KR20160125332A - A novel process for crosslinked hyaluronic acid derivatives - Google Patents

Abstract

Provided in the present invention is a novel method for manufacturing cross-linked hyaluronic acid derivatives in an effective manner, by washing the cross-linked hyaluronic acid derivatives using a washing device installed with a washing basket. The washing device removes a cross-linking agent used for a reaction after a cross-linking reaction by mixing a cross-linking agent and hyaluronic acid.

Description

[Technical Field] The present invention relates to a novel process for crosslinking hyaluronic acid derivatives,

The present invention relates to a method for effectively washing a crosslinked hyaluronic acid derivative in a gel state in an aseptic state, and a method for quickly and uniformly washing a crosslinking agent used in a crosslinking reaction while maintaining the shape of a hyaluronic acid crosslinked product cut to a predetermined size, The present invention relates to an apparatus and a method for easily recovering water.

Hyaluronic acid is composed of D-type glucuronic acid and N-acetylglucosamine, and the glycosidic bonds of β-1,4 and β-1,3 are alternately and repeatedly It is a linearly linked biopolymer of disaccharide that is widely distributed throughout the human body in connective tissue, epithelial tissue, and nerve tissue. Since hyaluronic acid is a bio-derived polymer, it has little side effects when applied to patients and has hydrophilic properties due to its chemical structure. It is also known as a biocompatible biocompatible material which has a high water content, has a buffering effect of physical impact on the joints, a lubrication effect on friction, and is involved in flexibility of the skin. In addition, it has a protective property against bacterial invasion from the outside, and has hyaluronidase in vivo, so that it is biodegraded when the human body is inserted.

Hyaluronic acid is the main component of the skin and is involved in skin recovery. Hyaluronic acid has excellent biocompatibility and viscoelasticity, and thus can be used for medical and medical purposes such as ophthalmic surgical aids, joint function improvers, drug delivery materials, postoperative adhesion inhibitors, wrinkle improving agents, shaping aids, cell culture scaffolds and eyelashes, Cosmetics, and the like. However, since it is easily decomposed in vivo or in acidic or alkaline conditions, its use is limited, and thus efforts to develop structurally stable hyaluronic acid derivatives have been widely carried out.

Recently, a cross-linked hyaluronic acid derivative composition has been actively studied for commercial purposes in order to increase the skin of the wrinkle-improving agent and the molding assistant. Typically, such a hyaluronic acid derivative composition is more resistant to degradation than a non-crosslinked hyaluronic acid composition, so that frequent re-sizing is not required, and a crosslinking agent is used to form hyaluronic acid molecules Linking.

  The crosslinked hyaluronic acid derivative is generally prepared by mixing hyaluronic acid and a crosslinking agent in an aqueous solution with stirring and chemically bonding the hyaluronic acid polymer chain to each other with a crosslinking agent. Here, when such a crosslinked hyaluronic acid derivative is administered in vivo, there is a great concern that the crosslinking agent component is recognized as a foreign substance in the living body and adversely affects such as causing an inflammatory reaction. In fact, according to a 2003 paper published by the US Food and Drug Administration (FDA) in cancer research, BDDE, the most commonly used crosslinking agent, has been reported to be dangerous if the residual amount is more than 2 ppm. It is an important issue. It is not easy to completely remove the crosslinking agent. The crosslinking agent of hyaluronic acid usually induces a crosslinking reaction in a liquid phase by mixing a crosslinking agent and hyaluronic acid in an aqueous alkaline solution. However, in general, since the reactivity is very low, It is necessary to increase the reaction concentration or to add a large amount of crosslinking agent. In this case, it is more difficult to completely remove the crosslinking agent added in the purification process.

U.S. Patent Application Publication 2003/0094719 A1 discloses a method for producing a crosslinked hyaluronic acid derivative by a simple method. A method of preparing a hyaluronic acid solution and adjusting the pH by using sodium hydroxide and then cross-linking with a multifunctional epoxy compound is provided. Preferred multifunctional epoxy compounds used as a crosslinking agent include 1,4-butanediol diglycidyl ether (BDDE), ethylene glycol diglycidyl ether (EGDGE), and sorbitol polyglycidyl ether. However, this invention does not disclose a method for efficiently removing the residual crosslinking agent after the crosslinking reaction.

On the other hand, U.S. Patent Application Publication No. 2006/0105022 A1 discloses that a cross-linked hyaluronic acid derivative gel is decomposed after it is used in the human body. At this time, since the residual cross-linking agent is concerned about side effects such as inflammatory reaction and the like, Discloses a method for producing a crosslinked hyaluronic acid derivative gel having a bonding ratio. Examples of crosslinking agents include divinyl sulfone, BDDE, and EGDGE. However, this invention also does not suggest a method for effective removal of the residual crosslinking agent after the crosslinking reaction.

U.S. Patent No. 8357795 discloses a method for removing unreacted crosslinking agent and dialyzing with a buffer solution for a maximum period of several days while filling a dialysis membrane with a crosslinking agent and stabilizing the bath in order to stabilize the pH near neutrality . This method is not suitable for mass production, and it is difficult to remove the crosslinking agent or it takes a long time.

  The present invention solves the problem that it is difficult to remove the crosslinking agent remaining after the crosslinking reaction of hyaluronic acid and takes long time or is not suitable for mass production in the conventional method, and thereby, the crosslinked hyaluronic acid derivative is effectively washed .

Since the cross - linking agents used in the cross - linking reaction of hyaluronic acid have a high possibility of side effects such as inflammation reaction in the human body, their removal is considered to be a very important problem. However, in the process of preparing a crosslinked hyaluronic acid derivative, it is very difficult to sufficiently remove the residual crosslinking agent after crosslinking reaction of the hyaluronic acid and the crosslinking agent.

  Accordingly, it is an object of the present invention to provide a method and apparatus for efficiently and massively washing a crosslinking agent remaining after a crosslinking reaction in a short time in the production of a crosslinked hyaluronic acid derivative and easily recovering the crosslinked crosslinked product.

To this end, the present invention provides a method for cleaning a crosslinked hyaluronic acid derivative using a cleaning device equipped with a basket.

  A method for efficiently washing a crosslinked hyaluronic acid derivative according to the present invention

Inserting a cross-section cut into a predetermined size into a wash basket formed with a mesh,

Fixing the cleaning basket to the inside of the cleaning tank,

Injecting the buffer solution into the washing tank,

Rotating the wash tank to wash the cross-linked product in the wash basket into the buffer solution,

Draining the washed buffer solution,

Repeatedly injecting the buffer solution, rotating the washing tank, discharging the buffer solution until the crosslinking agent used in the reaction is completely removed,

And recovering the washed cross-linked hyaluronic acid derivative.

It has been a classic method to wash the crosslinked hyaluronic acid derivative to completely remove the crosslinking agent and to make a hyaluronic acid derivative in a gel state through dialysis for a long time.

However, the present invention provides a simple and efficient method of removing the crosslinking agent using a cleaning device equipped with a mesh-formed cleaning basket. In the present invention, the cross-linking agent is cut to a predetermined size and the cross-linking agent used in the reaction is washed while maintaining the size thereof. At this time, the alkali component (usually, sodium hydroxide) used in the cross-linking reaction is simultaneously removed. The washing buffer solution may be a conventional solution, for example, phosphate buffered saline (PBS) or the like.

  If the crosslinked material in the lump state is not cut or is cut too large, it takes a long time to perform the washing and the uniformity may not be uniformly progressed to the inside of the crosslinked material. On the contrary, if the material is cut too small, There may be losses. Therefore, the hyaluronic acid derivative can be produced efficiently by removing the cross-linking agent used in the reaction while maintaining the shape and cutting the hyaluronic acid derivative of 0.5-1.0 cm in length and easily into a cubic shape by using a cutter have.

    This process enhances the efficiency and convenience of washing, and the formation of particles of homogeneous sized hyaluronic acid derivatives is advantageous.

The method of washing the crosslinked hyaluronic acid using the washing apparatus of the present invention can be carried out while maintaining the size and shape of the crosslinked hyaluronic acid derivative having a constant size and shape in an aseptic state and also can be carried out without loss of the crosslinked hyaluronic acid derivative It is possible to recover easily and efficiently within the time. The process according to the invention also has the advantage that the product of the final crosslinked hyaluronic acid after removal of the crosslinking agent can be obtained in large quantities with reproducible reproducibility in homogeneous quality. Also, the removal of the alkali component (usually, sodium hydroxide) used in the crosslinking reaction can be performed at the same time.

1 is a front view of a cleaning apparatus according to the present invention.
2 is a plan view of the cleaning apparatus of Fig.
Figure 3 is a front view of the washing tank in the washing apparatus of Figure 1;
4 is a right side sectional view of the washing tank of Fig.
Figure 5 is a top view of the washing tank of Figure 3;
Figure 6 is a front view of the wash basket in the wash tank of Figure 3;
Figure 7 is a top view of the wash basket in the wash tank of Figure 3;

DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS Hereinafter, the present invention will be described in detail with reference to the accompanying drawings. However, the present invention is not limited or limited by the following description.

Fig. 1 is a front view of the cleaning apparatus according to the present invention, Fig. 2 is a plan view of the cleaning apparatus of Fig. 1, Fig. 3 is a front view of the cleaning tank in the cleaning apparatus of Fig. Fig. 5 is a plan view of the cleaning tank of Fig. 3, Fig. 6 is a front view of the cleaning basket in the cleaning tank of Fig. 3, and Fig. 7 is a plan view of the cleaning basket in the cleaning tank of Fig.

), 컨트롤 패널(

)을 포함한다.
1 to 7, a washing apparatus for washing a crosslinked hyaluronic acid derivative includes a washing tank (1), a washing basket (4), a rotating motor

), The control panel (

).

)로 이루어져 있다. 세척 바스켓(④)의 옆면과 밑면은 전체에 메쉬(mesh)가 형성되어 있어 완충 용액의 출입은 자유롭지만 가교된 히알루론산 유도체는 세척 바스켓에서 빠져나가지 않도록 제공된다. 세척 바스켓은 세척 탱크(①)의 상부와 하부에 장착할 수 있으며, 세척 탱크(①)에 탈부착이 용이하다.
The cleaning basket (4) has a hollow hexahedron or a cylindrical shape, preferably a cylindrical shape, and the upper surface is a cleaning basket cover

). The sides and bottom of the cleaning basket (④) are provided with a mesh on the whole so that the buffer solution is free to go in and out but the crosslinked hyaluronic acid derivative is provided so as not to escape from the cleaning basket. The cleaning basket can be mounted on the top and bottom of the cleaning tank (①), and it is easy to attach and detach it to the cleaning tank (①).

The mesh of the cleaning basket (4) is suitably 500 탆 to 4,000 탆, preferably 1,500 탆 to 2,500 탆. If the mesh is too small, the efficiency of washing is low because the buffer solution is difficult to get in and out of the washing process. If the mesh is too large, the crosslinked product cut out to a certain size may escape from the washing basket (④) The resulting fragments of the crosslinked hyaluronic acid derivative may escape and the yield may be reduced.

  The cleaning tank (1) may be a hexahedron, a cylinder, or a double cone type.

)를 이용하여 360도 회전이 가능하며, 필요에 따라 회전 각도는 1도~360도로 설정하여 사용할 수 있다. 회전 속도의 설정이 가능하고, 회전 시간도 설정이 가능하다. 완충용액을 채워 일정 시간 회전하여 세척하면 완충용액을 배출하고, 새로운 완충용액을 주입하고 세척하는 작업을 반복하여 세척한다.The cleaning tank (1) is a rotary motor

), And the rotation angle can be set to 1 degree ~ 360 degrees as needed. The rotation speed can be set and the rotation time can be set. After filling the buffer solution and rotating it for a certain period of time, the buffer solution is drained, and a new buffer solution is injected and washed repeatedly.

Also, the rotation speed of the washing tank (1) proceeds at a slow speed of 1 rpm to 10 rpm, preferably 2 rpm to 6 rpm, so that the shape of the hyaluronic acid derivative crosslinked in the buffer solution is not crushed and is not damaged. If the rotating speed is high, it is pulverized and passes through the mesh of the washing basket (4) and is lost, so that the yield can be reduced. The rotation time is 10 to 90 minutes, preferably 30 to 50 minutes, and the efficiency of washing is improved by repeating the discharge and injection of the buffer solution. It was confirmed that the number of times of washing for normal washing was 3 to 5 times, and the crosslinking agent was completely removed. Therefore, the crosslinking agent can be easily removed in a short time, thereby providing a suitable method for mass production.

)을 이용하여 이송하고, 배출은 배출부 패럴(⑫)을 이용하여 진행한다. Injection and release of the buffer solution can proceed easily through the ferrule. The buffer solution prepared in a separate container contains the injection section ferrule (⑬ or ⑭ or ⑮ or

), And the discharge is carried out by using the discharge section paralel (12).

,

)를 통해 세척 탱크의 내부 진행 사항을 확인하며 작업할 수 있다.Also,

,

) To check the internal progress of the washing tank.

  Although the present invention has been described and illustrated in detail, it is clearly understood that the same is by way of illustration and example only and is not to be taken by way of limitation in the scope of the appended claims. The present invention is not limited thereto.

주입부 패럴

,

사이트 글라스

컨트롤 패널

회전 모터

안전 커버

세척 바스켓 커버

세척 바스켓 손잡이 ① Cleaning tank
④ Cleaning basket
⑦ Cleaning tank cover
⑫ Discharge Paralel
⑬, ⑭, ⑮,

Injection part

,

Sight Glass

Control panel

Rotation motor

Safety cover

Cleaning basket cover

Wash basket handle

Claims (4)

A washing device for mixing the crosslinking agent and hyaluronic acid to remove the crosslinking agent used in the reaction after the crosslinking reaction,
In this cleaning apparatus,
The washing device has a rotating motor, the rotating motor is connected to the washing tank, the washing basket is equipped with a washing basket, and the washing basket is formed with a mesh on the whole surface so that the washing buffer solution in the washing tank is freely And has a structure capable of flowing out. The washing apparatus according to claim 1, wherein the mesh of the washing basket is 500 mu m to 4,000 mu m. The method of any one of claims 1 to 3, wherein the washing tank is washed by rotating the washing tank equipped with the washing basket by 1 to 360 degrees, discharging the buffer solution, injecting a new buffer solution, Of a hyaluronic acid derivative The cleaning method of a crosslinked hyaluronic acid derivative according to claim 3, wherein the washing tank is rotated at a rotational speed of 1 rpm to 10 rpm and a washing time of 10 minutes to 90 minutes

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