1,4-Polyisoprene dispersions and active pharmaceutical ingredients and their uses

Abstract

The present invention relates to 1,4-polyisoprene dispersions, pharmaceutical active ingredients, and uses thereof. The dispersions remain stable in the gastric acid environment of mammals, do not precipitate 1,4-polyisoprene aggregates, are non-oral toxic to mammals, and are allergen-free. The 1,4-polyisoprene dispersions include 1,4-polyisoprene solution dispersions and 1,4-polyisoprene emulsion dispersions. The 1,4-polyisoprene dispersions can be used as the pharmaceutical active ingredient in the manufacture of drugs for treating atherosclerotic cardiovascular disease, type 2 diabetes, hypercholesterolemia, hypertriglyceridemia, fatty liver, colitis, obesity, polycystic ovary syndrome, Alzheimer's disease, and other conditions.

Description

[Technical Field]

[0001] The present application belongs to the technical field of medicinal materials, and specifically relates to 1,4-polyisoprene dispersions and active pharmaceutical ingredients and their uses. [Background technology]

[0002] Chronic metabolic diseases are metabolic syndromes, which mainly include atherosclerotic heart and cerebrovascular diseases (coronary heart disease and stroke), type 2 diabetes, hypercholesterolemia, hypertriglyceridemia, fatty liver, chronic enteritis, polycystic ovary syndrome, and obesity. Of these, obesity, hyperlipidemia, fatty liver, polycystic ovary syndrome, and chronic enteritis do not have as high a mortality rate as heart and cerebrovascular diseases and diabetes, but they have a high incidence rate among adults and a large number of patients, which seriously impacts health.

[0003] As a conventional technology for treating chronic metabolic diseases, many drugs have been developed for various symptoms. For example, in the case of atherosclerotic heart disease, conventional drugs include statin drugs for lipid lowering and atherosclerotic plaque stabilization, PCSK9 inhibitors, antiplatelet drugs such as aspirin and clopidogrel, beta-receptor blockers, and RAS inhibitors.

[0004] In the prior art, research and development of drugs for various chronic metabolic diseases has also been carried out. CN101505594A discloses drugs that can be used to treat various metabolic disorders, such as insulin resistance syndrome, diabetes, polycystic ovary syndrome, hyperlipidemia, fatty liver disease, cachexia, obesity, atherosclerosis, and arteriosclerosis.

[0005] Nevertheless, chronic metabolic diseases remain a significant cause of mortality, prevalence, and impact, and we need to develop more and better drugs to address these diseases. Summary of the Invention [Problem to be solved by the invention]

[0006] This application provides a 1,4-polyisoprene dispersion and a pharmaceutical active ingredient, as well as uses thereof. Taking advantage of these properties, the 1,4-polyisoprene dispersion is stable in the gastric acid environment of mammals, does not precipitate 1,4-polyisoprene aggregates, is non-oral toxic to mammals, and is free of allergens, it is used to manufacture drugs for treating chronic metabolic diseases, including atherosclerotic cardiovascular disease, type 2 diabetes, hypercholesterolemia, hypertriglyceridemia, fatty liver, colitis, obesity, and polycystic ovary syndrome. [Means for solving the problem]

[0007] In a first aspect, the present application provides a 1,4-polyisoprene dispersion that remains stable in the acidic stomach environment of a mammal, does not precipitate 1,4-polyisoprene aggregates, and is non-oral toxic and allergen-free in mammals.

[0008] In the present application, 1,4-polyisoprene is present in a dispersed state in the 1,4-polyisoprene dispersion. The present application utilizes the 1,4-polyisoprene dispersion as a pharmaceutical active ingredient, which is stable in the gastric acid environment of mammals, does not precipitate 1,4-polyisoprene aggregates, is non-oral toxic to mammals, and is free of allergens, to manufacture drugs for treating chronic metabolic diseases, including atherosclerotic cardiovascular disease, type 2 diabetes, hypercholesterolemia, hypertriglyceridemia, fatty liver, colitis, obesity, and polycystic ovary syndrome.

[0009] The preferred technical solutions of the present application are disclosed below, which are not intended to limit the technical solutions provided by the present application. Through the following preferred technical solutions, the objectives of the present application can be better achieved and beneficial effects can be realized.

[0010] As a preferred technical solution of the present application, the 1,4-polyisoprene dispersion includes a 1,4-polyisoprene solution dispersion and a 1,4-polyisoprene emulsion dispersion; The components of the 1,4-polyisoprene solution dispersion include 1,4-polyisoprene and a solvent, The components of the 1,4-polyisoprene emulsion dispersion include 1,4-polyisoprene, an emulsifier, and water.

[0011] In the preferred technical solution of the present application, the polymerization degree of the 1,4-polyisoprene is 6 or more, and there is no upper limit, and it may be, for example, 6, 10, 20, 50, 100, 200, 500, 1000, 2000, 5000, 8000, 10000, or 100000.

[0012] 1,4-Polyisoprene with a degree of polymerization of less than 6 has a certain degree of volatility, a pungent odor, and has an inhibitory effect on the growth of in vitro cultured cells, so 1,4-polyisoprene with a degree of polymerization of 6 or more is selected to prepare the 1,4-polyisoprene dispersion.

[0013] In this application, there are no particular restrictions on the cis / trans structure or origin of 1,4-polyisoprene; it can be produced from natural latex or obtained by artificial synthesis. Artificially synthesized 1,4-polyisoprene contains both cis and trans structures, with the cis structure predominating. Most natural rubber derived from plants such as Hevea brasiliensis or fungi has a cis structure. Natural rubber produced from Eucommia has a trans structure, i.e., it has been found to be trans 1,4-polyisoprene.

[0014] In a preferred technical solution of the present application, the solvent is selected from any one or a combination of at least two of fatty acids, fatty alcohols, ester compounds, and lipid compounds.

[0015] Preferably, the ester-based compound includes any one of a monohydric alcohol ester, a dihydric alcohol ester, and a polyhydric alcohol ester, or a combination of at least two of them.

[0016] In the present application, the monohydric alcohol ester is obtained by reacting a fatty acid with a monohydric alcohol, the dihydric alcohol ester is obtained by reacting a fatty acid with a dihydric alcohol, and the polyhydric alcohol ester is obtained by reacting a fatty acid with a polyhydric alcohol.

[0017] Preferably, the monohydric alcohol ester comprises a fatty acid ethyl ester.

[0018] Preferably, the fatty acid ethyl ester comprises any one or a combination of at least two of ethyl laurate, ethyl palmitate, ethyl stearate, ethyl oleate, ethyl linoleate, ethyl linolenate, ethyl eicosapentaenoate (EPA), or ethyl docosahexaenoate (DHA).

[0019] Preferably, the polyhydric alcohol ester comprises a glycerin ester.

[0020] Preferably, the glycerol ester comprises any one or a combination of at least two of a monoglyceride, a diglyceride or a triglyceride.

[0021] Preferably, the monoglycerides and diglycerides are obtained by reacting lauric acid, palmitic acid, palmitic acid, oleic acid, linoleic acid, linolenic acid with glycerol.

[0022] Preferably, the triglyceride is an artificially synthesized C6-C 12 Contains medium chain fatty acid triglycerides and / or coconut oil, a natural triglyceride.

[0023] Preferably, the lipid compound comprises any one or a combination of at least two of a phospholipid, a glycolipid, a sphingolipid, a steroid, or squalene.

[0024] Preferably, the lipid compound comprises squalene.

[0025] The reason for selecting the above solvent for the 1,4-polyisoprene solution dispersion system of the present invention is mainly that the solvent is highly safe, has good solubility for 1,4-polyisoprene, and has low viscosity.

[0026] Among monohydric alcohol esters, ethanol esters are preferable because they are safer than methanol esters. Among ethanol esters, fatty acids with 12 or more carbon atoms are selected because the solubility of 1,4-polyisoprene decreases when the carbon chain of the fatty acid has fewer than 12 carbon atoms. For fatty acids with 18 or more carbon atoms, the higher the degree of unsaturation, the better the fluidity at room temperature. Therefore, the monohydric alcohol ester is preferably any one or a combination of at least two of ethyl laurate, ethyl palmitate, ethyl stearate, ethyl oleate, ethyl linoleate, ethyl linolenate, ethyl eicosapentaenoate (EPA), and ethyl docosahexaenoate (DHA).

[0027] Among polyhydric alcohol esters, glycerol esters are safe, and glycerin esters include monoglycerides, diglycerides, and triglycerides. 12 ~C 18Monoglycerides and diglycerides produced from fatty acids and glycerol have small molecular weights, low viscosities, and good solubility of 1,4-polyisoprene. Therefore, monoglycerides or diglycerides obtained by reacting glycerol with any of lauric acid, palmitic acid, palmitic acid, oleic acid, linoleic acid, and linolenic acid are preferred. Triglycerides have three ester bonds and contain three fatty acids. When the fatty acid is a long-chain fatty acid with more than 12 carbon atoms, the triglycerides generally have large molecular weights, high viscosities, and poor solubility of 1,4-polyisoprene. Conversely, when the fatty acid is a short-chain fatty acid with less than 6 carbon atoms, the triglycerides generally have strong non-polar molecules and poor solubility of 1,4-polyisoprene. Therefore, medium-chain fatty acid triglycerides having 6 to 12 carbon atoms (for example, 6, 8, 10, or 12) have just the right molecular weight, just the right viscosity, and good solubility for 1,4-polyisoprene. Medium-chain fatty acid triglycerides having 6 to 12 carbon atoms include artificially synthesized medium-chain fatty acid glycerol esters and coconut oil, which is a natural fat and oil.

[0028] Preferably, the lipid compounds include phospholipids, glycolipids, sphingolipids, steroids, squalene, etc., which are mostly solid at room temperature and have poor solubility in 1,4-polyisoprene. Therefore, it is preferable to use squalene as the solvent, which is liquid at room temperature and is a non-polar molecule like 1,4-polyisoprene.

[0029] As a preferred technical solution of the present application, the mass percent content of 1,4-polyisoprene in the 1,4-polyisoprene solution dispersion is 60% or less and not 0 (for example, it may be 0.05%, 0.1%, 0.5%, 1%, 2%, 5%, 7%, 10%, 12%, 15%, 18%, 20%, 30%, 40%, 50%, or 60%, etc.), and is preferably 0.2% to 20%.

[0030] In 1,4-polyisoprene solution dispersions, as the mass percent content of 1,4-polyisoprene increases, the solution becomes increasingly viscous, making it difficult to manufacture and use. Selecting a solvent with a low molecular weight and high fluidity can reduce the solution viscosity to some extent. While it is difficult to manufacture 1,4-polyisoprene solution dispersions with a content of 60% or more using conventional techniques, research has shown that a content of 20% or less provides good manufacturing efficiency and fluidity. Therefore, the upper limit is set to 60%, preferably 20%. While there is no lower limit for the 1,4-polyisoprene content, as the 1,4-polyisoprene content decreases, a larger amount of solution must be applied to mammals or cells to achieve therapeutic drug activity, making it less convenient to use.

[0031] In the present application, there are no particular limitations on the method for producing a 1,4-polyisoprene solution dispersion, as long as a system in which 1,4-polyisoprene molecules are completely dispersed in a solvent can be obtained. Examples include, but are not limited to, the following: Lumped 1,4-polyisoprene extracted from Para rubber trees is frozen in liquid nitrogen to obtain a brittle solid, which is then crushed in a high-speed crusher and the crushed particles are sieved through a 20-mesh sieve. The crushed particles are then added to a solvent and constantly stirred until the 1,4-polyisoprene is completely dissolved in the solvent, yielding the 1,4-polyisoprene solution dispersion.

[0032] The 1,4-polyisoprene is crushed in a high-speed crusher and the crushed particles are passed through a 20-mesh sieve in order to accelerate the dissolution of the 1,4-polyisoprene.

[0033] In a preferred technical solution of the present application, the emulsifier is selected from any one or a combination of at least two of cationic surfactants, anionic surfactants, zwitterionic surfactants, and nonionic surfactants.

[0034] Preferably, the nonionic surfactant is polyoxyethylene (20) sorbitan monolaurate (polysorbate 20), polyoxyethylene (20) sorbitan monopalmitate (polysorbate 40), polyoxyethylene (20) sorbitan monostearate (polysorbate 60), polyoxyethylene (20) sorbitan monooleate (polysorbate 80), decaglycerol monolaurate (Q-12S), decaglycerol monomyristate (Q-14S), decaglycerol monopalmitate (Q-16S), decaglycerol monooleate (Q-17S), decaglycerol monostearate (Q-18S) or C 12 ~C 18 The present invention includes any one or a combination of at least two of the following mono-fatty acid sucrose esters:

[0035] In the prior art, as many as a million types of surfactants are known, with a vast variety of types and varying properties. In this application, the preferred surfactants are selected from common low- or non-toxic surfactants, and are those types that have been experimentally proven to be capable of maintaining the stability of 1,4-polyisoprene emulsion dispersion systems in a strongly acidic environment of pH = 1. There may be other available emulsifiers (including combinations of two or more emulsifiers) that have not yet been tested or screened, and these can be discovered and screened by simply repeating simple tests. Emulsion dispersion systems composed of these potential emulsifiers and 1,4-polyisoprene are also within the scope of protection of this patent.

[0036] As a preferred technical solution of the present application, the mass percentage content of 1,4-polyisoprene in the 1,4-polyisoprene emulsion system is 80% or less and is not 0 (for example, it may be 0.05%, 0.1%, 0.5%, 1%, 2%, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 60%, 70%, or 80%), and more preferably 0.5% to 70%.

[0037] In 1,4-polyisoprene emulsion dispersion systems, the higher the mass percent content of 1,4-polyisoprene, the worse the emulsion's fluidity and stability. A mass percent content of 80% 1,4-polyisoprene is the maximum stable emulsion content that can be produced by the present invention under conventional technology, but the fluidity is poor. When the content is reduced to 70%, both the stability and fluidity of the emulsion become good. Therefore, the upper limit of the 1,4-polyisoprene content in the emulsion is set to 80%, preferably 70%. While there is no lower limit for the 1,4-polyisoprene content, as the 1,4-polyisoprene content decreases, a larger amount of emulsion must be used in mammals or cells to achieve therapeutic drug activity, reducing ease of use. Research has shown that a content of 0.5% is the lower limit for acceptable ease of use, so the lower mass percent content of 1,4-polyisoprene is preferably 0.5%.

[0038] In a preferred technical solution of the present application, the average particle size of the emulsion droplets in the 1,4-polyisoprene emulsion system is 10 μm or less, and may be, for example, 0.1 μm, 0.5 μm, 1 μm, 2 μm, 3 μm, 4 μm, 5 μm, 6 μm, 7 μm, 8 μm, 9 μm, or 10 μm.

[0039] The smaller the particle size of 1,4-polyisoprene emulsion dispersion, the better the dispersion effect. Research has shown that when the average particle size of the emulsion droplets in 1,4-polyisoprene emulsion is 10 μm or less, obvious medicinal effects can be achieved.

[0040] In addition, the 1,4-polyisoprene emulsion dispersion system that can be used as a pharmaceutical active ingredient in the present application is different from natural latex extracted from Para rubber trees in conventional technology, industrial concentrated latex produced from natural latex as a raw material, and artificially synthesized 1,4-polyisoprene latex, and these substances have significant differences in components and properties, so they cannot be mixed or used interchangeably.

[0041] The components and characteristics of the 1,4-polyisoprene emulsion dispersion of the present application are as follows: 1) The ingredients are few and well-defined, consisting of 1,4-polyisoprene, water, and an emulsifier. 2) It can maintain stability in a strongly acidic environment of pH=1, and furthermore, it remains dispersed in the acidic environment of a mammal's stomach, ensuring that 1,4-polyisoprene aggregates do not precipitate. 3) It does not contain any allergens and does not cause allergies in humans or other mammals. 4) All ingredients are safe for mammals and have no oral toxicity.

[0042] The components and characteristics of natural latex are as follows: 1) Its composition is complex and unclear. In addition to 1,4-polyisoprene and water, it contains five other major substances: proteins, lipids, water-soluble substances, acetone-soluble substances, and inorganic salts. The protein content accounts for 1% to 2% of the raw latex mass. Of this, approximately one-fifth of the protein is distributed on the surface of the rubber particles, forming a protective layer. Another three-fifth is dispersed in the whey, and the remainder is present in the lower layer of the latex. In addition to major proteins such as rubber protein and α-globulin, the protein component also contains large amounts of enzymes, including coagulation enzymes, oxidases, peroxidases, reductases, proteases, and phospholipases. The lipid content is approximately 1%, consisting of complex compounds such as triglycerides, fatty acids, waxes, sterols, sterol esters, and phospholipids. The acetone-soluble substance content is approximately 1%, consisting mainly of oleic acid, linoleic acid, palmitic acid, sterols, and sterol esters. The content of water-soluble substances is 1% to 2%, and its main components are querachitol, inositol isomers, sucrose, glucose, galactose, fructose, and pentose. The inorganic salts are mainly potassium, calcium, magnesium, copper, iron, phosphate ions, sodium, and small amounts of sulfate ions, chloride ions, aluminum, manganese, nickel, rubidium, etc., accounting for 0.3% to 0.7% of the latex mass.

[0043] 2) It is alkaline and stable in an alkaline environment, but when it comes into contact with a weak acid, 1,4-polyisoprene precipitates and solidifies.

[0044] 3) The amino acid sequence is highly homologous to the agglutinin (UDA) derived from the common stinging nettle (Urtica dioica), has strong sensitizing properties, and contains the rubber protein hev1, which is also the main allergen in rubber products.

[0045] Natural rubber latex has a significant problem of coagulating when exposed to acid, in addition to obvious sensitization, and therefore cannot be used as an active ingredient in pharmaceuticals.

[0046] The ingredients and characteristics of concentrated natural latex and synthetic latex are as follows: 1) The ingredients are complex and unclear. In addition to the complex ingredients of natural latex, concentrated latex further contains various industrial chemical stabilizers, such as ammonia, TMTD (tetramethylthiuram disulfide) / ZnO, and in some cases, formaldehyde, sodium sulfite, hydroxylamine, hydrogen peroxide, sodium carbonate, urea, organic amines (e.g., methylamine), and sodium pentachlorophenol. Some low-ammonia or low-protein concentrated latexes further contain 2,2'-thiobis(4,6-dichlorophenol), 1,4-dioxanone and its derivatives, Stmktol LB219, dihydroxypropyl dodecanoate (BeKa100), sodium polyethylene glycol monocetyl ether sulfate (trade name Levenol WX), ethoxylated fatty alcohols, ethoxylated alkylphenols, ethoxylated long-chain alkylamines, N-(2-hydroxy)propyl-3-trimethylamine chloride chitosan, triazine derivatives, and the like. Artificial synthetic latex is polyisoprene latex obtained by polymerizing industrial chemical polyisoprene as a monomer with the action of a catalyst and initiator. Not only do various stabilizers contain added catalysts (neodymium (III) oxide, tri-n-butyl phosphate, methylaluminoxane, etc.), solvents (e.g., toluene, n-hexane, etc., which generally have a certain degree of toxicity), residual monomers and oligomers (polyisoprene with a very low degree of polymerization), and about 5% to 10% non-1,4-polyisoprene products (non-target products), the latex also contains.

[0047] 2) In an acidic environment, 1,4-polyisoprene quickly precipitates and coagulates. Concentrated natural latex is a product of concentrating natural latex through centrifugation. It inherits the natural latex characteristic of coagulating when exposed to acid, and this characteristic is reinforced as an essential attribute in the subsequent production and processing of latex products. Concentrated latex undergoes a process of coagulation from a fluid latex state to form a shaped form, whether it is used to produce membrane products such as latex gloves, condoms, and balloons, or sponge products such as latex mattresses and pillow inserts. This process requires the addition of a coagulant. Coagulants are primarily organic acids, and in some cases, divalent metal salts are also added. For example, when producing medical latex products, coagulants used to concentrate latex include organic acids such as formic acid, acetic acid, lactic acid, cyclohexylamine acetate, and boric acid. In the Dunlop process for producing latex mattresses, the coagulant used to concentrate latex is hydrofluoric acid, produced by hydrolysis of sodium silicofluoride, while in the Talalay process, the coagulant used to concentrate latex is carbon dioxide, which is added after vacuum foaming and freezing, using the acidity of the carbon dioxide to gradually coagulate the concentrated latex and shape it. Artificial synthetic latex is used in place of natural concentrated latex to produce various latex products, and in order to be compatible with the production process, it must have the same ability to coagulate when exposed to acid as natural latex.

[0048] 3) Natural concentrated latex still contains the allergen, which is the rubber protein hev1.

[0049] 4) Both concentrated natural latex and synthetic latex contain large amounts of substances that are orally toxic to mammals. Whether it is concentrated natural latex or synthetic latex, various stabilizers that are orally toxic must be added. Synthetic latex also contains catalysts, solvents, raw material monomers, raw material oligomers, N-nitrosamines and their precursors (potent carcinogens), making it more orally toxic.

[0050] Therefore, natural concentrated latex and synthetic latex contain large amounts of substances that are orally toxic, and their tendency to coagulate when exposed to acid remains a major problem.The 1,4-polyisoprene emulsion dispersion system provided by the present application remains dispersed in mammalian gastric juice and therefore exerts pharmacological activity.

[0051] In the present application, there is no particular limitation on the method for producing the 1,4-polyisoprene emulsion dispersion system, and examples of the method for producing the same include, but are not limited to, the following. Natural rubber emulsion collected from Para rubber trees is centrifuged at high speed at room temperature to obtain an upper layer of emulsion (removing substances other than rubber particles in the natural rubber emulsion, including some protein allergen components), an emulsifier is added, and then water is added and stirred until uniform. After that, alkaline protease isoenzyme is added (degrading the protein allergen components remaining in the rubber particles to purify 1,4-polyisoprene), the mixture is incubated, and then centrifuged again at high speed to obtain an upper layer of emulsion (1,4-polyisoprene coated with the emulsifier), which is then diluted with water to a specific mass concentration to obtain the 1,4-polyisoprene emulsion dispersion.

[0052] In a second aspect, the present application provides a pharmaceutical active ingredient comprising the 1,4-polyisoprene dispersion according to the first aspect.

[0053] In a third aspect, the present application provides a medicament comprising an active pharmaceutical ingredient according to the second aspect.

[0054] Preferably, the drug comprises a drug for treating a chronic metabolic disease; Preferably, the chronic metabolic diseases include atherosclerotic cardiovascular and cerebrovascular diseases, type 2 diabetes, hypercholesterolemia, hypertriglyceridemia, fatty liver, colitis, polycystic ovary syndrome and obesity.

[0055] Chronic metabolic diseases caused by long-term nutritional and energy surplus, such as atherosclerotic cardiovascular disease, type 2 diabetes, hypercholesterolemia, hypertriglyceridemia, obesity, polycystic ovary syndrome, and chronic enteritis, are all chronic metabolic diseases. Related research has revealed that nutrient surplus induces cellular metabolic reprogramming, and these metabolic products further promote the conversion of tissue-resident macrophages to the pro-inflammatory M1 type, which in turn recruits and concentrates more inflammatory cells in the tissue, ultimately leading to the development of chronic inflammatory lesions in tissues and organs and the development and progression of disease. Therefore, developing drugs that target macrophage polarization, inhibit macrophage foam cell transformation, inhibit their M1 polarization, and promote their conversion to the M2 type, is an important direction for the development of therapeutic drugs for chronic metabolic diseases.

[0056] The active pharmaceutical ingredient provided by the present application can effectively inhibit foam cell transformation of macrophages, inhibit their conversion to M1 type, and promote their conversion to M2 type. The active pharmaceutical ingredient provided by the present application can prevent and treat atherosclerotic cardiovascular disease, type 2 diabetes, hypercholesterolemia, hypertriglyceridemia, fatty liver, colitis, polycystic ovary syndrome, and obesity.

[0057] In a fourth aspect, the present application provides the use of a medicament in the treatment of Alzheimer's disease, the medicament comprising an active pharmaceutical ingredient according to the second aspect, the active pharmaceutical ingredient comprising the 1,4-polyisoprene dispersion according to the first aspect.

[0058] L-glutamate (L-glutamate), also known as glutamate, is an endogenous neurotransmitter abundant in the mammalian brain and spinal cord. It plays an important role in neuronal development, differentiation, and migration, and is closely related to synaptic initiation, development, plasticity, and learning and memory processes. Under normal physiological conditions, glutamate binds to glutamate receptors (GluRs) on the postsynaptic membrane, maintaining normal neuronal activity. Some pathological stimuli can cause excessively high extracellular glutamate concentrations, resulting in excessive calcium ion influx, which causes persistent neuronal depolarization and excessive activation of GluRs on the postsynaptic membrane. This can further induce adverse events, such as excitotoxicity, cell apoptosis, and oxidative stress, leading to neuronal degeneration and eventual death. Recent research has demonstrated that excessive levels of extracellular glutamate are closely related to many neuropsychiatric disorders, including Alzheimer's disease (AD).

[0059] A drug containing the 1,4-polyisoprene dispersion system provided by the present application as an active ingredient has a relieving effect on L-glutamic acid (L-GLU)-induced neuronal apoptosis and can be used as a drug for treating Alzheimer's disease. [Effects of the Invention]

[0060] Compared with the prior art, the present invention has the following beneficial effects: (1) The 1,4-polyisoprene dispersion system provided by the present application can be used as an active pharmaceutical ingredient, and is stable in the gastric acid environment of mammals, does not precipitate 1,4-polyisoprene aggregates, is non-toxic to mammals when administered orally, and does not contain any allergens. (2) The 1,4-polyisoprene dispersion provided herein can be used as an active pharmaceutical ingredient in the manufacture of drugs for treating chronic metabolic diseases, including atherosclerotic cardiovascular disease, type 2 diabetes, hypercholesterolemia, hypertriglyceridemia, fatty liver, colitis, polycystic ovary syndrome, and obesity. The 1,4-polyisoprene dispersion significantly inhibits foam cell transformation of macrophages, inhibits macrophage polarization into M1 type, promotes their polarization into M2 type, and alleviates inflammatory responses. This has significant effects on the prevention and treatment of atherosclerotic cardiovascular disease, as well as on type 2 diabetes, hypercholesterolemia, hypertriglyceridemia, fatty liver, colitis, obesity, and polycystic ovary syndrome. (3) The polyisoprene solution dispersion and polyisoprene emulsion dispersion provided herein have a rescue effect on L-glutamic acid (L-GLU)-induced neuronal apoptosis, and the rescue effect is dose-dependent, with significant rescue effects at polyisoprene mass fractions of 20 ppm and 40 ppm (P<0.05). The polyisoprene solution dispersion and polyisoprene emulsion dispersion have a significant inhibitory effect on neuronal apoptosis (P<0.05). Furthermore, the polyisoprene solution dispersion and polyisoprene emulsion dispersion can effectively improve the cognitive function of AD model mice, with mice treated with the polyisoprene dispersion showing a significant shortening of escape latency (P<0.05) and a significant increase in the number of crossings to the target platform and the number of crossings to the target quadrant (P<0.05). Therefore, the polyisoprene solution dispersion and polyisoprene emulsion dispersion can be used as an active ingredient in the preparation of drugs for the treatment of Alzheimer's disease. [Brief explanation of the drawings]

[0061] [Figure 1] 1 is an optical microscope photograph of the 1,4-polyisoprene emulsion dispersion provided in Example 2. [Figure 2] 1 shows ordinary light microscope photographs of foam macrophages in each of the seven experimental groups provided in Use Example 1. [Figure 3]1 is a bar graph showing the test results of foam macrophage content in total cells of the seven experimental groups provided in Use Example 1. [Figure 4] FIG. 10 is a graph showing the test results of the expression levels of macrophage M1 type marker gene IL-1β after treatment with LPS in the seven experimental groups provided in Use Example 2. [Figure 5] FIG. 10 is a graph showing the test results of the expression levels of the macrophage M1 marker gene iNOS after treatment with LPS in the seven experimental groups provided in Use Example 2. [Figure 6] FIG. 10 is a graph showing the test results of the expression levels of macrophage M2 type marker gene IL-10 mRNA after treatment with LPS in the seven experimental groups provided in Use Example 2. [Figure 7] FIG. 10 is a graph showing the test results of the expression levels of the macrophage M2 marker gene CD206 after treatment with LPS in the seven experimental groups provided in Use Example 2. [Figure 8] This is a graph showing the results of detecting the size of aortic plaques in mice in the four experimental groups provided in Example 3. [Figure 9] 1 is a bar graph showing the area of ​​aortic plaque in mice in the four experimental groups provided in Use Example 3 as a percentage of the area of ​​the mouse aorta. [Figure 10] This is a graph showing the results of detecting the size of aortic root plaque in mice in the four experimental groups provided in Use Example 3. [Figure 11] 1 is a bar graph of aortic root plaque area in mice from the four experimental groups provided in Use Example 3. [Figure 12] This is a graph showing the results of detecting the size of aortic plaques in mice in the four experimental groups provided in Use Example 4. [Figure 13] 1 is a bar graph showing the area of ​​aortic plaque in mice in the four experimental groups provided in Use Example 4 as a percentage of the area of ​​the mouse aorta. [Figure 14] This is a graph showing the results of detecting the size of aortic root plaque in mice in the four experimental groups provided in Use Example 4. [Figure 15]1 is a bar graph of aortic root plaque area in mice from the four experimental groups provided in Use Example 4. [Figure 16] This is a graph showing the test results of blood glucose content of mice in the four experimental groups provided in Use Example 5. [Figure 17] This is a graph showing the test results of serum glucose content in mice from the four experimental groups provided in Use Example 5. [Figure 18] This is a graph showing the test results of the glycated serum protein content of mice in the four experimental groups provided in Use Example 5. [Figure 19] This is a test result diagram of high density lipoprotein cholesterol content in mice of the four experimental groups provided in Use Example 6. [Figure 20] This is a test result diagram of low-density lipoprotein cholesterol content in mice of the four experimental groups provided in Use Example 6. [Figure 21] This is a test result diagram of the total cholesterol content of mice in the four experimental groups provided in Use Example 6. [Figure 22] FIG. 10 is a graph showing the test results of triglyceride content in mice of the four experimental groups provided in Use Example 6. [Figure 23] FIG. 10 is a graph showing the test results of lipoprotein lipase content in mice of the four experimental groups provided in Use Example 6. [Figure 24] This is a graph showing the test results of hepatic lipase content in mice from the four experimental groups provided in Use Example 6. [Figure 25] FIG. 10 is a graph showing the test results of the adipose triglyceride hydrolase content of mice in the four experimental groups provided in Use Example 6. [Figure 26] FIG. 10 is a graph showing the test results of hormone-sensitive triglyceride lipase content in mice of the four experimental groups provided in Use Example 6. [Figure 27] 10 shows liver photographs of mice from the four experimental groups provided in Use Example 7. [Figure 28] 10 is a test result of liver mass as a percentage of total mouse mass for the four experimental groups provided in Use Example 7. [Figure 29] 10 shows micrographs of liver sections from mice in the four experimental groups provided in Use Example 7 after H&E staining. [Figure 30] 10 shows micrographs of liver sections from mice in the four experimental groups provided in Use Example 7 after H&E staining. [Figure 31] This is a graph showing the test results of triglyceride content in liver tissue of mice from the four experimental groups provided in Use Example 7. [Figure 32] 10 shows the test results of the DAI scores of the four experimental groups provided in Example 8. [Figure 33] This is a graph showing the test results of the weight loss rates of mice in the four experimental groups provided in Example 8. [Figure 34] 10 is an optical photograph of the mouse colon after euthanasia of mice from the four experimental groups provided in Use Example 8. [Figure 35] This is a graph showing the test results of the length of the mouse colon after euthanasia of mice from the four experimental groups provided in Example 8. [Figure 36] This is a graph showing the test results of the weight of the mouse colon after euthanasia of mice from the four experimental groups provided in Example 8. [Figure 37] This is a graph showing the test results of the body weight of rats in the four experimental groups provided in Use Example 9. [Figure 38] This is a test result diagram of serum testosterone content in rats from the four experimental groups provided in Use Example 9. [Figure 39] This is a test result diagram of fasting blood glucose content of rats in the four experimental groups provided in Use Example 9. [Figure 40] This is a test result diagram of blood glucose content at different times for rats in four experimental groups provided in Use Example 9. [Figure 41] FIG. 10 is a graph showing test results of the area under the glucose tolerance curve of rats in the four experimental groups provided in Use Example 9. [Figure 42] This is a test result diagram of anti-Mullerian hormone AMH / GAPDH (mRNA) levels in the blood of rats in the four experimental groups provided in Use Example 9. [Figure 43] This is a graph showing the test results of IL-1β expression levels in rats in the four experimental groups provided in Use Example 9. [Figure 44] This is a graph showing the test results of TNF-α expression levels in rats in the four experimental groups provided in Use Example 9. [Figure 45] 1 shows the test results of the body weight of mice in the four experimental groups provided in Use Example 10. [Figure 46] 1 shows the fat mass test results of mice in the four experimental groups provided in Use Example 10. [Figure 47] This is a graph showing the test results of adipose tissue morphology in mice from the four experimental groups provided in Example 10. [Figure 48] 1 shows the test results of the average adipocyte area of ​​mice in the four experimental groups provided in Use Example 10. [Figure 49] 10 is a photograph of the stomach contents of mice in the natural latex group provided in Example 11. [Figure 50] 10 is a photograph of the stomach contents of mice in the rubber solution group provided in Example 11. [Figure 51] 10 is a photograph of the stomach contents of mice in the rubber emulsion group provided in Example 11. [Figure 52] 1 is a photograph of the state of 15 groups of 1,4-polyisoprene emulsion dispersion systems under acidic conditions provided in Use Example 12. [Figure 53] In Example 13, these are test results of the rescue effect of 12 experimental groups on L-glutamic acid-induced neuronal apoptosis in a cell model in which neuronal apoptosis is induced using L-glutamic acid. [Figure 54] 1 shows the results of detecting cell apoptosis using a flow cytometer for each of the four experimental groups in Use Example 14. [Figure 55] FIG. 11 is a statistical diagram showing the proportion of late apoptotic cells in the four experimental groups according to Use Example 14. [Figure 56]1 shows the movement trajectories of mice in a mouse maze model test for four experimental groups according to Use Example 15. [Figure 57] FIG. 10 is a statistical diagram of the escape latency of mice in four experimental groups according to Use Example 15. [Figure 58] FIG. 11 is a statistical diagram of the number of times mice in the four experimental groups crossed the target platform according to Use Example 15. [Figure 59] FIG. 10 is a statistical diagram of the number of times mice in the four experimental groups passed through the target quadrant according to Use Example 15. DETAILED DESCRIPTION OF THE INVENTION

[0062] The technical solutions of the present application will be further described below with reference to the drawings and specific embodiments. Those skilled in the art will appreciate that the above examples are only intended to facilitate understanding of the present application and should not be considered as specific limitations on the present application.

[0063] The origins of some of the components in the following Examples and Comparative Examples are as follows: Bulk cis-1,4-polyisoprene (abbreviated as bulk 1,4-polyisoprene) is obtained by adding 0.1 mol / L hydrochloric acid dropwise to fresh natural rubber emulsion produced from Para rubber trees to obtain coagulated natural rubber blocks, which are then rinsed with water, squeezed to remove moisture, and dried in the sun to obtain bulk 1,4-polyisoprene.

[0064] Bulk trans-1,4-polyisoprene can be obtained by collecting dried eucommia bark and leaves, mechanically crushing them, immersing them in n-hexane, concentrating the extract by rotary evaporation, adding excess 75% ethanol to precipitate a gel-like substance, dissolving the gel-like substance again in n-hexane, adding 75% ethanol again to precipitate, and allowing the solvent to escape.

[0065] Example 1 In this example, a cis-1,4-polyisoprene solution dispersion (hereinafter referred to as rubber solution) having a mass percent content of 2% is provided, and the manufacturing method of the 1,4-polyisoprene solution dispersion is as follows. The bulk 1,4-polyisoprene was immersed in liquid nitrogen to freeze it, and then pulverized in a high-speed pulverizer. The pulverized particles were passed through a 20-mesh sieve, and 2.0 g of the pulverized particles were weighed out. 98.0 g of ethyl oleate was added and the mixture was stirred constantly at 40°C until the 1,4-polyisoprene particles were completely dissolved, yielding the 1,4-polyisoprene solution dispersion, i.e., the rubber solution.

[0066] Example 2 In this example, a 1,4-polyisoprene emulsion dispersion (hereinafter referred to as rubber emulsion) having a mass percent content of 10% is provided, and the preparation method of the 1,4-polyisoprene emulsion dispersion is as follows. Fresh natural rubber emulsion extracted from Hevea brasiliensis is centrifuged at room temperature at high speed to obtain an upper layer of emulsion, to which 1% by weight of the emulsifier polyoxyethylene (20) sorbitan monooleate (purchased from Merck) is added, followed by addition of water and stirring until homogeneous. Alkaline protease isoenzyme is added (to decompose the protein emulsifier in natural latex that can cause allergies in humans), and the mixture is incubated at 37°C for 4 hours. After that, the mixture is centrifuged again at high speed to obtain an upper layer of emulsion, which is diluted with water until the mass percent content of 1,4-polyisoprene is 10% and the mass percent content of the emulsifier polyoxyethylene (20) sorbitan monooleate is 0.2%, thereby obtaining the 1,4-polyisoprene emulsion dispersion system, i.e., rubber emulsion.

[0067] The 1,4-polyisoprene emulsion dispersion provided by this example was characterized using an optical microscope, and the test results are shown in Figure 1. From Figure 1, it was found that the average particle size of the 1,4-polyisoprene emulsion dispersion provided by this example was 10 μm or less.

[0068] Example 3 This example provides a trans-1,4-polyisoprene solution dispersion (hereinafter referred to as "trans-rubber solution") with a mass percent content of 2%, and its preparation method is the same as that of Example 1. Except for replacing bulk 1,4-polyisoprene with bulk trans-1,4-polyisoprene, the other conditions are the same as those of Example 1.

[0069] (Comparative Example 1) In this comparative example, rat feed containing 1,4-polyisoprene particles (hereinafter referred to as rubber particles) is provided, and the method for producing the rat feed containing 1,4-polyisoprene particles is as follows. The bulk 1,4-polyisoprene was frozen in liquid nitrogen, crushed in a high-speed grinder, and passed through a 20-mesh sieve. The crushed particles were then added to powdered rat chow (purchased from Jiangsu Collaborative Medical Biotechnology Co., Ltd.) at a ratio of 0.5‰ and mixed until uniform. At this ratio, the amount of 1,4-polyisoprene ingested by the mice via chow per day was equivalent to the amount of 1,4-polyisoprene ingested by the mice via intragastric administration in the experimental group. The mixed chow was then granulated in a feed granulator to produce mouse chow containing rubber particles, referred to as "rubber particles."

[0070] (Example 1: Inhibitory effect on foam cell transformation of macrophages) Foam cell formation in the human macrophage cell line RAW264.7 was induced using oxidized low-density lipoprotein (oxLDL). Macrophages were then treated with a rubber solution (2% by weight of 1,4-polyisoprene dissolved in ethyl oleate), a trans-rubber solution (2% by weight of trans-1,4-polyisoprene dissolved in ethyl oleate), or a rubber emulsion (10% by weight of 1,4-polyisoprene and 0.2% by weight of the emulsifier polyoxyethylene(20)sorbitan monooleate) for 24 hours, and then stained with Oil Red O to detect the level of foam cell formation.

[0071] In the experiment, the subjects were divided into seven groups as shown in Table 1 below. [Table 1]

[0072] The morphology of the macrophages provided in the seven experimental groups was examined using an optical microscope. The test results are shown in Figure 2 (Figure 2(A) is a photograph of macrophages provided in the control group, Figure 2(B) is a photograph of macrophages provided in the oxLDL group, Figure 2(C) is a photograph of macrophages provided in the oxLDL + rubber solution group, Figure 2(D) is a photograph of macrophages provided in the oxLDL + trans-rubber solution group, Figure 2(E) is a photograph of macrophages provided in the oxLDL + vehicle group, Figure 2(F) is a photograph of macrophages provided in the oxLDL + rubber emulsion group, and Figure 2(G) is a photograph of macrophages provided in the oxLDL + emulsifier group). Statistics were also conducted on the percentage of foam macrophages out of the total number of cells in the seven groups, and Figure 3 is a bar graph of the foam macrophage content. In Figure 3, ns indicates no statistically significant difference, and an asterisk (*) indicates a significant difference (the same applies below).

[0073] Figures 2 and 3 show that compared to the oxLDL group, the foaming of macrophages was significantly suppressed in both the rubber solution group and rubber emulsion group containing cis and trans structures, resulting in a lower degree of foaming, whereas the solvent group and emulsifier group had no significant effect on the foaming of macrophages, resulting in a higher degree of foaming.

[0074] Furthermore, Figures 2 and 3 show that the cis / trans structure of 1,4-polyisoprene does not significantly affect drug activity.

[0075] (Example 2: Effect on macrophage polarization) The human macrophage cell line RAW264.7 was treated with the rubber solution provided in Example 1, the rubber emulsion provided in Example 2, or the trans-rubber solution provided in Example 3 for 24 hours, and then polarization of RAW264.7 was induced with lipopolysaccharide (LPS) for 10 hours, and the expression of marker genes associated with macrophage polarization was measured using real-time fluorescent quantitative PCR.

[0076] In the experiment, the subjects were divided into seven groups as shown in Table 2 below. [Table 2]

[0077] The mRNA levels of the seven experimental groups were measured using a fluorescent quantitative PCR system. The expression levels of the macrophage M1 marker genes IL-1β and iNOS after LPS treatment are shown in Figures 4 and 5, respectively. The expression levels of the macrophage M2 marker genes CD206 and IL-10 mRNA after LPS treatment are shown in Figures 6 and 7, respectively.

[0078] As can be seen from Figures 4-7, after treatment with LPS, the expression of macrophage M1 phenotype marker genes IL-1β and iNOS was upregulated, while the expression of M2 phenotype marker genes IL-10 and CD206 was downregulated. Compared to the LPS group, the rubber solution and rubber emulsion significantly reduced the expression levels of IL-1β and iNOS mRNA, and increased the expression levels of CD206 and IL-10 mRNA. Both the rubber solution group and the trans-rubber solution group demonstrated activity, with no statistically significant difference. Both the rubber solution and rubber emulsion containing cis and trans structures inhibited macrophage M1 polarization and promoted its polarization to M2, alleviating inflammatory responses. However, the solvent and emulsifier had no significant effect on macrophage polarization.

[0079] Furthermore, Figures 4 to 7 show that the cis / trans structure of 1,4-polyisoprene does not significantly affect drug activity.

[0080] (Example 3: Prevention research on atherosclerosis) The use case was a prevention study to study the preventive effect of drugs on atherosclerosis by initiating drug intervention at a time when atherosclerotic plaques had not yet formed in experimental mice.

[0081] Twenty-eight 8-week-old male apoE- / - mice were randomly divided into four groups, seven mice per group: control group, atherosclerosis model group (abbreviated as model group), model + rubber solution group, and model + rubber particles group.

[0082] The control group was fed a normal diet for 16 weeks, while the other three groups were fed a high-fat diet (21% fat, 0.5% cholesterol) for 16 weeks. The atherosclerosis model group, model + rubber solution group, and model + rubber particle group received daily intragastric administration of 3g / kg body weight of ethyl oleate, rubber solution, and ethyl oleate, respectively. Rubber particles were also added to the mouse feed of the model + rubber particle group. The grouping information for the experimental mice is shown in Table 3 below. [Table 3]

[0083] After the experiment, the mouse aortas and roots were harvested. The aortic roots were stained with Oil Red O. The positive areas were counted using ImageJ to determine the size of atherosclerotic plaques in the four experimental groups. The entire mouse aortas were stained with Oil Red O. The results of the detection of aortic plaque size are shown in Figure 8 (Figure 8(A) shows the aortic plaque size of the control group, Figure 8(B) shows the atherosclerosis model group, Figure 8(C) shows the aortic plaque size of the model + rubber solution group, and Figure 8(D) shows the aortic plaque size of the model + rubber particle group). The percentage of the positive area (plaque area) in the total aortic area is shown in Figure 9.

[0084] 8 and 9 show that the rubber solution significantly inhibited the expansion of plaque in atherosclerotic mice, while the plaque content in the rubber particle-fed group did not change significantly.

[0085] The root of the entire mouse aorta was stained with Oil Red O, and the results of detecting the size of the mouse aortic plaque are shown in Figure 10 (Figure 10(A) shows the results of detecting the size of the aortic root plaque in the control group of mice, Figure 10(B) shows the results of detecting the size of the aortic root plaque in the atherosclerosis model group of mice, Figure 10(C) shows the results of detecting the size of the aortic root plaque in the model + rubber solution group of mice, and Figure 10(D) shows the results of detecting the size of the aortic root plaque in the model + rubber particle group of mice), and Figure 11 is a bar graph of the positive area (plaque area).

[0086] 10 and 11 show that the rubber solution significantly inhibited the expansion of plaque in atherosclerotic mice, and that there was no significant change in plaque content in the rubber particle-fed group.

[0087] The development and progression of atherosclerotic plaques are important factors influencing cardiovascular and cerebrovascular diseases. This experiment demonstrated that rubber solution had a preventive effect on mice in which atherosclerotic plaques had not yet formed, slowing the rate of atherosclerosis formation and reducing plaque volume. Rubber particles did not show any preventive effect.

[0088] (Example 4: Therapeutic research into atherosclerosis) The use case was a therapeutic study to study the therapeutic effects of drugs on atherosclerosis by initiating drug intervention after atherosclerotic plaques had already formed in experimental mice.

[0089] Twenty-eight 8-week-old male apoE- / - mice were randomly divided into four groups, seven mice per group: control group, atherosclerosis model group, model + rubber solution group, and model + rubber particles group.

[0090] The control group was fed a normal diet for 16 weeks, while the other three groups were fed a high-fat diet (21% fat, 0.5% cholesterol) for 16 weeks. From week 12, the atherosclerosis model group, model + rubber solution group, and model + rubber particle group received daily intragastric administration of ethyl oleate, rubber solution, and ethyl oleate at doses of 3g / kg body weight, respectively. Rubber particles were also added to the high-fat diet of the model + rubber particle group from week 12. The grouping information for the experimental mice is shown in Table 4 below. [Table 4]

[0091] After the experiment was completed, the aorta and root of the mice were collected. The aortic root of the mice was then stained with Oil Red O. The positive areas were counted using the ImageJ program to detect the size of atherosclerotic plaques in the four experimental groups.

[0092] The entire mouse aorta was stained with Oil Red O, and the results of detecting the size of the mouse aortic plaque are shown in Figure 12 (Figure 12(A) shows the results of detecting the size of the aortic plaque in the control group of mice, Figure 12(B) shows the results of detecting the size of the aortic plaque in the atherosclerosis model group of mice, Figure 12(C) shows the results of detecting the size of the aortic plaque in the model + rubber solution group of mice, and Figure 12(D) shows the results of detecting the size of the aortic plaque in the model + rubber particle group of mice). The percentage of the area of ​​the positive region (plaque area) to the area of ​​the mouse aorta is shown in Figure 13.

[0093] 12 and 13 show that the rubber solution significantly inhibited the expansion of plaque in atherosclerotic mice, and there was no significant change in the plaque content in the rubber particle-fed group.

[0094] The root of the entire mouse aorta was stained with Oil Red O, and the results of detecting the size of the mouse aortic plaque are shown in Figure 14 (Figure 14(A) shows the results of detecting the size of the aortic root plaque in the control group of mice, Figure 14(B) shows the results of detecting the size of the aortic root plaque in the atherosclerosis model group of mice, Figure 14(C) shows the results of detecting the size of the aortic root plaque in the model + rubber solution group of mice, and Figure 14(D) shows the results of detecting the size of the aortic root plaque in the model + rubber particle group of mice), and Figure 15 is a bar graph of the positive area (plaque area).

[0095] 14 and 15 show that the rubber solution significantly inhibited the expansion of plaque in atherosclerotic mice, and that there was no significant change in plaque content in the rubber particle-fed group.

[0096] The development and progression of atherosclerotic plaques are important factors influencing cardiovascular and cerebrovascular diseases. This experiment demonstrated that rubber solution had a therapeutic effect on mice with already formed atherosclerotic plaques, slowing the rate of atherosclerosis formation and reducing plaque volume. Rubber particles did not show any therapeutic effect.

[0097] (Example 5: Treatment research for type 2 diabetes) Five 7-week-old SPF male m / m mice were used as the control group, and 15 7-week-old SPF male db / db mice were randomly divided into three groups, with five mice per group: type 2 diabetes group, type 2 diabetes + rubber solution group, and type 2 diabetes + rubber particle group. Random blood glucose measurements were taken for two days, and if the blood glucose level on the two days was 16.7 mmol / L or higher, the model was successfully established.

[0098] After the model was successfully established, the type 2 diabetes + rubber solution group was given rubber solution intragastrically at a dose of 3g / kg body weight once daily for 12 consecutive weeks, while the type 2 diabetes + rubber particle group was given ethyl oleate intragastrically at a dose of 3g / kg body weight once daily, while simultaneously replacing their feed with feed containing rubber particles, which was also given intragastrically for 12 consecutive weeks.

[0099] The following tests were conducted on mice in the control group, type 2 diabetes group, type 2 diabetes + rubber solution group, and type 2 diabetes + rubber particle group. (1) Blood glucose (FBG) content was measured. Mice were fasted for more than 12 hours without water restriction. At the time of measurement, the tails of the mice were disinfected with alcohol, blood was collected from the tip of the tail vein, and the second drop of blood was collected using a Roche blood glucose meter to measure blood glucose.

[0100] (2) Measurement of serum glucose (GLU) and glycated serum protein (GSP) contents. Whole blood was collected from mice to prepare serum, and the contents of glucose and glycated serum protein in the mouse serum were measured using an automatic biochemical analyzer.

[0101] The test results are shown in Figures 16 to 18. Figures 16 to 18 show that the rubber solution reduced blood sugar, serum glucose, and glycated serum protein content in diabetic model mice, indicating that it had a therapeutic effect on diabetes, but the rubber particles had no therapeutic effect on diabetes.

[0102] (Example 6: Treatment research for hypertriglyceridemia and hypercholesterolemia) Twenty 7-week-old apoe- / - mice were divided into four groups (5 mice per group): control, model, model + rubber solution, and model + rubber particles. The experiment lasted 8 weeks, with the first 4 weeks spent establishing the hyperlipidemia mouse model and the last 4 weeks spent on drug treatment.

[0103] During the eight-week feeding period, the control group was fed a basal diet, while the other three groups were always fed a high-sugar, high-fat diet, catalog number D12108C, purchased from Research Diets.

[0104] From the 5th to 8th weeks, the control group, model group, model + rubber solution group, and model + rubber particle group were intragastrically administered pure water, ethyl oleate, rubber solution, and ethyl oleate at a dose of 3g / kg body weight once a day for 4 consecutive weeks.From the 5th to 8th weeks, the feed of the model + rubber particle group was replaced with feed containing rubber particles.

[0105] The contents of total cholesterol (TC), triglyceride (TG), high-density lipoprotein cholesterol (HDL-C), and low-density lipoprotein cholesterol (LDL-C) in the serum of mice in each group were measured using an automatic biochemical analyzer.

[0106] The levels of triglyceride-related hydrolases in the serum of mice from each group, including adipose triglyceride hydrolase (ATGL), lipoprotein lipase (LPL), hepatic lipase (HL), and hormone-sensitive triglyceride lipase (HSL), were measured using enzyme-linked immunosorbent assay (ELISA).

[0107] Triglyceride hydrolase (ATGL) and hormone-sensitive triglyceride lipase (HSL) are the most important lipolytic enzymes in mammals, accounting for more than 95% of triglyceride hydrolase activity in white adipose tissue. Triglyceride (TG) degradation is primarily mediated by ATGL and HSL, which are key rate-limiting enzymes in fat hydrolysis. Hepatic lipase (HL) is primarily synthesized and secreted by hepatic parenchymal cells and is involved in lipid metabolism.

[0108] 19-22 show that the rubber solution reduces the total cholesterol, triglyceride, and low-density lipoprotein cholesterol content in blood lipids, and increases the high-density lipoprotein cholesterol content, while the rubber particles have no significant effect on blood lipids. The results show that the rubber solution has a significant therapeutic effect on hypercholesterolemia and hypertriglyceridemia, while the rubber particles have no therapeutic effect on hypercholesterolemia and hypertriglyceridemia.

[0109] Figures 23-26 show that the rubber solution significantly increased the levels of triglyceride metabolism-related enzymes adipose triglyceride hydrolase (ATGL), lipoprotein lipase (LPL), and hepatic lipase (HL), while significantly reduced the level of hormone-sensitive triglyceride lipase (HSL). This indicates that the rubber particles had no significant effect on triglyceride metabolism-related enzymes. This indicates that the rubber solution can promote the hydrolysis of TG. In other words, the rubber solution had a significant therapeutic effect on hypertriglyceridemia, while the rubber particles had no therapeutic effect on hypertriglyceridemia.

[0110] (Example 7: Treatment research for fatty liver) Twenty 6-week-old male C57BL / 6 mice were randomly divided into four groups, five per group: control group, fatty liver model group (abbreviated as model group), model + rubber solution group, and model + rubber particles group.

[0111] The control group was fed a basic diet for 14 weeks, while the other three groups were fed a high-fat diet (21% fat, 0.15% cholesterol) for 14 weeks, except for the model + rubber particle group, whose diet contained rubber particles. Simultaneously, the control, fatty liver model, model + rubber solution, and model + rubber particle groups were intragastrically administered pure water, ethyl oleate, rubber solution, and ethyl oleate at a dose of 3g / kg body weight once daily for 14 weeks.

[0112] After the experiment was completed, the livers and tissues of the mice were collected, photographed, and weighed.

[0113] Photographs of the liver morphology of mice in each group and statistical charts of the liver mass as a percentage of the total mass of the mice are shown in Figures 27 and 28, respectively (Figure 27(A) is a photograph of the liver of a mouse in the control group, Figure 27(B) is a photograph of the liver of a mouse in the model group, Figure 27(C) is a photograph of the liver of a mouse in the model + rubber solution group, and Figure 27(D) is a photograph of the liver of a mouse in the model + rubber particle group). Figure 27 shows that, compared to the control group, the model group successfully established fatty liver, and the liver fat accumulation of mice in the rubber solution group was significantly suppressed, while the liver fat of the rubber particle group did not change significantly compared to the model group. Figure 28 shows that the weight of the model group increased significantly compared to the control group, and the weight increase of fatty liver caused by lipid accumulation was significantly reversed in the rubber solution group, while the weight of the rubber particle group did not change significantly compared to the model group.

[0114] Furthermore, sections of liver tissue from mice in each group were prepared and stained with H&E and oil red, respectively. The H&E-stained and oil red-stained liver sections were examined under an optical microscope. The examination results are shown in Figures 29 and 30. Figures 29(A) and 30(A) are photographs of liver sections from mice in the control group after H&E and oil red staining, respectively. Figures 29(B) and 30(B) are photographs of liver sections from mice in the model group after H&E and oil red staining, respectively. Figures 29(C) and 30(C) are photographs of liver sections from mice in the model + rubber solution group after H&E and oil red staining, respectively. Figures 29(D) and 30(D) are photographs of liver sections from mice in the model + rubber particle group after H&E and oil red staining, respectively.

[0115] Figure 29 shows that H&E staining reveals the morphology of hepatocytes. Compared to the control group, the fatty liver model group had a large number of necrotic cells forming necrotic cores. Compared to the model group, the rubber solution group had intact hepatocyte structure, closely packed, no obvious cell necrosis, and no necrotic cores. The fatty liver lesions were significantly reversed, while the rubber particle group had a large number of necrotic cells forming necrotic cores, similar to the lesions in the model group.

[0116] Oil red staining showed the distribution of fat in hepatocytes, as shown in Figure 30. Compared with the model group, the rubber solution significantly suppressed fat gain in mice with fatty liver induced by a high-fat diet, and there was no significant change in fat content in the rubber particle-fed group.

[0117] The triglyceride content in liver tissue is a quantitative indicator of the degree of fatty liver lesions. The triglyceride (TG) content in the liver tissue of mice from each group was measured using a tissue triglyceride (TG) enzyme measurement kit to quantify the degree of fatty liver lesions, and the test results are shown in Figure 31. Figure 31 shows that, compared to the model group, intragastric administration of the rubber solution significantly reduced the triglyceride content in liver tissue, while there was no significant change in triglyceride content in the rubber particle-fed group.

[0118] From the above, it was found that the rubber solution can significantly inhibit the progression of fatty liver caused by a high-fat diet and has a therapeutic effect on fatty liver, but the rubber particles do not have this effect.

[0119] (Example 8: Treatment research on colitis) Thirty-two 8-week-old male mice were randomly divided into four groups of 8 mice each: a control group, a colitis model group (abbreviated as model group), a model + rubber solution group, and a model + rubber particles group.

[0120] The experimental period lasted for seven days. The control group was fed normal chow, while the other three groups had their chow supplemented with 4% dextran sulfate sodium (DSS) to induce colitis. The model + rubber particle group, however, had rubber particles added to its chow. Simultaneously, the control, colitis model, model + rubber solution, and model + rubber particle groups received 3g / kg body weight of pure water, ethyl oleate, rubber solution, and ethyl oleate intragastrically once daily for seven days.

[0121] During the experiment, the mice were weighed and recorded daily, and their feces were collected daily to determine the condition of the feces and whether or not there was any occult blood present. A DAI score was determined based on these data.

[0122] The disease activity index (DAI) was calculated by a comprehensive score based on three indices: the percentage of mouse weight loss, stool consistency, and stool bleeding. The criteria for the DAI score are shown in Table 5. [Table 5]

[0123] In the table above, normal stool refers to stool that is well-formed, muddy stool refers to partially formed paste-like stool that does not stick to the anus, and watery stool refers to stool with a high water content that can stick to the anus.

[0124] The DAI score quantifies the severity of colitis lesions in mice. The higher the DAI score, the more severe the colitis lesions. The DAI score results are shown in Figure 32. Figure 32 shows that the DAI score of the model + rubber solution group was lower than that of the colitis model group, but there was no significant difference between the model + rubber particle group and the colitis model group. This indicates that the rubber solution has the effect of alleviating colitis symptoms in mice, but the rubber particles do not have this effect.

[0125] The weight loss of mice, even under constant conditions, is also an indicator of the severity of the colitis lesions in the mice. Figure 33 shows the weight loss rate of mice in the four experimental groups. Figure 33 shows that the weight of mice in the colitis model group gradually decreased over the course of the study. The weight loss of mice in the model + rubber solution group decreased more slowly than that of the colitis model group, and the degree of weight loss in the model + rubber particle group was not significantly different from that of the colitis model group. This indicates that the rubber solution has the effect of alleviating colitis symptoms in mice, while the rubber particles do not.

[0126] The length and weight of the mouse colon were also quantitative indicators of the severity of colitis. The more severe the colitis in the mouse, the shorter the length and weight of the colon. After 7 days, the mice were euthanized and their colons were collected. Photographs of these specimens are shown in Figure 34. The length and weight of the colons of each group were measured, with the results shown in Figures 35 and 36, respectively. Figures 34-36 show that the length and weight of the colons of the colitis model group were reduced compared to the control group. However, the model + rubber solution group had a smaller effect on the colons of the mice than the colitis model group, and there was no significant difference in the length or weight of the colons of the mice in the colitis group between the model and rubber particle groups. This indicates that the rubber solution is effective in treating colitis in mice, while the rubber particles do not.

[0127] (Example 9: Treatment research for polycystic ovary syndrome) Twenty 4-week-old female SD rats were randomly divided into four groups, five per group: a control group, a polycystic ovary syndrome model group (abbreviated as model group), a model + rubber solution group, and a model + rubber particle group.

[0128] The experimental period lasted three weeks. During the experiment, rats in the other three groups, excluding the control group, were intragastrically administered letrozole once daily for three weeks to establish a polycystic ovary syndrome (PCOS) model. At the same time, the control group, PCOS model group, model + rubber solution group, and model + rubber particle group were intragastrically administered pure water, ethyl oleate, rubber solution, and ethyl oleate at 3g / kg body weight once daily for three weeks. The diet of the model + rubber particle group was replaced with diet containing rubber particles.

[0129] After the intragastric administration, the rats were weighed, blood was collected from the tail vein, and serum was prepared to measure testosterone content. The rats were fasted for 12 hours, and blood was collected from the tail vein and measured for fasting blood glucose using a blood glucose meter. The rats were then intraperitoneally injected with glucose (10 g / 100 mL) at a dose of 2 g / kg. Blood was collected from the tail vein at 15, 30, 60, 90, and 120 minutes after injection, and blood glucose levels were measured using a blood glucose meter. The rats were then euthanized, and ovarian tissue was removed. RNA was extracted from the tissue, and mRNA expression levels of anti-Müllerian hormone (AMH), IL-1β, and TNF-α were measured using a real-time fluorescent quantitative PCR system.

[0130] Once the rat polycystic ovary syndrome model was established, the rats' weight gradually increased. The extent of weight gain can reflect the severity of polycystic ovary syndrome (PCOS) lesions. The weights of the four groups of rats were measured, and the test results are shown in Figure 37. Figure 37 shows that intragastric administration of the rubber solution significantly reduced the weight gain caused by PCOS, while feeding the rubber particles had no significant effect on the rats' weight.

[0131] One of the important characteristics of polycystic ovary syndrome is elevated androgen levels, and androgen testosterone levels can reflect the severity of polycystic ovary syndrome lesions. The serum testosterone content of the rats in the above four groups was measured, and the test results shown in Figure 38 show that intragastric administration of rubber solution significantly inhibited the increase in serum testosterone in polycystic ovary syndrome model rats, and the serum testosterone content of the rubber particle feeding group did not change significantly compared to the model group.

[0132] Impaired glucose tolerance is a common metabolic disorder associated with polycystic ovary syndrome (PCOS), and the severity of impaired glucose tolerance can reflect the severity of the disease. The fasting blood glucose levels, blood glucose content at different times, and glucose tolerance of the four rat groups were measured, and the test results are shown in Figures 39-41. Figures 39-41 show that the rats in the PCOS model group developed increased fasting blood glucose levels and impaired glucose tolerance. However, after intragastric administration of a rubber solution, fasting blood glucose levels and impaired glucose tolerance were significantly alleviated, while feeding the rats with rubber particles had no significant effect.

[0133] Elevated anti-Mullerian hormone (AMH) levels indicate impaired follicular development in patients with polycystic ovary syndrome. Anti-Mullerian hormone (AMH) / GAPDH (mRNA) levels in the blood of rats from the above four groups were measured (normalized using the GAPDH internal reference gene), and the test results are shown in Figure 42. Figure 42 shows that the AMH mRNA levels in the ovarian tissue of rats from the polycystic ovary syndrome model group were significantly elevated, and that it was the intragastric administration of rubber solution, not feeding rubber particles, that reduced the AMH mRNA levels.

[0134] The ongoing chronic inflammatory response in the ovarian tissue of patients with polycystic ovary syndrome (PCOS) and increased expression of the chronic inflammatory factors IL-1β and TNF-α can be used as an indicator of the severity of the disease. The expression levels of IL-1β and TNF-α in the blood of rats from the four groups were measured, and the test results are shown in Figures 43 and 44, respectively. Figures 43 and 44 show that, compared with the control group, rats in the PCOS model group had significantly increased expression of IL-1β and TNF-α in ovarian tissue. After intragastric administration of a rubber solution, inflammatory factor expression in the rat ovaries was significantly downregulated, while inflammatory factor expression in the rubber particle-fed group remained unchanged.

[0135] Based on the above findings, a comprehensive comparison of multiple indicators of rats with polycystic ovary syndrome, including body weight, serum testosterone levels, fasting blood glucose, glucose tolerance, anti-Mullerian hormone (AMH) levels in ovarian tissue, and expression levels of inflammatory factors IL-1β and TNF-α, showed that the rubber solution had a therapeutic effect on polycystic ovary syndrome, while the rubber particles did not.

[0136] (Example 10: Obesity treatment research) Twenty-four 6-week-old male C57BL / 6 mice were randomly divided into four groups (6 mice per group): control, high-fat model, high-fat + rubber solution, and high-fat + rubber particles. The average weight of the mice in the four groups was 20±2 g.

[0137] The experimental period was 18 weeks. The control group was always fed a basal diet, while the other three groups were always fed a high-fat diet (35% fat content), except for the high-fat + rubber particle group, whose diet also contained rubber particles. The control group, high-fat model group, high-fat + rubber solution group, and high-fat + rubber particle group were intragastrically administered pure water, ethyl oleate, rubber solution, and ethyl oleate at a dose of 3g / kg body weight once daily for 18 weeks.

[0138] The body weight of the mice was recorded weekly, and at the end of the experiment, the fat mass of the mice was recorded and analyzed using a mouse body fat scale. The mice were then euthanized, and the adipose tissue around the epididymis was collected and sectioned. H&E staining was then performed, and the area per adipocyte was calculated.

[0139] The body weight and fat mass measurements of the mice in the four experimental groups are shown in Figures 45 and 46, respectively. Figure 45 shows that the body weight of the mice in the high-fat model group was significantly higher than that of the control group, and that intragastric administration of rubber solution significantly inhibited the weight gain induced by the high-fat diet, while feeding rubber particles did not have this inhibitory effect. Figure 46 shows that the fat mass data is consistent with the body weight data, and that rubber solution significantly inhibited the high-fat-induced increase in fat mass in the mice, but feeding rubber particles did not have this inhibitory effect.

[0140] The adipose tissue morphology of the mice was examined using an optical microscope, and the test results are shown in Figure 47 (Figure 47(A) is a photograph of the adipose tissue morphology of the control group, Figure 47(B) is a photograph of the adipose tissue morphology of the high-fat model group, Figure 47(C) is a photograph of the adipose tissue morphology of the model + rubber solution group, and Figure 47(D) is a photograph of the adipose tissue morphology of the model + rubber particle group). As can be seen from Figure 47, the adipocytes of the mice in the control group were small in volume and arranged in an orderly fashion. The obese mice in the high-fat group had enlarged adipocytes, some of which were infiltrated with inflammatory cells. The rubber solution significantly suppressed the increase in adipocytes induced by a high-fat diet. The cell morphology of the rubber particle group was similar to that of the high-fat group.

[0141] An optical microscope was used to measure the average area of ​​the mice's fat cells, and the test results are shown in Figure 48. From Figure 48, by statistically calculating the area of ​​each fat cell, it was found that the area per fat cell in the high fat group was 2.8 times that of the control group, and the rubber solution significantly inhibited the trend of fat cell area expansion, with the area per fat cell being only 1.2 times that of the control group, indicating that there was no significant change in the rubber particle group compared to the high fat group.

[0142] A comprehensive comparison of data such as body weight, fat weight, fat histopathological staining, and area per adipocyte of the four groups of mice revealed that the rubber solution was able to significantly suppress high-fat diet-induced obesity and had a therapeutic effect on obesity, while the rubber particles did not have this effect.

[0143] (Example 11: Study on the state of natural rubber latex, rubber emulsion, and rubber solution in the stomach of mice) Fifteen 6-week-old male C57BL / 6 mice were randomly divided into three groups (5 mice per group): natural latex group, rubber emulsion group, and rubber solution group. The experiment lasted for 7 days and the mice were fed a basal diet.

[0144] Mice in the natural latex group, rubber emulsion group, and rubber solution group were intragastrically administered 3 g / kg of natural latex extracted from Hevea brasiliensis, the rubber emulsion provided by the present application, and the rubber solution provided by the present application, respectively, once daily for 7 days. Feces were collected daily and examined under a dissecting microscope. After the final intragastric administration, the dose was increased to 15 g / kg. The mice were euthanized within 30 minutes of the end of the intragastric administration, and the stomach contents were removed and examined under a dissecting microscope.

[0145] The stomach contents of the mice in the natural latex group, rubber solution group, and rubber emulsion group were examined, and the results are shown in Figures 49 to 51. Figures 49 to 51 show that rubber aggregates were observed in the stomach contents of the mice in the natural latex group, but no rubber aggregates were observed in the stomach contents of the mice in the rubber emulsion group or rubber solution group.

[0146] This showed that when natural rubber latex enters the stomach of a mouse, it directly collects and coagulates into large rubber lumps, which are unable to remain dispersed and have no medicinal activity, and are then excreted from the body through the digestive tract. However, when the rubber emulsion and rubber solution provided by the present application enters the digestive tract of a mouse, they do not coagulate into large rubber lumps and always remain dispersed, thereby maintaining medicinal activity.

[0147] (Example 12: Study on the stability of rubber emulsions prepared with different emulsifiers in strong and weak acidic environments) The inventors have verified the stability under acidic conditions of conventional natural latex extracted from rubber trees, industrial concentrated latex produced from natural latex (including high ammonia-concentrated latex, low ammonia-concentrated latex, and low protein-concentrated latex), and synthetic 1,4-polyisoprene latex, and to compare these with the 1,4-polyisoprene emulsion system produced with the preferred emulsifier of the present application, they simulated the gastric acid environment of a mammal in vitro using hydrochloric acid solutions of pH 1 and pH 3 to confirm the stability of rubber emulsions produced with different emulsifiers. The former represents the strongly acidic environment of gastric juice, and the latter represents the weakly acidic environment of gastric juice.

[0148] For the method of preparing the 1,4-polyisoprene emulsion system with the preferred emulsifier of the present application, reference can be made to the method provided in Example 2, except that the emulsifier is different.

[0149] Is it stable under weakly acidic conditions of pH 3? Add 5 mL of pH 3 hydrochloric acid solution (1.0 mmol / L) to a glass dish, add 0.1 mL of 1,4-polyisoprene emulsion dropwise using a pipette, and stir until homogenous. Observe whether a solid precipitates. If a solid precipitates, the 1,4-polyisoprene emulsion system is unstable under weakly acidic conditions of pH 3. Otherwise, it is stable. Is it stable under strong acidic conditions at pH=1? Add 5 mL of pH=1 hydrochloric acid solution (0.1 mol / L) to a glass dish, add 0.1 mL of 1,4-polyisoprene emulsion dropwise using a pipette, stir until homogenous, and observe whether a solid precipitates. If a solid precipitates, the 1,4-polyisoprene emulsion system is unstable under strong acidic conditions at pH=1. Otherwise, it is stable.

[0150] See Table 6 below for detailed descriptions and test results for different 1,4-polyisoprene emulsion dispersions. [Table 6] JPEG2026508634000008.jpg219170

[0151] Figure 52 shows some test results. A is a photograph of natural latex from the Para rubber tree being added dropwise to hydrochloric acid at pH 3. B is a photograph of high ammonia concentration natural latex being added dropwise to hydrochloric acid at pH=3. C is a photograph of low ammonia concentration natural latex being added dropwise to hydrochloric acid at pH=3. D is a photograph of low protein concentrated natural latex being added dropwise to hydrochloric acid at pH=3. E is a photograph of synthetic isoprene latex added dropwise to hydrochloric acid at pH=3. F is a photograph of a rubber emulsion prepared in this application using polyoxyethylene (20) sorbitan monolaurate (polysorbate 20) as an emulsifier, added dropwise to hydrochloric acid at pH 1. G is a photograph of a rubber emulsion prepared in this application using polyoxyethylene (20) sorbitan monopalmitate (polysorbate 40) as an emulsifier, added dropwise to hydrochloric acid at pH 1. H is a photograph of a rubber emulsion prepared in this application using polyoxyethylene (20) sorbitan monostearate (polysorbate 60) as an emulsifier, added dropwise to hydrochloric acid at pH 1. I is a photograph of a rubber emulsion prepared in Example 2 of the present application using polyoxyethylene (20) sorbitan monooleate (polysorbate 80) as an emulsifier, which was added dropwise to hydrochloric acid at pH 1. J is a photograph of a rubber emulsion prepared in this application using decaglycerol monolaurate (Q-12S) as an emulsifier, which was added dropwise to hydrochloric acid at pH 1. K is a photograph of a rubber emulsion prepared in this application using decaglycerol monomyristate ester (Q-14S) as an emulsifier, which was added dropwise to hydrochloric acid at pH 1. L is a photograph of a rubber emulsion prepared in this application using decaglycerol monopalmitate (Q-16S) as an emulsifier, which was added dropwise to hydrochloric acid at pH 1. M is the result of adding dropwise the rubber emulsion prepared in this application using decaglycerol monooleate (Q-17S) as an emulsifier to hydrochloric acid at pH=1. N is a photograph of a rubber emulsion prepared in this application using decaglycerol monostearate (Q-18S) as an emulsifier, which was added dropwise to hydrochloric acid at pH 1. O is a photograph of a rubber emulsion prepared in this application using sucrose monostearate as an emulsifier, which was added dropwise to hydrochloric acid at pH 1.

[0152] From Table 6 and Figure 50, it can be seen that the conventional natural latex extracted from Para rubber trees, industrial concentrated latex (including high ammonia concentrated latex, low ammonia concentrated latex, and low protein concentrated latex) produced from natural latex, and artificially synthesized 1,4-polyisoprene latex are all unstable under acidic conditions of pH=1 and pH=3, and 1,4-polyisoprene coagulates and precipitates. This means that it coagulates and precipitates even in mammalian gastric juice, cannot maintain a dispersed state, and does not exert its pharmacological activity.

[0153] In the present application, emulsion systems prepared using polyoxyethylene (20) sorbitan monolaurate (polysorbate 20), polyoxyethylene (20) sorbitan monopalmitate (polysorbate 40), polyoxyethylene (20) sorbitan monostearate (polysorbate 60), polyoxyethylene (20) sorbitan monooleate (polysorbate 80), decaglycerol monolaurate (Q-12S), decaglycerol monomyristate (Q-14S), decaglycerol monopalmitate (Q-16S), decaglycerol monooleate (Q-17S), decaglycerol monostearate (Q-18S), or sucrose monostearate as emulsifiers are preferred, and all of these emulsion systems remain stable under acidic conditions of pH 1.0 and pH 3.0. This indicates that the rubber emulsion thus prepared maintains a dispersed state even in the gastric juice of mammals and exerts pharmacological activity.

[0154] (Example 13: Rescue of polyisoprene dispersions on MTT cell survival in a neuronal apoptosis model) We present a model of L-glutamate-induced neuronal apoptosis. L-glutamate (L-glutamate), also known as glutamate, is an endogenous neurotransmitter abundant in the mammalian brain and spinal cord. It plays an important role in neuronal development, differentiation, and migration, as well as in synaptic initiation, development, plasticity, and learning and memory processes. Under normal physiological conditions, glutamate maintains normal neuronal activity by binding to glutamate receptors (GluRs) on the postsynaptic membrane. Some pathological stimuli can increase extracellular glutamate concentrations, resulting in excessive calcium ion influx, leading to persistent neuronal depolarization and excessive activation of GluRs on the postsynaptic membrane. This can further induce adverse events, such as excitotoxicity, cell apoptosis, and oxidative stress, leading to neuronal degeneration and eventual death. Recent studies have demonstrated that many neuropsychiatric disorders, including Alzheimer's disease (AD), are closely related to excessive extracellular glutamate concentrations. Therefore, glutamate-induced neuronal apoptosis models are widely used in disease research and drug screening for Alzheimer's disease (AD).

[0155] In this example, the neuronal cells used were the HT-22 cell line derived from mouse hippocampal neurons. The rescue effects of different concentrations of polyisoprene dispersions (rubber solution and emulsion) on apoptotic HT-22 cells were compared by measuring the MTT cell viability index. MTT is 3-(4,5-dimethylthiazol-2)-2,5-diphenyltetrazolium bromide, and the MTT method is a method for detecting cell viability and growth.

[0156] The grouping information is as follows: Twelve groups were set up: 1. control group, 2. L-GLU modeling group, 3-7. rubber solution groups (with polyisoprene mass fractions of 4 ppm, 10 ppm, 20 ppm, 40 ppm, and 100 ppm, respectively. The polyisoprene in these five groups was derived from the 1,4-polyisoprene solution dispersion with a mass percentage content of 2% prepared in Example 1), and 8-12. polyisoprene emulsion treatment groups (with polyisoprene mass fractions of 4 ppm, 10 ppm, 20 ppm, 40 ppm, and 100 ppm, respectively. The polyisoprene in these five groups was derived from the 1,4-polyisoprene emulsion dispersion with a mass percentage content of 10% prepared in Example 2, which was then diluted 5 times with pure water to obtain a 2% 1,4-polyisoprene emulsion dispersion).

[0157] The cell additives for the 12 groups and their final concentrations in the medium are shown in Table 7 below. [Table 7]

[0158] The experimental steps are as follows. (1) Culturing: A cell suspension was prepared in a culture medium containing 10% fetal bovine serum and seeded into a 96-well plate at 5,000 cells per well, with the volume of the cell suspension in each well being 200 μL. (2) According to the above grouping information, the corresponding substances were added to each of the 12 groups of cells, and eight replicate wells were prepared for each group, followed by incubation for 24 hours. (3) After the incubation was completed, 20 μL of MTT solution (5 mg / mL, prepared in serum- and antibiotic-free medium) was added to each well and incubated for 4 hours. The incubation was then stopped, the medium was carefully aspirated from the wells, 150 μL of DMSO was added to each well, and the wells were shaken at room temperature for 15 minutes to thoroughly dissolve the crystals. (4) A wavelength of 550 nm was selected, and the absorbance of each well was measured using a microplate reader, and the results were recorded.

[0159] The results are analyzed as follows: The experimental results are shown in Figure 53. The results demonstrate that polyisoprene, in either a solution or emulsion dispersion, rescues L-glutamic acid (L-GLU)-induced neuronal apoptosis, and that the rescue effect is dose-dependent. That is, polyisoprene has a significant rescue effect at mass fractions of 20 ppm and 40 ppm (P<0.05), but there is no significant difference in the rescue effect at mass fractions below 20 ppm or above 40 ppm (P>0.05).

[0160] (Example 14: Reduction of the proportion of late apoptotic cells in a neuronal apoptosis model using polyisoprene dispersions) We present a model of L-glutamate-induced neuronal apoptosis. L-glutamate (L-glutamate), also known as glutamate, is an endogenous neurotransmitter abundant in the mammalian brain and spinal cord. It plays an important role in neuronal development, differentiation, and migration, as well as in synaptic initiation, development, plasticity, and learning and memory processes. Under normal physiological conditions, glutamate maintains normal neuronal activity by binding to glutamate receptors (GluRs) on the postsynaptic membrane. Some pathological stimuli can increase extracellular glutamate concentrations, resulting in excessive calcium ion influx, leading to persistent neuronal depolarization and excessive activation of GluRs on the postsynaptic membrane. This can further induce adverse events, such as excitotoxicity, cell apoptosis, and oxidative stress, leading to neuronal degeneration and eventual death. Recent studies have demonstrated that many neuropsychiatric disorders, including Alzheimer's disease (AD), are closely related to excessive extracellular glutamate concentrations. Therefore, glutamate-induced neuronal apoptosis models are widely used in disease research and drug screening for Alzheimer's disease (AD).

[0161] In this example, the neuronal cells used were the HT-22 cell line derived from mouse hippocampal neurons. The percentage of HT-22 cells in the late apoptotic stage was measured to detect the rescue effect of polyisoprene dispersions (rubber solution and emulsion).

[0162] The grouping information is as follows: In this example, four groups of HT-22 mouse hippocampal neurons were prepared, with each group of cells being prepared in three replicate wells.

[0163] Group 1 is a control group in which no substances other than the medium were added to the cells. Group 2 was an L-GLU modeling group to which 20 mM L-glutamic acid was added to generate a neuronal apoptosis model. Group 3 is a rubber solution group containing 20 mM L-glutamic acid and 20 ppm (20 μg / mL) of polyisoprene by mass fraction, which was derived from the 1,4-polyisoprene solution dispersion prepared in Example 1 with a 2% mass percentage content. Group 4 is a rubber emulsion group containing 20 mM L-glutamic acid and 20 ppm (20 μg / mL) of polyisoprene by mass fraction. The polyisoprene was derived from the 1,4-polyisoprene emulsion dispersion with a mass percentage content of 10% prepared in Example 2, and then diluted 5 times with pure water to obtain a 2% 1,4-polyisoprene emulsion dispersion. The additives for the four groups of cells and their final concentrations in the medium are shown in Table 8 below. [Table 8]

[0164] The specific experimental procedures are as follows. (1) HT-22 cells were seeded in 6-well plates, with each group consisting of three replicate wells. Once the cells began to adhere to the wall, they were treated with a polyisoprene solution dispersion, a polyisoprene emulsion dispersion, and 20 mM L-Glu for 24 hours. The mass fraction of polyisoprene in the cell culture medium was 20 ppm, or 20 μg / mL. The control group was left untreated, while the L-Glu modeling group was treated with 20 mM L-glutamic acid alone for 24 hours. (2) After the treatment was completed, the cells were digested with trypsin (containing no EDTA) and then washed twice with pre-chilled PBS. (3) After washing, the cells were incubated with Annexin V-PE and PI working solution for 20 minutes in the dark at room temperature, and then cell apoptosis was detected using a flow cytometer. Three replicate wells of cells from each group were each detected once. Typical detection results are shown in Figure 2. (4) The proportion of late apoptotic cells in each group was counted and statistically analyzed, and the results are shown in Figure 3.

[0165] The results are analyzed as follows: In Figure 54, the light grey dots in the upper right quadrant of the crosshairs in each image represent late apoptotic cells, and the corresponding numbers indicate the percentage of late apoptotic cells.

[0166] As can be seen from Figure 55, after treatment with L-glutamic acid, the proportion of late apoptotic cells in the model group significantly increased, but after treatment with 20 μg / mL polyisoprene, the proportion of late apoptotic cells significantly decreased, indicating that polyisoprene has a significant apoptosis-inhibiting effect on the neurons in this model (P<0.05). Whether polyisoprene is derived from a solution dispersion system or an emulsion dispersion system, it has the same effect and there is no difference.

[0167] (Example 15: In vivo animal experiment using a mouse water maze model) The animal model, grouping information, and test method are as follows: In this experiment, male C57BL / 6J APP / PS1 mice were used, which are double transgenic AD model mice. The temperature in the animal room was maintained at 24±2°C, and a 12-hour light-dark cycle was applied. Food intake, water intake, and body weight were monitored daily throughout the experiment. Male C57BL / 6J APP / PS1 mice (2 months old, weighing approximately 20 g) and their wild-type (WT) littermate control mice were purchased from GemPharmatech Co. Ltd. (Nanjing, Jiangsu Province, China). All animals had unlimited access to food and water.

[0168] C57BL / 6J wild-type mice fed a normal diet served as the normal control group (WT group, 12 mice / group).

[0169] Thirty-six APP / PS1 mice will be randomly divided into the following three groups (12 mice / group) and will receive the following treatments. The AD model group consists of APP / PS1 mice fed a normal diet. In the rubber solution group, APP / PS1 mice were fed with a normal diet and were intragastrically administered 100 μL of the 1,4-polyisoprene solution dispersion system with a mass percentage content of 2% prepared in Example 1 every day. In the rubber emulsion group, APP / PS1 mice were fed a normal diet and daily intragastrically administered 100 μL of 2% 1,4-polyisoprene emulsion dispersion. The 2% 1,4-polyisoprene emulsion dispersion was obtained by diluting the 10% 1,4-polyisoprene emulsion dispersion prepared in Example 2 five times with pure water.

[0170] All mice were kept and treated for 7 months. After the treatment, they were subjected to a water maze test, during which their movement trajectories were recorded to intuitively observe changes in spatial memory in each group of mice.

[0171] The results are analyzed as follows: The results of the maze model test conducted on the mice of the above four experimental groups are shown in Figures 56 to 59. Figure 56 records the movement trajectories of the mice of each group, with the upper right quadrant being the target quadrant. Figure 57 is a statistical diagram of the escape latency of the mice of each group. Figure 58 is a statistical diagram of the number of times the mice of each group crossed the target platform. Figure 59 is a statistical diagram of the number of times the mice of each group crossed the target quadrant.

[0172] As can be seen from Figures 56-59, compared to the AD model mice, the mice treated with the polyisoprene dispersion system had a significantly shorter escape latency (P<0.05), and the number of crossings to the target platform and the number of crossings to the target quadrant were both significantly increased (P<0.05). This suggests that polyisoprene significantly improved the cognitive function of the AD model mice and may be developed as a therapeutic drug for AD. Polyisoprene, whether dispersed in a solution or emulsion, has AD therapeutic activity.

[0173] Based on the above findings, the present application has demonstrated that 1,4-polyisoprene, when dispersed in a solution dispersion system and an emulsion dispersion system, remains stable in the gastric acid environment of mammals, does not precipitate 1,4-polyisoprene aggregates, is non-oral toxic to mammals, and is allergen-free. Dispersed 1,4-polyisoprene significantly inhibits macrophage foam cell formation, inhibits macrophage polarization into M1 type, promotes macrophage polarization into M2 type, and alleviates inflammatory responses. This has significant benefits for the prevention and treatment of atherosclerotic cardiovascular and cerebrovascular diseases, as well as for type 2 diabetes, hypercholesterolemia, hypertriglyceridemia, fatty liver, colitis, polycystic ovary syndrome, obesity, and Alzheimer's disease. It also has a reversible effect on L-glutamate (L-GLU)-induced neuronal apoptosis, significantly inhibits neuronal apoptosis, and effectively improves cognitive function in AD model mice.

[0174] Although we have described the detailed process of the present application through the above examples in this specification, we do not claim that the present application is limited to the above detailed process, that is, the present application can only be carried out by the above detailed process. It should be understood by those skilled in the art that any improvements to the present application, such as equivalent replacement of each raw material of the present application product, especially solvents and emulsifiers, addition of auxiliary ingredients, selection of specific methods, etc., are also within the scope of protection and disclosure of the present application.

Claims

1. 1,4-polyisoprene dispersion, It remains stable in the gastric acid environment of mammals, does not precipitate 1,4-polyisoprene aggregates, is non-toxic to mammals by oral administration, and does not contain any allergens. 1,4-Polyisoprene dispersion.

2. the 1,4-polyisoprene dispersions include 1,4-polyisoprene solution dispersions and 1,4-polyisoprene emulsion dispersions; The components of the 1,4-polyisoprene solution dispersion include 1,4-polyisoprene and a solvent, 10. The 1,4-polyisoprene dispersion of claim 1, wherein the components of the 1,4-polyisoprene emulsion dispersion include 1,4-polyisoprene, an emulsifier, and water.

3. 3. The 1,4-polyisoprene dispersion according to claim 1, wherein the degree of polymerization of the 1,4-polyisoprene is 6 or more.

4. The solvent is selected from any one or a combination of at least two of a fatty acid, a fatty alcohol, an ester compound, or a lipid compound, Preferably, the ester-based compound includes any one of a monohydric alcohol ester, a dihydric alcohol ester, and a polyhydric alcohol ester, or a combination of at least two of them; The 1,4-polyisoprene dispersion according to claim 2 or 3, wherein the lipid compound preferably comprises any one or a combination of at least two of a phospholipid, a glycolipid, a sphingolipid, a steroid, or squalene.

5. 5. The 1,4-polyisoprene dispersion of any one of claims 2 to 4, wherein the weight percent content of 1,4-polyisoprene in the 1,4-polyisoprene solution dispersion is less than or equal to 60% and is not 0, preferably between 0.2% and 20%.

6. the emulsifier is selected from one or at least two components of a cationic surfactant, an anionic surfactant, a zwitterionic surfactant, and a nonionic surfactant; Preferably, the nonionic surfactant is polyoxyethylene (20) sorbitan monolaurate, polyoxyethylene (20) sorbitan monopalmitate, polyoxyethylene (20) sorbitan monostearate, polyoxyethylene (20) sorbitan monooleate, decaglycerol monolaurate, decaglycerol monomyristate, decaglycerol monopalmitate, decaglycerol monooleate, decaglycerol monostearate or C １２ ~C １８ 6. The 1,4-polyisoprene dispersion of any one of claims 2 to 5, comprising any one or a combination of at least two of the mono-fatty acid sucrose esters.

7. The mass percent content of 1,4-polyisoprene in the 1,4-polyisoprene emulsion dispersion is 80% or less and not 0, preferably 0.5% to 70%; 7. The 1,4-polyisoprene dispersion according to any one of claims 2 to 6, wherein the average particle size of the emulsion droplets in the 1,4-polyisoprene emulsion dispersion is 10 μm or less.

8. A pharmaceutical active ingredient comprising the 1,4-polyisoprene dispersion of any one of claims 1 to 7.

9. A drug, 10. A pharmaceutical composition comprising the active pharmaceutical ingredient of claim 9, Preferably, the drug comprises a drug for treating a chronic metabolic disease; Preferably, the chronic metabolic diseases include atherosclerotic cardiovascular and cerebrovascular diseases, type 2 diabetes, hypercholesterolemia, hypertriglyceridemia, fatty liver, colitis, polycystic ovary syndrome and obesity; Drugs.

10. 1. Use of a drug in the treatment of Alzheimer's disease, The drug comprises the active pharmaceutical ingredient of claim 8, wherein the active pharmaceutical ingredient comprises the 1,4-polyisoprene dispersion of any one of claims 1 to 7.

Images