KR101262321B1 - Hydrogel with improved tissue adhesion prevention - Google Patents

Abstract

PURPOSE: Hydrogel is provided to have improved tissue adhesion inhibition power, and to perform a simple procedure by injection as the hydrogel is in a sol state at room temperature. CONSTITUTION: Hydrogel is manufactured by the cross-link reaction of the following: 100 parts by weight of modified polyethylene-polypropylene-polyethylene copolymer in which one end is substituted with lactic acid and the other end is substituted with ethylene glycol; 1-2 parts by weight of complex cross-linking agents in which more than two kinds of cross-linking agents selected from hyaluronic acid, adipic acid dihydrazid, and sodium alginate are combined; and 0.1-1 parts by weight of more than one divalent metal compound selected from the group consisting of calcium, magnesium, barium, zinc, strontium, and iron(II). The copolymer has the content of lactic acid of 4-7 mol%, and the content of ethylene glycol of 3-6 mol%.

Description

Hydrogel with improved tissue adhesion prevention

The present invention relates to a hydrogel that is easy to apply in vivo and has excellent biocompatibility, and is useful as a tissue adhesion inhibitor for surgical surgery or a skin cosmetic filler because it is injected in vivo and has excellent efficacy of preventing adhesion between tissues.

Surgical laparotomy is one of routine operations. Adhesion of organs and tissues, which commonly occurs after surgery, is one of the natural phenomena that occur during the proliferation and regeneration of damaged tissue cells. However, reoperation may be necessary for postoperative dysfunction and adhesion detachment, which may be life-threatening. Tissue adhesion is inevitable by abdominal or pelvic surgery and occurs in 93% of all patients after abdominal surgery. Therefore, in order to minimize the problem of tissue adhesion after surgery, there is a method of inhibiting adhesion between tissues after surgery using an anti-adhesion agent. Representative examples include a film form and a gel form. As a problem, the film form has a high probability of developing new adhesions at the sutures of surrounding tissues, and is difficult to apply to complex and conduit-type surgical sites. The gel form is easily broken down and absorbed in vivo before the wound is healed, and thus has low efficacy as an anti-adhesion agent. Therefore, there is a need for a new product that supplements the disadvantages of conventional tissue adhesion inhibitors.

Meanwhile, a hydrogel prepared by crosslinking a mixture of sodium alginate with a polyethylene (PEO) -polypropylene (PPO) -polyethylene (PEO) copolymer as a tissue adhesion preventing material and using calcium chloride and other divalent cations is known. Journal of the Korean Surgical Society 71 (4), pp. 280-289 (2006); Journal of Biomedical Materials Research. Part A. 72 (3), pp. 306-316 (2005)] However, the hydrogel is applied to the tissue as a hydrophilic material and easily absorbed into the body, and has a poor adhesion, and flows out of the applied tissue so that the hydrogel can function as an anti-adhesion agent or a cosmetic filler. There is a limit to expression.

Therefore, there is an urgent need to develop a new material capable of stably adhering to tissues in vivo as a tissue adhesion inhibitor for a desired period of time.

An object of the present invention is to provide a hydrogel which is injected in vivo to improve the anti-adhesion between tissues and adhesion to tissues.

In addition, another object of the present invention is to provide a use of the hydrogel as an anti-adhesion agent for surgery or skin filler.

In order to solve the above problems, the present invention

Iii) 100 parts by weight of a polyethylene (PEO) -polypropylene (PPO) -polyethylene (PEO) copolymer modified at one end thereof with lactic acid and the other end with ethylene glycol;

Ii) 1-2 parts by weight of a crosslinking agent in which at least two crosslinking agents selected from hyaluronic acid (HA), adipic acid dihydrazide (ADH), and sodium alginate (SA) are combined; And

V) 00.1 to 0.1 parts by weight of at least one valence 2 metal compound selected from the group consisting of calcium, magnesium, barium, zinc, strontium and iron (II);

It is characterized by a hydrogel prepared by crosslinking reaction.

In addition, the present invention is characterized in that the hydrogel is used as a surgical tissue adhesion inhibitor or skin cosmetic filler.

The polyethylene (PEO) -polypropylene (PPO) -polyethylene (PEO) copolymer of the present invention is used for producing hydrogel, and both ends of the copolymer are modified by lactic acid and ethylene glycol. Compared to the above, it is not easily absorbed at the time of injection into the body and stably adheres to the desired wound site and tissue, thereby maintaining the function as an anti-adhesion agent or a filler agent. Therefore, the hydrogel of the present invention has a useful effect as a filler for preventing tissue adhesion or skin care.

In addition, since the hydrogel of the present invention exists in a sol state at room temperature, a simple procedure by injection injection is possible.

Hereinafter, the present invention will be described in detail.

The hydrogel according to the present invention is a composite crosslinking agent in which both ends are modified polyethylene (PEO) -polypropylene (PPO) -polyethylene (PEO) copolymers each substituted by lactic acid and ethylene glycol, and at least two crosslinking agents. And a crosslinking reaction of a valence 2 metal compound. That is, the modified PEO-PPO-PEO terpolymer and the crosslinking agent crosslink under a water solvent to form a crosslinked polymer network structure, and the pores present in the network structure are filled with water filled as a solvent. To form a hydrogel. The hydrogel of the present invention can be injected in vivo and easily adhered to desired tissues, and it is also possible to maintain stable for a long time at the adhered tissue site. Therefore, the hydrogel of the present invention is stably performed in vivo for several months when applied as a tissue adhesion inhibitor or skin filler.

Hereinafter, each component used to prepare the hydrogel according to the present invention will be described in more detail.

As mentioned above, the polyethylene-polypropylene-polyethylene copolymer (hereinafter, referred to as 'PEO-PPO-PEO copolymer') has a high hydrophilicity and weak adhesion to tissues, and thus has many problems for application as a material for anti-adhesion and cosmetic filler. There is. Accordingly, in the present invention, both ends of the PEO-PPO-PEO copolymer are modified with lactic acid and ethylene glycol to impart hydrophobicity and adhesion, so that they can be applied to a living body stably for a long time without being easily absorbed in the body. It could be maintained.

If the present invention briefly describes a method for producing a modified PEO-PPO-PEO copolymer used for the preparation of a hydrogel, the present invention is not limited by the following copolymer modification method.

First, the PEO-PPO-PEO copolymer is reacted with lactic acid or a derivative thereof to replace one end with lactic acid, and then with ethylene glycol to replace the other end with ethylene glycol to modify the PEO-PPO-PEO air. Prepare coalescing. The reaction for substituting the lactic acid is performed under mildly acidic conditions of pH 5 to 6, at room temperature (25 to 30 ° C.) under conditions in which organic bases such as triethylamine, alcohol dehydrogenase and nicotine amide (NAD) coenzyme are present. Proceed. In addition, the reaction for substituting the ethylene glycol is carried out under conditions of pH 9 to 10 and room temperature (25 to 30 ℃).

As a modifier used for the lactic acid substitution reaction, the lactic acid or a derivative thereof includes lactic acid or an alkali metal or earth metal salt thereof, and in some cases, the carbon skeleton of the lactic acid includes OH, NO ₂ , CN, C _{1- 20} alkyl or the like may be introduced as a substituent. Specifically, the lactic acid or derivatives thereof may include lactic acid, sodium lactate, calcium lactate, calcium stearyl lactate, and the like. In the ethylene glycol substitution reaction, ethylene glycol having a molecular weight in the range of 40 to 90 may be used as a modifier. In addition, alcohol dehydrogenase and enzymes having a similar function may be used for the lactic acid substitution, and the present invention does not place any particular limitation on the selection of these enzymes. Alcohol dehydrogenase ™ (Sigma Aldrich, ≥300 units / mg proten), Aldehyde Dehydrogenase ™ (Sigma Aldrich, Potassium-activated), β-Nicotinamide adenine, which are commercially available products, can be used in the present invention. dinucleotide hydrate ™ (Sigma-Aldrich). In order to maintain the weakly acidic conditions (pH 5-6) for lactic acid substitution, inorganic acids (sulfuric acid, phosphoric acid, nitric acid, hydrochloric acid) and organic acids (formic acid, acetic acid, citric acid, ascorbic acid), etc., may be added at a concentration of 0.01 N to 0.1 N. It can be diluted. In addition, in order to maintain basic conditions (pH 9.0 to 10.0) for ethylene glycol substitution, sodium hydroxide, potassium hydroxide, barium hydroxide and the like can be used.

The lactic acid or derivatives thereof, or ethylene glycol, used as a modifier of the PEO-PPO-PEO copolymer in the above, may be used in the range of 5 to 10 parts by weight based on 100 parts by weight of the PEO-PPO-PEO copolymer. PEO-PPO-PEO copolymers modified within the above range can be identified through nuclear magnetic resonance (NMR) or infrared (IR) spectroscopy. For example, according to NMR analysis, the peak corresponding to lactic acid is observed at 1.6 ± 0.2 ppm, and when calculated as an integral value, the content of lactic acid substituted in the PEO-PPO-PEO copolymer is in the range of 4 to 7 mol%. . In the NMR analysis, a peak corresponding to ethylene glycol is observed at 2.2 ± 0.2 ppm, and when calculated as an integral value, the content of ethylene glycol substituted in the PEO-PPO-PEO copolymer is in the range of 3 to 6 mol%. Maintaining the substitution rate range of the lactic acid and ethylene glycol is suitable as a material for producing a hydrogel in which the adhesive force and tissue adhesion prevention force of the present invention are improved. In other words, when the content of the lactic acid substituted in the PEO-PPO-PEO copolymer is less than the above range, the copolymer is hydrophilic, so that the copolymer is prepared as a hydrogel and easily absorbed into the body when applied to a living body. On the other hand, when the content of ethylene glycol substituted in the PEO-PPO-PEO copolymer is less than the above range, the adhesive strength of the hydrogel is poor, and when applied to a living body there is a problem that it is not easily adhered to the tissue flows down. Therefore, in order to produce a hydrogel having improved adhesion and tissue adhesion preventing properties, the substitution rate of lactic acid and ethylene glycol substituted at the ends of the PEO-PPO-PEO copolymer is preferably maintained within the above range.

In addition, the biocompatible hydrogel of the present invention has a property of being in a sol state at room temperature (27 ± 3 ° C.) and changing to a gel state in vivo, and thus easily prepared by injection. It can be injected into a living body. In addition, since it has the property of changing to a gel state in vivo, it is possible to adhere to a desired tissue site and to stably stay in a gel state for a long time. As a result, the wound and tissue are separated by the gel adhered to the tissue to prevent adhesion between the tissues, or to impart a sense of volume to the skin. The optimal hydrogel for application to the present invention is a hydrogel having a low critical solution temperature (LCST) of 24 to 30 ° C., measured under the condition that the concentration of solids is 20 to 25% by weight (w / w).

In the present invention, the low low critical solution temperature (LCST) of the hydrogel was controlled through the PEO unit content and the weight average molecular weight of the PEO-PPO-PEO copolymer. That is, the present invention uses a PEO-PPO-PEO copolymer having a content of PEO units of 65 to 85% by weight and a weight average molecular weight of 7,000 to 16,000 g / mol. More preferably, two or more PEO-PPO-PEO copolymers satisfying the PEO unit content and the weight average molecular weight proposed above are mixed and used in an appropriate weight ratio. That is, a PEO-PPO-PEO copolymer having a relatively high PEO monomer content and a relatively low weight average molecular weight and a PEO-PPO-PEO copolymer having a relatively low PEO monomer content and a relatively high weight average molecular weight are used. will be. Specifically, PEO-PPO-PEO copolymer (hereinafter, referred to as 'copolymer 1') having a content of PEO unit 74 to 82% by weight and a weight average molecular weight (Mw) of 7,500 to 12,000 g / mol Weight percent; Copolymer consisting of 5 to 30% by weight of PEO-PPO-PEO copolymer (hereinafter referred to as 'copolymer 2') having a content of PEO units of 69 to 73% by weight and a weight average molecular weight of 12,500 to 16,000 g / mol Mixtures can be used.

In addition, the present invention uses a crosslinking agent in which two or more crosslinking agent compounds selected from hyaluronic acid (HA), adipic dihydrazide (ADH), and sodium alginate (SA) are combined as a crosslinking agent for preparing a hydrogel. Specific examples of the composite crosslinking agent include hyaluronic acid-adipic acid dihydrazide-sodium alginate (HA-ADH-SA), hyaluronic acid-adipic acid dihydrazide-hyaluronic acid (HA-ADH-HA), sodium At least one selected from the group consisting of a crosslinking agent consisting of alginate-adipic acid dihydrazide-sodium alginate (SA-ADH-SA) and the like can be used. In the composite crosslinking agent, hyaluronic acid-adipic acid dihydrazide-sodium alginate (HA-ADH-SA) is preferably used. The crosslinking agent may further include a crosslinking reactive group such as a carboxylate group and a sulfate group.

Conventionally, hyaluronic acid (HA), sodium alginate (SA), and divalent cationic metal compounds as crosslinking agents have been used as a mixture of PEO-PPO-PEO copolymers, but these crosslinking agent compounds are absorbed and discharged within 24 hours in the body. There is a limit in expressing a function as an anti-adhesion agent. In contrast, in the present invention, a crosslinker compound having two or more crosslinking compound chemically bonded to each other is used, and the terminal amine group of the crosslinker is crosslinked with the terminal carboxyl group of the modified PEO-PPO-PEO copolymer. As a result, the time of absorption and excretion in the body can be maintained for 7 days or more, and the adhesion to tissues or organs can be improved, and it is possible to sufficiently express the function in the body as an anti-adhesion agent or a filler.

The composite crosslinking agent used in the present invention may be prepared by combining two or more crosslinking compound compounds selected from hyaluronic acid (HA), adipic acid dihydrazide (ADH), and sodium alginate (SA). The coupling reaction can be carried out using a conventional coupling agent such as 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride (EDC). For example, the coupling reaction between hyaluronic acid (HA) and adipic dihydrazide (ADH) can be carried out using a conventional binder under acidic conditions of pH 4-5. And the coupling reaction of sodium alginate (SA) and adipic dihydrazide (ADH) can be carried out using a conventional binder in the basic conditions of pH 8-9. The composite crosslinker prepared by the above coupling reaction is prepared with a mixture of HA-ADH-SA, HA-ADH-HA, SA-ADH-SA, and the like. When the composition of the mixture of the prepared crosslinking agent was confirmed by FT-IR and ¹ H-NMR, 80 to 90 wt% of HA-ADH-SA, 0.1 to 1.0 wt% of HA-ADH-HA, and 19.9 to SA-ADH-SA 9 weight percent. In the present invention, a mixture of these composite crosslinks may be used separately, but may be used directly in the preparation of a hydrogel as a composite crosslinked mixture without undergoing a separate separation process.

The crosslinking agent is used in the range of 0.5 to 3 parts by weight, preferably 1 to 2 parts by weight, based on 100 parts by weight of the modified PEO-PPO-PEO copolymer used as the substrate. If the amount of the composite crosslinking agent is too small, the polymer network structure may not be formed. If the amount of the crosslinking agent is too high, the pore volume in the polymer network structure is reduced, thereby making it impossible to prepare a temperature sensitive hydrogel.

In the present invention, a metal compound of valence 2 is used to promote the formation of the polymer network structure of the modified first copolymer and the modified second copolymer and the crosslinking agent. The valence 2 metal compound uses at least one selected from compounds containing divalent metals such as calcium, magnesium, barium, zinc, strontium, and iron (II). Specifically, halides, sulfates, nitrates, or acetates containing a divalent metal selected from calcium, magnesium, barium, zinc, strontium, or iron (II) may be used. The metal compound of valence 2 is used in the range of 0.1 to 1 part by weight of the metal compound of valence 2 based on 100 parts by weight of the modified PEO-PPO-PEO copolymer used as the substrate. At this time, if the amount of the metal compound of valence 2 is too small, the polymer network structure may not be formed. If the amount is too high, the pore volume in the polymer network structure is reduced to form a temperature sensitive characteristic and a partial gel to function. Rather, it can be degraded. In addition, in using the valence 2 metal compound, the concentration is adjusted to be used in the range of 0.001 to 0.01 M concentration. The reason is that if the concentration of the metal compound of valence 2 is too low, the volume of the entire copolymer may be increased, and the concentration of the copolymer may be lowered, and thus the temperature sensitive characteristic may be degraded. This is because the compound may suddenly gel. When the metal compound of valence 2 is used in the range of 0.1 to 1 part by weight based on 100 parts by weight of the modified PEO-PPO-PEO copolymer by adjusting the valence 2 metal compound to a concentration of 0.001 to 0.01 M as proposed by the present invention. After the completion of the crosslinking reaction, a partially semi-solid gel is formed having a weight average molecular weight in the range of 90,000 to 130,000 g / mol.

Referring to the method for producing a hydrogel according to the present invention in detail.

First, the modified PEO-PPO-PEO copolymer is placed in sterile water and stirred. The amount of sterile water used may be adjusted to an amount that maintains the concentration of the copolymer solution in the range of 20-25% (w / w). At this time, if the amount of the sterile water is used too small, that is, if the concentration of the copolymer solution is too high, the time for dissolving the copolymer is prolonged as well as the gelation phenomenon may occur due to a partly rapid reaction during blending. Also, if the amount of sterile water is used too much, that is, if the concentration of the copolymer solution is too dilute, the low critical solution temperature (LCST) may increase or not exist and at the same time, the crosslinking may not be performed smoothly, and thus the polymer network structure may not be formed. have. At this time, the stirring temperature of the copolymer solution is suitable 4 ~ 10 ℃, if the stirring temperature is too high, small bubbles are generated during the dissolution process and the viscosity is high, it is difficult to add the next material, and also the modified first air Coupling of the carboxyl group of the copolymer and the modified second copolymer with the amine group of the crosslinking agent cannot be performed smoothly. If the stirring temperature is too low, freezing occurs and a smooth reaction cannot occur. Stirring speed is suitable 80 ~ 150 rpm, if the stirring speed is too slow, the stirring is not, if the stirring speed is too fast, bubbles are generated and the next process becomes difficult.

A crosslinking reaction is performed by adding a complex crosslinking agent and a metal compound of valence 2 to the copolymer solution. While stirring the copolymer solution, a crosslinking compound and a metal compound of valence 2 are added, and if the stirring temperature is too high, the crosslinking reaction may be promoted by the heat of reaction generated during the crosslinking reaction, thereby not forming a desired network structure. Therefore, it is appropriate to maintain the crosslinking reaction at a relatively low temperature of 4 to 10 ° C. and to maintain a stirring speed of 80 to 150 rpm. When the addition is completed, the internal temperature of the reactor is gradually increased to increase the reaction of the unreacted amine group and the carboxyl group to a maximum of 40 ° C. to crosslink. If the temperature of 40 ℃ is maintained for a long time, the hyaluronic structure of the composite crosslinking agent may be deformed, and at this time, the temperature increase rate is preferably maintained at 0.1 to 5 ℃ / 10 seconds. After completing the crosslinking reaction while maintaining the maximum temperature, it is better to cool slowly by applying vacuum pressure to naturally remove bubbles generated during the reaction. At this time, the vacuum pressure is maintained in the range of 0.07 to 0.08 MPa, and the cooling rate is cooled to the minimum temperature of 4 ℃ while maintaining 0.1 to 5 ℃ / 10 seconds. Even during the cooling process, the stirring speed of the reactor is preferably maintained at 80 to 150 rpm.

In addition, the hydrogel according to the present invention has a characteristic that the critical solution temperature (LCST) measured under the condition that the concentration of the solid content is 20 to 25% by weight (w / w) is 24 to 30 ° C. In addition, the weight average molecular weight (Mw) of the polymer immediately before adding the mixed solution of the crosslinking compound and the valence 2 metal compound was 7,000 ± 15,000 g / mol, and the weight average molecular weight (Mw) of the polymer after the completion of the crosslinking reaction was 90,000 to 130,000 g. It has a characteristic of / mol. In addition, the hydrogel is present in the dermal gel (gel) for more than one month. Copolymer containing a conventional polyethylene oxide is present in the body in the form of a gel for only a few days in the body, but is absorbed in the body, the hydrogel of the present invention is made of a specific composition that will be able to exist more stably in the body for a longer time .

The present invention will now be described in more detail with reference to the following examples, but the present invention is not limited thereto.

[Manufacturing Example]

Preparation Example 1 Preparation of Composite Crosslinking Agent

To 250 g of sterile water, 4 g of hyaluronic acid having an average molecular weight of 2 million daltons, 12 g of 1,4-linked glucopyranosideuronic acid and 60 g of 1,4-linked α-D-mannopranosideuronic acid Dilution adjusted to pH 4-5. 4 g of adipic dihydrazide and 80 ug of EDC binder were added thereto, stirred at a temperature of 10 ° C. to 15 ° C. for about 24 hours, and the reaction was terminated by adding an excess of ethanol and isopropyl alcohol. Dried.

After diluting the combination of hyaluronic acid and adipic acid prepared above by 10 weight times in sterile water, 20 g of sodium alginate and 80 ug of EDC binder were added under weakly basic pH (pH 8-9), and 10 ° C. to 15 ° C. After stirring for 24 hours at the temperature to terminate the reaction by adding an excess of ethanol and isopropyl alcohol, the fibrous composite crosslinker was separated by a filter and dried at room temperature. Then, the mixture was diluted 2 times by weight in sterile water, purified using a dialysis membrane of 3500 Da, and then frozen at -70 ° C and lyophilized at -42 ° C.

The composition of the prepared composite crosslinker was confirmed by FT-IR and ¹ H-NMR. As a result, 90% by weight of hyaluronic acid (HA) -adipic acid dihydrazide (ADH) -sodium alginate (SA), 1% of hyaluronic acid (HA) -adipic acid dihydrazide (ADH) -hyaluronic acid (HA) %, And a mixture of 9 wt% sodium alginate (SA) -adipic acid dihydrazide (ADH) -sodium alginate (SA). The prepared composite crosslinked mixture was used directly in the hydrogel preparation of the following examples as a composite crosslinked mixture without undergoing a separate separation process.

Preparation Example 2 Preparation of Modified PEO 70-1 Copolymer

To 100 g of sterile water, 60 g of PEO-PPO-PEO copolymer (hereinafter, referred to as 'PEO 70-1') having a content of PEO unit of 70% by weight on average and a weight average molecular weight of 12,309 g / mol is used. Diluted to 15 parts by weight. 3 g of calcium lactate hydrate ([CH ₃ CH (OH) COO] ₂ Ca.xH ₂ O, molecular weight 220 g / mol), 880 µl of triethylamine, alcohol dehydrogenase (trade name) 880 μl of Saccharomyces cerevisiae) and 880 μl of nicotinamide (NAD) were added and stirred at room temperature around 30 ° C. for 24 hours. In addition, a PEO-PPO-PEO copolymer in which lactic acid was substituted was filtered using a membrane (0.5 μm, 90 mm) made of polytetrafluoroethylene (PTFE).

To 100 g of sterile water, 15 g of PEO-PPO-PEO copolymer substituted with lactic acid, 2.5 g of ethylene glycol (weight average molecular weight 70), 1N NaOH were added thereto, adjusted to PH 9-10 and stirred at 60 ° C. for 2 hours. Reformate. And filtered using a membrane of polytetrafluoroethylene (PTFE) (0.5 ㎛, 90 mm) with NaCl 5% (w / v) solution, and then freeze and freeze-dried at -70 ℃, one end of the lactic A modified PEO 70-1 copolymer was prepared by replacing with acid and replacing the other end with ethylene glycol.

The substitution rate of the lactic acid and the substitution rate of ethylene glycol were measured through a nuclear magnetic resonance analyzer (NMR) for the modified PEO 70-1 copolymer. As a result, the substitution rate of the lactic acid was 4 mol% based on the NMR integral value, and the substitution rate of ethylene glycol was 3 mol%.

Preparation Example 3 Preparation of Modified PEO 80-1 Copolymer

The same method as Preparation Example 2 was carried out using a PEO-PPO-PEO copolymer (hereinafter, referred to as 'PEO 80-1') having an average content of PEO units of 80 wt% and a weight average molecular weight of 7,972 g / mol. One end was substituted with lactic acid, and the other end was substituted with ethylene glycol to prepare a modified PEO 80-1 copolymer.

For the modified PEO 80-1 copolymer, the substitution rate of lactic acid and the substitution rate of ethylene glycol were measured through a nuclear magnetic resonance analyzer (NMR). As a result, the substitution rate of the lactic acid was 4 mol% based on the NMR integral value, and the substitution rate of ethylene glycol was 3 mol%.

Comparative Production Example 1. Preparation of Unmodified PEO 70-2 Copolymer

A PEO-PPO-PEO copolymer (hereinafter, referred to as 'PEO 70-2') having an average content of PEO units of 70% by weight and a weight average molecular weight of 12,309 g / mol was used.

Comparative Preparation Example 2 Preparation of Unmodified PEO 80-2 Copolymer

A PEO-PPO-PEO copolymer (hereinafter, referred to as 'PEO 80-2') having an average content of PEO units of 80 wt% and a weight average molecular weight of 7,972 g / mol was used.

[Example]

Example 1 Preparation of Hydrogels

To react with the composition ratio shown in Table 1 below to prepare a hydrogel.

Specifically, 90 g of a PEO-PPO-PEO copolymer modified in 250 g of sterilized purified water while maintaining an internal temperature of 10 ° C. and a stirring speed of 100 rpm, 5.1 g of the composite crosslinking agent prepared in Preparation Example 5, and an aqueous solution of calcium chloride 0.34 g was added and stirred.

The temperature of the reactor was increased to a temperature increase rate of 1 ° C./10 sec, up to 40 ° C., and stirred for 1 hour to complete the crosslinking reaction. Upon completion of the crosslinking reaction, the reactor temperature was reduced to a cooling rate of 1 ° C./10 sec while cooling to a minimum of 4 ° C. while maintaining a vacuum pressure of 0.08 MPa. Thereafter, the reaction was terminated by stirring for 24 hours while maintaining the reactor internal temperature at 10 to 15 ° C. and a stirring speed of 100 rpm.

Hydrogel prepared by the above method was stabilized by storage at 4 ℃ for 24 hours, and then sterilized by irradiation of 5 to 12 kGy gamma rays for 4 to 8 hours.

division Copolymer (% by weight) Additives, parts by weight ¹⁾ PEO 70-1 PEO 70-2 PEO 80-1 PEO 80-2 Composite crosslinker ²⁾ Cross-linking agent
Mixture ³⁾ Chloride
calcium Example 1 100 - 0 - One - 0.1 Example 2 90 - 10 - One - 0.1 Example 3 80 - 20 - One - 0.1 Example 4 70 - 30 - One - 0.1 Comparative Example 1 - 100 - 0 One - 0.1 Comparative Example 2 - 90 - 10 One - 0.1 Comparative Example 3 - 80 - 20 One - 0.1 Comparative Example 4 - 70 - 30 One - 0.1 Comparative Example 5 - 100 - 0 - One 0.1 Comparative Example 6 - 90 - 10 - One 0.1 Comparative Example 7 - 80 - 20 - One 0.1 Comparative Example 8 - 70 - 30 - One 0.1 1) Additives are expressed in parts by weight of the additive based on 100 parts by weight of the copolymer.
2) Composite crosslinking agent: Composite crosslinking agent obtained in Preparation Example 1
3) Crosslinking agent mixture: HA (hyaluronic acid) / SA (sodium alginate) = 1/1 weight ratio crosslinking agent mixture

[Experimental Example]

Experimental Example 1. Measurement of adhesion and hydrophilicity of the hydrogel

The adhesive strength and hydrophilicity of each of the hydrogels prepared in Examples and Comparative Examples were measured and shown in Table 2 below.

division Adhesion (kgf / cm, 36 ℃) ¹⁾ Hydrophilicity ²⁾ Example 1 0.017 ± 0.005 34 ± 1 ° Example 2 0.020 ± 0.005 35 ± 1 ° Example 3 0.018 ± 0.005 32 ± 1 ° Example 4 0.015 ± 0.005 30 ± 1 ° Comparative Example 1 0.007 ± 0.002 16 ± 1 ° Comparative Example 2 0.010 ± 0.002 17 ± 1 ° Comparative Example 3 0.008 ± 0.002 15 ± 1 ° Comparative Example 4 0.005 ± 0.002 14 ± 1 ° Comparative Example 5 0.005 ± 0.0015 15 ± 1 ° Comparative Example 6 0.006 ± 0.0015 16 ± 1 ° Comparative Example 7 0.004 ± 0.0015 13 ± 1 ° Comparative Example 8 0.003 ± 0.0015 10 ± 1 ° 1) Adhesive force: Attach the sample to 1 * 1 cm of ABS material by using 0.1 kgf / cm tensioning device and pull it vertically at room temperature 36 ℃ to measure the adhesive force.
2) Hydrophilicity: The prepared hydrogel was dried at room temperature to prepare a 0.1 mm thick film, and a drop of 15 ° C distilled water was added dropwise to measure the contact angle after 30 seconds.

According to the results of Table 2, the hydrogels of Examples 1 to 4 according to the present invention can be stably adhered to the desired tissue in vivo because the viscosity at 36 ° C. is maintained at a high temperature. . On the contrary, the hydrogels of Comparative Examples 1 to 8 have a weak viscosity at 36 ° C. so that they can be easily separated from the desired tissue in vivo.

In addition, according to the results of Table 2, it was found that the hydrophilicity and adhesive force of the hydrogel is closely related. In particular, the hydrogel of Comparative Example 4 maintained a higher hydrophilicity than Example 2, and it can be seen that the viscosity at 36 ° C was kept particularly low.

Experimental Example 2 LCST of Hydrogel

In order to measure the LCST (Lower Critical Solution Temperature) of the temperature-sensitive hydrogel in vitro , the experiment was carried out by the following method. The sol-gel was measured by increasing 1 ° C. per 30 minutes in an 11% CO ₂ incubator. Average data were measured within the statistical range using 10 or more samples per temperature, and the results are shown in Table 3 below.

Experimental Example 3. Animal Experiment

In order to check whether the hydrogel of the present invention is present in vivo and adhered to tissues, animal experiments were conducted in the following manner.

The experimental animals used male New Zealand white rabbits 2 months old. The animals were anesthetized with ethyl ether, and then the incision was made. A cut of 1 cubic centimeter was made on the epidermal part of the abdominal wall, and a scar was formed using a surgical instrument. Then, the hydrogels prepared in Examples 1 to 4 or Comparative Examples 1 to 8 were applied to the wound area at about twice the area and then sutured. After 7 days, the surgical site was resectioned to determine the presence of hydrogel and adhesion of tissue.

Table 3 shows the presence of hydrogels and whether tissues are adhered to the naked eye, and Table 4 shows the determination of tissue adhesion as a Hooker score and a Knightly score.

division LCST (℃)
in vitro 7 days later
Hydrogel presence 7 days later
Tissue adhesion Example 1 23 ± 1 ◎ Do not coalesce Example 2 27 ± 1 ◎ Do not coalesce Example 3 32 ± 1 ◎ Do not coalesce Example 4 36 ± 1 ◎ Do not coalesce Comparative Example 1 19 ± 2 ● Partial adhesion Comparative Example 2 23 ± 2 ● Partial adhesion Comparative Example 3 26 ± 2 ● Partial adhesion Comparative Example 4 31 ± 2 ● Partial adhesion Comparative Example 5 17 ± 2 × Adhesion occurs Comparative Example 6 20 ± 2 × Adhesion occurs Comparative Example 7 25 ± 2 × Adhesion occurs Comparative Example 8 28 ± 2 × Adhesion occurs Determine the presence of a hydrogel:
(Double-circle): It means that a hydrogel exists as a gel uniformly around tissue.
●: Means that the hydrogel is non-uniformly partly around the tissue as a gel.
×× means that no hydrogel is present around the tissue.

According to the LCST test results of Table 3, the hydrogels prepared in Examples 1 to 4 according to the present invention were compared to Comparative Examples 1 to 4 and Comparative Examples 5 to 8. And less variation in adhesion occurrence and more stable physical properties can be confirmed through the table. In addition, as shown in Table 2, the hydrogels of Comparative Examples 1 to 4 and Comparative Examples 5 to 8 have low adhesion in the in vivo temperature range, as well as high or unstable LCST. Can be.

In addition, according to animal experiments, the hydrogels prepared in Examples 1 to 4 according to the present invention, when the hydrogel was present after 7 days after opening, no adhesion of tissues occurred, but the hydrogels prepared in Comparative Examples 1 to 4 When 7 days later, the hydrogel was not present or partially present even if the hydrogel was present, and in Comparative Examples 5 to 8, no hydrogel was present and coalescence occurred. Therefore, it can be seen that it does not function as an effective tissue adhesion inhibitor because it does not show an even distribution as in Examples 1 to 4.

division Hooker score Knightly score Example 1 1.24 ± 0.74 1.27 ± 0.75 Example 2 0.70 ± 0.86 1.01 ± 1.06 Example 3 1.21 ± 0.71 1.23 ± 0.84 Example 4 1.51 ± 0.84 1.81 ± 0.81 Comparative Example 1 2.74 ± 0.87 3.21 ± 1.00 Comparative Example 2 2.42 ± 0.78 2.98 ± 0.91 Comparative Example 3 2.55 ± 0.91 3.10 ± 1.01 Comparative Example 4 2.91 ± 0.81 3.76 ± 0.90 Comparative Example 5 2.97 ± 0.85 3.42 ± 1.21 Comparative Example 6 2.58 ± 0.79 3.01 ± 0.87 Comparative Example 7 2.89 ± 0.89 3.25 ± 1.04 Comparative Example 8 2.98 ± 0.97 3.89 ± 0.95

As described above, the hydrogel of the present invention has a function of preventing biocompatibility, temperature sensitivity, and tissue adhesion, and thus may be widely used as an anti-adhesion agent or a filler for skin molding used in surgical procedures.

Claims (11)

V) 100 parts by weight of a polyethylene-polypropylene-polyethylene copolymer modified at one end thereof with a lactic acid and at the other end thereof with ethylene glycol;
Ii) 1-2 parts by weight of a crosslinking agent combined with at least two crosslinking agents selected from hyaluronic acid, adipic dihydrazide and sodium alginate, and
Iii) 0.1 to 1 part by weight of at least one valence 2 metal compound selected from the group consisting of calcium, magnesium, barium, zinc, strontium and iron (II)
Hydrogel, characterized in that prepared by the crosslinking reaction.
The method of claim 1,
The copolymer is a hydrogel, characterized in that the content of 4 to 7 mol%, the content of ethylene glycol is 3 to 6 mol%.
The method of claim 1,
The copolymer is a hydrogel, characterized in that the content of polyethylene (PEO) unit is 65 to 85% by weight, the weight average molecular weight of 7,000 to 16,000 g / mol.
The method of claim 3, wherein
The copolymer has a polyethylene (PEO) unit content of 74 to 82% by weight and a weight average molecular weight of 7,500 to 12,000 g / mol of polyethylene-polypropylene-polyethylene copolymer 70 to 95% by weight; Hydrocarbon, characterized in that the copolymer mixture consisting of 5 to 30% by weight of polyethylene-polypropylene-polyethylene copolymer having a content of polyethylene (PEO) unit of 69 to 73% by weight and a weight average molecular weight of 12,500 to 16,000 g / mol Gel.
The method of claim 1,
The crosslinking agent is a crosslinking agent of hyaluronic acid-adipic acid dihydrazide-sodium alginate, a crosslinking agent of hyaluronic acid-adipic acid dihydrazide-hyaluronic acid, and sodium alginate-adipic acid dihydrazide-sodium alginate. Hydrogel, characterized in that at least one member selected from the group consisting of a crosslinking agent.
The method of claim 1,
The composite crosslinking agent is a hydrogel, characterized in that the composite crosslinking agent of hyaluronic acid-adipic acid dihydrazide-sodium alginate.
The method of claim 1,
The valence 2 metal compound is Mg ²⁺ , Ca ²⁺ , Sr ²⁺ , Ba ²⁺ , Be ²⁺ , Cr ²⁺ , Co ²⁺ , Cu ²⁺ , Fe ²⁺ , Mn ²⁺ , Sn ^{2+ Hydrogen} , characterized in that the metal compound containing at least one metal selected from the group consisting of, Ni ²⁺ , and Zn ²⁺ .
The method of claim 1,
The hydrogel has a low critical solution temperature (LCST) at a concentration of 20 to 35% by weight (w / w) of solids, characterized in that the hydrogel is 24 to 30 ℃.
The method of claim 1,
The hydrogel is characterized in that the hydrogel is prepared in powder form.
A tissue adhesive agent for surgical surgery, characterized in that the hydrogel of any one selected from claim 1 to claim 9.
A skin cosmetic filler, characterized in that the hydrogel of any one selected from claim 1 to claim 9.