Chlorinated trimethyl amino hydroxypropyl hydroxybutyl chitosan derivative as well as preparation method and application thereof

Abstract

The invention discloses a chlorinated trimethyl amino hydroxypropyl hydroxybutyl chitosan derivative as well as a preparation method and application thereof, and belongs to the field of biomedical materials. According to the preparation method, chitosan is subjected to trimethyl amino hydroxypropyl group chloride modification so as to obtain trimethyl amino hydroxypropyl chitosan chloride, and then hydroxybutylation modification is carried out so as to obtain the modified chitosan; or the chitosan is firstly subjected to hydroxybutylation modification and then subjected to chlorinated trimethylamino hydroxypropyl group modification, and the chitosan is obtained; or chitosan is subjected to hydroxybutyl and chlorinated trimethylamino hydroxypropyl group modification at the same time to obtain the product. The chitosan derivative disclosed by the invention is a positively charged high-molecular polymer and non-amphoteric polyelectrolyte, does not have a self-compounding phenomenon caused by positive and negative charge attraction, and can be compatible with an osmotic pressure regulator of a strong-polarity ionic compound to obtain a temperature-sensitive solution with good performance. The material can be used as a temperature-sensitive cell carrier, a tissue engineering scaffold material, a drug controlled release material and a body surface superficial layer and deep layer tissue or in-vivo implant material.

Description

Technical Field

[0001] The invention belongs to the field of biomedical materials, and in particular relates to a trimethylaminohydroxypropylhydroxybutyl chloride chitosan derivative and a preparation method and application thereof. Background Art

[0002] Chitosan is a natural high molecular weight cationic polysaccharide with the advantages of good biocompatibility, no immunogenicity, no toxic side effects, and good biosafety. It also has multiple functions such as hemostasis, analgesia, antibacterial, and anti-infection, and can promote wound healing. It is easy to make into membranes and has good performance for preparing various dressings. After decades of continuous development, chitosan materials have gone through the first three generations of materials and have now developed to the fourth stage.

[0003] The first generation of chitosan materials is chitin extracted from the shells of crustaceans such as shrimps and crabs, and chitosan obtained by further deacetylation. Chitin is insoluble in general common solvents, and chitosan is soluble in some dilute acid solutions, but both are insoluble in neutral aqueous solutions. The second generation of chitosan materials is made into water-soluble materials by grafting soluble groups on the molecules of chitosan or chitosan. Representative materials are derivatives such as carboxymethyl chitosan, carboxymethyl chitosan, and chitosan quaternary ammonium salts. This type of material solves the problem that the first generation of chitosan materials are insoluble in water. In the medical field, it solves the problem that the acidic solution of the first generation of chitosan materials cannot enter the body for use, greatly expanding the application of chitosan materials in the human body.

[0004] The third generation of chitosan materials is made by grafting intelligent groups on the molecules of chitin or chitosan to make it an intelligent chitosan biomaterial, including temperature-sensitive intelligent materials, pH-sensitive biomaterials, pressure-sensitive biomaterials, magnetic-sensitive biomaterials, etc. Among them, the chitosan material obtained by grafting hydroxybutyl has been widely used for its excellent temperature sensitivity. Hydroxybutyl chitosan material not only solves the problem of water insolubility of the first generation of biomaterials, but also has temperature-sensitive properties. That is, its aqueous solution is in a solution state at low temperature (lower than its gel temperature), and after gradually heating up, it can be converted into a non-flowing gel state after being higher than its gel temperature, such as reaching body temperature of 37°C. This feature allows the material to be fixed and play a role for a long time after acting on the affected area. It can be used on wounds of almost any shape, including flat, wrinkled, gap-shaped, etc. After use, it is converted into a non-flowing gel attached to the wound surface, and will not be lost due to changes in body position, thereby achieving a long-term treatment effect, solving the problem that the second generation of carboxymethyl chitosan and other materials have high fluidity and are easy to leave the wound area that needs a barrier. Therefore, hydroxybutyl chitosan material has good wound healing and other biological activities.

[0005] The invention patent application with publication number CN 113908329 A discloses a hydroxybutyl chitosan hydrogel dressing and a preparation method thereof, wherein the auxiliary materials used in combination with hydroxybutyl chitosan are polyols containing three or more hydroxyl groups, such as glycerol, mannitol, sorbitol, etc., while highly polar ionic osmotic pressure regulators commonly used in pharmacopoeias, such as sodium chloride, potassium chloride, calcium chloride, magnesium chloride, potassium nitrate, sodium sulfate, etc., cannot be used in combination with hydroxybutyl chitosan preparations under normal conditions. This is because hydroxybutyl chitosan is compatible with the above-mentioned highly polar substances, and is a uniform and transparent flowing solution at low temperature, but becomes a white opaque gel when heated. The gel strength is low, and it is easy to break under the action of external force, and will not solidify again unless it is homogenized again at low temperature.

[0006] And if only aqueous solution is used, or the solution prepared by the compatibility of polyols such as glycerol and mannitol, it can reach low temperature and be easy to flow, and after being warmed up to near body temperature, it is converted into a non-flowing gel, and the gel strength is relatively large, and the gel is not scattered. But after the hydroxybutyl chitosan hydrogel dressing using this type of formula is used for human body, especially in the position with more body fluids, such as the peritoneal cavity and the wound surface position with more exudate, it will gradually shrink and precipitate, and can only play a short-term gel effect. And when used as a cell culture medium, the phenomenon of precipitation will also occur, that is, the matching degree of the hydroxybutyl chitosan hydrogel dressing and the physiological environment of human body also has certain defects. Summary of the invention

[0007] In view of the above problems, an object of the present invention is to provide a chitosan derivative grafted with trimethylamino hydroxypropyl chloride groups and hydroxybutyl groups, which can be combined with an osmotic pressure regulator of a strongly polar ionic compound (such as sodium chloride, potassium chloride, calcium chloride, magnesium chloride, potassium nitrate, sodium sulfate, etc.) to obtain a temperature-sensitive solution with good performance, and when forming a gel at body temperature, it also presents a uniform and transparent state, has high gel strength, does not have the problem of whitening and fragmentation, and can be well matched with the physiological environment of the human body.

[0008] The trimethylamino hydroxypropyl hydroxybutyl chitosan derivative of the present invention has the following structure:

[0009]

[0010] Wherein, R1, R2, R3, and R4 are one of the following groups:

[0011] H, hydroxybutyl, trimethylaminohydroxypropyl chloride, acetyl.

[0012] Furthermore, R1 and R2 are H or hydroxybutyl; and R3 and R4 are H, acetyl, trimethylaminohydroxypropyl chloride or hydroxybutyl.

[0013] The structural formula of the group is as follows:

[0014] Hydroxybutyl: CH3CH2CHOHCH2-; Acetyl:

[0015] Trimethylamino Hydroxypropyl Chloride:

[0016] The substitution degree of the hydroxybutyl group is 1.0-4.0; the substitution degree of the trimethylaminohydroxypropyl chloride group is 0.1-2.0; and the substitution degree of the acetyl group is 0.001-0.3.

[0017] Preferably, the degree of substitution of hydroxybutyl groups is 1.5-3.0; the degree of substitution of trimethylaminohydroxypropyl chloride groups is 0.2-1.5; and the degree of substitution of acetyl groups is 0.001-0.1.

[0018] Above-mentioned hydroxybutyl chitosan derivative forms solution with pure water, or with polyol compatibility such as glycerol, mannitol, even with the osmotic pressure regulating substance compatibility of strong polar ionic compound (such as sodium chloride, potassium chloride, calcium chloride, magnesium chloride, potassium nitrate, sodium sulfate, phosphate buffer etc.), all can obtain the temperature-sensitive solution with good performance. That is, it is a solution that is easy to flow at low temperature, and it is a gel that does not flow at body temperature. And the solution at low temperature is homogeneous and transparent, and the gel at body temperature is also homogeneous, and transparency is very high, and gel strength is relatively large, does not exist the problem of bleaching and scattered, and can have good matching with human physiological environment. This material can also have good compatibility with cell culture fluid, has the effect as cell carrier, and is the cell carrier with temperature sensitivity that effect is more excellent.

[0019] Furthermore, the present invention also provides a hydrogel comprising the above-mentioned trimethylamino hydroxypropyl hydroxybutyl chitosan derivative, comprising the following components by mass: trimethylamino hydroxypropyl hydroxybutyl chitosan derivative: 0.1%-10%; osmotic pressure regulator: 0.1-10%; water: the balance.

[0020] The osmotic pressure regulator is a polyol such as glycerol and mannitol, or a strongly polar ionic compound such as sodium chloride, potassium chloride, calcium chloride, magnesium chloride, potassium nitrate, sodium sulfate, and phosphate buffer.

[0021] Another object of the present invention is to provide a method for preparing the chitosan derivative. The chitosan derivative can be prepared by any of the following methods.

[0022] Method 1: First, chitosan is modified with trimethylaminohydroxypropyl chloride to obtain trimethylaminohydroxypropyl chloride chitosan, and then hydroxybutylated to obtain hydroxybutyl chitosan derivatives. The detailed steps are as follows:

[0023] (1) dissolving or dispersing chitosan with a solvent, adding a trimethylaminohydroxypropyl chloride group modification reagent, then adding an alkaline solution or water, reacting at a certain temperature for a certain time, and performing preliminary purification to obtain trimethylaminohydroxypropyl chloride chitosan; (2) dissolving or dispersing trimethylaminohydroxypropyl chloride chitosan with a solvent, adding 1,2-butylene oxide and an alkaline solution, and reacting at a certain temperature for a certain time; (3) after the reaction is completed, stabilizing the reaction system at room temperature, placing it in a dialysis bag for dialysis for a certain time, taking it out and freeze-drying it to obtain the final trimethylaminohydroxypropyl chloride hydroxybutyl chitosan derivative.

[0024] Scheme 2: Modify chitosan by hydroxybutylation to obtain hydroxybutyl-substituted chitosan; modify the hydroxybutyl chitosan with trimethylaminohydroxypropyl chloride to obtain the final chitosan derivative, as shown in the following details:

[0025] (1) chitosan is modified by hydroxybutylation according to the method disclosed in patent CN 102276756 A to obtain hydroxybutyl chitosan; (2) hydroxybutyl chitosan is dissolved or dispersed in a solvent, a trimethylaminohydroxypropyl chloride group modification reagent is added, an alkaline solution or water is added, and the reaction is carried out at a certain temperature for a certain time; (3) after the reaction is completed, the reaction system is stabilized at room temperature, placed in a dialysis bag for dialysis for a certain time, taken out and freeze-dried to obtain the final trimethylaminohydroxypropyl chloride hydroxybutyl chitosan derivative.

[0026] Scheme 3: Modify chitosan with hydroxybutyl and trimethylaminohydroxypropyl chloride groups simultaneously to obtain the final chitosan derivative, as shown in the following details:

[0027] (1) dissolving or dispersing chitosan with a solvent, adding 1,2-butylene oxide and trimethylaminohydroxypropyl chloride group modification reagent respectively, adding an alkaline solution, and reacting at a certain temperature for a certain time; (2) after the reaction is completed, stabilizing the reaction system at room temperature, placing it in a dialysis bag for dialysis for a certain time, taking it out and freeze-drying it to obtain the final trimethylaminohydroxypropyl chloride hydroxybutyl chitosan derivative.

[0028] The solvent for dispersing chitosan or hydroxybutyl chitosan is a combination of one or more of water, sodium hydroxide solution, potassium hydroxide solution, urea solution, guanidine hydrochloride solution, hydrochloric acid, nitric acid, methanol, formic acid, ethanol, acetic acid, acetonitrile, n-propanol, isopropanol, n-butanol, acetone, dimethyl sulfoxide, N-methylpyrrolidone, NN-dimethylformamide, N,N-dimethylacetamide, and dioxane.

[0029] The reagents used for modifying the trimethylaminohydroxypropyl chloride group include, but are not limited to: 3-chloro-2-hydroxypropyltrimethylammonium chloride and 2,3-epoxypropyltrimethylammonium chloride.

[0030] The alkali is sodium hydroxide or potassium hydroxide.

[0031] The molar concentration of the alkaline solution is 0.01-15 mol / L.

[0032] The reaction temperature is 0°C-100°C.

[0033] The reaction time is 1h-120h.

[0034] The advantages of the present invention are:

[0035] 1. The chitosan derivative of the present invention can form a solution with pure water, or be compatible with polyols such as glycerol and mannitol, or even with osmotic pressure regulating substances of strongly polar ionic compounds (such as sodium chloride, potassium chloride, calcium chloride, magnesium chloride, potassium nitrate, sodium sulfate, etc.), and can obtain a temperature-sensitive solution with good performance. That is, it is a solution that is easy to flow at low temperatures and a gel that does not flow at body temperature. Moreover, the solution at low temperatures is uniform and transparent, and the gel at body temperature is also uniform, with high transparency, high gel strength, no whitening and fragmentation problems, and can be well matched with the physiological environment of the human body.

[0036] 2. The chitosan derivative of the present invention is a positively charged high molecular polymer, in which the amino group and trimethylamino hydroxypropyl chloride group are both positively charged groups, and is a non-amphoteric polyelectrolyte, and does not have the self-complexation phenomenon caused by the attraction of positive and negative charges.

[0037] 3. The chitosan derivative of the present invention can also have good compatibility with cell culture fluid, and is a cell carrier and tissue engineering scaffold material with better effect and temperature sensitivity.

[0038] 4. The chitosan derivative of the present invention can be mixed with drugs in various dosage forms, including liquid preparations, powder preparations, etc., and after mixing, it is still a gel under physiological conditions, there is no precipitation problem, and the effect of slow release can be achieved.

[0039] 5. The chitosan derivative of the present invention can meet the requirements of further filtration sterilization or wet heat sterilization after preparing the solution, and will not cause the material to precipitate due to temperature fluctuations, resulting in the inability to filter or agglomeration after wet heat sterilization.

[0040] 6. The chitosan derivatives of the present invention can be used for superficial body surface, deep body surface tissue or implanted in the body. They can be used for chronic wound care, packing and hemostasis of deep wounds, tissue adhesion prevention and hemostasis after implantation or placement in the body. BRIEF DESCRIPTION OF THE DRAWINGS

[0041] Figure 1 This is the infrared spectrum of the sample of Example 3 of the present invention.

[0042] Figure 2 This is the infrared spectrum of the sample of Example 7 of the present invention.

[0043] Figure 3 This is the liquid nuclear magnetic resonance hydrogen spectrum of the sample in Example 3 of the present invention.

[0044] Figure 4 This is the liquid nuclear magnetic resonance hydrogen spectrum of the sample in Example 7 of the present invention.

[0045] Figure 5 This is the acid solution titration curve of the sample in Example 3 of the present invention.

[0046] Figure 6 The following are photos of the samples of Example 3 (label 17) and Example 7 (label 18) of the present invention, which were respectively dissolved in 0.85% sodium chloride solution at low temperature to prepare a 3% concentration solution at a temperature between 2°C and 8°C.

[0047] Figure 7 The following are photos of the samples of Example 3 (label 17) and Example 7 (label 18) of the present invention after being dissolved in 0.85% sodium chloride solution at low temperature to prepare a 3% concentration solution and then heated to 37°C.

[0048] Figure 8 This is a photograph of the state of the hydroxybutyl chitosan sample after it was dissolved in 0.85% sodium chloride solution at low temperature to prepare a 3% solution and then heated to 37°C.

[0049] Fig. 9 This is the gel temperature test curve of sample ② in Example 10.

[0050] Fig.10 This is the gel temperature test curve of sample ④ in Example 10. DETAILED DESCRIPTION

[0051] The present invention will be further described in detail below with reference to the accompanying drawings and through specific embodiments.

[0052] Embodiment 1:

[0053] (1) Disperse 20 g of chitosan in 200 mL of 20% sodium hydroxide solution, stir for 2 h to make it evenly dispersed, add 20 g of 3-chloro-2-hydroxypropyltrimethylammonium chloride and 400 mL of purified water, react at 50° C. for 24 h, adjust the pH value of the reacted material to neutral with hydrochloric acid, dialyze and freeze-dry to obtain 24.3 g of hydroxypropyltrimethylammonium chloride chitosan.

[0054] (2) Disperse 20 g of hydroxypropyltrimethylammonium chloride chitosan in 100 mL of 50% sodium hydroxide solution, stir for 2 h to make it evenly dispersed, then add 100 mL of isopropanol and 200 mL of 1,2-butylene oxide, and react at 30 °C for 48 h.

[0055] (3) The pH value of the reacted material was adjusted to neutral with hydrochloric acid, and 28.5 g of trimethylaminohydroxypropylhydroxybutyl chitosan chloride was obtained through dialysis and freeze-drying.

[0056] Embodiment 2:

[0057] (1) Hydroxybutyl chitosan was prepared according to the method disclosed in patent application CN 102276756 A.

[0058] (2) Take 40 g of hydroxybutyl chitosan and dissolve it in 2000 mL of 0.1 mol / L acetic acid solution. After it is evenly dissolved, add 200 mL of 40% sodium hydroxide solution and then add 40 g of 2,3-epoxypropyltrimethylammonium chloride. React at 50° C. for 24 h.

[0059] (3) The pH value of the reacted material was adjusted to neutral with a dilute hydrochloric acid solution, and 48.3 g of trimethylaminohydroxypropylhydroxybutyl chitosan chloride was obtained through dialysis and freeze-drying.

[0060] Embodiment 3:

[0061] (1) Disperse 100 g of chitosan in 800 mL of 50% sodium hydroxide solution and stir for 2 h to make it uniformly dispersed. First, add 1000 mL of 1,2-butylene oxide and stir to mix evenly. Then, add 200 g of 2,3-epoxypropyltrimethylammonium chloride, 1000 mL of purified water and 1000 mL of isopropanol and react at 25 °C for 120 h.

[0062] (2) The pH value of the reacted material was adjusted to neutral with hydrochloric acid, and 24.4 g of trimethylaminohydroxypropylhydroxybutyl chitosan chloride was obtained through dialysis and freeze-drying.

[0063] Embodiment 4:

[0064] (1) Disperse 50 g of chitosan in 1000 mL of an aqueous solution containing 11% potassium hydroxide and 8% urea, stir to make the dispersion uniform, freeze overnight and then thaw at low temperature to obtain a uniform solution, add 50 g of 3-chloro-2-hydroxypropyl-trimethylammonium chloride and 500 mL of 1,2-butylene oxide, and react at 25°C for 48 h.

[0065] (2) The pH value of the reacted material was adjusted to neutral with hydrochloric acid, and 72.1 g of trimethylaminohydroxypropylhydroxybutyl chitosan chloride was obtained through dialysis and freeze-drying.

[0066] Embodiment 5:

[0067] (1) Hydroxybutyl chitosan was prepared according to the method disclosed in patent application CN 102276756 A.

[0068] (2) Take 10 g of hydroxybutyl chitosan and dissolve it in 100 mL of dimethyl sulfoxide. After it is evenly dissolved, add 10 g of sodium hydroxide and then add 10 g of 2,3-epoxypropyltrimethylammonium chloride, and react at 60° C. for 24 h.

[0069] (3) The pH value of the reacted material was adjusted to neutral with a dilute hydrochloric acid solution, and 12.6 g of trimethylaminohydroxypropylhydroxybutyl chitosan chloride was obtained through dialysis and freeze-drying.

[0070] Embodiment 6:

[0071] (1) Hydroxybutyl chitosan was prepared according to the method disclosed in patent application CN 102276756 A.

[0072] (2) Take 10 g of hydroxybutyl chitosan and dissolve it in 100 mL of N,N-dimethylformamide. After it is evenly dissolved, add 10 g of sodium hydroxide and then add 10 g of 2,3-epoxypropyltrimethylammonium chloride, and react at 40°C for 48 h.

[0073] (3) The pH value of the reacted material was adjusted to neutral with a dilute hydrochloric acid solution, and 13.5 g of trimethylaminohydroxypropylhydroxybutyl chitosan chloride was obtained through dialysis and freeze-drying.

[0074] Embodiment 7:

[0075] According to the method of Example 1, by increasing the amount of 3-chloro-2-hydroxypropyltrimethylammonium chloride added to 40 g and extending the reaction time to 48 h, 29.2 g of product was finally obtained.

[0076] Example 8: Product structure identification and functional testing

[0077] The samples prepared in Example 3 and Example 7 were subjected to infrared spectroscopy tests respectively. Figure 1 and Figure 2 The peak positions of the two samples are exactly the same, but the intensities of some peaks are different. The peak positions of the two samples are as follows: 1061cm -1 The absorption peaks on the left and right are the stretching vibration absorption peaks of the ether bond (C—O) generated by the primary alcohol, indicating that the hydroxybutylation or trimethylamino hydroxypropylation reaction mainly occurs at the C6 position. -1 Around, 2926cm -1 Around, 2875cm -1 The three peaks on the left and right and 1462cm -1 The absorption peaks on the left and right are the stretching vibration and bending vibration absorption peaks of the C-H bond, indicating the presence of hydroxybutyl. -1 The absorption peaks around 1480 cm-1 are broader than those of hydroxybutyl chitosan alone, which is the original absorption peak of trimethylaminohydroxypropyl chloride. -1The absorption peak of hydroxybutyl chitosan is about 1460 cm -1 The effect of superposition of the absorption peaks on the left and right indicates the presence of trimethylaminohydroxypropyl chloride. -1 The strong and broad absorption peaks on the left and right are the stretching vibration absorption peaks of O—H and N—H.

[0078] The samples prepared in Example 3 and Example 7 were dissolved in a D2O solution containing 20% ​​DCl and subjected to H NMR spectrum testing. The spectra were as shown in FIG. Figure 3 and Figure 4 . Due to the acidic solvent used, the chemical shift of the peak will be offset, but the relative position and peak type of the peak are basically the same. Taking the liquid nuclear magnetic hydrogen spectrum of the sample in Example 7 as an example for illustration, -1.02ppm is the absorption peak of the methyl hydrogen atom on the hydroxybutyl group, and -0.45ppm is the absorption peak of the methylene hydrogen atom on the hydroxybutyl group. The presence of the two indicates the presence of a hydroxybutyl group in the sample. 1.34ppm is the absorption peak of the hydrogen atom on the methyl group directly connected to the nitrogen atom on the trimethylamino hydroxypropyl chloride group, indicating the presence of a trimethylamino hydroxypropyl chloride group in the sample. Figure 4 and Figure 3 Compared with the above, the substitution degree of trimethylamino hydroxypropyl chloride group in the sample increases, and the peak at 1.34ppm increases accordingly. The multiple superimposed absorption peaks between 1ppm-2.5ppm (except the peak at 1.34ppm) are the superposition of the absorption peaks of the hydrogen atoms on the carbons at positions 3, 4, 5, and 6 on the chitosan sugar ring and the methylene and methine carbon atoms directly connected to the nitrogen atom on the trimethylamino hydroxypropyl chloride group. The absorption peak at 3.19ppm is the absorption peak of the hydrogen atom connected to the carbon at position 1 on the chitosan sugar ring.

[0079] Example 9: Acid-base titration test

[0080] The trimethylaminohydroxypropylhydroxybutyl chitosan material prepared in Example 3 was dissolved in excess hydrochloric acid and titrated with a standard sodium hydroxide solution using a fully automatic potentiometric titrator. The titration curve was as follows: Figure 5 As shown. The solid line is the relationship curve between pH value and the volume consumed by the standard sodium hydroxide solution, and the dotted line is the relationship curve between pH value and volume derivative and the volume consumed by the standard sodium hydroxide solution. It can be seen from the spectrum that there are two jump points in the curve, the first jump is relatively gentle, and the second jump is relatively steep. They are the end point of hydrochloric acid titration and the end point of amino titration, respectively. It can be calculated that the degree of substitution of trimethylaminohydroxypropyl chloride is 1.08.

[0081] Example 10: Solution property test

[0082] The trimethylamino hydroxypropyl hydroxybutyl chitosan material in Example 3 was dissolved in purified water and 0.85% sodium chloride solution, respectively, with a concentration of 3%, and was recorded as sample ① and sample ②. The trimethylamino hydroxypropyl hydroxybutyl chitosan material in Example 7 was dissolved in purified water and 0.85% sodium chloride solution, respectively, with a concentration of 3%, and was recorded as sample ③ and sample ④. And the hydroxybutyl chitosan prepared by the method disclosed in patent application CN 102276756 A was dissolved in purified water and 0.85% sodium chloride solution as a control, with a concentration of 3%, and was recorded as sample ⑤ and sample ⑥.

[0083]

[0084] The above data show that, depending on the sample preparation process, a thermosensitive gel that can solidify at 37°C can be obtained, and a thermosensitive solution that is fluid at 37°C can also be obtained, and the gel temperature is higher than 37°C.

[0085] Example 11: Preparation and performance testing of hydrogel

[0086] The sample ② in Example 10 was tested for osmotic pressure of 295 mOsmol / kg by an osmometer, and for gel temperature of 16.4°C by a rheometer. The intersection of the storage modulus (G') and the loss modulus (G") is the gel temperature. Fig. 9 The osmotic pressure of sample ④ in Example 10 was tested by an osmotic pressure meter and the osmotic pressure was 305 mOsmol / kg. The gel temperature was tested by a rheometer and the gel temperature was 37.2°C. Fig.10 As shown in the figure, it can be seen that the gel temperature of the above sample ② is 16.4℃, which is suitable for dressings, fillings or scaffold materials for use in the body and on the surface of the body, and can be used as a drug or cell carrier to load relevant therapeutic drugs or therapeutic cells to further enhance the therapeutic effect. The gel temperature of sample ④ is 37.2℃, which is higher than normal body temperature. It is a flowing liquid at body temperature and can be used as a wound site on the surface or in the body where flowing liquid is required, such as intra-articular injection, liquid dressing application on the surface of the body, etc.

[0087] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention rather than to limit it. Ordinary technicians in the relevant field can still modify or replace the specific implementation methods of the present invention with equivalents by referring to the above embodiments. Any modifications or equivalent replacements that do not depart from the spirit and scope of the present invention are within the scope of protection of the claims of the present invention.

Claims

1. A trimethylamino hydroxypropyl hydroxybutyl chitosan derivative, characterized in that: It has the following structure: Wherein, R1, R2, R3, and R4 are one of the following groups: H, hydroxybutyl, trimethylaminohydroxypropyl chloride, acetyl.

2. The chitosan derivative according to claim 1, characterized in that: The R1 and R2 are H or hydroxybutyl; the R3 and R4 are H, acetyl, trimethylaminohydroxypropyl chloride or hydroxybutyl.

3. The chitosan derivative according to claim 1, characterized in that: The substitution degree of the hydroxybutyl group is 1.0-4.0; the substitution degree of the trimethylaminohydroxypropyl chloride group is 0.1-2.

0.

4. A hydrogel comprising the trimethylaminohydroxypropylhydroxybutyl chitosan derivative according to claim 1, characterized in that: The invention comprises the following components by mass: 0.1%-10% of trimethylaminohydroxypropyl chitosan derivative; 0.1%-10% of osmotic pressure regulator; and water as the balance. The osmotic pressure regulator is glycerol, mannitol or sodium chloride, potassium chloride, calcium chloride, magnesium chloride, potassium nitrate, sodium sulfate, and phosphate buffer.

5. A method for preparing the trimethylamino hydroxypropyl hydroxybutyl chitosan derivative according to claim 1, characterized in that: The following steps are involved: (1) dissolving or dispersing chitosan with a solvent, adding a trimethylaminohydroxypropyl chloride group modification reagent, then adding an alkaline solution or water, reacting at a certain temperature for a certain time, and performing preliminary purification to obtain trimethylaminohydroxypropyl chloride chitosan; (2) dissolving or dispersing trimethylaminohydroxypropyl chloride chitosan with a solvent, adding 1,2-butylene oxide and an alkaline solution, and reacting at a certain temperature for a certain time; (3) after the reaction is completed, stabilizing the reaction system at room temperature, placing it in a dialysis bag for dialysis for a certain time, taking it out and freeze-drying it to obtain the final trimethylaminohydroxypropyl chloride hydroxybutyl chitosan derivative.

6. A method for preparing the trimethylamino hydroxypropyl hydroxybutyl chitosan derivative according to claim 1, characterized in that: The following steps are involved: (1) chitosan is modified by hydroxybutylation according to the method disclosed in patent CN 102276756A to obtain hydroxybutyl chitosan; (2) hydroxybutyl chitosan is dissolved or dispersed in a solvent, a trimethylaminohydroxypropyl chloride group modification reagent is added, an alkaline solution or water is added, and the reaction is carried out at a certain temperature for a certain time; (3) after the reaction is completed, the reaction system is stabilized at room temperature, placed in a dialysis bag for dialysis for a certain time, taken out and freeze-dried to obtain the final trimethylaminohydroxypropyl chloride hydroxybutyl chitosan derivative.

7. A method for preparing the trimethylaminohydroxypropylhydroxybutyl chitosan derivative according to claim 1, characterized in that: The following steps are involved: (1) dissolving or dispersing chitosan with a solvent, adding 1,2-butylene oxide and trimethylaminohydroxypropyl chloride group modification reagent respectively, adding an alkaline solution, and reacting at a certain temperature for a certain time; (2) after the reaction is completed, stabilizing the reaction system at room temperature, placing it in a dialysis bag for dialysis for a certain time, taking it out and freeze-drying it to obtain the final trimethylaminohydroxypropyl chloride hydroxybutyl chitosan derivative.

8. The method for preparing the trimethylamino hydroxypropyl hydroxybutyl chitosan derivative according to any one of claims 5 to 7, characterized in that The solvent for dissolving or dispersing chitosan or hydroxybutyl chitosan is a combination of one or more of water, sodium hydroxide solution, potassium hydroxide solution, urea solution, guanidine hydrochloride solution, hydrochloric acid, nitric acid, methanol, formic acid, ethanol, acetic acid, acetonitrile, n-propanol, isopropanol, n-butanol, acetone, dimethyl sulfoxide, N-methylpyrrolidone, N,N-dimethylformamide, N,N-dimethylacetamide, and dioxane.

9. The method for preparing the trimethylamino hydroxypropyl hydroxybutyl chitosan derivative according to any one of claims 5 to 7, characterized in that The trimethylaminohydroxypropyl chloride group modification agent is 3-chloro-2-hydroxypropyltrimethylammonium chloride or 2,3-epoxypropyltrimethylammonium chloride.

10. The method for preparing the trimethylamino hydroxypropyl hydroxybutyl chitosan derivative according to any one of claims 5 to 7, characterized in that The alkali is sodium hydroxide or potassium hydroxide; the molar concentration of the alkali solution is 0.01-15 mol / L; the reaction temperature is 0° C.-100° C.; and the reaction time is 1 h-120 h.

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