylic solid phase composition may also comprise one or more initiators in an amount of up to 3% -p, for example benzoyl peroxide, and / or one or more radiopaque agents in an amount between 20-25% -p with respect to the total weight of the solid phase belonging, by way of illustration and without limiting the scope of the invention: barium sulfate, zirconium dioxide, tantalum oxide, strontium oxide and organic compounds.

Another particular embodiment of the invention is a self-curable formulation of the invention comprising a liquid phase composition with monomers of eugenyl methacrylate (EgMA) and methyl methacrylate

(MMA) in varying proportions, with the compound (4-N, N-dimethylaminophenyl) -methanol (DMOH) as activator, and a solid phase composition based on poly (methyl methacrylate) (PMMA), or similarly with ethoxyethylene methacrylate without or with zinc oxide (see Examples 7 and 8).

These acrylic compositions - liquid and solid - of the formulation of the invention once mixed allow their handling and application for a controlled time and harden or cure, by means of a radical polymerization process of the liquid phase, to give rise to polymeric systems that are They adapt perfectly to dental and bone cavities where they are applied. Finally, another object of the invention is the use of the self-curable formulation of the invention, by mixing the liquid and solid phase compositions that compose it and its direct application or by injection and finally curing "in situ" for reconstruction. , temporary or permanent, dental and bone. Another particular embodiment is the use of the self-curable formulation of the invention in which bone reconstruction consists of a vertebra fixation or biomechanical fixation of osteoporotic fractures in minimally invasive surgery in the field of traumatology and orthopedic surgery.

DESCRIPTION OF THE FIGURES Figure 1.- Diagram of the 95% confidence limit for the values of the reactivity ratios of the EgMA / EMA and EEgMA / EMA copolymers determined by the treatment proposed by Tidwell and Mortimer. Figure 2.- Composition diagram of the EgMA / EMA and EEgMA / EMA systems. The curves correspond to the theoretical diagrams obtained from the values r (EgMA) = 1.48 yr (EMA) = 0.55 for the first system and from the values r (EEgMA) = 1, 22 yr (EMA) = 0.42 for the second system. The points correspond to the composition values in the copolymer obtained experimentally.

Figure 3.- Comparison of dose-response curves for Eugenol (Eg) and eugenyl methacrylate (EgMA) and ethoxyethylene methacrylate (EEgMA) monomers obtained in the MTT test. Each point represents the mean ± standard deviation (n = 8).

Figure 4.- ESEM images (scanning electron microscope) of the colonization of human fibroblasts on the TMX control and on PEMA, PEgMA, PEEgMA, EgMA / EMA 50/50 copolymer and EEgMA / EMA 50/50 copolymer, at 24 and 48 hours after planting.

Figure 5.- Results of the MTT cytotoxicity test for the TMX control and for the systems studied, PEMA, PEgMA (designated Eg), copolymers EgMA / EMA (designated E Eg), PEEgMA (designated EEg) and copolymers EEgMA / EMA (designated E EEg). The mean ± standard deviation (n = 16) is represented. Extracts are collected over a period of 7 days (*: p <0.05, **: p <0.01, ***: p <0.001). Figure 6.- Results of the Alamar Blue test obtained for the fibroblast cultures maintained for 21 days on samples of the PEgMA family (A) and the PEEgMA family (B). The results represent the mean ± standard deviation (n = 16; *: p <0.05, **: p <0.01, ***: p <0.001).

EXAMPLES OF REALIZATION

Example 1.- Synthesis of eugenyl methacrylate (EgMA).

Eugenol is introduced into a three-mouth flask and dissolved in diethyl ether at room temperature.

An equimolecular amount of triethylamine is then added to the solution as the reaction catalyst and constant stirring is maintained. An equimolecular amount of methacryloyl chloride is then added dropwise under a nitrogen atmosphere. Stirring is maintained and the mixture is allowed to react for 48 hours at room temperature. The reaction mixture is filtered to separate the amine hydrochloride formed. Finally, the reaction solid is isolated by solvent extraction under reduced pressure. Eugenyl methacrylate (EgMA) is purified by chromatographic column using 10/90 ethyl acetate / hexane as the mobile phase. The resulting product is characterized by nuclear magnetic resonance

(NMR) using deuterated chloroform (Cl ₃ DC) (5% w / v) as solvent and tetramethylsilane (TMS) as internal reference.

The proton resonance spectrum ( ¹ H-NMR) in CI ₃ DC shows the following signals: δ _H between 7.0 and 6.7 (3H of the aromatic ring), 6.4 and 5.7 (2H, CH ₂ = C (CH ₃ ) CO), between 6.0 and 5.8 (IH, CH = CH ₂ ), 5.1 (2H, CH = CH ₂ ), 3.8 (3H CH ₃ OPh), 3, 4

(2H, CH ₂ Ph) 2.1 (3H, CH ₂ = C (CH ₃ ) CO).

The carbon resonance spectrum 13 ( ¹³ C-NMR) in Cl ₃ DC shows the following signals: δ _c 165.7 (TOO), 151.3 (O-Ar, CHDCH ₃ ), 139.0 (Ar, C ^ OCO), 138.1 (p-Ar, C-CH ₂ -CH =), 137.2 (CH = CH ₂ ), 135.7 (= C (CH ₃ ) -COO), 126.9 (C ^β H ₂ ) = C), 122.4 (ra-Ar, CC _AΓ -O-CO), 120.6 (o-Ar, CC _Ar -O-CO), 116.1 (CH = CH ₂ ), 112.8 (m-Ar, CC _Ar -OMe), 55.7 (CH ₃ O-Ph), 40.1 (CH ₂ -Ph), 18.4 (C ⁰¹ H ₃ ). The purity of the analyzed product is greater than 98% and its chemical structure is shown in the following Figure:

Chemical formula of eugenyl methacrylate (EgMA)

Example 2.- Synthesis of ethoxyieugenyl methacrylate (EEgMA.).

The synthesis of ethoxyethylene methacrylate proceeds in two steps. First, the synthesis of 2-eugenyl ethanol is carried out by applying a Williamson-type reaction by treating Eugenol with 2-chloroethanol in a hydroalcoholic medium according to the following protocol: a solution of Eugenol in ethanol is prepared and mixed with a solution of excess 50% KOH with respect to the Eugenol and 0.5% KI as a co-catalyst, maintaining nitrogen flow. The reaction medium is refluxed for 24 h. The reaction product is purified by chromatographic column with ethyl acetate / hexane 30/70. The second step carries with it the esterification reaction of 2-eugenyl ethanol which is carried out by dissolving it in diethyl ether in a three-mouth flask. An equimolecular amount of triethylamine is added to the solution as the reaction catalyst and constant stirring is maintained. An equimolecular amount of methacryloyl chloride is then added dropwise under a nitrogen atmosphere. Stirring is maintained and allowed to react for 48 h at room temperature. The reaction mixture is filtered to separate the amine hydrochloride formed. Finally, the reaction solid is isolated by solvent extraction under reduced pressure. Etoxieugenyl methacrylate (EEgMA) is purified by chromatographic column with 10/90 ethyl acetate / hexane. The resulting product is characterized by nuclear magnetic resonance (NMR) using deuterated chloroform (CI ₃ DC) as solvent and tetramethylsilane (TMS) as internal reference.

The proton resonance spectrum ( ¹ H-NMR) in CI ₃ DC shows the following signals: δ _H between 6.9 and 6.6 (3H of the aromatic ring), 6.1 and 5.5 (2H, CH ₂ = C (CH ₃ ) CO), 5.8 (IH, CH = CH ₂ ), 5.1 (2H, CH = CH ₂ ), 4.5 (2H, OCH ₂ CH ₂ OPh), 4.2 ( 2H, OCH ₂ CH ₂ OPh), 3.8 (3H CH ₃ OPh), 3.3 (2H, CH ₂ Ph) 1.9 (3H, CH ₂ = C (CH ₃ ) CO).

The carbon resonance spectrum 13 ( ¹³ C-NMR) in Cl ₃ DC shows the following signals: δ _c 167.3 (COO), 149.8 (O-Ar, CHDCH ₃ ), 146.3 (p-Ar , C-CH ₂ -CH =), 137.4 (CH = CH ₂ ), 136.0 (Ar, CHDCO), 134.1 (= C (CH ₃ ) -COO), 126.0 (C ^β H ₂ = C), 120.5 (m-Ar, CC _Ar -CH ₂ ), 115.7 (CH = CH ₂ ), 115.0 (o-Ar, CC _Ar - O-CO), 112, 6 (m-Ar, CC _Ar -OCH ₃ ), 67, 5 (CH ₂ CH ₂ -OPh), 63, 2

(CH ₂ -CH ₂ -OPh), 55, 7 (CH ₃ O-Ph), 39, 8 (CH ₂ -Ph), 18, 0 (C ⁰¹ H ₃ ).

The purity of the analyzed product was greater than 98% and its chemical structure is shown in the following Figure:

Chemical formula of ethoxyieugenyl methacrylate (EEgMA)

Example 3.- Preparation of poly (eugenyl methacrylate) (PEgMA) and poly (ethoxygiene) methacrylate (PEEgMA) polymers.

The polymerization reaction of the corresponding Eugenol derivative is carried out in dissolution of the monomer in toluene (IM) and azobisisobutyronitrile (AIBN) is used as the radical initiator at a concentration of 1% -p with respect to the monomer. The reaction temperature is 50 ° C and the reaction time is 24 hours to obtain high conversion polymers or the reaction time is adjusted to achieve conversions below 10% -p. At the end of the reaction, the reaction mixture is precipitated in hexane, the precipitated solid is filtered, washed successively and dried until constant weighing. The polymers obtained at conversion less than 10% -p are soluble white powders and are characterized by proton nuclear magnetic resonance ( ¹ H-NMR).

The ¹ H-NMR spectrum in Cl ₃ DC of the PEgMA shows the following resonance bands: δ _H between 7.2 and 6.4 (3H of the aromatic ring), 5.8 (IH, CH = CH ₂ ), 5 , 0 (2H, CH = CH ₂ ), 3.6 (3H CH ₃ OPh), 3.2 (2H, CH ₂ Ph) 2.3 (2H, -CH ₂ -C (CH ₃ ) C00) and 1 , 1- 0.9 (3H, -CH ₂ -C (CH ₃ ) COO). The ¹ H-NMR spectrum in Cl ₃ DC of the PEEgMA shows the following resonance bands: δ _H between 6.9 and 6.5 (3H of the aromatic ring), 5.8 (IH, CH = CH ₂ ), 5 , 0 (2H, CH = CH ₂ ), 4.3-4.1 (2H, COOCH ₂ ), 4.1-3.9 (2H, CH ₂ CH ₂ OPh), 3.7 (3H, CH ₃ OPh), 3.2 (2H, CH ₂ Ph), 2.0-1.7 (2H, -CH ₂ -C (CH ₃ ) COO), 1.1-0.9 (3H, -CH ₂ - C (CH ₃ ) COO).

The presence of the bands corresponding to the allylic protons in both spectra and their good correlation with the bands of the aromatic protons, indicate that at low conversions only the acrylic double bonds participate in the polymerization reaction, giving rise to hydrocarbon macromolecular chains with residues of Eugenol as hanging side groups.

When the reaction is carried out at high conversion (t = 24 h), insoluble products are obtained, reflecting the participation of a good fraction of allylic double bonds once the acrylics have been consumed. As the viscosity increases, the mobility of the macro-radicals decreases and some of the allylic double bonds may participate in graft or cross-linking reactions. The reaction yields are 60% and 70% for PEgMA and PEEgMA, respectively. In order to know the soluble fraction of the polymers thus obtained, the reaction products are subjected to extraction in Soxhlet with toluene for 48 hours and percentages of soluble polymer of 1.06 and 0.6% are obtained for the PEgMA and PEEgMA respectively.

Example 4.- Preparation of copolymers carrying Eugenol.

Eugenyl methacrylate-ethyl methacrylate (EgMA / EMA) copolymers and ethoxy-ethylene-ethyl methacrylate methacrylate copolymers are prepared (EEgMA / EMA). The copolymerization reactions are carried out in a range of monomer composition in the feed between 10/90 and 90/10. The copolymerization reaction is carried out under the same conditions as those described for the polymerization of the eugenyl derivatives in EXAMPLE 3, with the exception of the reaction time, which is adjusted for each feed composition in order to obtain a conversion less than 10% The characterization of the copolymers is performed by proton nuclear magnetic resonance ( ¹ H-NMR).

The ¹ H-NMR (CI ₃ DC) spectra of the EgMA / EMA copolymers show the following resonance bands: 7.0-6.6 (3H, aromatic protons of EgMA), 5.9 (IH, CH = CH ₂ of EgMA), 5.0 (2H, CH = CH ₂ of EgMA), 4.0 (2H, -COOCH ₂ of EMA), 3.7 (3H, CH ₃ OPh of EgMA), 3.3 (2H , CH ₂ Ph of EgMA), 2.0 (2H, - CH ₂ -C (CH ₃ ) COO of EgMA), 1.9-1.8 (2H, -CH ₂ -C (CH ₃ ) EMA COO ), 1.22 (3H, CH ₂ CH ₃ of the EMA), 1.1-0.9 (3H, -CH ₂ -C (CH ₃ ) COO of the EMA and EgMA). The ¹ H-NMR (Cl ₃ DC) spectra of the EEgMA / EMA copolymers show the following resonance bands: 7.0-6.6 (3H, aromatic protons of the EEgMA), 5.9 (IH, CH = CH ₂ of the EEgMA), 5.0 (2H, CH = CH ₂ of the EEgMA), 4.5-4.0 (4H, OCH ₂ -CH ₂ -O of the EEgMA), 3.9 (2H, -COOCH ₂ of the EMA), 3.7 (3H, CH ₃ OPh of EEgMA), 3.3 (2H, CH ₂ Ph of EEgMA), 2.0 (2H, -CH ₂ - C (CH ₃ ) COO of EEgMA), 1 , 9-1.8 (2H, -CH ₂ -C (CH ₃ ) COO of the EMA), 1.2 (3H, CH ₂ CH ₃ of the EMA), 1.1-0.9 (3H, -CH ₂ -C (CH ₃ ) COO of EMA and EgMA).

The composition of the different copolymers is calculated from their corresponding ¹ H NMR spectra taking into account the signal of the allylic protons CH = CH ₂ (δ _H , 5.3-4.7) corresponding to the monomer unit of the derivative of Eugenol and the signal of the protons COOCH ₂ oxyethylene (δ _H , 4.2-3.8) corresponding to the EMA unit.

The reactivity ratios are calculated from the composition values using both the Finemann-Ross and Kelen-Tüdos linear methods, as well as the non-linear methods of Tidwell-Mortimer and Levenberg-Marquardt. The values of the reactivity ratios are shown in Table I.

Table I Values of reactivity ratios for EgMA / EMA and EEgMA / EMA copolymers calculated from linear and nonlinear methods.

Also in Figure FIG.1 the diagram of 95% confidence obtained by application of the mathematical treatment proposed by Tidwell and Mortimer is shown. From the values of the reactivity parameters it can be deduced that both macromolecular radicals whose active end in growth is a unit of EMA and a unit of the acrylic derivative of eugenyl have a greater reactivity against the carrier monomer of Eugenol. The product of the reactivity ratios (rix r ₂ ) less than the unit in both systems indicates that the copolymers prepared under the reaction conditions above These have a distribution of predominantly random units.

From the values of the reactivity ratios obtained by the non-linear methods and applying the composition equation of the copolymer, the composition diagrams shown in Figure FIG.2 are obtained, where the curve corresponds to the application of the equation of composition and the points correspond to the values of the compositions, obtained experimentally. A good fit can be observed between the experimental points and the theoretical diagram. This diagram indicates that radical copolymerization of these systems occurs primarily at random.

Example 5.- Cytotoxicity of EgMA and EEgMA. compared to that of Eugenol.

For this test primary culture cells of human fibroblasts cultured at 37 ° C are used. The culture medium is Minimal Essential Medium Eagle (MEM) modified with HEPES and enriched with 10% fetal bovine serum (FBS), 200 rtiM L-glutamine, 100 units / ml penicillin and 100 μg / ml streptomycin. The culture medium is changed at selected time intervals. Thermanox® (TMX) is used as a negative control. The corresponding monomer is mixed with the Tween 80 surfactant in a 3: 1 weight ratio. The mixture is dispersed in serum-free medium to obtain a solution of the mixture containing 0.0075% -p of monomer and 0.025% -p of surfactant. This solution is diluted successively with serum free medium. Human fibroblasts are seeded at a density of 11 x 10 ⁴ cells / ml in complete medium in a 96-well culture plate and incubated until confluence. After 24 hours of incubation, the medium is replaced with the corresponding dilution and incubated at 37 ° C in an atmosphere of humidified air with 5% CO2 for 24 h. A solution of 3- (4, 5-dimethylthiazol-2-yl) -2.5-diphenyltetrazolium (MTT) bromide in a warm phosphate buffer (PBS) solution (0.5 mg / ml) is prepared and the plates are incubated at 37 ° C for 4 h. Excess medium and MTT are extracted and dimethylsulfoxide (DMSO) is added to all wells in order to dissolve the MTT taken by the cells. This is mixed for 10 min. and absorbance is measured with a Biotek ELX808IU detector using a test wavelength 570 nm and a reference wavelength of 630 nm. Cell viability is calculated from equation (1), where D0 _M , D0 _B , and D0 _c are the optical densities of formazan production for the sample, the blank (MEM without cells) and the control (Tween solution 80 in serum-free and unenriched MEM). The relative cellular dose-viability curve is represented for each compound in order to determine the concentration of compound that produces a 50% MTT-formazan formation depression, which is called ICs ₀ .

Relative cell viability = 100 x (D0 _M - D0 _B ) / D0 _c

(D

The cytotoxic effects of Eugenol, EgMA and EEgMA on the relative viability of human fibroblasts are shown in Figure FIG. 3. The IC ₅₀ values obtained were 3.70 for EgMA, 1.83 for EEgMA and 2.60 for Eugenol. These differences are not very significant if it is taken into account that the Eugenol derivative will be added in small quantities to high molecular weight polymers and that any proportion of unreacted monomer will come from the majority component, a monomer methacrylic with an alkyl side substituent (eg MMA or EMA).

Example 6. Biocompatibility of PEgMA, PEEgMA. and copolymers EgMA / EMA and EEgMA / EMA.

In a first stage, the biocompatibility of polymers and copolymers is evaluated by direct contact of the cells with the corresponding material. The corresponding material is placed in a 24-well plate (in duplicate) that are seeded with human fibroblasts at a density of 14 x 10 ⁴ cells / ml and incubated at 37 ° C for 24 hours. The cells are then fixed with 1.5% glutaraldehyde buffered with a 0.1 M phosphate buffer. The dried samples are coated with a gold layer before being examined by scanning electron microscopy (ESEM) using a voltage of 15 KeV The results of this test after 24 and 48 h of incubation, for Eugenol-bearing polymers and copolymers are shown in Figure FIG. 4. It can be seen that the cells adopt a normal morphology and appear well extended on the surface of any of the Eugenol-bearing polymers or copolymers, indicating the formation of stable adhesions and contacts, and this fact can be considered as a sign of good Biocompatibility for all these materials.

The cytotoxicity from any extract of the material is analyzed by the MTT test. For this test, disks (10 mm in diameter and 1 mm thick) of the corresponding polymers and copolymers obtained at high conversion by reaction of the monomer or mass monomers are used, using AIBN as a radical initiator, at 50 ° C temperature and during a reaction time of 24 hours. The discs are washed and dried before use. The disks thus obtained and the negative control TMX is immersed in 5 ml of FBS-free MEM. Then introduced into a rotary mixer at 37 ⁰ C, the medium at different times (1, 2 and 7 days) is removed and replaced with another 5 ml of fresh medium. All extracts are obtained under sterile conditions. Human fibroblasts are seeded at a density of 11 x 10 ⁴ cells / ml in complete medium in a sterile 96-well culture plate and incubated at confluence. Then, the medium is replaced with the corresponding eluted extract and incubated at 37 ° C in a humidified air atmosphere with 5% CO2 for 24 hours. A solution of MTT in a warm phosphate buffer (PBS) solution is prepared and filtered before use. 10 µl of MTT is added to each well to give a final concentration of 0.5 mg / ml, and the plates are incubated at 37 ° C for 4 hours. Excess medium and MTT are removed and 100 µl of dimethylsulfoxide (DMSO) is added to all wells in order to dissolve the MTT taken by the cells. This is mixed for 10 min. and absorbance is measured with a Biotek ELX808IU plate reader using a test wavelength 570 nm and a reference wavelength of 630 nm. The results are normalized with respect to the negative control (TMX = 100%) and the statistical study of the analysis of variance (ANOVA) is carried out for p <0.05. The results of this test are shown in Figure FIG. 5, in which it can be observed that cell viability is not affected by the presence of the extracts of any of the polymers or copolymers released in a time of 7 days, reaching values of cellular viability greater than 90% of the TMX control in all copolymers.

Cell growth and proliferation on Eugenol-bearing polymers and copolymers is analyzed at through the Alamar Blue essay. For this test, human fibroblasts are planted at a density of 14 x 10 ⁴ cells / ml for 24 hours on the surfaces of the materials to be analyzed that are placed in a 24-well culture plate. Then they are added

2 ml of Alamar Blue dye (a 10% solution of Alamar Blue in DMEM medium free of phenol red) to each specimen. After 4 h of incubation, 100 μl (n = 4) for each sample is transferred to a 96-well plate and the absorbance at 490 nm is measured in a Biotek ELX808IU plate reader. The samples are washed with PBS twice to monitor the cells on the materials. This step is performed at 2, 7, 14 and 21 days. The results are statistically analyzed by analysis of variance (ANOVA) with respect to the PEMA using the significance levels of p <0.05, p <0.01 and p <0.001. As can be seen in Figure FIG. 6, in all polymers and copolymers, fibroblasts proliferate between days 1 and

3 similar to the TMX control, and after this time a decrease in proliferation is observed. The statistical analysis of the results with respect to PEMA shows that on days 1, 3 and 7 some copolymer compositions have significantly higher cell proliferation values compared to PEMA. On the third day all EgMA / EMA copolymers exhibit significantly higher cell proliferation values as well as EEg / EMA copolymers of composition 20/80 to 50/50. On the seventh day, when proliferation has begun to decrease in almost all materials, in a majority of copolymer compositions there is a significantly higher proliferation compared to PEMA. Thus, it can be affirmed that the derivatives of Eugenol They provide an improvement in proliferation during the initial periods with respect to PEMA.

Example 7.- Self-healing acrylic formulations of eugenyl methacrylate with acrylic and polyacrylic components.

Self-healing compositions of acrylic systems are formulated using a two-phase system as mentioned in the Detailed Description of the Invention. One of the phases is a liquid composed of the monomers of eugenyl methacrylate (EgMA) and methyl methacrylate (MMA) in varying proportions, using the compound (4-N, N-dimethylaminophenyl) -methanol (DMOH) as activator.

The second phase consists of a solid based on poly (methyl methacrylate) (PMMA). It is possible to use this component directly (for example commercial PMMA pearls such as Bonar, or the beads of a "Self-curing Resin, etc.) or the solid component of other commercial formulations such as DuraLay. As a solid phase in Example 7, they have been used the pearls of the "Self-curing Resin" and the solid phase of the Duralay commercial formulation, which are referred to herein as Res. Autopol. and DuraLay respectively.

The specific composition of the systems considered is described in Table II where the following abbreviations are used:

Res. Autopol = DuraLay PMMA beads = DuraLay commercial solid phase MMA = EgMA methyl methacrylate = Eugenyl methacrylate

DMOH = (4-N, N-Dimethylaminophenyl) -methanol S: L = Solid: liquid ratio Table II Compositions of self-healing acrylic formulations based on PMMA. in the presence of eugenyl methacrylate.

For each Example, a total of 40 g of solid phase and the corresponding amount of liquid are used according to the solid: liquid ratio used. The corresponding solid and liquid phases of each formulation are mixed and the resulting paste is introduced into a Teflon mold. A thermocouple is placed in the center of the mold at a height of 3 mm in the internal cavity. The time is taken from the beginning of the mixing of the two components and the temperature is recorded. An average of two measurements is made for each formulation. Teflon mold features are described in (Pascual B, Vázquez B, Gurruchaga M, Goñi I, Geneva MP, Gil FJ, Planell JA, Levenfeld B, San Román J. "New aspects of the effect of size and size distribution on the setting parameters and mechanical properties of acrylic bone cemente ". Biomaterials 1996, 17, 509-516).

Curing parameters are determined according to ISO 5833 (International Standard ISO 5833. Implants for Surgery-Acrylic Resins Cements. 1992).

The time of the pasty state (t _p ) represents the time in which the two phases are mixed forming the paste prior to its introduction to the mold, a moment that is considered when the paste does not adhere to the surgical glove.

The setting time (tf) is determined as the time at which the cement mass temperature is the arithmetic mean of the maximum temperature in ° C and the ambient temperature, 23 ± 1 ° C.

The working time (t _t ) is calculated as the difference between the setting time and the time of the pasty state. The peak or maximum temperature (T _max ) is defined as the maximum temperature reached during the polymerization reaction.

The values of the curing parameters for the Eugenol carrier formulations whose compositions are reflected in Table II are shown in Table III.

Table III Maximum temperature values (T ³ ^ _13x ), pasty state times (t _p ), setting (t _f ), and work (t _t ) obtained in curing acrylic formulations with eugenyl methacrylate.

The setting and working time values of the formulations prepared in the presence of the Eugenol derivative in Examples I and II are higher than those obtained with the control formulations, Controls I and II. However, the setting times in Examples III and IV decrease slightly with respect to the DuraLay commercial control, Controls III and IV, but without changing the working times. The peak temperatures of formulations containing eugenyl methacrylate are lower in all cases (about 10/5 ° C depending on the solid phase) to those of the corresponding controls, which represents an important and significant benefit from a point of view biological.

Similar results are obtained using ethoxyieugenyl methacrylate in the same proportions as eugenyl methacrylate.

Example 8. Self-healing acrylic formulations of eugenyl methacrylate with acrylic and polyacrylic components in the presence of zinc oxide.

Autocurable compositions of acrylic systems are formulated using a liquid phase composed of the monomers of eugenyl methacrylate (EgMA) and methyl methacrylate (MMA), using as compound the compound (4- N, N-dimethylaminophenyl) -methanol (DMOH).

The solid phase is composed of zinc oxide (ZnO) particles and poly (methyl methacrylate) (Plexigum, Merck) particles, in different proportions, using benzoyl peroxide (BPO) as the radical initiator. The specific compositions of the formulations tested in Example 8 are listed in Table IV where the following abbreviations are used: BPO = Benzoyl Peroxide PMMA = Poly (methyl methacrylate) beads ZnO = Zinc Oxide

MMA = Methyl Methacrylate

EgMA = Eugenyl Methacrylate

DMOH = (4-N, N-Dimethylaminophenyl) -methanol

S: L = Solid ratio: liquid

Table IV Compositions of self-healing acrylic formulations of

ZnO / PMMA in the presence of eugenyl methacrylate.

The preparation of the formulation and the determination of the curing parameters are carried out as explained in Example 7. The results are shown in table V.

Table V. Maximum temperature values (T ² S _113x ), pasty state times (t _p ), setting (t _f ), and work (t _t ) obtained in curing acrylic formulations with eugenyl methacrylate and in the presence of zinc oxide particles.

The most outstanding of the values of the curing parameters set forth in Table V is the low peak temperature obtained for all formulations, close to 40-45 ° C. Similar results are obtained using ethoxyethylene methacrylate in the same proportions as eugenyl methacrylate.

Claims

1.- Eugenol-derived acrylic monomer characterized by the general formula (I):

Where: n is 0, 2, 6 or 11

2. - Eugenol-derived acrylic monomer according to claim 1 characterized by the general formula Ia:

Where :

R2 is a methyl, ethyl, propyl, isopropyl, butyl, isobutyl or tert-butyl radical.

3.- Eugenol-derived acrylic monomer according to claim 2 characterized in that it is eugenyl methacrylate (EgMA):

4. - Eugenol-derived acrylic monomer according to claim 1 characterized by the general formula Ib:

Where: n is 2, 6 or 11.

5. Acrylic monomer derived from Eugenol according to claim 4, characterized in that it is ethoxyieugenyl methacrylate (EEgMA):

6. Procedure for obtaining an acrylic derivative of Eugenol according to claim 1 to 3 characterized in that it is carried out under mild conditions and because it comprises the following steps: i) Eugenol is dissolved with triethylamine in stoichiometric amounts, using diethyl ether as a solvent at room temperature, ii) under a nitrogen atmosphere, a stoichiometric amount of methacryloyl chloride is added dropwise to the solution of i) and Let it react for 48 hours at room temperature, and iii) the acrylic monomer is isolated and purified as a reaction product.

7. - Procedure for obtaining an acrylic monomer derived from Eugenol according to claim 1, 3 and 4 characterized in that it is carried out under mild conditions and that it comprises the following steps: i) synthesis of 1-hydroxy, n-alkyl Eugenol by treatment of Eugenol with the corresponding OC, Gϋ-chloro-alkyl alcohol in a hydroalcoholic medium, ii) the corresponding 1-hydroxy, n-alkyl Eugenol is dissolved with triethylamine in stoichiometric amounts, using diethyl ether as solvent, iü) under an atmosphere of nitrogen, a stoichiometric amount of methacryloyl chloride is added dropwise to the solution of ii) and allowed to react for 48 h at room temperature, and iv) the acrylic monomer is isolated and purified as a reaction product.

8. Method of obtaining according to claims 6 and 7 characterized in that the purification of the acrylic monomer of Eugenol is carried out in a chromatographic column of silica.

9. Eugenol carrier polymer characterized in that it comprises an acrylic monomer derived from Eugenol according to claims 1 to 5.

10.- Eugenol carrier polymer according to claim 9 characterized in that the Eugenol acrylic monomer belongs to the general formula Ia:

Where :

11. Polymer according to claim 10 characterized in that the polymer is poly (eugenyl methacrylate) (PEgMA).

12. Eugenol carrier polymer according to claim 9 characterized in that the Eugenol acrylic monomer belongs to the general formula Ib:

Where: n is 2, 6 or 11.

13. Polymer according to claim 12, characterized in that the polymer is poly (ethoxyieuylene methacrylate) (PEEgMA).

14. Eugenol carrier polymer according to claim 9 characterized in that it is a copolymer comprising an acrylic monomer derived from Eugenol of general formula

(I) according to claim 1 and a second acrylic monomer other than the previous one.

15. Eugenol carrier polymer according to claim 14, characterized in that the second acrylic monomer belongs to the following group: methyl methacrylate (MMA) or ethyl methacrylate (EMA).

16.- Eugenol carrier polymer according to claim 15, characterized in that the copolymer belongs to the following group: eugenyl methacrylate-ethyl methacrylate (EgMA / EMA) copolymer and ethoxyieugenyl-ethyl methacrylate (EEgMA / EMA) copolymer.

17. Process for obtaining a Eugenol carrier polymer according to claims 9 to 16, characterized in that it comprises a radical polymerization step of any of the monomeric compounds of the general formula

(I), and is carried out by dissolving the corresponding monomer in toluene, using azobisisobutyronitrile

(AIBN) as a radical initiator and at a temperature of 50-60 ° C.

18. Method for obtaining a Eugenol carrier polymer according to claims 9 to 16, characterized in that it comprises a copolymerization step in the presence of a radical initiator of any of the compounds of general formula (I) as the first monomer, with a monomer different acrylic as second monomer.

19. Method of obtaining a Eugenol carrier polymer according to claim 18 characterized in that the Second acrylic monomer belongs to the following group: methyl methacrylate (MMA) or ethyl methacrylate (EMA).

20. Self-curable formulation characterized in that it comprises a liquid phase composition with acrylic monomers derived from Eugenol according to claims 1 to 5 and a solid phase composition with acrylic polymers derived or not from Eugenol.

21. Self-curable formulation according to claim 20 characterized in that the liquid phase self-healing acrylic composition comprises one or more of the acrylic monomers derived from Eugenol, in an amount between 20-60% -p with respect to the total weight of the phase liquid, and a second acrylic monomer in an amount between 80-40% -p with respect to the total weight of the liquid phase.

22. Self-curable formulation according to claim 21, characterized in that the second acrylic monomer belongs to the following group: methyl methacrylate (MMA) or ethyl methacrylate (EMA).

23. Self-curable formulation according to claims 20 to 22, characterized in that the liquid phase composition also comprises an aromatic tertiary amine as an activator in an amount between 0.5-2.5% -p, preferably 2% -p, and one or more inhibitors in an amount of up to 0.01% -p.

24. Self-curable formulation according to claim 23 characterized in that the inhibitor belongs to the family of quinones.

25. Self-curable formulation according to claim 20 characterized in that the solid-phase self-curing acrylic composition comprises particles of prepolymerized poly (methyl methacrylate) (PMMA), or prepolymerized poly (ethyl methacrylate) (PEMA) or polymers derived from Eugenol according to claims 9 to 16 or mixtures of the same with other thieves, present in an amount between 20-80% -p with respect to the total weight of the solid phase.

26. Self-curable formulation according to claim 25 characterized in that the self-curing acrylic solid phase composition comprises zinc oxide (ZnO) in an amount between 50-80% -p with respect to the total weight of the solid phase.

27. Self-curable formulation according to claims 25 and 26 characterized in that the solid-phase self-healing acrylic composition comprises one or more initiators in an amount of up to 3% -py / or one or more radiopaque agents in an amount between 20-25% -p with respect to the total weight of the solid phase.

28. Self-curable formulation according to claim 27 characterized in that the initiator is benzoyl peroxide.

29. Self-curable formulation according to claim 27, characterized in that the radiopaque agents are selected from the following group: barium sulfate, zirconium dioxide, tantalum oxide, strontium oxide and organic compounds.

30. Self-curable formulation according to claim 20 characterized in that it comprises a liquid phase composition with monomers of eugenyl methacrylate (EgMA) and methyl methacrylate (MMA) in varying proportions, with the compound (4-N, N-dimethylaminophenyl) - methanol (DMOH) as an activator, and a solid phase composition based on poly (methyl methacrylate) (PMMA) or similarly with ethoxyieugenyl methacrylate without or with zinc oxide.

31.- Use of the self-curable formulation according to claims 20 to 30, by mixing the liquid and solid phase compositions and their direct application or by injection and finally curing "in situ" for reconstruction, temporary or permanent, dental and bone.

32.- Use of the formulation according to claim 31 characterized in that the bone reconstruction consists of a vertebra fixation or biomechanical fixation of osteoporotic fractures in minimally invasive surgery in the field of traumatology and orthopedic surgery.