Nano-delivery carrier

Abstract

Provided is a stevioside derivative. The stevioside derivative is a compound represented by general formula Ia, or a salt thereof, or a hydrate thereof, or any mixture of the compound represented by general formula Ia, the salt thereof, and the hydrate thereof. In general formula Ia, R1 and R2 are independent of each other. R1 is selected from hydroxyl or glycosyl, and R2 is selected from n-valent rhamnose, rhamnopyranose, mannose, mannopyranose, galactose, galactopyranose, or combinations thereof, or oligosaccharides formed by means of covalent bonding with other sugars, wherein 1 ≤ n ≤ 10, for example, n is 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10. The derivative or an in-vivo metabolite thereof has balanced hydrophilicity and lipophilicity and a multi-triggerable group and can serve as a delivery carrier and / or targeted guiding agent for an active ingredient. Further provided is a preparation method for the derivative and a use of the derivative.

Description

Nano delivery carrier TECHNICAL FIELD

[0001] The present application relates to the field of daily chemical and pharmaceutical technology. Specifically, it relates to a steviol glycoside derivative with targeting effect, a preparation method and uses thereof. BACKGROUND

[0002] Steviol glycoside is a natural sweetener extracted from the herbaceous plant Stevia rebaudiana (Bertoni) of the Asteraceae family. The use of Stevia rebaudiana as a medicinal herb or steviol glycoside as a sugar substitute has a history of several hundred years in South America. Stevioside as a medicinal adjuvant is a mixture of steviol glycosides mainly containing stevioside, with a steviol glycoside content of ≥95.0%; and as a food additive, steviol glycoside requires a glycoside content of ≥90.0%. In addition, stevioside, rebaudioside-A, B, C, D, E, M, and AM are listed as food additives in the GRAS (Generally Recognized As Safe, GRAS) material list in the United States. The general structure of steviol glycoside compounds is shown in (1):

[0003] G is selected from the following groups:

[0004] Steviol glycoside compounds belong to the class of cembranoid-type tetracyclic diterpenes in terms of structure. They have a hydrophobic parent nucleus of steviol and a highly hydrophilic polyvalent sugar residue structure in terms of chemical structure. This polyvalent effect makes it easy for such compounds to spontaneously assemble into supramolecular nanocomplexes through synergistic interactions with other molecules, or to specifically recognize lectins in the body, and thus can be used as a nano delivery carrier in any delivery scenario.

[0005] In 2019, the first GalNAc-siRNA conjugate Givlaari successfully achieved liver-targeted delivery of siRNA, thanks to the RNA delivery technology modified by N-acetylgalactosamine (GalNac). Drawing on the successful experience of targeted delivery of GalNac-modified lipid nanoparticles, glycosylation-targeted delivery has become a global hotspot in the application of mRNA, tumor immunotherapy, and the preparation of sugar conjugate prodrugs for targeted therapy.

[0006] There are three ways to achieve targeted delivery by glycosylation: one is to connect glycoprotein on the nanocarrier, to achieve targeting by specific recognition of glycoprotein sugar residues on the cell surface lectin, but it may cause immune response and degradation risk; in addition, due to the large molecular weight of glycoprotein, the particle size distribution of the nanocarrier must be limited in consideration of the targeting efficiency, and the drug loading capacity is the main defect of this way. The second is to design a glycosylated prodrug of the drug molecule, and the drug molecule is covalently coupled with the sugar residue, and there is a high degree of uncertainty whether the prodrug molecule can release the original drug after reaching the target site, in addition, it involves complex synthesis steps. The third is to glycosylate the lipid or high molecular material, and then prepare a nanocarrier with other materials in a certain ratio, so as to guide the recognition of sugar residues to the surface of the nanocarrier, and improve the highly specific recognition ability of the nanocarrier, and avoid the immune response or original drug release problem of the former.

[0007] At present, the recognition sugar residues introduced by any of the above ways are finally delivered by traumatic administration, such as intravenous injection or subcutaneous injection, which reduces the patient's compliance for long-term treatment of chronic diseases, increases the economic and nursing burden of patients and doctors, and also increases the risk of cross infection.

[0008] The steviol glycoside compound is limited by the single type of sugar residue in the existing structure, and when it is used as a drug to trigger release, the intestinal flora may be unpredictably changed due to factors such as disease itself, surgery, medication (antibiotics) or dietary habits, thereby directly affecting the targeted release of drugs in the colon and most of the drugs are excreted outside the body through feces.

[0009] The present application aims to improve the targeting of steviol glycoside nanocarriers, and a series of derivatives are designed and synthesized, and through a large number of in vitro experiments and in vivo studies, steviol glycoside derivatives with excellent permeability and regulated by multiple flora are selected to achieve targeted delivery and release of active ingredients.

[0010] SUMMARY

[0011] The purpose of the present application is to improve the specific targeting recognition ability of steviol glycoside nanocarriers by modifying the structure of steviol glycoside compounds, while ensuring the safety, self-assembly ability, solubility and ultra-small nanometer size of the modified compounds. Another aspect of the present application is to design new compounds as nanocarrier materials, which can be triggered by enzymes produced by multiple microbial flora in vivo to realize online editing of steviol glycoside derivatives or delivery carriers, so that their permeability is also significantly improved after in situ assembly with active ingredients, realizing efficient and safe targeted therapy.

[0012] Terminology:

[0013] The term "targeting directing agent" refers to a compound or component that is capable of directing an active ingredient to a target site (e.g., a target tissue and / or organ, a cell, a cell surface lectin, a receptor, a gut flora, an enzyme, etc.). In some embodiments, the targeting directing agent can be prepared as a separate component in a kit (e.g., stored in a separate container) and mixed with the active ingredient at the time of use. In some embodiments, the targeting directing agent can be prepared as a mixture with the active ingredient stored in the same container in a kit. In the broadest sense, the targeting directing agent can also be part of a delivery system used as a delivery vehicle. In some embodiments, the steviol glycoside derivatives of the present application can be used as targeting directing agents.

[0014] The term "carrier material" or "adjuvant" or "functional carrier material" refers to a material that, upon assembly, aids or carries another substance(s) into a biological entity, which can affect the odor, solubility, stability, mucosal irritation, rate of hydration, release, absorption, tissue distribution, metabolism, or clearance of the delivered substance during delivery or in the biological entity. In some embodiments, the steviol glycoside derivatives of the present application can be used as carriers, or both as targeting directing agents and as carriers.

[0015] The term "enantiomeric excess" or "ee value" refers to a measure of how much of one enantiomer is present compared to the other enantiomer, and for a mixture of R and S enantiomers, the enantiomeric excess is calculated as ee value = ([R] - [S] / [R] + [S]) x 100%, where R and S are the respective molar or mass fractions of the enantiomers in the mixture.

[0016] The term "diastereomeric excess" or "de value" refers to a measure of how much of one diastereomer is present compared to the other diastereomer, and is defined similarly to enantiomeric excess. Thus, for a mixture of diastereomers D1 and D2, the diastereomeric excess is calculated as de value = ([D1] - [D2] / [D1] + [D2]) x 100%, where D1 and D2 are the respective molar or mass fractions of the diastereomers in the mixture.

[0017] It is possible to determine diastereomeric excess and / or enantiomeric excess using analytical techniques, including routine protocols of nuclear magnetic resonance spectroscopy, high performance liquid chromatography based on chiral analytical columns, supercritical fluid chromatography, optical rotatory measurements, etc., as would be understood by one skilled in the art.

[0018] The term "n-valent" rhamnose, rhamnopyranose, mannose, mannopyranose, galactose, galactopyranose refers to the total number of rhamnose, rhamnopyranose, mannose, mannopyranose, galactose, galactopyranose residues attached to the carrier hydrophobic backbone. In some embodiments, n can have a value of, for example, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or higher. The rhamnose, rhamnopyranose, mannose, mannopyranose, galactose, galactopyranose residues containing n-valent rhamnose, rhamnopyranose, mannose, mannopyranose, galactose, galactopyranose residues are targeting recognition groups. In some embodiments, the targeting recognition groups, once recognized, can facilitate the absorption, transport, release, or accumulation of the active ingredient at the target site (e.g., target tissue and / or organ), thereby enabling differential pharmacokinetic profiles and more effective treatment and / or more precise diagnosis of the disease.

[0019] The term "steviol glycoside derivative" can refer to a substance obtained by structural modification of a steviol glycoside compound and containing a targeting recognition group. In some embodiments, the structural modification can include, for example, glycosylation modification. In some embodiments, the steviol glycoside derivative obtained by structural modification advantageously has one or more desired activities or advantages, which can be identified by, for example, the effect of the steviol glycoside derivative obtained after including the glycosylation modification on the active ingredient. In some embodiments, the effect can include, for example, preparing the obtained steviol glycoside derivative into a composition or complex (e.g., nanocomplex) with the active ingredient, and then selecting the steviol glycoside derivative that preferably shows any one or more of the following advantages compared to a control (e.g., a composition or complex containing the same active ingredient constructed without the substance containing the targeting recognition group): being able to be specifically delivered to a specific target site (e.g., target tissue and / or organ where the effect is exerted) or showing a higher drug concentration or drug exposure at a specific target site (e.g., target tissue and / or organ where the effect is exerted) compared to a non-target site, being able to achieve multiple targeted delivery, showing improved permeability, solubility, stability, drug loading, encapsulation efficiency, delivery system reliability, pharmacokinetic / pharmacodynamic specificity, and / or showing differential pharmacokinetic profiles, showing reduced administration dose, first-pass metabolism, intrinsic cytotoxicity, systemic toxicity, and / or non-target organ toxicity, etc. The term "hyperbranched" refers to a class of highly branched three-dimensional molecules, which have a branched structure that makes them have abundant terminal functional groups, are easy to modify, and are more conducive to the formation of functional materials of self-assembled complex objects between molecules through non-covalent bonds. Such molecules can be small molecules or high molecular weight polymers.

[0020] The term "supramolecular nanocomplex" can refer to a multi-molecular group assembled by non-covalent interaction of various components, preferably with a particle size in the range of 1-500 nm, more preferably in the range of 1-100 nm, for example 1 nm, 5 nm, 10 nm, 20 nm, 30 nm, 40 nm, 50 nm, 60 nm, 70 nm, 80 nm, 90 nm, 100 nm, 110 nm, 120 nm, 130 nm, 140 nm, 150 nm, 160 nm, 170 nm, 180 nm, 190 nm, 200 nm, 250 nm, 300 nm, 350 nm, 400 nm, 450 nm, 500 nm, or any range therebetween. The steviol glycoside derivative of the present application can be used to construct a supramolecular nanocomplex with active ingredients, which preferably has a hyperbranched structure, and is more physically and chemically stable. When part of the group is metabolized by enzymes, the intermediate of the carrier after metabolism still has a hyperbranched structure, and can quickly reassemble with active ingredients, i.e. has the functions of online editing and self-repairing. In some embodiments, the supramolecular nanocomplex of the present application preferably has one or more advantages compared to a control (e.g. a composition not comprising the steviol glycoside derivative of the present application or a commercially available product containing the same active ingredient). In some embodiments, the properties and / or advantages of the supramolecular nanocomplex can be determined by comparing one or more aspects such as drug loading concentration, nanocomplex properties, particle size, PDI, Zeta potential, stability, dissolution characteristics, tissue distribution, therapeutic activity (e.g. tumor therapeutic activity) and side effects (e.g. systemic toxicity). In some embodiments, the different components in the supramolecular nanocomplex of the present application can exhibit a synergistic effect that is far superior to the sum of the performance of each single component.

[0021] The technical solution of the present application is as follows:

[0022] 1. A steviol glycoside derivative characterized by a compound represented by general formula Ia, or a salt thereof, or a hydrate thereof, or any mixture of the compound represented by general formula Ia, the salt thereof and the hydrate thereof,

[0023] In formula Ia, R1 and R2 are independent, R1 is selected from hydroxyl or glycosyl, and R2 is selected from n-valent rhamnose, rhamnopyranose, mannose, mannopyranose, galactose, galactopyranose, or a combination thereof, or an oligosaccharide formed by covalent linkage of other sugars, wherein 1≤n≤10, for example n is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10.

[0024] 2. The steviol glycoside derivative according to item 1, characterized in that the absolute configuration of the rhamnose, rhamnopyranose, mannose, mannopyranose, galactose, galactopyranose is D or L, and the relative configuration of the terminal carbon is α or β.

[0025] 3. The steviol glycoside derivative according to any one of items 1-2, characterized in that the absolute configuration of the rhamnose is preferably L-form, the relative configuration of the terminal carbon is preferably a-form; the absolute configuration of the mannose is D-form, the relative configuration of the terminal carbon is β-form; the absolute configuration of the galactose is preferably D-form, the relative configuration of the terminal carbon is β-form.

[0026] 4. The steviol glycoside derivative according to any one of items 1-3, characterized in that the glycosyl group is selected from glucopyranose, galactopyranose, mannopyranose, deoxyglucopyranose, rhamnopyranose, xylopyranose, glucosaminopyranose, oligosaccharide of 1 to 9 arbitrary number of arbitrary pyranose monosaccharides polymerized at any position.

[0027] 5. The steviol glycoside derivative according to any one of items 1-4, characterized in that the absolute configuration of the glycosyl group is D-form or L-form, the relative configuration of the terminal carbon is a-form or β-form.

[0028] 6. The steviol glycoside derivative according to any one of items 1-5, characterized in that R1 is selected from hydroxyl,

[0029] 7. The steviol glycoside derivative according to any one of items 1-6, characterized in that the optical purity of the compound represented by the general formula la is not limited, R and / or S configuration, with any enantiomeric excess value or any diastereomeric excess value.

[0030] 8. The steviol glycoside derivative is selected from at least one compound in the group consisting of the following, or a salt thereof, or a hydrate thereof, or an in vivo metabolite thereof:

[0031] (1) (2S, 3S, 4S, 5S, 6R)-3, 4, 5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R, 4aS, 6aR, 9R, 11aR, 11bS)-9-hydroxy-4, 11-dimethyltetradecahydro-6a, 9- methanocyclohepta[a]naphthalene-4-carboxylate (STL-Man),

[0032] (2) (2S, 3S, 4S, 5S, 6R)-3, 4, 5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R, 4aS, 6aR, 9S, 11aR, 11bS)-4, 11b-dimethyl-8-methylene-9-(((2S, 3R, 4S, 5S, 6R)-3, 4, 5-trihydroxy-6- (hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetradecahydro-6a, 9- methanocyclohepta[a]naphthalene-4-carboxylate (RBSD-Acid-Man),

[0033] (3) (2S,3S,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-((2S,3R,4S,5S,6R)-4,5-dihydroxy-6-(hydroxymethyl)-3-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetrahydro-1 H-pyran-2-yl)oxy)-4,11 b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohepta[a]naphthalene-4-carboxylate (STVB-Man),

[0034] (4) (2S,3S,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-((2S,3R,4S,5S,6R)-4,5-dihydroxy-6-(hydroxymethyl)-3-(((2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2-yl)oxy)-4,11 b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohepta[a]naphthalene-4-carboxylate (DCS-A-Acid-Man),

[0035] (5) (2S,3S,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9R,11aR,11bS)-9-((2S,3R,4S,5R,6R)-5-hydroxy-6-(hydroxymethyl)-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-3-(((2S,3R,4S,5R)-3,4,5-trihydroxytetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2-yl)oxy)-4,11 b-dimethyltetradecahydro-6a,9-methanocyclohepta[a]naphthalene-4-carboxylate (RBDS-F-Acid-Man),

[0036] (6) (2S,3S,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-(((2S,3R,4S,5R,6R)-5-hydroxy-6-(hydroxymethyl)-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-3-(((2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2-yl)oxy)-4,11b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclo- hepta[a]naphthalene-4-carboxylate (DCS-B-Man),

[0037] (7) (2S,3S,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-(((2S,3R,4S,5R,6R)-3-(((2S,3R,4R,5S,6S)-3,5-dihydroxy-6-methyl-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2-yl)oxy)-5-hydroxy-6-(hydroxymethyl)-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetrahydro-1H-pyran-2-yl)oxy)-4,11b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclo- hepta[a]naphthalene-4-carboxylate (RBDS-H-Acid-Man),

[0038] (8) (2S,3S,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-(((2S,3R,4S,5R,6R)-5-hydroxy-6-(hydroxymethyl)-3,4-bis((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetrahydro-1H-pyran-2-yl)oxy)-4,11b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclo- hepta[a]naphthalene-4-carboxylate (RBDS-B-Man),

[0039] (9) a steviol glycoside-Man derivative;

[0040] (10) (2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9R,11aR,11bS)-9-hydroxy-4,11-dimethyltetradecahydro-6a,9- methylenocyclohepta[a]naphthalene-4-carboxylate (STL-Rha),

[0041] (11) (2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-4,11b-dimethyl-8-methylene-9-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6- (hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetradecahydro-6a,9- methylenocyclohepta[a]naphthalene-4-carboxylate (RBSD-Acid-Rha),

[0042] (12) (2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-(((2S,3R,4S,5S,6R)-4,5-dihydroxy-6-(hydroxymethyl)-3-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetrahydro-1H-pyran-2-yl)oxy)-4,11b-dimethyl-8-methylene tetradecahydro-6a,9-methylenecyclohepta[a]naphthalene-4-carboxylate (STVB-Rha),

[0043] (13) (2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-(((2S,3R,4S,5S,6R)-4,5-dihydroxy-6-(hydroxymethyl)-3-(((2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2-yl)oxy)-4,11b-dimethyl-8-methylenetetradecahydro-6a,9-methylenecyclohepta[a]naphthalene-4-carboxylate (DCS-A-Acid-Rha)

[0044] (14) (2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-(((2S,3R,4S,5R,6R)-5-hydroxy-6-(hydroxymethyl)-3,4-bis((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetrahydro-1 H-pyran-2-yl)oxy)-4,11 b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohexa[a]naphthalene-4-carboxylate (RBDS-B-Rha),

[0045] (15) (2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-(((2S,3R,4S,5R,6R)-5-hydroxy-6-(hydroxymethyl)-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-3-(((2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2-yl)oxy)-4,11 b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohexa[a]naphthalene-4-carboxylate (DCS-B-Rha),

[0046] (16) (2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9R,11aR,11bS)-9-((2S,3R,4S,5R,6R)-5-hydroxy-6-(hydroxymethyl)-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-3-(((2S,3R,4S,5R)-3,4,5-trihydroxytetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2-yl oxy)-4,11 b-dimethyltetradecahydro-6a,9-methanocyclohexa[a]naphthalene-4-carboxylate (RBDS-F-Acid-Rha),

[0047] (17) (2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-(((2S,3R,4S,5R,6R)-3-(((2S,3R,4R,5S,6S)-3,5-dihydroxy-6-methyl-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetrahydro-1H-pyran-2-yl)oxy)-5-hydroxy-6-(hydroxymethyl)-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetrahydro-1H-pyran-2-yl)oxy)-4,11b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohexa[a]naphthalene-4-carboxylate (RBDS-H-Acid-Rha),

[0048] (18) a steviol-Rha derivative,

[0049] (19) (2S,3R,4S,5R,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-hydroxy-4,11b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohexa[a]naphthalene-4-carboxylate (STL-Gal),

[0050] (20) (2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-4,11b-dimethyl-8-methylenetetradecahydro-9-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy) -6a,9-methanocyclohexa[a]naphthalene-4-carboxylate (RBSD-Acid-Gal),

[0051] (21) (2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-(((2S,3R,4S,5S,6R)-4,5-dihydroxy-6-(hydroxymethyl)-3-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetrahydro-1H-pyran-2-yl)oxy)-4,11b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohexa[a]naphthalene-4-carboxylate (STVB-Gal),

[0052] (22) (2S,3R,4S,5R,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2- yl (4R,4aS,6aR,9S,11aR,11bS)-9-((2S,3R,4S,5R,6R)-5-hydroxy-6- (hydroxymethyl)-3,4-bis((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6- (hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetrahydro-1H-pyran-2- yl)oxy)-4,11b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohepta[a]naphthalene-4-carboxylate (RBDS-B-Gal),

[0053] (23) (2S,3R,4S,5R)-3,4,5-trihydroxytetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-(((2S,3R,4S,5R,6R)-5-hydroxy-6-(hydroxymethyl)-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-3-(((2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2-yl)oxy)-4,11b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohepta[a]naphthalene-4-carboxylate (DCS-A-Acid-Gal),

[0054] (24) (2S,3R,4S,5R,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2- yl (4R,4aS,6aR,9S,11aR,11bS)-9-((2S,3R,4S,5R,6R)-5-hydroxy-6- (hydroxymethyl)-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6- (hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-3-(((2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2- yl)oxy)-4,11b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohepta[a]naphthalene-4-carboxylate (DCS-B-Gal),

[0055] (25) (2S,3R,4S,5R,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-(((2S,3R,4S,5R,6R)-5-hydroxy-6-(hydroxymethyl)-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-3-(((2S,3R,4S,5R)-3,4,5-trihydroxytetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2-yl)oxy)-4,11b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohexa[a]naphthalene-4-carboxylate (RBDS-F-Acid-Gal),

[0056] (26) (2S,3R,4S,5R,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-(((2S,3R,4S,5R,6R)-3-(((2S,3R,4R,5S,6S)-3,5-dihydroxy-6-methyl-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2-yl)oxy)-5-hydroxy-6-(hydroxymethyl)-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetrahydro-1H-pyran-2-yl)oxy)-4,11b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohexa[a]naphthalene-4-carboxylate (RBDS-H-Acid-Gal),

[0057] (27) a derivative of steviol glycoside-Gal.

[0058] 9. A method for preparing the steviol glycoside derivative as described in any one of items 1-8, characterized by comprising the following steps:

[0059] (1) Step 1: subjecting steviol glycoside compound 1 or steviol to a hydrolysis reaction to obtain compound 2 or a combination, wherein when compound 1 is in the form of a carboxylic acid, directly proceed to Step 2;

[0060] (2) Step 2: protecting the R1 group in compound 2 or a combination with a protecting group P to obtain compound 3 or a combination;

[0061] (3) Step 3: subjecting the activated sugar precursor to glycosidation reaction with compound 3 or a combination to obtain compound 4 or a combination; the activated sugar precursor is preferably a glycosyl halide, a glycosyl halide imidate ester;

[0062] (4) Step 4: deprotection reaction of compound 4 or combination to obtain a compound or combination of compounds of structural formula 1a;

[0063] wherein the steviol glycoside compound 1 is a compound of the general formula la as defined in items 1-7, or is R1in the aforementioned formula 2 is selected from hydroxyl, or combination.

[0064] 10. The preparation method according to item 9, wherein the hydrolysis reaction is a reaction in a solvent (preferably water) under basic conditions (preferably sodium hydroxide or potassium hydroxide or lithium hydroxide), wherein compound 1 is selected from stevioside (STVS), rebaudioside E (RBDS-E), rebaudioside A (RBDS-A), dulcoside A (DCS-A), rubusoside (RBSD), rubusoside acid (RBSD-Acid), steviolbioside (STVB), rebaudioside B / D / M / I / O / N / J (RBDS-B / D / M / O / N / J), rebaudioside C / K (RBDS-C / K), dulcoside B (DCS-B), rebaudioside F (RBDS-F), rebaudioside H (RBDS-H), steviol (STL), or isosteviol (ISTL), or a combination of any one or more thereof; when compound 1 is steviolbioside (STVB), rebaudioside B (RBDS-B), dulcoside B (DCS-B), or steviol (STL) any one or more thereof, the preparation method does not require step 1.

[0065] 11. The method of any one of claims 9-10, wherein the protecting group in step 2 is a protecting group for a sugar or a hydroxyl group, preferably a silyl group or an acetyl group, preferably the sugar or hydroxyl group is fully acetylated, for example the compound 3 after protection is selected from RBDS-B-Ac, STVB-Ac, RBSD-Acid-Ac, DCS-A-Acid-Ac, RBDS-F-Acid-Ac, RBDS-H-Acid-Ac, DCS-B-Ac, or STL-Ac, or a combination thereof, and the structures are shown below,

[0066] wherein Ac represents an acetyl group.

[0067] 12. The method of any one of claims 9-11, wherein the glycosylation reaction in step 3 is a reaction of compound 2 with an activated sugar precursor of the same structure as defined for R2 to obtain compound 4, and the glycosylation reaction is performed under conditions known in the art; preferably the activated sugar precursor is a glycosyl halide, a glycosyl halogenated imidazolide ester.

[0068] 13. The method of any one of claims 9-12, wherein when the protecting group P is an acetyl group, the step 4 is a deprotection reaction using an alcohol solvent (such as methanol or ethanol) under a strong base condition (such as potassium tert-butoxide or sodium methoxide or sodium ethoxide).

[0069] 14. A pharmaceutical composition or delivery kit comprising the steviol glycoside derivative of any one of claims 1-8, a delivery vehicle, and an active ingredient.

[0070] 15. The pharmaceutical composition or delivery kit of claim 14, wherein the active ingredient is a bioactive agent, a chemical active agent, or an adjuvant.

[0071] 16. The pharmaceutical composition or delivery kit of claim 14 or 15, wherein the active ingredient is selected from the group consisting of a protein / polypeptide, a polysaccharide / oligosaccharide, a nucleic acid or nucleic acid fragment, a nanoparticle (such as a liposome, a supramolecular nanocomplex, a polymeric micelle, an inorganic nanoparticle), a lipid, a nutritional element, an organic small molecule compound, a phage particle, a superparamagnetic substance, a vaccine, a cell, or any combination thereof.

[0072] 17. The pharmaceutical composition or delivery kit of any one of claims 14-16, wherein the delivery vehicle is selected from the group consisting of one or more of a lipid, a phosphatidylcholine, a cholesterol and derivatives thereof, a glycoside and derivatives thereof, a soy protein, an apolipoprotein, an inorganic nanocarrier, a polymer, a glycolipid.

[0073] 18. The pharmaceutical composition or delivery kit according to any one of items 14-17, wherein the mass ratio of the active ingredient to the sum of (steviol glycoside derivative + delivery vehicle) is 1 :0.5 to 1 :25 (preferably 1 :0.75, 1 :1, 1 :1.5, 1 :2, 1 :2.5, 1 :3, 1 :5, 1 :10, 1 :15, 1 :20, 1 :25).

[0074] 19. The pharmaceutical composition or delivery kit according to any one of items 14-18, wherein the steviol glycoside derivative comprises 5% to 100% of the total mass of (steviol glycoside derivative + delivery vehicle), preferably 7% to 100% (preferably 7% to 70%, 7% to 65%, 10% to 60%, 12.5 to 100%, 15% to 95%, 17.5% to 90%, 20% to 80%, 25% to 75%, 30% to 70%, 35% to 65%, 40% to 60%).

[0075] 20. The pharmaceutical composition or delivery kit according to any one of items 14-19, wherein the active ingredient, steviol glycoside derivative and delivery vehicle are present independently or are mixed together.

[0076] 21. The pharmaceutical composition or delivery kit according to any one of items 14-20, wherein the active ingredient, steviol glycoside derivative and delivery vehicle are in a form suitable for oral, sublingual, buccal, mucosal (e.g. esophageal, gastric, intraduodenal, colonic, rectal, vaginal, cervical, anal), subcutaneous, intramuscular, intravenous, intraarterial, dermal, pulmonary, pharyngeal, tracheal, bronchial, intranasal, otic, ocular administration.

[0077] 22. The pharmaceutical composition or delivery kit according to any one of items 14-21, wherein the active ingredient, steviol glycoside derivative and / or the delivery vehicle are in the form of micelles, liposomes, nanoparticles, microspheres, microcapsules, solid dispersions, molecular compositions or hydrogels.

[0078] 23. The pharmaceutical composition or delivery kit according to any one of items 14-22, wherein when the active ingredient, steviol glycoside derivative and delivery vehicle are present independently, they are assembled by mixing and dissolving at the time of use; when the active ingredient, steviol glycoside derivative and delivery vehicle are mixed together, the active ingredient, targeting agent and delivery vehicle in solid form are assembled by dissolving.

[0079] 24. The pharmaceutical composition or delivery kit according to any one of items 14-23, wherein the assembly of the active ingredient, steviol glycoside derivative and delivery vehicle in the delivery system is in situ assembly.

[0080] 25. The pharmaceutical composition or delivery kit according to item 24, wherein the in situ assembly is self-assembly in the gastrointestinal tract, self-assembly in the mucosa, self-assembly in the blood, self-assembly in the lymphatic vessels or lymph nodes, self-assembly in the intrathecal space or self-assembly in target cells.

[0081] 26. The pharmaceutical composition or delivery kit according to any one of items 24-25, wherein the in situ assembly is the in situ assembly of the steviol glycoside derivative according to any one of items 1-8 with a metabolite of an enzyme, a microorganism metabolism and the active ingredient and / or the delivery vehicle or a metabolite thereof.

[0082] 27. The pharmaceutical composition or delivery kit according to any one of items 14-26, wherein the active ingredient and / or the steviol glycoside derivative and / or the delivery vehicle is in a dosage form selected from the group consisting of tablets, capsules, oral solutions, drops, gels, granules, emulsions, creams, injections, eye drops, inhalants, sprays, aerosols or patches.

[0083] 28. The pharmaceutical composition or delivery kit according to any one of items 14-27, wherein the delivery system further comprises a polymer, preferably the polymer has groups that are dissociable under physiological conditions and / or has more than 10 groups that provide hydrogen donors or hydrogen acceptors, more preferably the polymer is selected from the group consisting of:

[0084] (1) Cellulose-based polymers such as hydroxypropyl methylcellulose (HPMC), low-substituted hydroxypropyl cellulose (L-HPC), hydroxypropyl methylcellulose acetate succinate (HPMCAS), sodium carboxymethylcellulose (CMC-Na) or hydroxyethyl cellulose;

[0085] (2) Synthetic polymers such as polyethylene glycol (PEG), polyvinyl alcohol (PVA), polyvinylpyrrolidone (PVP), copolyvidone (PVP-VA64), polyvinyl acetate-polyethylene glycol graft copolymer (Soluplus), polyglutamic acid (PGA), polydopamine (PDA), polylactic acid (PLA), poly-L-lactic acid (PLLA), poly(lactic-co-glycolic acid) (PLGA), sodium polyacrylate, siRNA and derivatives (alkyl chains to improve lipophilicity), mRNA and derivatives (alkyl chains to improve lipophilicity) or antisense oligonucleotides and derivatives (alkyl chains to improve lipophilicity);

[0086] (3) Polysaccharides and derivatives thereof (such as carboxymethylation, sulfonylation, phosphorylation, acylation or hydroxypropylation, cationic, anionic or other derivatives), such as carrageenan, guar gum, gum arabic, locust bean gum, konjac gum, agar, gelatin, pectin, gellan gum, hyaluronic acid (HA), aminodextran, chondroitin sulfate, dermatan sulfate, heparin, keratan sulfate, heparan sulfate, sodium alginate, propylene glycol alginate, agar, fucoidan, cyclodextrin and derivatives, chitosan and derivatives (such as acylation, carboxylation, alkylation and quaternization, etc.); and

[0087] (4) Proteins and polypeptides: soy protein, vegetable protein or bovine serum albumin, casein; and

[0088] (5) Any combination of (1)-(4).

[0089] 29. The pharmaceutical composition or delivery kit according to any one of items 14-28, wherein the delivery system further comprises an additive selected from the group consisting of an excipient, a diluent, a binder, a disintegrant, a lubricant, a flavoring agent, a pH adjusting agent, an osmotic pressure adjusting agent, a thickening agent, a plasticizer, a coloring agent, a film-forming agent, a preservative or a solvent, and any combination thereof.

[0090] 30. The pharmaceutical composition or delivery kit according to any one of items 14-29, for use selected from the group consisting of:

[0091] (1) for the delivery of an active ingredient to a mammal (such as a rodent, a bovine, a porcine, a canine, a feline, a primate, a human) or a bird (such as a chicken, a duck, a goose);

[0092] (2) for the modulation of the release of an active ingredient or a nutritional ingredient during the delivery of the active ingredient or the nutritional ingredient to a mammal (such as a rodent, a bovine, a porcine, a canine, a feline, a primate, a human) or a bird (such as a chicken, a duck, a goose);

[0093] (3) for use as a solubilizer in agrochemistry, especially in formulation products comprising a pesticide, a herbicide, a fungicide or an insecticide, especially as a solubilizer for spray, drench or dip mixtures of crop protection agents;

[0094] (4) for use as a preservative or a preservative or a flavoring agent for meat, vegetables, fruits, food, beverages;

[0095] (5) for use in cosmetics, fine chemical products for solubilization, stability enhancement, penetration promotion;

[0096] (6) for use as an adjuvant for biological agents.

[0097] In some embodiments, R1in structural formula Ia can be the corresponding sugar residue already present in steviol glycosides, for example, as shown in Table 1 below:

[0098] Table 1. R1groups in structural formula Ia are sugar groups listed in the table

[0099] In some embodiments, the steviol glycoside derivatives and in vivo metabolites that can act as carriers and / or targeting directing agents are selected from one or more combinations of the following compounds in Table 2:

[0100] Table 2. Steviol glycoside derivatives and in vivo metabolites as carrier materials

[0101] The steviol glycoside derivatives that can act as carriers and / or targeting directing agents described herein can be a single compound having the structure of formula Ia and / or hydrates and / or salts thereof, or a mixture of multiple compounds having the structure of formula Ia in any ratio and / or hydrates thereof.

[0102] The steviol glycoside derivatives that can act as carriers and / or targeting directing agents described herein, the compounds having the structure of formula Ia are not limited in optical purity, the compounds having the structure of formula Ia have any enantiomeric excess value or any diastereomeric excess value.

[0103] In another aspect of the present application, a method for preparing a compound having the structure of formula Ia is disclosed, the method comprising the following steps:

[0104] (1) Step 1: subjecting a steviol glycoside compound 1 or a steviol (combination 1) to a hydrolysis reaction to obtain a compound 2 or a combination 2, wherein when the compound 1 is in the form of a carboxylic acid, directly proceed to Step 2;

[0105] (2) Step 2: protecting the R1group in the compound 2 or the combination 2 with a protecting group P to obtain a compound 3 or a combination 3;

[0106] (3) Step 3: subjecting the compound 3 or the combination 3 to a glycosylation reaction with an activated sugar precursor to obtain a compound 4 or a combination 4; the activated sugar precursor is preferably a sugar halide, a sugar haloimide ester;

[0107] (4) Step 4: subjecting the compound 4 or the combination 4 to a deprotection reaction to obtain a compound or a combination of multiple compound derivatives having the structure of formula Ia.

[0108] wherein R1or R2in the compound 1, the compound 2, the compound 3 and the compound 4 are as defined in the compound having the structure of formula Ia.

[0109] wherein step 1 employs steviol glycosides (combination 1), combination 2 is one or more combinations of STL, RBSD-Acid, STVB, RBDS-B, DCS-A-Acid, DCS-B, RBDS-F-Acid, and RBDS-H-Acid, etc.; combination 3 is a combination of acetylated products of combination 2; and combination 4 is a combination of mannose esters or galactose esters or rhamnose esters of combination 3.

[0110] The specific reaction scheme is as follows:

[0111] Without limitation, step one is a hydrolysis step. Most known steviol glycosides contain a glycosidic bond in their structure, for example, the commercially available steviol glycosides stevioside and rebaudioside A (RBDS-A) or dulcoside A (DCS-A) or rubusoside (RBSD) have the following structures. To obtain the compound of Formula la described in the present application, the steviol glycoside containing a glycosidic bond needs to be subjected to a hydrolysis reaction.

[0112] If the skilled person has obtained a steviol glycoside that is a carboxylic acid structure, for example, steviolbioside (STVB), rebaudioside B (RBDS-B), dulcoside B (DCS-B), steviol, rubusoside acid form (RBSD-Acid), rebaudioside H acid form (RBDS-H-Acid), and rebaudioside F acid form (RBDS-F-Acid) have the following structures, then this step of hydrolysis reaction is not required.

[0113] The selection of the hydrolysis conditions can be performed according to known ester hydrolysis conditions in the art, for example, sodium hydroxide or potassium hydroxide can be used to perform the reaction in water.

[0114] Without limitation, step two is a protection step. The selection of the protecting group can be performed according to known suitable protecting groups for protecting sugars in the art, for example, silyl groups, acetyl groups, etc. In some embodiments, the sugar group is fully acetylated, for example, RBDS-B-Ac, STVB-Ac, RBSD-Acid-Ac, DCS-A-Acid-Ac, STL-Ac, DCS-B-Ac, RBDS-F-Acid-Ac, and RBDS-H-Acid-Ac, as shown in the following structures,

[0115] The conditions for adding the protecting group can be carried out according to the known techniques in the art, for example, the conditions for adding acetyl group, using acetic anhydride, sodium acetate, and carrying out the reaction under heating conditions.

[0116] Non-limitatively, step three is a condensation step, which requires compound 2 to react with the sugar carrying R2 group to form a glycoside. The conditions for the glycosylation reaction can be carried out according to the known techniques in the art.

[0117] Non-limitatively, step four is a deprotection step, and the conditions for removing the protecting group can be carried out according to the known techniques in the art, for example, the conditions for removing acetyl group, using alcoholic solvents such as methanol or ethanol, and carrying out the deprotection reaction in the presence of strong bases such as potassium tert-butoxide or sodium methoxide or sodium ethoxide.

[0118] The steviol glycoside derivative described in the present application can be used as a targeting guide and as a delivery carrier at the same time.

[0119] The active ingredient delivery system containing the steviol glycoside derivative of the present application can more reliably deliver the active ingredient to a specific site than the delivery system constructed by other delivery carriers, and through the specific recognition of different sugar residues by various intestinal flora, triggers the in-line editing or in-situ assembly of the steviol glycoside derivative or the delivery carrier, promotes the absorption and transport of the drug in the target organ / tissue, thereby achieving the treatment and diagnosis of diseases, and thus can provide differentiated pharmacokinetic characteristics to meet the treatment or prevention of different diseases.

[0120] The active substance delivery system containing the steviol glycoside derivative of the present application has the effect of delivering the active agent to the desired biological system, target area, and can improve the utilization rate of the active agent compared with the delivery system without using the steviol glycoside derivative. The delivery can be improved by delivering more active agent to the target area over a period of time or delivering the active agent to the target area at a specific time period (such as faster action or delayed delivery) or over a period of time (such as continuous delivery).

[0121] In some embodiments, the additive is a pharmaceutically acceptable additive, such as those described in Remington: The Science and Practiice of Pharmacy (Adeboye Adejare, 23rd edition, 2020, Academic Press), Pharmaceutical Formulation (Fang Liang, 8th edition, 2016, People's Medical Publishing House), which are incorporated herein by reference.

[0122] Advantages of the invention

[0123] Compared with the existing steviol glycoside compounds, the steviol glycoside derivatives provided by the present application have multiple specific target recognition groups or multiple intestinal flora trigger groups, and can carry active ingredients to target tissues / organs or cells after being used alone or assembled into a targeted nanocomplex with other carriers, so that targeted treatment or precise treatment is realized.

[0124] After the steviol glycoside derivatives provided by the present application are assembled into nanoparticles with active ingredients and orally taken, the nanoparticles can tolerate the influence of complex contents or extreme pH in the gastrointestinal tract, are transported to the lower half of the intestinal tract, are edited online and assembled in situ by means of intestinal microflora or enzymes generated by the intestinal microflora, are changed from nanocomplexes with low permeability into nanocomplexes with high permeability, and the permeation and absorption of drugs in the rectum are promoted.

[0125] The nanocomplexes assembled by the steviol glycoside derivatives provided by the present application, active ingredients and other carriers have the superiority of ultra-small size, and the nanocomplexes after assembly are more likely to realize targeted delivery of the lymphatic system, so that the major clinical needs of existing autoimmune diseases, organ transplantation anti-rejection treatment, malignant tumor lymphatic metastasis, anti-infection, anti-chronic inflammation, inflammatory bowel disease, tumor immunotherapy, systemic toxicity caused by systemic administration through intravenous injection, poor curative effect, and many complications are solved.

[0126] Compared with lipid nanoparticles, inorganic nanocarriers and polymer nanocarriers, the nanocomplexes constructed by the steviol glycoside derivatives provided by the present application only need simple synthesis steps, solve the pain points of low encapsulation efficiency and low drug loading of other nanocarriers, intrinsic cytotoxicity, inability to tolerate the complex environmental effects of the gastrointestinal tract, difficulty in controlling particle size distribution, and industrialization, and have broad application prospects and clinical transformation value. BRIEF DESCRIPTION OF DRAWINGS

[0127] FIG. 1 is a mass spectrum diagram of a STVB intermediate prepared in Example 1 in LC-MS / MS negative ion mode;

[0128] FIG. 2 is an H-NMR spectrum of a STVB intermediate prepared in Example 1; 1

[0129] FIG. 3 is a mass spectrum diagram of a STVB-Ac intermediate prepared in Example 1 in LC-MS / MS negative ion mode;

[0130] FIG. 4 is an H-NMR spectrum of a Man-4 intermediate prepared in Example 1; 1

[0131] FIG. 5 is a mass spectrum diagram of a STVB-Man compound prepared in Example 1 in LC-MS / MS negative ion mode;

[0132] FIG. 6 is an H-NMR spectrum of a STVB-Man compound prepared in Example 1; 1 ​​H-NMR spectrum;

[0133] Figure 7 LC-MS / MS mass spectrum of RBDS-B intermediate prepared in Example 2 in negative ion mode;

[0134] Figure 8 H-NMR spectrum of RBDS-B intermediate prepared in Example 2; 1 H-NMR spectrum;

[0135] Figure 9 LC-MS / MS mass spectrum of RBDS-B-Ac intermediate prepared in Example 2 in negative ion mode;

[0136] Figure 10 H-NMR spectrum of Rha-3 intermediate prepared in Example 2; 1 H-NMR spectrum;

[0137] Figure 11 LC-MS / MS mass spectrum of RBDS-B-Rha compound prepared in Example 2 in negative ion mode;

[0138] Figure 12 H-NMR spectrum of RBDS-B-Rha compound prepared in Example 2; 1 H-NMR spectrum;

[0139] Figure 13 LC-MS / MS mass spectrum of DCS-A-Acid intermediate prepared in Example 3 in negative ion mode;

[0140] Figure 14 H-NMR spectrum of DCS-A-Acid intermediate prepared in Example 3; 1 H-NMR spectrum;

[0141] Figure 15 LC-MS / MS mass spectrum of DCS-A-Acid-Ac intermediate prepared in Example 3 in negative ion mode;

[0142] Figure 16 H-NMR spectrum of Gal-2 intermediate prepared in Example 3; 1 H-NMR spectrum;

[0143] Figure 17 LC-MS / MS mass spectrum of DCS-A-Acid-Gal prepared in Example 3 in negative ion mode;

[0144] Figure 18 H-NMR spectrum of DCS-A-Acid-Gal compound prepared in Example 3; 1 H-NMR spectrum;

[0145] Figure 19 LC-MS / MS mass spectrum of DCS-B-Man compound prepared in Example 4 in negative ion mode;

[0146] Figure 20 H-NMR spectrum of DCS-B-Man compound prepared in Example 4; 1 H-NMR spectrum;

[0147] Figure 21 Mass spectrum of the DCS-B-Rha compound prepared in Example 5 in negative ion mode by LC-MS / MS;

[0148] Figure 22. DCS-B-Rha compound prepared in Example 5 1 H-NMR spectrum;

[0149] Figure 23 Mass spectrum of the STVB-Gal compound prepared in Example 6 in negative ion mode by LC-MS / MS;

[0150] Figure 24. STVB-Gal compound prepared in Example 6 1 H-NMR spectrum;

[0151] Figure 25 shows the cumulative release rates of Comparative Example 1 and F1-F5 in 80% human plasma;

[0152] Figure 26 Comparative 2DSC plots;

[0153] Figure 27 Carrier metabolic pathway 1;

[0154] Figure 28. Carrier metabolic pathway 2. Detailed Implementation

[0155] There are no particular restrictions on the raw materials, reagents, and solvents used in this invention; commercially available conventional raw materials, reagents, and solvents can be used.

[0156] Instruments and methods used for data collection:

[0157] Acquisition of nuclear magnetic resonance hydrogen spectrum (NMR) 1 The HNMR data were obtained using a Bruker AVANCE 600 instrument with a resonance frequency of 600 MHz and deuterated DMSO as the solvent.

[0158] The high-resolution mass spectrometry data involved in this invention were detected using a Waters Xevo TQ-S triple quadrupole mass spectrometer, and the liquid chromatography conditions were as follows:

[0159] The chromatographic column was an Agilent ZOBAX SB-C18, 2.1 × 50 mm, 1.8 μm;

[0160] Mobile phase A: 0.1% formic acid-water solution; Mobile phase B: 0.08% formic acid-acetonitrile solution;

[0161] Detection wavelength: 210nm;

[0162] Flow rate: 0.7 mL / min;

[0163] Injection volume: 5 μL;

[0164] Column temperature: 60℃;

[0165] Table 3. Gradient elution program for detection of free acid intermediates / derivatized products

[0166] Table 4. Gradient elution program for acetylated intermediates

[0167] The mass spectrometry detection conditions were: capillary voltage: 2.80 KV; drying gas temperature 350℃; positive / negative ion scan mode, ion source was ESI source.

[0168] The purity of each derivative was detected by high performance liquid chromatography, and the chromatographic conditions were as follows in Table 5:

[0169] Table 5. Chromatographic conditions

[0170] The main peak purity was calculated by principal component self-control method.

[0171] It should be noted that the meaning or significance of the numerical values or numerical endpoints involved in the technical solutions of the present application is not limited to the numbers themselves, and those skilled in the art can understand that they include the allowable error range widely accepted in the art, such as experimental error, measurement error, statistical error and random error, etc., and these error ranges are included in the scope of the present application.

[0172] In order to further illustrate the present application, specific examples will be described below, but the following examples do not constitute any limitation on the scope of protection of the present application.

[0173] Example 1. Preparation of STVB-Man

[0174] (1) Synthesis of STVB-Ac

[0175] In a 500 mL three-necked flask, 25 g of stevioside and 200 mL of 5% potassium hydroxide aqueous solution were added, the reaction system was heated to 60℃, and the reaction was stirred for 6 hours. The reaction system was cooled to room temperature, 6N hydrochloric acid was added to adjust the pH value of the system to 5-6, and the solid was precipitated. The filter cake was washed twice with 100 mL of purified water, the filter cake was collected, and the filter cake was dried at 40℃ under reduced pressure for 6 hours to obtain 18 g of STVB as a white solid, with a yield of 90.2%, an HPLC purity of 98.37%, and a negative ion mode mass-to-charge ratio of 641.3198 [MW-H] by LC-MS / MS, as shown in Figure 1, - 1 ​H-NMR(600MHz,d-DMSO+D2O)δ5.099(s,1H),4.745(s,1H),4.470-4.457(d,J=7.8Hz,1H),4.372-4.359(d ,J=7.8Hz,1H),3.609-3.570(m,2H),3.492-3.428(m,2H),3.400-3.370(m,1H),3.232-3.204(m,1H),3.1 84-3.117(m,3H), 3.060-2.985(m,3H), 2.210-1.966(m,4H), 1.882-1.696(m,6H), 1.520-1.467(m,3H), 1.399-1.316(m,3H), 1.090(s,3H), 0.991-0.967(m,1H), 0.923-0.885(m,5H), 0.794-0.747(m,1H), see Figure 2 for details.

[0176] In a 500 mL three-necked flask, 18 g of STVB, 42 g of acetic anhydride, and 5.5 g of sodium acetate were added. The reaction system was heated to 140 °C and stirred for 6 hours. The reaction system was then cooled to room temperature, 400 mL of dichloromethane was added, and the mixture was washed three times with 200 mL of purified water. The collected organic phase was evaporated to dryness under reduced pressure at 40 °C to obtain 23 g of STVB-Ac as a light brown solid. The yield was 87.6%, and the main peak mass-to-charge ratio in LC-MS / MS anion mode was 935.4012 [MW-H]. - And the mass-to-charge ratio is 980.9836[MW+2Na-H] - See Figure 3 for details.

[0177] (2) Preparation of Man-4 mannose activation precursor

[0178] In a 500 mL three-necked flask, 18 g of D-mannose (Man), 204 g of acetic anhydride, and 19.66 g of sodium acetate were added. The system was heated to 140 °C and stirred for 8 hours. Heating was stopped, and the mixture was allowed to cool naturally to room temperature. The system was then diluted with 500 mL of purified water and extracted three times with 300 mL of dichloromethane. The organic phases were combined, washed three times with 500 mL of purified water, and then washed once with 500 mL of saturated sodium chloride solution. The organic phase was separated, dried over anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure at 40 °C to obtain 35 g of a yellow oily substance, which was compound Man-1, with a yield of 89.7%.

[0179] In a 500 mL three-necked flask, 35 g of Man-1 and 200 mL of dichloromethane were added. While stirring at room temperature, 36 g of acetic acid solution of hydrogen bromide (33%) was added dropwise. After the addition was complete, stirring was continued at room temperature for 12 hours. The system was then poured into 400 mL of ice water and extracted three times with 400 mL of dichloromethane. The organic phases were combined and washed three times with 200 mL of purified water, and then washed once with 400 mL of saturated sodium chloride aqueous solution. The organic phase was separated, dried with anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure at 40 °C to obtain 32 g of a brownish-red oily substance as the compound. The yield was 86.8%.

[0180] In a 500mL three-necked flask, add 30g of Man-2, 300mL of acetone and 15mL of purified water. Add 26g of silver carbonate in portions. After the addition is complete, stir at room temperature for 5 hours, filter, and concentrate the filtrate under reduced pressure at 40℃ to obtain 23g of yellow oily substance, which is Man-3. Yield: 90.5%.

[0181] In a 500 mL three-necked flask, 20 g of Man-3, 200 mL of dichloromethane, 83 g of trichloroacetonitrile, and 1.74 g of 1,8-diazabicycloundec-7-ene (DBU) were added. The reaction mixture was stirred at room temperature for 5 hours. The mixture was then concentrated under reduced pressure at 40 °C. The residue was purified by preparative chromatography using silica gel column chromatography under medium pressure, with dichloromethane and ethyl acetate as eluents, using a gradient elution to give 25 g of a pale yellow oil, which was Man-4. Yield: 88.4%. 1 H-NMR (600MHz, CDCl3) δ 8.809 (s, 1H), 6.301 (d, 1H), 5.490–5.458 (m, 1H), 5.422 (d, 2H), 4.265 (d, 1H), 4.187 (s, 1H), 4.166–4.087 (m, 1H), 2.210 (s, 3H), 2.088 (d, 6H), 2.018 (s, 3H), see Figure 4 for details.

[0182] (3) Preparation of STVB-Man

[0183] In a 500mL three-necked flask, add 10g of Man-4, 200mL of dichloromethane, 21g of STVB-Ac, and 20g of [unspecified ingredient]. Molecular sieves were used, and 1 g of trimethylsilyl trifluoromethanesulfonate (TMSOTf) was added dropwise at -20℃. After the addition was complete, the mixture was kept at this temperature and stirred for 5 hours. Then, 6.5 g of triethylamine was added dropwise, and the mixture was naturally heated to room temperature and stirred for 1 hour. The mixture was filtered, and the filtrate was concentrated under reduced pressure at 40℃ to obtain 38.5 g of a brown oily substance, which was the crude STVB-Ac-Man product.

[0184] In a 500 mL flask, 38 g of STVB-Ac-Man crude product, 300 mL of methanol, 5.27 g of sodium methoxide were added, stirred at room temperature for 5 hours, and the system was concentrated under reduced pressure at 40°C to obtain a foaming oil as STVB-Man crude product, which was purified by medium pressure preparative chromatography to obtain 11 g of white solid as STVB-Man, yield: 67.3%, main peak LC-MS / MS mass-to-charge ratio in negative ion mode: 803.3758 [MW-H] - and 849.3783 [MW+Na-H] - , as shown in Figure 5, 1 HNMR (600MHz, d-DMSO) δ 5.623-5.614 (d, 1H), 5.165-5.154 (d, 1H), 5.133 (s, 1H), 5.114-5.108 (d, 1H), 5.052-5.043 (d, 1H), 4.886 (s, 2H), 4.763 (s, 1H), 4.670-4.657 (d, 1H), 4.621-4.603 (t, 1H), 4.508-4.493 (m, 2H), 4.441-4.421 (m, 2H), 4.064-4.045 (t, 1H), 3.708-3.548 (m, 4H), 3.480-3.403 (m, 5H), 3.231-3.135 (m, 4H), 3.117-3.000 (m, 5H), 2.944-2.904 (m, 1H), 2.116-1.975 (m, 4H), 1.862-1.713 (m, 6H), 1.515-1.363 (m, 6H), 1.110 (s, 3H), 1.016-0.916 (m, 3H), 0.882 (s, 3H), 0.806-0.762 (m, 1H), as shown in Figure 6.

[0185] Example 2. Preparation of RBDS-B-Rha

[0186] (1) Preparation of RBDS-B-Ac

[0187] In a 500 mL flask, 38 g of STVB-Ac-Man crude product, 300 mL of methanol, 5.27 g of sodium methoxide were added, stirred at room temperature for 5 hours, and the system was concentrated under reduced pressure at 40°C to obtain a foaming oil as STVB-Man crude product, which was purified by medium pressure preparative chromatography to obtain 11 g of white solid as STVB-Man, yield: 67.3%, main peak LC-MS / MS mass-to-charge ratio in negative ion mode: 803.3758 [MW-H] - and 849.3783 [MW+Na-H] - , as shown in Figure 5, 1 HNMR (600MHz, d-DMSO) δ 5.623-5.614 (d, 1H), 5.165-5.154 (d, 1H), 5.133 (s, 1H), 5.114-5.108 (d, 1H), 5.052-5.043 (d, 1H), 4.886 (s, 2H), 4.763 (s, 1H), 4.670-4.657 (d, 1H), 4.621-4.603 (t, 1H), 4.508-4.493 (m, 2H), 4.441-4.421 (m, 2H), 4.064-4.045 (t, 1H), 3.708-3.548 (m, 4H), 3.480-3.403 (m, 5H), 3.231-3.135 (m, 4H), 3.117-3.000 (m, 5H), 2.944-2.904 (m, 1H), 2.116-1.975 (m, 4H), 1.862-1.713 (m, 6H), 1.515-1.363 (m, 6H), 1.110 (s, 3H), 1.016-0.916 (m, 3H), 0.882 (s, 3H), 0.806-0.762 (m, 1H), as shown in Figure 6.

[0185] Example 2. Preparation of RBDS-B-Rha

[0186] (1) Preparation of RBDS-B-Ac

[0187] In a 500 mL flask, 38 g of STVB-Ac-Man crude product, 300 mL of methanol, 5.27 g of sodium methoxide were added, stirred at room temperature for 5 hours, and the system was concentrated under reduced pressure at 40°C to obtain a foaming oil as STVB-Man crude product, which was purified by medium pressure preparative chromatography to obtain 11 g of white solid as STVB-Man, yield: 67.3%, main peak LC-MS / MS mass-to-charge ratio in negative ion mode: 803.3758 [MW-H] - and 849.3783 [MW+Na-H] - , as shown in Figure 5, 1 HNMR (600MHz, d-DMSO) δ 5.623-5.614 (d, 1H), 5.165-5.154 (d, 1H), 5.133 (s, 1H), 5.114-5.108 (d, 1H), 5.052-5.043 (d, 1H), 4.886 (s, 2H), 4.763 (s, 1H), 4.670-4.657 (d, 1H), 4.621-4.603 (t, 1H), 4.508-4.493 (m, 2H), 4.441-4.421 (m, 2H), 4.064-4.045 (t, 1H), 3.708-3.548 (m, 4H), 3.480-3.403 (m, 5H), 3.231-3.135 (m, 4H), 3.117-3.000 (m, 5H), 2.944-2.904 (m, 1H), 2.116-1.975 (m, 4H), 1.862-1.713 (m, 6H), 1.515-1.363 (m, 6H), 1.110 (s, 3H), 1.016-0.916 (m, 3H), 0.882 (s, 3H), 0.806-0.762 (m, 1H), as shown in Figure 6.- See Figure 7 for details. 1 H NMR (600 MHz, d-DMSO) δ 11.957 (s, 1H), 5.610-5.601 (d, J = 5.4, 1H), 5.156-5.135 (m, 2H), 5.084-5.076 (d, J = 4.8, 1H), 5.021-5.012 (d, J = 5.4, 1H), 4.846 (s, 2H), 4.767 (s, 1H), 4.672-4.659 (d, J = 7.8, 1H), 4.596-4.579 (m, 1H), 4.509-4.491 (m, 2H), 4.442-4.408 (m, 2H), 4.033 (m, 1H), 3.714-3.545 (m, 4H), 3.482-3.392 (m, 4H), 3.233-2.986 (m, 9H), 2.945-2.918 (m, 1H), 2.078-1.932 (m, 4H), 1.874-1.698 (m, 6H), 1.529-1.344 (m, 6H), 1.112 (s, 3H), 1.025-0.928 (m, 3H), 0.844 (s, 3H), 0.807-0.767 (m, 1H). See Figure 8 for details.

[0188] In a 500 mL three-necked flask, 18 g of rebaudioside B (RBDS-B), 45 g of acetic anhydride and 4.5 g of sodium acetate were added, the reaction system was warmed to 140°C, and was kept stirring for 6 hours. The reaction system was reduced to room temperature, 400 mL of dichloromethane was added, the organic phase was washed with 200 mL of purified water for three times, the organic phase was collected, and was concentrated at 40°C to obtain 27 g of RBDS-B-Ac as a light yellow solid. LC-MS / MS main peak negative ion mode mass to charge ratio 1223.4819 [MW-H] - and mass to charge ratio 1310.4568 [MW+2Na+MeCN-H] - See Figure 9 for details.

[0189] (2) Preparation of rhamnose activated precursor Rha-3

[0190] In a 500 mL flask, 25 g of L-rhamnose (Rha) was added, 150 mL of pyridine, the system was cooled to 0°C, 100 g of acetic anhydride was added dropwise under stirring, after the dropwise addition was completed, it was stirred at room temperature for 24 hours, 200 mL of purified water was added, then extracted with 300 mL of ethyl acetate three times, the organic phase was combined and washed once with 200 mL of saturated sodium chloride aqueous solution, washed three times with 200 mL of saturated sodium bicarbonate aqueous solution, the organic phase was separated, dried with anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure at 40°C to obtain 43 g of compound Rha-1 in the form of yellow oil, yield: 84.3%.

[0191] In a 500 mL flask, 40 g of Rha-1 was added, 250 mL of tetrahydrofuran, 20 g of benzylamine, stirred at room temperature for 24 hours, concentrated under reduced pressure at 40°C, the residue was added to 500 mL of dichloromethane, washed twice with 100 mL of 1N hydrochloric acid, washed three times with 200 mL of saturated sodium bicarbonate aqueous solution, the organic phase was separated, dried with anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure at 40°C to obtain 27.5 g of compound Rha-2 in the form of yellow oil, yield: 77.6%.

[0192] In a 500 mL flask, 25 g of Rha-2 was added, 250 mL of acetone, 24 g of potassium carbonate, 35 g of 2,2,2-trifluoro-N-phenyliminoacetyl chloride, stirred at room temperature for 24 hours, filtered, and the filtrate was concentrated under reduced pressure at 40°C. The residue was purified by silica gel column using medium pressure preparative liquid phase, the eluent was petroleum ether and ethyl acetate, and the online gradient elution was performed to obtain 31 g of Rha-3 in the form of light yellow oil, yield: 79.4%, 1 H NMR (600 MHz, CDCI3) δ 7.333-7.316 (t, 2H), 7.160-7.137 (t, 1H), 6.894-6.878 (d, 2H), 6.170 (m, 1H), 5.152-5.056 (m, 3H), 4.080-4.031 (m, 1H), 2.054 (s, 3H), 2.039 (s, 3H), 2.032 (s, 3H), 1.106-1.071 (d, 3H), see Figure 10.

[0193] (3) Preparation of RBDS-B-Rha

[0194] In a 500 mL flask, 10 g of Rha-3 was added, 200 mL of dichloromethane, 25 g of RBDS-B-Ac, 25 g of Molecular sieves were used, and 1.5 g of TMSOTf was added dropwise at -10℃. After the addition was complete, the mixture was kept warm and stirred for 8 hours. Then, 6.0 g of triethylamine was added dropwise. The mixture was naturally heated to room temperature and stirred for another 14 hours. The mixture was filtered, and the filtrate was concentrated under reduced pressure at 40℃ to obtain 45 g of a brown oily substance, which was crude RBDS-B-Ac-Rha.

[0195] In a 500 mL three-necked flask, 45 g of crude RBDS-B-Ac-Rha, 300 mL of methanol, and 8.0 g of potassium tert-butoxide were added. The mixture was stirred at room temperature for 16 hours. The system was then concentrated under reduced pressure at 40 °C to obtain a foaming oily substance, which was crude RBDS-B-Rha. After medium-pressure separation and purification, 9.31 g of white solid, RBDS-B-Rha, was obtained, with a yield of 48.0%. The mass-to-charge ratio of the main peak in negative ion mode on LC-MS / MS was 949.4298 [MW-H]. - and 995.3783[MW+2Na-H] - See Figure 11 for details. 1 H NMR(600MHz,d-DMSO)δ5.630-5.622(d,1H),5.285-5.272(d,1H),5.236-5.226 (d,1H),5.149-5.138(d,1H),5.108-5.100(d,1H),5.058(s,1H),5.035-5.026 (d,1H),4.988-4.973(m,3H),4.904(s,1H),4.744(s,1H),4.688-4.669(t,1H) ,4.645-4.632(d,1H),4.610-4.592(t,1H),4.511-4.465(m,3H),4.434-4.421( d,1H),4.110-4.091(t,1H),3.715-3.676(m,2H),3.639-3.603(m,2H),3.562- 3.396(m,5H),3.270-3.231(m,1H),3.149-3.117(m,6H),3.088-3.025(m,4H), 3.003-2.964(m,1H), 2.113-1.690(m,10H), 1.497-1.334(m,6H), 1.147(s,3H), 1.073-0.910(m,3H), 0.874(s,3H), 0.807-0.788(m,1H), 0.765(d,3H), see Figure 12 for details.

[0196] Example 3. Synthesis of DCS-A-Gal

[0197] (1) Preparation of DCS-A-Acid-Ac

[0198] In a 500 mL three-necked flask, 30 g of DCS-A and 250 mL of 5% potassium hydroxide aqueous solution were added, the reaction system was warmed to 85-90°C, and the reaction was stirred for 6 hours. The reaction system was cooled to room temperature, 6N hydrochloric acid was added to adjust the pH value of the system to 5-6, and the solid was precipitated. The filter cake was washed twice with 100 mL of purified water, the filter cake was collected, and dried under reduced pressure at 40°C to obtain 18 g of DCS-A-Acid as a brown solid with a purity of 90.51% by HPLC, and the main peak of LC-MS / MS was 625.3297[MW-H] in negative ion mode. - and 1251.6632[2MW-H] - , as shown in Figure 13, 1 H NMR (600 MHz, d-DMSO) δ 11.886 (s, 1H), 5.781 (s, 1H), 5.307-4.961 (m, 4H), 4.705 (s, 1H), 4.504-4.230 (m, 3H), 3.428-3.370 (m, 4H), 3.254-3.124 (m, 4H), 3.061-2.986 (m, 3H), 2.123-1.967 (m, 4H), 1.882-1.704 (m, 6H), 1.520-1.467 (m, 3H), 1.390-1.317 (m, 3H), 1.090 (s, 3H), 1.007-0.909 (m, 9H), 0.885-0.747 (m, 1H), as shown in Figure 14.

[0199] In a 500 mL three-necked flask, 30 g of DCS-A and 250 mL of 5% potassium hydroxide aqueous solution were added, the reaction system was warmed to 85-90°C, and the reaction was stirred for 6 hours. The reaction system was cooled to room temperature, 6N hydrochloric acid was added to adjust the pH value of the system to 5-6, and the solid was precipitated. The filter cake was washed twice with 100 mL of purified water, the filter cake was collected, and dried under reduced pressure at 40°C to obtain 18 g of DCS-A-Acid as a brown solid with a purity of 90.51% by HPLC, and the main peak of LC-MS / MS was 625.3297[MW-H] in negative ion mode. - and 923.3894[MW+2Na-H] - , as shown in Figure 15.

[0200] (2) Preparation of galactose-activated precursor Gal-2

[0201] In a 500 mL three-necked flask, 30 g of D-galactose (Gal) was added, 150 g of acetic anhydride and 10 g of sodium acetate were added, after completion of addition, the system was heated to 120°C and stirred for 8 hours, the heating was stopped, and the system was naturally cooled to room temperature, then the system was diluted into 500 mL of purified water, and then extracted with 300 mL of dichloromethane three times, the organic phase was combined and washed with 500 mL of purified water three times, and then washed with 500 mL of saturated sodium chloride aqueous solution once, the organic phase was separated, dried with anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure at 40°C to obtain 55 g of yellow oil as compound Gal-1, yield: 84.6%.

[0202] In a 500 mL three-necked flask, 50 g of Gal-1, 250 mL of dichloromethane were added, and 55 g of hydrogen bromide in acetic acid solution (33%) was added dropwise while stirring at room temperature, after completion of dropwise addition, the stirring was continued at room temperature for 16 hours, the system was poured into 400 mL of ice water, extracted with 400 mL of dichloromethane three times, the organic phase was combined and washed with 200 mL of purified water three times, and then washed with 400 mL of saturated sodium chloride aqueous solution once, the organic phase was separated, dried with anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure at 40°C to obtain 41 g of brown oil as compound Gal-2, yield: 78.2%, 1 H NMR (600 MHz, CDCl3) δ 6.688-6.681 (d, 1H), 5.527-5.522 (d, 1H,), 5.439-5.417 (m, 1H), 5.040-5.018 (m, 1H), 4.469-4.425 (m, 1H), 4.242-4.159 (m, 2H), 2.129 (s, 3H), 2.093 (s, 6H), 2.033 (s, 3H), see Figure 16.

[0203] (3) Preparation of DCS-A-Acid-Gal

[0204] In a 500 mL three-necked flask, 30 g of D-galactose (Gal) was added, 150 g of acetic anhydride and 10 g of sodium acetate were added, after completion of addition, the system was heated to 120°C and stirred for 8 hours, the heating was stopped, and the system was naturally cooled to room temperature, then the system was diluted into 500 mL of purified water, and then extracted with 300 mL of dichloromethane three times, the organic phase was combined and washed with 500 mL of purified water three times, and then washed with 500 mL of saturated sodium chloride aqueous solution once, the organic phase was separated, dried with anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure at 40°C to obtain 55 g of yellow oil as compound Gal-1, yield: 84.6%. In a 500 mL three-necked flask, 50 g of Gal-1, 250 mL of dichloromethane were added, and 55 g of hydrogen bromide in acetic acid solution (33%) was added dropwise while stirring at room temperature, after completion of dropwise addition, the stirring was continued at room temperature for 16 hours, the system was poured into 400 mL of ice water, extracted with 400 mL of dichloromethane three times, the organic phase was combined and washed with 200 mL of purified water three times, and then washed with 400 mL of saturated sodium chloride aqueous solution once, the organic phase was separated, dried with anhydrous sodium sulfate, filtered, and the filtrate was concentrated under reduced pressure at 40°C to obtain 41 g of brown oil as compound Gal-2, yield: 78.2%,

[0205] In a 500 mL three-necked flask, 45 g of DCS-A-Acid-Gal-Ac crude product, 300 mL of methanol, 8.0 g of potassium tert-butoxide were added, stirred at room temperature for 16 hours, and the system was concentrated under reduced pressure at 40°C to obtain a foaming oil as DCS-A-Acid-Gal crude product. After purification by medium pressure preparative chromatography, 8.89 g of white solid was obtained as DCS-A-Acid-Gal, yield: 32.9%, LC-MS / MS main peak mass-to-charge ratio in negative ion mode was 787.3796 [MW-H] - and mass-to-charge ratio 439.1916 [MW / 2+2Na-1] - as shown in Figure 17, 1 H NMR (600 MHz, d-DMSO) δ 6.257 (d, 1H), 5.622 (d, 1H), 5.335 (s, 1H), 5.073-4.976 (m, 3H), 4.893 (s, 1H), 4.737 (s, 1H), 4.691 (s, 1H), 4.497-4.129 (m, 4H), 3.696-3.593 (m, 3H), 3.498-3.379 (m, 5H), 3.238-2.992 (m, 10H), 2.123-1.995 (m, 4H), 1.911-1.710 (m, 5H), 1.608-1.352 (m, 9H), 1.056-0.918 (m, 7H), 0.835-0.735 (m, 4H), as shown in Figure 18

[0206] Example 4. Preparation of DCS-B-Man

[0207] DCS-B-Man was synthesized according to the synthesis route and method of STVB-Man in Example 1, and 8 g of white solid was obtained, yield: 57.7%, LC-MS / MS main peak mass-to-charge ratio in negative ion mode was 949.4291 [MW-H] - and mass-to-charge ratio 474.2373 [MW / 2-H] - as shown in Figure 19, 1H NMR (600 MHz, d-DMSO) δ 5.698 (d, 1H), 5.353-5.340 (d, 1H), 5.304-5.294 (d, 1H), 5.217-5.129 (m, 4H), 5.094 (m, 2H), 4.972 (s, 1H), 4.812 (s, 1H), 4.756-4.660 (m, 3H), 4.575-4.489 (m, 2H), 4.178-4.159 (t, 1H), 3.783-3.671 (m, 4H), 3.601-3.464 (m, 7H), 3.337-3.240 (m, 9H), 3.189-3.071 (m, 4H), 2.180-1.789 (m, 10H), 1.564-1.423 (m, 6H), 1.293 (s, 3H), 1.212-0.978 (m, 5H), 0.942-0.871 (m, 1H), 0.851 (s, 3H), 0.851-0.834 (m, 1H), see Figure 20.

[0208] Example 5. Preparation of DCS-B-Rha

[0209] DCS-B-Rha was synthesized according to the synthetic route and method of RBDS-B-Rha in Example 2, to give white solid 8.5 g, yield: 60.1%. LC-MS / MS main peak mass to charge ratio 933.4391 [MW-H] in negative ion mode and 979.4902 [MW+2Na-H] in positive ion mode, see Figure 21. - - 1 H NMR (600 MHz, d-DMSO + D20) δ 5.873 (d, 1H), 5.288 (d, 1H), 5.202 (d, 1H), 5.154 (s, 1H), 5.073 (s, 1H), 4.971 (m, 1H), 4.234-4.207 (m, 1H), 4.104-4.031 (m, 2H), 4.022-3.858 (m, 2H), 3.787-3.746 (m, 7H), 3.607-3.197 (m, 8H), 2.172-2.110 (m, 2H), 2.066-1.905 (m, 5H), 1.859-1.575 (m, 9H), 1.499-1.426 (m, 2H), 1.289-1.257 (m, 1H), 1.150 (d, 3H), 1.132 (s, 3H), 1.095 (s, 3H), 0.980 (m, 4H), see Figure 22.

[0210] Example 6. Preparation of STVB-Gal ​​

[0211] STVB-Gal was prepared according to the synthetic route and preparation method of DCS-A-Acid-Gal in Example 3, to give white solid 9.4 g, yield: 54.3%. LC-MS / MS main peak mass to charge ratio 803.3746 [MW-H] - and mass to charge ratio 447.1799 [MW / 2-H] - , as shown in Figure 23, 1 H NMR (600 MHz, d-DMSO) δ 5.607-5.598 (d, 1H), 5.149-5.138 (d, 1H), 5.117 (s, 1H), 5.098-5.090 (d, 1H), 5.036-5.027 (d, 1H), 4.870 (s, 2H), 4.747 (s, 1H), 4.655-4.641 (d, 1H), 4.605-4.587 (t, 1H), 4.490-4.478 (m, 2H), 4.425-4.405 (m, 2H), 4.048-4.029 (t, 1H), 3.692-3.532 (m, 4H), 3.464-3.387 (m, 5H), 3.215-3.119 m, 4H), 3.101-2.984 (m, 5H), 2.928-2.888 (m, 1H), 2.100-1.959 (m, 4H), 1.846-1.679 (m, 6H), 1.499-1.347 (m, 6H), 1.094 (s, 3H), 1.000-0.900 (m, 3H), 0.866 (s, 3H), 0.790-0.746 (m, 1H), as shown in Figure 24.

[0212] Example 7. Preparation of Steviosin (STVN)-Man

[0213] STVN-Man derivatives were prepared using steviol glycosides or crude steviol glycosides extract as starting material.

[0214] The content of each steviol glycoside in the STVN used in this experiment was as follows: RBDS-A: 45.22%, STVS: 19.34%, RBDS-C: 14.56%, DCS-A: 1.98%, RBSD: 1.44%, RBDS-B: 2.18%, STVB: 1.69%, RBDS-F: 2.77%, RBDS-H: 1.76%, RBDS-E: 0.85%, RBDS-O: 0.68%, RBDS-D: 1.82%, STL: 0.38%, and RBDS-M: 2.92%, total steviol glycoside content: 97.21% (not including STL), unknown steviol glycoside about 2.41%.

[0215] Take about 25 g of the above STVN, put it in a 500 mL three-necked flask, and prepare the crude STVN-Man according to the synthesis route and method of STVB-Man in Example 1. After purification by medium pressure preparative chromatography, 12.7 g of white solid is obtained, with a yield of about 70.1%.

[0216] The peak mass spectrum identification results of each component in STVN-Man are shown in the following table.

[0217] Table 6. LC / MS / MS mass spectrum identification of each component peak of STVN-Man - from large to small in polarity

[0218] Note 1: Absolute error.

[0219] Example 8. Preparation of STVN-Rha

[0220] STVN-Rha is prepared using the same amount of STVN as in Example 7.

[0221] Take about 25 g of the above STVN, put it in a 500 mL three-necked flask, and prepare the crude STVN-Man according to the synthesis route and method of STVB-Man in Example 1. After purification by medium pressure preparative chromatography, 12.7 g of white solid is obtained, with a yield of about 70.1%.

[0222] The peak mass spectrum identification results of each component in STVN-Rha are shown in the following table.

[0223] Table 7. LC / MS / MS mass spectrum identification of each component peak of STVN-Rha - from large to small in polarity

[0224] Note 1: Absolute error.

[0225] Example 9. Preparation of STVN-Gal

[0226] STVN-Gal is prepared using the same amount of STVN as in Example 7.

[0227] Take about 25 g of the above STVN, put it in a 500 mL three-necked flask, and prepare the crude STVN-Man according to the synthesis route and method of STVB-Man in Example 1. After purification by medium pressure preparative chromatography, 12.7 g of white solid is obtained, with a yield of about 70.1%.

[0228] The peak mass spectrum identification results of each component in STVN-Gal are shown in the following table.

[0229] Table 8. LC / MS / MS mass spectrum identification of each component peak of STVN-Gal - from large to small in polarity

[0230] Note 1: Absolute error.

[0231] Example 10. Solubility investigation in common solvents (liquids)

[0232] According to the preparation process of each derivative and the common solvents used as carriers, the solubility of the above derivatives in the following solvents (liquids) was investigated according to the solubility test method of Chinese Pharmacopoeia 2020 Edition Fourteen (2).

[0233] Each derivative or its composition was weighed into fine powder and placed in a certain amount of the following solvent at 25°C ± 2°C. Every 5 minutes, it was shaken vigorously for 30 seconds and observed for 30 minutes. If no visible solute particles were observed, it was considered completely dissolved. The results are shown in Table 9 below:

[0234] Table 9: Solubility test results of each derivative or composition (25°C)

[0235] According to the solubility investigation results of the above carrier derivatives or their compositions, the solubility of each steviol glycoside derivative in the above medium is basically the same as that of its glucose derivative. The solubility of the rhamnose derivative of the same steviol glycoside is lower than that of its mannose or galactose derivative. The different sugar derivatives of STVN also show similar trends.

[0236] Example 11. Steviol glycoside derivatives as delivery carriers and / or targeting guides

[0237] Curcumin (CCM, molecular weight: 368.38; melting point: 179°C-182°C; slightly soluble in hot water, insoluble in cold water) was used as the target delivery material. STVB-Man:RBDS-A, (1:3, g / g), DSC-B-Man, STVN-Rha:RBDS-A (1:3, g / g), STVN-Gal, and STVN-Man were used as delivery carriers and / or targeting guides. The nanocomposites containing CCM were prepared according to the following prescription and preparation process.

[0238] (1) Composition of excipients or carriers in the prescription

[0239] Table 10. Prescription composition of supramolecular nanocomposites

[0240] (2) Preparation process

[0241] CCM powder was weighed, and THF was added to dissolve it to prepare a solution containing about 20% to 30% CCM. The solution was filtered and prepared for use. The carrier or derivative and the polymer in the prescription were weighed, and the carrier or derivative in each prescription was dissolved in 20 mL of 0.078% citric acid aqueous solution. The pH of the aqueous solution was controlled at 2.8 to 3.2. If the carrier or derivative was not dissolved, heating (temperature not exceeding 60°C) was used to aid dissolution. After the solution was clear, the polymer Soluplus was added to dissolve it. After the solution was clear, it was naturally cooled to room temperature, and the initial weight was recorded. While stirring, the THF solution containing CCM was added dropwise. After the dropwise addition was completed, the stirring was continued at room temperature until the total weight was substantially consistent with the weight before the dropwise addition. The stirring was stopped, and centrifugation was performed at 13000 rpm for 5 minutes. The supernatant aqueous solution was taken, and the CCM concentration, particle size, and Zeta potential were detected. The supernatant was sealed in a vial and placed at room temperature to investigate the physical stability.

[0242] (3) Characteristic evaluation and result detection

[0243] 1) CCM concentration, particle size, Zeta potential, and physical stability

[0244] Table 11. Detection results of CCM supramolecular nanocomplexes of Comparative Example 1 and F1-F5

[0245] Comparative Example 1 and F1-F5 were placed at room temperature for 24 hours, and no precipitation or delamination was observed. They were all uniform yellow solutions. The particle size of Comparative Example 1 was greater than 100 nm, and the particle sizes of F1-F5 were between 57 nm and 63 nm. The PDI of each prescription was less than 0.3 and was basically neutral.

[0246] According to the determination results of the CCM concentration in the supernatant of Comparative Example 1 and F1-F5, the prescriptions were the same except for the different steviol glycoside derivatives. Under the same preparation process, the assembly ability and the solubilization ability of each derivative after assembly were not worse than those of Comparative Example 1. The improvement of the solubility of each derivative was 4590 times to 5320 times that of curcumin powder (the solubility of curcumin in water at 25°C was about 13.76 μg / mL), and that of Comparative Example 1 was 2607 times.

[0247] 2) Drug loading and encapsulation efficiency calculation

[0248] 3 mL of the supramolecular nanocomplex solution prepared in each of the above Comparative Example 1 and F1-F5 was taken into a vial with a known weight, and the weight was recorded. The vial was placed in a freeze dryer for 24 hours, taken out, sealed with a rubber plug, and the weight of the solid was recorded again. The drug loading and encapsulation efficiency were calculated, and the results are shown in the table below.

[0249] The drug loading calculation formula is as follows:

[0250] Drug loading % = drug content in unit volume (mg) / solid content in unit volume (mg / ml) x 100.

[0251] The drug loading of Comparative Example 1, F1-F5 was 13.3%, 23.8%, 27.0%, 24.3%, 27.6% and 26.5%, respectively. Under the same conditions of the amount of carrier or derivative, other high molecular polymers in the prescription and preparation process, the drug loading of F1-F5 was more than 10% higher than that of Comparative Example 1.

[0252] The encapsulation efficiency was calculated according to the following formula:

[0253] EE = [amount of encapsulated drug (mg) / initial amount (mg)] x 100%.

[0254] The encapsulation efficiency of Comparative Example 1, F1-F5 was 51.15%, 90.24%, 103.3%, 94.54%, 104.6% and 100.1%, respectively. Using CCM as the active ingredient, the encapsulation efficiency of the carrier derivative on CCM was more than 90%.

[0255] 3) Resolubilization after lyophilization

[0256] Weighed each prescription lyophilized powder, containing about 35 mg of CCM, placed in a 4 mL centrifuge tube, respectively, 10 mL of 0.1 M hydrochloric acid or pH 6.8 phosphate buffer was used for resolubilization, placed in a room temperature shaker for 30 minutes, 13000 rpm centrifugation for 10 minutes, the supernatant was detected for content and particle size distribution, according to the measured content to calculate the solubilization percentage of lyophilized powder after resolubilization, the results were shown in the following table.

[0257] Table 12. Resolubilization results of Comparative Example 1 and F1-F5 after lyophilization

[0258] According to the above results, the dissolution percentage of the freeze-dried powder of Comparative Example 1, F3, F4 after reconstitution in 0.1M hydrochloric acid is slightly lower than that of other formulations, which are 78.32%, 86.18% and 88.33% respectively, and the dissolution percentage of other formulations after reconstitution in 0.1M hydrochloric acid is greater than 90%, indicating that the supramolecular nanocomplexes prepared by Comparative Example 1 and Formulations F1-F5 can withstand a strong acid environment. The particle size distribution of the supernatant after reconstitution of each formulation in 0.1M hydrochloric acid solution is basically consistent with that before freeze-drying; the reconstitution phenomenon of each formulation in pH 6.8 phosphate buffer is as follows: Comparative Example 1 and F1 are slightly turbid after reconstitution, and the dissolution percentages after reconstitution are 84.21% and 89.42% respectively, F2 and F5 are clear after reconstitution and are basically completely dissolved, F3 and F4 are clear after reconstitution, but there is a small amount of suspended matter after shaking, and the dissolution percentages are 85.37% and 90.88% respectively. The particle size detection of the supernatant after reconstitution of each formulation in pH 6.8 phosphate buffer is basically consistent with that before freeze-drying. According to the above results, whether it is the supramolecular nanocomplex solution prepared by Comparative Example 1 or Formulations F1-F5, it can be basically reconstituted after freeze-drying, and the particle size does not change significantly after reconstitution, and the physical stability is good.

[0259] 4) Investigation of cumulative dissolution in pre-meal biological media

[0260] Dissolution medium: pre-meal simulated gastric fluid (FaSSGF) and pre-meal simulated intestinal fluid (FaSSIF-V2)

[0261] Fasted State Simulated Gastric Fluid (FaSSGF) Preparation of FaSSGF: weigh 2g of sodium chloride, 0.043g of sodium taurocholate, 0.015g of lecithin, add 800ml of degassed deionized water, ultrasonic at 40-50°C to dissolve, adjust pH to 1.60 with 1M HCl, add 0.1g of pepsin, add degassed deionized water to 1L, confirm pH again and adjust to 1.60, and it is obtained.

[0262] Fasted State Simulated Intestinal Fluid (FaSSIF) Preparation of FaSSIF-V2: add 63ml of conditioning solution (preparation of conditioning solution: weigh 8.8056g of maleic acid, 8.8062g of sodium hydroxide, 9.9762g of sodium chloride, 6.618g of sodium taurocholate, and 0.551g of lecithin, add 1000ml of degassed deionized water, ultrasonic at 40-50°C to dissolve) to 187ml of FaSSGF, and control the pH at 6.50.

[0263] Dissolution method:

[0264] Small cup method, paddle method, rotation speed: 100rpm, temperature: 37°C;

[0265] Medium and volume: 187 mL FaSSGF / 250 mL FaSSIF-V2

[0266] Dosage: 70 mg per dissolution cup, the volume of supramolecular nanosolution was calculated according to the actual content.

[0267] Sampling points: 15, 30, 45, 60, 90, 120, 180 (min), and the sampling was stopped at 30 min, and then the simulated intestinal fluid was adjusted.

[0268] Sampling and sample processing method: 4 mL was taken, and 4 mL of the same medium at the same temperature was added to the dissolution cup. The sample was centrifuged at 15000 rpm for 5 min at 37°C.

[0269] 0.5 mL of the supernatant after centrifugation was taken, 2.5 mL of methanol-water (90:10, V / V) was added for quantitative dilution, and the content of CCM was detected by liquid chromatography. The cumulative dissolution at each sampling time point was calculated. The results are shown in the following table:

[0270] Table 13. Cumulative dissolution in pre-meal biological medium

[0271] The cumulative dissolution of each prescription prepared from the derivative in the pre-meal simulated intestinal fluid for 180 min was close to or more than 90%, and there was no significant precipitation after being transferred to the simulated intestinal fluid. It is indicated that the dissolution behaviors of the supramolecular nanocomplexes constructed by different derivatives in the same dissolution medium are slightly different, especially in the pre-meal simulated gastric fluid. After being transferred to the pre-meal simulated intestinal fluid, the cumulative dissolution of each prescription is not less than that of the comparative example 1. The nanocomplexes constructed by the comparative example 1 and each derivative can tolerate the dilution and influence of the dissolution medium, and most of the nanocomplexes can still maintain the complete nanostructure.

[0272] 5) Cumulative release in 80% human plasma

[0273] The human plasma was naturally thawed at room temperature, and diluted with pH 7.4 phosphate buffer to prepare 80% human plasma.

[0274] F2-F5 were diluted with pH 7.4 phosphate buffer to the same concentration solution as the comparative example 1 according to the actual detection content.

[0275] Precisely pipette 1 mL of the diluted solution of Comparative Example 1 and F1-F5 (about 35.8 mg / mL) into a dialysis bag (relative molecular mass 50000 Dalton) which has been soaked in distilled water, tie the bag opening, use 50 mL of 80% human plasma as the release medium, and stir in a 37°C water bath (100 rpm / min). Prepare 3 samples for each prescription. Take 0.5 mL at the sampling time points (0.5, 1, 2, 4, 6, 8, 12) and add 1.5 mL of methanol to precipitate the protein, vortex, centrifuge (13000 rpm / 10 min), take 1 mL of the supernatant, dilute with 1 mL of pH 3.0 phosphate-methanol (10:90, V / V), and detect the CCM content to calculate the cumulative release rate. Add the same volume of release medium at the same temperature after each sampling. The cumulative release curve is shown in Figure 25:

[0276] As shown by the cumulative release rate in 80% human plasma, the release rate of F3 and Comparative Example 1 is slow, and the cumulative release rate is less than 80% at 12 hours; the release rate of F4 and F5 is the fastest, and the cumulative release rate is close to complete release at 6 hours; the cumulative release rates of F1 and F2 at 12 hours are 94.2% and 91.8%, respectively, which are slightly slower than those of F4 and F5. The above supramolecular nanoparticles are different in carrier composition, but are the same in other aspects, and supramolecular nanoparticles with different release rates can be obtained by adjusting the carrier composition.

[0277] Example 12 Hemolytic experiment

[0278] The small size and unique physical and chemical properties of nanoparticles can lead to different interactions with red blood cells than those observed in conventional drugs. Referring to the Technical Guidelines for Drug Irritability, Allergy and Hemolysis Research and Nano Lett. 2008 August; 8 (8): 2180-2187, an in vitro test tube method is used for hemolysis evaluation.

[0279] (1) Investigation of the particle size stability of CCM supramolecular nanocomplexes

[0280] Take 5 mL of the supramolecular nanocomplex solution prepared from each prescription of Comparative Example 1 and F1-F5, mix with an equal volume of 0.9% normal saline, 5% glucose injection, and pH 7.4 phosphate buffer, incubate at 37°C, measure the particle size at 0, 2, 4, and 6 hr, observe whether there is precipitation or turbidity, and plot the particle size at different time points against time. Evaluate the particle size stability of CCM supramolecular self-assembled nanosolutions in 0.9% normal saline, 5% glucose injection, and pH 7.4 phosphate buffer. Record the observed phenomena at 2 hr and 4 hr, and take 1 mL of each sample and centrifuge at 15000 rpm / 37°C to observe whether there is precipitation after centrifugation. If no obvious turbidity is observed at the 6 hr sampling point, measure the particle size at 6 hr and compare with that at 0 hr.

[0281] According to the stability results, obvious turbidity appeared in pH 7.4 phosphate buffer for 2 hours, while no obvious suspension, precipitation or turbidity appeared in 0.9% normal saline and 5% glucose injection until 6 hours. The particle size detection results at 6 hours changed within ±3 nm compared with the particle size at 0 hours, indicating that the prepared CCM supramolecular nanocomplex had good physical stability.

[0282] Therefore, 0.9% normal saline or 5% glucose injection can be selected for dilution for hemolytic evaluation experiments.

[0283] (2) Hemolysis experiment

[0284] Test drug: Supramolecular nanocomplex solution of Comparative Example 1 and F3-F5 prepared in Example 10

[0285] Experimental animal: male Japanese white rabbits, 2.08 kg, one; source: Beijing Changyang Xishan Breeding Farm, production license number: SCXK-(Jing) 2022-0002. The animals were raised in a 100,000-level environment by a breeder with animal feeding qualifications.

[0286] 1) Preparation of red blood cell suspension for experiment

[0287] Before blood collection, the animal was fixed, the hair on one side of the rabbit ear was shaved, and the blood collection site was disinfected with iodophor. 11.5 mL of rabbit blood was taken and placed in a glass bead triangular flask for shaking for 10 minutes to remove fibrinogen and make the blood defibrinated. About 10 times the amount of 5% glucose injection was added, shaken, centrifuged at 3000 r / min for 5 min, and the supernatant was removed. The precipitate was washed with 5% glucose injection according to the above method for 2-3 times until the supernatant was not red. 4 mL of red blood cells were taken and prepared into a 4% red blood cell suspension with 5% glucose injection, ready for use.

[0288] 2) Experimental method

[0289] Test solution: The F3-F5 formula was diluted with 5% glucose injection to prepare a series of concentration solutions, such as 0.5, 0.125, 0.0625 mg / mL (calculated as CCM), and each concentration was prepared in triplicate as the test solution. 500 μL of test solution and 500 μL of 4% red blood cell suspension were mixed, vortexed for 10 seconds, incubated at 37°C for 3 hours, centrifuged at 5000 r / min for 5 minutes, and 150 μL of supernatant was taken in a 96-well plate. The absorbance A was measured at 540 nm by an enzyme marker. 5% glucose injection was used as the negative control (n=3), and 500 μL of deionized water was used instead of the above test solution and 4% red blood cell suspension 500 μL, and the rest of the operation was the same as the test operation, as the positive control, to calculate the hemolysis rate.

[0290] Hemolysis rate (%) = (A value of test sample solution - A value of negative control group) / (A value of positive control - A value of negative control) x 100, and the results are shown in the following table:

[0291] Table 14. Hemolysis rate of Comparative Example 1 and F3-F5 (n = 3, %)

[0292] According to the above results, the hemolysis rates of Comparative Example 1 and F3-F5 are all less than 5%, and the hemolysis rates of the derivative groups are basically the same as those of Comparative Example 1, indicating that the above carrier derivatives will not cause hemolysis if injected after dilution.

[0293] Preparation of Comparative Example 2 CCM solid dispersion

[0294] CCM powder is difficult to dissolve in aqueous medium, therefore, spray drying method is used to prepare a solid dispersion of CCM and high molecular polymer hydroxypropyl methyl cellulose acetate succinate (HPMCAS LG) at a mass ratio of 1:3.

[0295] The preparation process of CCM solid dispersion is as follows: take CCM 10.0 g and HPMCAS LG 30.0 g, dissolve in methanol-dichloromethane (5:1) 420 ml, spray drying: inlet air temperature 100°C, outlet air temperature 60°C, liquid inlet speed 4.0, air volume 7.0, atomization pressure 0.08 Mpa. Collect the CCM solid dispersion, dry in a vacuum drying oven at 45°C for 4 hours, control the loss on drying (LOD) ≤2%, residual solvent methanol ≤0.3%, dichloromethane ≤0.06%. Detect the content of CCM in the solid dispersion by HPLC method, and confirm whether the solid dispersion is completely formed by DSC.

[0296] (1) The content of CCM in CCM-HPMCAS solid dispersion is 21.78% (g / g).

[0297] (2) Solubility determination of CCM-HPMCAS solid dispersion in pH 7.4 phosphate buffer

[0298] Take pH 7.4 phosphate buffer 30 mL (n = 2), add solid dispersion of CCM:HPMCAS LG = 1:3 (w / w) respectively, place in a shaking bed at 37°C / 180 rpm, take samples at 0.5 hr, 1 hr, 3 hr, 6 hr and 24 hr, place the samples in 2 mL centrifuge tubes, centrifuge at 37°C / 13000 rpm for 5 min, and detect the content of CCM in the supernatant.

[0299] Table 15. Solubility of CCM-HPMCAS solid dispersion

[0300] According to the above results, after CCM was prepared into a solid dispersion with HPMCAS LG (1:3, w / w) and incubated in phosphate buffer for 6 hours, the solubility was 192 μg / mL, which was 32 times that of CCM powder; incubated for 24 hours, as the time was prolonged, the interaction between CCM molecules led to the formation of molecular accumulation, and the solubility decreased to 103 μg / mL, but was still 20.6 times that of CCM powder. The DSC graphs of CCM powder, physical mixture of CCM and HPMCAS, and CCM-HPMCAS solid dispersion are shown in Figure 26:

[0301] According to the DSC results, the endothermic peak of CCM powder completely disappeared in the solid dispersion graph c, indicating that CCM-HPMCAS LG (1:3, w / w) could completely form a solid dispersion.

[0302] Example 13 Investigation of the permeability of supramolecular nanocomplexes constructed by different derivatives

[0303] In vitro permeability evaluation was performed using isolated intestinal tissues. In order to simultaneously investigate the influence of intestinal flora on carrier metabolism, rat ileum tissues were taken, and the corresponding colon contents were collected.

[0304] SD clean male rats were taken after overnight fasting, weighed (223±8g), anesthetized with a certain amount of ether on a cotton ball, and then sacrificed by decapitation. The limbs of the rats were fixed, the skin was cut along the midline of the abdomen, the abdominal muscle layer was clamped with forceps, and an opening of about 10 cm was cut with scissors. The stomach pylorus was cut down 1 cm with scissors, the mesentery was separated by sudden separation method along the duodenum, jejunum and ileum, and the intestinal segment was cut off at the ascending colon 2 cm. The ileum and jejunum each accounted for half of the rat small intestine, and the isolated ileum segment was rinsed with 4℃ physiological saline to clean the outer surface. The ileum was cut along the mesentery, and then washed with 4℃ physiological saline. Under the microscope, avoid the part with vesicle cells, cut into 1 cm intestinal segments, and lay them flat on the receiving chamber inlet of the diffusion cell with the intestinal mucus layer facing up, so that they completely cover the receiving chamber inlet. Then, the supply chamber was vertically placed on the receiving chamber, and clamped with a clamp (after fixing, do not adjust the supply chamber randomly to avoid displacement of the intestinal tissue). The magnet was gently inserted into the receiving pool at the sampling port, and 37℃ pH 7.4 phosphate buffer was added as the receiving solution, with the liquid level at the scale line of the receiving chamber sampling port. Slowly release the bubbles, and check if there is any leakage.

[0305] Scheme A: the comparative example 1, F1-F5 were each diluted one time with pH 7.4 phosphate buffer as the supply chamber solution;

[0306] Option B: Dissect and collect intestinal tissue descending from the cecum. Cut the intestinal segment along the mesentery and place approximately 0.5g of intestinal contents into a 10ml sample tube. Add approximately 4mL of pH 7.4 phosphate buffer and gently vortex to mix (minimize air contact, as most intestinal flora are anaerobic). Take Comparative Example 1 and F1-F5 and dilute them by half with a suspension or paste containing rectal contents, respectively, as components for the supply chamber. For Comparative Example 2, weigh an appropriate amount of CCM-HPMCAS solid dispersion (containing approximately 20mg of CCM), add 5mL of pH 7.4 phosphate buffer, vortex to mix, and take 2mL of each. Dilute them by half with a rectal contents suspension and pH 7.4 phosphate buffer, respectively. Place both diluted suspensions into the supply chamber.

[0307] Set the circulating water bath temperature to 37℃, turn on the circulating water switch, and after maintaining the temperature and stirring for 5 minutes, add the supply liquid / contents that have been incubated at 37℃ to the supply tank, and start timing. Sampling times: 2 hours, 4 hours, 5 hours, 6 hours, and 7 hours;

[0308] Sampling volume: 500 μL, and simultaneously add an isothermal solution of the same volume.

[0309] Add one volume of methanol to the sample solution, vortex for 30 seconds, and centrifuge at 3500 rpm for 10 minutes. Take 20 μL of the supernatant and determine the CCM content by HPLC.

[0310] The cumulative transmittance Q (μg / cm³) at each sampling time point is calculated using the following formula. 2 Plot a graph with time (hr) on the x-axis and Q on the y-axis. Perform linear regression on the three consecutive points with the largest slope, and calculate T using the linear regression equation. lag and J max or J ss Plot the median time (hr) on the x-axis and J on the y-axis. The J after J reaches a steady state over time is taken as J. max or J ss Calculate the apparent permeability coefficient (P) based on J. app ,cm·s -1 )

[0311] J=dQ / dt (2)

[0312] P app = J / ( AC0) (3)

[0313] In the formula:

[0314] Cn: CCM concentration in the receiving chamber at the nth sampling point, μg·mL -1 ;

[0315] J: absorption rate, pg / (cm 2 ·hr);

[0316] A: effective membrane area (0.19625 cm 2 );

[0317] Co: initial concentration of CCM supplied to the supply chamber (pg·mL －1 ) in the supply chamber.

[0318] The results are as follows:

[0319] Table 16. Absorption rate J, apparent permeability P app of Comparative Example 1 and F1-F5

[0320] Each experimental group reached a steady state in 4-7 hours, so the J of each experimental group described above is J ss .

[0321] According to the above results, the absorption rate J and apparent permeability P app of Comparative Example 1, F1-F5 and the direct colon content group were significantly higher than those of the pH 7.4 phosphate buffer group, which indicates that the direct colon microflora and enzymes can gradually metabolize part of the sugar groups on the carrier and induce the reassembly of the metabolic products into nanocomplexes with better permeability. Compared with Comparative Example 2, the STVN-CCM nanocomplexes increased the apparent permeability P app of CCM by 10.8 times (pH 7.4 phosphate buffer) and 19.6 times (direct colon content). The apparent permeability P app of F1-F5 in the pH 7.4 phosphate buffer group was (2.11, 5.08, 7.58, 7.31 and 6.46) x 10 -6 (cm s -1 ), and the apparent permeability P app of F1-F5 in the direct colon content group was (16.22, 19.44, 24.76, 16.79 and 17.02) x 10 - 6 (cm s -1 ), which was significantly improved compared with the absorption rate of Comparative Example 1 + direct colon content group.

[0322] Microbial metabolism of each steviol glycoside derivative

[0323] Take the content of the supply chamber at the end of the 7 hour sampling of F2 prescription in Example 12, add 3 mL of 75% methanol-pH 3.0 phosphate buffer mixed solvent, vortex mix for 20 seconds, centrifuge at 13000 rpm for 5 minutes, take 5 μL of supernatant for tandem mass spectrometry analysis, according to the mass spectrometry scan results of LC / MS / MS metabolites, the following is analyzed:

[0324] Table 16. Microbial metabolic analysis in rat cecum contents

[0325] According to the detected mass spectrum signal, it is speculated that the above metabolic process is shown in Figures 27-28.

[0326] The above specific examples further illustrate the purpose, technical solutions and beneficial effects of the present application. It should be understood that the above description is only a specific embodiment of the present application and is not intended to limit the present application. Any modification, equivalent replacement, improvement, etc. made within the spirit and principles of the present application shall be included in the protection scope of the present application.

Claims

1. A steviol glycoside derivative characterized in that a compound of the formula Ia, or a salt thereof, or a hydrate thereof, or any mixture of a compound of the formula Ia, a salt thereof and a hydrate thereof, In formula Ia, R1, R2 are independently selected, R1 is selected from hydroxyl or glycosyl, R2 is selected from n-valent rhamnose, rhamnopyranose, mannose, manno-pyranose, galactose, galactopyranose, or a combination thereof, or an oligosaccharide formed by covalent linkage of other saccharides, wherein 1≤n≤10, for example n is 1, 2, 3, 4, 5, 6, 7, 8, 9, 10.

2. The steviol glycoside derivative of claim 1, having any one of the following characteristics: (1) the absolute configuration of the rhamnose, rhamnopyranose, mannose, manno-pyranose, galactose, galactopyranose is D or L, and the relative configuration of the terminal carbon is a or b; and (2) the absolute configuration of the rhamnose is L, and the relative configuration of the terminal carbon is a; the absolute configuration of the mannose is D, and the relative configuration of the terminal carbon is b; the absolute configuration of the galactose is D, and the relative configuration of the terminal carbon is b.

3. The steviol glycoside derivative of claim 1 or 2, characterized in that the glycosyl is selected from glucopyranose, galactopyranose, manno-pyranose, deoxyglucopyranose, rhamnopyranose, xylopyranose, glucosamino-pyranose, an oligosaccharide formed by polymerization of 1 to 9 arbitrary number of arbitrary pyranose monosaccharides at arbitrary positions.

4. The steviol glycoside derivative of any one of claims 1-3, wherein said R1 is selected from the group consisting of hydroxy, 5. The steviol glycoside derivative of any one of claims 1-4, wherein the optical purity of the compound of formula Ia is not limited, R and / or S configuration, with any enantiomeric excess value or any diastereomeric excess value.

6. A steviol glycoside derivative, which is at least one compound selected from the group consisting of: (1) (2S, 3S, 4S, 5S, 6R)-3, 4, 5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R, 4aS, 6aR, 9R, 11aR, 11bS)-9-hydroxy-4, 11-dimethyltetradecahydro-6a, 9-methanocyclohepta[a]naphthalene-4-carboxylate (STL-Man), (2) (2S, 3S, 4S, 5S, 6R)-3, 4, 5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R, 4aS, 6aR, 9S, 11aR, 11bS)-4, 11b-dimethyl-8-methylene-9-(((2S, 3R, 4S, 5S, 6R)-3, 4, 5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetradecahydro-6a, 9-methanocyclohepta[a]naphthalene-4-carboxylate (RBSD-Acid-Man), (3) (2S, 3S, 4S, 5S, 6R)-3, 4, 5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R, 4aS, 6aR, 9R, 11aR, 11bS)-9-hydroxy-4, 11-dimethyl-8-methylene-tetradecahydro-6a, 9-methanocyclohepta[a]naphthalene-4-carboxylate (STL-Man-8-Methylene), (4) (2S, 3S, 4S, 5S, 6R)-3, 4, 5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R, 4aS, 6aR, 9S, 11aR, 11bS)-4, 11b-dimethyl-8-methylene-9-(((2S, 3R, 4S, 5S, 6R)-3, 4, 5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetradecahydro-6a, 9-methanocyclohepta[a]naphthalene-4-carboxylate (RBSD-Acid-Man-8-Methylene), or a salt thereof, or a hydrate thereof, or an in vivo metabolite thereof. ​ ​ (3) (2S,3S,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-((2S,3R,4S,5S,6R)-4,5-dihydroxy-6-(hydroxymethyl)-3-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetrahydro-1 H-pyran-2-yl)oxy)-4,11 b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohepta[a]naphthalene-4-carboxylate (STVB-Man), (4) (2S,3S,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-((2S,3R,4S,5S,6R)-4,5-dihydroxy-6-(hydroxymethyl)-3-(((2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2-yl)oxy)-4,11 b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohepta[a]naphthalene-4-carboxylate (DCS-A-Acid-Man), (5) (2S,3S,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9R,11aR,11bS)-9-((2S,3R,4S,5R,6R)-5-hydroxy-6-(hydroxymethyl)-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-3-(((2S,3R,4S,5R)-3,4,5-trihydroxytetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2-yl)oxy)-4,11 b-dimethyltetradecahydro-6a,9-methanocyclohepta[a]naphthalene-4-carboxylate (RBDS-F-Acid-Man), (6) (2S,3S,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-(((2S,3R,4S,5R,6R)-5-hydroxy-6-(hydroxymethyl)-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-3-(((2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2-yl)oxy)-4,11b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohepta[a]naphthalene-4-carboxylate (DCS-B-Man), (7) (2S,3S,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-(((2S,3R,4S,5R,6R)-3-(((2S,3R,4R,5S,6S)-3,5-dihydroxy-6-methyl-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2-yl)oxy)-5-hydroxy-6-(hydroxymethyl)-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2-yl)oxy)-4,11b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohepta[a]naphthalene-4-carboxylate (RBDS-H-Acid-Man), (8) (2S,3S,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-(((2S,3R,4S,5R,6R)-5-hydroxy-6-(hydroxymethyl)-3,4-bis((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2-yl)oxy)-4,11b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohepta[a]naphthalene-4-carboxylate (RBDS-B-Man), (9) a steviol glycoside-Man derivative; (10) (2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9R,11aR,11bS)-9-hydroxy-4,11-dimethyltetradecahydro-6a,9-methanocyclohepta[a]naphthalene-4-carboxylate (STL-Rha), (11) (2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-4,11b-dimethyl-8-methylene-9-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetradecahydro-6a,9-methanocyclohepta[a]naphthalene-4-carboxylate (RBDS-A-Acid-Rha), naphthalene-4-carboxylate (RBSD-A-Acid-Rha), (12) (2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-(((2S,3R,4S,5S,6R)-4,5-dihydroxy-6-(hydroxymethyl)-3-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetrahydro-1H-pyran-2-yl)oxy)-4,11b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohepta[a]naphthalene-4-carboxylate (STVB-Rha), (13) (2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-(((2S,3R,4S,5S,6R)-4,5-dihydroxy-6-(hydroxymethyl)-3-(((2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2-yl)oxy)-4,11b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohepta[a]naphthalene-4-carboxylate (DCS-A-Acid-Rha), (14) (2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-(((2S,3R,4S,5R,6R)-5-hydroxy-6-(hydroxymethyl)-3,4-bis((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetrahydro-1H-pyran-2-yl)oxy)-4,11b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohepta[a]naphthalene-4-carboxylate (RBDS-B-Rha), (15) (2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-(((2S,3R,4S,5R,6R)-5-hydroxy-6-(hydroxymethyl)-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-3-(((2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2-yl)oxy)-4,11b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohexa[a]naphthalene-4-carboxylate (DCS-B-Rha), (16) (2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9R,11aR,11bS)-9-((2S,3R,4S,5R,6R)-5-hydroxy-6-(hydroxymethyl)-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-3-(((2S,3R,4S,5R)-3,4,5-trihydroxytetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2-yl)oxy)-4,11b-dimethyltetradecahydro-6a,9-methanocyclohexa[a]naphthalene-4-carboxylate (RBDS-F-Acid-Rha), (17) (2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-(((2S,3R,4S,5R,6R)-3-(((2S,3R,4R,5S,6S)-3,5-dihydroxy-6-methyl-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetrahydro-1H-pyran-2-yl)oxy)-5-hydroxy-6-(hydroxymethyl)-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetrahydro-1H-pyran-2-yl)oxy)-4,11b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohexa[a]naphthalene-4-carboxylate (RBDS-H-Acid-Rha), (18) a steviol-Rha derivative, ​ (19) (2S,3R,4S,5R,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-hydroxy-4,11b-dimethyl-8-methylenetetrahydro-6a,9- methanocyclohepta[a]naphthalene-4-carboxylate (STL-Gal), (20) (2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-4,11b-dimethyl-8-methylen-9-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetradecahydro-6a,9- methanocyclohepta[a]naphthalene-4-carboxylate (RBSD-Acid-Gal), (21) (2S,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-(((2S,3R,4S,5S,6R)-4,5-dihydroxy-6-(hydroxymethyl)-3-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetrahydro-1H-pyran-2-yl)oxy)-4,11b-dimethyl-8-methylenetetradecahydro-6a,9- methanocyclohepta[a]naphthalene-4-carboxylate (STVB-Gal), (22) (2S,3R,4S,5R,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-((2S,3R,4S,5R,6R)-5-hydroxy-6-(hydroxymethyl)-3,4-bis((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetrahydro-1H-pyran-2-yl)oxy)-4,11b-dimethyl-8-methylenetetradecahydro-6a,9- methanocyclohepta[a]naphthalene-4-carboxylate (RBDS-B-Gal), (23) (2S,3R,4S,5R)-3,4,5-trihydroxytetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-(((2S,3R,4S,5R,6R)-5-hydroxy-6-(hydroxymethyl)-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-3-(((2S,3R,4S,5R)-3,4,5-trihydroxytetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2-yl)oxy)-4,11b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohexa[a]naphthalene-4-carboxylate (DCS-A-Acid-Gal), (24) (2S,3R,4S,5R,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-((2S,3R,4S,5R,6R)-5-hydroxy-6-(hydroxymethyl)-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-3-(((2S,3R,4S,5R)-3,4,5-trihydroxytetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2-yl)oxy)-4,11b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohexa[a]naphthalene-4-carboxylate (DCS-B-Gal), (24) (2S,3R,4S,5R,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-((2S,3R,4S,5R,6R)-5-hydroxy-6-(hydroxymethyl)-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-3-(((2S,3R,4S,5R)-3,4,5-trihydroxytetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2-yl)oxy)-4,11b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohexa[a]naphthalene-4-carboxylate (DCS-B-Gal), (24) (2S,3R,4S,5R,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-((2S,3R,4S,5R,6R)-5-hydroxy-6-(hydroxymethyl)-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)-3-(((2S,3R,4S,5R)-3,4,5-trihydroxytetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2-yl)oxy)-4,11b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohexa[a]naphthalene-4-carboxylate (DCS-B-Gal), (26) (2S,3R,4S,5R,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl (4R,4aS,6aR,9S,11aR,11bS)-9-(((2S,3R,4S,5R,6R)-3-(((2S,3R,4R,5S,6S)-3,5-dihydroxy-6-methyl-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetrahydro-2H-pyran-2-yl)oxy)-5-hydroxy-6-(hydroxymethyl)-4-(((2S,3R,4S,5S,6R)-3,4,5-trihydroxy-6-(hydroxymethyl)tetrahydro-2H-pyran-2-yl)oxy)tetrahydro-1H-pyran-2-yl)oxy)-4,11b-dimethyl-8-methylenetetradecahydro-6a,9-methanocyclohepta[a]naphthalene-4-carboxylate (RBDS-H-Acid-Gal), (27) derivatives of steviol-Gal.

7. A method of preparing the steviol glycoside derivative of any one of claims 1-6, characterized in that comprising the following steps: (1) Step 1: subjecting steviol glycosides 1 or steviol to a hydrolysis reaction to obtain compound 2 or a combination, wherein when compound 1 is in carboxylic acid form, proceed directly to Step 2; (2) Step 2: protecting the R1 group in compound 2 or a combination with a protecting group P to obtain compound 3 or a combination; (3) Step 3: subjecting the activated sugar precursor to glycosidation reaction with compound 3 or a combination to obtain compound 4 or a combination; the activated sugar precursor is preferably a glycosyl halide, a glycosyl halide imidate ester; (4) Step 4: subjecting compound 4 or a combination to a deprotection reaction to obtain compound of structural formula 1a or a combination of compounds, wherein the structure of the steviol glycoside compound 1 is R1in the foregoing Formula 2 is selected from the group consisting of hydroxyl, or a combination.

8. The method of claim 7, wherein The hydrolysis reaction is a reaction in a solvent (preferably water) under basic conditions (preferably sodium hydroxide or potassium hydroxide or lithium hydroxide), wherein compound 1 is selected from stevioside (STVS), rebaudioside E (RBDS-E), rebaudioside A (RBDS-A), dulcoside A (DCS-A), rubusoside (RBSD), acid of rubusoside (RBSD-Acid), steviolbioside (STVB), rebaudioside B / D / M / I / O / N / J (RBDS-B / D / M / O / N / J), rebaudioside C / K (RBDS-C / K), dulcoside B (DCS-B), rebaudioside F (RBDS-F), steviol (STL) or isosteviol (ISTL), or a combination of any one or more of the above; when compound 1 is any one or more of steviolbioside (STVB), rebaudioside B (RBDS-B), dulcoside B (DCS-B), steviol, the preparation method does not need step 1.

9. The production method according to claim 7 or 8, characterized by The protecting group in step 2 is a protecting group for sugar or hydroxyl group, preferably silyl or acetyl, preferably the sugar or hydroxyl group is fully acetylated protected, for example the protected compound 3 is selected from RBDS-B-Ac, STVB-Ac, RBSD-Acid-Ac, DCS-A-Acid-Ac, RBDS-F-Acid-Ac, RBDS-H-Acid-Ac, DCS-B-Ac or STL-Ac or a combination of the above, the structural formulae are shown as follows, respectively, Wherein: Ac represents acetyl.

10. The method of any one of claims 7-9, wherein: (1) the glycosylation reaction of step 3 is a reaction of compound 2 with an activated sugar precursor of the same structure as the defined R2substituent to obtain compound 4; the preferred activated sugar precursor is a glycosyl halide, a glycosyl halogenated imidazole ester, and / or (2) when the protecting group P is acetyl, the step 4 is a deprotection reaction using an alcohol solvent (such as methanol or ethanol) under strong base conditions (such as potassium tert-butoxide or sodium methoxide or sodium ethoxide).

11. A pharmaceutical composition or delivery kit characterized in that The steviol glycoside derivative, the delivery carrier and the active ingredient according to any one of claims 1-6.

12. The pharmaceutical composition or delivery kit according to claim 11, having any one of the following features: (1) the active ingredient is a bioactive agent, a chemical active agent or an adjuvant; (2) the active ingredient is selected from the group consisting of: a protein / polypeptide, a polysaccharide / oligosaccharide, a nucleic acid or nucleic acid fragment, a nanoparticle (such as a liposome, a supramolecular nanocomplex, a polymeric micelle, an inorganic nanoparticle), a lipid, a nutritional element, an organic small molecule compound, a bacteriophage particle, a superparamagnetic substance, a vaccine, a cell, or any combination thereof; (3) the delivery carrier is selected from the group consisting of: one or more of a lipid, a lecithin, a cholesterol and its derivatives, a glycoside and its derivatives, a soy protein, an apolipoprotein, an inorganic nanocarrier, a polymer, a glycolipid; (4) the mass ratio of the active ingredient to the sum of (steviol glycoside derivative + delivery vehicle) is 1:0.5 to 1:25 (preferably 1:0.75, 1:1, 1:1.5, 1:2, 1:2.5, 1:3, 1:5, 1:10, 1:15, 1:20, 1:25), the steviol glycoside derivative accounts for 5% to 100% of the total mass of (steviol glycoside derivative + delivery vehicle), preferably 7% to 100% (preferably 7% to 70%, 7% to 65%, 10% to 60%, 12.5 to 100%, 15% to 95%, 17.5% to 90%, 20% to 80%, 25% to 75%, 30% to 70%, 35% to 65%, 40% to 60%); (5) the active ingredient, steviol glycoside derivative, and delivery vehicle are present independently or in admixture; (6) the pharmaceutical composition or delivery kit is in a form suitable for oral, sublingual, buccal, cavity mucosal (e.g., esophageal, gastric, duodenal, colonic, rectal, vaginal, cervical, anal), subcutaneous, intramuscular, intravenous, intraarterial, dermal, pulmonary, pharyngeal, tracheal, bronchial, intranasal, otic, ocular administration; (7) the active ingredient, steviol glycoside derivative, and / or delivery vehicle are assembled in micelles, liposomes, nanoparticles, microspheres, microcapsules, solid dispersions, molecular assemblies, or hydrogels; (8) when the active ingredient, steviol glycoside derivative, and delivery vehicle are present independently, they are assembled in situ upon mixing and dissolution; when the active ingredient, steviol glycoside derivative, and delivery vehicle are present in admixture, the active ingredient, steviol glycoside derivative, and delivery vehicle are assembled upon dissolution from a solid state; (9) the active ingredient, steviol glycoside derivative, and delivery vehicle are assembled in situ; (10) the active ingredient, steviol glycoside derivative, and delivery vehicle are self-assembled in the gastrointestinal tract, locally in the mucosa, in the blood, in the lymphatic vessels or lymph nodes, intrathecally, or in target cells; (11) the active ingredient, steviol glycoside derivative, and delivery vehicle are assembled in situ, wherein the in situ assembly is the metabolic product of the steviol glycoside derivative upon enzymatic or microbial metabolism and the active ingredient and / or delivery vehicle or their metabolites; (12) the pharmaceutical composition or delivery kit is in a dosage form selected from the group consisting of tablets, capsules, oral solutions, drops, gels, granules, emulsions, creams, injections, eye drops, inhalers, sprays, aerosols, or patches.

13. The pharmaceutical composition or delivery kit of claim 11 or 12, further comprising a polymer, preferably the polymer has groups that dissociate under physiological conditions and / or has more than 10 groups that provide hydrogen donors or hydrogen acceptors, more preferably the polymer is selected from the group consisting of: ​ (1) Cellulosic polymers such as hydroxypropyl methylcellulose (HPMC), low-substituted hydroxypropyl cellulose (L-HPC), hydroxypropyl methylcellulose acetate succinate (HPMCAS), sodium carboxymethylcellulose (CMC-Na), or hydroxyethyl cellulose; (2) Synthetic polymers: such as polyethylene glycol (PEG), polyvinyl alcohol (PVA), polyvinylpyrrolidone (PVP), copovidone (PVP-VA64), polyvinyl acetate-polyethylene glycol graft copolymer (Soluplus), polyglutamic acid (PGA), polydopamine (PDA), polylactic acid (PLA), poly-L-lactic acid (PLLA), poly(lactic-co-glycolic acid) (PLGA), sodium polyacrylate, siRNA and derivatives (improved lipophilic alkyl chain), mRNA and derivatives (improved lipophilic alkyl chain), or antisense oligonucleotides and derivatives (improved lipophilic alkyl chain); (3) Polysaccharides and derivatives thereof (such as carboxymethylation, sulfonylation, phosphorylation, acylation, or hydroxypropylation, cationic, anionic, or other derivatives), such as carrageenan, guar gum, gum arabic, locust bean gum, konjac gum, agar, gelatin, pectin, gellan gum, hyaluronic acid (HA), aminodextran, chondroitin sulfate, dermatan sulfate, heparin, keratan sulfate, heparan sulfate, sodium alginate, propylene glycol alginate, agar, fucoidan, cyclodextrin and derivatives, chitosan and derivatives (such as acylation, carboxylation, alkylation, and quaternization, etc.); and (4) Proteins and polypeptides: soy protein, vegetable protein, or bovine serum albumin, casein; and (5) Any combination of (1)-(4).

14. The pharmaceutical composition or delivery kit according to any one of claims 11-13, further comprising an additive selected from the group consisting of an excipient, a diluent, a binder, a disintegrant, a lubricant, a flavoring agent, a pH adjuster, an osmotic pressure adjuster, a thickening agent, a plasticizer, a coloring agent, a film-forming agent, a preservative, or a solvent, and any combination thereof.

15. The pharmaceutical composition or delivery kit according to any one of claims 11-14, for use in a group consisting of: (1) for delivery of an active ingredient to a mammal (such as a rodent, a cow, a pig, a dog, a cat, a primate, a human) or a bird (such as a chicken, a duck, a goose); (2) for the regulation of the release of an active ingredient or a nutritional ingredient during the delivery of the active ingredient or the nutritional ingredient to a mammal (such as a rodent, a cow, a pig, a dog, a cat, a primate, a human) or a bird (such as a chicken, a duck, a goose); (3) for use as a solubilizer in agrochemistry, especially in formulations containing a pesticide, a herbicide, a fungicide, or an insecticide, especially as a solubilizer for spray, drench, or dip mixtures of crop protection agents; (4) for use as a preservative or a preservative or a flavoring agent for meat, vegetables, fruits, food, beverages; (5) for use in cosmetics, fine chemical products for solubilization, stability improvement, penetration promotion; (6) for use as an adjuvant for biological agents. ​

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