A polysaccharide composition, nano-polysaccharide and preparation method and application thereof

Abstract

The present application relates to the technical field of medicine, in particular to a polysaccharide composition, nano-polysaccharide and its preparation method and application.The polysaccharide composition comprises polysaccharide and lipid material;the weight ratio of polysaccharide to lipid material is 2-50:1.The composition disclosed in the present application can change the mode of polysaccharide and cell interaction, pharmacokinetics and tissue distribution behavior, improve the cell uptake of polysaccharide, improve the oral bioavailability, delay the clearance of polysaccharide from blood circulation, prolong the residence time in vivo, thereby significantly improving the in vitro and in vivo activity and efficacy of active polysaccharide.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical technology, specifically to a polysaccharide composition, nanopolysaccharides, their preparation methods and applications. Background Technology

[0002] Polysaccharides are important biopolymers composed of more than 10 monosaccharide molecules that have undergone dehydration condensation and are linked by glycosidic bonds. Their structure can be represented by the general formula (C6H12H2O). 10 O5)n indicates that the glycosidic bonds are usually (1→3), (1→4), and (1→6)-, as well as α and β configurations. Polysaccharides typically contain more than 100 monosaccharides, with most containing tens of thousands or even millions, making them complex and large biomolecules.

[0003] Polysaccharides are essential components of living organisms, abundant in nature and widely found in almost all plants (ginseng, astragalus, wolfberry), animals (heparin, chondroitin sulfate, chitosan), fungi (shiitake mushrooms, Ganoderma lucidum, Polyporus umbellatus), microorganisms, and other organisms. Some polysaccharides are major components of plant and animal cell walls, such as peptidoglycan and cellulose, while others are important storage nutrients in plants and animals, such as starch and glycogen. In recent years, 21 polysaccharide drugs in six major categories have been marketed in China. The diversity of polysaccharides stems from their structural diversity, including the number, type, linkage sequence, position, and number of sugar chains among the monosaccharides that make them up. This structural diversity, in turn, determines the diversity of their pharmacological activities. Pharmacological and clinical studies have discovered that hundreds of polysaccharides possess various activities, including immunomodulatory, antitumor, lipid-lowering, hypoglycemic, anti-inflammatory, antibacterial, antioxidant, antiviral, and anti-radiation activities. At the same time, plant polysaccharides can also improve food flavor, assist enzyme catalysis processes, and reduce the toxicity, teratogenicity, and potential carcinogenicity of synthetic chemicals.

[0004] Nanotechnology is an emerging field, and many nanomaterials have been used in biomedicine to prevent and treat various diseases. Nanoparticles are particles with a size in the nanometer range. Drugs can usually be encapsulated in the internal matrix of nanoparticles or adsorbed onto the surface of nanoparticles; some drugs can also form nanoparticles themselves. Nanomedicines may exceed 100 nm, but are usually less than 500 nm. Due to the unique physicochemical and biological properties of nanoparticles, the encapsulated drugs can more easily cross blood vessels and enter the body's circulation, and can even cross biological barriers such as the blood-brain barrier, exhibiting strong biopermeability and facilitating absorption. In addition, by changing the particle size of nanoparticles or modifying their surface, active or passive targeted drug delivery can be achieved, with sustained-release effects and improved bioavailability. Nanoparticle formulations can achieve drug delivery via multiple routes, including oral, injection, transdermal, pulmonary, and intraocular administration. After appropriate nano-sizing, the activity and bioavailability of drugs are usually improved to varying degrees, while their toxic side effects are reduced. Therefore, nanotechnology is increasingly being used in the field of polysaccharides.

[0005] Current research on polysaccharide nanoparticles mainly focuses on the following aspects: 1) synthesizing polysaccharide selenium nanoparticles by combining with selenium through reactions with selenite solution; 2) forming nano-silver through the reducing silver nitrate reaction of polysaccharides, or forming nano-gold through hybridization with metal elements such as gold; 3) chelating with metal ions and polyphenols; 4) forming nanoscale complexes with proteins or peptides, and then loading drugs; 5) modifying hydrophilic polysaccharides with fatty acids to make them amphiphilic polysaccharides, which, like other amphiphilic polymers, are used to load poorly soluble drugs to form micelles or nanoparticles; 6) combining positively charged or negatively charged polysaccharides with small molecules of opposite charge (called ion crosslinking agents) through electrostatic adsorption and ion bridging to form polysaccharide nanodelivery carriers, which are then coupled with the drug to be loaded to form polysaccharide nanodelivery systems, such as chitosan and sodium tripolyphosphate, sodium alginate and calcium ions. All of these methods use polysaccharides as carrier materials; the polysaccharides merely play the role of excipients and do not contribute to the administration or enhancement of the active polysaccharide.

[0006] Compared to polysaccharides used as excipients (such as chitosan and sodium alginate), many polysaccharides possess definite pharmacological activities and therapeutic effects on diseases, making them functionally active polysaccharides. However, due to their complex structures, large molecular weights, and high water solubility, polysaccharides exhibit highly flexible chain segments and conformations in aqueous solutions, making it difficult for them to cross the intestinal epithelial barrier and be absorbed into the bloodstream after oral administration. Once in the bloodstream, they are easily phagocytosed by the reticuloendothelial system (liver, spleen, etc.), leading to rapid elimination and a short residence time in the body. Furthermore, their strong hydrophilicity hinders cellular uptake. The understanding of the structure-activity relationship of active polysaccharides is currently almost nonexistent. When polysaccharides are used as carriers, various structural modifications and alterations can be made to better load drugs, such as grafting or modifying hydrophobic groups or molecules (e.g., stearoyl) onto surface functional groups to impart amphiphilicity and improve assembly capabilities. However, when polysaccharides are used as active ingredients for health and disease intervention or treatment, any chemical modification may affect or even alter their activity, thus introducing unpredictable risks to their application. Because active polysaccharides exert their physiological effects by interacting with the body, cells, and targets through their surface functional groups. However, chemical modification of polysaccharide macromolecules is poorly controllable, resulting in significant batch-to-batch variations, and subsequent separation and purification are costly.

[0007] Currently, research on drug delivery systems that encapsulate or design polysaccharides as active drugs is scarce. Existing research primarily focuses on encapsulating polysaccharides as water-soluble drugs using existing formulation techniques. Examples include encapsulating Ganoderma lucidum polysaccharides within chitosan carrier materials using ionogel methods; encapsulating solutions of Dioscorea opposita polysaccharides and Angelica sinensis polysaccharides as the inner aqueous phase in W / O / W double emulsions or within the core of PLGA nanocapsules using polylactic acid-glycolic acid copolymer (PLGA) as the carrier material; and encapsulating polysaccharide solutions within the inner aqueous phase of liposomes. However, these methods suffer from complex preparation processes, low encapsulation efficiency, and low drug loading, making practical applications difficult. While some studies have used polysaccharide solutions as the inner aqueous phase to prepare water-in-oil emulsions, these emulsions face significant limitations in practical and clinical applications.

[0008] In summary, nano-sizing holds promise for enhancing the activity of polysaccharides. However, to date, no nano-sizing technology for active polysaccharides with practical application value (e.g., high drug loading capacity and simple preparation process) has been found, either domestically or internationally, nor have any corresponding products been developed. Therefore, providing an active polysaccharide composition, nano-polysaccharides, and their preparation method is a technical challenge that urgently needs to be addressed by those skilled in the art. Summary of the Invention

[0009] In view of this, the present invention discloses a polysaccharide composition, nano-active polysaccharide, preparation method thereof, and application thereof.

[0010] Although polysaccharides are generally hydrophilic macromolecules, they likely contain some relatively hydrophobic regions. Monosaccharides are linked through different glycosidic bond types (α- and β-) and bonding mechanisms, forming a wide variety of complex natural polysaccharides. Polysaccharide chains are rich in hydroxyl groups, and they readily self-assemble into unique structures through amino bonds, such as the triple helix structures of polysaccharides from wolfberry and carob. Amylose exists in three crystalline forms: types A and B exhibit a regular left-handed double helix structure, which transforms into a single helix structure in dimethyl sulfoxide (DMSO); type V amylose exists as a left-handed single helix structure. The polymorphs of starch share the same helical structure, with six glucose units forming a helix 1.3 nm in diameter and 0.8 nm in tilt distance. Based on its size, it is speculated that this helix can accommodate a single fatty acid chain segment.

[0011] Based on the embodiments of the present invention, it is speculated that most active polysaccharides possess similar helical structures to varying degrees within their molecules, thereby forming hydrophobic cavities or hydrophobic regions. When a lipid adjuvant encounters these hydrophobic regions, the hydrophobic effect drives the interaction between the lipid adjuvant and these hydrophobic regions, or / and the host-guest interaction between the helical cavity and the lipid adjuvant fatty chain segment, inducing an adaptive adjustment in the conformation of the polysaccharide molecule, spontaneously undergoing a disordered-to-order transition, and through the bridging effect of the hydrophobic regions, multiple polysaccharide molecules form relatively stable nanopolysaccharides.

[0012] To achieve the above objectives, the present invention adopts the following technical solution:

[0013] A polysaccharide composition comprising a polysaccharide and a lipid material; wherein,

[0014] The polysaccharides include animal polysaccharides, plant polysaccharides, marine polysaccharides, fungal polysaccharides, and bacterial polysaccharides; the polysaccharides can be pure polysaccharides with a narrow molecular weight distribution, total polysaccharides containing different molecular weights, or complex polysaccharides formed by mixing different polysaccharides (including those from different sources).

[0015] The lipid material is selected from medium-chain and long-chain fatty alcohols (C6-C50) and their esters and ethers, medium-chain and long-chain fatty amines (C6-C50) and their amides formed with acids, medium-chain and long-chain fatty acids (C6-C50) and their esters and derivatives, amides of medium-chain and long-chain fatty acids (C6-C50) and their derivatives, amphiphilic materials containing medium-chain and long-chain fatty chains (C6-C50), long-chain lipophilic materials with a molecular weight <10000 and amphiphilic polymers with lipophilic segments thereof, terpenoids with side-chain fatty chains and their glycosides, other small molecules with fatty chains and amphiphilic molecules with lipophilic moieties thereof, and the fatty chains of the lipid material are saturated fatty chains or contain one, two or more unsaturated bonds;

[0016] The weight ratio of the polysaccharide to the lipid material is 2-50:1.

[0017] Furthermore, the weight ratio of the polysaccharide to the lipid material is 3-30:1.

[0018] Furthermore, the lipid material is selected from medium-chain and long-chain fatty alcohols (C8-C30) and their esters and ethers, medium-chain and long-chain fatty amines (C8-C30) and their amides formed with acids, medium-chain and long-chain fatty acids (C6-30) and their esters and derivatives, amides of medium-chain and long-chain fatty acids (C8-C30) and their derivatives, amphiphilic materials containing medium-chain and long-chain fatty chains (C8-C30), long-chain lipophilic materials with a molecular weight <5000, and amphiphilic polymers with lipophilic segments thereof.

[0019] Furthermore, the lipid material is selected from medium-chain and long-chain fatty alcohols (C8-C18) and their esters and ethers, medium-chain and long-chain fatty amines (C8-C18) and their amides formed with acids, medium-chain and long-chain fatty acids (C8-C18) and their esters and derivatives, amides of medium-chain and long-chain fatty acids (C8-C18) and their derivatives, amphiphilic materials containing medium-chain and long-chain fatty chains (C8-C18), long-chain lipophilic materials with a molecular weight <3000, and amphiphilic polymers with lipophilic segments thereof.

[0020] It is worth noting that the inventors unexpectedly discovered during their research that, under the induction of certain lipid adjuvants, polysaccharide macromolecules can self-assemble into nanostructures with good stability, allowing for oral and intravenous administration. This led to in-depth and systematic research. The results showed that many lipid materials can induce polysaccharide macromolecules to assemble into nanoaggregates (nanopolysaccharides) in aqueous solution, including medium-chain and long-chain fatty alcohols (C6-C50) and their esters and ethers, medium-chain and long-chain fatty amines (C6-C50) and their amides formed with acids, medium-chain and long-chain fatty acids (C6-C50) and their esters and derivatives, amides of medium-chain and long-chain fatty acids (C6-C50) and their derivatives, amphiphilic materials containing medium-chain and long-chain fatty acid chains (C6-C50), and materials with a molecular weight <1000. Long-chain lipophilic materials (including PLA, PLGA, PCL) and amphiphilic polymers with lipophilic segments thereof, terpenoids (cholesterol, cholesterol, protopanaxadiol, protopanatriol, etc.) and their glycosides (such as ginsenoside Rb1 and ginsenoside Rg1) with fatty chains (not less than 5 carbon atoms), other small molecules with fatty chains (not less than 5 carbon atoms, including carbon atoms transesterified by ester bonds) (such as vitamin E), and amphiphilic molecules with lipophilic moieties thereof (such as TPGS), etc. The fatty chains of these lipid materials can be saturated fatty chains or contain one, two, or more unsaturated bonds.

[0021] The types of polysaccharides suitable for inducing the formation of nanoaggregates (or nanoassemblies) by these lipid adjuvants are very broad, covering various active polysaccharides from plant, animal, and fungal sources, such as: Astragalus polysaccharide, Plantago asiatica polysaccharide, Trametes versicolor polysaccharide, Lycium barbarum polysaccharide, soybean polysaccharide, Lentinus edodes polysaccharide, Angelica sinensis polysaccharide, Polyporus umbellatus polysaccharide, Tremella fuciformis polysaccharide, Aloe vera polysaccharide, Dioscorea opposita polysaccharide, Fritillaria cirrhosa polysaccharide, Cordyceps sinensis polysaccharide, Eucommia ulmoides polysaccharide, Achyranthes bidentata polysaccharide, Polygonatum sibiricum polysaccharide, Bletilla striata polysaccharide, Ginseng polysaccharide, Gardenia jasminoides polysaccharide, Dendrobium nobile polysaccharide, Lilium brownii polysaccharide, etc. Different proportions of lipid adjuvants can induce the formation of these polysaccharide nanoassemblies. Whether it is a pure polysaccharide with a very narrow molecular weight distribution, a total polysaccharide containing different molecular weights, or a complex polysaccharide formed by mixing different polysaccharides (including those from different sources), lipid adjuvants can induce the formation of these polysaccharide nanoassemblies. Of course, different polysaccharides may have their own most suitable lipid inducers and the most suitable polysaccharide-excipient ratio. Combinations of different lipid adjuvants may also yield better results than preparations using a single lipid adjuvant.

[0022] Furthermore, the polysaccharide composition also includes pharmaceutically acceptable excipients, including commonly used injectable adjuvants (pH adjusters, surfactants, suspending agents, drug delay adjuvants, isotonic modifiers, local analgesics, antibacterial agents), binders, disintegrants, lubricants, tablet fillers / diluents, and tablet lubricants.

[0023] A second object of the present invention is to provide a nanopolysaccharide. The nanopolysaccharide is prepared from the polysaccharide composition described above.

[0024] To achieve the above objectives, the present invention adopts the following technical solution:

[0025] A nanopolysaccharide is obtained by dissolving a lipid material in a water-miscible organic solvent, adding it to a polysaccharide solution under stirring or ultrasound to induce the self-assembly of the polysaccharide, and then removing the organic solvent, so that the lipid material and polysaccharide assemble into nanoparticles with a particle size of 20-1000 nm.

[0026] It should be noted that the nanopolysaccharides can alter the way polysaccharides interact with cells, their pharmacokinetics and tissue distribution behavior, increase cellular uptake of polysaccharides, delay the clearance of polysaccharides from the bloodstream, and prolong their retention time in the body, thereby significantly improving the in vitro and in vivo activity and efficacy of the active polysaccharides.

[0027] Furthermore, the preparation method of the nanopolysaccharide specifically includes the following steps:

[0028] 1) Dissolve the lipid material in ethanol, methanol, acetone, isopropanol or a mixture thereof to obtain a lipid material solution, and dissolve the polysaccharide in water to obtain a polysaccharide solution;

[0029] 2) The lipid material solution is added to the polysaccharide solution under stirring or sonication, and the solvent is removed by evaporation to obtain nano-polysaccharides with an average particle size of 20-1000 nm.

[0030] 3) If the obtained nanopolysaccharide has a large particle size or a wide particle size distribution, it can be further homogenized under high pressure to reduce the particle size and / or particle size distribution;

[0031] 4) The obtained nano-polysaccharides can be further spray-dried or freeze-dried to become solid powders to suit different applications.

[0032] Furthermore, the evaporation in step 2) is one or a combination of natural evaporation, heated evaporation, and reduced pressure evaporation, and the homogenization pressure in step 3) is 500-4000 bar.

[0033] For amphiphilic lipid materials, this invention also provides a second technical solution for preparing nanopolysaccharides. This involves directly adding the amphiphilic lipid material to an aqueous solution of the polysaccharide, stirring to hydrate it, and then using ultrasound (with heating if necessary) to accelerate its diffusion into or between polysaccharide molecules. This promotes its interaction with the polysaccharide molecules, inducing the polysaccharide molecules to assemble into nano-aggregates. The advantage of this technical solution is that it does not use organic solvents. The disadvantages are that the preparation process is slightly slower, and the resulting nanopolysaccharides have a slightly larger particle size and a slightly wider particle size distribution than those obtained using the first technical solution. However, high-pressure homogenization can effectively reduce the particle size and particle size distribution of the nanopolysaccharides.

[0034] This invention also seeks protection for the use of the described nanopolysaccharide in the preparation of food, health products, and pharmaceutical products.

[0035] Furthermore, the nanopolysaccharides can be further loaded with lipid-soluble small molecules or protein, polypeptide, and nucleic acid macromolecules to become nanoparticles with health care and therapeutic effects, enabling in vivo delivery of poorly soluble drugs and macromolecular drugs and synergistic effects based on nano-effects, or synergistic effects with polysaccharides; and the application routes of the nanopolysaccharides include oral administration, injection, mucosa, cavity, wound, or topical application.

[0036] Furthermore, the nanopolysaccharides are used in drugs with anti-inflammatory, antioxidant, (adjunctive) antitumor, (adjunctive) anticardiovascular and cerebrovascular disease, neuroprotective, (adjunctive) antifibrotic, lipid-lowering, antihypertensive, hypoglycemic, anti-ADHD, and anti-scarring properties.

[0037] Compared with existing technologies, this invention discloses a polysaccharide composition, nanopolysaccharides, their preparation methods, and applications. It prepares active polysaccharide nanoparticles through hydrophobic interactions induced by lipid chaperone molecules, offering the following significant advantages:

[0038] 1) Low-cost and safe excipients: The selected liposome adjuvants are mostly physiologically inert and commonly used pharmaceutical excipients, offering low cost and good safety. 2) Extremely simple preparation process, facilitating industrial production: The alcoholic solution of the lipid adjuvant is added to the polysaccharide solution under ultrasonic or stirring conditions, and the organic solvent is removed (high-pressure homogenization can be used to reduce particle size if necessary). 3) Extremely high polysaccharide drug loading capacity: Only a small amount of lipid material is used in the preparation process; the main component of the nanopolysaccharide is polysaccharide. 4) Does not alter the polysaccharide structure, perfectly preserving the pharmacological activity of the polysaccharide: This invention induces the nano-assembly of polysaccharides through the hydrophobic interaction of the added lipid adjuvant. The formation of nanopolysaccharides relies solely on physical interactions, without altering the chemical structure of the polysaccharide, thus preserving its activity. 5) Significantly enhances polysaccharide activity: Nanoparticles significantly increase the uptake of polysaccharides by tissue cells (by more than 5 times), thereby significantly improving the in vitro activity of polysaccharides. 6) Significantly improves pharmacokinetic behavior: Nanoparticles of carob polysaccharides have a higher AUC in blood than free polysaccharides, significantly prolonging their in vivo retention. 7) Significantly improves in vivo efficacy: Nanoparticles can increase uptake by tissue cells while prolonging their in vivo retention. Whether administered orally or intravenously, nanoparticles significantly improve efficacy compared to free polysaccharides. 8) Expected to significantly improve polysaccharide absorption and oral bioavailability: Oral absorption of many small molecule drugs is improved after being prepared as nanoparticles. The in vivo efficacy of oral administration of nanoparticles is significantly better than that of polysaccharide solutions, suggesting that nanoparticles are expected to significantly improve the oral absorption and bioavailability of polysaccharides. Attached Figure Description

[0039] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the description of the embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are only embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on the provided drawings without creative effort.

[0040] Figure 1 These are photographs of the particle size and particle size distribution of nanocarob polysaccharide CSPS@OA NPS (polysaccharide-oleic acid = 15:1).

[0041] Figure 2 This is a transmission electron microscope image of carob nanopolysaccharide.

[0042] Figure 3 This is a curve showing the changes in particle size and polydispersity index of carob nanopolysaccharides over time during room temperature storage.

[0043] Figure 4 The curves show the changes in particle size and polydispersity index of nanocarob polysaccharides over time during incubation at 37°C in different physiological media.

[0044] Figure 5 The study investigated the effects of different concentrations of free polysaccharides (A) and nanopolysaccharides (B) on the survival rate of RAW 264.7 macrophages, and the effects of free polysaccharides and nanopolysaccharides on the release of NO (C), IL-6 (D), IL-10 (E), and TNF-α (F) after 48 hours of treatment with RAW 264.7 macrophages (n=3). (**** P < 0.0001 vs. control group;) # P < 0.05; ## P < 0.01; ### P < 0.001; #### P < 0.01 vs. LPS group; && P < 0.01).

[0045] Figure 6 The results showed the cough latency (A), number of coughs within 2 minutes (B), secretion levels of phenol red (C), IL-10 (D), TNF-α (E), and IL-6 (F) (n=10) in mice with a lung inflammation-cough model induced by lipopolysaccharide (LPS) and ammonia, after oral and intravenous injection of free polysaccharides and nanopolysaccharides, and the lung histopathological sections (G). (**** P< 0.0001 vs. control group; # P < 0.05; ## P < 0.01; ### P < 0.001; #### P < 0.01 vs. LPS group; && P < 0.01).

[0046] Figure 7 The images show fluorescence images (A) and semi-quantitative analysis (C) of Caco-2 cells taking up FITC-labeled free polysaccharides (B). Blue represents 4,6-diamidino-2-phenylindole (DAPI), and green represents FITC-labeled free polysaccharides or nanopolysaccharides.

[0047] Figure 8 The curves show the changes in blood drug concentration over time after intravenous injection of IR783-labeled free polysaccharide and nanopolysaccharide in mice (n=3).

[0048] Figure 9 This is a comparison of Congo Red experimental results for carob polysaccharide CSPS and nanocarob polysaccharide CSPS@OA NPS.

[0049] Figure 10Comparison of FT-IR spectra of carob polysaccharide CSPS and nano carob polysaccharide CSPS@OA NPS. Detailed Implementation

[0050] The technical solutions in the embodiments of the present invention will be clearly and completely described below. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative effort are within the scope of protection of the present invention.

[0051] The term "embodiment" used herein, as an example, is not necessarily to be construed as superior to or better than other embodiments. Performance testing in the embodiments of this application, unless otherwise specified, employs conventional testing methods in the art. It should be understood that the terminology used in this application is merely for describing particular implementations and is not intended to limit the scope of this disclosure.

[0052] Unless otherwise stated, the technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application pertains; other experimental methods and technical means not specifically mentioned herein refer to experimental methods and technical means commonly used by one of ordinary skill in the art.

[0053] To better illustrate the content of this application, numerous specific details are provided in the following detailed embodiments. Those skilled in the art should understand that this application can be implemented even without certain specific details. In the embodiments, some methods, means, instruments, and devices well-known to those skilled in the art are not described in detail in order to highlight the main points of this application.

[0054] Without conflict, the technical features disclosed in the embodiments of this application can be combined arbitrarily, and the resulting technical solution belongs to the content disclosed in the embodiments of this application.

[0055] To better understand the present invention, the following embodiments are provided for further detailed description of the present invention, but they should not be construed as limiting the present invention. Any non-essential improvements and adjustments made by those skilled in the art based on the above-described invention are also considered to fall within the protection scope of the present invention.

[0056] Example 1. Nano-Astragalus polysaccharide with glycosides of terpenoid compounds having side-chain fatty chains as excipients

[0057] Astragalus polysaccharide (APS) was accurately weighed and dissolved in water to prepare a 5 mg / mL polysaccharide solution. 10 mg of ginsenoside Rg1, ginsenoside Rb1, and total ginsenosides were weighed and dispersed in 10 mL of the polysaccharide solution. The mixture was sonicated at 37℃, and the particle size, PDI, and potential were measured at 5, 10, 20, 30, and 60 min (see Tables 1, 2, and 3). Nano-Astragalus polysaccharide was successfully prepared.

[0058] Table 1. Particle size, PDI, and potential of Rg1-APS (weight ratio 1:5) nano-Astragalus polysaccharides obtained by ultrasound at different times.

[0059] Time (min) Particle size PDI Potential 5 125.1±78.67 0.333±0.099 -11.2±9.38 10 290±16.05 0.345±0.018 -20.7±1.8 20 153.3±8.122 0.262±0.026 -21.6±5.41 30 140.5±35.59 0.23±0.055 -20.6±1.05 60 189.9±16.6 0.243±0.015 -22.6±1.05

[0060] Table 2. Particle size, PDI, and potential of Rb1-APS (weight ratio 1:5) nano-Astragalus polysaccharides obtained by ultrasound at different times.

[0061] Time (min) Particle size PDI Potential 5 189.4±98.59 0.212±0.137 -19.6±2.23 10 165.8±66.97 0.225±0.054 -21.5±2.25 20 110.4±58.9 0.244±0.103 -22.6±1.01 30 208.2±87.18 0.288±0.091 -16.9±0.551 60 172.6±32.21 0.295±0.091 -21.1±2.19

[0062] Table 3. Particle size, PDI, and potential of ginsenoside-APS (weight ratio 1:5) nano-Astragalus polysaccharides obtained by ultrasound at different times.

[0063] Time (min) Particle size PDI Potential 5 189.7±40.54 0.28±0.026 -33.6±2.31 30 230.7±78.67 0.284±0.077 -29.1±1.74 Homogenize 20 times at 1400 bar 162±2.178 0.249±0.025 -33.3±4.77

[0064] Example 2. Nano-Astragalus polysaccharide with fatty acids, fatty alcohols, fatty amines and their amides as excipients (polysaccharide-excipient ratio = 5:1, by weight)

[0065] A 5 mg / mL Astragalus polysaccharide aqueous solution was prepared. Separately weighed amounts of neodecanoic acid, oleic acid, linoleic acid, ricinoleic acid, linolenic acid, stearic acid, arachidic acid, octanol, lauryl alcohol, stearyl alcohol, dodecylamine, stearylamine, and stearoylethanolamine were prepared into 4 mg / mL methanol solutions. 0.5 mL of the methanol solution (2 mg excipients) was then added dropwise to 2 mL of the polysaccharide solution (10 mg polysaccharide) under ultrasonic conditions. Ultrasonic treatment continued for 5 min, and the organic phase was removed by rotary evaporation under reduced pressure (homogenization was performed 10 times at 1400 bar if necessary). Particle size, PDI, and potential were measured. The results are shown in Table 4. Nanoparticles were successfully obtained.

[0066] Table 4. Particle size, PDI, and potential of nano-Astragalus polysaccharides with fatty acids, fatty alcohols, fatty amines, and their amides as excipients.

[0067] Particle size (nm) PDI Potential Astragalus polysaccharide-neodecanic acid 279.2±13.701 0.321±0.053 -23.1±0.4 Astragalus polysaccharide-caprylic acid 223.1±156.23 0.312±0.123 -14.2±1.15 Astragalus polysaccharide-decanedic acid 225.9±2.065 0.304±0.038 -5.88±0.248 Astragalus polysaccharide-oleic acid 192.6±7.257 0.21±0.017 -35.6±4.36 Astragalus polysaccharide-linoleic acid 197.0±7.142 0.168±0.043 -51.7±4.85 Astragalus polysaccharide-ricinoleic acid 184.4±4.916 0.209±0.046 -40.2±1.05 Astragalus polysaccharide-linolenic acid 360.8±32.76 0.397±0.074 -59.8±2.44 Homogenize 10 times at 1400 bar 249.4±3.156 0.081±0.096 -28.8±1.15 Astragalus polysaccharide-arachidic acid 539.0±42.96 0.470±0.07 -57.3±0.404 Homogenize 10 times at 1400 bar 374.3±9.853 0.201±0.05 -32.9±8.1 Astragalus polysaccharide-stearic acid 487.9±23.05 0.17±0.126 -22.9±0.737 Homogenize 10 times at 1400 bar 403.2±18.14 0.113±0.033 -40.1±2.2 Astragalus polysaccharide-octanol 297.7±2.757 0.253±0.064 -26.5±3.23 Astragalus polysaccharide-laurol 273.2±4.366 0.264±0.076 -27.9±3.01 Astragalus polysaccharide-stearyl 323.6±6.233 0.287±0.084 -24.6±2.56 Astragalus polysaccharide-dodecylamine 287.3±5.336 0.236±0.055 -8.2±0.34 Astragalus polysaccharide-stearylamine 362.5±7.442 0.341±0.099 -7.6±0.41 Astragalus polysaccharide-stearoylethanolamine 297.5±8.215 0.289±0.087 -26.2±2.88

[0068] Example 3. Nanoparticles of polysaccharides with fatty acid esters as excipients

[0069] A 5 mg / mL Astragalus polysaccharide aqueous solution was prepared. Peceol™ monooleate, Maisine® CC monolinoleate, Lauroglycol™ 90 propylene glycol monolaurate, and Labrafac MC60 monocaprylate (mainly composed of monoesters and a small amount of diesters) were weighed and prepared into 4 mg / mL ethanol solutions. 0.5 mL of each ethanol solution (2 mg excipient) was added dropwise to 2 mL of the polysaccharide solution (10 mg polysaccharide) under ultrasonic conditions. The mixture was sonicated for 5 min, and the organic phase was removed by rotary evaporation at 45 °C. The particle size, PDI, and potential were measured and are shown in Table 5. Nano-polysaccharides were successfully obtained.

[0070] Aqueous solutions of different polysaccharides (Polygonatum polysaccharide, Ginseng polysaccharide, Gardenia polysaccharide, Fritillaria polysaccharide, Aloe polysaccharide, Dioscorea polysaccharide, Astragalus polysaccharide, Polyporus umbellatus polysaccharide, Carob polysaccharide, etc.) with a concentration of 5 mg / mL were prepared. 4 mg / mL ethanol solutions were prepared by weighing glyceryl mono- and dicaprylic / capric esters (Labrafac MC60, mainly composed of monoesters and a small amount of diesters), lauryl laurate, pentyl octanoate, and lauryl palmitate. 0.25 mL–0.5 mL of each ethanol solution was added dropwise to 2 mL of the polysaccharide solution under ultrasonic conditions. Ultrasonic treatment was continued for 5 min, and the organic phase was removed by rotary evaporation at 45 °C. Particle size, PDI, and potential were measured (see Table 5). Nano-polysaccharides were successfully obtained in all cases.

[0071] Table 5. Particle size, PDI, and potential of nanopolysaccharides with fatty acid esters, edible oils, and fat-soluble small molecules as excipients.

[0072] Category mass ratio Particle size (nm) PDI Potential (mV) Astragalus Polysaccharide - Maisine® 5:1 222.3±6.21 0.145±0.02 -60.3±1.86 Astragalus polysaccharide-Peceol™ 5:1 217.4±7.15 0.151±0.051 -31.4±1.71 Astragalus polysaccharide-Lauroglycol™ 5:1 240.1±13.34 0.155±0.025 -38.0±1.95 Plantain polysaccharide - Maisine® 5:1 214.2±4.90 0.177±0.034 -19.8±1.72 Peceol™ (Plantago asiatica polysaccharide) 5:1 212.1±7.27 0.144±0.073 -20.3±1.59 Lauroglycol™ 5:1 195.0±7.55 0.221±0.036 -24.4±0.473 Yunzhi Polysaccharide - Maisine® 5:1 206.4±0.85 0.164±0.031 -23.6±0.40 Peceol™ (Yunzhi Polysaccharide) 5:1 180.8±1.945 0.128±0.076 -21.6±1.08 Lauroglycol™ 5:1 200.9±3.55 0.124±0.022 -28.9±1.53 Goji Berry Polysaccharide - Maisine® 5:1 630.5±21.33 0.193±0.122 -34.4±1.71 Peceol™ (goji berry polysaccharide) 5:1 677.0±49.34 0.353±0.201 -37.0±0.346 Lauroglycol™ (goji berry polysaccharide) 5:1 280.8±6.62 0.111±0.031 -24.7±1.95 Soybean Polysaccharide - Maisine® 5:1 199.0±2.88 0.130±0.011 -32.2±3.05 Soybean Polysaccharide - Peceol™ 5:1 180.3±3.95 0.123±0.022 -45.6±2.05 Soybean polysaccharide-Lauroglycol™ 5:1 206.2±4.96 0.230±0.052 -41.7±2.37 Lentinan-Maisine® 5:1 283.0±9.51 0.112±0.036 -31.0±0.666 Peceol™ (Lentinol Polysaccharide) 5:1 322.8±8.56 0.057±0.072 -27.1±1.35 Lentinan-Lauroglycol™ 5:1 309.5±30.8 0.301±0.066 -33.4±0.924 Polygonatum Polysaccharide - Labrafac MC60 8:1 225.8±13.76 0.233±0.056 -25.4±1.69 Ginseng Polysaccharide - Labrafac MC60 8:1 186.1±9.76 0.201±0.045 -20.1±1.87 Gardenia polysaccharide - Labrafac MC60 8:1 165.8±8.72 0.186±0.051 -29.5±3.01 Fritillaria polysaccharide - Labrafac MC60 8:1 158.9±6.32 0.145±0.042 -24.8±2.37 Aloe polysaccharide - Labrafac MC60 8:1 215.2±8.24 0.198±0.065 -31.1±2.15 Yam polysaccharide - Labrafac MC60 8:1 167.2±6.01 0.176±0.052 -28.0±1.77 Astragalus polysaccharide-laurate laurate 10:1 256.0±5.32 0.211±0.078 -30.1±3.44 Polysaccharide-Lauryl Laurate 10:1 267.3±5.95 0.231±0.085 -32.5±4.23 Carob polysaccharide-laurate ester 10:1 198.4±5.23 0.243±0.061 -26.6±3.68 Astragalus polysaccharide-octanoic acid pentyl ester 10:1 281.0±8.76 0.238±0.071 -24.6±3.31 Polysaccharide from Poria cocos - Amyl octanoate 10:1 301.8±9.26 0.221±0.101 -23.2±2.43 Carob polysaccharide-caprylate 10:1 188.4±3.66 0.176±0.065 -30.2±3.77 Astragalus polysaccharide-lauryl oleate 10:1 231.4±6.01 0.188±0.074 -36.1±5.32 Polysaccharide-Lauryl Oleate 10:1 264.0±9.16 0.268±0.097 -27.2±4.15 Carob polysaccharide-lauryl oleate 10:1 165.6±3.55 0.163±0.036 -33.6±5.01

[0073] Example 4. Nanoparticles of polysaccharide with edible oil and fat-soluble small molecules as excipients (polysaccharide-excipient ratio = 8:1, by weight)

[0074] Edible oils containing medium-chain and long-chain fatty acid glycerides, such as refined soybean oil, olive oil, and corn oil, were accurately weighed and dissolved in ethanol to prepare a 10 mg / mL solution. Astragalus polysaccharide, Cordyceps polysaccharide, and carob polysaccharide were also accurately weighed and dissolved in water to prepare a 10 mg / mL solution. 0.5 mL of each ethanol solution (5 mg excipient) was slowly dripped into 4 mL of the polysaccharide solution (40 mg polysaccharide) under ultrasonic treatment in a 250 W water bath. The organic solvent was removed by rotary evaporation at 45 °C. Particle size, PDI, and Zeta potential were measured. The results are shown in Table 6 below, indicating that nano-Cordyceps polysaccharides were successfully prepared.

[0075] Accurately weigh 5 mg of protopanaxadiol (PPD, a lipid-soluble aglycone of ginsenosides), protopanaxadiol, vitamin E, cholesterol, and cholesterol alcohol, and dissolve them in anhydrous ethanol to prepare a 10 mg / mL solution. In 0.5 mL of this solution, accurately weigh 50 mg of cordyceps polysaccharide, carob polysaccharide, shiitake mushroom polysaccharide, and eucommia polysaccharide, and dissolve them in 5 mL of water as the aqueous phase. Slowly add 0.5 mL of the ethanol solution to 4 mL of the polysaccharide solution using ultrasonication. Remove the organic solvent by rotary evaporation at 45 °C. Detect the particle size, PDI, and Zeta potential. The results are shown in Table 6 below, indicating that nano-polysaccharides were successfully prepared.

[0076] Table 6. Particle size, PDI, and potential of nanopolysaccharides with edible oils and fat-soluble small molecules as excipients.

[0077] Particle size (nm) PDI Potential (mV) Astragalus polysaccharide - refined soybean oil 288.3±11.76 0.211±0.067 -26.3±2.01 Astragalus polysaccharide-olive oil 253.7±8.33 0.167±0.053 -24.7±3.82 Astragalus polysaccharide-corn oil 310.4±13.57 0.268±0.088 -27.6±3.53 Astragalus polysaccharide - Vitamin E 155.3±5.31 0.187±0.065 -32.4±2.36 Cordyceps polysaccharide-vitamin E 178.5±6.22 0.164±0.071 -29.6±3.21 Eucommia polysaccharides - Vitamin E 232.8±8.73 0.221±0.032 -34.1±4.43 Carob polysaccharide-VE 146.3±2.37 0.134±0.03 -35.6±2.01 Carob polysaccharide-VE, after one week of storage 157.9±2.10 0.201±0.02 -31.5±1.59 Carob polysaccharide-cholesterol 214.0±6.38 0.185±0.093 -32.6±3.20 Carob polysaccharide-cholesterol 252.0±10.73 0.226±0.097 -29.5±4.45 Eucommia polysaccharide-cholesterol 473.0±18.54 0.341±0.215 -24.7±3.98 Lentinan-PPD 265.2±0.764 0.067±0.023 -22.3±0.839 Carob polysaccharide-VE, after one week of storage 157.9±2.10 0.201±0.02 -31.5±1.59

[0078] Example 5. Nano-Astragalus polysaccharides and nano-Lycium barbarum polysaccharides using amphiphilic molecules with fatty chain segments as excipients.

[0079] Prepare an aqueous solution of Astragalus polysaccharide with a concentration of 5 mg / mL, and weigh it as follows: polyoxyethylene castor oil EL, TPGS, and linoleyl polyoxyethylene-6 glycerol ester Labrafil. ® M2125 CS, octanoic acid, caprylic / capric acid, polyethylene glycol glycerol ester (Labrasol®), Maize 59, and Plannico P188 were prepared into an ethanol solution. Separately, egg yolk lecithin, soybean lecithin, dioleoylphosphatidylcholine, and DSPE-PEG2000 were dissolved in acetone. 0.5 mL of either the ethanol or acetone solution was added dropwise to 2 mL of the polysaccharide solution under stirring. The mixture was then sonicated for 5 min, and the organic phase was removed by rotary evaporation at 45°C. The particle size, PDI, and potential were measured (see Table 7). Nano-polysaccharides were successfully obtained in both cases.

[0080] Prepare a 5 mg / mL solution of Lycium barbarum polysaccharide by taking PLA. 1000 -PEG 1000 PCL 1000 -PEG 1000 PLGA 1000 -PEG 1000 Several ethanol solutions were prepared to a concentration of 4 mg / mL. 0.25 mL of the ethanol solution (1 mg excipient) was then added dropwise to 2 mL of polysaccharide solution (10 mg polysaccharide) under ultrasonic conditions. The mixture was then sonicated for 5 min, and the organic phase was removed by rotary evaporation at 45 °C. The particle size, PDI, and potential were measured and are shown in Table 7. Nano-polysaccharides were successfully obtained in all cases.

[0081] Table 7. Particle size, PDI, and potential of amphiphilic molecules with fatty acid segments induced in Astragalus polysaccharide and Lycium barbarum polysaccharide nanoassemblies

[0082] weight ratio Particle size (nm) PDI Potential (mV) Astragalus polysaccharide-polyoxyethylene castor oil EL 5:1 221.2±35.56 0.334±0.034 -21.5±1.31 Astragalus polysaccharide-TPGS 5:1 299.2±16.9 0.319±0.039 -26.5±2.22 Astragalus polysaccharide-egg yolk phospholipid 10:1 215.8±8.25 0.281±0.042 -27.5±2.17 Astragalus polysaccharide-soybean lecithin 15:1 254.6±8.11 0.232±0.063 -11.2±0.63 Astragalus polysaccharide-DOPC 15:1 203.6±6.32 0.189±0.054 -13.8±0.56 <![CDATA[Astragalus polysaccharide-DSPE-PEG 2000 > 10:1 187.4±7.24 0.196±0.061 -23.5±3.11 Astragalus polysaccharide-Labrafil M2125 CS 5:1 203.9±0.46 0.164±0.025 -23.6±2.26 Astragalus polysaccharide - Labrasol 5:1 182.5±5.34 0.364±0.084 -25.3±0.643 Astragalus polysaccharide - Maize 59 8:1 282.3±26.21 0.264±0.771 -21.5±1.31 Astragalus polysaccharide-P188 6:1 295.9±63.12 0.322±0.133 -27.9±2.68 <![CDATA[Lycium barbarum polysaccharide-PLA 1000 -PEG 1000 > 10:1 123.9±3.021 0.136±0.028 -30.8±3.35 <![CDATA[Lycium barbarum polysaccharide - PCL 1000 -PEG 1000 > 10:1 131.2±3.674 0.112±0.013 -32.1±3.87 <![CDATA[Lycium barbarum polysaccharide-PLGA 1000 -PEG 1000 > 10:1 158.0±4.011 0.098±0.026 -34.6±4.21

[0083] Example 6: Nanopolysaccharide with long-chain fat-soluble material as excipient (polysaccharide-excipient = 5:1, weight ratio)

[0084] Materials: Polylactic acid (PLA) (molecular weight 2000), polycaprolactone (PCL) (molecular weight 1000), polylactide-glycolic acid copolymer (PLGA) (molecular weight 500)

[0085] A polysaccharide aqueous solution with a concentration of 5 mg / mL was prepared. A certain amount of PLA, PCL, and PLGA were prepared into an ethanol solution with a concentration of 4 mg / mL. 0.5 mL of the ethanol solution (2 mg of excipient) was added dropwise to 2 mL of the polysaccharide solution (10 mg of polysaccharide) under ultrasonic conditions. The mixture was sonicated for another 5 min. The organic phase was removed by rotary evaporation at 45 °C. The particle size, PDI, and potential were measured and are shown in Table 8. After standing at room temperature for 2 days, the particle size, PDI, and potential were measured again and are shown in Table 9. Nano-polysaccharides were successfully obtained and showed good stability.

[0086] Table 8. Nanoparticles of polysaccharides using PLA (MW2000), PCL (MW1000), and poly(lactic-co-glycolic acid) copolymer PLGA (MW600) as excipients.

[0087] Particle size PDI Potential Astragalus polysaccharide-PLA 269.3±5.2 0.162±0.031 -14.3±0.781 Astragalus polysaccharide-PCL 157.6±2.307 0.227±0.018 -17.5±1.45 Astragalus polysaccharide-PLGA 149.3±0.116 0.182±0.001 -15.4±1.76 Plantain polysaccharide-PLA 218.2±2.203 0.098±0.031 -15.2±0.361 Plantain polysaccharide-PCL 131.5±1.721 0.206±0.028 -14.5±0.551 Plantain polysaccharide-PLGA 174.1±1.29 0.151±0.034 -11.4±0.513 Yunzhi Polysaccharide-PLA 259.4±2.816 0.086±0.067 -14.9±0.3 Yunzhi Polysaccharide-PCL 149.2±3.107 0.139±0.032 -11.6±1.27 Yunzhi Polysaccharide-PLGA 166±5.297 0.067±0.029 -9.55±0.466 Lycium barbarum polysaccharide-PLA 245.9±11.14 0.199±0.03 -22.6±1.66 Lycium barbarum polysaccharide-PCL 114.2±3.404 0.145±0.016 -24±1.49 Lycium barbarum polysaccharide-PLGA 207.8±4.54 0.076±0.046 -24±1.06 Soybean polysaccharide-PLA 247.6±4.709 0.128±0.092 -16.8±1.91 Soybean polysaccharide-PCL 142.3±1.21 0.208±0.022 -26.3±0.436 Soybean polysaccharide-PLGA 159.6±1.358 0.136±0.007 -16.8±0.611 Lentinan-PLA 335.2±11.42 0.231±0.037 -17.2±0.458 Lentinan-PCL 189.2±2.458 0.342±0.034 -21.8±1.81 Lentinan-PLGA 199.5±8.866 0.231±0.044 -15.1±3.27 Angelica polysaccharide-PLA 331.3±17.57 0.195±0.134 -16.4±1.06 Angelica polysaccharide-PCL 194.1±9.277 0.202±0.053 -16.2±0.265 Angelica polysaccharide-PLGA 354.1±7.582 0.074±0.067 -14.5±1.17

[0088] Table 9. Particle size, particle size distribution, and potential of nanopolysaccharides using PLA (MW2000), PCL (MW1000), and poly(lactic-co-glycolic acid) copolymer PLGA (MW600) as excipients after being left at room temperature for 2 days.

[0089] Category Particle size PDI Potential Astragalus polysaccharide-PLA 301.3±15.23 0.208±0.033 -17.6±0.85 Astragalus polysaccharide-PCL 184.2±6.757 0.144±0.102 -22.9±1.44 Astragalus polysaccharide-PLGA 172.5±11.15 0.22±0.009 -19.3±0.764 Plantain polysaccharide-PLA 264.3±24.46 0.154±0.117 -17.5±0.557 Plantain polysaccharide-PCL 177.3±9.708 0.268±0.05 -9.9±0.529 Plantain polysaccharide-PLGA 208.2±6.453 0.164±0.055 -11.0±0.656 Yunzhi Polysaccharide-PLA 315.7±6.793 0.227±0.051 -14.1±0.265 Yunzhi Polysaccharide-PCL 197.5±14.35 0.203±0.015 -9.81±0.683 Yunzhi Polysaccharide-PLGA 166.1±2.04 0.129±0.02 -9.70±0.976 Lycium barbarum polysaccharide-PLA 255.0±6.20 0.206±0.019 -34.5±2.50 Lycium barbarum polysaccharide-PCL 125.7±5.326 0.193±0.025 -22.6±1.00 Lycium barbarum polysaccharide-PLGA 234.9±7.404 0.103±0.009 -17.4±0.764 Soybean polysaccharide-PLA 273.9±12.97 0.215±0.123 -21.2±1.80 Soybean polysaccharide-PCL 157.6±2.85 0.203±0.028 -24.4±0.20 Soybean Polysaccharide-PLGA 170.3±7.39 0.168±0.068 -21.6±0.503 Lentinan-PLA 395.5±22.0 0.237±0.044 -15.0±0.608 Lentinan-PCL 196.9±6.088 0.202±0.145 -22.2±1.96 Lentinan-PLGA 188.9±8.888 0.272±0.012 -16.5±0.702 Angelica polysaccharide-PLA 385.7±11.98 0.242±0.186 -16.2±0.586 Angelica polysaccharide-PCL 171.3±4.386 0.219±0.079 -24.2±1.71 Angelica polysaccharide-PLGA 327.8±8.041 0.189±0.0126 -18.2±0.862

[0090] Example 7: Long-chain fat-soluble material is excipient nanopolysaccharide (polysaccharide-excipient = 10:1, mass ratio)

[0091] A polysaccharide aqueous solution with a concentration of 5 mg / mL was prepared. A certain amount of PLA, PCL, and PLGA were prepared into an ethanol solution with a concentration of 4 mg / mL. 0.25 mL of the ethanol solution (1 mg of excipient) was added dropwise to 2 mL of the polysaccharide solution (10 mg of polysaccharide) under ultrasonic conditions. Ultrasonic treatment was continued for 5 min. The organic phase was removed by rotary evaporation at 45 °C. The particle size, PDI, and potential were measured and are shown in Table-10. Nano-polysaccharides were successfully obtained.

[0092] Table 10 Particle size and particle size distribution of nanopolysaccharides with PLA, PCL, and poly(lactic-co-glycolic acid) copolymer (PLGA) as excipients at other molecular weights

[0093] Particle size PDI <![CDATA[Astragalus polysaccharide - PLA 5000 > 281.3±7.921 0.216±0.045 <![CDATA[Astragalus polysaccharide - PCL 5000 > 258.3±9.861 0.208±0.021 <![CDATA[Astragalus polysaccharide - PLGA 5000 > 301.2±6.314 0.231±0.037 <![CDATA[Plantago polysaccharide - PLA 3500 > 236.1±4.357 0.167±0.053 <![CDATA[Plantago polysaccharide - PCL 3500 > 223.5±3.231 0.187±0.031 <![CDATA[Plantago polysaccharide - PLGA 3500 > 196.3±2.23 0.181±0.036 <![CDATA[Coriolus versicolor polysaccharide-PLA 1000 > 203.4±2.002 0.126±0.052 <![CDATA[Coriolus versicolor polysaccharide - PCL 2000 > 169.4±3.110 0.148±0.037 <![CDATA[Polysaccharide-K-PLGA 1000 > 149.3±2.206 0.087±0.031 <![CDATA[Soybean polysaccharide - PLA 1000 > 207.6±3.361 0.152±0.042 <![CDATA[Soybean polysaccharide - PCL 1000 > 137.3±2.26 0.183±0.032 <![CDATA[Soybean polysaccharide - PLGA 1000 > 167.6±2.351 0.206±0.058 <![CDATA[Lentinan-PLA 1000 > 215.2±7.43 0.211±0.041 <![CDATA[Lentinan-PCL 2000 > 193.1±3.008 0.243±0.051 <![CDATA[Lentinan-PLGA 1000 > 215.7±6.637 0.221±0.039 <![CDATA[Angelica polysaccharide - PLA 1000 > 233.1±8.306 0.251±0.084 <![CDATA[Angelica polysaccharide - PCL 2000 > 187.1±7.451 0.218±0.096 <![CDATA[Angelica polysaccharide - PLGA 1000 > 283.1±6.035 0.178±0.067

[0094] Example 8: Nano-Astragalus polysaccharides in different proportions, using Labrafac MC60, Labrafil M2125 CS, and linoleic acid as excipients.

[0095] A 5 mg / mL Astragalus polysaccharide (APS) aqueous solution was prepared. A 4 mg / mL ethanol solution was prepared from Labrafac MC60, Labrafil M2125 CS, and linoleic acid. 0.5 mL of the ethanol solution (containing 2 mg of excipients) was then added dropwise to 2 mL (containing 10 mg of polysaccharide), 4 mL (containing 20 mg of polysaccharide), 8 mL (containing 40 mg of polysaccharide), and 20 mL (containing 100 mg of polysaccharide) polysaccharide solutions under ultrasonic conditions. The solutions were then sonicated for 5 min, and the organic phase was removed by rotary evaporation at 45 °C. Particle size, PDI, and potential were measured. The results are shown in Tables 11, 12, and 13, indicating that nano-polysaccharides were successfully prepared in all cases.

[0096] Table 11 Particle size, PDI, and potential of different proportions of nano-Astragalus polysaccharides with Labrafac MC60 as an excipient.

[0097] Polysaccharide-excipient ratio Particle size (nm) PDI Potential (mV) 2：1 171.9±1.935 0.242±0.002 -51±2.34 3：1 206.3±5.147 0.166±0.02 -31.7±8.56 5：1 225.8±13.76 0.233±0.056 -25.4±1.69 10：1 196.2±7.919 0.211±0.074 -16.6±0.577 20：1 191.5±14.01 0.232±0.04 -42.8±1.44 50：1 265.6±8.585 0.230±0.107 -36.7±1.37

[0098] Table 12 Particle size, PDI, and potential of Labrafil M2125 CS as an excipient with different proportions of nano-Astragalus polysaccharides

[0099] Drug loading ratio Particle size (nm) PDI Potential (mV) 2：1 214.0±6.133 0.150±0.050 -71.7±12.5 3：1 198.5±2.967 0.136±0.038 -36.3±2.86 5：1 203.9±0.4583 0.164±0.025 -23.6±2.00 10：1 160.3±3.318 0.217±0.088 -37.1±3.16 20：1 157.1±5.98 0.169±0.052 -35.1±1.6 50：1 229.4±8.062 0.304±0.094 -19.8±1.80

[0100] Table 13 Particle size, PDI, and potential of nano-Astragalus polysaccharides with different proportions and linoleic acid as an excipient.

[0101] Drug loading ratio Particle size (nm) PDI Potential (mV) 2：1 223.0±8.046 0.149±0.034 -71.1±2.55 3：1 217.7±9.672 0.24±0.02 -47.8±1.65 5：1 197.0±7.142 0.168±0.043 -51.7±4.85 10：1 232.6±19.55 0.198±0.149 -56.6±4.7 20：1 231.7±4.45 0.242±0.122 -40.9±2.9 30：1 208.3±4.02 0.232±0.135 -38.6±3.3 50：1 203.4±11.37 0.269±0.078 -34.1±3.51

[0102] The particle size was measured again after being placed at room temperature for one week. The results are shown in Tables 14, 15 and 16. It can be seen that the nano-astragalus polysaccharides prepared by the three excipients have good particle size stability when placed at room temperature.

[0103] Table 14. Particle size, PDI, and potential of different proportions of Astragalus membranaceus polysaccharide with different proportions of Labrafac MC60 as excipients after 7 days of storage.

[0104] Drug loading ratio Particle size (nm) PDI Potential (mV) 2：1 172.3±4.403 0.256±0.035 -36.8±0.608 3：1 220.4±7.255 0.155±0.059 -10.9±1.61 5：1 211.6±6.646 0.193±0.047 -31.3±3.81 10：1 203.6±17.85 0.232±0.18 -12.9±1.72 20：1 158.0±16.36 0.243±0.024 -24.5±4.80 50：1 241.6±7.936 0.208±0.135 -16.8±1.34

[0105] Table 15. Particle size, PDI, and potential of Labrafil M2125 CS as an excipient with different proportions of nano-Astragalus polysaccharides after 7 days.

[0106] Drug loading ratio Particle size (nm) PDI Potential (mV) 2：1 201.8±2.358 0.170±0.026 -75.7±6.24 3：1 214.8±3.233 0.113±0.050 -25.1±4.39 5：1 192.3±2.25 0.117±0.041 -26.3±3.0 10：1 156.6±6.529 0.289±0.024 -57.4±6.62 20：1 161.8±4.751 0.209±0.047 -37.6±5.76

[0107] Table 16. Particle size, PDI, and potential of nano-sized astragalus polysaccharides with different proportions and linoleic acid as excipients after 7 days of storage.

[0108] Drug loading ratio Particle size (nm) PDI Potential (mV) 2：1 237.9±8.632 0.169±0.019 -49.3±1.58 3：1 250.3±39.04 0.274±0.050 -30.6±6.77 5：1 206.8±9.3 0.183±0.013 -25.5±1.19 10：1 256.5±21.92 0.239±0.072 -65±1.47 20：1 264.1±7.00 0.253±0.176 -29.9±1.95 30：1 213.4±7.074 0.277±0.062 -27.4±1.45 50：1 211.2±3.007 0.212±0.031 -21.3±4.11

[0109] Example 9: Caprylic / Capric Triglyceride Induces Multiple Nanoparticles (Polysaccharide-Excipient = 5:1)

[0110] Prepare aqueous solutions of polysaccharides from Astragalus membranaceus, Plantago asiatica, Trametes versicolor, Lentinus edodes, soybean, Angelica sinensis, and Lycium barbarum at a concentration of 5 mg / mL. Prepare methanol solutions of 4 mg / mL Labrafac™ Lipophile WL 1349 (caprylic / capric acid medium-chain triglycerides) at a concentration of 0.5 mL each. 载 =2mg) was infused into 2mL (m) under ultrasonic conditions. 药 =10mg) of polysaccharide solution, continue sonication for 5 min, remove organic phase by rotary evaporation at 45 ℃, and measure particle size, PDI and potential. The results are shown in Table-17. It can be seen that nano polysaccharides were successfully prepared.

[0111] After homogenization at 1400 bar for 10 cycles, the particle size, PDI, and potential were measured. The results are shown in Table 18. It can be seen that high-pressure homogenization can be used to reduce the average particle size and particle size distribution of the obtained nanopolysaccharides if necessary.

[0112] Table 17 Particle size, PDI, and potential of nano-Astragalus polysaccharides using caprylic / capric acid medium-chain triglycerides as excipients

[0113] Particle size (nm) PDI Potential (mV) Astragalus polysaccharides 235.1±9.902 0.231±0.064 -14.3±0.557 Plantain polysaccharide 244.8±11.57 0.272±0.047 -14.3±0.500 Yunzhi polysaccharide 262.9±7.702 0.190±0.125 -15.0±0.681 Lentinan 288.5±8.182 0.293±0.128 -22.5±0.907 Lycium barbarum polysaccharides 208.2±4.652 0.234±0.023 -32.4±1.01 Angelica polysaccharides 241.9±2.887 0.217±0.035 -14.3±0.557

[0114] Table 18. Particle size, PDI, and potential of nano-Astragalus polysaccharide with medium-chain triglycerides as excipient after homogenization at 1400 bar for 10 cycles.

[0115] Particle size (nm) PDI Potential (mV) Astragalus polysaccharides 204.7±6.240 0.181±0.051 -28.2±1.85 Plantain polysaccharide 185.0±1.041 0.046±0.045 -24.2±2.61 Yunzhi polysaccharide 177.1±2.255 0.126±0.041 -37.0±1.05 Lentinan 181.7±2.615 0.179±0.015 -35.8±1.51 Lycium barbarum polysaccharides 174.3±6.199 0.162±0.025 -44.6±2.38 Angelica polysaccharides 208.9±14.41 0.183±0.032 -27.7±0.902

[0116] Example 10: Various nanopolysaccharides with stearic acid as an excipient

[0117] Aqueous solutions of polysaccharides from Astragalus membranaceus, Plantago asiatica, Trametes versicolor, Lentinus edodes, soybean, Angelica sinensis, and Lycium barbarum were prepared at a concentration of 5 mg / mL. A methanol solution of 4 mg / mL was prepared by taking a certain amount of stearic acid. 0.5 mL of the methanol solution was added dropwise to 2 mL of the polysaccharide solution under ultrasonic conditions. The mixture was ultrasonicated for another 5 min, and the organic phase was removed by rotary evaporation at 45 °C. The particle size, PDI, and potential were measured. The results are shown in Table 19. It can be seen that nano-polysaccharides were successfully prepared in all cases.

[0118] After homogenization at 1400 bar for 10 cycles, the particle size, PDI, and potential were measured. The results are shown in Table 20. It can be seen that high-pressure homogenization can be used to reduce the average particle size and particle size distribution of the obtained nanopolysaccharides if necessary.

[0119] Table 19 Particle size, PDI, and potential of nanopolysaccharides with stearic acid as an excipient (polysaccharide-excipient = 5:1)

[0120] Particle size (nm) PDI Potential (mV) Astragalus polysaccharide NPs 487.9±23.05 0.117±0.126 -22.9±0.737 Plantain polysaccharide NPs 517.2±26.75 0.234±0.051 -12.3±0.755 Yunzhi polysaccharide NPs 484.5±9.58 0.063±0.051 -11.4±0.896 Lentinan NPs 437.3±10.80 0.154±0.062 -26.4±0.231 Angelica polysaccharide NPs 409.7±15.56 0.211±0.018 -13.2±0.361 Soybean polysaccharides NPs 317.4±8.62 0.156±0.066 -17.6±1.04

[0121] Table 20. Particle size, PDI, and potential of nanoassemblies with stearic acid as an excipient after 10 homogenization cycles at 1404 bar.

[0122] Category Particle size (nm) PDI Potential (mV) Astragalus polysaccharide NPs 303.2±18.14 0.113±0.033 -40.1±2.20 Plantain polysaccharide NPs 305.2±14.21 0.201±0.066 -32.9±1.35 Yunzhi polysaccharide NPs 328.8±4.706 0.128±0.098 -28.1±1.83 Lentinan NPs 336.1±14.55 0.112±0.061 -46.3±2.65 Angelica polysaccharide NPs 286.8±17.39 0.092±0.049 -28.1±1.95 Soybean polysaccharides NPs 264.0±6.274 0.171±0.125 -31.1±2.91

[0123] Example 11 Nanoassemblies of multiple polysaccharides with oleic acid as an excipient (polysaccharide: excipient = 5:1)

[0124] 40 mg of polysaccharide was weighed and dissolved in 4 mL of deionized water. After sonication for 5 min, the resulting solution was a polysaccharide solution (10 mg / mL). 54 mg of oleic acid was dissolved in 3 mL of acetone. 444 μl of each solution (approximately 8 mg in total) was added dropwise to 4 mL of the polysaccharide solution under sonication. After sonication for another 5 min, the organic solvent was removed by rotary evaporation under reduced pressure at 45°C. The solution was homogenized 10 times at 1400 bar. The particle size, PDI, and potential were measured. The results are shown in Table 21. It can be seen that nano-polysaccharides were successfully prepared.

[0125] Table 21 Particle size, PDI, and potential of nanopolysaccharides with oleic acid as an excipient

[0126] After homogenization Size (nm) PDI Zeta (mV) Soybean polysaccharides 141.9±2.364 0.194 -3.72 Plantain polysaccharide 169.4±5.519 0.184 -2.68 Yunzhi polysaccharide 156.0±1.644 0.170 -0.97 Astragalus polysaccharides 207.1±8.334 0.219 -2.40 Lentinan 171.0±6.191 0.179 +0.728 Angelica polysaccharides 107.4±1.453 0.200 +0.683

[0127] Example 12 Nanoparticles loaded with protopanaxadiol PPD using TPGS as an excipient

[0128] Accurately weigh 5 mg of protopanaxadiol (PPD) and dissolve it in 0.5 mL of anhydrous ethanol. Accurately weigh 30 mg of polysaccharide and 5 mg of TPGS and dissolve them together in 5 mL of water to form the aqueous phase. Slowly drip the ethanol solution into the aqueous phase using ultrasound, and remove the organic solvent by rotary evaporation at 45 °C. The particle size, PDI, and Zeta potential were measured, and the results are shown in Table 22 below. It can be seen that nano-polysaccharides were successfully prepared.

[0129] Table 22 Particle size, PDI, and potential of nanopolysaccharides loaded with PPD using TPGS as an excipient

[0130] Size (nm) PDI Zeta (mV) Soybean polysaccharide-PPD-TPGS (6:1:1) 176.7±2.007 0.216±0.011 -0.052±0.042 Lentinan-PPD-TPGS (6:1:1) 149.7±1.124 0.204±0.017 -9.19±2.13 Angelica polysaccharide-PPD-TPGS (6:1:1) 193.0±5.250 0.268±0.015 -8.43±0.552 Yunzhi polysaccharide-PPD-TPGS (6:1:1) 209.3±2.579 0.205±0.023 -7.82±0.658 Plantain seed polysaccharide-PPD-TPGS (6:1:1) 201.2±1.069 0.241±0.021 -8.91±0.737 Astragalus polysaccharide-PPD-TPGS (6:1:1) 242.4±47.09 0.356±0.105 -6.48±1.85 Lentinan-PPD-TPGS (5:1:0.5) 203.6±2.879 0.185±0.022 -9.40±0.090 Lentinan-PPD (4:1) 265.2±0.764 0.067±0.023 -22.3±0.839

[0131] Example 13: Nano-soybean polysaccharides and nano-astragalus polysaccharides with TPGS as excipients were further loaded with ginsenoside CK.

[0132] 5 mg of ginsenoside CK was accurately weighed and dissolved in 0.5 mL of anhydrous ethanol. 30 mg of soybean polysaccharide or astragalus polysaccharide and 5 mg of TPGS were accurately weighed and dissolved together in 5 mL of water to form the aqueous phase. The ethanol solution was slowly added dropwise to the aqueous phase under ultrasonication. The organic solvent was removed by rotary evaporation under reduced pressure at 45 °C. The particle size, PDI, and Zeta potential were measured. The results are shown in Table 23, indicating that nano-polysaccharides were successfully prepared and successfully loaded with ginsenoside CK.

[0133] Table 23 Particle size, PDI, and potential of nano-soybean polysaccharides and nano-astragalus polysaccharides loaded with ginsenoside CK using TPGS as excipients.

[0134] Size (nm) PDI Zeta (mV) Soybean polysaccharide-CK-TPGS (5:1:1) 435±1.153 0.122±0.016 -6.51±0.190 Homogenize 10 times at 1400 bar 321.6±15.01 0.064±0.069 -6.48±1.85 Astragalus polysaccharide-CK-TPGS (5:1:1) 203.6±2.879 0.185±0.022 -9.40±0.090 Homogenize 10 times at 1400 bar 165.2±0.764 0.067±0.023 -22.3±0.839

[0135] Example 14: Nanoparticles of ginsenoside CK loaded with oleic acid as an excipient (polysaccharide-CK-oleic acid = 8:1:2)

[0136] 5 mg of ginsenoside CK and 10 mg of oleic acid were accurately weighed and dissolved in 1 mL of anhydrous ethanol. 40 mg of polysaccharide was accurately weighed and dissolved in 5 mL of water. The ethanol solution was slowly added dropwise to the polysaccharide solution under ultrasonication at 150 W. The ethanol was removed by rotary evaporation under reduced pressure at 45 °C. The particle size, PDI, and Zeta potential were detected. The results are shown in Table 24. It can be seen that nano-polysaccharides were successfully prepared and successfully loaded with ginsenoside CK.

[0137] Table 24 Particle size, PDI, and potential of nanopolysaccharides loaded with PPD using oleic acid as an excipient

[0138] Size (nm) PDI Zeta (mV) Lentinan-CK-Oleic Acid 205.3±4.506 0.220±0.044 -(25.1±1.19) Angelica polysaccharide-CK-oleic acid 317.7±7.991 0.256±0.025 -(20.9±0.709) Yunzhi polysaccharide-CK-oleic acid 221.9±5.765 0.258±0.012 -(2.17±0.260) Psyllium polysaccharide-CK-oleic acid 237.9±3.062 0.254±0.043 -(20.0±0.173) Yunzhi polysaccharide-CK-oleic acid (4:1:1) 210.4±0.586 0.135±0.037 -(16.1±0.424)

[0139] Example 15: Ginsenoside Rh2 loaded onto nano-polysaccharide with oleic acid as an excipient (polysaccharide-oleic acid-Rh2=5:2:1)

[0140] 5 mg of Rh2 and 10 mg of oleic acid were accurately weighed and dissolved in 500 μL of anhydrous ethanol using ultrasonication. 25 mg of soybean polysaccharide was dissolved in 5 mL of water. The ethanol solution was added dropwise to the polysaccharide solution under ultrasonication in a 200 W water bath. The ethanol was removed by rotary evaporation under reduced pressure at 45 °C. The particle size, PDI, and Zeta potential were measured. The results are shown in Table 25, indicating that nano-polysaccharides were successfully prepared and successfully loaded with ginsenoside Rh2.

[0141] Table 25 Particle size, PDI, and potential of nanopolysaccharides loaded with ginsenoside Rh2 using oleic acid as an excipient.

[0142] Size (nm) PDI Zeta (mV) Soybean polysaccharides 206±8.669 0.172±0.054 -14.0±1.35 Astragalus polysaccharides 309±8.942 0.064±0.03 -15.2±0.608 Lentinan 155.5±2.71 0.244±0.03 -18.2±1.24 Plantain polysaccharide 158.7±3.002 0.202±0.015 -14.5±1.19 Angelica polysaccharides 286.2±4.967 0.112±0.031 -12.7±1.30 Yunzhi polysaccharide 138.6±1.55 0.18±0.036 -12.8±0.656

[0143] Example 16: Ginsenoside Rh2 loaded onto nano-polysaccharides with oleic acid as an excipient (polysaccharide-oleic acid-Rh2=4:1:1)

[0144] 5 mg of ginsenoside Rh2 and 5 mg of oleic acid were accurately weighed and dissolved in 600 μL of anhydrous ethanol using ultrasonication. 20 mg of soybean polysaccharide was dissolved in 5 mL of water. The ethanol solution was added dropwise to the polysaccharide solution under ultrasonication at 250 W. The organic solvent was removed by rotary evaporation under reduced pressure at 45 °C. The particle size, PDI, and Zeta potential were measured. The results are shown in Table 26, indicating that nano-polysaccharides were successfully prepared and successfully loaded with ginsenoside Rh2.

[0145] Table 26. Particle size, PDI, and potential of nanopolysaccharides loaded with ginsenoside Rh2 using oleic acid as an excipient.

[0146] Size (nm) PDI Zeta (mV) Lentinan 135.5±2.751 0.233±0.008 -19.0±1.05 Yunzhi polysaccharide 142.9±2.155 0.223±0.025 -13.3±0.451

[0147] Example 17: Nanoparticles of ginsenoside Rh2 loaded with oleic acid as an excipient (polysaccharide-oleic acid-Rh2=4:1:2, drug loading 28.57%)

[0148] 10 mg of Rh2 and 5 mg of oleic acid were accurately weighed and dissolved in 600 μL of anhydrous ethanol by ultrasonication. 20 mg of soybean polysaccharide was dissolved in 5 mL of water. The ethanol solution was added dropwise to the polysaccharide solution under ultrasonication. The ethanol was removed by rotary evaporation under reduced pressure at 45 °C. The particle size, PDI, and Zeta potential were detected. The results are shown in Table 27 below. It can be seen that nano-polysaccharides were successfully prepared and successfully loaded with ginsenoside Rh2.

[0149] After being stored at room temperature for one week, there were no significant changes in particle size and PDI (Table 26), indicating that the drug-loaded nanopolysaccharides have good stability.

[0150] Table 27 Particle size, PDI, and potential of oleic acid-based nanopolysaccharides loaded with Rh2

[0151] Size (nm) PDI Zeta (mV) freshly prepared Lentinan 209.3±5.677 0.239±0.073 -10.4±1.66 Yunzhi polysaccharide 211.1±1.795 0.295±0.032 -14.9±0.651 Leave at room temperature for 1 week Yunzhi polysaccharide 173.2±4.173 0.247±0.062 -11.2±0.306 Lentinan 160.0±3.502 0.262±0.04 -17.0±2.48

[0152] Example 18 Preparation of nano-lily polysaccharides with oleic acid as an excipient and preparation of composite excipient-composite polysaccharide nanoaggregates

[0153] Accurately weigh 150 mg of lily polysaccharide (sugar content >80%), astragalus polysaccharide, and shiitake polysaccharide, and prepare 5 mL of 50 mg / mL polysaccharide aqueous solution with deionized water. Accurately measure a certain amount of oleic acid (OA), medium-chain triglycerides of caprylic / capric acid and capric acid (Labrafac™ Lipophile WL 1349), polycaprolactone (PCL) with a molecular weight of 1000, and polylactic acid (PLA) with a molecular weight of 1000, and dissolve them in ethanol to prepare a solution with a concentration of 10 mg / mL. Take an appropriate amount of ethanol solution and add it dropwise to the polysaccharide solution under ultrasonic conditions of 250 W. Remove the ethanol by rotary evaporation under reduced pressure at 45 °C. Detect the particle size, PDI, and Zeta potential. The results are shown in Table 28. It can be seen that nano-polysaccharides can be successfully prepared without single polysaccharides or single lipid excipients. Nano-polysaccharides can also be successfully prepared with single polysaccharides on composite excipients, composite polysaccharides on single excipients, and composite polysaccharides on composite excipients.

[0154] Table 28 Particle size, PDI, and potential of lily polysaccharide and composite excipients-composite polysaccharide nanoassemblies

[0155] mass ratio Particle size PDI Zate Lily polysaccharide-oleic acid 10:1 172.6±2.4 0.163±0.04 -31.2 Lily polysaccharide-medium chain triglyceride 10:1 141.5±2.0 0.146±0.03 -28.5 Lily polysaccharide-PCL 10:1 209.2±3.3 0.182±0.05 -30.7 Lily polysaccharide-oleic acid-medium chain triglyceride-PCL 30:1:1:1 156.2±2.3 0.153±0.06 -32.6 Lily polysaccharide-Astragalus polysaccharide-Oleic acid 4:4:1 188.9±3.5 0.187±0.06 -30.2 Lily polysaccharide-Astragalus polysaccharide-PLA 5:5:1 214.6±3.1 0.201±0.06 -33.7 Lily polysaccharide-Astragalus polysaccharide-Lentinula polysaccharide-Oleic acid-Medium chain triglyceride-PLA 8:8:8:1:1:1 201.5±3.3 0.177±0.05 -29.4

[0156] Example 19 Preparation of nano-carob polysaccharides and mixed polysaccharide nanoparticles: oleic acid induced

[0157] Methods: Carob polysaccharide of uniform molecular weight (PDI < 1.6, polysaccharide content > 93%, referred to as carob polysaccharide in this invention, abbreviated as CSPS) was accurately weighed and dissolved in deionized water to prepare a 10 mg / mL solution. Oleic acid (OA) of 7.5 mg, 10 mg, and 15 mg was accurately measured and dissolved in 1 mL of methanol. The solutions were then added dropwise to the carob polysaccharide solution under ultrasonication at 250 W. The methanol was removed by rotary evaporation under reduced pressure at 35 °C to obtain CSPS nanopolysaccharides (CSPS@OA NPS). Particle size, PDI, and Zeta potential were measured, and the results are shown in Table 28. It can be seen that nanopolysaccharides were successfully prepared using these methods. The particle size and particle size distribution of the carob polysaccharide nanopolysaccharide CSPS@OA NPS with a polysaccharide-oleic acid ratio of 15:1 are shown in the appendix. Figure 1 .

[0158] Total carob polysaccharide (the effective polysaccharide fraction containing CSPS, with a polysaccharide content of approximately 82%) was used to replace carob polysaccharide and prepared using the same method. The results are shown in Table 29. It can be seen that nano-polysaccharides were successfully prepared in both cases, and it seems that it is easier to prepare than pure carob polysaccharide.

[0159] Equal amounts of carob polysaccharide, astragalus polysaccharide, and wolfberry polysaccharide were mixed, and nano-polysaccharides were prepared by mixing the total polysaccharide with oleic acid at a ratio of 9:1 (mass ratio). The results are shown in Table 28, indicating that nano-polysaccharides were successfully obtained by mixing polysaccharides.

[0160] Table 29 Particle size, PDI, and potential of carob polysaccharide and its complex polysaccharide nanoassemblies with oleic acid as an excipient.

[0161] Carob polysaccharide-oleic acid mass ratio Particle size PDI Zate 10:1 158.8±1.7 0.231±0.05 -38.4 Carob polysaccharide-oleic acid 15:1 122.5±2.5 0.235±0.03 -29.7 20:1 109.4±1.2 0.330±0.02 -26.4 10:1 146.2±1.3 0.161±0.04 -37.8 Carob total polysaccharide-oleic acid 15:1 132.9±2.1 0.157±0.03 -35.1 20:1 115.3±1.7 0.232±0.05 -32.6 Carob polysaccharides-Astragalus polysaccharides-Oleic acid 4.5:4.5:1 168.5±3.8 0.191±0.07 -30.5 Total polysaccharides from carob, astragalus, wolfberry, and oleic acid 3:3:3:1 183.6±4.3 0.230±0.09 -35.6

[0162] Example 20 Morphology of nanocarob polysaccharide CSPS@OA NPS

[0163] Carob polysaccharide CSPS and carob nanopolysaccharide CSPS@OA NPS were diluted with pure water to a polysaccharide content of approximately 1 mg / mL. Approximately 10 μL of each was dropped onto a 300-mesh copper grid and allowed to air dry. The grids were then stained with 2% (w / v) uranyl acetate for 2 minutes. The morphology of carob polysaccharide CSPS and carob nanopolysaccharide CSPS@OA NPS was then observed under an accelerating voltage of 120 kV. (See attached...) Figure 2 As shown, carob nanopolysaccharide CSPS@OA NPS exhibits a regular spherical shape under a microscope.

[0164] Example 21: Room temperature and particle size stability of nanocarob polysaccharide CSPS@OA NPS in physiological media

[0165] To understand the room-temperature stability and particle size stability of carob nanopolysaccharides in physiological media, freshly prepared CSPS@OA NPS were stored at 4°C, and samples were taken at specific time points (1, 3, 5, 7, 9, 11, 13, and 15 days) to evaluate particle size and PDI. Results are attached. Figure 3 It can be seen that the particle size and PDI of carob nanopolysaccharide did not change significantly during the storage process, indicating good stability.

[0166] To investigate the particle size stability of carob nanopolysaccharides in physiological media, CSPS@OA NPS were mixed with equal volumes of 1.8% NaCl, 2×PBS, 10% glucose, or four times the volume of artificial gastric fluid, artificial intestinal fluid, or mouse plasma at 37°C. Samples were taken at different time intervals to measure the particle size and PDI value. The results are attached. Figure 4 As shown, carob nanopolysaccharides did not exhibit significant changes in particle size and PDI in the six physiological media tested, demonstrating good stability and suitability for direct oral or intravenous administration.

[0167] Example 22 Freeze-drying and reconstitution reconstruction of nanocarob polysaccharide CSPS@OA NPS

[0168] Freshly prepared CSPS@OA NPS was pre-frozen at -20℃ and then freeze-dried. 30 mg of the freeze-dried powder was added to 3 mL of pure water and shaken. It was found that it dissolved rapidly. The particle size after reconstitution was measured to be 142±3.2 mm, and the PDI value was 0.261±0.11, which was almost the same as the particle size and PDI value before freeze-drying. This shows that the nanocarob polysaccharide CSPS@OA NPS can be freeze-dried without any freeze-drying protectant, which is convenient for long-term storage. It can be restored to nanopolysaccharide by adding water and shaking before use.

[0169] Example 23 FITC fluorescent labeling of carob polysaccharide CSPS

[0170] 100 mg of carob polysaccharide (CSPS) was dissolved in 5 mL of water, mixed with 2.5 mL of 2.6 M NaOH solution and 40 μL of ammonium hydroxide, and stirred at 40 °C for 2 h. The mixture was then transferred to a dialysis bag with a molecular weight cutoff of 8000-14000 and dialyzed against water at 25 °C for 48 h, followed by freeze-drying. The freeze-dried residue was dissolved in 5.0 mL of 30% ammonium hydroxide and amination was performed at 40 °C for 90 min. The reaction mixture was dialyzed against water for 48 h (molecular weight cutoff 8000-14000) and then freeze-dried. The freeze-dried sample was dissolved in 5 mL of 0.5 M sodium bicarbonate solution, and 20 mg of FITC was added. The mixture was stirred at room temperature in the dark for 24 h, and anhydrous ethanol was added to 80% (V / V) to precipitate the product. The precipitate was centrifuged at 5000 rpm for 10 min, collected, dissolved in water, dialyzed against water in the dark for 24 h, and then freeze-dried to obtain the CSPS-FITC fluorescently labeled product.

[0171] Example 24 Near-infrared fluorescent labeling of carob polysaccharide CSPS

[0172] 100 mg of carob polysaccharide (CSPS) was dissolved in 5 mL of water, mixed with 2.5 mL of 2.6 M NaOH solution and 40 μL of ammonium hydroxide, and stirred at 40 °C for 2 h. The mixture was then transferred to a dialysis bag with a molecular weight cutoff of 8000-14000, dialyzed against water at 25 °C for 48 h, and lyophilized. The lyophilized residue was dissolved in 5.0 mL of 30% ammonium hydroxide, amination was performed at 40 °C for 90 min, and the reaction mixture was dialyzed against water for 48 h (molecular weight cutoff 8000-14000) and then lyophilized. The lyophilized powder was dissolved in 5 mL of 0.5 M sodium bicarbonate solution, 4 mg of IR783 was added, and the mixture was stirred in the dark at room temperature for 24 h. Anhydrous ethanol was added to 80% (V / V) to precipitate the product. The precipitate was centrifuged at 5000 rpm for 10 min, collected, dissolved in water, dialyzed against water in the dark for 24 h, and lyophilized to obtain the CSPS-IR783 fluorescently labeled product.

[0173] Example 25: In vitro anti-inflammatory activity study of Astragalus polysaccharide APS and nano-Astragalus polysaccharide APS@OA NPS

[0174] RAW264.7 cells in logarithmic growth phase were seeded in 96-well plates (density 1×10⁻⁶). 5 Cells were cultured for 1 day. Astragalus polysaccharide solution and nano-Astragalus polysaccharide were diluted with fresh DMEM medium to different concentrations (0.1, 0.5, 1.0, 2.0, 5.0, 10, 20, 50, 100 μg / mL CSPS) and added to designated wells (200 μL each). After incubation for 48 h, 10 μL of CCK-8 reagent was added to each well, followed by 2 h of incubation. Absorbance was measured at 450 nm using a microplate reader. Cell viability (%) was determined using the following formula: Cell inhibition rate (%) = (1 - OD1 / OD2) × 100%, OD1: sample group; OD2: blank group. The CCK-8 experiment results showed that, within 24 hours, free polysaccharide and nano-polysaccharide in the range of 0-300 μg / mL had no effect on the proliferation ability of RAW 264.7 cells (see attached diagram). Figure 5 Therefore, free polysaccharides and nanopolysaccharides at concentrations of 10, 100, and 300 μg / mL were selected to evaluate their in vitro anti-inflammatory effects on RAW 264.7 cells.

[0175] RAW 264.7 macrophage inflammation models are widely used to evaluate the anti-inflammatory activity of drugs. RAW264.7 cells in logarithmic growth phase were seeded into 96-well plates at a density of 1 × 10⁻⁶ cells / well. 5 Cells were cultured for 24 hours, and then stimulated with 1 μg / mL LPS for 24 hours to obtain a cellular inflammation model. The supernatant was discarded, and blank DMEM medium, polysaccharide solutions (10, 100, and 300 μg / mL, diluted with fresh DMEM), and nanopolysaccharides (10, 100, and 300 μg / mL, diluted with fresh DMEM) were added to each well. After incubation for 48 hours, 50 μL of Griess solution was seeded into each well, and the cells were cultured for 15 minutes. NO levels in the cells were measured using a commercial assay kit following the instructions on the kit. Similarly, the concentrations of tumor necrosis factor-α (TNF-α), interleukin-12, and IL-6 (IL-6) in the cells were assessed using an enzyme-linked immunosorbent assay (ELISA) kit following the instructions on the kit.

[0176] NO, IL-6, and TNF-α are important inflammatory markers, while IL-10 is an important anti-inflammatory marker. The production of pro-inflammatory factors in RAW 264.7 cells after incubation with free polysaccharides and nanopolysaccharides for 24 hours was detected using an ELISA kit. Figure 5C-5F analysis showed that free polysaccharides significantly reduced the secretion of NO, IL-6, and TNF-α, and significantly increased the production of IL-10 in a dose-dependent manner, exhibiting good anti-inflammatory activity. At the same concentration (100 μg / mL or 300 μg / mL), nano-polysaccharides significantly reduced NO (P<0.05), IL-6 (P<0.01), and TNF-α (P<0.01) levels, and significantly increased IL-10 levels (P<0.05) compared to free polysaccharides, indicating that nano-assembly greatly enhanced the anti-inflammatory activity of the polysaccharides.

[0177] Example 26: In vivo antitussive, expectorant, and anti-inflammatory activities of lily polysaccharide and nano-lily polysaccharide.

[0178] Ninety male ICR mice, weighing 21±2g, were used. Inflammation and coughing were induced in the mice using endotracheal intubation combined with lipopolysaccharide (LPS) and ammonia inhalation. Mice were adaptively fed for 3 days and then randomly divided into 9 groups (n=10 per group), with the administration regimens shown in Table-30. The nanopolysaccharide used in this study was the nano-lily polysaccharide with oleic acid as an excipient, as described in Example 18. The polysaccharide solution for intravenous injection was dispersed in physiological saline, and the nanopolysaccharide was adjusted to isotonicity with sodium chloride before administration.

[0179] Table 30 Grouping and Dosing Regimens

[0180] Grouping Mouse type Dosing regimen dose Blank control normal mice Normal saline, orally 0.2 mL Model comparison Model mice Normal saline, orally Cough-relieving positive control Model mice Phenylacetyl phosphate tablets (oral administration) 60 mg / kg Positive control for expectoration Model mice Ammonium chloride tablets (oral administration) 500 mg / kg Anti-inflammatory positive control Model mice Dexamethasone (iv) 5 mg / kg Polysaccharide solution (oral administration) Model mice Polysaccharide solution (oral administration) 100 mg / kg Nanoparticles (oral administration) Model mice Nanoparticles (oral administration) 100 mg / kg Polysaccharide solution (intravenous injection) Model mice Polysaccharide solution (intravenous injection) 50 mg / kg Nanoparticles (intravenous injection) Model mice Nanoparticles (intravenous injection) 50 mg / kg

[0181] Except for the blank control group, LPS (0.4 mg / mL) (10 μL / 10 g mouse body weight) was administered via air tube intubation on days 1, 7, and 15. On days 7 and 14, 0.2 mL of 25% ammonia solution was placed on a cotton ball and placed in a 1 L inverted beaker. Except for the blank control group, each mouse was quickly placed in the beaker for 2 minutes. The model was considered successfully established if the mice in the model group exhibited frequent neck extension, mouth opening, back arching, abdominal muscle twitching, and coughing more than 20 times within 2 minutes.

[0182] Phenylacetyl phosphate tablets (60 mg / kg) were used as a positive control for treating cough. One hour after the last administration, each mouse was placed in a 500 mL glass jar saturated with 0.2 mL of 25% ammonia water, and the latency and frequency of cough within 2 minutes were observed and recorded.

[0183] Subsequently, the expectorant efficacy was immediately evaluated on the same mice using the phenol red secretion method, with ammonium chloride as the positive control. After intraperitoneal injection of 3.5% phenol red solution for 30 min, the mice were anesthetized with 1% sodium pentobarbital (sodium pentobarbital), and the trachea and a section of bronchus were immediately dissected. The trachea was immersed in 1 mL of 5% sodium bicarbonate solution, sonicated for 15 min, and centrifuged at 3000 rpm for 10 min. The absorbance of the supernatant was measured at 546 nm using an ELISA reader, and the phenol red content was calculated. A phenol red standard curve was plotted according to the literature.

[0184] Finally, the anti-inflammatory activity was evaluated by inducing acute airway inflammation in mice with ammonia. After the mice were sacrificed, the right lung was collected, washed with pre-cooled PBS, homogenized using a high-throughput tissue homogenizer (lung tissue:PBS = 1g:3mL), centrifuged at 1000 rpm for 10 min at 4°C, and the supernatant was collected. IL-6, IL-12, and TNF-α were measured by ELISA. Simultaneously, the left lung was fixed by immersion in 4% formaldehyde solution for 24 hours, dehydrated with ethanol solution, embedded in paraffin, sectioned at 4 μm, and stained with hematoxylin and eosin (H&E) for pathological observation.

[0185] Results: The in vivo antitussive effects of free polysaccharides and nanopolysaccharides were compared using an ammonia-induced cough model. The results are shown in the attached figure. Figure 6 As shown in A and 6B. After 14 days of treatment, compared with the model control group, the number of coughs decreased by 67.87% with oral administration of free polysaccharides and by 88.97% with injection. The number of coughs decreased by 78.82% with oral administration of nano-polysaccharides and by 97.21% with injection, both significantly better than polysaccharide solutions (P<0.05) and also significantly better than phenylpropyl phosphate positive control drugs (P<0.05 and P<0.01). Simultaneously, the cough latency period was also significantly prolonged. Figure 6 B) Oral and intravenous administration of free polysaccharides prolonged the cough latency by 373.57% and 549.32%, respectively. The prolongation of the cough latency by nanopolysaccharides was even more significant, being 1.44 times (oral) and 1.54 times (intravenous) that of free polysaccharides (both P<0.01), and significantly superior to phenylpropyl phosphate positive control drugs (P<0.05 and P<0.01). Whether administered orally or intravenously, nanopolysaccharides were significantly more effective than polysaccharide solutions in treating cough (P<0.05, P<0.01). Whether considering cough latency (polysaccharide P<0.05, nanopolysaccharide P<0.01) or the number of coughs within 2 minutes (both groups P<0.01), intravenous administration was significantly more effective than oral administration. Since large polysaccharides are rarely absorbed into the circulation intact, this suggests that polysaccharides may have multiple targets in vivo, in addition to the intestine, or that polysaccharides may have multiple pharmacologically active domains, and that degradation fragments with these active domains can be absorbed into the circulation to exert their pharmacological activity.

[0186] The expectorant activity of free polysaccharides and nanopolysaccharides was detected by examining phenol red secretion in mice. The results are shown in the attached figure. Figure 6 As shown in Figure C. Compared with the model control group, free polysaccharides significantly increased phenol red secretion in rats by 32.08% (oral administration) and 57.08% (intravenous administration), respectively. Nanoparticles further promoted phenol red secretion, with secretion levels 1.87 times that of oral administration and 1.62 times that of intravenous administration of the polysaccharide solution. The results showed that at a dose of 100 mg / kg, free polysaccharides had a strong expectorant effect in vivo, and the expectorant effect of nanoparticles was significantly better than that of free polysaccharides (P<0.05, P<0.01).

[0187] IL-10, TNF-α, and IL-6 are important pro-inflammatory chemokines and cytokines that play a crucial role in adaptive and innate immunity. Due to their pro-inflammatory properties, IL-10, TNF-α, and IL-6 directly participate in the recruitment and activation of inflammatory cells at sites of injury. (See attached image) Figure 6 As shown in Figures D, 6E, and 6F, compared with the model group, the levels of IL-10 in the lung tissue of rats in other groups were significantly increased (P<0.01, 0.001), TNF-α levels were significantly decreased (P<0.0001), and IL-6 levels were significantly decreased (P<0.01, 0.0001). The in vivo anti-inflammatory capacity was in the following order: oral administration of free polysaccharides < intravenous administration of free polysaccharides ≈ oral administration of nanopolysaccharides < intravenous administration of nanopolysaccharides (iv), with intravenously administered nanopolysaccharides exhibiting the strongest in vivo anti-inflammatory capacity. Regardless of whether it was administered orally or intravenously, the in vivo anti-inflammatory activity of nanopolysaccharides was significantly higher than that of free polysaccharides (P<0.05, P<0.01).

[0188] H&E staining showed pathological changes in mouse lung tissue (attached) Figure 6 G). In the control group, the alveolar structure of the lung tissue was clear, with no inflammatory cell infiltration and no alveolar fusion. In the model group, the alveolar walls of the rats' lung tissue showed significant thickening, alveolar mucosal dysfunction, disordered alveolar structure, and a small amount of alveolar fusion. Diffuse infiltration of lymphocytes and neutrophils was observed, with localized minor hemorrhage. In the free polysaccharide oral administration group, the alveolar walls were significantly thickened, accompanied by a small amount of lymphocyte and neutrophil infiltration. In the free polysaccharide injection group, a small amount of alveolar wall thickening was observed, accompanied by a small amount of lymphocyte, neutrophil, and macrophage infiltration. The alveolar structure of the nanopolysaccharide oral administration group was clear. In the nanopolysaccharide injection group, no alveolar wall thickening was observed, inflammatory cell infiltration of the bronchial walls was reduced, bronchial smooth muscle was thinned, and the alveolar structure remained intact. It can be concluded that nanopolysaccharides can better repair damage caused by lung inflammation than free polysaccharides.

[0189] Example 27 In vitro cellular uptake study of carob polysaccharide CSPS and nanocarob polysaccharide CSPS@OA NPS

[0190] Caco-2 cells in logarithmic growth phase were seeded into 24-well plates (1×10⁻⁶ cells / year).6 Cells were cultured at 37°C and 5% CO2 for 24 hours. FITC-labeled free polysaccharide (FITC-CSPS) solution and FITC-CSPS@OA NPS nanopolysaccharide were diluted to 100 µg / mL with serum-free medium and added to wells. The cells were incubated for 1 h, 3 h, and 6 h, respectively. The medium was discarded, and DAPI (5 µg / mL, 0.4 mL) was added to fix the cells in 0.4 mL PBS containing 4% (w / v) paraformaldehyde. Cell uptake of FITC-CSPS and FITC-CSPS@OANPS was observed using a fluorescence inverted microscope. Semi-quantitative fluorescence analysis and co-localization analysis were performed using ImageJ software.

[0191] The results are as follows. Figure 7 As shown, Caco-2 cells exhibited time-dependent cellular uptake of both free polysaccharides and nanopolysaccharides. However, at the same concentration of 100 μg / mL, cellular uptake of nanopolysaccharides was significantly enhanced by more than 5 times compared to free polysaccharides (P < 0.0001), particularly at hours 3 and 6. This part of the experiment demonstrates that cellular uptake of nanopolysaccharides is substantially increased compared to free polysaccharides. This finding partially explains why nanopolysaccharides exhibit better anti-inflammatory activity than polysaccharide solutions.

[0192] Example 28 Pharmacokinetic Study of Carob Polysaccharide CSPS and Nanocarob Polysaccharide CSPS@OA NPS

[0193] Mouse blank plasma was mixed with near-infrared fluorescently labeled carob polysaccharide solution (CSPS-IR783) to prepare a series of solutions with polysaccharide concentrations of 1, 2.5, 5, 10, 20, 40, 80, 160, 320 and 640 μg / mL, and a standard curve was obtained.

[0194] Sixty-six ICR mice were randomly divided into two groups, and administered near-infrared fluorescently labeled carob polysaccharide solution (CSPS-IR783) and carob nanopolysaccharide (CSPS-IR783@OA NPs) via tail vein, respectively, at a dose of 50 mg / kg. At 0.5 h, 1 h, 2 h, 3 h, 4 h, 6 h, 8 h, 10 h, 12, 24 h, and 48 h after administration, blood samples were collected from three mice in each group. Heparinized blood samples were centrifuged at 3500 rpm for 10 min at 4℃, and plasma samples were collected. These samples were mixed with physiological saline (1:4, volume ratio), centrifuged at 3500 rpm for 10 min, and the supernatant was collected. Fluorescence intensity was measured using an enzyme-linked immunosorbent assay (ELISA) reader (excitation wavelength 745 nm, emission wavelength 820 nm). Polysaccharide concentration was calculated from the standard curve, and a plasma polysaccharide concentration versus time curve was plotted to calculate pharmacokinetic parameters.

[0195] Results: See attached. Figure 8 As shown, after a single intravenous injection of the polysaccharide solution, the free polysaccharide was rapidly cleared from the circulation, while the clearance rate of the nanopolysaccharide was slightly slower. Pharmacokinetic parameters calculated by DAS 2.0 software are shown in Table 31. The metabolism of free polysaccharide and nanopolysaccharide in mice approximates a two-compartment model, with a fitting constant R0. 2 > 0.9. Compared with free polysaccharides, although Cmax MRT(0→∞) and Vd only increased slightly, nanopolysaccharides significantly increased AUC(0→t) (1.17 times, P<0.0001) due to the significant prolongation of t(1 / 2)α and t(1 / 2)β. This result partially explains why nanopolysaccharides have better in vivo antitussive, expectorant and anti-inflammatory effects than free polysaccharides when administered intravenously.

[0196] Table 31 Pharmacokinetic parameters of mice after intravenous injection of IR783-labeled free polysaccharide and nanopolysaccharide

[0197] parameter CSPS-IR783 solution CSPS-IR783@OA NPs <![CDATA[T （1 / 2） a (h)]]> 0.239±0.01 6.104±0.03 <![CDATA[T （1 / 2） β (h)]]> 3.203±0.02 69.315±0.11 <![CDATA[AUC (0→t) (mg h L -1 )]]> 255.197±0.14 553.119±1.35**** <![CDATA[AUC (0→∞) (mg h L -1 )]]> 255.319±0.25 602.181±2.17**** <![CDATA[CL (L h -1 kg -1 )]]> 0.071±0.281 0.033±0.296 <![CDATA[Vd (L kg -1 )]]> 0.506±0.52 0.655±0.416 MRT (0→∞) (h) 8.197±0.02 9.649±0.05 <![CDATA[Cmax (mg L -1 )]]> 48±2.16 57±1.88

[0198] Example 29 Congo Red Analysis of Carob Polysaccharide CSPS and Nanocarob Polysaccharide CSPS@OA NPS

[0199] Congo red is readily soluble in ethanol and water and is a powdered acidic dye. It is primarily used to detect whether polysaccharides possess a triple-helix structure. The main principle is that it can form complexes with polysaccharides possessing a triple-helix structure. Detection using ultraviolet spectroscopy reveals a relationship between the maximum absorption wavelength and the concentration of NaOH. Specifically, when the NaOH concentration exceeds 0.2 mol / L, the maximum absorption wavelength shows a significant decrease. Therefore, based on the change in the maximum absorption wavelength, we can determine whether a polysaccharide possesses a triple-helix structure.

[0200] Methods: Accurately weigh 10 mg of carob polysaccharide nanoparticles (CSPS@OA NPs) and dissolve them in 5 ml of ultrapure water to prepare a 2 mg / ml solution. Weigh an appropriate amount of Congo red and dissolve it in ultrapure water to prepare a 160 μg / ml Congo red solution. Prepare NaOH solutions with concentrations of 0.4, 0.8, 1.2, 1.6, and 2.0 mol / L for later use. Separately, take 10 test tubes and add carob polysaccharide nanoparticles (CSPS@OA NPS), Congo red solution, and NaOH solution to 5 test tubes in a 2:1:1 ratio, making the final polysaccharide concentration 1 mg / ml. Add ultrapure water, Congo red solution, and NaOH solution to 5 test tubes in the same 2:1:1 ratio. The final concentration of Congo red in each test tube was 40 μg / ml, and the final concentrations of NaOH were 0.1, 0.2, 0.3, 0.4, and 0.5 mol / L, respectively. The mixtures were thoroughly shaken and allowed to stand at room temperature for 10 min. Ultrapure water was used as a blank control. The maximum absorption wavelengths at different NaOH concentrations were recorded using a UV spectrophotometer at wavelengths of 400-600 nm.

[0201] A graph was plotted with NaOH concentration (mol / L) and maximum absorption wavelength (nm) on the x and y axes, respectively. The results are attached. Figure 9 As shown, the maximum absorption wavelength of the complex formed by carob polysaccharide and Congo red increases slightly in the range of 0-0.2 mol / L sodium hydroxide and decreases sharply in the range of 0.2-0.3 mol / L, proving that carob polysaccharide has a typical triple helix structure. After carob polysaccharide was prepared into nano-polysaccharide, the maximum absorption wavelength of the complex formed with Congo red was consistent with that of free carob polysaccharide, indicating that the preparation process of nano-polysaccharide did not affect the triple helix higher-order structure of carob polysaccharide at all.

[0202] Example 30: Comparison of Fourier Transform Infrared Spectra of Carob Polysaccharide CSPS and Nanocarob Polysaccharide CSPS@OA NPS

[0203] Accurately weigh 5.0 mg of CSPS and CSPS@OA NPs samples, add an appropriate amount of dry KBr, grind to prepare a uniform transparent pellet, and analyze using Fourier transform infrared spectroscopy in the range of 4000 cm⁻¹ to 400 cm⁻¹. The results are shown in the appendix. Figure 10 .

[0204] Fourier transform infrared spectroscopy (FT-IR) is a method for identifying or quantifying molecular structures, and is also a commonly used technique for analyzing the fine structures of macromolecules such as polysaccharides. (From the attached...) Figure 10 It can be seen that the FT-IR overlap between nanopolysaccharides and free polysaccharides is excellent, proving that the preparation process of nanopolysaccharides does not affect the fine structure of polysaccharides.

[0205] Example 31. Preparation of nano-polysaccharides by direct dispersion method using amphiphilic lipid materials as excipients.

[0206] Accurately weighed carob total polysaccharides were dissolved in water to prepare a 10 mg / mL polysaccharide solution. 6 mg each of TPGS, PLA-1000-PEG2000, PCL1000-PEG2000, and PLGA1000-PEG2000 were weighed and added to 5 mL of the polysaccharide solution respectively. The mixture was magnetically stirred for 5 minutes, then sonicated in a 250 W water bath for 10 minutes. The particle size and particle size distribution (PDI) were measured, and the results are shown in Table 32. The mixture was then homogenized at 1400 bar for 10 cycles, and the particle size and PDI were measured again, with the results also shown in Table 32. This indicates that for lipid materials containing PEG segments, these materials can also be dispersed in a polysaccharide solution and stirred and sonicated to obtain polysaccharide nanoassemblies. High-pressure homogenization can further reduce the particle size and particle size distribution of the obtained nanopolysaccharides.

[0207] Table 32 Particle size, PDI, and potential of nanopolysaccharides prepared by direct dispersion method using amphiphilic lipid materials as excipients.

[0208] mass ratio Particle size PDI Total polysaccharides from carob - TPGS 3:25 265.2±78.67 0.343±0.102 After high-pressure homogenization 10 times 131.5±4.67 0.142±0.004 <![CDATA[Carob total polysaccharide-(PLA 1000 -PEG 2000 )]]> 3:25 345.0±56.65 0.337±0.108 After high-pressure homogenization 10 times 163.2±5.13 0.172±0.005 <![CDATA[Carob Total Polysaccharide-(PCL 1000 -PEG 2000 )]]> 3:25 413.5±78.12 0.363±0.097 After high-pressure homogenization 10 times 238.3±5.03 0.197±0.012 <![CDATA[Carob total polysaccharide-(PLGA 1000 -PEG 2000 )]]> 3:25 441.5±95.23 0.377±0.113 After high-pressure homogenization 10 times 166.2±7.61 0.203±0.005

[0209] Example 32. Nanoparticles loaded with macromolecular drugs

[0210] Accurately weigh total carob polysaccharides and dissolve them in water to prepare a 10 mg / mL polysaccharide solution. Precisely prepare 1 mL solutions of 5 mg insulin, 5 mg bevacizumab, and 5 mg rituximab, and add each solution to 4 mL of the polysaccharide solution and mix well to obtain three 5 mL aliquots of the mixture. Separately, dissolve 3 mg oleic acid and 3 mg TGPS in 0.5 mL of ethanol. Slowly add the ethanol solution dropwise to the above mixtures while gently stirring at 100 rpm. Remove the ethanol by rotary evaporation under reduced pressure at 35 °C. Measure the particle size and particle size distribution. The results are shown in Table 33. It can be seen that nanopolysaccharides can successfully load molecular drugs such as peptides and proteins.

[0211] After being placed in a 35℃ refrigerator for 3 days, the particle size and particle size distribution of the nanopolysaccharide loaded with macromolecular drugs were re-measured. The results are shown in Table 32. It can be seen that the obtained nanopolysaccharide loaded with macromolecular drugs has good storage stability.

[0212] Table 33 Drug-loaded nanopolysaccharides obtained by loading peptides and proteins with nanopolysaccharides and their particle size and PDI after loading.

[0213] mass ratio Particle size (nm) PDI Carob total polysaccharides-insulin-oleic acid-TPGS 40:5:3:3 183.5±4.86 0.215±0.05 After being placed at 4℃ for 3 days 178.5±4.36 0.202±0.04 Carob total polysaccharides-bevacizumab-oleic acid-TPGS 40:5:3:3 221.6±5.33 0.183±0.04 After being placed at 4℃ for 3 days 235.2±4.76 0.191±0.04 Carob total polysaccharides - rituximab - oleic acid - TPGS 40:5:3:3 246.0±6.11 0.207±0.08 After being placed at 4℃ for 3 days 233.8±8.26 0.237±0.009

[0214] The above description of the disclosed embodiments enables those skilled in the art to make or use the invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the general principles defined herein may be implemented in other embodiments without departing from the spirit or scope of the invention. Therefore, the invention is not to be limited to the embodiments shown herein, but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.

Claims

1. A nanopolysaccharide, characterized in that, The nanopolysaccharide is composed of polysaccharide and lipid materials; The polysaccharides are selected from Astragalus polysaccharides, carob polysaccharides, plantain seed polysaccharides, wolfberry polysaccharides, soybean polysaccharides, angelica polysaccharides, aloe polysaccharides, yam polysaccharides, fritillaria polysaccharides, eucommia polysaccharides, polygonatum polysaccharides, ginseng polysaccharides, gardenia polysaccharides, lily polysaccharides, shiitake mushroom polysaccharides, turkey tail polysaccharides, polyporus umbellatus polysaccharides, and cordyceps polysaccharides. The lipid material is selected from C6-C50 medium-chain and long-chain fatty alcohols and their esters and ethers, C6-C50 medium-chain and long-chain fatty amines and their amides formed with acids, C6-C50 medium-chain and long-chain fatty acids and their esters, C6-C50 medium-chain and long-chain fatty acid amides, and amphiphilic materials containing C6-C50 medium-chain and long-chain fatty chains. Long-chain lipophilic materials with a chain length of <10000 and amphiphilic polymers thereof with lipophilic segments, wherein the lipid material has a saturated lipid chain or contains one or more unsaturated bonds. The weight ratio of the polysaccharide to the lipid material is 2-50:1; The nanopolysaccharide is prepared by dissolving lipid material in a water-miscible organic solvent, adding it to the polysaccharide solution under stirring or ultrasound to induce the self-assembly of the polysaccharide, and then removing the organic solvent. The lipid material and polysaccharide are then assembled into nanoparticles with a particle size of 20-1000 nm. Alternatively, the lipid material may be an amphiphilic lipid material, which is obtained by dispersing the amphiphilic lipid material in an aqueous solution of polysaccharide, stirring, and sonicating.

2. The nanopolysaccharide according to claim 1, characterized in that, The lipid material is selected from C8-C30 medium-chain and long-chain fatty alcohols and their esters and ethers, C8-C30 medium-chain and long-chain fatty amines and their amides formed with acids, C6-30 medium-chain and long-chain fatty acids and their esters, C8-C30 medium-chain and long-chain fatty acid amides, amphiphilic materials containing C8-C30 medium-chain and long-chain fatty chains, long-chain lipophilic materials with a molecular weight <5000, and amphiphilic polymers with lipophilic segments thereof.

3. The nanopolysaccharide according to claim 2, characterized in that, The lipid material is selected from C8-C18 medium-chain and long-chain fatty alcohols and their esters and ethers, C8-C18 medium-chain and long-chain fatty amines and their amides formed with acids, C8-C18 medium-chain and long-chain fatty acids and their esters, C8-C18 medium-chain and long-chain fatty acid amides, amphiphilic materials containing C8-C18 medium-chain and long-chain fatty chains, long-chain lipophilic materials with a molecular weight <3000, and amphiphilic polymers with lipophilic segments thereof.

4. The nanopolysaccharide according to claim 1, characterized in that, The nanopolysaccharide also includes pharmaceutically acceptable excipients, including binders, disintegrants, lubricants, and diluents.

5. A method for preparing nanopolysaccharides as described in any one of claims 1-4, characterized in that, The method specifically includes the following steps: 1) The lipid material is dissolved in ethanol, methanol, acetone, isopropanol or a mixture thereof to obtain a lipid material solution, and the polysaccharide is dissolved in water to obtain a polysaccharide solution. The lipid material solution is added to the polysaccharide solution under stirring or sonication, and the solvent is evaporated to obtain nano-polysaccharides with an average particle size of 20-1000 nm. Alternatively: dissolve the polysaccharide in water to obtain a polysaccharide solution, disperse the amphiphilic lipid material in the polysaccharide aqueous solution, and then stir and sonicate to obtain nano-polysaccharides; 2) If the obtained nanopolysaccharide has a large particle size or a wide particle size distribution, it can be further homogenized under high pressure to reduce the particle size; 3) The obtained nano-polysaccharides can be further spray-dried or freeze-dried to become solid powders to suit different applications.

6. The method for preparing nanopolysaccharides according to claim 5, characterized in that, The evaporation in step 1) is one or a combination of natural evaporation, heated evaporation, and reduced pressure evaporation, and the homogenization pressure in step 2) is 500-4000 bar.

7. The application of the nanopolysaccharide as described in any one of claims 1-4 in the preparation of food and pharmaceutical products.

8. The application according to claim 7, characterized in that, The aforementioned nanopolysaccharides can be further loaded with lipid-soluble small molecules or macromolecules such as proteins, peptides, and nucleic acids to become nanoparticles with health-care and therapeutic effects.

9. The application according to claim 7, characterized in that, The nanopolysaccharide solution and its dried powder can also be used as intermediates to further process into various forms of food or pharmaceutical products, including oral liquids, suspensions, gels, granules, tablets, capsules, inhalation preparations, spray preparations, injection solutions, lyophilized powders for injection, ointments, patches, and microneedles, for oral, injection, mucosal, cavity, and external use.

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