A sponge bone needle with surface-adsorbed nanoparticles, and a preparation method and use thereof

Abstract

The application discloses a sponge bone needle with surface-adsorbed nanoparticles and a preparation method and application thereof, and comprises modified bee sponge bone needles with active silicon hydroxyl groups on surfaces as carriers and mesoporous silica nanoparticles loaded on the active silicon hydroxyl groups, and the average particle size of the mesoporous silica nanoparticles is 90-1000 nm. The sponge bone needle can pierce into the cuticle to open the skin barrier, and can be retained in the cuticle for a long time to form a large number of sustained microchannels (not less than 72 h), release the nanoparticles carrying more drugs, and enter the skin to release the drugs to play a role.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of sponge spicules, and particularly relates to a sponge spicule with surface-adsorbed nanoparticles and a preparation method and application thereof. BACKGROUND

[0002] With the increasing demand for patient-friendly therapies, transdermal drug delivery, a forward-looking drug delivery strategy, is continuously developed to compensate for the limitations of oral and injection drug delivery. However, due to the presence of the stratum corneum, transdermal drug delivery is not easy. It is generally believed that only a small number of drug molecules with moderate oil-water partition coefficients (molecular weight not higher than 500 Dalton) can be effectively absorbed through the skin. Therefore, skin penetration enhancement technology has become a hot spot in the field of transdermal drug delivery.

[0003] Current skin penetration enhancement technologies are divided into physical penetration enhancement and chemical penetration enhancement. Microneedle technology is a popular physical penetration enhancement technology in recent years. Microneedle technology can open a large number of microchannels on the surface of the skin with minimal damage, allowing drugs and drug carriers to transport into the skin through the microchannels.

[0004] Sponge Haliclona sp. Spicules (SHS) is a new type of siliceous microneedle that can be used for transdermal drug delivery. The morphological structure of the sponge spicule is free double-pointed, and the spicule tip is relatively sharp. It has a smooth surface and its size does not change easily, has high mechanical strength, can continuously open the skin channel for large-area use, and can maintain good efficacy after acting on the skin surface for 72 hours. At the same time, the damage of the sponge spicule to the skin is almost negligible, so it can effectively avoid problems such as bleeding and infection. While significantly enhancing the transdermal absorption capacity of hydrophilic macromolecules, it can also ensure the safety and controllability of the drug delivery process, and is a very promising penetration enhancement technology. However, SHS also has certain limitations: for hydrophilic drugs, as the molecular weight of the drug increases, the penetration enhancement effect of SHS on the drug will gradually decrease, and lipophilic drugs cannot penetrate into the deep skin through the hydrophilic channels opened by SHS. SUMMARY

[0005] The present application aims to overcome the defects of the prior art and provide a sponge spicule with surface-adsorbed nanoparticles.

[0006] Another object of the present application is to provide a preparation method of the sponge spicule with surface-adsorbed nanoparticles.

[0007] Still another object of the present application is to provide the use of the sponge spicule with surface-adsorbed nanoparticles.

[0008] The technical solution of the present application is as follows:

[0009] A sponge skeletal structure with surface adsorbed nanoparticles includes a modified honeycomb sponge skeletal structure with active silanol groups on its surface as a carrier and mesoporous silica nanoparticles loaded thereon through the active silanol groups, wherein the average particle size of the mesoporous silica nanoparticles is 90-1000 nm.

[0010] The modified bee sponge spicules mentioned above were prepared by washing bee sponge spicules and then reacting them sequentially with NaOH solution and hydrochloric acid solution.

[0011] The above-mentioned mesoporous silica nanoparticles were prepared by reacting CTAB, NaOH and TEOS, and then modified with APTES.

[0012] The modified bee sponge spicules and mesoporous silica nanoparticles mentioned above are linked by a reaction with a citric acid-trisodium citrate aqueous solution.

[0013] In a preferred embodiment of the present invention, the pH of the citric acid-trisodium citrate aqueous solution is 4.

[0014] The preparation method of the above-mentioned sponge bone needles with surface adsorbed nanoparticles includes the following steps:

[0015] (1) Mesoporous silica nanoparticles were prepared using CTAB, NaOH and TEOS;

[0016] (2) Modify the above mesoporous silica nanoparticles with APTES to obtain APTES-modified mesoporous silica nanoparticles.

[0017] (3) The cleaned bee sponge spicules were treated with NaOH solution and hydrochloric acid solution in sequence to obtain modified bee sponge spicules with active silanol groups on the surface.

[0018] (4) The APTES-modified mesoporous silica nanoparticles obtained in step (2) and the modified bee sponge spicules obtained in step (3) are reacted with an aqueous solution of citric acid-trisodium citrate, and then washed and dried to obtain the final product.

[0019] In a preferred embodiment of the present invention, step (1) is as follows: CTAB and NaOH are added to distilled water, mixed evenly, heated to 80°C and stirred continuously. The ratio of CTAB, NaOH and distilled water is 1g:0.25-0.5g:480mL. Then, 5-10 times the mass of TEOS of CTAB is added dropwise. After stirring at 1300-1500rpm for 2-3 hours at 80°C, the obtained white precipitate is washed with distilled water and methanol in sequence, then dried, and finally CTAB is removed by calcination to obtain mesoporous silica nanoparticles.

[0020] In a preferred embodiment of the present invention, step (2) is as follows: the mesoporous silica nanoparticles obtained in step (1) are added to distilled water to obtain an MSNs suspension; then 175 μL / mL of APTES aqueous solution is added to the above MSNs suspension under dry nitrogen purging, and then the reaction is stirred and refluxed at 77°C under a nitrogen atmosphere. The reaction product is washed with ethanol and dried to obtain APTES-modified mesoporous silica nanoparticles.

[0021] In a preferred embodiment of the present invention, step (3) is as follows: the bee sponge spicules are ultrasonically cleaned with water, ethanol and acetone in sequence, and dried to obtain a clean SHS. Then, it is added to a 5 mol / L NaOH solution and stirred for 30 min, and washed with distilled water. Then, it is added to a 10% hydrochloric acid solution and stirred for 30 min, and washed with distilled water. After drying, the modified bee sponge spicules with active silanol groups on the surface are obtained.

[0022] In a preferred embodiment of the present invention, step (4) is as follows: prepare an aqueous solution of citric acid-trisodium citrate with pH 4, add the APTES-modified mesoporous silica nanoparticles obtained in step (2) and the modified honeycomb sponge spicules obtained in step (3) in a mass ratio of 7:1, stir and react, wash the resulting reaction product with distilled water and dry it to obtain the sponge spicules with surface adsorbed nanoparticles.

[0023] More preferably, the citric acid-trisodium citrate aqueous solution is prepared from a 0.1 mol / L citric acid aqueous solution and a 0.1 mol / L trisodium citrate aqueous solution.

[0024] The above-mentioned sponge bone needles with surface adsorbed nanoparticles are used as transdermal drug delivery carriers.

[0025] A transdermal drug delivery carrier comprising a sponge bone needle having the aforementioned surface-adsorbed nanoparticles.

[0026] The beneficial effects of this invention are:

[0027] 1. This invention can penetrate the stratum corneum to open the skin barrier and remain in the stratum corneum for a long time to form a large number of continuous microchannels (not less than 72 hours), releasing nanoparticles carrying more drugs to enter the skin and release the drugs to exert their effects.

[0028] 2. This invention can utilize the ability of mesoporous silica nanoparticles to simultaneously load multiple drugs, thereby enabling the simultaneous administration of different types of drugs and potentially achieving synergistic treatment of diseases with multiple drugs.

[0029] 3. This invention can load different types of drug molecules into mesoporous silica nanoparticles, improving the solubility or stability of the drugs. The drugs will be inserted into the skin along with this invention and will not be affected by the physicochemical properties of the drugs themselves, and then released from the mesopores of the mesoporous silica nanoparticles.

[0030] 4. After the present invention is applied to the skin, it will automatically detach from the skin as the keratinocytes shed physiologically, and the skin barrier can recover on its own without causing serious damage to the skin.

[0031] 5. The difference between this invention and the previously prepared mesoporous bee sponge bone needle (mSHS) is that after mSHS is inserted into the skin, the drug is released from its mesoporous layer, while after this invention is inserted into the skin, the nanoparticles can first diffuse to a deeper layer of the skin before the drug is released. Attached Figure Description

[0032] Figure 1 The images shown are scanning electron microscope (SEM) images of the modified bee sponge spicules in Embodiment 1 of the present invention, wherein: a) is the morphology of the modified bee sponge spicules under a 500x magnification SEM field, b) is the morphology of the modified bee sponge spicules under a 5000x magnification SEM field, and c) is the morphology of the modified bee sponge spicules under a 10000x magnification SEM field.

[0033] Figure 2 The images shown are scanning electron microscope (SEM) images of mesoporous silica nanoparticles in Example 1 of this invention, wherein: a is MSNs prepared under condition A with an average diameter of 90 nm; b is MSNs prepared under condition B with an average diameter of 120 nm; c is MSNs prepared under condition C with an average diameter of 500 nm; and d is MSNs prepared under condition D with an average diameter of 1000 nm.

[0034] Figure 3 The images shown are scanning electron microscope (SEM) images of mesoporous silica nanoparticles before and after modification in Example 1 of this invention. Specifically, MSNs (i.e., MSNs500) prepared under experimental condition C are modified with amino groups using a post-grafting method. a is MSNs500 before modification, and b is MSNs500 after modification.

[0035] Figure 4 This is a scanning electron microscope image of the bee sponge spicules (nSHS) with surface adsorption of mesoporous nanoparticles obtained in Example 1 of the present invention.

[0036] Figure 5 This is a graph showing the changes in the content of coumarin 6 in different skin layers of nSHS after 16 hours of in vitro transdermal transdermal transmission following loading with coumarin 6 in Example 2 of the present invention.

[0037] Figure 6This demonstrates that in Embodiment 3 of the present invention, the nSHS can simultaneously carry two different drugs. Detailed Implementation

[0038] The technical solution of the present invention will be further explained and described below with reference to specific embodiments and accompanying drawings.

[0039] Example 1

[0040] (1) Preparation of mesoporous silica nanoparticles (MSNs): Sodium hydroxide and N-hexadecyltrimethylammonium bromide (CTAB) were added to 480 mL of distilled water according to Table 1. The resulting solution was heated to 80 °C and stirred continuously. Then, tetraethyl orthosilicate (TEOS) was added dropwise to the above solution, and the mixture was stirred vigorously at 80 °C for 2-3 h. The resulting white precipitate was washed three times with distilled water and methanol, and then dried in an oven for 8 h. Finally, the product was placed in a muffle furnace and heated to 550 °C at 4 °C / min for 3 h to remove CTAB and obtain MSNs (e.g., ...). Figure 2 and Figure 3 (As shown).

[0041] The experimental conditions and material usage for this step are shown in Table 1.

[0042] Table 1 Experimental conditions and material usage for MSNs preparation

[0043]

[0044]

[0045] (2) MSNs modification: 70 mg of the MSNs prepared in step (1) was added to 75 mL of distilled water to prepare an MSNs suspension, which was then added to a three-necked flask. 2 mL of a 175 μL / mL LAPTES aqueous solution was prepared and added to the MSNs suspension under dry nitrogen purging. The mixture was then refluxed at 77 °C under nitrogen atmosphere with magnetic stirring (500 rpm) for 14 h, washed three times with ethanol, and dried at 60 °C for 8 h. Figure 3 As shown, this modification process does not change the shape and average diameter of MSNs.

[0046] (3) SHS Modification: SHS was ultrasonically cleaned sequentially with 500 mL of water, 500 mL of ethanol, and 400 mL of acetone, and then dried to obtain clean SHS. 8 g of the clean SHS was added to 400 mL of 5 mol / L NaOH solution and stirred for 30 min, then washed with distilled water. The product was then added to 400 mL of 10% hydrochloric acid solution and stirred for 30 min. The resulting product was washed with distilled water and dried to obtain modified SHS with active silanol groups on the surface.Figure 1 As shown, the modified SHS has a smooth surface, but exhibits a porous structure with a diameter of approximately 100 nm.

[0047] (4) Combination: Using citric acid monohydrate, trisodium citrate, and distilled water as raw materials, 0.1 mol / L citric acid aqueous solution and 0.1 mol / L trisodium citrate aqueous solution were prepared. 32.75 mL of 0.1 mol / L citric acid aqueous solution was added to 17.25 mL of 0.1 mol / L trisodium citrate aqueous solution to prepare 50 mL of citric acid-trisodium citrate aqueous solution with pH 4. 70 mg of modified MSN-NH2 and 10 mg of modified SHS were added to 50 mL of citric acid-trisodium citrate aqueous solution with pH 4, and the mixture was stirred for 8 h (200 rpm). The resulting product was washed three times with distilled water and dried at 60 °C for 8 h to obtain the product as shown. Figure 4 The honeycomb sponge spicules (nSHS) shown are surface-adsorbed mesoporous silica nanoparticles.

[0048] Table 2 Results of MSNs prepared under different experimental conditions and material dosages

[0049]

[0050]

[0051] The comparative experimental results above show that increasing the amount of both NaOH and TEOS during MSN preparation leads to larger MSN sizes. This is because TEOS serves as the silicon source in MSN preparation; increasing the amount of TEOS provides more silicon, resulting in larger MSNs. Furthermore, it can be observed that as the MSN size increases, its shape shifts from spherical to more disc-like.

[0052] Studies have shown that increasing CTAB concentration is accompanied by an increase in the particle size of synthesized silica. NaOH, as an acid-base regulator, results in a limited number of negatively charged adsorbed silicon precursors during the reaction when the alkalinity is weak, leading to poor binding with surfactant micelles and the formation of amorphous silica products. However, in strongly alkaline conditions, the rapid hydrolysis of TEOS also hinders the formation of long-chain oligomeric silica particles. This can result in a significant portion of the silicon source failing to fully bind with CTAB micelles, leading to the formation of solid silica. Furthermore, the interaction between different morphologies of CTAB micelles and silicon precursors is unlikely to reach equilibrium within a short timeframe, resulting in particles with varying morphologies. As reaction time increases, the hydrophobic properties of the micelles modify the nanoparticles. Sufficient time is also allowed for close packing within the nanoparticles, resulting in uniformly morphological mesoporous silica nanoparticles.

[0053] Therefore, the amounts of CTAB, NaOH, and TEOS, as well as the reaction temperature and time, are all very important for the preparation of MSNs. To prepare MSNs with uniform morphology, stable size, and large specific surface area, it is necessary to pay close attention to each preparation condition.

[0054] Example 2

[0055] In this embodiment, coumarin 6 was used as a lipid-soluble model drug to study the drug loading and transdermal delivery effects of the nSHS prepared in Example 1 on coumarin 6.

[0056] (1) nSHS loading method: Weigh 10 mg of nSHS into a 1.5 mL EP tube, then add 600 μL of 40 mg / mL coumarin 6 solution (solvent: dichloromethane), vortex to mix thoroughly, then wrap the centrifuge tube with aluminum foil and place it on a rotary mixer to mix for 12 h (70 rpm). Remove the centrifuge tube from the rotary mixer and let it stand for 5 min to allow the nSHS to settle naturally to the bottom of the centrifuge tube. Centrifuge the centrifuge tube in a centrifuge (1500 rpm, 5 min), and remove the supernatant as completely as possible with a pipette. Wrap the centrifuge tube with the drug removed from the supernatant in aluminum foil and freeze-dry it in a freeze dryer (or air dry it in a fume hood) to obtain nSHS@coumarin 6.

[0057] (2) Quantitative determination of nSHS@coumarin 6 loading: nSHS@coumarin 6 was placed in a certain amount of ethanol solution and shaken overnight in a shaker (180 rpm, 28℃). 1 mL of the extracted solution was taken, and the fluorescence value was measured using an ELISA reader. The result was substituted into the standard curve (standard curve of coumarin 6 in ethanol solution: y = 15468x + 282.68, R0). 2 =0.999. y is the fluorescence value, x is the concentration of coumarin 6 (in μg / mL), and then the drug loading of the corresponding nSHS is calculated based on the coumarin 6 content in the ethanol solution (as shown in Table 3).

[0058] Table 3. Nanoparticle size and corresponding drug loading of nSHS-carried nanoparticles.

[0059]

[0060] It can be observed that as the particle size of the nanoparticles carried by nSHS increases, its drug loading also increases accordingly.

[0061] (3) Study on the in vitro transdermal permeation enhancement effect of nSHS@coumarin 6:

[0062] This embodiment uses ex vivo pig skin as a skin model for the experiment.

[0063] First, select pigskin from the back of a pig without obvious external injuries. Then, carefully remove the subcutaneous fat tissue with a scalpel. Next, use an electric shaver to shave off the hair on the pigskin, reducing its length to less than 5mm. Wash the treated pigskin with ultrapure water and store it at -20°C for later use. Thaw the skin at room temperature before use. Drill a piece of pigskin of the same diameter using a 40mm diameter circular punch and attach it to the Franz transdermal diffusion device.

[0064] Transdermal resistance testing of isolated skin was used to determine the conductivity of the skin to ensure the integrity of the skin barrier. Specifically, a waveform generator was used to measure the current passing through pig skin at 100mV and 100Hz. Pig skin with a current less than 5A was considered intact and allowed to proceed to the next stage of the experiment. The effective penetration area of ​​the Franz diffusion cell was 1.77 cm², and the receptor volume was 12 mL.

[0065] Skin was placed atop a vertical Franz diffusion cell, and the receptor cell was filled with 0.2M PBS solution (pH 7.4) containing 1% Tween 80. A small rotor was then added to the receptor cell. The in vitro transdermal experiments were conducted in six groups, with three replicates per group.

[0066] nSHS@Coumarin 6 group: Take a certain amount of nSHS powder (according to step (1) above in this embodiment), add 1 mL of 0.2M PBS to dissolve it, and add 200 μL to the administration cavity to make the coumarin 6 dosage 200 μg, so that it is evenly distributed on the pigskin surface; use a household electric massager (apply a force of about 0.3N, a speed of about 300 rpm) to apply nSHS locally to the skin for 2 minutes. Then seal the administration cavity with sealing film.

[0067] SHS+MSNs@Coumarin 6 group: 100 μL of PBS (0.2 M, pH = 7.4) containing 10 mg SHS was placed in the administration cavity and evenly distributed on the pigskin surface. The SHS was applied topically to the skin for 2 minutes using a household electric massager (approximately 0.3 N of force, approximately 300 rpm). After application, the skin was washed three times with 0.2 M PBS solution to remove residual SHS. Then, 200 μL of MSNs1000 powder suspension loaded with coumarin 6 (containing 200 μg of coumarin 6) was added to the administration cavity, evenly distributed on the pigskin surface, and the administration cavity was sealed with a sealing film.

[0068] mSHS@Coumarin 6 group: 100 μL of PBS (0.2 M, pH = 7.4) solution containing 10 mg mSHS@coumarin 6 was placed in the administration cavity to distribute it evenly on the surface of the pig skin; mSHS@coumarin 6 was applied locally to the skin for 2 minutes using a household electric massager (applying a force of about 0.3 N and a rotation speed of about 300 rpm / min) to massage the mSHS@coumarin 6 as evenly as possible on the skin surface; finally, the administration cavity was sealed with a sealing film; the mSHS is a mesoporous sponge spicule, see CN 112237567 A "A mesoporous sponge spicule and its preparation method and use".

[0069] SHS + Coumarin 6 group: 100 μL of PBS (0.2 M, pH = 7.4) containing 10 mg SHS was placed in the administration cavity and evenly distributed on the pigskin surface. The SHS was applied topically to the skin for 2 minutes using a household electric massager (approximately 0.3 N of force, approximately 300 rpm). After application, the skin was washed three times with 0.2 M PBS solution to remove residual SHS. Then, 200 μL of saturated coumarin 6 solution (30% ethanol) was added to the administration cavity, ensuring even distribution on the pigskin surface. The administration cavity was then sealed with a sealing film.

[0070] Control group: 200 μL of saturated coumarin 6 solution (solvent is 30% ethanol) was added into the administration cavity, and the coumarin 6 solution was evenly distributed on the surface of the pig skin. Then the administration cavity was sealed with sealing film.

[0071] Blank group: 200 μL of 0.2 M PBS solution (pH = 7.4) containing 1% Tween 80 was added to the administration cavity and evenly distributed on the pigskin surface. The administration cavity was then sealed with sealing film.

[0072] The Franz diffusion cells of each group were placed in a water bath at 37±0.5℃ and the rotation speed was set to 600 rpm / min to ensure uniform dispersion of the drug in the receptor cell. The receptor cell outlet was then sealed with a sealing film and allowed to permeate for 16 hours in the dark. Each group was repeated at least three times. After transdermal treatment, the residual drug in the administration cavity was washed five times with PBS solution (control group) or ethanol solution. The skin was then removed, wrapped in aluminum foil to protect it from light, and kept for later use. One mL of the solution from the receptor cell was taken to determine the content of coumarin 6 in the receptor cell.

[0073] The liquid in the receptor pool was removed, and the content of coumarin 6 that had penetrated into the subcutaneous tissue was measured. Then, the content of coumarin 6 in each layer of the skin was measured using the tape peeling method.

[0074] The content of coumarin 6 in various dermal layers was determined using the adhesive tape peeling method. Skin removed after transdermal treatment was fixed to the peeling device, and the stratum corneum (SC) was peeled 10 times consecutively using Scotch® transparent tape. The peeled tapes were collected and placed in brown glass bottles according to the following protocol: the first strip was designated as stratum corneum 1, the second to fifth strips as stratum corneum 2-5, and the sixth to tenth strips as stratum corneum 6-10. After the stratum corneum collection was completed, the active epidermal layer was collected by separating it from the dermis using a scalpel and placing the separated active epidermal layer in a brown glass bottle. Finally, two dermal skin samples were drilled using a punch (d=5mm), shredded with a scalpel, and placed in brown glass bottles. 4 mL of methanol solution was added to each of the five brown glass bottles, and the bottles were then shaken overnight (25°C, 180 rpm) to fully extract coumarin 6 from the skin. After extraction, the extract in the glass bottle was centrifuged (5000 rpm, 5 min), and the supernatant was used to determine the content of coumarin 6.

[0075] Extracts from the stratum corneum, active epidermis, and dermis, as well as solutions from the receiving cell, were placed under an ELISA reader to measure their corresponding fluorescence values ​​(excitation wavelength 466 nm, emission wavelength 504 nm). The results were then substituted into a standard curve to obtain the content of coumarin 6 in different epidermal layers.

[0076] The experimental results are shown in Figure 5 Quantitative analysis of the experimental results showed that the transdermal permeation rates of the nSHS90@coumarin 6 group (31.26±5.29 μg), nSHS500@coumarin 6 group (35.41±5.42 μg), and nSHS1000@coumarin 6 group (59.76±21.13 μg) were all higher than those of the SHS+coumarin 6 group (1.88±0.59 μg) and the control group (1.19±0.17 μg). With consistent coumarin 6 dosage, the total permeation rate of the nSHS@coumarin 6 group showed a trend of increasing with the increase of the adsorbed MSN particle size. The transdermal penetration of the nSHS1000@coumarin 6 group was approximately 50.22 times that of the control group. The drug permeation rate of the nSHS1000@coumarin 6 group was 29.88% ± 10.57%, while the drug permeation rate of the control group was only 0.59% ± 0.08%. There was no significant difference in the total permeation amount between the SHS+coumarin 6 group and the control group. This indicates that lipophilic drugs generally have difficulty directly penetrating the skin, and SHS alone has no significant permeation-enhancing effect when used to deliver lipophilic drugs.

[0077] The experimental results show that, compared with the direct application of the drug and the application of SHS, nSHS carrying coumarin 6 significantly increased the total amount of coumarin 6 transdermally, but it was still concentrated in the stratum corneum and epidermis. However, its effect on promoting the transdermal delivery of coumarin 6 was still quite significant.

[0078] Secondly, this embodiment compares SHS-adsorbed MSNs with SHS-mixed MSNs. The total amount of permeation in the nSHS@coumarin 6 group was higher than that in the SHS+MSNs@coumarin 6 group (19.69±5.66μg). This shows that the administration method of applying SHS massage first and then adding MSNs@coumarin 6 cannot achieve the permeation-enhancing effect of nSHS.

[0079] Meanwhile, this embodiment compared nSHS with the previously prepared mesoporous bone needle (mSHS) and found that nSHS can greatly increase the drug loading of coumarin 6 and increase the transdermal delivery of coumarin 6.

[0080] Therefore, experiments have shown that nSHS can improve drug solubility, increase drug loading, and promote the transdermal absorption of poorly soluble drugs. Furthermore, the total amount of poorly soluble drugs permeating increases with the increase in the particle size of the adsorbed MSNs.

[0081] Example 3

[0082] In this embodiment, fluorescein isothiocyanate (FITC) and protoporphyrin IX (PpIX) were used as model drugs to study the feasibility of the nSHS prepared in Example 1 carrying multiple drugs simultaneously.

[0083] (1) Drug loading

[0084] Weigh 70 mg of MSNs1000 into a 2 mL EP tube, then add 1.5 mL of an aqueous solution containing 5 mg of FITC; vortex to mix thoroughly, then wrap the centrifuge tube with aluminum foil and place it on a rotary mixer to mix for 12 h (70 rpm); remove the centrifuge tube from the rotary mixer and let it stand for 5 min to allow the MSNs to settle naturally at the bottom of the centrifuge tube; centrifuge the centrifuge tube in a centrifuge (1500 rpm, 5 min), and after centrifugation, use a pipette to remove the supernatant as completely as possible; wrap the centrifuge tube with the removed supernatant in aluminum foil and freeze-dry it in a freeze dryer (or place it in a fume hood to air dry naturally) to obtain MSNs@FITC.

[0085] Weigh 70 mg of MSNs1000 into a 2 mL EP tube, then add 1 mL of 5 mg / mL PpIX solution (DMSO solvent); vortex to mix thoroughly, then wrap the centrifuge tube with aluminum foil and place it on a rotary mixer to mix for 12 h (70 rpm); remove the centrifuge tube from the rotary mixer and let it stand for 5 min to allow the MSNs to settle naturally at the bottom of the centrifuge tube; centrifuge the centrifuge tube in a centrifuge (1500 rpm, 5 min), and after centrifugation, use a pipette to remove the supernatant as completely as possible; wrap the centrifuge tube after removing the drug supernatant with aluminum foil and freeze-dry it in a freeze dryer (or place it in a fume hood to air dry naturally) to obtain MSNs@PpIX.

[0086] (2) MSNs modifiers

[0087] To prepare an MSNs suspension, 30 mg of MSNs@FITC and 30 mg of MSNs@PpIX were added to 75 mL of distilled water and the solution was placed in a three-necked flask. 2 mL of a 175 μL / mL APTES aqueous solution was prepared and added to the MSNs suspension under dry nitrogen purging. The mixture was then refluxed at 77 °C under nitrogen atmosphere with magnetic stirring (500 rpm) for 14 h. After washing three times with ethanol, the solution was freeze-dried.

[0088] (3) SHS modification

[0089] 0.5 g of SHS was ultrasonically cleaned sequentially with 50 mL of water, 50 mL of ethanol, and 50 mL of acetone, and then dried to obtain SHS with a clean surface. The clean SHS was then added to 200 mL of 5 mol / L NaOH solution and stirred for 30 min, followed by washing with distilled water. The product was then added to 200 mL of 10% hydrochloric acid solution and stirred for 30 min. The resulting product was washed with distilled water and dried to obtain modified SHS with active silanol groups on its surface.

[0090] (4) Combination

[0091] Using citric acid monohydrate, trisodium citrate, and distilled water as raw materials, 0.1 mol / L citric acid aqueous solution and 0.1 mol / L trisodium citrate aqueous solution were prepared. 32.75 mL of 0.1 mol / L citric acid aqueous solution was added to 17.25 mL of 0.1 mol / L trisodium citrate aqueous solution to prepare 50 mL of citric acid-trisodium citrate aqueous solution with pH 4. 60 mg of modified MSN-NH2 and 10 mg of modified SHS were added to 50 mL of citric acid-trisodium citrate aqueous solution with pH 4, and the mixture was stirred for 8 h (200 rpm). The resulting product was washed three times with distilled water and then freeze-dried in a freeze dryer to obtain MSNs-NH2-SHS (abbreviated as nSHS).

[0092] When the nSHS, which successfully carried two drugs, was observed under a fluorescence confocal microscope, different colors were observed on the surface of the bone needles in different channels. Figure 6 (FITC is green, PpIX is red), proving that nSHS can carry two different drugs at the same time.

[0093] The above description is merely a preferred embodiment of the present invention, and therefore should not be construed as limiting the scope of the present invention. All equivalent changes and modifications made in accordance with the scope of the patent and the contents of the specification should still fall within the scope of the present invention.

Claims

1. A sponge bone needle with surface adsorbed nanoparticles, characterized in that: The invention includes modified bee sponge spicules with active silanol groups on their surface as a carrier, and mesoporous silica nanoparticles loaded thereon through the active silanol groups, wherein the average particle size of the mesoporous silica nanoparticles is 90-1000 nm. The modified bee sponge spicules mentioned above were prepared by washing bee sponge spicules and then reacting them sequentially with NaOH solution and hydrochloric acid solution. The above-mentioned mesoporous silica nanoparticles were prepared by reacting CTAB, NaOH and TEOS, and then modified with APTES. The modified bee sponge spicules and mesoporous silica nanoparticles are linked by a reaction with a citric acid-trisodium citrate aqueous solution at pH 4. Its preparation method includes the following steps: (1) Mesoporous silica nanoparticles were prepared using CTAB, NaOH and TEOS; (2) Modify the above mesoporous silica nanoparticles with APTES to obtain APTES-modified mesoporous silica nanoparticles. (3) The cleaned bee sponge spicules were treated with NaOH solution and hydrochloric acid solution in sequence to obtain modified bee sponge spicules with active silanol groups on the surface; (4) The APTES-modified mesoporous silica nanoparticles obtained in step (2) and the modified bee sponge spicules obtained in step (3) are reacted with a citric acid-trisodium citrate aqueous solution, and then washed and dried to obtain the final product.

2. The sponge bone needle with surface adsorption of nanoparticles as described in claim 1, characterized in that: Step (1) is as follows: CTAB and NaOH are added to distilled water, mixed evenly, heated to 80°C and stirred continuously. The ratio of CTAB, NaOH and distilled water is 1g: 0.25-0.5g: 480mL. Then, 5-10 times the mass of TEOS of CTAB is added dropwise. After stirring at 1300-1500rpm for 2-3 hours at 80°C, the obtained white precipitate is washed with distilled water and methanol in sequence, then dried, and finally CTAB is removed by calcination to obtain mesoporous silica nanoparticles.

3. The sponge bone needle with surface adsorbed nanoparticles as described in claim 1, characterized in that: Step (2) is as follows: the mesoporous silica nanoparticles obtained in step (1) are added to distilled water to obtain an MSNs suspension; then 175 μL / mL of APTES aqueous solution is added to the above MSNs suspension under dry nitrogen purging, and then the reaction is stirred and refluxed at 77°C under a nitrogen atmosphere. The reaction product is washed with ethanol and dried to obtain APTES modified mesoporous silica nanoparticles.

4. The sponge bone needle with surface adsorbed nanoparticles as described in claim 1, characterized in that: Step (3) is as follows: the bee sponge spicules are ultrasonically cleaned with water, ethanol and acetone in sequence, and dried to obtain a clean SHS. Then, it is added to a 5 mol / L NaOH solution and stirred for 30 min, then washed with distilled water. After that, it is added to a 10% hydrochloric acid solution and stirred for 30 min, then washed with distilled water. After drying, the modified bee sponge spicules with active silanol groups on the surface are obtained.

5. A sponge needle with surface-adsorbed nanoparticles as described in claim 1, characterized in that: Step (4) is as follows: Prepare a citric acid-trisodium citrate aqueous solution with pH 4, add the APTES-modified mesoporous silica nanoparticles obtained in step (2) and the modified honeycomb sponge spicules obtained in step (3) in a mass ratio of 7:1, stir and react, wash the resulting reaction product with distilled water and dry it to obtain the sponge spicules with surface adsorbed nanoparticles.

6. The sponge bone needle with surface adsorbed nanoparticles as described in claim 5, characterized in that: The citric acid-trisodium citrate aqueous solution is prepared by mixing 0.1 mol / L citric acid aqueous solution and 0.1 mol / L trisodium citrate aqueous solution.

7. A transdermal drug delivery carrier, characterized in that: Sponge bone needles having surface-adsorbed nanoparticles as described in any one of claims 1 to 6.

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