Ginseng tissue culture adventitious root wall-broken pure powder tablet rich in rare ginsenoside and preparation method of ginseng tissue culture adventitious root wall-broken pure powder tablet

Abstract

The invention provides a ginseng tissue culture adventitious root wall-broken pure powder tablet rich in rare ginsenoside and a preparation method of the ginseng tissue culture adventitious root wall-broken pure powder tablet rich in rare ginsenoside. The method comprises the steps that fresh ginseng tissue culture adventitious roots are subjected to heat-moisture treatment for 60-100 min at the temperature of 110-121 DEG C and then treated through freeze-drying processes of pre-freezing, sublimation drying and desorption drying till the water content is smaller than 5%, and the ginseng tissue culture adventitious root wall-broken pure powder tablet rich in rare ginsenoside is obtained. Then wall-breaking and crushing at the low temperature of-20 to-30 DEG C until the particle size D90 is less than 20 microns, adding 0.1-1 per thousand of mogroside or stevioside, mixing, granulating by a dry method, and tabletting to obtain a finished product. The content of rare ginsenoside is remarkably increased through heat-moisture treatment, and the synergistic effect of freeze drying, low-temperature wall breaking and dry granulation is combined, so that the tablet is fine and smooth in taking, free of slag and granular feeling, free of radiation residues, only added with a trace amount of a natural sweetening agent, outstanding in pure natural property, suitable for large-scale production, high in efficacy and excellent in taste, and suitable for large-scale production. The marketization requirements of functional foods are met.

Description

Technical Field

[0001] This application relates to a ginseng tissue culture adventitious root cell wall-broken pure powder tablet rich in rare ginsenosides and its preparation method, belonging to the field of functional food technology. Background Technology

[0002] Ginseng (Panax ginseng CAMeyer), a perennial medicinal plant belonging to the genus Panax in the family Araliaceae, is mainly distributed in Northeast China, Korea, Japan, and other regions. It is a traditional and valuable food and medicine, with ginsenosides as its core active ingredient. Numerous types of ginsenosides have been identified, with Rh1, Rg2, Rb2, Ro, and Rc being common types. However, rare ginsenosides such as Rg3 and Rh2, due to their extremely low content, have long been underutilized. Compared to common ginsenosides, rare ginsenosides possess superior pharmacological activities, including anti-cancer properties, reduction of radiotherapy toxicity, enhancement of immunity, regulation of hormone levels, vasodilation, protection of brain nerve cells against dementia, and anti-platelet aggregation. Rg3, in particular, has been shown to have low toxicity and is recognized as a high-quality component combining medicinal and tonic effects. Therefore, increasing the content and utilization rate of rare ginsenosides in ginseng has become a research hotspot and important direction in the field of ginseng deep processing.

[0003] To overcome the limitations of natural ginseng resources, such as dependence on climate and soil conditions, susceptibility to pests and diseases, pesticide residue risks, long production cycles, and unstable quality, plant tissue culture technology has emerged. Ginseng tissue-cultured adventitious roots utilize the totipotency of plant cells, inducing callus formation through explants, and cultivating adventitious roots through processes such as proliferation and induction. Large-scale production using bioreactors offers advantages such as being unrestricted by the natural environment, free from viral contamination and pesticide residues, ensuring uniform quality, and a short production cycle. This provides a new path for the green, environmentally friendly, sustainable, and high-value production of ginseng, and has become an important development direction for the industrialization of ginseng raw materials.

[0004] Despite the significant advantages of ginseng tissue culture adventitious roots, efficiently increasing their content of rare ginsenosides and preparing products that meet market demands remains a key technical challenge. Existing technologies for improving ginsenoside hydrolysis efficiency and increasing rare ginsenoside content mainly include enzymatic methods, acid-base methods, microwave methods, and ultrasonic methods. While these methods can achieve the desired results to some extent, they generally suffer from drawbacks such as demanding reaction conditions, numerous byproducts, complex operating procedures, and high production costs, hindering their industrial-scale application. Furthermore, Chinese patent application CN119563871A – "A Ginseng Product with High Rare Saponin Content and Its Preparation Method" – discloses a method of irradiating fresh ginseng with Co-60 γ-rays followed by steaming. While this method can increase the rare ginsenoside content, the irradiation treatment can easily lead to residual radiation in the product, posing safety hazards and limiting its application in the food industry.

[0005] Another Chinese patent application, CN119908294A, describes a method for converting ginsenosides in ginseng adventitious roots. This method uses a bottom-addition of water under high temperature and pressure for 2-5 hours to improve the conversion rate and increase the content of rare ginsenosides. However, it only focuses on the conversion efficiency of rare ginsenosides. If prolonged high-temperature treatment is used, it can easily lead to the accumulation of bitter substances and the loss of natural aroma, making it difficult to use directly in finished food products.

[0006] Meanwhile, in the manufacturing process of ginseng tablets, regardless of whether ginseng powder, ginseng extract, or ginseng extract powder is used as raw material, excipients are generally added to improve certain properties of the powder, extract, or extract powder, ensuring that the compressed tablets meet relevant quality requirements. However, the addition of excipients not only destroys the product's natural properties but may also affect the absorption and utilization of ginseng's active ingredients.

[0007] Therefore, developing a ginseng tissue culture adventitious root product that does not require the addition of large amounts of excipients, has no risk of radiation residue, can effectively increase the content of rare ginsenosides, has a good taste, and is easy to carry and consume has become an urgent technical need in the field of functional food technology, and is of great significance to promoting the high-quality development of the ginseng deep processing industry. Summary of the Invention

[0008] To address the aforementioned issues, a method for preparing ginseng tissue culture adventitious root cell-wall-broken pure powder tablets rich in rare ginsenosides is provided. Through a synergistic process of wet heat treatment, freeze-drying, low-temperature cell-wall-breaking pulverization, and dry granulation, the content of rare ginsenosides is significantly increased. The product has a delicate texture without any grainy feel, moderate sweetness, no obvious bitterness, and a good overall taste. It retains the natural aroma and active ingredients of ginseng, has no radiation residue, requires only trace amounts of natural sweeteners, eliminates the need for large amounts of excipients, has a controllable production process, and ensures stable product quality, making it suitable for sublingual administration and large-scale production.

[0009] According to one aspect of this application, a method for preparing pure powder tablets of ginseng tissue-cultured adventitious roots rich in rare ginsenosides is provided, comprising the following steps: S1. Pretreatment: The fresh adventitious roots of ginseng tissue culture are subjected to moist heat treatment at 110-121℃ for 60-100 minutes; S2. Freeze-drying: Adventitious roots of ginseng tissue culture treated with S1 are subjected to freeze-drying and double drying until the moisture content is <5%; S3. Cell wall breaking and pulverization: Pulverize the dried ginseng tissue culture adventitious roots; S4. Mixing: Add natural sweetener to the pulverized powder and mix well; S5. Dry granulation: Dry granulation is performed using a dry granulator; S6. Tableting: The tablets are prepared by using a tablet press to obtain the pure powder tablets of ginseng tissue culture adventitious roots rich in rare ginsenosides.

[0010] Optionally, in step S1, the wet heat treatment is carried out in a saturated steam environment with a pressure of 0.14-0.20 MPa.

[0011] Specifically, the wet heat treatment facilitates the hydrolysis of glycosidic bonds in common ginsenosides (highly glycosylated precursors), thereby effectively increasing the content of rare ginsenosides. The high-pressure wet heat environment causes a phase transition in the phospholipid bilayer of ginseng cell walls and increases permeability. At the same time, the steam condensation water swells the pectin components, leading to the loosening and breakage of the fiber structure, and the full release of precursor saponins such as Rb1 and Rd from the cells.

[0012] At high temperatures, glycosidic bonds become unstable and undergo hydrolysis, dehydration, and cyclization reactions, thereby converting native saponins (such as Rb1, Rc, Rd, etc.) into rare saponins (such as Rg3, Rk1, Rg5, etc.). Additionally, the trace organic acids present in ginseng tissue culture adventitious root tissues or culture media can provide a small amount of H+ at high temperatures. + This further promotes the acid-catalyzed hydrolysis of glycosidic bonds and accelerates the conversion process.

[0013] Optionally, in step S2, the freezing temperature is -30°C to -45°C, and the time is 2-5 hours.

[0014] Specifically, by using a specific temperature freezing process, the moisture in the material can be quickly frozen into small ice crystals, avoiding the damage to the ginseng fiber structure and active ingredients caused by the large ice crystals formed by slow freezing. If the freezing temperature is too high or the time is insufficient, the unfrozen free water will lead to incomplete drying, and excessive moisture will affect the stability of the product.

[0015] Optionally, in step S2, the dual drying is sublimation drying and desorption drying; the parameters for sublimation drying are: vacuum degree <100Pa, temperature 20-30℃, and drying time 10-15h; the parameters for desorption drying are: temperature 40-60℃ and drying time 2-5h.

[0016] Specifically, sublimation drying is carried out under high vacuum and low temperature conditions, which allows ice crystals to directly sublimate into water vapor, avoiding material shrinkage, loss of active ingredients and generation of bitter substances caused by liquid water evaporation. The time limit ensures that more than 90% of free water is removed. Then, analytical drying is used to specifically remove the bound water adsorbed inside the material, reducing the moisture content to <5%, which significantly improves the product's storage stability and prevents mold and saponin degradation.

[0017] The two processes work synergistically to achieve deep moisture removal while fully preserving the natural aroma and rare saponin activity of ginseng. Compared to a single drying process, the material has a fuller texture and more brittle fibers, providing a high-quality material foundation for subsequent cell wall breaking and pulverization.

[0018] Optionally, in step S3, the pulverization is low-temperature cell wall breaking pulverization, with a pulverization temperature of -20℃ to -30℃ and a pulverization time of 60-120min; the particle size D90 of the powder is <20um.

[0019] Specifically, the low temperature limit makes the ginseng fiber structure more brittle, reducing the energy consumption of grinding, while avoiding the oxidation and decomposition of saponins caused by the heat generated during grinding; the limited grinding time ensures that the material is fully broken down, and the ultra-fine powder standard with a particle size D90 < 20um not only ensures that there is no grainy feeling when taking it, but also improves the absorption and utilization rate of effective ingredients through the large specific surface area.

[0020] The low-temperature cell wall breaking pulverization and the previous freeze-drying work synergistically. After freeze-drying, the fiber of the material becomes brittle. Combined with low-temperature pulverization, the cell wall breaking rate can reach >99.9%, which is much higher than that of conventional pulverization processes, further enhancing the release and absorption of effective ingredients.

[0021] Optionally, in step S4, the natural sweetener is one or more of mogrosides and steviol glycosides, and the amount added is 0.1-1‰; the mixing speed is 15-30 rpm, and the mixing time is 10-30 min.

[0022] Monk fruit glycosides and steviol glycosides are natural, low-calorie sweeteners with high safety. Adding even a tiny amount precisely neutralizes the bitterness of ginseng without masking its natural aroma, aligning with modern health-conscious consumer trends. This trace addition does not affect saponin content or stability, while significantly improving palatability, resulting in high consumer preference and addressing the pain point of the bitter taste in traditional ginseng products.

[0023] Optionally, in step S5, the parameters for dry granulation are: roller pressure 20-80 KN / cm, roller gap 1.0-3.0 mm, tube wheel speed 5-10 rpm, and screen aperture 1-3 mm.

[0024] Granules produced by traditional wet granulation or fluidized bed granulation often have a heavy, unrefined feel during sublingual administration after compression. This is mainly because the large specific surface area of ​​ultrafine powders leads to extremely strong capillary forces and numerous liquid bridges when water or alcohol of varying concentrations is added. During subsequent drying, these strong capillary forces tightly bind the particles together, forming a dense structure. Simultaneously, after water evaporation, numerous rigid solid bridges remain between the particles, formed by the crystallization of water-soluble substances or the close contact of the particles. Water or alcohol of varying concentrations, acting as low-viscosity binders, cannot provide a flexible bonding network; their bonding mechanism naturally tends to form a rigid structure under ultrafine powder conditions.

[0025] The dry granulation method in this application avoids the problem of water / alcohol forming liquid bridges with ultrafine powder in wet granulation, which leads to the formation of a dense and rigid structure after drying. The resulting granules maintain an ultrafine particle size after disintegration. In conjunction with low-temperature cell wall breaking and pulverization, it ensures a delicate taste. At the same time, no subsequent drying step is required, which shortens the production cycle and reduces energy consumption.

[0026] Optionally, in step S6, the hardness of the tablets after compression is 50-150 N.

[0027] According to another aspect of this application, a pure powder tablet of ginseng tissue culture adventitious root with broken cell walls, rich in rare ginsenosides, is also provided. It is prepared by any of the above-described preparation methods, with ginseng tissue culture adventitious root as the sole raw material matrix, and contains 0.1-1‰ of natural sweetener.

[0028] Optionally, the ginseng tissue culture adventitious root cell wall-breaking pure powder tablets have a total saponin content ≥8.8%, a rare ginsenoside content ≥4.0%, and a particle size D90 <20um.

[0029] The beneficial effects of this application include, but are not limited to: 1. This application involves subjecting ginseng tissue culture adventitious roots to specific wet heat treatment, which significantly increases the content of rare ginsenosides while maintaining a stable total saponin content. Compared to enzymatic and acid-base methods, this process is simple, has mild reaction conditions, and produces no byproducts. Compared to Co-60 gamma ray irradiation, it completely avoids the risk of radiation residue, resulting in a safer and more compliant product that is more suitable for application in the functional food field.

[0030] 2. The product of this application achieves a breakthrough optimization in taste. By using freeze-drying to inhibit the Maillard reaction, the formation of bitter substances is reduced while preserving the natural aroma of ginseng. At the same time, low-temperature cell wall breaking and pulverization to D90 < 20um, combined with dry granulation to avoid liquid bridging effect, makes the product delicate and free of granules when taken orally. In addition, a trace amount of natural sweetener is added to precisely neutralize the bitterness, resulting in a moderate sweetness. This solves the core pain point of the rough and bitter taste of traditional ginseng tablets, and significantly improves consumer acceptance.

[0031] 3. This application uses ginseng tissue culture adventitious roots as the sole raw material matrix, with only trace amounts of natural sweeteners added. No excipients, binders, or other auxiliary materials are required, thus preserving the natural and pure properties of the product. The high cell wall breakage rate and ultra-fine powder form facilitate an increase in the contact area between the active ingredients and the human intestines, promoting absorption and utilization. Compared to products without cell wall breakage or with added excipients, the efficacy is more direct and efficient.

[0032] 4. The parameters of each step in the preparation process of this application are clear and highly controllable, making it suitable for industrial mass production; the freeze-drying process ensures that the product moisture content is less than 5%, making it less prone to mold growth and degradation of active ingredients during storage, significantly extending the shelf life, and resulting in product quality stability that is superior to existing similar products. Attached Figure Description

[0033] The accompanying drawings, which are included to provide a further understanding of this application and form part of this application, illustrate exemplary embodiments and are used to explain this application, but do not constitute an undue limitation of this application. In the drawings: Figure 1 This is a scanning electron microscope image of the pure powder of ginseng tissue culture adventitious roots after cell wall disruption in Example 1 of this application (1000x view on the left and 500x view on the right). Figure 2 This is a particle size distribution diagram of the ginseng tissue culture adventitious roots after cell wall disruption and pulverization in Example 1 of this application. Detailed Implementation

[0034] The present application is described in detail below with reference to the embodiments, but the present application is not limited to these embodiments.

[0035] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of skill in the art. The reagents and raw materials used in this invention are readily available through conventional means, and unless otherwise specified, they shall be used in accordance with conventional methods in the art or as per the product instructions. Furthermore, any methods and materials similar to or equivalent to those described herein may be applied to the methods of this invention. The preferred embodiments and materials described in this patent are for illustrative purposes only.

[0036] The detection of rare ginsenosides referred to in this invention covers 22 specific components, namely: 20(s) ginsenoside Rg3, 20(R) ginsenoside Rg3, (s-type) ginsenoside Rh2, (R-type) ginsenoside Rh2, ginsenoside Rk1, ginsenoside Rg5, ginsenoside CK, ginsenoside Rf, ginsenoside Rc, ginsenoside Rb2, ginsenoside Rb3, ginsenoside F3, ginsenoside Rg2, ginsenoside Rh1, ginsenoside Rd, ginsenoside F1, ginsenoside Fe, ginsenoside F2, ginsenoside Rg6, ginsenoside Rk3, ginsenoside Rk2, and ginsenoside Rh3.

[0037] Among them, the contents of six components—20(s) ginsenoside Rg3, 20(R) ginsenoside Rg3, (s-type) ginsenoside Rh2, (R-type) ginsenoside Rh2, ginsenoside Rk1, and ginsenoside Rg5—account for more than 80% of the total content of rare ginsenosides and are the core components for efficacy. The fresh adventitious roots of ginseng tissue culture were purchased from Shandong Kedilian Plant Stem Cell Co., Ltd.

[0038] Example 1 S1 Pretreatment: Fresh adventitious roots of cultured ginseng tissue culture were placed in a moist heat sterilizer and treated with saturated steam pressure of 0.20 MPa and 121°C for 60 min.

[0039] S2 Freeze-drying: Freeze-drying is adopted, with a pre-freezing temperature of -30℃ and a time of 2 hours, a sublimation drying temperature of 20℃ and a time of 12 hours, and a desorption drying temperature of 50℃ and a time of 2 hours, until the moisture content is <5%.

[0040] S3 Cell Wall Crushing: Crushed using a vibratory mill at a temperature of -30℃ for 60 minutes, resulting in a powder particle size D90 < 20 μm.

[0041] S4 Mixing: Add 0.5‰ of mogroside and mix using a three-dimensional mixer at 15 rpm for 30 minutes.

[0042] S5 Dry Granulation: Adjust the process parameters of the dry granulator to 30KN / cm between rollers, 1.5mm between rollers, 5rpm for tube wheel speed, and 2mm for screen aperture to perform dry granulation.

[0043] S6 tableting: Adjust the tableting machine to control the tablet hardness to 70N.

[0044] Example 2 S1 Pretreatment: The cultured ginseng tissue culture adventitious roots were placed in a moist heat sterilizer and treated with saturated steam pressure of 0.17 MPa and 115°C for 90 min.

[0045] S2 Freeze-drying: Freeze-drying is adopted, with a pre-freezing temperature of -30℃ for 2 hours, a sublimation drying temperature of 20℃ for 12 hours, and a desorption drying temperature of 50℃ for 2 hours, until the moisture content is less than 5%.

[0046] S3 Cell Wall Crushing: Crushed using a vibratory mill at a temperature of -30℃ for 60 minutes, resulting in a powder particle size D90 < 20 μm.

[0047] S4 Mixing: Add 0.5‰ of mogrosides and mix using a three-dimensional mixer at 15 rpm for 30 minutes.

[0048] S5 Dry Granulation: Adjust the process parameters of the dry granulator to 50KN / cm between rollers, 1.5mm between rollers, 5rpm for tube wheel speed, and 2mm for screen aperture to perform dry granulation.

[0049] S6 tableting: Adjust the tableting machine to control the tablet hardness to 90N.

[0050] Example 3 S1 Pretreatment: The cultured ginseng tissue culture adventitious roots were placed in a moist heat sterilizer and treated with saturated steam pressure of 0.14 MPa, 110°C, for 100 min.

[0051] S2 Freeze-drying: Freeze-drying is adopted, with a pre-freezing temperature of -30℃ for 2 hours, a sublimation drying temperature of 20℃ for 12 hours, and a desorption drying temperature of 50℃ for 2 hours, until the moisture content is less than 5%.

[0052] S3 Cell Wall Breaking and Grinding: Grinding is carried out using a vibratory mill at a temperature of -30℃ for 60 minutes.

[0053] S4 Mixing: Add 0.7‰ steviol glycosides and mix using a three-dimensional mixer at 15 rpm for 30 min.

[0054] S5 Dry Granulation: Adjust the process parameters of the dry granulator to 80KN / cm between rollers, 1.5mm between rollers, 5rpm for tube wheel speed, and 2mm for screen aperture to perform dry granulation.

[0055] S6 tableting: Adjust the tableting machine to control the tablet hardness to 100N.

[0056] Comparative Example 1 The difference between Comparative Example 1 and Example 1 is that the pretreatment temperature in step S1 is 100°C and the time is 120 min, while the other contents are the same as in Example 1.

[0057] Comparative Example 2 The difference between Comparative Example 2 and Example 1 is in step S5 granulation: wet granulation is used, 50% ethanol is added and wet-mixed for 3 minutes, a 2mm nylon screen is installed on the oscillating granulator, and the granules are dried at 55°C until the moisture content is <5%. The granules are then sized. All other contents are the same as in Example 1.

[0058] Comparative Example 3 The difference between Comparative Example 3 and Example 1 is that no mogrosides are added during the mixing process in step S4; otherwise, the contents are the same as in Example 1.

[0059] Comparative Example 4 The difference between Comparative Example 4 and Example 1 is that step S2 uses hot air circulation drying, the drying temperature is 50°C, the drying time is 36 hours, until the moisture content is <5%, and the other contents are the same as Example 1.

[0060] Comparative Example 5 The difference between Comparative Example 5 and Example 1 is that the pretreatment time in step S1 is 3 hours, while the rest is the same as Example 1.

[0061] Experimental Example The products prepared in Examples 1-3 and Comparative Examples 1-4 were tested according to the following testing methods, and the results are shown in Table 1.

[0062] 1. Particle size detection method: Take one tablet of ginseng tissue culture adventitious root after compression, place it in 50mL of purified water to disintegrate naturally, and detect the particle size (D90) according to Appendix B of GB / T 34778-2017 "Matcha".

[0063] 2. Detection method for total ginsenosides: The determination was carried out in accordance with the standard method of GB / T 19506-2009 "Geographical Indication Product Jilin Changbai Mountain Ginseng".

[0064] 3. Detection Methods for Rare Ginsenoside Content 3.1 Principle After extraction and other pretreatments, the samples were separated by a C18 column, detected by high performance liquid chromatography-ultraviolet detector, and the content of each component of ginsenosides was quantitatively determined by external standard method.

[0065] 3.2 Reagents and Materials Unless otherwise stated, all reagents used in this method are of analytical grade, and the water is Grade I water as specified in GB / T6682.

[0066] 3.2.1 Reagents Acetonitrile and methanol were both of chromatographic grade. 3.2.2 Preparation of reagents 70% methanol solution: Measure 70 mL of methanol, dilute with water and bring the volume to 100 mL.

[0067] 3.2.3 Standard Products Preparation of stock solution (0.8 mg / ml): Weigh out 8.00 mg of each of the following ginsenosides: Re, Rg1, Rb1, Rf, Rc, Rb2, Rb3, F3, Rg2, Rh1, Rd, F1, Fe, F2, and F3. Twenty-four standards, including Rg6, ginsenoside Rk3, 20(s) ginsenoside Rg3, 20(R) ginsenoside Rg3, ginsenoside Rk1, ginsenoside Rg5, ginsenoside CK, (s-type) ginsenoside Rh2, (R-type) ginsenoside Rh2, ginsenoside Rk2, ginsenoside Rh3, and protopanaxadiol, were placed in a 10ml volumetric flask, sonicated for 30 minutes, and then diluted to volume with methanol.

[0068] Preparation of working solution (32ug / ml): Accurately pipette 1ml of stock solution (0.8mg / ml) into a 25ml volumetric flask, dilute to volume with methanol, and filter through a 0.22um filter membrane for later use.

[0069] 3.3 Instruments and Equipment High performance liquid chromatograph: with ultraviolet detector.

[0070] 3.4 Analysis Steps 3.4.1 Sample Preparation Accurately weigh 50 mg into a 10 mL volumetric flask, add 70% methanol solution, and dilute to volume. Sonicate for 30 minutes, then filter through a 0.22 μm filter membrane for later use.

[0071] 3.4.2 Instrument Reference Conditions A) Chromatographic column: C18 column, column length 250 mm, column inner diameter 4.6 mm, column packing particle size 5 μm, or equivalent; B) Mobile phase: a: acetonitrile; b: water, filtered through a 0.45 μm microporous membrane; C) The gradient elution program is as follows: 0–20 min (23%–58.5% acetonitrile, 0.7 mL / min), 20–33 min (58.5%–78% acetonitrile, 0.7 mL / min), 33–34 min (78%–79.5% acetonitrile, 1.0 mL / min), 34–41 min (79.5%–90% acetonitrile, 1.0 mL / min), 41–46 min (90%–100% acetonitrile, 1.0 mL / min), 46–56 min (100% acetonitrile, 1.0 mL / min), 56–62 min (23% acetonitrile, 1.0 mL / min), and 62–63 min (23% acetonitrile, 0.7 mL / min). D) Column temperature: 30℃; E) Detection wavelength: 203nm; F) Injection volume: 10 μL.

[0072] 3.5 Presentation of Analysis Results The content of each component of ginsenosides in the sample was calculated according to formula (1): X= (1) In the formula: X — The content of each component of ginsenosides in the sample, in milligrams per kilogram (mg / kg) or milligrams per liter (mg / L); A1—Peak area of ​​each ginsenoside component in the sample A2 – Peak area of ​​each ginsenoside component in the standard. ρ — Concentration of each ginsenoside component in the standard (ug / ml) V — Final volume of the sample solution, in milliliters (mL); M — Sample mass, in grams (g); The content of ginsenosides in the sample is obtained by adding up the components.

[0073] 4. Sensory evaluation methods Ten assessors trained and qualified according to GB / T 16291.1-2012 standard were selected to conduct blind tests at room temperature of 23±2℃: each person took one sample, held it in their mouth for 60 seconds, spat it out, rinsed their mouth and rested for 5 minutes before tasting the next sample. The samples were scored according to the following dimensions, and the results are expressed as the arithmetic mean (accurate to one decimal place): Scoring criteria: Fineness: using a 5-point scale (1 = noticeable graininess, 5 = no graininess); Taste: Rate according to the intensity of sweetness preference (1 = very strong, 5 = none); Acceptance (liking): A 5-point acceptance scale is used (1 = strong aversion, 5 = strong liking).

[0074] 5. Bitterness Detection Experiment - Method for Determining the Content of Bitter Substances (5-Hydroxymethylfurfural, HMF) 5.1 Principle HMF in the sample was extracted with 70% methanol, separated by a C18 column, detected by high performance liquid chromatography-ultraviolet detector at a wavelength of 284 nm, and quantified by external standard method.

[0075] 5.2 Reagents and Materials Reagents: Acetonitrile, methanol (chromatographic grade); 70% methanol solution (same as 3.2.2); Standard: HMF standard (purity ≥99.0%); Stock solution preparation (1.0 mg / mL): Weigh 10.0 mg of HMF standard into a 10 mL volumetric flask, dilute to volume with methanol, and sonicate for 10 minutes; Working solution preparation (20 μg / mL): Accurately pipette 0.2 mL of stock solution into a 10 mL volumetric flask, dilute to volume with methanol, and filter through a 0.22 μm filter membrane for later use.

[0076] 5.3 Instruments and Equipment High-performance liquid chromatograph (with UV detector), electronic balance (accuracy 0.01 mg), ultrasonic cleaner.

[0077] 5.4 Analysis Steps Sample preparation: Accurately weigh 50 mg of sample into a 10 mL volumetric flask, add 70% methanol solution to make up to volume, extract by ultrasonication for 20 minutes, and filter through a 0.22 μm filter membrane for later use; Instrument reference conditions: chromatographic column: C18 column (column length 250 mm, column inner diameter 4.6 mm, column packing particle size 5 μm); mobile phase: acetonitrile:water = 20:80 (volume ratio); flow rate 1.0 mL / min; column temperature 30 ℃; detection wavelength 284 nm; injection volume 10 μL.

[0078] 5.5 Result Calculation Quantification was performed using the external standard method, with units expressed in mg / g.

[0079] Table 1. Physicochemical indicators and taste evaluation results of each embodiment and comparative example.

[0080] First, by Figure 1 As can be seen, in the 1000x view on the left, the particles are irregularly fragmented, with no dense, rigid agglomeration structure on the surface, and the overall morphology is loose and porous; there is no obvious complete cell wall outline, which proves that the cell wall breaking effect is good; in the 500x view on the right, the particles are evenly distributed, and the particle size is concentrated in the range of 2-10μm; there are no large-sized clumps, indicating that the process effectively controls the particle size of the cell wall breaking and pulverization.

[0081] Depend on Figure 2 It can be seen that the particle size distribution after cell wall breaking and pulverization is effectively controlled within D90 < 20 μm. The smaller the distribution span value, the more concentrated the particle size distribution. The span of this product = 2.912 indicates that the particle size distribution is relatively narrow and there is no obvious agglomeration of large particles, reflecting that the low temperature cell wall breaking and pulverization process has good stability and uniformity.

[0082] Furthermore, by comparing Examples 1-3 and Comparative Example 1, the wet heat treatment at 110-121℃ for 60-120 min as specified in this application, compared with the treatment at 100℃ for 120 min, resulted in a significant increase in the content of rare ginsenosides to 4.79%-4.91% while maintaining a relatively stable total saponin content (8.83%-8.92%). This represents an increase of over 200% in the content of rare ginsenosides, resulting in a marked improvement in the efficacy of ginseng tissue culture adventitious root slices and a lower content of bitter substances.

[0083] By comparing Comparative Example 2 with Example 1, the dry granulation method of this application avoids the rigid agglomeration of particles caused by the liquid bridge effect in wet granulation (the particle size D90 of Comparative Example 2 reaches 50μm). The particle size D90 of the products in the examples is all <20μm, and the fineness score reaches 4.5-4.6 points, which is significantly better than the 1.5 points of Comparative Example 2. The products obtained by this application have a more delicate taste and are more popular with people.

[0084] Comparing Comparative Example 3 with Example 1, the addition of mogrosides had virtually no effect on particle size and saponin content. However, the trace amount of addition significantly improved the taste of ginseng tissue culture adventitious root slices, making them more appealing to consumers.

[0085] Comparing Comparative Example 4 with Example 1, the ginseng tissue culture adventitious root slices obtained by drying in a hot air circulating oven had a bitter taste. The freeze-drying and double drying methods specified in this application avoid the Maillard reaction caused by hot air drying (Comparative Example 4 had a bitter taste, score 3.1). The product of this application has a better taste and significantly higher preference. At the same time, the material fibers are more brittle after freeze-drying, which is conducive to subsequent cell wall breaking and pulverization, ensuring particle size uniformity. Although Comparative Example 5 (pretreatment for 3 hours) further increased the content of rare ginsenosides, the HMF content increased significantly, and the bitterness affected the taste of the final product, resulting in a decrease in overall preference.

[0086] In summary, this invention, through various synergistic processes, successfully prepared ginseng tissue culture adventitious root cell wall-broken pure powder tablets with high content of rare ginsenosides, delicate taste, moderate sweetness, and suitable for sublingual administration. The technical solution is feasible and easy to industrialize.

[0087] The above description is merely a preferred embodiment of this application and is not intended to limit this application. Any modifications, equivalent substitutions, and improvements made within the spirit and principles of this application should be included within the protection scope of this application.

Claims

1. A method for preparing a broken cell wall pure powder tablet of a Panax ginseng tissue culture adventitious root rich in rare ginsenosides, characterized in that, Includes the following steps: S1. Pretreatment: The fresh adventitious roots of ginseng tissue culture are subjected to moist heat treatment at 110-121℃ for 60-100 minutes; S2. Freeze-drying: Adventitious roots of ginseng tissue culture treated with S1 are subjected to freeze-drying and double drying until the moisture content is <5%; S3. Cell wall breaking and pulverization: Pulverize the dried ginseng tissue culture adventitious roots; S4. Mixing: Add natural sweetener to the pulverized powder and mix well; S5. Dry granulation: Dry granulation is performed using a dry granulator; S6. Tableting: The tablets are prepared by using a tablet press to obtain the pure powder tablets of ginseng tissue culture adventitious roots rich in rare ginsenosides.

2. The production method according to claim 1, characterized by, In step S1, the wet heat treatment is carried out in a saturated steam environment with a pressure of 0.14-0.20 MPa.

3. The preparation method according to claim 1, characterized in that, In step S2, the freezing temperature is -30℃ to -45℃, and the time is 2-5 hours.

4. The production method according to claim 1, characterized by, In step S2, the double drying process consists of sublimation drying and desorption drying; The parameters for the sublimation drying are: vacuum degree <100Pa, temperature 20-30℃, and drying time 10-15h; the parameters for the analytical drying are: temperature 40-60℃ and drying time 2-5h.

5. The preparation method according to claim 1, characterized in that, In step S3, the pulverization is low-temperature cell wall breaking pulverization, with a pulverization temperature of -20℃ to -30℃ and a pulverization time of 60-120 min; The particle size of the powder, D90, is less than 20 μm.

6. The method of claim 1, wherein, In step S4, the natural sweetener is one or more of mogrosides and steviol glycosides, and the amount added is 0.1-1‰. Mix at 15-30 rpm for 10-30 minutes.

7. The preparation method according to claim 1, characterized in that, In step S5, the parameters for dry granulation are: roller pressure 20-80KN / cm, roller gap 1.0-3.0mm, tube wheel speed 5-10rpm, and screen aperture 1-3mm.

8. The method of claim 1, wherein, In step S6, the hardness of the tablets after compression is 50-150N.

9. A broken cell wall pure powder tablet of tissue culture adventitious root of Panax ginseng rich in rare ginsenoside, characterized in that, Prepared by any of the preparation methods described in claims 1-8, using ginseng tissue culture adventitious roots as the sole raw material matrix, and containing 0.1-1‰ of a natural sweetener.

10. The ginseng tissue culture adventitious root cell wall-breaking pure powder tablets according to claim 9, characterized in that, Its total saponin content is ≥8.8%, rare ginsenoside content is ≥4.79%, and particle size D90 <20um.

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