Method for preparing microcapsule structure of ergothioneine with probiotic and oral dosage form

Abstract

The present application relates to a microcapsule structure containing ergothioneine and probiotics, a preparation method, uses and oral dosage forms thereof, comprising: a core layer having a plurality of pores, which are communicated with each other, not communicated, partially communicated or partially not communicated, and an attached layer formed by spraying a concentrated liquid to part or all of the surface of the core layer and the pores, the concentrated liquid containing a plurality of probiotics, and the core layer being formed by mushroom fruiting body particles. By increasing the content of polysaccharides, polyphenols and ergothioneine contained in each microcapsule structure, the concentration of the medicinal active ingredients with health care effects for the human body is ensured, and the inconvenience of having to eat a large amount of mushrooms, the difficulty of digestion and absorption or the disadvantage of overeating in the past are solved. The present application also provides a preparation method, uses and oral dosage forms.

Description

Technical Field

[0001] This invention relates to a microcapsule structure, specifically a microcapsule structure formed by a core layer of mushroom fruiting body particles, and an attachment layer containing polysaccharides, polyphenols, and ergothioneine formed on the surface of the core layer, as well as the preparation method, uses, and oral dosage form thereof. Background Technology

[0002] Mushrooms have long been a popular food among health supplements, prized for their rich content of various functional components that offer numerous health benefits, including boosting energy, enhancing immunity, fighting cancer, and regulating blood sugar and lipids. Among these functional components, ergothioneine is gaining increasing attention for its powerful antioxidant and anti-inflammatory properties. Furthermore, the polysaccharides in mushrooms also offer numerous health benefits, such as lowering blood sugar and cholesterol, regulating immunity, and promoting intestinal motility by adjusting gut microbiota.

[0003] Ergothioneine cannot be synthesized in animals or plants and can only be obtained through diet. Consuming mushrooms is the most effective way to obtain ergothioneine. However, to achieve a concentration with health benefits, a large amount of mushrooms needs to be consumed. Mushrooms are high in potassium, which is unsuitable for kidney patients with high blood potassium or gout patients with high uric acid. Furthermore, mushrooms have thick cell walls that are difficult for the body to digest; excessive consumption can burden the digestive system and cause discomfort. Additionally, ergothioneine is highly water-soluble, and improper cooking methods can easily lead to its loss.

[0004] Therefore, this invention describes how to effectively improve the inconvenience of directly consuming ergothioneine through cooking mushrooms in the traditional way by means of innovative invention design, so that users can more easily consume enough ergothioneine, polyphenols and polysaccharides and other probiotics to improve intestinal function and reduce the occurrence of constipation. Summary of the Invention

[0005] To address the inconvenience or excessive consumption of mushrooms in the past, this invention provides a microcapsule structure containing prebiotics, its preparation method, uses, and oral dosage form. By increasing the content of polysaccharides, polyphenols, and ergothioneine in each microcapsule structure, this invention ensures that the microcapsules contain a sufficient concentration of pharmaceutically active ingredients and reduces the disadvantages associated with directly consuming mushrooms.

[0006] The main objective of this invention is to provide a microcapsule structure of ergothioneine containing prebiotics, comprising: at least one core layer and an attachment layer, wherein the core layer has a plurality of pores, which may be interconnected, non-interconnected, partially interconnected, or partially non-interconnected, and the attachment layer is formed by spraying a concentrated liquid onto part or all of the surfaces of the core layer and the pores, wherein the weight ratio of the attachment layer to the core layer is 1:0.5 to 1:2.2.

[0007] In one embodiment of the present invention, the concentrated liquid contains polysaccharides, polyphenols and ergothioneine, and the core layer is formed of mushroom fruiting body particles.

[0008] In one embodiment of the present invention, the core layer is an irregular structure, a sphere, an ellipsoid, a cylinder, or a polyhedron.

[0009] In one embodiment of the present invention, the ergothioneine microcapsule structure containing prebiotics is composed of multiple core layers having the same or different particle sizes.

[0010] In one embodiment of the present invention, the core layer is obtained by freeze-drying.

[0011] Based on its main objective, this invention also provides the use of a prebiotic-containing ergothioneine microcapsule structure in the preparation of ergothioneine microcapsules for improving constipation and clearing accumulated stool.

[0012] Based on its main objective, the present invention further provides a microcapsule structure of ergothioneine containing prebiotics, comprising: at least one core layer and an attachment layer, wherein the core layer has a plurality of pores that are interconnected, non-interconnected, partially interconnected, or partially non-interconnected with each other, and the attachment layer is coated on part or all of the surfaces of both the core layer and the pores, and the attachment layer comprises an ergothioneine layer.

[0013] Based on its main objective, this invention also provides a method for preparing a microcapsule structure containing prebiotics of ergothioneine. The method comprises the following steps: (a) using a first mushroom fruiting body particle as a core layer, the core layer having multiple pores; and (b) spraying a concentrated liquid onto part or all of the surfaces of the core layer and the pores through a granulation step to form an adhesion layer, wherein the concentrated liquid contains polysaccharides, polyphenols and ergothioneine, and the weight ratio of the adhesion layer to the core layer is 1:0.5 to 1:2.2.

[0014] In one embodiment of the present invention, the concentrated liquid is made from a second mushroom fruiting body by sequentially passing through a second pulverizing step, an extraction step, and a concentration step.

[0015] In one embodiment of the present invention, the second pulverizing step involves adding the second mushroom fruiting body to a solvent to form a mushroom mixture, and then pulverizing it. The mushroom mixture comprises 10 to 80 parts by weight of the mushroom fruiting body and the remainder is solvent, which is pure water or a 20% to 95% ethanol solution.

[0016] In one embodiment of the present invention, the extraction step involves extracting the pulverized mushroom mixture through a hot water extraction step or an ultrasonic extraction step to obtain a first extract. The extraction time is 0.5 to 24 hours. The first extract is then centrifuged to collect a supernatant, which is a second extract.

[0017] In one embodiment of the present invention, the concentration step involves concentrating the second extract through a vacuum concentration step or a heating concentration step to obtain the concentrated liquid, and the ratio of the weight of the concentrated liquid to the weight of the second mushroom fruiting body is in the range of 0.1 to 0.3.

[0018] In one embodiment of the present invention, the mushroom fruiting body particles are made by sequentially passing a first mushroom fruiting body through a freeze-drying step and a dry grinding step.

[0019] In one embodiment of the present invention, the freeze-drying step involves freezing the first mushroom fruiting body at a freezing temperature of -20°C to -80°C.

[0020] In one embodiment of the present invention, the dry grinding step utilizes a dry grinding technique to grind the frozen first mushroom fruiting body into mushroom fruiting body particles.

[0021] In one embodiment of the present invention, the fruiting body particles of the mushroom are obtained by sieving through a first mesh screen with a mesh size of 80.

[0022] In one embodiment of the present invention, the ergothioneine microcapsule structure containing prebiotics is obtained by sieving through a second mesh screen with a mesh size of 60.

[0023] In one embodiment of the present invention, the first or second fungal fruiting body is shiitake mushroom, enoki mushroom, king oyster mushroom, corn mushroom, or a combination thereof.

[0024] Based on its main objective, the present invention also provides an oral dosage form comprising a plurality of prebiotic-containing ergothioneine microcapsule structures, wherein the prebiotic-containing ergothioneine microcapsule structures are prepared by a method described above for preparing prebiotic-containing ergothioneine microcapsule structures, and the oral dosage form is selected from gelatin capsules and non-gelatin capsules.

[0025] Therefore, this invention utilizes a core layer formed by mushroom fruiting body particles, and a microcapsule structure containing polysaccharides, polyphenols, and ergothioneine formed on the surface of the core layer. By purposefully increasing the surface area or filling capacity of the attachment layer, and by increasing the content of polysaccharides, polyphenols, and ergothioneine in each microcapsule structure, the invention ensures that the microcapsule structure has a concentration of medicinal active ingredients that have health benefits for the human body, thus solving the inconvenience or excessive consumption of mushrooms in the past. Attached Figure Description

[0026] Figure 1 : A schematic diagram of the overall structure of ergothioneine microcapsules containing prebiotics according to a preferred embodiment of the present invention.

[0027] Figure 2A : A cross-sectional view of the core layer of a preferred embodiment of the present invention.

[0028] Figure 2B A cross-sectional view (a) of the structure of a prebiotic-containing ergothioneine microcapsule according to a preferred embodiment of the present invention.

[0029] Figure 2C : A cross-sectional view (II) of the structure of ergothioneine microcapsules containing prebiotics according to a preferred embodiment of the present invention.

[0030] Figure 3 (A) is a magnified view of the core layer observed using a scanning electron microscope.

[0031] Figure 3 (B) is an enlarged view of the structure of ergothioneine microcapsules containing prebiotics observed using a scanning electron microscope.

[0032] Figure 4 The present invention provides a preferred embodiment of a method for preparing ergothioneine microcapsule structures containing prebiotics.

[0033] Figure 5 (A) and (B): These are data graphs showing the results of a questionnaire survey on bowel movements before and after taking the ergothioneine microcapsule structure containing prebiotics of the present invention and a regular mushroom powder capsule.

[0034] Figure 6 : Graph showing experimental data on the preparation of concentrated liquids using different types of mushrooms and the testing of their ergothioneine content. Detailed Implementation

[0035] To facilitate understanding of the technical features, content, advantages, and effects of this invention, the invention will be described in detail below with reference to the accompanying drawings. The drawings used are for illustrative purposes only and to assist in the description. They may not represent the actual proportions and precise configurations of the invention after implementation. Therefore, the proportions and configurations of the accompanying drawings should not be used to interpret or limit the scope of the invention in actual implementation. This should be stated in advance.

[0036] To make the description of the present invention more detailed and complete, the following illustrative descriptions of the embodiments and specific examples of the present invention are provided; however, these are not the only forms of implementing or using the specific examples of the present invention.

[0037] Throughout this specification, unless otherwise specified, the singular form is to be understood to include the plural form as well. Furthermore, unless otherwise specified, all terms used in this specification are to be understood to have the meaning commonly used in the field. Therefore, unless otherwise defined, all technical and scientific terms used in this specification have the same meaning as commonly understood by those skilled in the art to which this invention pertains.

[0038] Please see Figure 1 As shown, the main objective of this invention is to provide a microcapsule structure containing prebiotics such as ergothioneine. This invention uses mushroom fruiting body particles as the core layer (11) and an attachment layer (13) formed by spraying a concentrated liquid onto the surface of both the core layer (11) and the pores (12). The concentrated liquid contains prebiotics such as polysaccharides, polyphenols, and ergothioneine. The multiple pores (12) can increase the surface area or filling capacity of the polysaccharides, polyphenols, and ergothioneine, thereby increasing the content of polysaccharides, polyphenols, and ergothioneine in each microcapsule structure. This invention intentionally selects the microcapsule structure so that the mushroom fruiting body particles, polysaccharides, polyphenols, and ergothioneine can be effectively released and the dissolution rate can be improved, which is beneficial to human absorption.

[0039] In one embodiment of the present invention, the core layer (11) can be an irregular structure, a sphere, an ellipsoid, a cylinder, or a polyhedron, and the irregular structure can be a structure with an irregular surface. The core layer (11) has a plurality of pores (12), and the pores (12) are interconnected, non-interconnected, partially interconnected, or partially non-interconnected. The adhesion layer (13) is formed by spraying a concentrated liquid onto part or all of the surfaces of both the core layer (11) and the pores (12), and part or all of the surfaces of the core layer (11) also include part or all of the surfaces of the pores (12), wherein the porous (12) and irregular surface structure of the core layer (11) can increase the adsorption capacity of the adhesion layer (13).

[0040] Please see Figure 2A As shown, in one embodiment of the present invention, the core layer (11) is a sphere and has a plurality of pores (12), which are not interconnected. Please refer to [further details omitted]. Figure 2B As shown, Figure 2B for Figure 1 A cross-sectional view along tangent A-A'. The core layer (11) forms an attachment layer (13) on the surface of its pores (12). The concentrated liquid contains prebiotics such as polysaccharides, polyphenols, and ergothioneine. Examples of the concentrated liquid, but not limited to, those extracted from dried powders of king oyster mushrooms, corn mushrooms, shiitake mushrooms, or enoki mushrooms, are included. The core layer (11) is formed from mushroom fruiting body particles, and examples of the core layer (11), but not limited to, those made from king oyster mushrooms or corn mushrooms, are included. Figure 3 (A) and Figure 3 As shown in (B), this is a magnified view of the core layer (11) and the ergothioneine microcapsule structure containing prebiotics observed using a scanning electron microscope (SEM), with a scale bar of 20.0 μm. Figure 3 In (A), the core layer (11) made from the fruiting body particles of *Pleurotus eryngii* was observed using a scanning electron microscope at a magnification of 2000x. The magnified image shows that the core layer (11) was freeze-dried to obtain an irregular structure and pores (12). The irregular structure is three-dimensional, and the pores (12) can be interconnected, disconnected, partially connected, or partially disconnected. Figure 3 In (B), a scanning electron microscope at 2000x magnification was used to observe the core layer (11) and the attachment layer (13) made of Pleurotus eryngii fruiting body particles in a single microcapsule structure. The magnified image shows that an attachment layer (13) forms on the surface of the core layer (11) and its pores (12). It is particularly noteworthy that the freeze-drying process of this invention reduces the destruction of the active ingredient structure in the core layer, thereby preserving its chemical structure and medicinal activity.

[0041] The ergothioneine microcapsule structure (1) containing probiotics can also be composed of multiple core layers (11) with the same or different particle sizes from multiple types of mushroom fruiting bodies. The microcapsules are sieved through 80-mesh and 60-mesh screens to obtain ergothioneine microcapsule structures with an average particle size of 180μm to 250μm, so as to indirectly control the weight ratio of the attachment layer (13) to the core layer (11) to be 1:0.5 to 1:2.2.

[0042] Please refer to this again. Figure 1 , Figure 2A and Figure 2BAs shown, based on its main objective, this invention also provides a microcapsule structure containing prebiotics, ergothioneine. This invention uses mushroom fruiting body particles as a core layer (11) and an adhesion layer (13) formed by spraying a concentrated liquid onto the surfaces of both the core layer (11) and the pores (12). The adhesion layer (13) covers the surfaces of both the core layer (11) and the pores (12). The concentrated liquid contains at least ergothioneine, therefore the adhesion layer (13) contains an ergothioneine layer formed from the aforementioned ergothioneine. Furthermore, the concentrated liquid may also contain prebiotics such as polysaccharides, polyphenols, and ergothioneine, therefore the adhesion layer (13) may contain a polysaccharide layer, a polyphenol layer, and an ergothioneine layer formed from polysaccharides, polyphenols, and ergothioneine, respectively. Please refer to [link to relevant documentation]. Figure 2C As shown, Figure 2C This is a cross-sectional view (II) of a preferred embodiment of the ergothioneine microcapsule structure containing prebiotics. The ergothioneine layer connects the polysaccharide layer and the polyphenol layer, and the ergothioneine, the polysaccharide layer, and the polyphenol layer are stacked. Multiple pores (12) can increase the surface area or filling capacity of the polysaccharides, polyphenols, and ergothioneine, thereby increasing the content of polysaccharides, polyphenols, and ergothioneine in each microcapsule structure. The present invention intentionally selects the microcapsule structure so that the mushroom fruiting body particles, polysaccharides, polyphenols, and ergothioneine can be effectively released and the dissolution rate can be improved, which is beneficial to human absorption.

[0043] In one embodiment of the present invention, the core layer (11) can be an irregular structure, a sphere, an ellipsoid, a cylinder, or a polyhedron, and the irregular structure can be a structure with an irregular surface. The core layer (11) has a plurality of pores (12), and the pores (12) are interconnected, non-interconnected, partially interconnected, or partially non-interconnected. The adhesion layer (13) is formed by spraying a concentrated liquid onto part or all of the surfaces of both the core layer (11) and the pores (12), so that the adhesion layer (13) covers part or all of the surfaces of both the core layer (11) and the pores (12). The part or all of the surface of the core layer (11) also includes part or all of the surface of the pores (12), wherein the porous structure (12) and irregular surface structure of the core layer (11) can increase the adsorption capacity of the adhesion layer (13).

[0044] Please see Figure 3 (A) and Figure 4As shown, based on its main objective, this invention also provides a method (2) for preparing a microcapsule structure containing prebiotics of ergothioneine, which is used to prepare a microcapsule structure containing prebiotics of ergothioneine. The preparation method includes the following steps: Step (a) providing a first mushroom fruiting body particle composed of at least one mushroom fruiting body particle as a core layer (11), the core layer (11) having multiple pores (12), and the mushroom fruiting body particle being prepared by sequentially passing a first mushroom fruiting body through a freeze-drying step and a dry grinding step, so that the core layer (11) is an irregular structure and has multiple pores (12) that are interconnected, non-interconnected, partially interconnected or partially non-interconnected. Please refer to [link to relevant documentation]. Figure 2A As shown. The freeze-drying step involves placing the first mushroom fruiting body in a vacuum at a freezing temperature of -20℃ to -80℃ to reduce residual liquid. Freeze-drying not only reduces the damage to the structure of the core layer (11), thus preserving the chemical structure and medicinal activity of the mushroom fruiting body particles, but also extends the shelf life for subsequent processing. The dry grinding step uses a dry grinding technology to grind the frozen first mushroom fruiting body into mushroom fruiting body particles. This dry grinding technology can generate particles through the high-speed impact of the first mushroom fruiting body and the blade, and can also discharge the ground particles using the centrifugal principle of a cyclone separator or an air jet mill. The core layer (11) can be made from, but is not limited to, king oyster mushrooms or corn mushrooms.

[0045] Please see Figure 3 (B) and Figure 4 As shown, in the preparation method of ergothioneine microcapsule structure containing prebiotics, step (b) involves spraying a concentrated liquid onto part or all of the surfaces of the core layer (11) and the pores (12) through a granulation step to form an adhesion layer (13). The porous structure (12) and irregular surface structure of the core layer (11) increase the adsorption capacity of the adhesion layer (13). The granulation step can be spray granulation, which can be performed using a spray dryer. The internal temperature and nozzle temperature of the spray dryer can be adjusted according to actual needs. Spray granulation not only yields free-flowing powder particles but also prevents the medicinal activity of the prebiotic-containing ergothioneine microcapsule structure from being destroyed during spray drying. The concentrated liquid contains prebiotics such as polysaccharides, polyphenols, and ergothioneine. The weight percentage of the adhering layer (13) to the core layer (11) is 1:0.5 to 1:2.2. In actual implementation, in addition to retaining a higher content of polysaccharides, polyphenols, and ergothioneine, the release time and concentration of these medicinal active ingredients in the human body are also maintained to achieve the maximum effect. The concentrated liquid can be exemplified by, but is not limited to, a concentrate extracted from dried powder of king oyster mushroom, corn mushroom, shiitake mushroom, or enoki mushroom.

[0046] In one embodiment of the present invention, the concentrated liquid is prepared by sequentially passing a second fungal fruiting body, consisting of at least one fungal fruiting body, through a second pulverizing step, an extraction step, and a concentration step.

[0047] The second pulverization step involves adding the second mushroom fruiting body to a solvent to form a mushroom mixture, followed by pulverization. The mushroom mixture, based on 100 parts by weight, includes 10-80 parts by weight of the mushroom fruiting body, with the remainder being solvent. The solvent can be pure water or a 20%-95% ethanol solution, preferably a 50%-70% ethanol solution. Pure water or ethanol are highly hydrophilic solvents. In practice, pure water or an ethanol solution of suitable concentration can be selected for extraction according to the different weight parts of the mushroom fruiting body. Examples of the second mushroom fruiting body include, but are not limited to, king oyster mushrooms, corn mushrooms, shiitake mushrooms, enoki mushrooms, or other mushrooms.

[0048] The extraction step involves extracting the pulverized mushroom mixture using either a hot water extraction step or an ultrasonic extraction step to obtain a first extract. The extraction time for this step is 0.5–24 hours. Hot water extraction is performed when the solvent is pure water, while ultrasonic extraction is performed when the solvent is ethanol. Both extraction steps involve heating the pulverized mushroom mixture at a temperature of 60°C–80°C for 0.5–24 hours to obtain a higher content of active ingredients. The first extract is then centrifuged to obtain a second extract. This centrifugation is performed at 4°C at 9000 rpm (Revolution(s) Per Minute) for 15 minutes. The supernatant is then collected; this supernatant is the second extract containing polysaccharides, polyphenols, and ergothioneine.

[0049] The concentration step involves concentrating the second extract through a vacuum concentration step or a heating concentration step to obtain the concentrated liquid. The vacuum concentration step and the heating concentration step are performed under specific pressure, temperature, and time conditions to remove excess solvent. The weight of the resulting concentrated liquid is 0.1 to 0.3 times the weight of the second mushroom fruiting body. This concentration step concentrates the second extract originally extracted from the second mushroom fruiting body to improve the content and concentration of effective components during the subsequent preparation of the attachment layer (13).

[0050] Based on its primary objective, this invention also provides an oral dosage form comprising multiple prebiotic-containing ergothioneine microcapsule structures (1). These prebiotic-containing ergothioneine microcapsule structures (1) are prepared by a method (2) for preparing prebiotic-containing ergothioneine microcapsule structures. This oral dosage form is selected from gelatin capsules and non-gelatin capsules. The materials for gelatin capsules can be selected from polyethylene glycol (PEG), sorbitol, glycerin, polypropylene glycol, and other polyols; the materials for non-gelatin capsules can be selected from starch, starch derivatives, cellulose, cellulose esters, cellulose ethers, etc. Cellulose nitrate, cellulose triacetate, cellulose phthalate acetate (CAP), methylcellulose, ethylcellulose, hydroxypropyl methylcellulose (HPMC), hydroxypropylcellulose (HPC), and hydroxypropyl methylcellulose phthalate (HPMCP) are all available options. In practice, the ergothioneine microcapsule structure containing prebiotics of the present invention can also be adapted to different types of oral dosage forms according to the user's needs. For example, considering concerns about allergies from animal sources, or the needs of religious or vegetarian consumers, non-gelatin capsule oral dosage forms can be used.

[0051] Based on its main objective, this invention also provides the use of a prebiotic-containing ergothioneine microcapsule structure in the preparation of ergothioneine microcapsules for improving constipation and clearing accumulated stool.

[0052] This trial recruited 24 volunteers with constipation problems, who were divided into an experimental group taking the ergothioneine microcapsule structure containing prebiotics as described in this invention, and a control group taking conventional mushroom powder capsules. The ergothioneine microcapsule structure of this invention consists of a core layer of mushroom fruiting body particles and an attachment layer formed by spraying a concentrated liquid onto the surface of this core layer. Conventional mushroom powder capsules consist of maltodextrin and concentrated mushroom liquid. The age distribution of the participants in each group was as follows: 2 men aged 40-50 years, 2 men aged 60-70 years, 4 women aged 40-50 years, and 4 women aged 60-70 years, totaling 12 participants. Before taking the capsules, the participants completed a gut age questionnaire (see Table 1) and their bowel movement data were collected. This gut age questionnaire was based on an assessment form created by Dr. Yoshimi Benno, Director of the Microbial Functional Analysis Laboratory at the RIKEN Institute in Japan. Next, this experiment lasted for four weeks. The capsules were taken after breakfast and dinner. The subjects in the experimental group took four ergothioneine capsules of this invention (500mg / capsule, each capsule containing multiple ergothioneine microcapsule structures containing prebiotics) every day, while the subjects in the control group took four ordinary mushroom powder capsules (500mg / capsule) every day. After four weeks of taking the capsules, the subjects in each group completed the intestinal age questionnaire according to their own physical sensations, and the relevant data on defecation were collected.

[0053] Table 1

[0054]

[0055]

[0056] The results of this experiment were obtained by statistically analyzing relevant data on defecation. Based on the purpose of this experiment, the following questions from the Intestinal Age Questionnaire were selected as the basis for assessing whether constipation had improved: (1) difficulty in defecation without straining, (2) feeling incomplete evacuation even after going to the toilet, (3) hard stools that are difficult to pass, and (4) stools that appear as small pellets. If the subject answered "yes" to any of the above questions, the number of "yes" answers was summed and a value was obtained according to the formula. The formula was the total number of times all subjects in each group answered "yes" to questions (1) to (4) / (number of subjects in each group x 4 questions). The degree of improvement was the value of the index before taking the capsules minus the value of the index after taking the capsules.

[0057] Test results: Please refer to Table 2 and... Figure 5 As shown in Table 2, the index values ​​and improvement rates of the experimental group and the control group before and after taking the capsules are presented. Figure 5 This is a graph showing the results of a questionnaire survey on bowel movements before and after taking the ergothioneine microcapsules containing prebiotics of this invention, compared to conventional mushroom powder capsules. The X-axis represents the bowel movements before and after taking the capsules, and the Y-axis represents the indicator values. The experimental results show that before taking the capsules, the indicator values ​​for the experimental group and the control group were 77% and 69%, respectively. After four weeks of taking the capsules, as... Figure 5 As shown in (A), the indicator values ​​of the control group after taking regular mushroom powder capsules remained at approximately 65%, while the improvement was only 4%. However, as... Figure 5 As shown in (B), the experimental group's index value decreased by 27%, and the improvement rate was 50%, indicating that the ergothioneine microcapsule structure containing prebiotics of this invention has a good effect on improving constipation and clearing accumulated stool. It should be noted that the functioning of the human digestive organs is affected not only by diet and lifestyle but also by aging with age, leading to constipation problems. However, the ergothioneine microcapsule structure containing prebiotics of this invention still has a good effect on improving constipation and clearing accumulated stool in men and women with an average age between 40 and 70 years old.

[0058] Table 2

[0059]

[0060] Next, in order to further understand the purpose, features, and desired effects of the present invention, the following describes the preparation method of the prebiotic-containing ergothioneine microcapsule structure mentioned in the present invention, to demonstrate that the prebiotic-containing ergothioneine microcapsule structure prepared by the present invention is rich in polysaccharides, polyphenols, and ergothioneine, and to further demonstrate the scope of practical application of the present invention, but is not intended to limit the scope of the present invention in any way.

[0061] Example 1: Using different types of mushrooms to prepare concentrated liquid and testing its ergothioneine content.

[0062] Preparation Process: Different types of mushrooms, including shiitake, enoki, king oyster, and corn mushrooms, are used as raw materials. 1 kg of dried powder from each type of mushroom is weighed and obtained. Then, 10 liters of 95% ethanol solution or pure water are added to each, and the mixture is homogenized for 5 minutes to obtain a homogenate. The homogenate is then extracted for 30 to 60 minutes using a food-grade ultrasonic extractor at a frequency of 35 kHz, a power of 180 W, and an extraction temperature of 60°C to obtain the first extract. The first extract is then centrifuged at 9000 rpm for 15 minutes at 4°C, and the supernatant is collected. This supernatant is the second extract from the shiitake, enoki, king oyster, and corn mushrooms. The second extract is then concentrated to 1 liter using a vacuum concentrator at a reduced pressure of 40°C to 60°C to obtain the concentrated liquid.

[0063] Ergothioneine content test:

[0064] Ergothioneine content in the concentrated liquid was analyzed using ultra-high pressure liquid chromatography (UPLC), and ergothioneine was quantified in each concentrated liquid using calibration curves prepared with ergothioneine standards of different concentrations. The analytical conditions for UPLC were as follows: System: Waters Acquity UPLC H-Class; Detector: Photodiode Array Detector, wavelength 254 nm; Column: HSS T3 (2.1 mm × 100 mm, 1.8 μm); Column temperature: 35 °C; Mobile phase solution: 0.1% formic acid aqueous solution; Flow rate: 0.3 mL / min; Sample injection volume: 10 μL.

[0065] Test results: such as Figure 6 As shown, Figure 6This figure presents experimental data on the preparation of concentrated liquids using different types of mushrooms and the testing of their ergothioneine content. The graph shows the ergothioneine content obtained after extraction with 95% ethanol solution or pure water, using various mushroom species including shiitake, enoki, king oyster, and corn mushrooms. The analysis results show that the ergothioneine content obtained by extraction with pure water is higher than that obtained with 95% ethanol solution. Furthermore, the concentrated liquid prepared from corn mushrooms has the highest ergothioneine content (approximately 2.3-5.0 g / L), followed by king oyster mushrooms (approximately 0.5-1.5 g / L), while enoki mushrooms (approximately 0.1-0.3 g / L) and shiitake mushrooms (approximately 0.01-0.07 g / L) have lower ergothioneine content.

[0066] Example 2: Analysis of ergothionein content in concentrated liquid prepared from fresh mushrooms using different concentrations and types of extraction solvents.

[0067] Preparation process: Fresh king oyster mushrooms (72% ± 3% moisture content) and fresh corn mushrooms (78% ± 3% moisture content) were used as raw materials. 1 kg of each type of fresh king oyster mushroom or fresh corn mushroom was weighed and then added to 2.5 liters of pure water, 50% ethanol solution, 70% ethanol solution, and 95% ethanol solution respectively as extraction solvents. The mixture was homogenized for 5 minutes to obtain a homogenate. The homogenate was then extracted for 30 to 60 minutes using a food-grade ultrasonic extractor at an ultrasonic frequency of 35 kHz, a power of 180 W, and an extraction temperature of 60°C to obtain the first extract. The first extract was then centrifuged at 9000 rpm for 15 minutes at 4°C. The supernatant was collected, which is the second extract of the fresh king oyster mushrooms or corn mushrooms. The second extract was passed through a vacuum concentrator at a vacuum temperature of 40°C to 60°C to obtain a concentrated liquid system of king oyster mushrooms that was 0.2 times the weight of the original fresh king oyster mushrooms, and the concentrated liquid obtained from corn mushrooms was concentrated to 0.15 times the weight of the original fresh corn mushrooms.

[0068] Ergothioneine content test:

[0069] Ergothioneine content in the concentrated liquid was analyzed using ultra-high pressure liquid chromatography (UPLC), and ergothioneine was quantified in each concentrated liquid using calibration curves prepared with ergothioneine standards of different concentrations. The analytical conditions for UPLC were as follows: System: Waters Acquity UPLC H-Class; Detector: Photodiode Array Detector, wavelength 254 nm; Column: HSS T3 (2.1 mm × 100 mm, 1.8 μm); Column temperature: 35 °C; Mobile phase solution: 0.1% formic acid aqueous solution; Flow rate: 0.3 mL / min; Sample injection volume: 10 μL.

[0070] Test Results: Table 3 shows the ergothioneine content obtained from extraction with different types of mushrooms, including fresh king oyster mushrooms and fresh corn mushrooms, using 2.5 liters of pure water, 50% ethanol solution, 70% ethanol solution, and 95% ethanol solution. The analysis results show that the ergothioneine content obtained using 50% ethanol solution and 70% ethanol solution is higher than that obtained using 95% ethanol solution and pure water.

[0071] Table 3:

[0072]

[0073] Example 3: Different types of mushrooms were used to prepare concentrated liquids and the contents of soluble solids, crude polysaccharides, and total polyphenols were tested.

[0074] Preparation process: Fresh king oyster mushrooms (72% ± 3% moisture content) and fresh corn mushrooms (78% ± 3% moisture content) were used as raw materials. 1 kg of each type of fresh king oyster mushroom or fresh corn mushroom was weighed and then 2.5 liters of 50% ethanol solution and 70% ethanol solution were added respectively as extraction solvents. The mixture was homogenized for 5 minutes to obtain a homogenate. The homogenate was then extracted for 30 to 60 minutes using a food-grade ultrasonic extractor at an ultrasonic frequency of 35 kHz, a power of 180 W, and an extraction temperature of 60°C to obtain the first extract. The first extract was then centrifuged at 9000 rpm for 15 minutes at 4°C. The supernatant was collected, which is the second extract of the fresh king oyster mushrooms or fresh corn mushrooms. The second extract was passed through a vacuum concentrator at a vacuum temperature of 40°C to 60°C to obtain a concentrated liquid system of king oyster mushrooms that was 0.2 times the weight of the original fresh king oyster mushrooms, and the concentrated liquid obtained from corn mushrooms was concentrated to 0.15 times the weight of the original fresh corn mushrooms.

[0075] Tests for soluble solids, crude polysaccharides, and total polyphenols:

[0076] 10 ml of concentrated liquids prepared from fresh king oyster mushrooms and fresh corn mushrooms were taken respectively and dried at 50°C to constant weight, which was used to determine their soluble solids content. The crude polysaccharide content was determined by analyzing the crude polysaccharide content of the concentrated liquids prepared from fresh king oyster mushrooms and fresh corn mushrooms using the phenol-sulfuric acid method.

[0077] Test results: As shown in Table 4, the concentrated liquid prepared from fresh king oyster mushrooms contained approximately 34.9±2.14 g / L of soluble solids, approximately 26.9±3.26 g / L of crude polysaccharides, and approximately 2.5±0.14 g / L of total polyphenols. In contrast, the concentrated liquid prepared from fresh corn mushrooms contained approximately 22.7±1.77 g / L of soluble solids, approximately 18.2±1.76 g / L of crude polysaccharides, and approximately 3.7±0.21 g / L of total polyphenols.

[0078] Table 4:

[0079]

[0080] Example 4: Different types of fresh mushroom fruiting bodies were used to prepare a porous core layer, and its yield, ergothioneine content, and crude polysaccharide content were tested.

[0081] Preparation process: Fresh king oyster mushrooms (moisture content 72% ± 3%) and fresh corn mushrooms (moisture content 78% ± 3%) were used as raw materials. 1 kg of fresh king oyster mushrooms or fresh corn mushrooms were weighed and obtained respectively. Then, the fresh king oyster mushrooms and fresh corn mushrooms were placed in a freezer at a temperature of -20℃ to -80℃ for 16 hours. The frozen fresh king oyster mushrooms and fresh corn mushrooms were then freeze-dried using a freeze dryer. After grinding with a dry grinder, the powder was sieved through an 80-mesh sieve, and the fruiting body particles (powder particle size less than 180μm) were collected as a porous core layer.

[0082] Yield, ergothionein, and crude polysaccharide content tests: Yield was determined by weighing the collected powder and calculating its percentage of the weight of 1 kg of fresh king oyster mushrooms or fresh corn mushrooms; crude polysaccharide content was analyzed by using the phenol-sulfuric acid method to analyze the content of crude polysaccharides in the concentrated liquids prepared from fresh king oyster mushrooms and fresh corn mushrooms; ergothionein content analysis was performed using ultra-high pressure liquid chromatography (UPLC) to analyze the ergothionein content in the obtained mushroom fruiting body particles, and the ergothionein in the mushroom fruiting body particles was quantified by using calibration curves prepared with ergothionein standards of different concentrations. The analytical conditions for the ultra-high pressure liquid chromatography (UHPLC) system were as follows: System: Waters acquity UPLC H-Class; Detector: Photodiode Array Detector, wavelength 254 nm; Column: HSS T3 (2.1 mm × 100 mm, 1.8 μm); Column temperature: 35 °C; Mobile phase solution: 0.1% formic acid aqueous solution; Flow rate: 0.3 mL / min; Sample injection volume: 10 μL.

[0083] Test results: The yield of fruiting body particles of *Pleurotus ostreatus* was 187.3±27.6 g / kg, and its ergothioneine and crude polysaccharide contents were 0.57±0.04% and 37.75±1.43%, respectively; while the yield of fruiting body particles of *Pleurotus ostreatus* was 262.2±41.1 g / kg, and its ergothioneine and crude polysaccharide contents were 0.39±0.31% and 39.85±1.3%, respectively.

[0084] Example 5: Preparation of Microcapsules

[0085] Preparation process: Fruiting body particles of king oyster mushroom or corn mushroom (powder particle size less than 180μm) are used as a porous core layer; the core layer powder is sprayed into the air through a granulation step, and then concentrated liquids prepared from king oyster mushroom and corn mushroom are respectively adhered to the core layer by spraying; then the powder is sieved through a 60-mesh sieve (powder particle size less than 250μm) to obtain the microcapsule structure.

[0086] Ergothioneine content determination: The ergothioneine content in the obtained microcapsule structures was analyzed using ultra-high pressure liquid chromatography (UPLC), and the ergothioneine in the microcapsule structures was quantified using calibration curves prepared with different concentrations of ergothioneine standards. The analytical conditions of the UPLC were as follows: System: Waters Acquity UPLC H-Class; Detector: Photodiode Array Detector, wavelength 254 nm; Column: HSS T3 (2.1 mm × 100 mm, 1.8 μm); Column temperature: 35 °C; Mobile phase solution: 0.1% formic acid aqueous solution; Flow rate: 0.3 mL / min; Sample injection volume: 10 μL

[0087] Test results: As shown in Table 5, with 1 part as 100 grams, Group 1: The microcapsule structure obtained by combining 1 part of *Mammillaria dichotoma* fruiting body particles with 1 part of *Pleurotus ostreatus* concentrated liquid has 0.69±0.03% ergothioneine; Group 2: The microcapsule structure obtained by combining 1 part of *Pleurotus ostreatus* fruiting body particles with 1 part of *Mammillaria dichotoma* concentrated liquid has 0.45±0.02% ergothioneine; Group 3: The microcapsule structure obtained by combining 2 parts of *Mammillaria dichotoma* fruiting body particles, 1 part of *Pleurotus ostreatus* fruiting body particles with 0.7 parts of *Mammillaria dichotoma* concentrated liquid and 0.7 parts of *Pleurotus ostreatus* concentrated liquid has 0.50±0.04% ergothioneine.

[0088] Table 5:

[0089]

[0090]

[0091] In summary, compared with existing technologies and products, the present invention has one of the following advantages:

[0092] One of the objectives of this invention is to address the inconvenience or excessive consumption of mushrooms in the past by using a microcapsule structure formed by a core layer of mushroom fruiting body particles and an attachment layer containing polysaccharides, polyphenols, and ergothioneine on the surface or pores of the core layer. This is achieved by intentionally increasing the surface area or filling capacity of the attachment layer and increasing the content of polysaccharides, polyphenols, and ergothioneine in each microcapsule structure.

[0093] One of the objectives of this invention is to provide the use of ergothioneine microcapsules for improving constipation and clearing accumulated stool. This invention uses a microcapsule structure consisting of a core layer formed by mushroom fruiting body particles and an attachment layer containing polysaccharides, polyphenols, and ergothioneine on the surface or pores of the core layer. This allows manufacturers to adjust the concentration of the pharmaceutically active ingredients in the microcapsules according to specific groups, frequency of administration, or intervals of administration to meet the needs of different physiological conditions at different ages.

[0094] One of the objectives of this invention is to purposefully select a specific preparation process sequence through a method for preparing ergothioneine microcapsules containing prebiotics, so that the weight ratio of the attachment layer to the core layer is controlled at 1:0.5 to 1:2.2. In addition to retaining a high content of polysaccharides, polyphenols and ergothioneine, this method also maintains the release time and concentration of these medicinal active ingredients in the human body and in the blood to achieve the maximum effect.

[0095] However, the above description is merely an embodiment of the present invention and should not be construed as limiting the scope of the present invention. Any simple equivalent changes and modifications made in accordance with the scope of the patent application and the contents of the patent specification shall still fall within the scope of the patent of the present invention.

[0096] Symbol Explanation

[0097] 1: Ergothioneine microcapsule structure containing prebiotics

[0098] 11: Core Layer

[0099] 12: Porosity

[0100] 13: Adhesion layer

[0101] a~b: Steps

[0102] 2: Preparation method of ergothioneine microcapsule structure containing prebiotics

Claims

1. A method for preparing a probiotic ergothioneine microcapsule structure (1) having an average particle size of 180 μm to 250 μm, the method comprising the steps of: (a) providing at least one first mushroom fruiting body particle as a core layer (11), each of the core layer (11) having a plurality of pores (12) that are in communication with each other, not in communication with each other, partially in communication with each other, or partially not in communication with each other, the core layer (11) comprising ergothioneine, the core layer (11) being an irregular structure, a sphere, an ellipsoid, a cylinder, or a polyhedron; (b) providing a concentrated liquid; and (c) providing a coating layer (13) on the core layer (11) to form the probiotic ergothioneine microcapsule structure (1), the coating layer (13) having a weight ratio of 1:0.5 to 1:2.2 relative to the core layer (11), the concentrated liquid comprising polysaccharides, polyphenols, and ergothioneine.

2. The method of claim 1, wherein the first mushroom fruiting body particle is prepared by sequentially subjecting a first mushroom fruiting body to a freeze-drying step and a dry grinding step, the first mushroom fruiting body being fresh Pleurotus eryngii or fresh Pleurocybella porosa, the first mushroom fruiting body particle having a particle size of less than 180 μm, the concentrated liquid is prepared by sequentially subjecting a second mushroom fruiting body to a second pulverization step, an extraction step, and a concentration step, the second mushroom fruiting body being fresh Pleurotus eryngii or fresh Pleurocybella porosa, and the freeze-drying step is performed by freezing the first mushroom fruiting body at a freezing temperature of -20 °C to -80 °C under vacuum. (b) forming an attachment layer (13) by spraying a concentrated liquid onto part or all of the surface of each of the core layer (11) and the aperture (12) through a granulation step, wherein, 3. The method of claim 2, wherein the extraction step is performed by subjecting the pulverized mushroom mixture to a hot water extraction step or an ultrasonic extraction step for 0.5 to 24 hours to obtain a first extraction liquid, and the first extraction liquid is subjected to a centrifugation step to collect a supernatant as a second extraction liquid.

4. The method of claim 3, wherein the concentration step is performed by subjecting the second extraction liquid to a reduced pressure concentration step or a heating concentration step to obtain the concentrated liquid, and a ratio of the weight of the concentrated liquid to the weight of the second mushroom fruiting body ranges from 0.1 to 0.

3.

2. The method for preparing ergothioneine microcapsule structures containing prebiotics as described in claim 1, wherein the second pulverizing step involves adding the second fungal fruiting body to a solvent to form a fungal mixture and then pulverizing it, wherein, 5. The method of claim 4, wherein the dry grinding step is performed by using a dry grinding technique to grind the frozen first mushroom fruiting body into the mushroom fruiting body particles.

6. The method of claim 5, wherein the mushroom fruiting body particles are further sieved through a first mesh screen having a mesh size of 80 mesh. ​ ​ ​ 7. The method of claim 6, wherein the probiotic-containing ergothioneine microcapsule structure (1) is further sieved through a second mesh screen having a mesh size of 60 mesh.

8. An oral dosage form comprising a plurality of probiotic-containing ergothioneine microcapsule structures (1) produced by the method of any one of claims 2 to 7, wherein the oral dosage form is selected from the group consisting of gelatin capsules and non-gelatin capsules.

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