阅读说明：本技术 烷基膦酰基改性胶原基材料及制备方法、改性胶原、应用 (Alkyl phosphono modified collagen-based material, preparation method, modified collagen and application ) 是由 樊璐露 钱松 于 2020-06-30 设计创作，主要内容包括：本发明属于生物制药技术领域,具体涉及一种烷基膦酰基改性胶原基材料及制备方法、改性胶原、应用。本烷基膦酰基改性胶原基材料的制备方法,包括：制备胶原溶液；对胶原溶液中的胶原进行烷基膦酰基改性反应；将反应产物进行透析；取透析液浇铸至模具中；冷冻干燥处理,得到烷基膦酰基改性胶原基材料。在胶原蛋白分子中引入含磷基团,可以对胶原蛋白的功能发挥起到重要的作用。(The invention belongs to the technical field of biological pharmacy, and particularly relates to an alkyl phosphono modified collagen-based material, a preparation method, modified collagen and application thereof. The preparation method of the alkyl phosphono modified collagen-based material comprises the following steps: preparing a collagen solution; performing an alkyl phosphono modification reaction on collagen in the collagen solution; dialyzing the reaction product; taking the dialysate and casting the dialysate into a mold; and (4) carrying out freeze drying treatment to obtain the alkyl phosphono modified collagen-based material. The introduction of phosphorus-containing groups into collagen molecules can play an important role in the function of collagen.)

1. A method of preparing an alkylphosphonyl-modified collagen-based material, comprising:

preparing a collagen solution;

performing an alkyl phosphono modification reaction on collagen in the collagen solution;

dialyzing the reaction product;

taking the dialysate and casting the dialysate into a mold;

and (4) carrying out freeze drying treatment to obtain the alkyl phosphono modified collagen-based material.

2. The production method according to claim 1,

the alkylphosphonyl modification reaction comprises:

mixing the collagen solution with a 4- (dialkylphosphono) butanoic acid solution;

adding a cross-linking agent, heating and stirring at constant temperature to perform alkyl phosphono group modification reaction.

3. The production method according to claim 2,

the crosslinking agent comprises: 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride and N-hydroxysuccinimide in a molar ratio of 1: (1-1.5).

4. The production method according to claim 2,

the alkyl substituent of the 4- (dialkyl phosphonyl) butyric acid comprises at least one of linear chain or branched chain alkyl with the carbon number of 1-20;

the 4- (dialkylphosphono) butanoic acid solution is an aqueous 4- (dialkylphosphono) butanoic acid solution, wherein

The concentration of 4- (dialkylphosphono) butanoic acid is from 0.01 to 1 mol/L.

5. The production method according to claim 1,

the collagen solution comprises: a collagen acid solution or a collagen aqueous solution; wherein

The concentration of collagen is 5-50 mg/mL.

6. The production method according to claim 5,

the collagen acid solution is a solution of collagen dissolved in acetic acid or hydrochloric acid; wherein

The concentration of acetic acid or hydrochloric acid is 0.01-0.1 mol/L.

7. The production method according to claim 2,

the mass ratio of the collagen to the 4- (dialkyl phosphonyl) butyric acid is 1: (0.05-5);

the molar ratio of 4- (dialkylphosphine) butyric acid to 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride is 1: (1-1.5).

8. An alkyl phosphono modified collagen characterized in that,

the molecular structural formula is as follows:

wherein

Col-is collagen, and R is C1-20 linear or branched alkyl.

9. An alkylphosphonyl-modified collagen-based material prepared by the method of claim 1.

10. Use of an alkylphosphonyl modified collagen-based material as claimed in claim 9 in wound repair.

Technical Field

The invention belongs to the technical field of biological pharmacy, and particularly relates to an alkyl phosphono modified collagen-based material, a preparation method, modified collagen and application thereof.

Background

Collagen is the main component of extracellular matrix, and is the most abundant and widely distributed protein in animals. The collagen has unique tissue structure and biological function, plays an important role in maintaining normal physiological functions of cells, tissues and organs and repairing damage, has the characteristics of low immunogenicity, biodegradability, good biocompatibility, good mechanical property and the like, and is a good wound repair material.

Phosphorus is an important element in the life process, and no living body containing no phosphorus is found in the life process. Phosphorus is important because most of the reactions involved in the enzymes in the living body are regulated by their phosphorylation and dephosphorylation, including proliferation and differentiation of cells, nerve activity, muscle contraction, metabolism, tumorigenesis, etc., and it can be said that the normal operation of physiological functions in the living body is not involved in the phosphorus-containing compounds.

Disclosure of Invention

The invention provides an alkyl phosphono modified collagen-based material, a preparation method, modified collagen and application.

In order to solve the above technical problems, the present invention provides a method for preparing an alkyl phosphono modified collagen-based material, comprising: preparing a collagen solution; performing an alkyl phosphono modification reaction on collagen in the collagen solution; dialyzing the reaction product; taking the dialysate and casting the dialysate into a mold; and (4) carrying out freeze drying treatment to obtain the alkyl phosphono modified collagen-based material.

In a second aspect, the present invention also provides an alkyl phosphono modified collagen, having a molecular structural formula:

wherein Col-is collagen, and R is linear chain or branched chain alkyl with carbon atom number of 1-20.

In a third aspect, the present invention also provides an alkyl phosphono modified collagen-based material prepared by the preparation method as described above.

In a fourth aspect, the present invention also provides a use of an alkyl phosphono modified collagen-based material as described above in wound repair.

The invention has the beneficial effects that the alkylphosphono modified collagen base material, the preparation method, the alkylphosphono modified collagen and the application thereof carry out alkylphosphono grafting modification on the collagen by adopting a chemical method, and a phosphorus-containing group is introduced into collagen molecules, so that the alkylphosphono modified collagen base material can play an important role in the function of the collagen, thereby leading the alkylphosphono modified collagen base material to be a functionalized collagen base wound repair material with excellent physical and chemical properties and controllable molding.

Additional features and advantages of the invention will be set forth in the description which follows, and in part will be obvious from the description, or may be learned by practice of the invention. The objectives and other advantages of the invention will be realized and attained by the structure particularly pointed out in the written description and claims hereof as well as the appended drawings.

In order to make the aforementioned and other objects, features and advantages of the present invention comprehensible, preferred embodiments accompanied with figures are described in detail below.

Drawings

In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the embodiments or the prior art will be briefly described below, and it is obvious that the drawings in the following description are some embodiments of the present invention, and other drawings can be obtained by those skilled in the art without creative efforts.

FIG. 1 is a flow diagram of a process for the preparation of alkylphosphonyl-modified collagen-based materials according to the invention.

Detailed Description

To make the objects, technical solutions and advantages of the embodiments of the present invention clearer, the technical solutions of the present invention will be clearly and completely described below with reference to the accompanying drawings, and it is apparent that the described embodiments are some, but not all embodiments of the present invention. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.

A first part: elucidating the specific technical scheme

In order to introduce a phosphorus-containing group into a collagen molecule, it plays an important role in the function of collagen. Referring to fig. 1, the present invention provides a method for preparing an alkyl phosphono modified collagen-based material, comprising: preparing a collagen solution; performing an alkyl phosphono modification reaction on collagen in the collagen solution; diluting the reaction product (i.e. the reaction solution after the reaction) and then dialyzing; taking the dialysate and casting the dialysate into a mold; and (4) carrying out freeze drying treatment to obtain the alkyl phosphono modified collagen-based material.

Optionally, the dialysis bag can be a cellulose membrane dialysis bag, and the residual molecular weight of the dialysis bag is 8-14KDa, optionally 10KDa and 12 KDa. Small molecule molecules, typically less than 8000, can be removed.

Optionally, the freeze-drying treatment comprises pre-freezing at-20 deg.C for 4 hr, and then freezing at-80 deg.C for 24 hr to obtain solid collagen material for storage.

As an alternative embodiment of the collagen solution.

Optionally, the collagen solution comprises: a collagen acid solution or a collagen aqueous solution; wherein the concentration of collagen is 5-50mg/mL, optionally 15mg/mL, 25mg/mL, 40 mg/mL. And the collagen acid solution is a solution of collagen dissolved in acetic acid or hydrochloric acid; wherein the concentration of acetic acid or hydrochloric acid is 0.01-0.1mol/L, optionally 0.04mol/L, 0.06mol/L, and 0.08 mol/L.

Alternatively, the collagen may be, for example, but not limited to, collagen extracted from animal tissue or recombinant collagen prepared by biomedical engineering techniques.

The collagen solution of the embodiment can be carried out in a certain range according to the reaction conditions, the collagen solution is a reaction medium, and different concentrations of the collagen solution can correspond to different reaction conditions, so that the reaction effect is improved.

As an alternative embodiment of the alkylphosphonyl modification reaction.

The alkylphosphonyl modification reaction comprises: mixing the collagen solution with a 4- (dialkylphosphono) butanoic acid solution; adding a cross-linking agent, heating and stirring at constant temperature to react the collagen with the 4- (dialkyl phosphonyl) butyric acid. The reaction formula of the alkyl phosphonyl group modification reaction is

Wherein Col-is collagen, and R is linear chain or branched chain alkyl with carbon atom number of 1-20. Specifically, the mass ratio of the collagen to the 4- (dialkylphosphono) butyric acid is 1: (0.05-5), which can be selected from (1: 0.1) or (1: 1) or (1: 3); the molar ratio of 4- (dialkylphosphine) butyric acid to 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride is 1: (1-1.5), optionally (1: 1.2) or (1: 1.4), collagen-based materials of different degrees of modification can be prepared.

Optionally, the cross-linking agent comprises: 1-ethyl-3- (3-dimethylaminopropyl) carbodiimide hydrochloride and N-hydroxysuccinimide in a molar ratio of 1: (1-1.5), optionally 1: 1.2 or 1: 1.3. the cross-linking agent is easy to dissolve in water, has low toxicity, is easy to remove in the post-treatment process, and can ensure that the reaction is more complete.

Optionally, the alkyl substituent of the 4- (dialkylphosphono) butanoic acid includes at least one of a linear or branched alkyl group having 1 to 20 carbon atoms, such as, but not limited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, sec-butyl, isobutyl, and the like.

Alternatively, the 4- (dialkylphosphono) butanoic acid solution is an aqueous 4- (dialkylphosphono) butanoic acid solution, wherein the concentration of 4- (dialkylphosphono) butanoic acid is 0.01 to 1mol/L, alternatively 0.05mol/L, 0.1mol/L,0.5mol/L, and is generally adjusted according to the modification degree requirement and the reaction concentration.

Optionally, the heating temperature of the constant-temperature heating and stirring is 25-60 ℃, optionally 35 ℃, 45 ℃ and 50 ℃, and the heating time is 12-72 hours, so that the reaction can be stably performed, the heating temperature is too low to cause incomplete reaction, and the heating temperature is too high to cause side reaction, even collagen denaturation.

The alkylphosphonyl modification reaction of the embodiment is to react collagen with 4- (dialkylphosphono) butyric acid under the action of a cross-linking agent, so that a phosphorus-containing group is introduced into a collagen molecule, which can play an important role in the function of collagen, for example, the activity of most enzymes in a living body is regulated by means of phosphorylation and dephosphorylation, and the activity of the enzymes can be regulated by introducing a phosphono group.

In addition, the aqueous solution of each substance in the present case may be an aqueous solution of each substance dissolved in deionized water.

Further, the invention provides an alkyl phosphono modified collagen, the molecular structural formula of which is:

col-is collagen, and R is C1-20 linear or branched alkyl.

Further, the present invention provides an alkylphosphonyl modified collagen-based material prepared by the preparation method as described above.

Further, the invention also provides an application of the alkyl phosphono modified collagen-based material in wound repair.

Optionally, the wound surface is such as, but not limited to, a scald wound surface, a burn wound surface, a damaged skin, and the like.

A second part: some examples are given below