Eugenol soluble microneedle patch and preparation method thereof

Abstract

The invention discloses a eugenol soluble microneedle patch and a preparation method thereof. The eugenol soluble microneedle patch is composed of a microneedle array and a backing, the microneedle array is prepared from an amphiphilic polymer, a water-soluble polymer and eugenol, and the mass ratio of the amphiphilic polymer to the water-soluble polymer is 1: 1-3: 1; the ratio of the total mass of the amphiphilic polymer and the water-soluble polymer to the mass of the eugenol is (1: 1)-(6: 1). After the amorphous solid dispersion is prepared by a hot-melt extrusion method, the amorphous solid dispersion is pressed into the microneedle patch by a hot-pressing method, and the prepared microneedle patch is good in mechanical property and excellent in treatment effect on acute pain. Compared with combination of a traditional solution casting method, a hot melt extrusion technology and a hot pressing technology, the preparation technology of the microneedle is simplified, industrial production prospects are achieved, no organic reagent is used in the preparation process of the microneedle, and the production process is environmentally friendly.

Description

Technical Field

[0001] The present invention belongs to the technical field of biomedicine, and in particular relates to an in-situ self-emulsifying eugenol soluble microneedle patch based on amorphous solid dispersion technology and a preparation method thereof. Background Art

[0002] Pain is a painful feeling accompanied by unpleasant emotions such as fear, tension, and uneasiness, and is accompanied by cardiovascular and respiratory changes. It is reported that the proportion of pain patients around the world has increased from 26.3% in 2009 to 32.1% in 2021. Acute pain includes gastric perforation, angina pectoris, dysmenorrhea, migraine and postoperative pain. It is a common disease. If it is not treated properly, it will cause damage to the body, such as insomnia and poor postoperative recovery. It will also have an impact on the psychology, leading to anxiety and even depression. It is easy to develop into chronic pain, causing more serious health problems.

[0003] Eugenol (EUG), as a natural liquid small molecule drug, has a good analgesic effect and is currently used as an endodontic analgesic in endodontic capping agents available in Japan. Studies have shown that eugenol can relieve acute pain by regulating glutamate receptors and inhibiting tumor necrosis factor α (TNF-α), and can provide a safe and effective new formulation option for the clinical treatment of acute pain. Due to the poor chemical stability of eugenol, the obvious oral first-pass effect in the liver, low bioavailability, and adverse gastrointestinal reactions, most current studies have developed eugenol into transdermal preparations. However, the high lipophilicity of eugenol will cause it to remain in the lipid epidermis after transdermal administration and not be absorbed by the human body, and the addition of penetration enhancers or surfactants is prone to side effects such as skin allergies, which limits the formulation development and clinical application of eugenol.

[0004] Compared with traditional oral and transdermal drug delivery preparations, soluble microneedles are a simple, effective, and well-tolerated transdermal drug delivery preparation. They are painless, non-invasive, and can be administered autonomously. They do not require professional management, which improves patient compliance. Soluble microneedles can bypass the first-pass metabolism of the liver and deliver drugs directly to the dermis, thereby improving the effective absorption of drugs. In addition, the solid form of soluble microneedles improves the stability of eugenol, eliminating the need for low-temperature storage and transportation, and reducing cost requirements. However, the traditional solution casting method often uses organic reagents when preparing microneedle patches for poorly soluble drugs, and the complex process not only prolongs the preparation time of the microneedles, but also causes a large amount of eugenol loss during the preparation period. Therefore, in order to improve the stability, release, and diffusion properties of soluble microneedles of eugenol, it is necessary to introduce other technologies to solidify eugenol while improving the solubility of eugenol. Summary of the invention

[0005] One of the purposes of the present invention is to provide a eugenol soluble microneedle patch, which consists of a microneedle array and a backing; the microneedle array is made of an amphiphilic polymer, a water-soluble polymer and eugenol, the mass ratio of the amphiphilic polymer to the water-soluble polymer is 1:1 to 3:1, preferably 1:1 to 1.5:1; the ratio of the total mass of the amphiphilic polymer and the water-soluble polymer to the mass of eugenol is 1:1 to 6:1, preferably 4:1.

[0006] Furthermore, the amphiphilic polymer is Soluplus, copovidone, hydroxypropyl cellulose, hypromellose, hypromellose phthalate or hypromellose acetate succinate, preferably copovidone, hypromellose or hypromellose acetate succinate.

[0007] Furthermore, the water-soluble polymer is polyvinyl pyrrolidone, polyvinyl alcohol, hyaluronic acid or polyacrylamide, preferably polyvinyl pyrrolidone or polyvinyl alcohol.

[0008] In the present invention, the amphiphilic polymer and the water-soluble polymer have the following characteristics:

[0009] (1) It can provide good mechanical properties for the microneedle tip;

[0010] (2) Suitable for hot melt extrusion and hot pressing preparation processes;

[0011] (3) As a component of an amorphous solid dispersion, it can accelerate the release of eugenol;

[0012] (4) An in-situ self-emulsifying drug delivery system can be formed during the dissolution process of the amorphous solid dispersion.

[0013] In one embodiment of the present invention, the backing is made of copolyvidone VA64 (PVP VA64) and polyvinyl alcohol MXP (PVA MXP), and the mass ratio of PVP VA64 to PVAMXP is 1:1 to 4:1, preferably 2:1.

[0014] In the present invention, the microneedle backing has the following characteristics:

[0015] (1) It has good toughness and will not break when pressed;

[0016] (2) The microneedle backing material is suitable for the preparation process of hot melt extrusion and hot pressing;

[0017] (3) It has good adhesion and the microneedles will not loosen after insertion.

[0018] The second object of the present invention is to provide a method for preparing the above-mentioned eugenol soluble microneedle patch, comprising the following steps:

[0019] Step 1, mixing the raw material components of the microneedle array evenly for use, and mixing the raw material components of the backing evenly for use;

[0020] Step 2, hot-melt extruding the microneedle array premix, and crushing the extrudate for standby use;

[0021] Step 3, hot-melt extruding the backing premix, and crushing the extrudate for standby use;

[0022] Step 4, filling the microneedle array extrudate powder obtained in step 2 into a microneedle array mold, and performing hot pressing to form a microneedle array;

[0023] Step 5, spreading the backing extrudate powder obtained in step 3 on the microneedle array mold of step 4, performing hot pressing to make a backing, and peeling it off after cooling to obtain a eugenol microneedle patch.

[0024] Furthermore, the process parameters of hot melt extrusion in step 2 are: extrusion temperature 120-200° C., screw speed 20-40 rpm.

[0025] Furthermore, the process parameters of hot melt extrusion in step 3 are: extrusion temperature 120-200° C., screw speed 20-40 rpm.

[0026] Furthermore, the process parameters of hot pressing in step 4 are: temperature 160-200° C., pressure 0.4-1 MPa.

[0027] Furthermore, the process parameters of hot pressing in step 5 are: temperature 120-160° C., pressure 0.4-0.8 MPa.

[0028] The present invention combines soluble microneedles with a self-emulsifying drug delivery system, uses hot melt extrusion technology to prepare eugenol into a solid self-emulsifying drug delivery system in the form of an amorphous solid dispersion (the oil phase forming the emulsion is eugenol, the surfactant is an amphiphilic polymer, and the aqueous phase is a dissolution medium), and then uses hot pressing to prepare the eugenol solid self-emulsifying drug delivery system into soluble microneedles.

[0029] The inventors have found that the presence of polymers in the amorphous solid dispersion can solidify eugenol, thereby maintaining the stability of eugenol, reducing the volatilization and oxidation of eugenol, and enhancing the dissolution and release of eugenol. The contents of the eugenol amorphous solid dispersion and the microneedle tip were measured respectively, and the results showed that the actual content of eugenol was close to the theoretical content, proving that eugenol has good stability during hot melt extrusion and hot pressing. In addition, eugenol also reduces the glass transition temperature (T g ), thereby improving the processing performance of the amorphous solid dispersion system, reducing the extrusion temperature, and making the extrusion process easier to implement.

[0030] Furthermore, the inventors also found in the study that the addition of eugenol can mediate the formation of a self-emulsifying drug delivery system during the release process, thereby prolonging the supersaturation time of the drug, improving the diffusion properties of eugenol, maintaining a high gradient drug concentration difference between the human lymphatic and other absorption systems and the dermis, promoting the absorption of drugs into the systemic circulation by the human body, and enhancing the bioavailability of drugs.

[0031] The present invention uses eugenol as a drug and combines hot melt extrusion and hot pressing to successfully prepare an in-situ self-emulsifying eugenol soluble microneedle patch based on amorphous solid dispersion technology. The soluble microneedles improve the bioavailability of eugenol from three aspects. First, the advantage of amorphous solid dispersions in enhancing the dissolution and diffusion of drugs in the dermis is utilized; second, the advantage of the self-emulsifying drug delivery system formed during the dissolution process can promote the diffusion and absorption of drugs in the dermis and prolong the supersaturation of drugs; third, the advantage of soluble microneedles in avoiding oral administration and retention in the lipid epidermis is utilized.

[0032] Beneficial effects:

[0033] (1) The present invention combines hot melt extrusion and hot pressing, and no solvent needs to be added during the preparation process, which is in line with the concept of green environmental protection. Compared with the traditional microneedle preparation method, the present invention simplifies the microneedle preparation process and has the prospect of industrial production.

[0034] (2) The present invention combines amorphous solid dispersion technology with a self-emulsifying drug delivery system, thereby increasing the solubility of the drug, promoting the release and diffusion of the drug through the dermal body fluid, prolonging the drug supersaturation time, significantly improving the transdermal absorption performance of eugenol, and enhancing the analgesic effect of eugenol. BRIEF DESCRIPTION OF THE DRAWINGS

[0035] Figure 1 These are stereomicroscope images and scanning electron microscope images of the eugenol soluble microneedles in Example 1.

[0036] Figure 2 This is the DSC characterization diagram of the eugenol-soluble microneedle tip sample in Example 1 and the various components of the tip and the tip extrudate.

[0037] Figure 3 This is the result of the mechanical strength test of the eugenol soluble microneedles in Example 1.

[0038] Figure 4 This is the in vitro dissolution study result of the eugenol soluble microneedles in Example 1.

[0039] Figure 5 The curves showing the change of particle size over time of the eugenol-VA64 solid dispersion and the eugenol soluble microneedle system in Example 1 under PBS 7.4 at 37°C.

[0040] Figure 6 It is a transmission electron microscope (TEM) image of the emulsion droplets formed by the eugenol soluble microneedles in Example 1 after dissolution in PBS 7.4 medium for 1 hour, as well as the corresponding particle size distribution and average diameter.

[0041] Figure 7 The transdermal release curves of the eugenol soluble microneedles in Example 1 and the eugenol emulsion in Comparative Example 1. DETAILED DESCRIPTION

[0042] The technical scheme of the present invention is further explained below by specific examples. It should be understood that the following examples are provided only for the purpose of illustration and are not intended to limit the scope of the present invention. Those skilled in the art may make various modifications and substitutions to the present invention without departing from the purpose and spirit of the present invention.

[0043] In addition, for the numerical range in the present invention, it is understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. The intermediate value in any stated value or stated range, and each smaller range between any other stated value or intermediate value in the range is also included in the present invention. The upper and lower limits of these smaller ranges can be independently included or excluded in the scope.

[0044] Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art to which the invention pertains. Although only preferred methods and materials are described herein, any methods and materials similar or equivalent to those described herein may also be used in the practice or testing of the present invention.

[0045] The words “include,” “including,” “have,” “contain,” etc. used in this document are open-ended terms, meaning including but not limited to.

[0046] Unless otherwise specified, the experimental methods used in the following examples are conventional methods.

[0047] Unless otherwise specified, the materials and reagents used in the following examples can be obtained from commercial sources.

[0048] Example 1

[0049] The preparation method of the soluble microneedles containing eugenol in this embodiment comprises the following steps:

[0050] (1) PVP VA64 (44%), PVP K90 (36%) and eugenol (20%) were mixed evenly and used as a needle tip premix; PVP VA64 and PVAMXP were weighed and mixed evenly in a mass ratio of 2:1 and used as a backing premix;

[0051] (2) hot-melt extruding the needle tip premix in step (1), with the extrusion temperature of zone 01 being 40° C., the temperature of zones 02 to 04 being 160° C., and the screw speed being 40 rpm. The extruded product is collected on an aluminum plate, cooled at room temperature, ground in a mortar, and passed through a 60-mesh sieve, and the powder under the sieve is taken for later use;

[0052] (3) The backing premix in step (1) is subjected to hot melt extrusion, the extrusion temperature of zone 01 is 40° C., the temperature of zones 02 to 04 is 180° C., and the screw speed is 40 rpm. The extruded product is collected on an aluminum plate, cooled at room temperature, ground in a mortar, and passed through a 60-mesh sieve, and the powder under the sieve is taken for later use;

[0053] (4) using a hot pressing method to evenly fill the sieved material in step (2) into the cavity array mold and retain it for 10 minutes, wherein the hot pressing temperature is 160° C. and the pressure is 0.6 MPa, then taking out the mold and removing the excess melt on the mold surface and cooling it;

[0054] (5) Using a hot pressing method, the sieved material in step (3) is evenly spread on the surface of the array mold and retained for 10 minutes, and the microneedle patch array mold is removed and cooled at room temperature and then peeled off to obtain a microneedle patch.

[0055] Example 2

[0056] The type and amount of water-soluble polymer in the needle tip component were investigated.

[0057] The preparation method of the soluble microneedles containing eugenol in this embodiment comprises the following steps:

[0058] (1) PVP VA64 (44%), PVA MXP (36%) and eugenol (20%) were mixed evenly and used as needle tip premix 1; PVP VA64 (32%), PVP K90 (48%) and eugenol (20%) were mixed evenly and used as needle tip premix 2; PVP VA64 and PVA MXP were weighed and mixed evenly in a mass ratio of 2:1 and used as backing premix;

[0059] (2) hot-melt extruding the needle tip premix in step (1), the extrusion temperature of zone 01 is 40° C., the temperatures of zones 02 to 04 are 160° C. and 190° C., respectively, and the screw speed is 40 rpm. The extruded product is collected on an aluminum plate, cooled at room temperature, ground in a mortar, and passed through a 60-mesh sieve, and the powder under the sieve is taken for later use;

[0060] (3) The material in step (1) is subjected to hot melt extrusion, the extrusion temperature of zone 01 is 40° C., the temperature of zones 02 to 04 is 180° C., and the screw speed is 40 rpm. The extruded product is collected on an aluminum plate, cooled at room temperature, ground in a mortar, and passed through a 60-mesh sieve, and the powder under the sieve is taken for later use;

[0061] (4) using a hot pressing method to evenly fill the two batches of materials screened in step (2) into the cavity array mold and retain them for 10 minutes, wherein the hot pressing temperatures are 160° C. and 200° C., and the pressures are 0.6 MPa and 1.0 MPa, respectively. Then, the mold is taken out and excess melt on the mold surface is removed and then cooled;

[0062] (5) The sieved material in step (3) is evenly spread on the surface of the array mold by hot pressing and retained for 10 minutes. The microneedle patch array mold is removed and cooled at room temperature and then peeled off to obtain two batches of microneedle patches.

[0063] Example 3

[0064] Investigation of the amount of drug in the needle tip component

[0065] The preparation method of the soluble microneedles containing eugenol in this embodiment comprises the following steps:

[0066] (1) PVP VA64 (38.5%), PVP K90 (31.5%) and eugenol (30%) were mixed evenly and used as a needle tip premix; PVP VA64 and PVA MXP were weighed and mixed evenly in a mass ratio of 2:1 and used as a backing premix;

[0067] (2) hot-melt extruding the needle tip premix in step (1), with the extrusion temperature of zone 01 being 40° C., the temperature of zones 02 to 04 being 160° C., and the screw speed being 40 rpm. The extruded product is collected on an aluminum plate, cooled at room temperature, ground in a mortar, and passed through a 60-mesh sieve, and the powder under the sieve is taken for later use;

[0068] (3) The backing premix in step (1) is subjected to hot melt extrusion, the extrusion temperature of zone 01 is 40° C., the temperature of zones 02 to 04 is 180° C., and the screw speed is 40 rpm. The extruded product is collected on an aluminum plate, cooled at room temperature, ground in a mortar, and passed through a 60-mesh sieve, and the powder under the sieve is taken for later use;

[0069] (4) using a hot pressing method to evenly fill the sieved material in step (2) into the cavity array mold and retain it for 10 minutes, wherein the hot pressing temperature is 160° C. and the pressure is 0.6 MPa, then taking out the mold and removing the excess melt on the mold surface and cooling it;

[0070] (5) Using a hot pressing method, the sieved material in step (3) is evenly spread on the surface of the array mold and retained for 10 minutes, and the microneedle patch array mold is removed and cooled at room temperature and then peeled off to obtain a microneedle patch.

[0071] Comparative Example 1

[0072] The preparation method of the eugenol emulsion of this comparative example comprises the following steps:

[0073] Accurately weigh 25 mg of eugenol, 55 mg of PVP VA64, and 45 mg of PVP K90 respectively, place them in 20 mL of pure water, and vortex for 10 minutes.

[0074] Comparative Example 2

[0075] The preparation method of the indomethacin solution of this comparative example comprises the following steps:

[0076] Accurately weigh 20 mg of indomethacin and place it in 20 mL of pure water. Vortex for 10 minutes.

[0077] Comparative Example 3

[0078] The preparation method of the eugenol-PVP K90 solid dispersion of this comparative example comprises the following steps:

[0079] 2g of eugenol and 8g of PVP K90 were accurately weighed respectively, stirred and mixed, and then put into a hot melt extruder. The extrusion temperature of zone 01 was 40°C, the temperatures of zones 02 to 04 were 190°C respectively, and the screw speed was 40rpm. The extruded product was collected on an aluminum plate, cooled at room temperature, ground in a mortar, and passed through a 60-mesh sieve to obtain the powder under the sieve.

[0080] Comparative Example 4

[0081] The preparation method of the eugenol-PVP VA64 solid dispersion of this comparative example comprises the following steps:

[0082] 2g of eugenol and 8g of PVP VA64 were accurately weighed respectively, stirred and mixed, and then put into a hot melt extruder. The extrusion temperature of zone 01 was 40°C, the temperatures of zones 02 to 04 were 160°C respectively, and the screw speed was 40rpm. The extruded product was collected on an aluminum plate, cooled at room temperature, ground in a mortar, and passed through a 60-mesh sieve to obtain the powder under the sieve.

[0083] The morphology and mechanical properties of the eugenol soluble microneedles of the above-mentioned Examples 1 to 3 were analyzed, and the results showed that the eugenol soluble microneedles prepared using needle tip premix 1 in Example 2 had poor mechanical properties and could not puncture the skin of KM mice; the eugenol soluble microneedles prepared using needle tip premix 2 in Example 2 were difficult to form during the preparation process; the eugenol soluble microneedles prepared in Example 3 had strong hygroscopicity, were difficult to preserve, and had poor formability; while the eugenol soluble microneedles prepared in Example 1 had good mechanical properties and good microneedle formability, and could be used for subsequent investigation experiments on eugenol soluble microneedles.

[0084] The samples prepared in the above embodiments and comparative examples are tested below.

[0085] 1. Morphology observation

[0086] Instruments: AXio200mV.16 stereo microscope (Zeiss, Germany), VEGA3 scanning electron microscope (TESCAN, Czech)

[0087] Light source for stereo microscope: normal light

[0088] Accelerating voltage of scanning electron microscope: 20 kV

[0089] The stereomicroscope and scanning electron microscope images of the eugenol soluble microneedles in Example 1 are as follows: Figure 1 As shown in the figure, the microneedle array is complete (20×20), the needle surface is flat, and it is a quadrangular pyramid with no obvious breakage or bending.

[0090] 2. Differential scanning calorimetry (DSC)

[0091] Instrument: DSC 3500 differential scanning calorimeter (Netzsch, Germany)

[0092] Sample plate type: solid plate

[0093] Temperature range: 20-200℃

[0094] Heating rate: 10℃ / min

[0095] The DSC detection spectra of the eugenol soluble microneedle tip sample, the tip components, and the tip extrudate in Example 1 are as follows: Figure 2 The DSC spectra of the prepared needle tip extrudate and microneedle tip sample both have a single T g , indicating that the needle tip extrudate remains in the form of a single homogeneous amorphous solid dispersion before and after hot pressing, that is, the eugenol molecules are uniformly dispersed in the PVP VA64-PVP K90 mixed polymer matrix.

[0096] 3. Mechanical strength inspection

[0097] After male KM mice were killed by cervical dislocation, the back hair of the mice was immediately shaved with a rechargeable animal depilatory machine, and then depilatory cream was applied and wiped clean with saline. After complete depilation, the back skin was cut immediately and the removed skin was laid flat on a clean table with the stratum corneum facing up. The eugenol-soluble microneedles in Example 1 were removed by pressing with the thumb for 30 seconds, and finally the pinhole array area was immediately stained with 0.5% methylene blue dye for 5 minutes, and the residual solution on the skin was wiped with an alcohol cotton pad. The morphology of the pinholes left by the microneedles on the skin was observed using an AXio200mV.16 stereo microscope (Zeiss, German).

[0098] The mechanical strength test results of the eugenol soluble microneedles in Example 1 are as follows: Figure 3 Compared with before puncture ( Figure 3 Middle a), on the back skin of KM mice in vitro, the microneedles formed a dot-like distribution consistent with their array (20×20, Figure 3 (b) The dyed skin also shows neatly arranged and regular blue dots ( Figure 3 Middle c), indicating that the microneedle has good mechanical strength and can pierce the skin smoothly.

[0099] 4. In vitro dissolution study

[0100] Instrument: RC806ADK intelligent dissolution instrument (TianDa TianFa, China)

[0101] Method: According to the second method (paddle method) of dissolution and release determination in the 2020 edition of the Chinese Pharmacopoeia, general chapter 0931, small cup method. Dissolution medium: 20 mL PBS7.4

[0102] Temperature: 37±0.5℃

[0103] Speed: 0rpm

[0104] Test solution: The cut foam block is adhered to the eugenol soluble microneedle backing in Example 1, and the buoyancy force is used to ensure that the tip of the microneedle is facing the dissolution medium. 1 mL of sample is taken at predetermined time intervals (3, 5, 10, 15, 20, 30, 45, 60, 120 and 180 min), and then an equal amount of fresh dissolution medium is added isothermally. The sample is filtered with a 0.45 μm polyethersulfone needle filter (25 mm × 0.45 μm, Shanghai Anpu Laboratory), and the eugenol concentration is analyzed by high performance liquid chromatography (HPLC). Each sample is measured in parallel 3 times.

[0105] Eugenol reference solution: Take an appropriate amount of eugenol reference, weigh accurately, and add methanol to prepare a solution containing 40 μg of eugenol per 1 mL.

[0106] Eugenol was determined by HPLC using a chromatographic column of C18, 4.6*250mm, 5μm; the mobile phase is methanol-purified water, the ratio of organic phase to aqueous phase is 72:28, the flow rate is 1mL / min, the column temperature is 30℃, the detection wavelength is 280nm, and the injection volume is 10μL. The above solution is placed in a high performance liquid chromatograph for peak area determination, thereby calculating its solubility.

[0107] Stereoscopic microscopic observation: In the same manner as the above-mentioned test solution, the eugenol-soluble microneedles of the four Examples 1 were respectively placed in a dissolution cup, and the microneedles were taken out at 10, 30, 60 and 180 min, respectively, and the dissolution of the microneedle tip was observed using an AXio200mV.16 stereo microscope (Zeiss, German).

[0108] The release rate of eugenol from the eugenol soluble microneedles in Example 1 was evaluated by in vitro dissolution study. The results are as follows: Figure 4 As shown in Figure a, the release amount of the microneedles in PBS 7.4 medium reached 35% after 10 minutes, and quickly released to 90% after 30 minutes, and then basically released completely (95%) after 60 minutes. At the same time, a stereo microscope was used to observe the dissolution of the microneedles at different release times. The results are shown in Figure 5. Figure 4 As shown in b, the microneedle partially dissolved 10 minutes after piercing the mouse skin, only a small amount of microneedle remained after 30 minutes, and only a flat backing was observed after 1 hour and 3 hours, which is consistent with the release rate curve, that is, the tip of the microneedle almost completely dissolved 1 hour after piercing the mouse skin. The DSC results show that the tip of the microneedle is an amorphous solid dispersion, and the dispersion effect of the polymer carrier can significantly increase the specific surface area of ​​eugenol, thereby effectively improving the solubility of eugenol.

[0109] 5. Confirmation of Self-emulsifying Drug Delivery System (SEDDS)

[0110] (1) Dynamic light scattering (DLS)

[0111] Instrument: NanoBrook Omni dynamic light scattering particle size analyzer (Brookhaven, America)

[0112] Under the same conditions as the above-mentioned in vitro dissolution study, 25 mg of the eugenol-PVP K90 solid dispersion powder in Comparative Example 3, 25 mg of the eugenol-PVP VA64 solid dispersion powder in Comparative Example 4, and a piece of eugenol soluble microneedle in Example 1 were respectively put into three dissolution cups. The dissolution medium was PBS 7.4, the medium volume was 20 mL, and the medium temperature was 37°C. 2 mL of samples were taken at 10, 30, 60 and 180 min, respectively, and then an equal amount of fresh dissolution medium was added at the same temperature. The sample was placed in a quartz fluorescent dish at 90° for testing, and the measurement was performed three times (1.5 min / time) to obtain the average value of the effective particle size, and the PDI value was recorded. The DLS measurement results are shown as follows. Figure 5 As shown in Table 1.

[0113] Table 1 PDI values ​​of eugenol-VA64 solid dispersion and the eugenol soluble microneedle system of Example 1 at different time points under PBS 7.4 at 37°C (n=3, mean ± SD)

[0114]

[0115] The eugenol-PVP K90 solid dispersion failed to measure effective particle size data due to its large and unstable particle size; the eugenol-PVP VA64 solid dispersion had a stable particle size within 3 hours, maintained in the range of 75-83 nm (the PDI at different sampling time points was less than 0.3). The eugenol soluble microneedles in Example 1 also had a stable particle size within 3 hours, maintained in the range of 65-71 nm (the PDI at different sampling time points was less than 0.3), showing a particle size range smaller than that of the eugenol-PVP VA64 solid dispersion system.

[0116] (2) Transmission electron microscopy (TEM) observation

[0117] Instrument: HT7700 transmission electron microscope (Hitachi, Japan)

[0118] The morphology of the emulsion droplets formed by the eugenol soluble microneedles in Example 1 during the in vitro dissolution release was observed by TEM. The accelerating voltage was 120 kV. Under the same conditions as the above in vitro dissolution study, 1 mL of the dissolution sample of the microneedles was collected at 1 hour. The TEM observation results are shown in Figure 2. Figure 6 The values ​​of the long and short diameters of the emulsion droplets are shown in Table 2.

[0119] Table 2 Values ​​of the long and short diameters of the emulsion droplets formed by the eugenol soluble microneedles in Example 1 in the TEM image

[0120]

[0121]

[0122] The long and short diameters of the emulsion droplets formed by the eugenol soluble microneedles in Example 1 are roughly the same, from which it can be inferred that the three-dimensional structure of the emulsion droplets is approximately spherical. The particle size data in Table 2 were further Gaussian fitted using Origin 2019b software (e.g. Figure 6 As shown in (c), the self-emulsifying system has a good particle size distribution, and the average particle size is 70.1±17.1 nm, which is basically consistent with the DLS measurement result.

[0123] 6. In vitro transdermal test

[0124] The in vitro transdermal performance of the eugenol soluble microneedles in Example 1 and the eugenol emulsion in Comparative Example 1 was studied using a SYT-103A1 multifunctional transdermal diffusion instrument (Yanji Aidi, China) to evaluate the skin permeability of the microneedles.

[0125] Microneedle group: A piece of eugenol soluble microneedle in Example 1 was cut into four small pieces for in vitro transdermal experiment investigation. The microneedle was cut into 4 parts of 10×10 arrays and inserted vertically into fresh skin respectively. The epidermis was then fixed between the receiving chamber and the supply chamber of the diffusion cell. At different intervals, 200 μL aliquots were taken out from the four diffusion cells respectively. Then the four aliquots taken out at the same time were collected in a tube and mixed evenly with 800 μL of methanol. An equal amount of PBS7.4 was supplemented. Finally, the amount of the drug was determined by the HPLC method in the above-mentioned in vitro dissolution investigation.

[0126] Control group: The selected skin piece not treated with microneedles was mounted on the diffusion cell with the epidermis facing upward. 1 mL of the eugenol emulsion in Comparative Example 1 was added to the donor chamber, the sample was treated with the microneedle permeation solution treatment method, and analyzed using the HPLC method in the in vitro dissolution study described above.

[0127] The results of in vitro transdermal permeation experiments were as follows Figure 7 As shown. In the first 30 minutes, the cumulative permeability of the eugenol soluble microneedles in Example 1 can reach 74%, and it has reached the permeation platform (84%) in 60 minutes, and maintains this permeability until the end of the study (3 hours). However, the cumulative permeability of the eugenol emulsion in Comparative Example 1 is only 33% at 3 hours. It can be seen that microneedles can effectively improve the in vitro transdermal performance of eugenol.

[0128] 7. Analgesic efficacy experiment

[0129] The analgesic efficacy of the microneedles was evaluated by observing the inhibitory effects of the eugenol emulsion in Comparative Example 1, the indomethacin solution in Comparative Example 2, and the eugenol soluble microneedles in Example 1 on the writhing reaction of mice induced by glacial acetic acid and the plantar reaction of mice induced by formalin.

[0130] In the acetic acid writhing test, the KM mice in the control group were intraperitoneally injected with acetic acid solution (0.6%, 10 mL / kg), and the number of writhing times of the mice within 30 minutes was observed and recorded. In the formalin plantar test, formalin (2.5%, 20 μL) was subcutaneously injected into the plantar surface of the left hind paw of the KM mice in the control group, and the total time that the animals licked or bit the injected paw in two stages (0-5 min in the first stage and 15-30 min in the second stage) was measured using a timer.

[0131] The KM mice in the eugenol emulsion group in the comparative example 1 and the eugenol soluble microneedle group in the embodiment 1 were subjected to back hair removal, that is, the hair on the back of the mice was shaved clean with a rechargeable animal hair removal machine, and then a depilatory cream was applied and wiped clean with saline. The prepared microneedles were pressed on the back of the mice with the thumb for 30 seconds and then released, and then the microneedles were fixed to the skin with medical tape. 1 mL of the eugenol emulsion in the comparative example 1 was transferred and applied to the back of the mice. The KM mice in the indomethacin group were gavaged with indomethacin suspension (10 mg / kg).

[0132] The results of the acetic acid writhing and formalin plantar reaction tests are shown in Tables 3 and 4, respectively.

[0133] Table 3 Results of acetic acid writhing test (n=3, mean ± SD)

[0134]

[0135] Table 4 Formalin plantar reaction test results (n = 3, mean ± SD value)

[0136]

[0137] Compared with the eugenol emulsion in Comparative Example 1 and the indomethacin solution in Comparative Example 2, the analgesic rate of the eugenol soluble microneedles in Example 1 was significantly improved, indicating that it has a good analgesic effect and has certain advantages over traditional oral and transdermal preparations.

Claims

1. A eugenol soluble microneedle patch, comprising a microneedle array and a backing, characterized in that: The microneedle array is made of an amphiphilic polymer, a water-soluble polymer and eugenol, the mass ratio of the amphiphilic polymer to the water-soluble polymer is 1:1-3:1; the ratio of the total mass of the amphiphilic polymer and the water-soluble polymer to the mass of eugenol is 1:1-6:

1.

2. The eugenol soluble microneedle patch according to claim 1, characterized in that: The amphiphilic polymer is Soluplus, copovidone, hydroxypropyl cellulose, hypromellose, hypromellose phthalate or hypromellose acetate succinate.

3. The eugenol soluble microneedle patch according to claim 1, characterized in that: The water-soluble polymer is polyvinyl pyrrolidone, polyvinyl alcohol, hyaluronic acid or polyacrylamide.

4. The method for preparing the eugenol soluble microneedle patch according to claim 1, characterized in that: The following steps are involved: Step 1, mixing the raw material components of the microneedle array evenly for use, and mixing the raw material components of the backing evenly for use; Step 2, hot-melt extruding the microneedle array premix, and crushing the extrudate for standby use; Step 3, hot-melt extruding the backing premix, and crushing the extrudate for standby use; Step 4, filling the microneedle array extrudate powder obtained in step 2 into a microneedle array mold, and performing hot pressing to form a microneedle array; Step 5, spreading the backing extrudate powder obtained in step 3 on the microneedle array mold of step 4, performing hot pressing to make a backing, and peeling it off after cooling to obtain a eugenol microneedle patch.

5. The preparation method according to claim 4, characterized in that: The process parameters of hot melt extrusion in step 2 are: extrusion temperature 120~200℃, screw speed 20~40rpm.

6. The preparation method according to claim 4, characterized in that: The process parameters of hot melt extrusion in step 3 are: extrusion temperature 120~200℃, screw speed 20~40rpm.

7. The preparation method according to claim 4, characterized in that: The process parameters of hot pressing in step 4 are: temperature 160~200℃, pressure 0.4~1MPa.

8. The preparation method according to claim 4, characterized in that: The process parameters of hot pressing in step 5 are: temperature 120~160℃, pressure 0.4~0.8MPa.

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