阅读说明：本技术 微针阵列及微针阵列的制造方法 (Microneedle array and method for producing microneedle array ) 是由 阪井正树 柿田浩辅 中务阳裕 森久容 于 2020-02-26 设计创作，主要内容包括：本发明的课题在于提供一种流感疫苗的稳定性良好,且流感疫苗的利用效率高的微针阵列及其制造方法。根据本发明,提供一种微针阵列,其具有片材部和存在于片材部上表面的多个针部,其为,针部含有水溶性高分子、流感疫苗及胍胺或其盐,通过针部溶解,将流感疫苗给药到体内的自溶型微针阵列。(The present invention addresses the problem of providing a microneedle array having good stability of an influenza vaccine and high utilization efficiency of the influenza vaccine, and a method for producing the microneedle array. According to the present invention, there is provided an autolytic microneedle array comprising a sheet portion and a plurality of needle portions present on an upper surface of the sheet portion, wherein the needle portions contain a water-soluble polymer, an influenza vaccine, and guanamine or a salt thereof, and the influenza vaccine is administered into the body by dissolution of the needle portions.)

1. A microneedle array having a sheet portion and a plurality of needle portions present on the upper surface of the sheet portion, which is an autolytic microneedle array for administering an influenza vaccine into the body by dissolving the needle portions, wherein the needle portions contain a water-soluble polymer, an influenza vaccine, and guanamine or a salt thereof.

2. The microneedle array of claim 1,

the content of the water-soluble polymer contained in the needle portion is 10 mass% or more and 99 mass% or less with respect to the solid content of the needle portion.

3. The microneedle array according to claim 1 or 2,

the content of guanamine or a salt thereof in the needle portion is 0.01 to 300 times relative to the content of the influenza vaccine.

4. The microneedle array according to any one of claims 1 to 3,

the needle portion contains a saccharide.

5. The method of manufacturing the microneedle array of any one of claims 1 to 4, comprising:

concentrating the influenza vaccine;

a step of forming a needle portion using the concentrated influenza vaccine obtained in the step; and

and forming the sheet portion.

6. The method of manufacturing a microneedle array according to claim 5,

the step of concentrating the influenza vaccine is a step of concentrating the influenza vaccine by centrifugation.

Technical Field

The present invention relates to a microneedle array and a method of manufacturing the same. The invention particularly relates to a microneedle array containing an influenza vaccine and a manufacturing method thereof.

Background

In recent years, a dissolving-type microneedle array has been developed in which a drug is contained in a base material made of a substance soluble in the living body. Since the needles of the microneedle array are thin and short, the stimulation to nerves is small. Thus, microneedle arrays are also referred to as "painless injections.

Patent document 1 describes a method for manufacturing a percutaneous delivery device, which includes: (i) providing a minute projection member having a plurality of minute projections; (ii) providing a biocompatible film-coated dosage form containing a bioactive agent; (iii) coating the microprotrusion member with a biocompatible coating agent to form a percutaneous delivery device; (iv) and packaging the transdermal delivery device under dry inert atmosphere conditions and/or partial vacuum. Patent document 1 describes that guanamine can be added as a counter ion for a biological agent. The microneedle array described in patent document 1 is not an autolytic microneedle array in which a needle portion is dissolved and a drug is administered into the body. On the other hand, patent document 2 describes a drug for stabilizing a protein containing guanamine.

Documents of the prior art

Patent document

Patent document 1: japanese Kohyo publication No. 2009-522288

Patent document 2: international publication WO08/132363

Disclosure of Invention

Technical problem to be solved by the invention

Influenza vaccines have been administered by subcutaneous and intramuscular injection. In this case, fear of the injection needle, pain at the time of injection, mental stress, and the like become problems. In order to solve these problems, microneedle array-based drug delivery has been proposed as a painless method. In particular, a microneedle array containing a vaccine is expected to be improved in effectiveness, and a microneedle array containing an influenza vaccine is expected to be present.

Since the microneedle array is a minute preparation, the volume containing the drug is small, and the drug needs to be contained at a higher concentration than an injection. Therefore, the distance between the drugs is reduced, and aggregation or reaction is likely to occur, which may deteriorate the stability of the drugs. The inventors of the present invention also obtained the result that the stability of the influenza vaccine was impaired in the microneedle array containing the influenza vaccine studied compared to the injection, and it was desired to improve the stability of the influenza vaccine.

Also, the drug contained in the microneedle array wastes a large amount of the drug if there is a portion that is not administered into the living body. Thus, the microneedle array requires the drug to be concentrated at the leading end portion of the needle.

The present invention addresses the problem of providing a microneedle array having good stability of an influenza vaccine and high utilization efficiency of the influenza vaccine, and a method for producing the microneedle array.

Means for solving the technical problem

As a result of intensive studies to solve the above problems, the present inventors have found that the stability of an influenza vaccine is improved over time and the tip filling performance of the influenza vaccine is improved by adding guanamine or a salt thereof to a needle portion containing the influenza vaccine in a microneedle array containing the influenza vaccine. The present invention has been completed based on the above findings.

That is, according to the present invention, the following inventions are provided.

(1) A microneedle array of the autolytic type having a sheet portion and a plurality of needle portions present on the upper surface of the sheet portion, wherein the needle portions contain a water-soluble polymer, an influenza vaccine, and guanamine or a salt thereof, and the influenza vaccine is administered into the body by dissolving the needle portions.

(2) The microneedle array according to (1), wherein a content of the water-soluble polymer contained in the needle portion is 10 mass% or more and 99 mass% or less with respect to a solid content of the needle portion.

(3) The microneedle array according to (1) or (2), wherein the content of guanamine or a salt thereof is 0.01 to 300 times that of an influenza vaccine.

(4) The microneedle array according to any one of (1) to (3), wherein the needle portion contains a saccharide.

(5) The method for manufacturing a microneedle array according to any one of (1) to (4), comprising:

concentrating the influenza vaccine;

forming a needle portion using the concentrated influenza vaccine obtained in the above step; and

and forming the sheet portion.

(6) The method of manufacturing a microneedle array according to (5), wherein the step of concentrating the influenza vaccine is a step of concentrating the influenza vaccine by centrifugation.

Effects of the invention

In the microneedle array containing the influenza vaccine, the stability of the influenza vaccine is good, and the utilization efficiency of the influenza vaccine is high.

Drawings

Fig. 1 shows a region from the tip of a microneedle to 600 μm and a region from the tip of a microneedle to 800 μm.

In fig. 2, fig. 2A is a perspective view of a conical microneedle, fig. 2B is a perspective view of a pyramidal microneedle, and fig. 2C is a cross-sectional view of a conical microneedle and a pyramidal microneedle.

Fig. 3 is a perspective view of a microneedle of another shape.

Fig. 4 is a perspective view of a microneedle of another shape.

Fig. 5 is a sectional view of the microneedle shown in fig. 3 and 4.

Fig. 6 is a perspective view of a microneedle of another shape.

Fig. 7 is a perspective view of a microneedle of another shape.

Fig. 8 is a sectional view of the microneedle shown in fig. 6 and 7.

Fig. 9 is a cross-sectional view of another microneedle shape in which the inclination (angle) of the side surface of the needle portion changes continuously.

In fig. 10, fig. 10A to C are process diagrams of a method of manufacturing a mold.

Fig. 11 is an enlarged view of the mold.

Fig. 12 is a sectional view showing another embodiment.

In fig. 13, fig. 13A to C are schematic views of a step of filling a liquid containing an influenza vaccine into a mold.

Fig. 14 is a perspective view showing the tip of the nozzle.

Fig. 15 is a partially enlarged view of the front end of the nozzle and the mold in filling.

Fig. 16 is a partial enlarged view of the moving nozzle tip and mold.

In fig. 17, fig. 17A to D are explanatory views showing another microneedle array formation step.

Fig. 18A to C are explanatory views showing another microneedle array formation step.

Fig. 19 is an explanatory view showing a peeling step.

Fig. 20 is an explanatory view showing another peeling step.

Fig. 21 is an explanatory view showing a microneedle array.

Fig. 22 (a) and (B) are a plan view and a side view of the master.

Fig. 23 is a schematic view of a filling device used in the examples.

Detailed Description

Hereinafter, embodiments of the present invention will be described in detail.

In the present specification, "containing a drug" means that the drug is contained in an amount that exerts a drug effect when the drug penetrates a body surface. "drug-free" means that the drug is not contained in an amount that exerts its pharmacological effects, and the range of the drug amount includes a range from the case where the drug is not contained at all to an amount that does not exert its pharmacological effects.

In the microneedle array of the present invention, the stability of the influenza vaccine can be improved by including guanamine in the needle portion, and the guanamine can be locally present at the tip of the needle portion. The effect that cannot be expected in the prior art is that the stability of the influenza vaccine can be improved by including guanamine in the needle part, and the guanamine is locally present at the tip of the needle part. The microneedle array of the present invention is not intended to use the coating type described in patent document 1. That is, the microneedle array of the present invention is an autolytic type in which a drug is administered into the body by dissolving the microneedle portion.

[ Structure of microneedle array ]

The microneedle array of the present invention is an autolyzed microneedle array having a sheet portion and a plurality of needle portions present on the upper surface of the sheet portion, wherein the needle portions contain an influenza vaccine, a water-soluble polymer, and guanamine or a salt thereof, and the drug, i.e., the influenza vaccine, is administered into the body by dissolving the needle portions.

In the present invention, "a plurality" means one or more.

In order to effectively administer a drug into the skin, the microneedle array of the present invention includes at least a sheet portion and a needle portion, and loads the needle portion with the drug.

The microneedle array of the present invention is a device in which a plurality of needle portions are arranged in an array on the upper surface side of a sheet portion. The needle portion is preferably disposed on the upper surface side of the sheet portion. The needle portion may be disposed directly on the upper surface of the sheet portion, or the needle portion may be disposed on the upper surface of a tapered portion disposed on the upper surface of the sheet portion.

The sheet portion is a base for supporting the needle portion, and has a planar shape like the sheet portion 116 shown in fig. 2 to 9. In this case, the upper surface of the sheet portion is a surface on which a plurality of needle portions are arranged in an array.

The area of the sheet portion is not particularly limited, but is preferably 0.005 to 1000mm²More preferably 0.1 to 800mm²More preferably 1 to 800mm²。

The thickness of the sheet portion is represented by the distance between the frustum portion or the surface in contact with the needle portion and the opposite surface. The thickness of the sheet portion is preferably 1 μm or more and 2000 μm or less, more preferably 3 μm or more and 1500 μm or less, and still more preferably 5 μm or more and 1000 μm or less.

The sheet portion preferably contains a water-soluble polymer. The sheet portion may be made of a water-soluble polymer, or may contain an additive (e.g., a disaccharide) other than the water-soluble polymer. In addition, the sheet portion preferably does not contain a drug.

The water-soluble polymer contained in the sheet portion is not particularly limited, and examples thereof include polysaccharides (e.g., hyaluronic acid, sodium hyaluronate, pullulan, dextran, dextrin, sodium chondroitin sulfate, carboxymethyl cellulose, hydroxypropyl cellulose, hydroxyethyl starch, hydroxypropyl methyl cellulose, polyvinylpyrrolidone, polyoxyethylene polyoxypropylene glycol, polyethylene glycol, and gum arabic), and proteins (e.g., gelatin). The above-mentioned components may be used singly or as a mixture of two or more kinds. Among the above, polysaccharides are preferable, hydroxyethyl starch, hydroxypropyl cellulose, hydroxypropyl methylcellulose, pullulan, dextran, sodium chondroitin sulfate, sodium hyaluronate, carboxymethyl cellulose, polyvinylpyrrolidone, polyoxyethylene polyoxypropylene glycol, polyethylene glycol, and polyvinyl alcohol are more preferable, and chondroitin sulfate and dextran are particularly preferable.

A disaccharide may be added to the sheet portion, and examples of the disaccharide include sucrose, lactulose, lactose, maltose, trehalose, cellobiose, and the like, and sucrose, maltose, and trehalose are particularly preferable.

The microneedle array is composed of a plurality of needle portions arranged in an array on the upper surface side of the sheet portion. The needle portion is a convex structure having a tip, and is not limited to a needle shape having a sharp tip, and may be a shape having a non-sharp tip.

Examples of the shape of the needle portion include a conical shape, a polygonal pyramid shape (quadrangular pyramid shape, etc.), a spindle shape, and the like. For example, the needle portion may have a shape like the needle portion 112 shown in fig. 2 to 9, the entire shape of the needle portion may be a conical shape or a polygonal cone shape (quadrangular pyramid shape or the like), or the inclination (angle) of the side surface of the needle portion may be continuously changed. Further, a multilayer structure of two or more layers in which the inclination (angle) of the side surface of the needle portion discontinuously changes may be adopted.

When the microneedle array of the present invention is applied to the skin, the needle portion is preferably inserted into the skin so that the upper surface of the sheet portion or a part thereof is in contact with the skin.

The height (length) of the needle portion is represented by the length of a perpendicular line that hangs from the tip of the needle portion to the tapered portion or the sheet portion (in the case where the tapered portion is not present). The height (length) of the needle portion is not particularly limited, but is preferably 50 μm or more and 3000 μm or less, more preferably 100 μm or more and 1500 μm or less, and further preferably 100 μm or more and 1000 μm or less. It is preferable that the length of the needle portion is 50 μm or more because transdermal administration of the drug is possible, and that the length of the needle portion is 3000 μm or less because pain due to contact of the needle portion with the nerve can be prevented and bleeding can be avoided.

The interface between the tapered portion (i.e., the needle portion when no tapered portion is present) and the sheet portion is referred to as a base portion. The distance between the farthest points in the base portion of one needle portion is preferably 50 μm or more and 2000 μm or less, more preferably 100 μm or more and 1500 μm or less, and still more preferably 100 μm or more and 1000 μm or less.

The number of the needle portions is preferably 1 to 2000, more preferably 3 to 1000, and further preferably 5 to 500 per microneedle array. In the case where each microneedle array contains 2 needle portions, the intervals of the needle portions are represented by the distances between the legs of the perpendicular lines hanging from the leading ends of the needle portions toward the frustum portion or the sheet portion (in the case where the frustum portion is not present). When each microneedle includes 3 or more needle portions, the distance between the legs of the perpendicular lines drawn from the tip end of each needle portion to the frustum portion or the sheet portion (in the case where the frustum portion is not present) with respect to the nearest needle portion is obtained for the interval of the arranged needle portions, and is represented by the average value thereof. The interval between the needle portions is preferably 0.1mm to 10mm, more preferably 0.2mm to 5mm, and still more preferably 0.3mm to 3 mm.

The needle portion contains an influenza vaccine, a water-soluble polymer, and guanamine or a salt thereof, and the water-soluble polymer is preferably a biosoluble substance so that the needle portion does not affect the human body even if left in the skin.

The water-soluble polymer contained in the needle portion is not particularly limited, and examples thereof include polysaccharides (e.g., hyaluronic acid, sodium hyaluronate, pullulan, dextran, dextrin, sodium chondroitin sulfate, carboxymethyl cellulose, hydroxypropyl cellulose, hydroxyethyl starch, hydroxypropyl methyl cellulose, polyvinylpyrrolidone, polyoxyethylene polyoxypropylene glycol, polyethylene glycol, and gum arabic), and proteins (e.g., gelatin). The above-mentioned components may be used singly or as a mixture of two or more kinds. Among the above, polysaccharides are preferable, hydroxyethyl starch, hydroxypropyl cellulose, hydroxypropyl methylcellulose, pullulan, dextran, sodium chondroitin sulfate, sodium hyaluronate, carboxymethyl cellulose, polyvinylpyrrolidone, polyoxyethylene polyoxypropylene glycol, polyethylene glycol, and polyvinyl alcohol are more preferable, and hydroxyethyl starch and dextran are particularly preferable. In addition, in order to prevent aggregation when mixed with a drug, uncharged polysaccharides are generally more preferable. The water-soluble polymer contained in the needle portion may be the same as or different from the water-soluble polymer contained in the sheet portion.

The content of the water-soluble polymer contained in the needle portion is preferably 10 mass% or more and 99 mass% or less, more preferably 10 mass% or more and 70 mass% or less, and further preferably 20 mass% or more and 50 mass% or less, with respect to the solid content of the needle portion.

The needle portion contains an influenza vaccine as a drug.

The influenza vaccine may contain a single virus antigen, or may contain two or more virus antigens. When a specific influenza virus is epidemic and rapidly produced and supplied to a specific vaccine strain, the vaccine preferably contains a single virus antigen. In the case where a broad range of viral strains are immunized by injection of the vaccine, it is preferred that the vaccine contains two or more viral antigens. The influenza vaccine may preferably comprise influenza a virus antigens, influenza B virus antigens or mixtures thereof. It is further preferred that the influenza vaccine comprises influenza a H1N1 antigen, influenza a H3N2 antigen, influenza B antigen or a mixture thereof. When two or more viral antigens are included in an influenza vaccine, the amount of antigen from each virus is not particularly limited, but it is preferable that the antigen from each virus is included in the vaccine in an equal amount.

The influenza vaccine and the vaccine stock solution may contain a pharmaceutically acceptable carrier as needed. As the pharmaceutically acceptable carrier, a carrier used for vaccine production can be used without limitation, and specifically, sugars, inorganic salts, buffered saline, glucose, water, glycerol, isotonic aqueous buffer, surfactants, emulsifiers, preservatives, isotonizing agents, pH adjusters and inactivating agents, and combinations of two or more thereof are appropriately blended.

Influenza vaccines and vaccine stocks may also contain immunopotentiators (adjuvants). Examples of adjuvants include mineral-containing compositions, oily emulsions, saponin compositions, virosomes, virus-like particles (VLPs), bacterial derivatives or microbial derivatives (non-toxic derivatives of intestinal bacterial lipopolysaccharides, lipid a derivatives, immunostimulatory oligonucleotides ADP ribosylating toxins and detoxified derivatives thereof), and the like.

The content of the influenza vaccine in the entire needle portion is not particularly limited, but the HA content per preparation is preferably 0.01 μ g to 200 μ g. More preferably 1. mu.g to 100. mu.g. HA is an abbreviation for hemoglobin (hemagglutinin).

The mass ratio of the drug to the water-soluble polymer in the needle portion is not particularly limited, but is preferably 1/0.5 to 1/10, more preferably 1/0.5 to 1/4.

In the present invention, the needle portion contains guanamine or a salt thereof. In addition, guanamine is also known as methylglucamine (N-methyl-D-glucamine).

Specific examples of the salt of guanamine include hydrochloride, sulfate, carboxylate, borate, methanesulfonate, p-toluenesulfonate, ascorbate and the like, and among them, hydrochloride is preferable.

The content of guanamine or a salt thereof in the needle portion is preferably 0.01 to 300 times that of the influenza vaccine, and more preferably 0.5 to 3 times that of the influenza vaccine.

The needle portion may also contain a soluble additive.

Examples of the soluble additive include saccharides.

As the saccharide, one or more of monosaccharides, disaccharides, oligosaccharides and polysaccharides can be used. Preferably, a disaccharide may be added to the needle portion. Examples of the disaccharide include sucrose, lactulose, lactose, maltose, trehalose, and cellulose sugar, and sucrose, maltose, and trehalose are particularly preferable.

When the needle portion contains the soluble supplement, the mass ratio of the drug (influenza vaccine) to the soluble supplement is not particularly limited, but is preferably 1/0.1 to 1/10, and more preferably 1/0.5 to 1/3.

Hereinafter, preferred embodiments of the present invention will be described with reference to the drawings, but the present invention is not limited thereto.

Fig. 2 to 9 show a partially enlarged microneedle 110 as a microneedle array. The microneedle array of the present invention is configured by forming a plurality of needle portions 112 on the surface of a sheet portion 116 (in the figure, only one needle portion 112, or one frustum portion 113 and one needle portion 112, which will be referred to as microneedles 110, is shown on the sheet portion 116).

In fig. 2A, the needle portion 112 has a conical shape, and in fig. 2B, the needle portion 112 has a quadrangular pyramid shape. In fig. 2C, H denotes the height of the needle portion 112, W denotes the diameter (width) of the needle portion 112, and T denotes the height (thickness) of the sheet portion 116.

Fig. 3 and 4 show a microneedle 110 having another shape in which a frustum portion 113 and a needle portion 112 are formed on a surface of a sheet portion 116. In fig. 3, the frustum portion 113 has a shape of a truncated cone, and the needle portion 112 has a shape of a cone. In fig. 4, the frustum portion 113 has a quadrangular pyramid shape, and the needle portion 112 has a quadrangular pyramid shape. However, the shape of the needle portion is not limited to these shapes.

Fig. 5 is a sectional view of the microneedle 110 shown in fig. 3 and 4. In fig. 5, H denotes the height of the needle portion 112, W denotes the diameter (width) of the base portion, and T denotes the height (thickness) of the sheet portion 116.

The microneedle array of the present invention is preferably in the shape of the microneedles 110 of fig. 5, according to the shape of the microneedles 110 of fig. 2C. With such a configuration, the volume of the entire needle portion becomes large, and more drug can be concentrated at the tip of the needle portion when manufacturing a microneedle array.

Fig. 6 and 7 illustrate a microneedle 110 having yet another shape.

The needle portion 1 st layer 112A shown in fig. 6 has a conical shape, and the needle portion 2 nd layer 112B has a columnar shape. The needle portion 1 st layer 112A shown in fig. 7 has a quadrangular pyramid shape, and the needle portion 2 nd layer 112B has a quadrangular prism shape. However, the shape of the needle portion is not limited to these shapes.

Fig. 8 is a sectional view of the microneedle 110 shown in fig. 6 and 7. In fig. 8, H denotes the height of the needle portion 112, W denotes the diameter (width) of the base portion, and T denotes the height (thickness) of the sheet portion 116.

Fig. 9 is a cross-sectional view of a microneedle having another shape in which the inclination (angle) of the side surface of the needle portion 112 changes continuously. In fig. 9, H denotes the height of the needle portion 112, and T denotes the height (thickness) of the sheet portion 116.

In the microneedle array of the present invention, the needle portions are preferably arranged at intervals of about 0.1 to 10 per 1mm of the row. More preferably, the microneedle array is per 1cm²Has 1-10000 micro-needles. By setting the density of the microneedles to 1 root/cm²In the above, the skin can be effectively perforated, and by setting the density of the microneedles to 10000 roots/cm²Hereinafter, the microneedle array can be sufficiently punctured. The density of the needle part is preferably 10 to 5000 roots/cm²More preferably 25 to 1000 roots/cm²Particularly preferably 25 to 400 roots/cm²。

The microneedle array of the present invention may be provided in a hermetically sealed form together with a desiccant. As the desiccant, a known desiccant (for example, silica gel, quicklime, calcium chloride, alumina, a sheet-like desiccant, or the like) can be used.

[ method for producing microneedle array ]

According to the present invention, there is provided a method for manufacturing a microneedle array of the present invention, including a step of concentrating an influenza vaccine, a step of forming a needle portion using the concentrated influenza vaccine obtained in the above step, and a step of forming a sheet portion.

The step of concentrating the influenza vaccine is preferably a step of concentrating the influenza vaccine by centrifugation.

In the present invention, the needle portion can be formed by filling a liquid containing the influenza vaccine, the water-soluble polymer, and guanamine or a salt thereof into a mold.

The microneedle array of the present invention can be produced, for example, by the method described in japanese patent application laid-open No. 2013-153866 or international publication No. WO 2014/077242.

(mold production)

Fig. 10A to 10C are process diagrams for manufacturing a mold (die). As shown in fig. 10A, a master for making a mold is first produced. There are two methods for making the master 11.

In the first method, a photoresist is coated on a Si substrate, and then exposure and development are performed. Then, an array of conical shaped portions (projections) 12 is formed on the surface of the original plate 11 by etching using RIE (reactive ion etching) or the like. In addition, when etching such as RIE is performed to form a conical portion on the surface of the original plate 11, the Si substrate can be obliquely etched while being rotated, thereby forming a conical shape. In the second method, an array of shape portions 12 such as rectangular pyramids is formed on the surface of the original plate 11 by machining using a cutting tool such as a diamond turning tool on a metal substrate such as Ni.

Next, a mold is manufactured. Specifically, as shown in fig. 10B, a mold 13 is formed from the master 11. As the method, the following 4 methods can be considered.

In the first method, a silicone resin obtained by adding a curing agent to PDMS (polydimethylsiloxane, for example, Sillgard184 (registered trademark) manufactured by Dow Corning corp.) is injected into the original plate 11, and the silicone resin is cured by heat treatment at 100 ℃. In the second method, a uv (ultraviolet) curable resin that is cured by irradiation with ultraviolet rays is injected into the original plate 11, and after irradiation with ultraviolet rays in a nitrogen atmosphere, the resin is peeled from the original plate 11. The third method is a method in which a solution obtained by dissolving a plastic resin such as polystyrene or PMMA (polymethyl methacrylate) in an organic solvent is poured into the original plate 11 coated with the release agent, and the organic solvent is evaporated and cured by drying the solution, and then the solution is released from the original plate 11. The fourth method is a method of producing a reverse product by Ni electroforming.

Thus, the mold 13 in which the needle-like recesses 15, which are the inverse shapes of the conical or pyramidal shape of the original plate 11, are two-dimensionally arranged is produced. The mold 13 thus produced is shown in fig. 10C.

Figure 11 shows another preferred form of mould 13. The needle-like recess 15 includes a tapered entrance portion 15A that is tapered in the depth direction from the surface of the die 13 and a distal end recess portion 15B that is tapered in the depth direction. By making the inlet portion 15A tapered, the water-soluble polymer solution can be easily filled into the needle-like recess 15.

Fig. 12 shows a form of the mold complex 18 more preferable in the production of the microneedle array. In fig. 12, the part (a) shows the mold composite 18. In fig. 12, the portion (B) is an enlarged view of the portion surrounded by the circle in the portion (a).

As shown in fig. 12(a), the mold composite 18 includes: a mold 13 having a vent hole 15C formed at the tip (bottom) of the needle-like recess 15; and a gas-permeable sheet 19 formed of a gas-permeable but liquid-impermeable material and attached to the back surface of the mold 13. The exhaust hole 15C is formed as a through hole penetrating the back surface of the mold 13. Here, the back surface of the mold 13 is a surface on which the vent hole 15C is formed. Therefore, the tip of the needle-like recess 15 communicates with the atmosphere through the gas discharge hole 15C and the gas permeable sheet 19.

By using such a mold complex 18, the polymer solution filled in the needle-like recesses 15 does not penetrate, and only air present in the needle-like recesses 15 can be discharged from the needle-like recesses 15. This makes it possible to transfer the shape of the needle-like recess 15 to the polymer well, and to form a finer needle portion.

The diameter D (diameter) of the vent hole 15C is preferably in the range of 1 to 50 μm. When the diameter D of the exhaust hole 15C is less than 1 μm, the function as an exhaust hole cannot be sufficiently exhibited. When the diameter D of the air vent 15C is larger than 50 μm, sharpness of the distal end of the molded microneedle is impaired.

As the gas-permeable sheet 19 formed of a gas-permeable but liquid-impermeable material, for example, a gas-permeable film (Sumitomo Electric Industries, ltd., product., Pouflon (registered trademark), FP-010) can be suitably used.

As a material used for the mold 13, an elastic material or a metal material can be used, and an elastic material is preferable, and a material having high gas permeability is more preferable. Oxygen permeability as a representative of gas permeability is preferably 1 × 10^-12(mL/s·m²Pa) or more, and more preferably 1X 10^-10(mL/s·m²Pa) or more. Further, 1mL was 10^-6m³. By setting the gas permeability to the above range, air present in the concave portion of the mold 13 can be discharged from the mold side, and a microneedle array with few defects can be manufactured. Specific examples of such a material include a silicone resin (for example, Silgard184 (registered trademark) manufactured by Dow Corning Corp., Shin-Etsu Chemical Co., Ltd., KE-1310ST (product model) manufactured by Ltd.), an ultraviolet curable resin, a plastic resin (for example, polystyrene or PMMA) melted or dissolved in a solvent, and the like. Among these materials, silicone rubber-based materials are preferable because they are durable against repeated pressure transfer and have good releasability from the material. Examples of the metallic material include Ni, Cu, Cr, Mo, W, Ir, Tr, Fe, Co, MgO, Ti, Zr, Hf, V, Nb, Ta, α -alumina, zirconia, stainless steel (e.g., STAVAX (trademark) material from Bohler-Uddeholm, Bohler-Uddeholm KK), and alloys thereof. As the material of the frame, the same material as that of the mold 13 can be used.

(Water-soluble Polymer solution)

In the present invention, it is preferable to prepare a water-soluble polymer solution containing an influenza vaccine and guanamine or a salt thereof for forming at least a part of the needle portion and a water-soluble polymer solution for forming the sheet portion.

The kind of the water-soluble polymer is as described in the present specification.

In any of the above-mentioned water-soluble polymer solutions, a disaccharide may be mixed, and the type of the disaccharide is as described in the present specification.

The solvent used for dissolution may be any solvent other than warm water as long as it is volatile, and Methyl Ethyl Ketone (MEK), ethanol, or the like may be used.

The water-soluble polymer solution containing a drug and guanamine or a salt thereof for forming at least a part of the needle portion is specifically a solution containing an influenza vaccine, a water-soluble polymer, and guanamine or a salt thereof.

The concentration of the influenza vaccine in the liquid containing the influenza vaccine, the water-soluble polymer, and guanamine or a salt thereof is not particularly limited, but is preferably 0.001mg/mL to 100mg/mL, and more preferably 0.1mg/mL to 20 mg/mL.

The concentration of guanamine in the liquid containing the influenza vaccine, the water-soluble polymer, and guanamine or a salt thereof is not particularly limited, but is preferably 0.001mg/mL to 100mg/mL, and more preferably 0.1mg/mL to 20 mg/mL.

The concentration of the water-soluble polymer in the liquid containing the influenza vaccine, the water-soluble polymer, and guanamine or a salt thereof is not particularly limited, but is preferably 1mg/mL to 100mg/mL, and more preferably 5mg/mL to 50 mg/mL.

(formation of needle portion)

As shown in fig. 13A, the mold 13 having the needle-like recesses 15 arranged two-dimensionally is disposed on the base 20. The mold 13 has 2 sets of a plurality of needle-like recesses 15 two-dimensionally arranged at 5 × 5. A liquid supply device 36 is prepared, and the liquid supply device 36 has a tank 30 for storing a water-soluble polymer solution 22 containing a drug and guanamine or a salt thereof, a pipe 32 connected to the tank, and a nozzle 34 connected to the tip of the pipe 32. In this example, the needle-like recesses 15 are two-dimensionally arranged at 5 × 5, but the number of needle-like recesses 15 is not limited to 5 × 5, and may be M × N two-dimensionally arranged (M and N each independently represent an arbitrary integer of 1 or more, preferably 2 to 30, more preferably 3 to 25, and still more preferably 3 to 20).

Fig. 14 is a schematic perspective view of the tip end of the nozzle. As shown in fig. 14, a lip portion 34A as a flat surface and a slit-shaped opening portion 34B are provided at the tip of the nozzle 34. The slit-shaped openings 34B allow the water-soluble polymer solution 22 containing the drug and guanamine or a salt thereof to be simultaneously filled into the plurality of needle-shaped recesses 15 constituting, for example, row 1. The size (length and width) of the opening 34B is appropriately selected according to the number of needle-like recesses 15 to be filled at one time. By increasing the length of the opening 34B, more needle-like recesses 15 can be filled with the polymer solution 22 containing the drug at one time. This can improve productivity.

As a material for the nozzle 34, an elastic material or a metal material may be used. Examples thereof include Teflon (registered trademark), Stainless Steel (SUS (Steel Special Use Stainless Steel)), titanium and the like.

As shown in fig. 13B, the position of the opening 34B of the nozzle 34 is adjusted above the needle-like recess 15. The lip 34A of the nozzle 34 contacts the surface of the mold 13. The water-soluble polymer solution 22 containing the drug and guanamine or a salt thereof is supplied from the liquid supply device 36 to the mold 13, and the water-soluble polymer solution 22 containing the drug and guanamine or a salt thereof is filled into the needle-like recess 15 from the opening 34B of the nozzle 34. In the present embodiment, a plurality of needle-like recesses 15 constituting row 1 are simultaneously filled with a water-soluble polymer solution 22 containing a drug and guanamine or a salt thereof. However, the needle-like recess 15 may be filled with the liquid one by one without being limited thereto.

When the mold 13 is made of a material having gas permeability, the water-soluble polymer solution 22 containing the drug and guanamine or a salt thereof can be sucked by sucking from the back surface of the mold 13, and the filling of the water-soluble polymer solution 22 containing the drug and guanamine or a salt thereof into the needle-like recess 15 can be promoted.

Referring to fig. 13B, after the filling step, as shown in fig. 13C, the nozzle 34 is moved to the needle-like recess 15 not filled with the water-soluble polymer solution 22 containing the drug and guanamine or a salt thereof by relatively moving the liquid supply device 36 in a direction perpendicular to the longitudinal direction of the opening 34B while bringing the lip 34A of the nozzle 34 into contact with the surface of the mold 13. The position of the opening 34B of the nozzle 34 is adjusted above the needle-like recess 15. In the present embodiment, the example of moving the nozzle 34 is described, but the mold 13 may be moved.

Since the lip 34A of the nozzle 34 moves in contact with the surface of the mold 13, the nozzle 34 can scrape off the water-soluble polymer solution 22 containing the drug and guanamine or a salt thereof remaining on the surface of the mold 13 other than the needle-like recess 15. The water-soluble polymer solution 22 containing the drug and guanamine or a salt thereof can be prevented from remaining in the mold 13 except the needle-like recess 15.

In order to reduce damage to the mold 13 and to suppress deformation due to compression of the mold 13 as much as possible, it is preferable that the pressing pressure of the nozzle 34 against the mold 13 during movement be as low as possible. At least one of the mold 13 and the nozzle 34 is preferably made of a material that is elastically and flexibly deformed so as not to leave the water-soluble polymer solution 22 containing the drug and guanamine or a salt thereof outside the needle-like recess 15 of the mold 13.

By repeating the filling step of fig. 13B and the moving step of fig. 13C, the water-soluble polymer solution 22 containing the drug and guanamine or a salt thereof is filled in the needle-like recesses 15 two-dimensionally arranged at 5 × 5. When the water-soluble polymer solution 22 containing the drug and guanamine or a salt thereof is filled into the needle-shaped recesses 15 two-dimensionally arranged at 5 × 5, the liquid supply device 36 moves to the adjacent needle-shaped recesses 15 two-dimensionally arranged at 5 × 5, and the filling step of fig. 13B and the moving step of fig. 13C are repeated. The adjacent needle-like recesses 15 two-dimensionally arranged at 5 × 5 are also filled with a water-soluble polymer solution 22 containing a drug and guanamine or a salt thereof.

The filling step and the moving step may be (1) a method of filling the needle-shaped recess 15 with the water-soluble polymer solution 22 containing the drug and the guanamine or the salt thereof while moving the nozzle 34, or (2) a method of temporarily stopping the nozzle 34 on the needle-shaped recess 15 during the movement of the nozzle 34, filling the needle-shaped recess 15 with the water-soluble polymer solution 22 containing the drug and the guanamine or the salt thereof, and moving the nozzle 34 again after the filling. Between the filling process and the moving process, the lip 34A of the nozzle 34 is in contact with the surface of the mold 13.

Fig. 15 is a partially enlarged view of the tip of the nozzle 34 and the mold 13 when the water-soluble polymer solution 22 containing the influenza vaccine and guanamine or a salt thereof is filled in the needle-like recess 15. As shown in fig. 15, the filling of the water-soluble polymer solution 22 containing the drug and guanamine or a salt thereof into the needle-like recess 15 can be promoted by applying a pressure P1 into the nozzle 34. Further, when the water-soluble polymer solution 22 containing the drug and guanamine or a salt thereof is filled into the needle-like recess 15, the pressing force P2 for bringing the nozzle 34 into contact with the surface of the mold 13 is preferably equal to or greater than the pressing force P1 in the nozzle 34. By setting the pressing force P2 to a pressing force P1, the water-soluble polymer solution 22 containing the drug and guanamine or a salt thereof can be prevented from leaking from the needle-like recesses 15 to the surface of the mold 13.

Fig. 16 is a partially enlarged view of the front end of the nozzle 34 and the mold 13 while the nozzle 34 is moving. When the nozzle 34 is moved relative to the mold 13, the pressing force P3 for bringing the nozzle 34 into contact with the surface of the mold 13 is preferably smaller than the pressing force P2 for bringing the nozzle 34 being filled into contact with the surface of the mold 13. This is to reduce damage to the mold 13 and to suppress deformation due to compression of the mold 13.

After the completion of filling the plurality of needle-like recesses 15 of 5 × 5, the nozzle 34 is moved to the adjacent plurality of needle-like recesses 15 of 5 × 5. The supply of the liquid is preferably stopped when the liquid moves to the adjacent needle-like recesses 15 of 5 × 5, and the supply of the water-soluble polymer solution 22 containing the drug and guanamine or a salt thereof is stopped. There is a distance from the needle recess 15 of the 5 th row to the needle recess 15 of the next row. If the water-soluble polymer solution 22 containing the drug and guanamine or a salt thereof is continuously supplied while the nozzle 34 is moved therebetween, the liquid pressure in the nozzle 34 may become excessively high. As a result, the water-soluble polymer solution 22 containing the drug and guanamine or a salt thereof may flow out from the nozzle 34 to the outside of the needle-shaped recess 15 of the mold 13, and in order to suppress this, it is preferable to detect the liquid pressure in the nozzle 34 and stop the supply of the water-soluble polymer solution 22 containing the drug and guanamine or a salt thereof when it is determined that the liquid pressure is too high.

In addition, although the method of supplying the drug and the water-soluble polymer solution of guanamine or a salt thereof using a dispenser having a nozzle has been described above, the application may be performed by bar coating, spin coating, spraying, or the like, in addition to the application using the dispenser.

In the present invention, it is preferable that a water-soluble polymer solution containing an influenza vaccine and guanamine or a salt thereof is supplied to the needle-shaped recess and then dried. That is, in the method for producing a microneedle array of the present invention, it is preferable that the filling step of filling a mold with a liquid containing an influenza vaccine, a water-soluble polymer, and guanamine or a salt thereof is followed by a drying step of drying the liquid.

Further, the method for manufacturing a microneedle array according to the present invention preferably includes a step of applying a water-soluble polymer solution to the mold after the drying step. That is, as a preferable example of the method for producing a microneedle array of the present invention, there is a method including: forming a part of the needle part by drying the needle part-forming mold filled with the first water-soluble polymer solution containing the influenza vaccine and guanamine or a salt thereof; and a step of filling a second water-soluble polymer solution on the upper surface of a part of the formed needle part and drying the filled needle part.

The conditions for drying the needle-forming mold filled with the first water-soluble polymer solution containing the influenza vaccine and guanamine or a salt thereof are preferably conditions under which the water content of the first water-soluble polymer solution is 20% or less after 30 to 300 minutes from the start of drying.

In particular, it is preferable that the drying is maintained at a temperature not higher than the temperature at which the drug does not lose efficacy, and that the water content of the water-soluble polymer solution is controlled to 20% or lower after 60 minutes or longer has elapsed from the start of the drying.

As the method of controlling the drying rate, for example, any means capable of delaying drying, such as temperature, humidity, drying air volume, use of a container, and volume and/or shape of a container, may be adopted.

The drying is preferably performed in a state where the container is covered with a mold for forming a needle portion filled with a first water-soluble polymer solution containing the drug or in a state where the container is stored.

The temperature during drying is preferably 1 to 45 ℃, and more preferably 1 to 40 ℃.

The relative humidity during drying is preferably 10 to 95%, more preferably 20 to 95%, and still more preferably 30 to 95%.

(formation of sheet portion)

Several modes of the step of forming the sheet portion will be described.

The step of forming the sheet portion will be described with reference to fig. 17A to 17D as embodiment 1. The needle-like recess 15 of the mold 13 is filled with a water-soluble polymer solution 22 containing a drug and guanamine or a salt thereof from a nozzle 34. Next, as shown in fig. 17B, the water-soluble polymer solution 22 containing the influenza vaccine and guanamine or a salt thereof is dried and solidified, thereby forming a drug-containing layer 120 in the needle-like recesses 15. Next, as shown in fig. 17C, a water-soluble polymer dissolving solution 24 is applied by a dispenser to the mold 13 on which the drug-containing layer 120 is formed. In addition to coating using a dispenser, bar coating, spin coating, spray coating, or the like may be used. Since the drug-containing layer 120 is solidified, diffusion of the drug into the water-soluble polymer solution 24 can be suppressed. Next, as shown in fig. 17D, the water-soluble polymer solution 24 is dried and solidified to form the microneedle array 1 including the plurality of needle portions 112, the truncated cone portion 113, and the sheet portion 116.

In embodiment 1, in order to facilitate filling of the water-soluble polymer solution 22 and the water-soluble polymer solution 24 containing the influenza vaccine and guanamine or a salt thereof into the needle-like recesses 15, it is also preferable to apply pressure from the front surface of the mold 13 and suction under reduced pressure from the back surface of the mold 13.

Next, the 2 nd embodiment will be described with reference to fig. 18A to 18C. As shown in fig. 18A, a water-soluble polymer solution 22 containing a drug and guanamine or a salt thereof is filled into the needle-like recess 15 of the mold 13 from the nozzle 34. Next, in the same manner as in fig. 17B, the water-soluble polymer solution 22 containing the drug and guanamine or a salt thereof is dried and solidified, thereby forming a drug-containing layer 120 in the needle-like recesses 15. Next, as shown in fig. 18B, a water-soluble polymer solution 24 is applied to the other support 29. The material of the support 29 is not limited, and for example, polyethylene terephthalate, polycarbonate, polypropylene, acrylic resin, triacetyl cellulose, glass, or the like can be used. Next, as shown in fig. 18C, the water-soluble polymer solution 24 formed on the support 29 is superimposed on the mold 13 having the drug-containing layer 120 formed on the needle-like recess 15. Thereby, the water-soluble polymer solution 24 is filled into the needle-like recess 15. Since the drug-containing layer 120 is solidified, diffusion of the drug into the water-soluble polymer solution 24 can be suppressed. Next, the water-soluble polymer solution 24 is dried and solidified to form a microneedle array including a plurality of needle portions 112, a truncated cone portion 113, and a sheet portion 116.

In embodiment 2, in order to promote the filling of the water-soluble polymer solution 24 into the needle-like recesses 15, it is also preferable to apply pressure from the front surface of the mold 13 and suction under reduced pressure from the back surface of the mold 13.

The method for drying the water-soluble polymer solution 24 may be any method as long as it is a step of volatilizing the solvent in the polymer solution. The method is not particularly limited, and for example, heating, blowing, or pressure reduction may be used. The drying treatment may be performed at 1 to 50 ℃ for 1 to 72 hours. In the case of blowing air, a method of blowing warm air of 0.1 to 10 m/sec is exemplified. The drying temperature is preferably a temperature at which the drug in the polymer solution 22 containing the drug is not thermally degraded.

(peeling)

The method of peeling the microneedle array from the mold 13 is not particularly limited. The needle portion is preferably not bent or bent at the time of peeling. Specifically, as shown in fig. 19, a sheet-like base material 40 having an adhesive layer formed thereon is attached to the microneedle array, and then peeled off while being turned over from the end portion of the base material 40. However, this method may cause the needle portion to bend. Therefore, as shown in fig. 20, a method of providing a suction pad (not shown) on the base material 40 on the microneedle array and vertically lifting the base material by air suction can be applied. The support 29 may be used as the substrate 40.

Fig. 21 shows the microneedle array 2 peeled off from the mold 13. The microneedle array 2 is composed of a base material 40, a needle portion 112 formed on the base material 40, a truncated cone portion 113, and a sheet portion 116. Although the needle portion 112 has a conical shape or a polygonal pyramid shape at least at the front end, the needle portion 112 is not limited to this shape.

The method for producing the microneedle array of the present invention is not particularly limited, but is preferably obtained by a production method including the following steps: (1) a step of producing a mold, (2) a step of preparing a water-soluble polymer solution containing an influenza vaccine and guanamine or a salt thereof, (3) a step of filling the liquid obtained in step (2) into the mold to form the upper end portion of the needle portion, (4) a step of filling the water-soluble polymer into the mold to form the lower end portion of the needle portion and the sheet portion, and (5) a step of peeling off the mold.

The present invention will be described in more detail below with reference to examples of the present invention. The materials, the amounts used, the ratios, the contents of the processes, the processing procedures, and the like described in the following examples can be modified as appropriate without departing from the spirit of the present invention. Therefore, the scope of the present invention should not be construed as being limited to the specific examples shown below.

Examples

Production of microneedle arrays containing influenza vaccine

(production of mold)

On the surface of a smooth Ni plate having one side of 40mm, 100 needles were ground into a two-dimensional square array at a pitch L1 of 1000 μm by a quadrangular shape on a truncated cone 50 having a diameter D1 of 800 μm at the bottom and a height H1 of 200 μm as shown in fig. 22, and a shape portion 12 having a needle-like structure in which a cone 52 having a height H2 of 834 μm is formed at a diameter D2 of 340 μm, thereby producing a master plate 11. A film was formed on the original plate 11 by using silicone rubber (SILASTIC MDX4-4210, manufactured by Dow Corning Corp.) to a thickness of 0.6mm, and the film was thermally cured and peeled off with the cone tip portion of the original plate 11 protruding 50 μm from the film surface. Thus, a silicone rubber reverse having a through-hole with a diameter of about 30 μm was produced. The silicone rubber reverse product was used as a mold, which had needle-like recesses two-dimensionally arranged in 10 rows × 10 lines at the center and a flat surface portion 30mm on one side circumscribed. The opening of the needle-like recess was wider than the mold, and the surface of the through-hole (vent hole) having a diameter of 30 μm was the back surface of the mold.

(preparation of influenza vaccine concentrate)

The influenza vaccine stock solution was placed in a dedicated ultracentrifugation container, and the influenza vaccine was sedimented by ultracentrifugation (conditions: 131, 491 Xg, 90 minutes, 4 ℃). The supernatant was discarded, PBS (phosphate buffered saline) was added instead of the supernatant to carry out vortexing, and the mixture was allowed to stand overnight at 4 ℃ to prepare an influenza vaccine concentrate.

(preparation of Water-soluble Polymer solution containing influenza vaccine and guanamine)

An aqueous solution prepared by mixing an influenza vaccine concentrate, a water-soluble polymer, guanamine (FUJIFILM Wako Pure Chemical Corporation), a saccharide, and Tween (registered trademark) 80(MERCK) was prepared as an influenza vaccine-containing water-soluble polymer solution. As the water-soluble polymer, hydroxyethyl starch (HES) (Fresenius Kabi) and Chondroitin Sulfate (CS) (Maruha Nichiro Corporation) were used. Sucrose (Suc) (FUJIFILM Wako Pure Chemical Corporation), trehalose (Tre) (forest land), and glucose (Japanese pharmacopoeia grade FUJIFILM Wako Pure Chemical Corporation) were used as the saccharides. The formulations of the solutions are shown in tables 1, 2 and 3 below. The influenza vaccine content in tables 1 and 3 was set to 9mg/1 mL. The influenza vaccine content in Table 2 was set to 10.3mg/1 mL.

(preparation of a solution of a Water-soluble Polymer forming the sheet portion)

Chondroitin sulfate (Maruha Nichiro Corporation) was dissolved in water to prepare a water-soluble polymer solution for forming a sheet portion.

(filling and drying of the drug-containing Polymer solution)

A filling device as shown in fig. 23 was used. The filling device comprises: an X-axis drive section 61 and a Z-axis drive section 62 that control the relative position coordinates of the mold and the nozzle, a liquid supply device 64(Musashi engineering co., ltd. manufactured micro quantitative dispenser SMP-III) to which a nozzle 63 can be attached, a suction table 65 to which a mold 69 is fixed, a laser displacement meter 66 (HL-C201A, manufactured by Panasonic Corporation) that measures the shape of the mold surface, a load cell 67(Kyowa Electronic Instruments co., ltd. manufactured LCX-a-500N) that measures the advancing pressure of the nozzle, and a control mechanism 68 that controls the Z-axis based on the data of the surface shape and the measured value of the pressing pressure.

A15 mm-side gas permeable film (Sumitomo Electric Industries, Ltd., Pouflon (registered trademark), FP-010) was placed on a horizontal suction table, and a mold was placed so that the surface was on the top. The pressure was reduced from the back side of the mold under a suction pressure of 90kPa, and the gas permeable film and the mold were fixed on a vacuum table.

An SUS (stainless steel) nozzle having a shape as shown in FIG. 14 was prepared, and a slit-like opening having a length of 12mm and a width of 0.2mm was formed in the center of a lip having a length of 20mm and a width of 2 mm. The nozzle was connected to a liquid supply. A water-soluble polymer solution containing 3mL of a drug and guanamine or a salt thereof was filled in the liquid supply device and the nozzle. The nozzle was adjusted so that the opening portion was parallel to the 1 st row composed of a plurality of needle-like recesses formed on the surface of the mold. At a position spaced at intervals of 2mm in the opposite direction to the row 2 with respect to the row 1, at 1.372X 10⁴Pa(0.14kgf/cm²) The pressure of (a) presses the nozzle against the mould. In the state of pressing the nozzle, the Z axis is controlled to make the variation of the pressing pressure within + -0.490 × 10⁴Pa(0.05kgf/cm²) The water-soluble polymer solution containing the drug and guanamine or a salt thereof was discharged from the opening at 0.15. mu.L/sec by a liquid supply apparatus for 20 seconds while moving at 0.5 mm/sec in a direction perpendicular to the longitudinal direction of the opening. The nozzle was removed from the mold by stopping the movement of the nozzle at positions spaced 2mm apart by the hole pattern of the two-dimensionally arranged needle-like recesses.

The mold filled with the water-soluble polymer solution containing the influenza vaccine was allowed to stand at a temperature of 23 ℃ and a relative humidity of 45%, and dried.

(formation and drying of sheet portion)

The mold filled with the water-soluble polymer solution containing the influenza vaccine is placed on a vacuum table and is placed under reduced pressure for suction, and the mold is spread over the water-soluble polymer solution forming the sheet portion by suction. After adding the water-soluble polymer solution for 60 minutes, the suction was stopped, and the mixture was allowed to stand at 23 ℃ under a relative humidity of 45% and dried.

(peeling)

The dried and solidified microneedle array was carefully peeled off from the mold to form a microneedle array containing an influenza vaccine. The microneedle comprises a truncated cone portion and a needle portion, and has a truncated cone structure in which the height of the needle portion is about 800 μm, the width of the base portion is about 320 μm, the height of the truncated cone portion is about 160 μm, the diameter of the upper bottom surface is about 320 μm, and the diameter of the lower bottom surface is about 780 μm, and the microneedle is square-shaped such that the thickness of the sheet portion is about 200 μm, the number of needles is 100, and the distance between the needles is about 1 mm.

< evaluation of microneedle array >

(quantification of influenza vaccine content in microneedles)

(a) Content in 600 μm microneedle from tip to front end

The needle portion of a microneedle having a needle length of 800 μm was cut at a position 800 μm from the tip of the microneedle by a cutter. The cut needle portion was recovered in a 1.5mL tube. 0.5mL of phosphate buffer was added to a 1.5mL tube containing the recovered needle portion, and the mixture was stirred to dissolve the needle portion. The needle lysate was diluted to an appropriate concentration with phosphate buffer, and the influenza vaccine content in the cut needle was quantified by ELISA (enzyme linked immunosorbent assay).

(b) Content in 800 μm microneedle from tip to front end

The needle residue cut at a length of 600 μm from the tip was cut at the boundary between the needle portion and the frustum portion by a cutter. The remaining portion of the cut needle was recovered into a 1.5mL tube. 0.5mL of phosphate buffer was added to a 1.5mL tube containing the recovered needle portion, and the mixture was stirred to dissolve the needle portion. The needle lysate was diluted to an appropriate concentration with phosphate buffer, and the content of influenza vaccine in the cut needle was quantified by ELISA.

The content of the remaining portion of the needle was determined and the content of the microneedle in which 600 μm was added from the needle tip determined in the above (a) was determined as the content of the microneedle 800 μm from the needle tip.

The region of the microneedle tip to 600 μm and the region of the microneedle tip to 800 μm are shown in fig. 1. In addition, the needle remnant in the above (b) corresponds to a region of 200 μm in FIG. 1 (i.e., a region from the tip to 800 μm from which the tip to 600 μm region was removed).

The results of obtaining the tip filling ratio of each microneedle array by the following formula are shown in tables 1, 2 and 3 below.

Front end filling rate (influenza vaccine content in 600 μm microneedle from tip to front end/800 μm microneedle from tip to front end)

(stability of influenza vaccine content in microneedle)

For the manufactured microneedle array containing the influenza vaccine, it was left standing at 35 ℃ for 3 days. And (5) taking out the microneedle array after standing, and investigating the content of the vaccine. The vaccine content was evaluated by ELISA.

The residual vaccine rate was determined as the content of the vaccine immediately after standing at 35 ℃ for 3 days.

[ Table 1]

[ Table 2]

[ Table 3]

Description of the symbols

1 microneedle array

2 microneedle array

110 microneedle

112 needle part

113 frustum part

116 sheet part

120 layer containing polio vaccine

122 polio vaccine free layer

W diameter (Width)

Height H

T height (thickness)

11 original edition

12 shaped part

13 mould

15 needle-like recess

D diameter (diameter)

18 mould composite

19 gas permeable sheet

20 base station

22 poliomyelitis-containing vaccine liquid

24 Water-soluble Polymer solution

29 support body

30 jar

32 piping

34 nozzle

34A lip

34B opening part

36 liquid supply device

P1 applied pressure

P2 pressing force

P3 pressing force

40 base material

50 truncated cone

52 cone

D1 diameter

D2 diameter

L1 pitch

Height H1

Height H2

61X-axis driving part

62Z-axis driving part

63 spray nozzle

64 liquid supply device

65 suction table

66 laser displacement meter

67 force cell

68 control mechanism

69 mould