Preparation method of high-purity sponge bone needle and application of high-purity sponge bone needle in permeation promotion

Abstract

The application discloses a preparation method of high-purity sponge bone needles and application of the high-purity sponge bone needles in penetration promotion, and belongs to the technical field of sponge bone needles. The method comprises the following steps: cutting natural sponges, soaking the sponges in deionized water and performing ultrasonic oscillation, filtering, and cleaning to obtain pretreated sponges; subjecting the pretreated sponges to alkaline protease enzymolysis to obtain primary enzymolysis products; subjecting the primary enzymolysis products to ginger protease enzymolysis to obtain secondary enzymolysis products; and performing vacuum freeze-drying to obtain the high-purity sponge bone needles; the natural sponges are subjected to enzymolysis treatment by alkaline protease and ginger protease to obtain the high-purity sponge bone needles; and the sponge bone needles are used in surface grafting of wheat oligopeptides and loading of giant salamander skin collagen, pomegranate seed extract, Honeysuckle extract, Gotu Kola extract, early celery extract, iris extract, oyster shell extract and calcium chloride mixed treatment materials, so that the repair effect on skin inflammation acne is improved.

Description

Technical Field

[0001] This invention relates to the field of sponge bone needle technology, specifically to a method for preparing high-purity sponge bone needles and their application in promoting permeation. Background Technology

[0002] In the field of skin care and beauty, transdermal absorption is a crucial concept, specifically referring to the process by which active ingredients in daily cosmetics act on the surface or deeper layers of the skin through a specific pathway. Unlike transdermal drug absorption, the transdermal absorption of functional cosmetic ingredients focuses more on effective accumulation and sustained action in the target skin layer, rather than systemic absorption. This process requires specialized penetration enhancement technologies, currently categorized into two main types: physical and chemical penetration enhancement. Among these, microneedling technology, as a representative method of physical penetration enhancement, has gained significant industry acclaim in recent years due to its remarkable effects. This technology creates numerous micron-sized channels in the stratum corneum through minimally invasive techniques, providing a direct delivery pathway for active ingredients and drug carriers to reach the deeper layers of the skin.

[0003] Among all permeation-enhancing technologies, microneedling is the most direct and safest. However, with traditional solid microneedles, the microchannels opened by the skin close quickly due to the skin's elasticity, limiting drug penetration time. While novel microneedle patches can remain on the skin for extended periods for continuous drug delivery, their array-like manufacturing process limits the delivery area and site. The minimally invasive nature of sponge-bone needles minimizes skin damage, virtually eliminating the risk of bleeding or infection, making them a safe and efficient new option for transdermal drug delivery.

[0004] Sponge spicules are typically extracted from natural sponges. However, in addition to the spicules we need, natural sponges contain a large number of other impurities, such as various organic matter, microorganisms, and some minerals. Completely removing these impurities during the extraction process is no easy task.

[0005] Acne is a common skin condition that severely impacts a patient's appearance and mental well-being. Most mainstream acne treatments on the market achieve their goal by adding oil-controlling or antibacterial ingredients. However, these ingredients often have significant drawbacks. The skin's absorption capacity is limited, and many oil-controlling or antibacterial ingredients cannot penetrate effectively, leaving residues on the skin's surface that can cause significant irritation. Long-term use can disrupt the normal flora balance of the skin's surface, leading to reduced treatment effectiveness.

[0006] Therefore, it is urgent to develop a new microneedle penetration-enhancing technology to repair skin inflammation and acne problems. Summary of the Invention

[0007] The purpose of this invention is to provide a method for preparing high-purity sponge bone needles and their application in promoting penetration, thereby providing a novel microneedle penetration-promoting technology to repair skin inflammation and acne problems.

[0008] The objective of this invention can be achieved through the following technical solutions:

[0009] A method for preparing high-purity sponge bone needles includes the following steps:

[0010] S1. Take natural sponge, cut it into pieces, soak it in deionized water and ultrasonically vibrate it at a frequency of 35-45Hz for 1-2 hours, filter it, take the sponge fragments and wash them with deionized water 4-5 times to obtain the pretreated sponge.

[0011] S2. Place the pretreated sponge in an extraction tank, add deionized water and alkaline protease, adjust the pH to 10 with sodium hydroxide-sodium hexametaphosphate solution, enzymatically hydrolyze at 40℃ for 4-6 hours, then inactivate the enzyme at 110℃ for 10 minutes, filter, wash the solid phase until neutral, and dry to obtain the first enzymatic hydrolysate; the sodium hydroxide-sodium hexametaphosphate solution is prepared by mixing 4-8g sodium hydroxide, 1.8-2.2g sodium hexametaphosphate, and 100mL deionized water evenly; the sodium hydroxide-sodium hexametaphosphate solution is used to adjust the pH to be suitable for alkaline protease, while simultaneously dissolving the tips of the sponge spicules with the alkali solution to form openings, thereby forming hollow channels. Simultaneously, under the synergistic effect of sodium hexametaphosphate, the OH groups can be effectively increased. - Increase concentration to increase double-layer potential and remove metal ion impurities from sponge spicules;

[0012] S3. Place the primary enzymatic hydrolysate in an extraction tank, add deionized water and ginger protease, add 1 mol / L hydrochloric acid solution to adjust the pH to 4.5, enzymatically hydrolyze at 40℃ for 2-3 hours, then inactivate the enzyme at 110℃ for 10 minutes, filter, take the solid phase and wash until neutral to obtain the secondary enzymatic hydrolysate, freeze-dry at -80℃ under vacuum for 6-10 hours until it becomes a loose powder, thus obtaining the high-purity sponge bone needle.

[0013] As a further aspect of the present invention, in step S2, the mass ratio of the pretreated sponge, deionized water and alkaline protease is 1-1.5:100:0.02-0.04.

[0014] As a further embodiment of the present invention, in step S3, the mass ratio of the primary enzymatic hydrolysate, deionized water and ginger protease is 1-1.2:100:0.0075-0.014.

[0015] High-purity sponge bone needles were prepared using the above-described preparation method.

[0016] The application of the high-purity sponge bone needles to enhance penetration specifically involves preparing a penetration-enhancing composition using sponge bone needles for repairing inflammatory acne skin. The preparation method of the penetration-enhancing composition includes the following steps:

[0017] A1. Take giant salamander skin collagen, pomegranate seed extract, honeysuckle extract, centella asiatica extract, early celery extract, iris extract, oyster shell extract, anhydrous ethanol, deionized water, and calcium chloride, mix them, and ultrasonically stir for 15-25 minutes to obtain material A.

[0018] A2. Take the high-purity sponge bone needles, N,N-dimethylformamide, and deionized water, mix them, and ultrasonically disperse them for 30-40 minutes. Add succinic anhydride and 3-aminopropyltriethoxysilane, heat and stir in a water bath at 50-60°C for 5-7 hours, centrifuge, take the solid phase for washing, freeze dry under vacuum, and obtain material B through surface carboxylation treatment.

[0019] A3. Take the material B, wheat oligopeptide and deionized water, mix them, sonicate for 20-30 minutes, centrifuge, take the solid phase, wash and dry it, and obtain material C by amidation reaction of wheat oligopeptide with carboxyl groups on the surface of sponge bone needles.

[0020] A4. Take the material C and material A, mix them, ultrasonically disperse for 30-40 minutes, maintain pressure under vacuum conditions of gauge pressure -0.1MPa for 30-40 minutes, filter, take the solid phase, and introduce the therapeutic agent containing a variety of skin inflammation and acne into the hollow cavity of the sponge bone needle through vacuum negative pressure treatment to obtain material D;

[0021] A5. Mix aluminum hydroxide, tartaric acid, and deionized water, stir for 20-30 minutes, add sodium polyacrylate, continue stirring for 30-40 minutes, add material D, and continue stirring for 5-10 minutes to obtain the penetration-enhancing composition.

[0022] As a further embodiment of the present invention, the preparation method of giant salamander skin collagen includes the following steps: removing fat and pigment from giant salamander skin, then taking 1g of giant salamander skin, adding 15mL of 0.5mol / L acetic acid, then adding 2.5mg of pepsin for extraction, filtration and centrifugation, taking the supernatant, adding 5mol / L NaCl solution to the supernatant to make the NaCl concentration 0.9mol / L, letting it stand for 24h to separate into layers, centrifuging to recover the flocculent matter, placing it in ultrapure water for dialyzing for three days, changing the water every 6 hours during this period, dialyzing out the acetic acid, and then freeze-drying to obtain giant salamander skin collagen.

[0023] As a further aspect of the present invention, in step A1, the mass ratio of the giant salamander skin collagen, pomegranate seed extract, honeysuckle extract, centella asiatica extract, early celery extract, iris extract, oyster shell extract, anhydrous ethanol, and deionized water is 25-35:8-10:4-6:18-26:4-6:2-4:3-5:40-50:60-80:0.4-0.8.

[0024] As a further embodiment of the present invention, in step A2, the ratio of the high-purity sponge bone needle, N,N-dimethylformamide, deionized water, succinic anhydride and 3-aminopropyltriethoxysilane is 0.6-0.8g: 2-3mL: 30mL: 4-5g: 8-10g.

[0025] As a further embodiment of the present invention, in step A3, the ratio of material B, wheat oligopeptide and deionized water is 0.4-0.8g:10-15mg:10-15mL.

[0026] As a further aspect of the present invention, in step A4, the mass ratio of material C to material A is 1:10.

[0027] As a further embodiment of the present invention, in step A5, the ratio of aluminum hydroxyl, tartaric acid, deionized water, sodium polyacrylate, and material D is 0.3-0.5g: 0.4-0.6g: 50mL: 8-10g: 1.5-2.2g.

[0028] The beneficial effects of this invention are:

[0029] This invention obtains high-purity sponge spicules by enzymatic hydrolysis of natural sponges using alkaline protease and ginger protease.

[0030] Furthermore, regarding the penetration enhancement, utilizing the hollow cavity structure of the sponge bone needle, a mixture of giant salamander skin collagen, pomegranate seed extract, honeysuckle extract, centella asiatica extract, early celery extract, iris extract, oyster shell extract, and calcium chloride is pressed into the sponge bone needle. Simultaneously, wheat oligopeptides are grafted onto the outside of the sponge bone needle to obtain a penetration enhancement composition.

[0031] Wheat oligopeptides can effectively inhibit the overexpression of inflammatory factors and reduce inflammatory responses. At the same time, their small molecule properties can penetrate into the dermis of the skin, directly act on cells at the site of inflammation, regulate immune responses, reduce skin sensitivity, protect cells from oxidative stress damage, and promote collagen synthesis to help repair damaged skin tissue.

[0032] When the sponge needles pierce the skin, they release calcium ions, increasing the calcium ion concentration in skin cells. This activates nitric oxide synthase in macrophages at the site of acne inflammation, which synthesizes low concentrations of nitric oxide from arginine and other amino acids found in giant salamander skin collagen. This repairs inflamed acne and promotes wound healing. Simultaneously, the synergistic effect of pomegranate seed extract, honeysuckle extract, centella asiatica extract, early celery extract, iris extract, and oyster shell extract further repairs the skin.

[0033] Furthermore, the chlorogenic acid in honeysuckle extract, the ellagic acid in pomegranate seed extract, and the flavonoids in iris extract inhibit Propionibacterium acnes from different angles: chlorogenic acid disrupts the bacterial cell membrane, ellagic acid inhibits the activity of bacterial metabolic enzymes, and iris flavonoids interfere with the bacterial reproductive cycle. The three work synergistically to enhance the antibacterial effect and reduce the inflammatory stimulation caused by bacteria. The zinc element in oyster shell extract further assists in inhibiting bacterial growth and regulates the skin's microecological balance. Iris extract and centella asiatica extract work synergistically to regulate keratin metabolism, promote normal keratin shedding, inhibit excessive proliferation of keratinocytes, and prevent follicle blockage. Celery extract reduces sebum secretion and improves the oily environment within the follicles. Through the synergistic effect of each component, the skin inflammation and acne problems are improved and repaired. Attached Figure Description

[0034] The invention will now be further described with reference to the accompanying drawings.

[0035] Figure 1 This is a magnified image of sponge spicules under a scanning electron microscope in Example 2 of the present invention.

[0036] Figure 2 These are Fourier transform infrared (FTIR) spectra of the products obtained in different steps during the preparation process of Example 2 of this invention. Detailed Implementation

[0037] The technical solutions of the present invention will be clearly and completely described below with reference to the accompanying drawings of the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0038] Preparation Example 1

[0039] A method for preparing high-purity sponge bone needles includes the following steps:

[0040] S1. Take natural sponge, cut it into pieces, soak it in deionized water and ultrasonically vibrate it at a frequency of 35Hz for 1 hour, filter it, take the sponge fragments and wash them with deionized water 4 times to obtain the pretreated sponge.

[0041] S2. Place the pretreated sponge in an extraction tank, add deionized water and alkaline protease, adjust the pH to 10 with sodium hydroxide-sodium hexametaphosphate solution, enzymatically hydrolyze at 40℃ for 4 hours, then inactivate the enzyme at 110℃ for 10 minutes, filter, wash the solid phase until neutral, and dry to obtain the first enzymatic hydrolysate; the mass ratio of the pretreated sponge, deionized water and alkaline protease is 1:100:0.02; the sodium hydroxide-sodium hexametaphosphate solution is prepared by mixing 4g sodium hydroxide, 1.8g sodium hexametaphosphate and 100mL deionized water evenly.

[0042] S3. Place the primary enzymatic hydrolysate in an extraction tank, add deionized water and ginger protease, add 1 mol / L hydrochloric acid solution to adjust the pH to 4.5, enzymatically hydrolyze at 40℃ for 2 h, then inactivate the enzyme at 110℃ for 10 min, filter, take the solid phase and wash until neutral to obtain the secondary enzymatic hydrolysate, freeze-dry at -80℃ under vacuum for 6 h until it becomes a loose powder, thus obtaining the high-purity sponge bone needle; the mass ratio of the primary enzymatic hydrolysate, deionized water and ginger protease is 1:100:0.0075.

[0043] Preparation Example 2

[0044] A method for preparing high-purity sponge bone needles includes the following steps:

[0045] S1. Take natural sponge, cut it into pieces, soak it in deionized water and ultrasonically vibrate it at a frequency of 40Hz for 1.5 hours. Filter it, take the sponge fragments and wash them with deionized water 5 times to obtain the pretreated sponge.

[0046] S2. Place the pretreated sponge in an extraction tank, add deionized water and alkaline protease, add sodium hydroxide-sodium hexametaphosphate solution to adjust the pH to 10, enzymatically hydrolyze at 40℃ for 5 hours, then inactivate the enzyme at 110℃ for 10 minutes, filter, take the solid phase, wash until neutral, and dry to obtain the first enzymatic hydrolysate; the mass ratio of the pretreated sponge, deionized water and alkaline protease is 1.25:100:0.03; the sodium hydroxide-sodium hexametaphosphate solution is prepared by mixing 6g sodium hydroxide, 2.0g sodium hexametaphosphate and 100mL deionized water evenly.

[0047] S3. Place the primary enzymatic hydrolysate in an extraction tank, add deionized water and ginger protease, add 1 mol / L hydrochloric acid solution to adjust the pH to 4.5, enzymatically hydrolyze at 40℃ for 2.5 h, then inactivate the enzyme at 110℃ for 10 min, filter, take the solid phase and wash until neutral to obtain the secondary enzymatic hydrolysate, freeze-dry at -80℃ under vacuum for 8 h until it becomes a loose powder, thus obtaining the high-purity sponge bone needle; the mass ratio of the primary enzymatic hydrolysate, deionized water and ginger protease is 1.1:100:0.01.

[0048] As attached Figure 1The image shown is a magnified view of the sponge bone needles prepared in this embodiment under a scanning electron microscope. The surface is smooth and the structure is intact.

[0049] Preparation Example 3

[0050] A method for preparing high-purity sponge bone needles includes the following steps:

[0051] S1. Take natural sponge, cut it into pieces, soak it in deionized water and ultrasonically vibrate it at a frequency of 45Hz for 2 hours, filter it, take the sponge fragments and wash them with deionized water 5 times to obtain the pretreated sponge.

[0052] S2. Place the pretreated sponge in an extraction tank, add deionized water and alkaline protease, add sodium hydroxide-sodium hexametaphosphate solution to adjust the pH to 10, enzymatically hydrolyze at 40℃ for 6 hours, then inactivate the enzyme at 110℃ for 10 minutes, filter, wash the solid phase until neutral, and dry to obtain the first enzymatic hydrolysate; the mass ratio of the pretreated sponge, deionized water and alkaline protease is 1.5:100:0.04; the sodium hydroxide-sodium hexametaphosphate solution is prepared by mixing 8g sodium hydroxide, 2.2g sodium hexametaphosphate and 100mL deionized water evenly.

[0053] S3. Place the primary enzymatic hydrolysate in an extraction tank, add deionized water and ginger protease, add 1 mol / L hydrochloric acid solution to adjust the pH to 4.5, enzymatically hydrolyze at 40℃ for 3 h, then inactivate the enzyme at 110℃ for 10 min, filter, take the solid phase and wash until neutral to obtain the secondary enzymatic hydrolysate, freeze-dry at -80℃ under vacuum for 10 h until it becomes a loose powder, thus obtaining the high-purity sponge bone needle; the mass ratio of the primary enzymatic hydrolysate, deionized water and ginger protease is 1.2:100:0.014.

[0054] Preparation Example 4

[0055] A method for preparing high-purity sponge bone needles includes the following steps:

[0056] S1. Take natural sponge, cut it into pieces, soak it in deionized water and ultrasonically vibrate it at a frequency of 40Hz for 1.5 hours. Filter it, take the sponge fragments and wash them with deionized water 5 times to obtain the pretreated sponge.

[0057] S2. Place the pretreated sponge in an extraction tank, add deionized water and alkaline protease, add sodium hydroxide-sodium hexametaphosphate solution to adjust the pH to 10, enzymatically hydrolyze at 40℃ for 5 hours, then inactivate the enzyme at 110℃ for 10 minutes, filter, take the solid phase and wash until neutral, freeze-dry under vacuum at -80℃ for 8 hours until loose powder is obtained, thus obtaining the high-purity sponge spicules; the mass ratio of the pretreated sponge, deionized water and alkaline protease is 1.25:100:0.03; the sodium hydroxide-sodium hexametaphosphate solution is prepared by mixing 6g sodium hydroxide, 2.0g sodium hexametaphosphate and 100mL deionized water evenly.

[0058] The silica content of the sponge bone needles prepared in Preparation Examples 1-4 was measured, as shown in Table 1 below.

[0059] Table 1

[0060]

[0061] As shown in Table 1, preparation examples 1-3 obtained high-purity sponge spicules after two enzymatic hydrolysis; preparation example 4 only underwent one enzymatic hydrolysis, resulting in a decrease in purity.

[0062] Example 1

[0063] A penetration-enhancing composition was prepared using the sponge bone needles obtained in Preparation Example 2 for repairing inflammatory acne. The preparation method of the penetration-enhancing composition includes the following steps:

[0064] A1. Take giant salamander skin collagen, pomegranate seed extract, honeysuckle extract, centella asiatica extract, early celery extract, iris extract, oyster shell extract, anhydrous ethanol, deionized water, and calcium chloride, mix them, and ultrasonically stir for 15 minutes to obtain material A; the mass ratio of the giant salamander skin collagen, pomegranate seed extract, honeysuckle extract, centella asiatica extract, early celery extract, iris extract, oyster shell extract, anhydrous ethanol, and deionized water is 25:8:4:18:4:2:3:40:60:0.4;

[0065] A2. Take the high-purity sponge spicules, N,N-dimethylformamide, and deionized water, mix them, and ultrasonically disperse them for 30 min. Add succinic anhydride and 3-aminopropyltriethoxysilane, heat and stir in a water bath at 50°C for 5 h, centrifuge, take the solid phase, wash, and freeze-dry under vacuum to obtain material B; the ratio of the high-purity sponge spicules, N,N-dimethylformamide, deionized water, succinic anhydride, and 3-aminopropyltriethoxysilane is 0.6 g: 2 mL: 30 mL: 4 g: 8 g;

[0066] A3. Take the material B, wheat oligopeptide and deionized water, mix them, sonicate for 20 min, centrifuge, take the solid phase, wash and dry to obtain material C; the ratio of material B, wheat oligopeptide and deionized water is 0.4g:10mg:10mL.

[0067] A4. Take the aforementioned material C and material A, mix them, ultrasonically disperse them for 30 minutes, maintain the pressure under a vacuum of -0.1 MPa for 30 minutes, filter them, and take the solid phase to obtain material D; the mass ratio of the aforementioned material C to material A is 1:10;

[0068] A5. Mix aluminum hydroxyl, tartaric acid, and deionized water, stir for 20 minutes, add sodium polyacrylate, continue stirring for 30 minutes, add material D, and continue stirring for 5 minutes to obtain the penetration-enhancing composition; the ratio of aluminum hydroxyl, tartaric acid, deionized water, sodium polyacrylate, and material D is 0.3g:0.4g:50mL:8g:1.5g.

[0069] Example 2

[0070] A penetration-enhancing composition was prepared using the sponge bone needles obtained in Preparation Example 2 for repairing inflammatory acne. The preparation method of the penetration-enhancing composition includes the following steps:

[0071] A1. Take giant salamander skin collagen, pomegranate seed extract, honeysuckle extract, centella asiatica extract, early celery extract, iris extract, oyster shell extract, anhydrous ethanol, deionized water, and calcium chloride, mix them, and ultrasonically stir for 20 minutes to obtain material A; the mass ratio of the giant salamander skin collagen, pomegranate seed extract, honeysuckle extract, centella asiatica extract, early celery extract, iris extract, oyster shell extract, anhydrous ethanol, and deionized water is 30:9:5:22:5:3:4:45:70:0.6;

[0072] A2. Take the high-purity sponge spicules, N,N-dimethylformamide, and deionized water, mix them, and ultrasonically disperse them for 35 min. Add succinic anhydride and 3-aminopropyltriethoxysilane, heat and stir in a water bath at 55℃ for 6 h, centrifuge, take the solid phase, wash, and freeze-dry under vacuum to obtain material B; the ratio of the high-purity sponge spicules, N,N-dimethylformamide, deionized water, succinic anhydride, and 3-aminopropyltriethoxysilane is 0.7 g: 2.5 mL: 30 mL: 4.5 g: 9 g;

[0073] A3. Take the material B, wheat oligopeptide and deionized water, mix them, sonicate for 25 min, centrifuge, take the solid phase, wash and dry to obtain material C; the ratio of material B, wheat oligopeptide and deionized water is 0.6g:12mg:12mL.

[0074] A4. Take the material C and material A, mix them, ultrasonically disperse them for 35 minutes, maintain the pressure under a vacuum of -0.1 MPa for 35 minutes, filter them, and take the solid phase to obtain material D; the mass ratio of material C to material A is 1:10.

[0075] A5. Mix aluminum hydroxyl, tartaric acid, and deionized water, stir for 25 minutes, add sodium polyacrylate, continue stirring for 35 minutes, add material D, and continue stirring for 8 minutes to obtain the penetration-enhancing composition; the ratio of aluminum hydroxyl, tartaric acid, deionized water, sodium polyacrylate, and material D is 0.4g:0.5g:50mL:9g:1.8g.

[0076] As attached Figure 2 As shown, the Fourier transform infrared (FTIR) spectra of the products obtained in different steps during the preparation process in this embodiment can indicate that wheat oligopeptides were successfully grafted onto the surface of the sponge spicules.

[0077] Example 3

[0078] A penetration-enhancing composition was prepared using the sponge bone needles obtained in Preparation Example 2 for repairing inflammatory acne. The preparation method of the penetration-enhancing composition includes the following steps:

[0079] A1. Take giant salamander skin collagen, pomegranate seed extract, honeysuckle extract, centella asiatica extract, early celery extract, iris extract, oyster shell extract, anhydrous ethanol, deionized water, and calcium chloride, mix them, and ultrasonically stir for 25 minutes to obtain material A; the mass ratio of the giant salamander skin collagen, pomegranate seed extract, honeysuckle extract, centella asiatica extract, early celery extract, iris extract, oyster shell extract, anhydrous ethanol, and deionized water is 35:10:6:26:6:4:5:50:80:0.8;

[0080] A2. Take the high-purity sponge spicules, N,N-dimethylformamide, and deionized water, mix them, and ultrasonically disperse them for 40 min. Add succinic anhydride and 3-aminopropyltriethoxysilane, heat and stir in a water bath at 60℃ for 7 h, centrifuge, take the solid phase, wash, and freeze-dry under vacuum to obtain material B; the ratio of the high-purity sponge spicules, N,N-dimethylformamide, deionized water, succinic anhydride, and 3-aminopropyltriethoxysilane is 0.8 g: 3 mL: 30 mL: 5 g: 10 g;

[0081] A3. Take the material B, wheat oligopeptide and deionized water, mix them, sonicate for 30 min, centrifuge, take the solid phase, wash and dry to obtain material C; the ratio of material B, wheat oligopeptide and deionized water is 0.8g:15mg:15mL.

[0082] A4. Take the aforementioned material C and material A, mix them, ultrasonically disperse them for 40 minutes, maintain the pressure under a vacuum of -0.1 MPa for 40 minutes, filter them, and take the solid phase to obtain material D; the mass ratio of the aforementioned material C to material A is 1:10;

[0083] A5. Mix aluminum hydroxyl, tartaric acid, and deionized water, stir for 30 minutes, add sodium polyacrylate, continue stirring for 40 minutes, add material D, and continue stirring for 10 minutes to obtain the penetration-enhancing composition; the ratio of aluminum hydroxyl, tartaric acid, deionized water, sodium polyacrylate, and material D is 0.5g:0.6g:50mL:10g:2.2g.

[0084] Comparative Example 1

[0085] This comparative example provides a penetration-enhancing composition. The only difference between this composition and Example 2 is that it does not contain giant salamander skin collagen. The reduced amount is allocated in parts to pomegranate seed extract, honeysuckle extract, centella asiatica extract, early celery extract, iris extract, oyster shell extract, and calcium chloride. The remaining components and their amounts remain unchanged.

[0086] Comparative Example 2

[0087] This comparative example provides a penetration-enhancing composition. The only difference between this composition and Example 2 is that it does not contain pomegranate seed extract. The reduced amount is allocated in parts to giant salamander skin collagen, honeysuckle extract, centella asiatica extract, early celery extract, iris extract, oyster shell extract, and calcium chloride. The remaining components and their amounts remain unchanged.

[0088] Comparative Example 3

[0089] This comparative example provides a penetration-enhancing composition. The only difference between this composition and Example 2 is that it does not contain honeysuckle extract. The reduced amount is allocated in parts to giant salamander skin collagen, pomegranate seed extract, centella asiatica extract, early celery extract, iris extract, oyster shell extract, and calcium chloride. The remaining components and their amounts remain unchanged.

[0090] Comparative Example 4

[0091] This comparative example provides a penetration-enhancing composition. The only difference between this composition and Example 2 is that it does not contain Centella asiatica extract. The reduced amount is allocated in parts to giant salamander skin collagen, pomegranate seed extract, honeysuckle extract, early celery extract, iris extract, oyster shell extract, and calcium chloride. The remaining components and their amounts remain unchanged.

[0092] Comparative Example 5

[0093] This comparative example provides a penetration-enhancing composition. The only difference between this composition and Example 2 is that it does not contain celery extract. The reduced amount is allocated in parts to giant salamander skin collagen, pomegranate seed extract, honeysuckle extract, centella asiatica extract, iris extract, oyster shell extract, and calcium chloride. The remaining components and their amounts remain unchanged.

[0094] Comparative Example 6

[0095] This comparative example provides a penetration-enhancing composition, which differs from Example 2 only in that it does not contain iris extract. The reduced amount is allocated in parts to giant salamander skin collagen, pomegranate seed extract, honeysuckle extract, centella asiatica extract, early celery extract, oyster shell extract, and calcium chloride, while the remaining components and amounts remain unchanged.

[0096] Comparative Example 7

[0097] This comparative example provides a penetration-enhancing composition. The only difference between this composition and Example 2 is that it does not contain oyster shell extract. The reduced amount is allocated in parts to giant salamander skin collagen, pomegranate seed extract, honeysuckle extract, centella asiatica extract, early celery extract, iris extract, and calcium chloride. The remaining components and their amounts remain unchanged.

[0098] Comparative Example 8

[0099] This comparative example provides a penetration-enhancing composition, which differs from Example 2 only in that it does not contain calcium chloride. The reduced amount is allocated in parts to giant salamander skin collagen, pomegranate seed extract, honeysuckle extract, centella asiatica extract, early celery extract, iris extract, and oyster shell extract, while the remaining components and amounts remain unchanged.

[0100] Comparative Example 9

[0101] The difference from Example 2 is that the preparation method of this penetration-enhancing composition includes the following steps:

[0102] A1. Take giant salamander skin collagen, pomegranate seed extract, honeysuckle extract, centella asiatica extract, early celery extract, iris extract, oyster shell extract, anhydrous ethanol, deionized water, and calcium chloride, mix them, and ultrasonically stir for 20 minutes to obtain material A.

[0103] A2. Take the high-purity sponge bone needles and material A, mix them, ultrasonically disperse them for 35 minutes, maintain the pressure for 35 minutes under a vacuum of -0.1 MPa, filter them, and take the solid phase to obtain material D; the mass ratio of the high-purity sponge bone needles to material A is 1:10.

[0104] A5. Mix aluminum hydroxyl, tartaric acid, and deionized water, stir for 25 minutes, add sodium polyacrylate, continue stirring for 35 minutes, add material D, and continue stirring for 8 minutes to obtain the penetration-enhancing composition; the ratio of aluminum hydroxyl, tartaric acid, deionized water, sodium polyacrylate, and material D is 0.4g:0.5g:50mL:9g:1.8g.

[0105] Comparative Example 10

[0106] The difference from Example 2 is that the preparation method of this penetration-enhancing composition includes the following steps:

[0107] A1. Take giant salamander skin collagen, pomegranate seed extract, honeysuckle extract, centella asiatica extract, early celery extract, iris extract, oyster shell extract, anhydrous ethanol, deionized water, and calcium chloride, mix them, and ultrasonically stir for 20 minutes to obtain material A.

[0108] A2. Mix aluminum hydroxyl, tartaric acid, and deionized water, stir for 25 minutes, add sodium polyacrylate, continue stirring for 35 minutes, add material A, and continue stirring for 8 minutes to obtain the penetration-enhancing composition; the ratio of aluminum hydroxyl, tartaric acid, deionized water, sodium polyacrylate, and material A is 0.4g:0.5g:50mL:9g:10g.

[0109] Performance testing

[0110] Test Example 1

[0111] One hundred and thirty 6-week-old ICR mice were randomly divided into 13 groups. The skin on the back of the mice was shaved (2cm x 2cm). From day 2 to 4 after shaving, 0.2 × 10⁻⁶ Propionibacterium acnes was injected into the skin on the back of the mice. 7 A mouse acne model was constructed using CFU / mL. Starting on day 5, the penetration-enhancing compositions prepared in Examples 1-3 and Comparative Examples 1-10 were applied by pressing and continued for 14 days. The acne area before and after application was compared, as shown in Table 2 below.

[0112] Table 2

[0113]

[0114] As shown in Table 2, the penetration-enhancing compositions prepared in Examples 1-3 of this application have good therapeutic effects on acne. The lack of synergistic effects between giant salamander skin collagen, pomegranate seed extract, honeysuckle extract, centella asiatica extract, celery extract, iris extract, oyster shell extract, and calcium chloride in Comparative Examples 1-8 resulted in a reduction in therapeutic effects. The absence of sponge spicule surface modification in Comparative Example 9 resulted in a reduction in therapeutic effects. The absence of sponge spicules in Comparative Example 10 led to poor penetration and a significant reduction in therapeutic effects.

[0115] Test Example 2: Treatment Effects on Volunteers

[0116] Test subjects: 130 volunteers with varying degrees of acne inflammation were randomly selected, aged 26-48 years. Among them, 68 had normal skin, 41 had oily skin, and 21 had dry skin. They were divided into 13 groups of 10, each corresponding to one of the penetration-enhancing compositions in the examples and comparative examples. The penetration-enhancing compositions prepared in Examples 1-3 and Comparative Examples 1-10 were applied to the inflamed acne areas of the patients.

[0117] Test method: Take 2g of the penetration-enhancing composition prepared in Examples 1-3 and Comparative Examples 1-10 above and apply it evenly to the acne inflammation area and within a radius of 3cm. Apply it once in the morning and once in the evening. After two weeks of continuous use, evaluate the effect of the penetration-enhancing composition. The test results are shown in Table 3.

[0118] Evaluation criteria for effectiveness:

[0119] Grade I: Significant repair effect, with marked improvement in acne-affected areas;

[0120] Grade II: Good repair effect, with significant improvement in acne areas;

[0121] Grade III: The repair effect is average, and there is no significant improvement in the acne area;

[0122] Grade IV: Poor repair effect, no improvement in acne areas.

[0123] Table 3

[0124]

[0125] As shown in Table 3, the penetration-enhancing compositions of Examples 1-3 were considered to have good repair effects; while the soothing and repairing effects of the penetration-enhancing compositions of Comparative Examples 1-9 were worse than those of Examples 1-3, with Comparative Example 10 being considered to have extremely poor effects.

[0126] The foregoing has described some embodiments of the present invention in detail, but the content described is only a preferred embodiment of the present invention and should not be considered as limiting the scope of the present invention. All equivalent changes and improvements made within the scope of the present invention should still fall within the patent coverage of the present invention.

Claims

1. A method for preparing high-purity sponge bone needles, characterized in that, Includes the following steps: S1. Take natural sponge, cut it into pieces, soak it in deionized water and ultrasonically vibrate it at a frequency of 35-45Hz for 1-2 hours, filter it, take the sponge fragments and wash them with deionized water 4-5 times to obtain the pretreated sponge. S2. Place the pretreated sponge in an extraction tank, add deionized water and alkaline protease, adjust the pH to 10, enzymatically hydrolyze at 40℃ for 4-6 hours, then inactivate the enzyme at 110℃ for 10 minutes, filter, take the solid phase, wash, and dry to obtain the first enzymatic hydrolysate. S3. Place the primary enzymatic hydrolysate in an extraction tank, add deionized water and ginger protease, adjust the pH to 4.5, enzymatically hydrolyze at 40℃ for 2-3 hours, then inactivate the enzyme at 110℃ for 10 minutes, filter, take the solid phase for washing, and obtain the secondary enzymatic hydrolysate. Freeze-dry at -80℃ under vacuum for 6-10 hours until it becomes a loose powder, thus obtaining the high-purity sponge bone needle. In step S2, the mass ratio of the pretreated sponge, deionized water, and alkaline protease is 1-1.5:100:0.02-0.

04. In step S3, the mass ratio of the primary enzymatic hydrolysate, deionized water, and ginger protease is 1-1.2:100:0.0075-0.

014.

2. A high-purity sponge bone needle prepared by the preparation method as described in claim 1.

3. The application of the high-purity sponge bone needle as described in claim 2 in the preparation of a permeation-enhancing composition, characterized in that, The method for preparing the penetration-enhancing composition includes the following steps: A1. Take giant salamander skin collagen, pomegranate seed extract, honeysuckle extract, centella asiatica extract, early celery extract, iris extract, oyster shell extract, anhydrous ethanol, deionized water, and calcium chloride, mix them, and ultrasonically stir for 15-25 minutes to obtain material A. A2. Take the high-purity sponge bone needles, N,N-dimethylformamide, and deionized water, mix them, and ultrasonically disperse them for 30-40 minutes. Add succinic anhydride and 3-aminopropyltriethoxysilane, heat and stir in a water bath at 50-60°C for 5-7 hours, centrifuge, wash the solid phase, and freeze-dry under vacuum to obtain material B. A3. Take the material B, wheat oligopeptides and deionized water, mix them, sonicate for 20-30 minutes, centrifuge, take the solid phase, wash and dry to obtain material C; A4. Take the material C and material A, mix them, ultrasonically disperse for 30-40 minutes, maintain pressure under vacuum conditions of gauge pressure -0.1MPa for 30-40 minutes, filter, and take the solid phase to obtain material D; A5. Mix aluminum hydroxide, tartaric acid, and deionized water, stir for 20-30 minutes, add sodium polyacrylate, continue stirring for 30-40 minutes, add material D, and continue stirring for 5-10 minutes to obtain the penetration-enhancing composition. In step A1, the mass ratio of the giant salamander skin collagen, pomegranate seed extract, honeysuckle extract, centella asiatica extract, early celery extract, iris extract, oyster shell extract, anhydrous ethanol, and deionized water is 25-35:8-10:4-6:18-26:4-6:2-4:3-5:40-50:60-80:0.4-0.8; In step A2, the ratio of the high-purity sponge bone needle, N,N-dimethylformamide, deionized water, succinic anhydride and 3-aminopropyltriethoxysilane is 0.6-0.8g: 2-3mL: 30mL: 4-5g: 8-10g; In step A3, the ratio of material B, wheat oligopeptide and deionized water is 0.4-0.8g: 10-15mg: 10-15mL; In step A4, the mass ratio of material C to material A is 1:10; In step A5, the ratio of aluminum hydroxyl, tartaric acid, deionized water, sodium polyacrylate, and material D is 0.3-0.5g: 0.4-0.6g: 50mL: 8-10g: 1.5-2.2g.

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