Novel compound as well as preparation method and application thereof

Abstract

The invention discloses a novel compound which can be used for artificial skin pore forming, enables the pore diameter of the artificial skin to be controllable, meets the micropore structure requirement of skin tissue fibroblasts, is simple in process, short in production cycle, low in cost and free of adverse effects such as excessive heavy metal, improves the functionality and clinical application potential of the artificial skin, and has broad application prospects. The defects existing in an existing artificial skin chambering technology are overcome.

Description

Technical Field

[0001] The present invention relates to the technical field of biomedical materials, and particularly relates to a new compound, a preparation method thereof, and an application thereof. Background Art

[0002] As a core material for wound repair and regenerative medicine, the porous structure of artificial skin is crucial for cell migration, nutrient transport, and vascularization. Currently, modulating the pore structure of materials (such as pore size, porosity, and connectivity) to mimic the functions of natural skin is a research hotspot in the field of tissue engineering. Since artificial skin is generally made of nanomaterials and its pore size is nanoscale pores, while the diameter of human skin fibroblasts is generally 75 - 200 μm, in order for human skin fibroblasts to drill into artificial skin for reproduction and thus create new skin, it is often necessary to expand the pores of artificial skin.

[0003] The existing artificial skin pore expansion technologies mainly include the following several types:

[0004] 1. Inorganic nanoparticle hybridization technology

[0005] Technical principle: By introducing inorganic precursors (such as TiO2 or SiO2) during the fermentation process of bacterial cellulose, and utilizing the biomineralization of microorganisms, inorganic nanoparticles are attached to the surface of cellulose fibers or embedded in the membrane structure. The addition of these inorganic particles can change the arrangement of fibers, forming a microporous structure with a pore size of 0.2 - 2 μm, but still cannot meet the microporous structure requirements of skin tissue fibroblasts.

[0006] 2. Electrospinning technology and fiber composite

[0007] Technical principle: Using electrospinning technology to composite bacterial cellulose with other polymers (such as EVOH or polypropylene), and by adjusting the fiber diameter (10 - 100 nm) and packing density, a hierarchical pore structure is formed. For example, the fiber packing pore formation mechanism of an EVOH nanofiber membrane can achieve uniform pores of 0.22 - 0.45 μm. This technology also cannot meet the microporous structure requirements of skin tissue fibroblasts, and the production cost and energy consumption are relatively high.

[0008] 3. Combination of dynamic fermentation and mechanical processing

[0009] Technical principle: Using a dynamic fermentation device (such as a rotary drum fermentation equipment), the three-dimensional network structure of fibers is changed through mechanical stirring or shear force, thereby expanding the pore size. Compared with static shallow pan fermentation, dynamic fermentation can shorten the production cycle and optimize the pore distribution, but its pore size cannot be controlled.

[0010] 4. Composite of template method and biodegradable materials

[0011] Technical principle: During the preparation of BC membrane, a degradable template (such as polylactic acid microspheres) is introduced, and micron-sized pores are formed through the subsequent degradation of the template. This method requires precise control of the template size and degradation conditions to match the target pore size, and the long degradation period results in an extended production cycle and excessively high production costs.

[0012] 5. Chemical modification and surface functionalization

[0013] Technical principle: By chemical modification (such as introducing active groups like amino groups and quaternary ammonium salts) or doping functional substances (such as silver ions), the surface properties of BC fibers are changed to promote crosslinking or depolymerization between fibers, thereby regulating the pore size. For example, silver ion doping can enhance antibacterial properties while forming larger pores through local depolymerization of the fiber structure. Disadvantage: Chemical modification is not easily fully reacted, and silver is a heavy metal, which is likely to cause heavy metal over-standard.

[0014] Therefore, there is an urgent need to provide a new technology to solve the problem of pore expansion of artificial skin. Summary of the invention

[0015] In response to the above problems, the present invention has developed a new compound that can be used for pore formation of artificial skin, enabling the pore size of artificial skin to be controllable, meeting the microporous structure requirements of skin tissue fibroblasts, with a simple process, short production cycle, low cost, and no adverse effects such as other heavy metal over-standard.

[0016] The new compound provided by the present invention has the following chemical structural formula:

[0017]

[0018] Among them, n = 2 - 8.

[0019] In some embodiments, the new compound includes at least one of the following compounds:

[0020]

[0021]

[0022] In some embodiments, the synthesis method of the new compound is to generate through the condensation reaction of the compound of formula A as follows:

[0023]

[0024] In some embodiments, the raw materials for the condensation reaction further include: amine compounds and polypeptide condensation reagents. The amine compounds include but are not limited to: at least one of triethylamine, ethylenediamine, 1,3-propanediamine, 1,4-butanediamine, 1,5-pentanediamine, 1,6-hexanediamine, 1,7-heptanediamine, 1,8-octanediamine.

[0025] In some embodiments, the compound of formula A is formed by the condensation hydrolysis reaction of the compounds of formula B and formula C as follows:

[0026]

[0027] The present invention also provides the application of the new compound in the preparation of biomedical nanomaterials.

[0028] In some embodiments, the biomedical nanomaterials include artificial skin.

[0029] The new compound of the present invention can be added during the synthesis of the biomedical nanomaterials, and then the new compound in the synthesis product can be removed by simple methods such as organic solvents (such as methanol, ethanol, acetone, etc.). By controlling the particle size of the new compound, the pore size of the biomedical nanomaterials can be regulated, thereby obtaining biomedical nanomaterials with a controllable pore size.

[0030] The new compound of the present invention is particularly suitable for creating pores in artificial skin, enabling the artificial skin of the nanomaterials to have more irregular micron-sized pores that can accommodate human skin fibroblasts, meeting the microporous structure requirements of human skin tissue fibroblasts, enhancing the functionality and clinical application potential of artificial skin, and solving the defects existing in the existing artificial skin pore expansion technology.

[0031] The present invention also provides a biomedical nanomaterial, which is perforated by the new compound provided by the present invention.

[0032] In some embodiments, the biomedical nanomaterials include artificial skin, such as microbial biofilms such as bacterial cellulose and fungal mycelia.

[0033] In some embodiments, the biomedical nanomaterials have micron-sized pores with a pore size of 75 - 200 μm. In clinical applications, these micron-sized pores can allow fibroblasts of the human skin to penetrate and multiply to create new skin.

[0034] The present invention also provides a method for preparing a biomedical nanomaterial, which includes the following steps:

[0035] 1) Put the new compound of the present invention into the fermentation raw materials of the biofilm and ferment to obtain the biofilm;

[0036] 2) Purify and remove impurities from the biofilm obtained in step 1) to obtain the biomedical nanomaterial.

[0037] In some embodiments, the fermentation raw materials of the biofilm include Acetobacter xylinum, sugar, and coconut water.

[0038] In some embodiments, the new compound is granular with a particle size of 75 - 200 μm.

[0039] The biomedical nanomaterial prepared by the above preparation method of the present invention is a nanofiber network membrane and can be used as artificial skin. During the preparation process, by controlling the particle size of the new compound, the pore size of the biomedical nanomaterial can be controlled, realizing the controllable pore size of the biomedical nanomaterial. Moreover, the new compound is easily removed during the later purification process of the biofilm.

[0040] In some embodiments, the particle size of the new compound can be obtained by conventional physical crushing, such as mechanical pulverization, etc., to obtain particles with the target particle size.

[0041] When the new compound input during the preparation process is micron-sized particles of 75 - 200 μm, the prepared biomedical nanomaterial can also have micron pores of 75 - 200 μm, thus having a microporous structure that allows human skin fibroblasts to penetrate and reproduce. When used in clinical treatments such as burn and chronic ulcer treatments, it can better promote skin regeneration after covering the wound surface.

[0042] Secondly, when the new compound is prepared into smaller particles and added to the fermentation of the biofilm, the bacterial corpses present in the nanostructure of the biofilm can be removed together when the new compound is removed by purification later, which helps to reduce the immunogenicity of the biofilm.

[0043] In addition, there is a certain chance of condensation reaction between the new compound of the present invention and the hydroxyl groups abundantly present in the biofilm structure. Therefore, more treatment methods can be derived in terms of pore formation, purification, and later degradation reaction treatment. Detailed implementation manners

[0045] The experimental methods used in the following examples are all conventional methods unless otherwise specified.

[0046] The materials, reagents, etc. used in the following examples can be obtained from commercial channels unless otherwise specified.

[0047] Example 1

[0048] Provide a synthesis method of a compound of formula I, the steps are as follows:

[0049] S1. Synthesize compound A by condensation hydrolysis reaction

[0050] Synthesis process: Add 1 equivalent of Compound B (3,8-dimethoxyformamido-6-phenylphenanthridine) into a reaction kettle, add 1.5 equivalents of Compound C (ethyl iodohexanoate), stir, and slowly heat to 130 °C overnight. Monitor by TLC. After the reaction is complete, cool the reaction solution to room temperature, add 2.0 equivalents of hydrobromic acid, stir, and slowly heat to 130 °C. Monitor by TLC until the reaction is complete. Finally, cool and filter to obtain the solid target product.

[0051]

[0052] S2. Synthesis of a new compound (Formula I) by condensation reaction

[0053] Synthesis process: Add 1 equivalent of the Formula A compound synthesized in Step S1 into a reaction kettle, add DMAC, stir to dissolve, then add 8.5 equivalents of triethylamine, 2.2 equivalents of HATU (polypeptide condensation reagent), and 0.5 equivalent of ethylenediamine. Stir and slowly heat to 50 °C for 2 h. Monitor the reaction process in real time using TLC. After the reaction is completed, cool the reaction solution to room temperature, add 5 - 10 times the amount of MTBE solvent. After the solid precipitates, filter to obtain the solid. Purify the obtained solid: Heat with ethanol for 2 - 4 h to completely dissolve it, then naturally cool to room temperature. Crystals will precipitate, filter to obtain the high-purity solid target product of Formula I.

[0054]

[0055] Characterization of Compound I:

[0056] Dark red solid, [α] 20 D +22.5 (c 0.2, CHCl3); From the electrospray ionization mass spectrometry signal m / z: 824.45 [M + H] + , the calculated molecular formula is C 52 H 56 O2N8; Infrared spectrum shows absorption peaks at 2542 cm -1 、1676 cm -1 、1565 cm -1 、indicating that the compound contains amide group, aniline group, and phenyl group. 1 1H nuclear magnetic resonance spectrum shows five aromatic hydrogen proton signals on the ring [δ 7.67 (1H, d, J = 7.54 Hz), 7.13 (1H, m), 7.11 (1H, m), 7.12 (1H, d, J = 7.6 Hz), 6.55 (1H, s)] and two characteristic hydrogen proton signals on the ring [δ 4.63 (1H, dd, J = 9.6 Hz) and 4.42 (1H, d, J = 9.6 Hz)], 13 13C nuclear magnetic resonance.13 CNMR: δ 14.9 (1C, s), 25.6 - 25.8 (2C, 25.7 (s), 25.7 (s)), 26.2 (1C, s), 29.3 - 29.4 (4C, 29.4 (s), 29.4 (s), 29.4 (s), 29.4 (s)), 34.4 (1C, s), 34.7 (1C, s), 36.6 (1C, s), 39.0 - 39.0 (2C, 39.0 (s), 39.0 (s)), 114.2 - 114.3 (4C, 114.2 (s), 114.2 (s), 114.2 (s), 114.2 (s)), 116.0 - 116.0 (4C, 116.0 (s), 116.0 (s), 116.0 (s), 116.0 (s)), 125.2 - 125.3 (4C, 125.2 (s), 125.2 (s), 125.2 (s), 125.2 (s)), 127.5 - 127.5 (4C, 127.5 (s), 127.5 (s), 127.5 (s), 127.5 (s)), 127.9 - 128.0 (4C, 127.9 (s), 127.9 (s)), 128.1 - 128.3 (2C, 128.2 (s), 128.2 (s)), 128.8 - 128.9 (4C, 128.8 (s), 128.8 (s)), 130.6 - 130.7 (2C, 130.7 (s), 130.7 (s)), 137.7 - 137.8 (4C, 137.8 (s), 137.8 (s), 137.8 (s), 137.8 (s)), 138.5 - 138.6 (2C, 138.5 (s), 138.5 (s)), 147.1 - 147.2 (4C, 147.1 (s), 147.1 (s), 147.1 (s), 147.1 (s)), 169.9 (1C, s), 172.8 (1C, s).

[0057] The above NMR data indicate that the compound is the product of Formula I.

[0058] Example 2

[0059] A method for synthesizing a compound of Formula II is provided. The difference from Example 1 is only that: in step S2, ethylenediamine is replaced with an equal amount of 1,3 - propanediamine, and the rest is the same as in Example 1, to obtain a high - purity solid target product of Formula II.

[0060]

[0061] Example 3

[0062] Provide a method for synthesizing a compound of formula III, which is only different from Example 1 in that: in step S2, ethylenediamine is replaced with an equal amount of 1,4-butanediamine, and the rest is the same as in Example 1, to obtain a high-purity solid target product of formula III.

[0063]

[0064] Example 4

[0065] Provide a method for synthesizing a compound of formula IV, which is only different from Example 1 in that: in step S2, ethylenediamine is replaced with an equal amount of 1,5-pentanediamine, and the rest is the same as in Example 1, to obtain a high-purity solid target product of formula IV.

[0066]

[0067] Example 5

[0068] Provide a method for synthesizing a compound of formula V, which is only different from Example 1 in that: in step S2, ethylenediamine is replaced with an equal amount of 1,6-hexanediamine, and the rest is the same as in Example 1, to obtain a high-purity solid target product of formula V.

[0069]

[0070] Example 6

[0071] Provide a method for synthesizing a compound of formula VI, which is only different from Example 1 in that: in step S2, ethylenediamine is replaced with an equal amount of 1,7-heptanediamine, and the rest is the same as in Example 1, to obtain a high-purity solid target product of formula VI.

[0072]

[0073] Example 7

[0074] Provide a method for synthesizing a compound of formula VII, which is only different from Example 1 in that: in step S2, ethylenediamine is replaced with an equal amount of 1,8-octanediamine, and the rest is the same as in Example 1, to obtain a high-purity solid target product of formula VII.

[0075]

[0076]

[0077] Use the pore-forming compounds of formula I to formula VII prepared in Examples 1 to 7 for pore formation of artificial skin biological substrates, and the specific steps are as follows:

[0078] 1) 100 g of the pore-forming compound was crushed with a grinder, and the powder obtained by sifting through a 100-mesh sieve was added into 10 L of coconut water, and the mixture was stirred slowly to allow the powder to be fully mixed to obtain a suspension;

[0079] 2) 100uL of Acetobacter xylinum and 5g of white sugar were added to the suspension of step 1), and fermented at 30-35°C for 7 days. At this time, a biofilm of about 3 mm thick was formed on the surface of the fermentation liquid. The biofilm was taken out and put into 10L of 0.1% sodium hydroxide solution, and slowly stirred for 1-2h to remove the bacterial corpses and the like mixed in the biofilm. Then, 2L of ethanol was used to soak the biofilm to remove the pore-forming compounds mixed in the biofilm. Finally, it was washed three times with clean water to obtain an artificial skin biological substrate with a microporous structure.

[0080] Through electron microscopic observation, the artificial skin biological substrates prepared with the pore-forming compounds of Formula I to Formula VII prepared in Examples 1 to 7 all have numerous irregular micropores of 75-200 micrometers.

[0081] Comparative Example 1

[0082] A preparation method of an artificial skin biological substrate comprises the following steps: adding 100uL of Acetobacter xylinum and 5g of white sugar into 10L of coconut water, fermenting at 30-35°C for 7 days, taking out the biofilm and adding it into 10L of 0.1% sodium hydroxide solution, slowly stirring for 1-2h to remove bacterial corpses and the like mixed in the biofilm, and finally washing with clean water for three times to obtain the artificial skin biological substrate.

[0083] Test example

[0084] The artificial skin biosubstrates prepared with the pore-forming compounds of formula I to formula VII prepared in Examples 1 to 7 and the biosubstrates prepared in Comparative Example 1 were subjected to animal experiments. The experimental method was as follows: 8 groups of experimental rabbits were anesthetized and a third-degree burn wound of 5cm*5cm was made at the same site. Among them, the rabbits in the experimental group were covered with the biosubstrates of Examples 1 to 7 after debridement, and the wound was fixed with gauze; the rabbits in the control group were covered with the biosubstrates of Comparative Example 1 after debridement, and the wound was fixed with gauze; during the experiment, the rabbits in each group were fed with the same routine. After 21 days, the skin recovery of the 8 groups of rabbits was observed. The results showed that the rabbits in the experimental group had grown new skin and the wound healed well; while the rabbits in the control group failed to grow new skin and the wound healed poorly.

[0085] This shows that the artificial skin biological substrate prepared with the pore-forming compounds of Formula I to Formula VII prepared in Examples 1 to 7 has a diameter of 75-200 μm, which is suitable for skin fibroblasts to penetrate and multiply. When used in clinical treatments such as burns and chronic ulcers, it can better promote skin regeneration after covering the wound surface.

[0086] 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 the protection scope of the present invention. Although the present invention has been described in detail with reference to the preferred embodiments, those of ordinary skill in the art should understand that the technical solutions of the present invention can be modified or equivalently replaced without departing from the essence and scope of the technical solutions of the present invention.

Claims

1. A new compound, characterized in that, It has the following chemical structural formula: Wherein, n = 2 - 8.

2. The synthesis method of the new compound according to claim 1, characterized in that, It is formed by the condensation reaction of the compound of formula A as follows:

3. The synthesis method according to claim 2, characterized in that, The raw materials for the condensation reaction further include: amine compounds and polypeptide condensation reagents.

4. The synthesis method according to claim 2, wherein The compound of formula A is formed by the condensation hydrolysis reaction of the compounds of formula B and formula C as follows:

5. The application of the new compound according to claim 1 in the preparation of biomedical nanomaterials.

6. The application according to claim 5, characterized in that, The biomedical nanomaterials include artificial skin.

7. A biomedical nanomaterial, characterized in that, The biomedical nanomaterials are pore-formed by the new compound according to claim 1.

8. The biomedical nanomaterial according to claim 7, wherein The biomedical nanomaterials include artificial skin.

9. The biomedical nanomaterial according to claim 7, wherein, The biomedical nanomaterials have micropores with a pore diameter of 75 - 200 μm.

10. A preparation method of a biomedical nanomaterial, characterized in that, It includes the following steps: 1) Put the new compound according to claim 1 into the fermentation raw materials of the biofilm, and ferment to obtain the biofilm; 2) Purify and remove impurities from the biofilm obtained in step 1) to obtain the biomedical nanomaterials.

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