A phosphatidylcholine structuring compound and a method for its preparation

Abstract

The present application relates to a kind of phospholipid choline structural compounds and its preparation method, the preparation method of the phospholipid choline structural compounds includes the following steps: with hydroxyethyl methacrylate and beta-mercaptoethylamine as raw material, intermediate M1 is obtained by reaction at room temperature;Intermediates M1 and acrylate are added to obtain intermediate M2;Intermediate M2 and 2-chloro-2-oxygen-1,3,2-dioxaphospholane are reacted to obtain intermediate M3;Then intermediate M3 and chain or cyclic tertiary amine are reacted to obtain phospholipid choline compound.The phospholipid choline compound obtained by the method of the present application is simple, mild, can be combined with many compound fragments on the market, and the product is rich and varied.Such phospholipid choline compound can be selectively delivered to the spleen site for the delivery of nucleic acid drugs.

Description

Technical Field

[0001] The present invention belongs to the technical field of medicinal chemistry, and specifically relates to a series of phosphatidylcholine structural compounds, a preparation method, and the application of the phosphatidylcholine compounds in targeted nucleic acid drug delivery. Background Art

[0002] Phospholipids are essential components of biological membranes. Their inherent hydrophilic / lipophilic properties allow them to spontaneously form a closed bilayer in water, serving as the membrane's backbone. Leveraging this property, Lehman et al. in the UK began using phospholipids as drug carriers in 1971. Since then, liposomes have attracted worldwide attention as a new formulation for targeted drug delivery systems. These liposomes offer advantages such as minimal toxicity and side effects, good tissue compatibility between their lipid bilayer and biological membranes, and ease of tissue absorption.

[0003] Phospholipids play an important role in liposomes, but the application of natural phospholipids has certain limitations due to their shortcomings such as susceptibility to oxidation and poor stability. Synthetic phospholipids have strong antioxidant capacity and are more ideal for preparing liposomes, which has important application significance in the field of drug carriers. Phosphatidylcholine is an amphiphilic molecule composed of a hydrophilic head and a hydrophobic tail. It is a type of phospholipid with a choline group inserted in the head. Lipid nanoparticles are one of the main carriers for the effective delivery of nucleic acid drugs. They can protect nucleic acid drugs from rapid degradation in the body, prolong their circulation time in the body, and enable more effective delivery of nucleic acid drugs into target cells. Among them, auxiliary phospholipids play a key role in the delivery of nucleic acid drugs. Therefore, the development of new auxiliary phospholipids is also of great significance in the research of nucleic acid drugs. Currently, two common choline compounds are distearoylphosphatidylcholine (DSPC) and dipalmitoylphosphatidylcholine (DPPC).

[0004] While synthetic phospholipids overcome the shortcomings of natural phospholipids, they also suffer from disadvantages such as rapid metabolism and a shorter duration of efficacy. The derivatization of synthetic phospholipids has advanced liposome technology in terms of targeting and long-circulation. Given the widespread application of DSPC and DPPC in liposome research and the increasing market demand, the development of novel synthetic methods for phosphatidylcholine structures is of paramount importance.

[0005] Patent (CN111057099A) discloses dipalmitoylphosphatidylcholine (DPPC) and its preparation method. This method uses the condensation of GPC and palmitic acid to produce DPPC. This method is limited to the preparation of DPPC and is unlikely to enrich the product portfolio.

[0006] Patent (CN114213458A) discloses a cyclobutylamine-terminated distearoylphosphatidylcholine and its preparation method. This method is limited to the synthesis of cyclobutylamine-terminated compounds, has significant limitations, and has a limited coverage.

[0007] Therefore, there is an urgent need in this field to develop a series of novel phosphatidylcholine preparation methods so as to better meet the market demand for phosphatidylcholine compounds. Summary of the Invention

[0008] The purpose of the present invention is to provide a series of phosphatidylcholine structural compounds and their preparation methods and applications. The novel phosphatidylcholine is mild in reaction during the synthesis process, simple to operate, and easy to purify. In addition, it has abundant synthetic intermediates and can be combined with a variety of chain or cyclic tertiary amines and ester groups to differentially synthesize various types of phosphatidylcholine compounds, which is conducive to enriching the phosphatidylcholine product library. In order to solve the above technical problems, the present invention provides the following technical solutions:

[0009] The present invention provides a phosphatidylcholine compound having a structure described in formula (I) or formula (II), and a pharmaceutically acceptable salt, tautomer or stereoisomer of the compound.

[0010] ,

[0011] wherein X is selected from -O- or -S-;

[0012] Ra is selected from C 3-8 Alkylene, wherein the carbon atoms in the alkylene are optionally substituted by 1-3 -O-, -S-, -NH-, provided that the -O-, -S-, -NH- and X groups or R1R2N- groups are not directly connected, and the -O-, -S-, -NH- groups are not directly connected to each other; optionally, one or more hydrogens in the Ra are replaced by Rb, and the Rb is selected from C 1-8 alkyl.

[0013] is a chain quaternary amine or a cyclic quaternary amine, R3, R4, and R5 are each independently selected from C 1-8 A branched or branched alkyl group, or a N atom, one of (R3, R4, R5) and its adjacent one together form a 5-membered or 6-membered heterocyclic ring; optionally, the 5-membered or 6-membered heterocyclic ring is substituted by one or more R6, wherein R6 is selected from C 1-6 A straight chain or branched chain alkyl group.

[0014] R1 and R2 are each independently selected from -H, C 2-8 A straight chain or branched alkyl group, the C 2-8The methylene contained in the straight or branched alkyl is optionally replaced by -S-, -O-, or -NH-; one or more hydrogen atoms of R1 and R2 are optionally replaced by -COOR7, -OCOR7, -COSR7, -SCOR7, -CONR7R8, or -NR7COR8; and R7 and R8 are each independently selected from -H, C 1-20 branched or straight chain alkyl, C 1-20 branched or straight chain alkenyl, C 1-20 wherein the branched or straight chain alkynyl group is selected from the group consisting of: -H, ...

[0015] Preferably, R1 and R2 are selected from -H or a group of the following structure, and R1 and R2 are not -H at the same time;

[0016] ;

[0017] More preferably, Has the following structure:

[0018] ,

[0019] The present invention provides a phosphatidylcholine compound of the structure described in formula (Ia) or (IIa), and a pharmaceutically acceptable salt, tautomer or stereoisomer of the compound, wherein R1, R2, R3, R4, R5, X and Rb are as defined above;

[0020] .

[0021] More preferably, the compound of formula (I) of the present invention has the structure shown below:

[0022] ,

[0023] ,

[0024] ,

[0025] ,

[0026] ,

[0027] ,

[0028] ,

[0029] ,

[0030] ;

[0031] Furthermore, the present invention also provides a use of a phosphatidylcholine compound of the structure described in formula (I), a pharmaceutically acceptable salt, tautomer or stereoisomer of the compound as a drug carrier. Preferably, the drug is selected from nucleic acid drugs.

[0032] Furthermore, the present invention also provides a method for preparing the phosphatidylcholine compound of the structure described in formula (Ia), the pharmaceutically acceptable salt, tautomer or stereoisomer of the compound, the method comprising the following steps:

[0033] S1. Hydroxyethyl methacrylate and β-mercaptoethylamine are reacted in a solvent at room temperature to obtain intermediate M1; the reaction formula is:

[0034] ;

[0035] S2. Intermediate M1 and An addition reaction or a substitution reaction is carried out to obtain the intermediate M2; the reaction formula is:

[0036] , said R9 is selected from C 0-6 The straight or branched alkyl group, the methylene group in R9 is optionally replaced by -S-, -O-, or -NH-, one or more hydrogen atoms in R9 are optionally replaced by -COOR7, -OCOR7, -CONR7R8, -NR7COR8, -COSR7, or -SCOR7, and R7 and R8 are each independently selected from -H, C 1-20 branched or straight chain alkyl, C 1-20 branched or straight chain alkenyl, C 1-20 a branched or straight-chain alkynyl group,

[0037] S3. Intermediate M2 reacts with 2-chloro-2-oxo-1,3,2-dioxaphospholane to obtain intermediate M3; the reaction formula is:

[0038] ;

[0039] S4. Intermediate M3 and Reaction to obtain the phosphatidylcholine compound shown in formula Ia; the reaction formula is:

[0040] ;

[0041] The definitions of R1, R2, R3, R4 and R5 are as described above.

[0042] Preferably, in step S1, the molar ratio of SM1 to SM2 should be between 1:1 and 1.2, and the optimal molar ratio is 1:1.

[0043] Preferably, in step S1, the reaction temperature should be between 10 and 30° C., and the reaction time should be 3 to 6 hours.

[0044] Preferably, in step S1, the solvent is anhydrous ethanol, and the volume ratio of hydroxyethyl methacrylate to the solvent is 1:5-10 (g / ml).

[0045] Preferably, in step S1, the post-treatment operation is: the reaction solution is concentrated under reduced pressure to remove the solvent to obtain a colorless oily crude product, which is directly used in the next reaction without further purification.

[0046] Preferably, in step S2, the molar ratio of M1 to SM3 should be between 1:2.2 and 3, and the optimal molar ratio is 1:2.5.

[0047] Preferably, in step S2, a solvent-free reaction is adopted, the reaction temperature should be between 70 and 80° C., and the reaction time should be between 40 and 60 hours.

[0048] Preferably, in step S2, 0.05 equivalents of BHT should be added as a reaction stabilizer (the reaction stabilizer can prevent polymerization from occurring during the reaction process).

[0049] Preferably, in step S2, 0.05 equivalents of a catalyst are added, and the catalyst is preferably acetic acid or boric acid.

[0050] Preferably, in step S3, the reaction occurs under the action of an acid binding agent, and the molar ratio of M2 to SM4 and the acid binding agent is 1:2:2.5. The acid binding agent is selected from an organic base or an inorganic base. Preferably, the organic base is selected from but not limited to one or more of triethylamine, diethylisopropylamine, pyridine, DMAP, and DBU, and the inorganic base is selected from one or more of sodium carbonate, potassium carbonate, sodium bicarbonate, and potassium bicarbonate.

[0051] Preferably, in step S3, the reaction is carried out in an organic inert aprotic solvent, preferably one or more of dry dichloromethane, tetrahydrofuran, DMF, DMSO, acetone, and dioxane, and the mass volume ratio of substrate to solvent is 1:10 (g / ml).

[0052] Preferably, in step S3, the reaction temperature should be between 0 and 10° C., and the reaction time should be between 1 and 3 hours.

[0053] Preferably, in step S3, there are two post-processing operations. The first post-processing operation is: due to the poor stability of the intermediate, it cannot be purified, so the reaction solution is directly concentrated under reduced pressure to remove the solvent to obtain a milky white oily crude product. After beating with ethyl acetate to obtain a white suspension, the solid is filtered to remove the mother liquor, and the crude product is directly used in the next reaction without further purification. The product has poor stability and cannot be characterized. If the acid binding agent used in step S3 is the same as the tertiary amine used in step S4, the second post-processing operation is adopted: the reaction solution does not need further post-treatment, and the reaction solution is directly used for step S4 feeding.

[0054] Preferably, in step S4, M3 and The molar ratio is 1:1.5~5, preferably 1:3.

[0055] Preferably, in step S4, the reaction is carried out in a solvent selected from one or more of dry acetonitrile, tetrahydrofuran, DMF, DMSO, acetone, and dioxane. The mass-to-volume ratio of substrate to solvent is 1:5-10 (g / ml).

[0056] Preferably, in step S4, the reaction temperature should be between 50 and 55° C., and the reaction time should be 20 to 50 hours.

[0057] Preferably, in step S4, the post-treatment operation is: the reaction solution is concentrated under reduced pressure to remove the solvent to obtain a pale yellow oily crude product, and the crude product is directly purified by column chromatography to obtain the target product.

[0058] Furthermore, the present invention provides a composition comprising a phosphatidylcholine compound having a structure described in the above formula (I) or formula (II), or a pharmaceutically acceptable salt, tautomer or stereoisomer of the compound.

[0059] Furthermore, the present invention provides the above composition, which further comprises other lipids.

[0060] Furthermore, the present invention provides the above composition, wherein the other lipids are selected from , cholesterol, DMG-PEG2000 or one or more.

[0061] Furthermore, the present invention provides the above composition, wherein the other lipids are selected from , cholesterol, DMG-PEG2000 or one or more.

[0062] Furthermore, the present invention provides the above composition, wherein the other lipids are selected from , cholesterol, DMG-PEG2000 or one or more.

[0063] Furthermore, the present invention provides the above composition, wherein the other lipids are selected from , cholesterol, DMG-PEG2000 or one or more.

[0064] Furthermore, the present invention provides the above composition, wherein the other lipids are selected from , cholesterol, DMG-PEG2000 or one or more.

[0065] Furthermore, the present invention provides the above-mentioned composition, wherein the molar percentages of the other lipids, the phosphatidylcholine compound of the structure described in formula (I) or formula (II), cholesterol, and DMG-PEG2000 are (40%~60%): (5%~15%): (22%~54.5%): (0.5%~1.5%).

[0066] Furthermore, the present invention provides the above composition, which further comprises cholesterol and DMG-PEG2000.

[0067] The C of the present invention 1-6 or C 1-8 The alkyl group is preferably selected from methyl, ethyl, n-propyl, isopropyl, n-butyl, 1-methylpropyl, 2-methylpropyl, tert-butyl, n-pentyl, n-hexyl, n-heptyl, n-octyl, .

[0068] The C of the present invention 1-20 branched or straight chain alkyl, C 1-20 branched or straight chain alkenyl, C 1-20 The number of carbon atoms of the branched or straight-chain alkynyl group is preferably 6-20, 8-20, 10-20, 12-20, 14-20, 16-20 or 18-20.

[0069] The alkyl group mentioned in the present invention refers to an alkane with a corresponding number of carbon atoms in which one hydrogen is replaced by another group, and the alkylene group refers to an alkane with a corresponding number of carbon atoms in which two hydrogens are replaced by other groups, such as the structure of a methylene group is -CH2-, and the structure of an ethylene group is -CH2CH2-.

[0070] The "optionally substituted" described in the present invention means that the substitution may or may not occur.

[0071] Compared with the prior art, the present invention has the following beneficial effects:

[0072] The synthesis process of the invention has mild reaction, simple operation and easy purification.

[0073] The present invention can differentially synthesize various types of phosphatidylcholine compounds, which is beneficial to enriching the phosphatidylcholine product library and providing many options and directions for the application of liposomes.

[0074] The phosphatidylcholine compound obtained by the method of the present invention has a simple reaction and mild conditions, and can be combined with numerous compound fragments on the market to produce rich and diverse products. Such phosphatidylcholine compounds can be used to encapsulate the delivery of nucleic acid drugs and can be selectively delivered to the spleen. BRIEF DESCRIPTION OF THE DRAWINGS

[0075] Figure 1 Schematic diagram of the reaction route for the synthesis of the phosphatidylcholine compound of the present invention;

[0076] Figure 2 Fluci expression in HEK-293T cells;

[0077] Figure 3 Fluci expression in Huh7 cells;

[0078] Figure 4 Formula 1 LNP and SM-102 LNP formulations for Compound (1) and Compound (4) lipids were expressed in Fluci;

[0079] Figure 5 Fluci expression levels in various organs of compound (1) and compound (4) lipid formulations 1 LNP and SM-102 LNP;

[0080] Figure 6 Fluci expression in the liver and spleen of the LNP formulation of Formula 1 containing compound (2), compound (13), compound (24), compound (5), compound (16) and compound (27);

[0081] Figure 7 The expression levels of Fluci in various organs of lipid formulation 1 LNP preparations of compound (2), compound (5), compound (13) and compound (16) are shown. DETAILED DESCRIPTION

[0082] In order to make the technical means, creative features, objectives and effects achieved by the present invention easier to understand, the present invention is further described below in conjunction with specific implementation methods.

[0083] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art to which this invention pertains. The terms used in this specification of the present invention are for the purpose of describing specific embodiments only and are not intended to limit the present invention. The term "and / or" as used herein includes any and all combinations of one or more of the associated listed items.

[0084] The present invention will be further described below with reference to the accompanying drawings and specific embodiments so that those skilled in the art can better understand the present invention and implement it. However, the embodiments are not intended to limit the present invention.

[0085] Unless otherwise specified, the experimental methods used in the following examples are conventional methods, and the materials and reagents used are all commercially available unless otherwise specified.

[0086] Example 1 Synthesis of a novel phosphatidylcholine compound (1)

[0087] The structural formula of the novel phosphatidylcholine compound (1) is as follows:

[0088] ;

[0089] Step S1:

[0090] ,

[0091] Under nitrogen, hydroxyethyl methacrylate (5.0 g, 1.0 eq.) and anhydrous ethanol (50 mL, 10V) were added sequentially to the reaction flask at room temperature. Mercaptoethylamine (2.96 g, 1.0 eq.) was added with stirring, and the mixture was stirred at room temperature for 3 h. TLC analysis (PE / DCM = 1 / 1) indicated complete consumption of the hydroxyethyl methacrylate starting material. The reaction solution was concentrated under reduced pressure to remove the solvent, yielding a colorless oily crude product (8.0 g, crude product), which was used directly in the next reaction. The synthesized intermediate compound M1 was subjected to nuclear magnetic resonance analysis, and its characterization data were as follows: 1H NMR (400 MHz, Chloroform-d) δ 4.31 (ddd, J =11.7, 5.3, 3.8 Hz, 1H), 4.22 – 4.15 (m, 1H), 3.81 (ddd, J = 5.3, 3.8, 1.2 Hz, 2H), 2.90 – 2.64(m,8H), 1.27 (d, J = 6.7 Hz, 3H).

[0092] Step S2:

[0093] ,

[0094] M1 (1.0 g, 1.0 eq.), SM3 (containing 0.05% BHT stabilizer) (3.87 g, 2.5 eq.), and acetic acid (14.49 mg, 0.05 eq.) were added sequentially to a reaction flask at room temperature. The mixture was heated to 70°C with stirring and reacted for 40 h. TLC (DCM / MeOH = 20 / 1) was performed to confirm that the starting materials were essentially consumed. The reaction solution was directly purified by silica gel column chromatography (DCM / MeOH = 1 / 0-100 / 1). 2.7 g of the desired product was obtained as a light yellow oil in a 65.97% yield. The synthesized intermediate compound M2 was subjected to nuclear magnetic resonance analysis, and its characterization data are as follows: 1H NMR (400 MHz, Chloroform-d) δ 5.43 – 5.27 (m, 8H),4.36 – 4.28 (m, 1H), 4.23 – 4.17 (m, 1H), 4.06 (t, J = 6.8 Hz, 4H),3.83 (t, J= 4.6 Hz, 2H), 2.86 – 2.57 (m, 16H), 2.44 (t, J = 7.2 Hz, 4H), 2.05 (q, J =6.8 Hz, 8H), 1.62 (p, J = 6.8 Hz, 4H),1.40 – 1.22 (m, 37H), 0.93 – 0.84 (m,6H). The synthesized intermediate compound M2 was subjected to high-resolution mass spectrometry analysis, and its characterization data are as follows: MS (m / z): [M+H] + = 848.69978.

[0095] Step S3:

[0096] ,

[0097] M2 (500 mg, 1.0 eq.) and anhydrous DCM (5 mL, 10 V) were added to the reaction flask in sequence. The mixture was cooled to 4°C in an ice bath, and 2-chloro-2-oxo-1,3,2-dioxaphospholane (167.95 mg, 2.0 eq.) was added. After the addition was complete, the mixture was stirred for 5 minutes. Triethylamine (149.10 mg, 2.5 eq.) was slowly added dropwise. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material 1 hour after the addition, indicating the reaction was complete. The reaction solution was concentrated under reduced pressure to remove the DCM, and EA (5 mL, 10 V) was added. The mother liquor was filtered and collected, and then concentrated under reduced pressure to yield 600 mg of a colorless oil. The crude product was unstable and was used directly in the next reaction without characterization.

[0098] Step S4:

[0099] ,

[0100] Under nitrogen, M3 (150 mg crude product, 1.0 eq.), anhydrous acetonitrile (0.75 mL, 5 V), and trimethylamine (2M in THF) (27.87 mg, 3.0 eq.) were added sequentially to the reaction flask at room temperature. The temperature was slowly raised to 50°C and stirred for 30 h. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly purified by silica gel column chromatography (DCM / MeOH = 1 / 0 to 5 / 1). The desired product was obtained as a white waxy product (33 mg) in a 20.72% yield. The synthesized product was subjected to nuclear magnetic resonance analysis and its characterization data are as follows: 1H NMR (400 MHz, Chloroform-d) δ 5.28 (dtt, J = 17.8,11.4, 5.1 Hz, 8H), 4.23 (dt, J = 10.2, 5.2 Hz, 2H), 3.99 (t, J = 6.8 Hz, 6H),3.92 – 3.83 (m, 2H), 3.66 (t, J = 4.2 Hz, 2H), 2.92 – 2.37 (m, 19H), 1.98 (q,J = 6.8 Hz, 8H), 1.54 (q, J =6.9 Hz, 4H), 1.34 – 1.11 (m, 39H), 0.82 (t, J =6.7 Hz, 6H). The synthesized product was subjected to high-resolution mass spectrometry analysis, and its characterization data are as follows: MS (m / z): [M+H] + =1013.67236.

[0101] Example 2 Synthesis of a novel phosphatidylcholine compound (2)

[0102] The structural formula of the novel phosphatidylcholine compound (2) is as follows:

[0103] ,

[0104] Step S4:

[0105] ,

[0106] Under nitrogen, M3 (600 mg crude product, 1.0 eq.), anhydrous acetonitrile (3.0 mL, 5V), and triethylamine (127.24 mg, 2.0 eq.) were added sequentially to the reaction flask at room temperature. The temperature was slowly raised to 50°C and stirred for 30 h. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly purified by silica gel column chromatography (DCM / MeOH = 1 / 0 to 5 / 1). The desired product (148 mg) was obtained as a white waxy product in a 22.30% yield. The synthesized product was subjected to nuclear magnetic resonance analysis and its characterization data are as follows: 1H NMR (400 MHz, Chloroform-d) δ 5.44 – 5.26 (m, 8H), 4.29 (t, J =4.8 Hz, 2H), 4.05 (t, J = 6.8 Hz, 6H), 3.95 (q, J = 7.1, 5.1 Hz,2H), 3.73 (t,J = 4.1 Hz, 2H), 2.92 – 2.58 (m, 15H), 2.50 (t, J = 7.2 Hz, 4H), 2.05 (q, J =6.8 Hz, 8H), 1.62 (p, J = 6.7Hz, 4H), 1.40 – 1.19 (m, 42H), 0.93 – 0.77 (m,6H). The synthesized product was subjected to high-resolution mass spectrometry analysis, and its characterization data are as follows: MS (m / z): [M+H] + =1055.75450.

[0107] Example 3 Synthesis of a novel phosphatidylcholine compound (3)

[0108] The structural formula of the novel phosphatidylcholine compound (3) is as follows:

[0109] ,

[0110] Step S4:

[0111] ,

[0112] Under nitrogen, M3 (150 mg crude product, 1.0 eq.), anhydrous acetonitrile (0.75 mL, 5V), and DIPEA (60.94 mg, 3.0 eq.) were added sequentially to a reaction flask at room temperature. The temperature was slowly raised to 50°C and stirred for 30 h. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly purified by silica gel column chromatography (DCM / MeOH = 1 / 0 to 5 / 1). The desired product was obtained as a white waxy product (63 mg) in a 36.99% yield. The synthesized product was subjected to nuclear magnetic resonance analysis and its characterization data were as follows: 1H NMR (400 MHz, Chloroform-d) δ 5.44 – 5.28 (m, 8H),4.40 – 4.22 (m,2H), 4.12 – 3.91 (m, 8H), 3.74 (t, J = 4.1 Hz, 2H), 3.00 – 2.50 (m, 20H),2.05 (q, J = 6.8 Hz, 9H), 1.61 (q, J = 7.0 Hz, 5H), 1.31 (ddt, J = 22.4,20.0, 6.5 Hz, 47H), 0.94 – 0.76 (m, 12H).

[0113] Example 4 Synthesis of a novel phosphatidylcholine compound (4)

[0114] The structural formula of the novel phosphatidylcholine compound (4) is as follows:

[0115] ,

[0116] Step S4:

[0117] ,

[0118] Under nitrogen, M3 (150 mg crude product, 1.0 eq.), anhydrous acetonitrile (0.75 mL, 5V), and N,N-diethylmethylamine (46.24 mg, 3.0 eq.) were added sequentially to the reaction flask at room temperature. The temperature was slowly raised to 50°C and stirred for 30 h. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly purified by silica gel column chromatography (DCM / MeOH = 1 / 0 to 5 / 1). The desired product was obtained as a white waxy substance (60 mg) in a 32.58% yield. The synthesized product was subjected to nuclear magnetic resonance analysis, and its characterization data are as follows: 1H NMR (400 MHz, Chloroform-d) δ 5.45 – 5.26 (m, 8H), 4.41 –4.22 (m, 2H), 4.13 – 3.92 (m, 8H), 3.74 (t, J = 4.2 Hz, 2H), 2.99 – 2.51 (m,20H), 2.05 (q, J = 6.9 Hz, 9H), 1.62 (t, J = 7.0 Hz, 4H), 1.39 – 1.20 (m,42H), 0.89 (t, J = 6.8 Hz, 6H). The synthesized product was subjected to high-resolution mass spectrometry analysis, and its characterization data are as follows: MS (m / z): [M+H] + = 1041.80881.

[0119] Example 5 Synthesis of a novel phosphatidylcholine compound (5)

[0120] The structural formula of the novel phosphatidylcholine compound (5) is as follows:

[0121] ,

[0122] Step S4:

[0123] ,

[0124] Under nitrogen, M3 (150 mg crude product, 1.0 eq.), anhydrous acetonitrile (0.75 mL, 5V), and N-methyldipropylamine (54.33 mg, 3.0 eq.) were added sequentially to the reaction flask at room temperature. The temperature was slowly raised to 50°C and stirred for 30 h. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly purified by silica gel column chromatography (DCM / MeOH = 1 / 0 to 5 / 1). The desired product was obtained as a white waxy product (58 mg, 34.50% yield). The synthesized product was subjected to nuclear magnetic resonance analysis and its characterization data are as follows: 1H NMR (400 MHz, Chloroform-d) δ 5.28 (dtt, J = 17.8, 11.4,5.1 Hz, 8H), 4.23 (dt, J =10.2, 5.2 Hz, 2H), 3.99 (t, J = 6.8 Hz, 6H), 3.92 –3.83 (m, 2H), 3.66 (t, J = 4.2 Hz, 2H), 2.92 – 2.37 (m, 19H), 1.98 (q, J =6.8 Hz, 8H), 1.54 (q, J = 6.9 Hz, 4H), 1.34 – 1.11 (m, 39H),0.82 (t, J = 6.7 Hz, 6H).

[0125] The synthesized product was subjected to high-resolution mass spectrometry analysis, and its characterization data are as follows:

[0126] MS (m / z) : [M+H] + = 1069.77214.

[0127] Example 6 Synthesis of a novel phosphatidylcholine compound (6)

[0128] The structural formula of the novel phosphatidylcholine compound (6) is as follows:

[0129] ,

[0130] Step S4:

[0131] ,

[0132] Under nitrogen, M3 (150 mg crude product, 1.0 eq.), anhydrous acetonitrile (0.75 mL, 5V), and N,N-dimethylbutylamine (56.78 mg, 3.0 eq.) were added sequentially to the reaction flask at room temperature. The temperature was slowly raised to 50°C and stirred for 30 h. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly purified by silica gel column chromatography (DCM / MeOH = 1 / 0 to 5 / 1). The desired product was obtained as a white waxy product (63 mg) in a 37.94% yield.

[0133] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0134] 1H NMR (400 MHz, Chloroform-d) δ 5.35 (tdt, J = 10.7, 8.4, 5.1 Hz,8H), 4.30 (d, J = 6.0 Hz, 2H), 4.07 (q, J = 6.6Hz, 6H), 3.95 (p, J = 4.6, 4.2Hz, 2H), 3.73 (t, J = 4.3 Hz, 2H), 2.96 – 2.40 (m, 19H), 2.05 (q, J = 6.8 Hz, 8H), 1.62 (t, J = 7.0 Hz, 4H), 1.40 – 1.18 (m, 38H),0.89 (t, J = 6.8 Hz, 6H).

[0135] The synthesized product was subjected to high-resolution mass spectrometry analysis, and its characterization data are as follows:

[0136] MS (m / z) : [M+H] + = 1055.76551.

[0137] Example 7 Synthesis of a novel phosphatidylcholine compound (7)

[0138] The structural formula of the novel phosphatidylcholine compound (7) is as follows:

[0139] ,

[0140] Step S4:

[0141] ,

[0142] Under nitrogen, M3 (150 mg crude product, 1.0 eq.), anhydrous acetonitrile (0.75 mL, 5V), and N,N-dimethyl-1-octylamine (78.00 mg, 3.0 eq.) were added sequentially to the reaction flask at room temperature. The temperature was slowly raised to 50°C and stirred for 30 h. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly purified by silica gel column chromatography (DCM / MeOH = 1 / 0 to 5 / 1). The desired product was obtained as a white waxy product (78 mg) in a 44.64% yield.

[0143] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0144] 1H NMR (400 MHz, Chloroform-d) δ 5.35 (tdd, J = 11.1, 6.4, 3.5 Hz, 8H), 4.30 (d, J = 6.6 Hz, 2H), 4.00 (dt, J = 43.6, 7.3 Hz, 8H), 3.73 (t, J =4.1 Hz, 2H), 2.97 – 2.43 (m, 20H), 2.05 (q, J = 6.8 Hz, 8H), 1.62 (t, J = 6.9Hz, 4H), 1.40 – 1.15 (m, 46H), 0.92 – 0.77(m, 12H).

[0145] The synthesized product was subjected to high-resolution mass spectrometry analysis, and its characterization data are as follows:

[0146] MS (m / z) : [M+H] + = 1111.80362.

[0147] Example 8 Synthesis of a novel phosphatidylcholine compound (8)

[0148] The structural formula of the novel phosphatidylcholine compound (8) is as follows:

[0149] ,

[0150] Step S4:

[0151] ,

[0152] Under nitrogen, M3 (150 mg crude product, 1.0 eq.), anhydrous acetonitrile (0.75 mL, 5V), and 1-methylpyrrolidine (40.92 mg, 3.0 eq.) were added sequentially to a reaction flask at room temperature. The temperature was slowly raised to 50°C and stirred for 30 h. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly purified by silica gel column chromatography (DCM / MeOH = 1 / 0 to 5 / 1). The desired product was obtained as a white waxy product (54.8 mg) in a 32.91% yield.

[0153] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0154] 1H NMR (400 MHz, Chloroform-d) δ 5.35 (tdd, J = 11.3, 6.5, 3.5 Hz, 8H), 4.40 – 4.23 (m, 2H), 4.05 (td, J = 6.8, 4.1 Hz, 6H), 3.97 – 3.68 (m, 4H),3.24 (s, 1H), 2.94 – 2.41 (m, 21H),2.35 – 2.16 (m, 1H), 2.05 (q, J = 6.8 Hz,8H), 1.62 (p, J = 6.8 Hz, 4H), 1.31 (ddt, J = 22.6, 19.5, 6.6 Hz, 38H), 0.89(t, J =6.7 Hz, 6H).

[0155] The synthesized product was subjected to high-resolution mass spectrometry analysis, and its characterization data are as follows:

[0156] MS (m / z) : [M+H] + = 1039.74359.

[0157] Example 9 Synthesis of a novel phosphatidylcholine compound (9)

[0158] The structural formula of the novel phosphatidylcholine compound (9) is as follows:

[0159] ,

[0160] Step S4:

[0161] ,

[0162] Under nitrogen, M3 (150 mg crude product, 1.0 eq.), anhydrous acetonitrile (0.75 mL, 5V), and N-methylpiperidine (46.76 mg, 3.0 eq.) were added sequentially to the reaction flask at room temperature. The temperature was slowly raised to 50°C and stirred for 30 h. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly purified by silica gel column chromatography (DCM / MeOH = 1 / 0 to 5 / 1). The desired product was obtained as a white waxy substance (42 mg) in a 25.36% yield.

[0163] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0164] 1H NMR (400 MHz, Chloroform-d) δ 5.35 (tdd, J = 11.1, 6.4, 3.5 Hz, 7H), 4.40 – 4.20 (m, 2H), 4.13 – 3.92 (m, 7H), 3.74 (t, J = 4.2 Hz, 2H), 3.02– 2.53 (m, 19H), 2.05 (q, J = 6.9 Hz, 8H), 1.62 (p, J = 6.8 Hz, 4H), 1.40 –1.21 (m, 38H), 0.89 (t, J = 6.7 Hz, 6H).

[0165] The synthesized product was subjected to high-resolution mass spectrometry analysis, and its characterization data are as follows:

[0166] MS (m / z) : [M+H] + = 1053.75037.

[0167] Example 10 Synthesis of a novel phosphatidylcholine compound (10)

[0168] The structural formula of the novel phosphatidylcholine compound (10) is as follows:

[0169] ,

[0170] Step S4:

[0171] ,

[0172] Under nitrogen, M3 (150 mg crude product, 1.0 eq.), anhydrous acetonitrile (0.75 mL, 5V), and N-ethylpiperidine (54.17 mg, 3.0 eq.) were added sequentially to the reaction flask at room temperature. The temperature was slowly raised to 50°C and stirred for 30 h. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly purified by silica gel column chromatography (DCM / MeOH = 1 / 0 to 5 / 1). The desired product was obtained as a white waxy substance (48 mg) in a 28.19% yield.

[0173] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0174] 1H NMR (400 MHz, Chloroform-d) δ 5.28 (tdd, J = 11.6, 10.3, 5.8, 3.2Hz, 8H), 4.31 – 4.15(m, 2H), 3.99 (t, J = 6.8 Hz, 6H), 3.93 – 3.84 (m, 2H), 3.66 (t, J = 4.2 Hz, 2H), 3.41 (s, 4H), 2.89 – 2.40 (m, 21H), 1.98 (q, J =6.9 Hz, 9H), 1.55 (t, J =7.0 Hz, 4H), 1.34 – 1.13 (m, 41H), 0.82 (t, J = 6.7Hz, 6H).

[0175] The synthesized product was subjected to high-resolution mass spectrometry analysis, and its characterization data are as follows:

[0176] MS (m / z) : [M+H] + = 1067.74502.

[0177] Example 11 Synthesis of a novel phosphatidylcholine compound (11)

[0178] The structural formula of the novel phosphatidylcholine compound (11) is as follows:

[0179] ,

[0180] Step S4:

[0181] ,

[0182] Under nitrogen, M3 (150 mg crude product, 1.0 eq.), anhydrous acetonitrile (0.75 mL, 5V), and N,N-dimethylcyclohexylamine (59.99 mg, 3.0 eq.) were added sequentially to the reaction flask at room temperature. The temperature was slowly raised to 50°C and stirred for 30 h. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly purified by silica gel column chromatography (DCM / MeOH = 1 / 0 to 5 / 1). The desired product was obtained as a white waxy substance (42 mg) in a 24.71% yield.

[0183] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0184] 1H NMR (400 MHz, Chloroform-d) δ 5.45 – 5.24 (m, 8H), 4.37 (s, 1H), 4.25 (d, J= 12.5 Hz, 1H), 4.15 – 3.92 (m, 8H), 3.75 (d, J = 4.3 Hz, 2H), 3.04– 2.54 (m, 19H), 2.05 (q, J = 6.9 Hz, 9H), 1.62 (t, J = 6.9 Hz, 4H), 1.39 –1.20 (m, 47H), 0.91 – 0.81 (m, 9H).

[0185] The synthesized product was subjected to high-resolution mass spectrometry analysis, and its characterization data are as follows:

[0186] MS (m / z) : [M+H] + = 1081.76979.

[0187] Example 12 Synthesis of a novel phosphatidylcholine compound (12)

[0188] The structural formula of the novel phosphatidylcholine compound (12) is as follows:

[0189] ,

[0190] Step S2:

[0191] ,

[0192] M1 (1.5 g, 1.0 eq.), octadecyl acrylate (5.17 g, 2.2 eq.), BHT (79.73 mg, 0.05 eq.), and acetic acid (21.73 mg, 0.05 eq.) were added sequentially to a reaction flask at room temperature. The mixture was heated to 70°C with stirring and allowed to react for 40 h. TLC (DCM / MeOH = 20 / 1) was performed to confirm that the starting materials were essentially consumed. The reaction solution was directly purified by silica gel column chromatography (DCM / MeOH = 1 / 0-100 / 1). 1.8 g of the desired product was obtained as a white wax in a 29.05% yield.

[0193] The synthesized intermediate compound M2 was subjected to nuclear magnetic resonance analysis, and its characterization data are as follows:

[0194] 1H NMR (400 MHz, Chloroform-d) δ 4.36 – 4.27 (m, 1H), 4.24 – 4.15 (m,1H), 4.06 (t, J = 6.8 Hz, 4H), 3.83 (t, J = 4.6Hz, 2H), 2.89 – 2.70 (m, 6H), 2.69 – 2.54 (m, 6H), 2.44 (t, J = 7.1 Hz, 4H), 1.61 (q, J = 5.5, 3.7 Hz, 7H), 1.26 (s, 66H), 0.93 – 0.83(m, 6H).

[0195] Step S3:

[0196] ,

[0197] M2 (500 mg, 1.0 eq.) and anhydrous DCM (5 mL, 10 V) were added to the reaction flask in sequence. The mixture was cooled to 4°C in an ice bath, and 2-chloro-2-oxo-1,3,2-dioxaphospholane (166.37 mg, 2.0 eq.) was added. After stirring for 5 minutes, triethylamine (149.10 mg, 2.5 eq.) was slowly added dropwise. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material 1 hour after the addition, indicating the reaction was complete. The reaction solution was concentrated under reduced pressure to remove the DCM, and EA (5 mL, 10 V) was added. The mother liquor was filtered and collected, and then concentrated under reduced pressure to yield 600 mg of a colorless oil. The crude product was unstable and was used directly in the next reaction without characterization.

[0198] Step S4:

[0199] ,

[0200] Under nitrogen, M3 (150 mg crude product, 1.0 eq.), anhydrous tetrahydrofuran (0.75 mL, 5 V), and trimethylamine (2M in THF) (0.24 mL, 3.0 eq.) were added sequentially to the reaction flask at room temperature. The temperature was slowly raised to 50°C and stirred for 30 h. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly purified by silica gel column chromatography (DCM / MeOH = 1 / 0 to 5 / 1). The desired product was obtained as a white solid (46 mg) in a 28.89% yield.

[0201] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0202] 1H NMR (400 MHz, Chloroform-d) δ 4.37 – 4.22 (m, 2H), 4.12 – 3.92 (m,7H), 3.78 – 3.70(m, 2H), 3.00 – 2.46 (m, 16H), 1.62 (p, J = 6.9 Hz, 4H), 1.26(s, 63H), 0.88 (t, J = 6.6 Hz, 6H).

[0203] Example 13 Synthesis of a novel phosphatidylcholine compound (13)

[0204] The structural formula of the novel phosphatidylcholine compound (13) is as follows:

[0205] ,

[0206] Step S4:

[0207] ,

[0208] Under nitrogen, M3 (150 mg crude product, 1.0 eq.), anhydrous tetrahydrofuran (0.75 mL, 5V), and triethylamine (47.32 mg, 3.0 eq.) were added sequentially to the reaction flask at room temperature. The temperature was slowly raised to 50°C and stirred for 30 h. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly purified by silica gel column chromatography (DCM / MeOH = 1 / 0 to 5 / 1). The desired product was obtained as a white waxy product (48 mg) in a 28.96% yield.

[0209] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0210] 1H NMR (400 MHz, Chloroform-d) δ 4.41 – 4.22 (m, 2H), 4.13 – 3.91 (m,7H), 3.74 (t, J = 4.2 Hz, 2H), 3.00 – 2.49 (m, 15H), 1.62 (t, J = 7.1 Hz, 4H), 1.26 (s, 66H), 0.88 (t, J = 6.7 Hz, 6H).

[0211] The synthesized product was subjected to high-resolution mass spectrometry analysis, and its characterization data are as follows:

[0212] MS (m / z) : [M+H] + = 1063.82195.

[0213] Example 14 Synthesis of a novel phosphatidylcholine compound (14)

[0214] The structural formula of the novel phosphatidylcholine compound (14) is as follows:

[0215] ,

[0216] Step S4:

[0217] ,

[0218] Under nitrogen, M3 (180 mg crude product, 1.0 eq.), anhydrous tetrahydrofuran (0.9 mL, 5V), and DIPEA (60.43 mg, 3.0 eq.) were added sequentially to the reaction flask at room temperature. The temperature was slowly raised to 50°C and stirred for 30 h. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly purified by silica gel column chromatography (DCM / MeOH = 1 / 0 to 5 / 1). The desired product (40 mg) was obtained as a white waxy product in a 23.51% yield.

[0219] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0220] 1H NMR (400 MHz, Chloroform-d) δ 4.31 (d, J = 34.3 Hz, 2H), 4.15 –3.91 (m, 7H), 3.75 (t, J = 4.3 Hz, 2H), 3.02 – 2.51 (m, 13H), 1.62 (t, J = 7.0Hz, 4H), 1.26 (s, 60H), 0.88 (t, J = 6.7 Hz, 6H).

[0221] Example 15 Synthesis of a novel phosphatidylcholine compound (15)

[0222] The structural formula of the novel phosphatidylcholine compound (15) is as follows:

[0223] ,

[0224] Step S4:

[0225] ,

[0226] Under nitrogen, M3 (180 mg crude product, 1.0 eq.), anhydrous tetrahydrofuran (0.9 mL, 5V), and N,N-diethylmethylamine (48.91 mg, 3.0 eq.) were added sequentially to the reaction flask at room temperature. The temperature was slowly raised to 50°C and stirred for 30 h. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly purified by silica gel column chromatography (DCM / MeOH = 1 / 0 to 5 / 1). The desired product was obtained as a white wax (39.5 mg) in a 20.12% yield.

[0227] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0228] 1H NMR (400 MHz, Chloroform-d) δ 4.37 – 4.24 (m, 2H), 4.06 (t, J =6.7 Hz, 6H), 3.95 (t, J = 6.7 Hz, 2H), 3.74 (t, J = 4.1 Hz, 2H), 2.98 – 2.41(m, 18H), 1.62 (t, J = 7.0 Hz, 4H), 1.26 (s, 71H), 0.88 (t, J = 6.8 Hz, 6H).

[0229] The synthesized product was subjected to high-resolution mass spectrometry analysis, and its characterization data are as follows:

[0230] MS (m / z) : [M+H] + = 1049.79375.

[0231] Example 16 Synthesis of a novel phosphatidylcholine compound (16)

[0232] The structural formula of the novel phosphatidylcholine compound (16) is as follows:

[0233] ,

[0234] Step S4:

[0235] ,

[0236] Under nitrogen, M3 (180 mg crude product, 1.0 eq.), anhydrous tetrahydrofuran (0.9 mL, 5V), and N-methyldipropylamine (64.65 mg, 3.0 eq.) were added sequentially to the reaction flask at room temperature. The temperature was slowly raised to 50°C and stirred for 30 h. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly purified by silica gel column chromatography (DCM / MeOH = 1 / 0 to 5 / 1). The desired product was obtained as a white wax (33.2 mg) in a 16.47% yield.

[0237] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0238] 1H NMR (400 MHz, Chloroform-d) δ 4.39– 4.22 (m, 2H), 4.06 (t, J = 6.8Hz, 6H), 3.95 (d, J = 7.3 Hz, 2H), 3.74 (t, J = 4.1 Hz, 2H), 2.98 – 2.47 (m,17H), 1.61 (q, J = 7.0 Hz, 4H), 1.26 (s, 70H), 0.88 (t, J = 6.7 Hz, 6H).

[0239] The synthesized product was subjected to high-resolution mass spectrometry analysis, and its characterization data are as follows:

[0240] MS (m / z) : [M+H] + = 1077.81804.

[0241] Example 17 Synthesis of a novel phosphatidylcholine compound (17)

[0242] The structural formula of the novel phosphatidylcholine compound (17) is as follows:

[0243] ,

[0244] Step S4:

[0245] ,

[0246] Under nitrogen, M3 (180 mg crude product, 1.0 eq.), anhydrous tetrahydrofuran (0.9 mL, 5V), and N,N-dimethylbutylamine (56.78 mg, 3.0 eq.) were added sequentially to the reaction flask at room temperature. The temperature was slowly raised to 50°C and stirred for 30 h. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly purified by silica gel column chromatography (DCM / MeOH = 1 / 0-5 / 1). The desired product was obtained as a white waxy product (29.1 mg) in a 14.63% yield.

[0247] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0248] 1H NMR (400 MHz, Chloroform-d) δ 4.36 – 4.23 (m, 2H), 4.06 (t, J =6.8 Hz, 6H), 3.95 (d, J = 7.9 Hz, 2H), 3.74 (t, J = 4.2 Hz, 2H), 2.95 – 2.41(m, 17H), 1.62 (p, J = 6.9 Hz, 4H), 1.26 (s, 68H), 0.88 (t, J = 6.7 Hz, 6H).

[0249] The synthesized product was subjected to high-resolution mass spectrometry analysis, and its characterization data are as follows:

[0250] MS (m / z) : [M+H] + = 1063.81467.

[0251] Example 18 Synthesis of a novel phosphatidylcholine compound (18)

[0252] The structural formula of the novel phosphatidylcholine compound (18) is as follows:

[0253] ,

[0254] Step S4:

[0255] ,

[0256] Under nitrogen, M3 (180 mg crude product, 1.0 eq.), anhydrous tetrahydrofuran (0.9 mL, 5V), and N,N-dimethyloctylamine (88.26 mg, 3.0 eq.) were added sequentially to the reaction flask at room temperature. The temperature was slowly raised to 50°C and stirred for 30 h. Samples were taken and checked by TLC (PE / EA = 1 / 1), confirming complete consumption of the starting material. The reaction solution was directly separated and purified by silica gel column chromatography (DCM / MeOH = 1 / 0-5 / 1). The desired product was obtained as a white wax (35.40 mg) in a 16.90% yield.

[0257] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0258] 1H NMR (400 MHz, Chloroform-d) δ 4.45 (dd, J = 11.0, 5.2 Hz, 1H), 4.17 – 3.93 (m, 8H), 3.76 – 3.61 (m, 2H), 3.24 – 2.47 (m, 17H), 2.40 – 1.91(m, 4H), 1.56 (p, J = 6.9 Hz, 4H), 1.31 – 1.06(m, 68H), 0.84 – 0.77 (m, 6H).

[0259] The synthesized product was subjected to high-resolution mass spectrometry analysis, and its characterization data are as follows:

[0260] MS (m / z) : [M+H] + = 1119.88601.

[0261] Example 19 Synthesis of a novel phosphatidylcholine compound (19)

[0262] The structural formula of the novel phosphatidylcholine compound (19) is as follows:

[0263] ,

[0264] Step S4:

[0265] ,

[0266] Under nitrogen, M3 (180 mg crude product, 1.0 eq.), anhydrous tetrahydrofuran (0.9 mL, 5V), and 1-methylpyrrolidine (47.78 mg, 3.0 eq.) were added sequentially to the reaction flask at room temperature. The temperature was slowly raised to 50°C and stirred for 30 h. Samples were collected and analyzed by TLC (PE / EA = 1 / 1), confirming complete consumption of the starting material. The reaction solution was directly separated and purified by silica gel column chromatography (DCM / MeOH = 1 / 0 to 5 / 1). The desired product was obtained as a white waxy product (45.90 mg) in a 23.43% yield.

[0267] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0268] 1H NMR (400 MHz, Chloroform-d) δ 4.41 – 3.89 (m, 9H), 3.78 – 3.61 (m,2H), 3.15 – 1.86(m, 19H), 1.54 (q, J = 7.1 Hz, 4H), 1.31 – 0.97 (m, 66H),0.81 (t, J = 6.7 Hz, 6H).

[0269] The synthesized product was subjected to high-resolution mass spectrometry analysis, and its characterization data are as follows:

[0270] MS (m / z) : [M+H] + = 1147.79604.

[0271] Example 20 Synthesis of a novel phosphatidylcholine compound (20)

[0272] The structural formula of the novel phosphatidylcholine compound (20) is as follows:

[0273] ,

[0274] Step S4:

[0275] ,

[0276] Under nitrogen, M3 (180 mg crude product, 1.0 eq.), anhydrous tetrahydrofuran (0.9 mL, 5V), and N-methylpiperidine (55.65 mg, 3.0 eq.) were added sequentially to the reaction flask at room temperature. The temperature was slowly raised to 50°C and stirred for 30 h. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly separated and purified by silica gel column chromatography (DCM / MeOH = 1 / 0-5 / 1). The desired product was obtained as a white wax (36.90 mg) in an 18.58% yield.

[0277] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0278] 1H NMR (400 MHz, Chloroform-d) δ 4.42 – 3.88 (m, 9H), 3.73 – 3.61 (m,2H), 3.10 – 2.49(m, 15H), 1.55 (t, J = 7.1 Hz, 4H), 1.19 (s, 66H), 0.81 (t, J= 6.8 Hz, 6H).

[0279] The synthesized product was subjected to high-resolution mass spectrometry analysis, and its characterization data are as follows:

[0280] MS (m / z) : [M+H] + = 1061.79010.

[0281] Example 21 Synthesis of a novel phosphatidylcholine compound (21)

[0282] The structural formula of the novel phosphatidylcholine compound (21) is as follows:

[0283] ,

[0284] Step S4:

[0285] ,

[0286] Under nitrogen, M3 (180 mg crude product, 1.0 eq.), anhydrous tetrahydrofuran (0.9 mL, 5V), and N-ethylpiperidine (63.52 mg, 3.0 eq.) were added sequentially to the reaction flask at room temperature. The temperature was slowly raised to 50°C and stirred for 30 h. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly separated and purified by silica gel column chromatography (DCM / MeOH = 1 / 0 to 5 / 1). The desired product was obtained as a white wax (37.60 mg) in an 18.69% yield.

[0287] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0288] 1H NMR (400 MHz, Chloroform-d) δ 4.46 – 4.35 (m, 1H), 4.15 – 3.91 (m,8H), 3.75 – 3.62(m, 2H), 3.20 – 2.62 (m, 16H), 1.94 (d, J = 6.4 Hz, 3H), 1.56(t, J = 7.1 Hz, 4H), 1.19 (s, 65H), 0.81 (t, J = 6.7 Hz, 6H).

[0289] Example 22 Synthesis of a novel phosphatidylcholine compound (22)

[0290] The structural formula of the novel phosphatidylcholine compound (22) is as follows:

[0291] ,

[0292] Step S4:

[0293] ,

[0294] Under nitrogen, M3 (180 mg crude product, 1.0 eq.), anhydrous tetrahydrofuran (0.9 mL, 5V), and N,N-dimethylcyclohexylamine (71.39 mg, 3.0 eq.) were added sequentially to the reaction flask at room temperature. The temperature was slowly raised to 50°C and stirred for 30 h. Samples were collected and checked by TLC (PE / EA = 1 / 1), confirming complete consumption of the starting material. The reaction solution was directly separated and purified by silica gel column chromatography (DCM / MeOH = 1 / 0 to 5 / 1). The desired product (40.80 mg) was obtained as a white waxy product in a 20.02% yield.

[0295] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0296] 1H NMR (400 MHz, Chloroform-d) δ 4.36 (d, J = 12.2 Hz, 1H), 4.20 – 3.87 (m, 8H), 3.75 – 3.61 (m, 2H), 3.42 (s, 1H), 3.09 – 2.35 (m, 17H), 1.55 (t, J = 7.1 Hz, 4H), 1.19 (s, 65H), 0.81 (t, J = 6.7 Hz, 6H).

[0297] The synthesized product was subjected to high-resolution mass spectrometry analysis, and its characterization data are as follows:

[0298] MS (m / z) : [M+H] + = 1089.83038.

[0299] Example 23 Synthesis of a novel phosphatidylcholine compound (23)

[0300] The structural formula of the novel phosphatidylcholine compound (23) is as follows:

[0301] ,

[0302] Step S3:

[0303] ,

[0304] M2 (150 mg, 1.0 eq.) and anhydrous DCM (1.5 mL, 10 V) were added to the reaction flask in sequence. The mixture was cooled to 4°C in an ice bath and 2-chloro-2-oxo-1,3,2-dioxaphospholane (51.35 mg, 2.0 eq.) was added. After the addition was complete, the mixture was stirred for 5 minutes. Trimethylamine (2.0 min THF) (0.25 mL, 2.5 eq.) was slowly added dropwise. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material 1 hour after the addition, indicating the reaction was complete. The reaction mixture was used directly in the next step without further purification. The crude product weight was estimated to be 160 mg.

[0305] Step S4:

[0306] ,

[0307] Under nitrogen, trimethylamine (2M in THF) (0.15 mL, 1.5 eq.) was added to the crude product solution M3 (160 mg, 1.0 eq.) in a reaction flask at room temperature with stirring. The temperature was slowly raised to 50°C and stirred for 30 h. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly purified by silica gel column chromatography (DCM / MeOH = 1 / 0-5 / 1). The desired product was obtained as a white solid (44.7 mg) in a 26.28% yield.

[0308] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0309] 1H NMR (400 MHz, Chloroform-d) δ 4.30 (td, J = 11.1, 10.4, 5.7 Hz, 2H), 4.05 (q, J = 5.4 Hz, 2H), 3.94 (s, 2H), 3.74(d, J = 5.4 Hz, 2H), 2.91 –2.78 (m, 9H), 2.74 – 2.41 (m, 10H), 1.66 – 1.50 (m, 4H), 1.26 (s, 60H), 0.91– 0.83 (m, 6H).

[0310] Example 24 Synthesis of a novel phosphatidylcholine compound (24)

[0311] The structural formula of the novel phosphatidylcholine compound (24) is as follows:

[0312] ,

[0313] Step S3:

[0314] ,

[0315] M2 (150 mg, 1.0 eq.) and anhydrous DCM (1.5 mL, 10 V) were added to the reaction flask in sequence. The mixture was cooled to 4°C in an ice bath and 2-chloro-2-oxo-1,3,2-dioxaphospholane (51.35 mg, 2.0 eq.) was added. After the addition was complete, the mixture was stirred for 5 minutes. Triethylamine (45.59 mg, 2.5 eq.) was slowly added dropwise. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material 1 hour after the addition, indicating the reaction was complete. The reaction mixture was used directly in the next step without further purification. The estimated crude product weight was 160 mg.

[0316] Step S4:

[0317] ,

[0318] Under nitrogen, triethylamine (25.88 mg, 1.5 eq.) was added to the crude product solution M3 (160 mg, 1.0 eq.) in a reaction flask at room temperature with stirring. The temperature was slowly raised to 50°C and stirred for 30 h. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly separated and purified by silica gel column chromatography (DCM / MeOH = 1 / 0-5 / 1). The desired product was obtained as a white solid (25 mg) in a 14.10% yield.

[0319] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0320] 1H NMR (400 MHz, Chloroform-d) δ 4.36 – 4.21 (m, 2H), 4.05 (q, J =5.6 Hz, 2H), 3.95 (s, 2H), 3.79 – 3.70 (m, 2H), 2.91 – 2.77 (m, 9H), 2.74 –2.46 (m, 9H), 1.56 (dq, J = 15.0, 7.6, 6.7 Hz, 4H), 1.26(s, 60H), 0.93 – 0.84(m, 6H).

[0321] Example 25 Synthesis of a novel phosphatidylcholine compound (25)

[0322] The structural formula of the novel phosphatidylcholine compound (25) is as follows:

[0323] ,

[0324] Step S3:

[0325] ,

[0326] M2 (150 mg, 1.0 eq.) and anhydrous DCM (1.5 mL, 10 V) were added to the reaction flask in sequence. The mixture was cooled to 4°C in an ice bath and 2-chloro-2-oxo-1,3,2-dioxaphospholane (51.35 mg, 2.0 eq.) was added. After the addition was complete, the mixture was stirred for 5 minutes. DIPEA (58.23 mg, 2.5 eq.) was slowly added dropwise. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material 1 hour after the addition, indicating the reaction was complete. The reaction mixture was used directly in the next step without further purification. The estimated crude product weight was 160 mg.

[0327] Step S4:

[0328] ,

[0329] Under nitrogen, the crude product solution M3 (160 mg, 1.0 eq.) was stirred in a reaction flask at room temperature. DIPEA (33.05 mg, 1.5 eq.) was added with stirring. The temperature was slowly raised to 50°C and stirred for 30 h. Samples were taken and checked by TLC (PE / EA = 1 / 1), confirming complete consumption of the starting material. The reaction solution was directly separated and purified by silica gel column chromatography (DCM / MeOH = 1 / 0-5 / 1). The desired product was obtained as a white solid (30 mg) in a 16.48% yield.

[0330] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0331] 1H NMR (400 MHz, Chloroform-d) δ 4.28 (d, J = 4.7 Hz, 2H), 4.14 – 3.89 (m, 4H), 3.72 (d, J = 7.1 Hz, 2H), 2.93 – 2.46 (m, 22H), 1.62 – 1.49 (m,4H), 1.26 (s, 52H), 0.88 (t, J= 6.7 Hz, 6H).

[0332] Example 26 Synthesis of a Novel Phosphatidylcholine Compound (26)

[0333] The structural formula of the novel phosphatidylcholine compound (26) is as follows:

[0334] ,

[0335] Step S4:

[0336] ,

[0337] Under nitrogen, M3 (160 mg crude product, 1.0 eq.), anhydrous tetrahydrofuran (0.8 mL, 5 V), and N,N-diethylmethylamine (44.59 mg, 3.0 eq.) were added sequentially to the reaction flask at room temperature. The temperature was slowly raised to 50°C and stirred for 30 h. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly purified by silica gel column chromatography (DCM / MeOH = 1 / 0 to 5 / 1). The desired product (64 mg) was obtained as a white waxy product in a 36.60% yield.

[0338] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0339] 1H NMR (400 MHz, Chloroform-d) δ 4.30(d, J = 5.9 Hz, 1H), 4.01 (dd, J= 46.5, 6.9 Hz, 4H), 3.79 – 3.65 (m, 3H), 2.96 – 2.47 (m, 20H), 1.65 – 1.50 (m, 4H), 1.26 (s, 62H), 0.88 (t, J = 6.7 Hz, 6H).

[0340] The synthesized product was subjected to high-resolution mass spectrometry analysis, and its characterization data are as follows:

[0341] MS (m / z) : [M+H]+ = 1025.68314.

[0342] Example 27 Synthesis of a Novel Phosphatidylcholine Compound (27)

[0343] The structural formula of the novel phosphatidylcholine compound (27) is as follows:

[0344] ,

[0345] Step S4:

[0346] ,

[0347] Under nitrogen, M3 (160 mg crude product, 1.0 eq.), anhydrous tetrahydrofuran (0.8 mL, 5 V), and N-methyldipropylamine (58.94 mg, 3.0 eq.) were added sequentially to the reaction flask at room temperature. The temperature was slowly raised to 50°C and stirred for 30 h. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly separated and purified by silica gel column chromatography (DCM / MeOH = 1 / 0 to 5 / 1). The desired product was obtained as a white wax (30.60 mg) in a 17.03% yield.

[0348] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0349] 1H NMR (400 MHz, Chloroform-d) δ 4.34 – 4.26 (m, 2H), 4.13 – 4.02 (m,2H), 3.96 (d, J = 7.6 Hz, 2H), 3.73 (dd, J = 6.0,3.0 Hz, 2H), 3.00 – 2.55 (m,19H), 2.01 (d, J = 6.4 Hz, 2H), 1.64 – 1.50 (m, 4H), 1.26 (s, 63H), 0.88 (t, J= 6.7 Hz, 6H).

[0350] Example 28 Synthesis of a Novel Phosphatidylcholine Compound (28)

[0351] The structural formula of the novel phosphatidylcholine compound (28) is as follows:

[0352] ,

[0353] Step S4:

[0354] ,

[0355] Under nitrogen, M3 (160 mg crude product, 1.0 eq.), anhydrous tetrahydrofuran (0.8 mL, 5 V), and N,N-dimethylbutylamine (51.76 mg, 3.0 eq.) were added sequentially to the reaction flask at room temperature. The temperature was slowly raised to 50°C and stirred for 30 h. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly purified by silica gel column chromatography (DCM / MeOH = 1 / 0 to 5 / 1). The desired product (31.80 mg) was obtained as a white wax in a 17.94% yield.

[0356] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0357] 1H NMR (400 MHz, Chloroform-d) δ 4.36 – 4.26 (m, 2H), 4.06 (q, J =5.7, 5.2 Hz, 2H), 3.99 – 3.90 (m, 2H), 3.73 (dd, J = 5.8, 2.9 Hz, 2H), 2.97 –2.55 (m, 20H), 2.01 (d, J = 6.2 Hz, 3H), 1.56 (p, J = 7.6, 6.9 Hz, 4H), 1.26 (s, 62H), 0.88 (t, J = 6.8 Hz, 6H).

[0358] Example 29 Synthesis of a novel phosphatidylcholine compound (29)

[0359] The structural formula of the novel phosphatidylcholine compound (29) is as follows:

[0360] ,

[0361] Step S3:

[0362] ,

[0363] M2 (150 mg, 1.0 eq.) and anhydrous tetrahydrofuran (1.5 mL, 10 V) were added to the reaction flask in sequence. The mixture was cooled to 4°C in an ice bath. 2-Chloro-2-oxo-1,3,2-dioxaphospholane (51.35 mg, 2.0 eq.) was added. After the addition was complete, the mixture was stirred for 5 minutes. N,N-dimethyloctylamine (70.86 mg, 2.5 eq.) was slowly added dropwise. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material 1 hour after the addition, indicating the reaction was complete. The reaction mixture was used directly in the next step without further purification. The estimated crude product weight was 160 mg.

[0364] Step S4:

[0365] ,

[0366] Under nitrogen, the crude product solution M3 (160 mg, 1.0 eq.) was stirred at room temperature in a reaction flask. N,N-dimethyloctylamine (40.23 mg, 1.5 eq.) was added. The temperature was slowly raised to 50°C and stirred for 30 hours. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly separated and purified by silica gel column chromatography (DCM / MeOH = 1 / 0-5 / 1). The desired product was obtained as a white solid (39.20 mg) in a 20.98% yield.

[0367] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0368] 1H NMR (400 MHz, Chloroform-d) δ 4.31 (t, J = 4.9 Hz, 2H), 4.13 –4.04 (m, 2H), 3.97 (d, J = 7.4 Hz, 2H), 3.78 – 3.69 (m, 2H), 2.88 (dp, J =14.9, 7.5, 6.9 Hz, 10H), 2.79 – 2.56 (m, 11H), 1.60 – 1.51 (m, 4H), 1.26 (s, 63H), 0.88 (t, J = 6.8 Hz, 6H).

[0369] Example 30 Synthesis of a novel phosphatidylcholine compound (30)

[0370] The structural formula of the novel phosphatidylcholine compound (30) is as follows:

[0371] ,

[0372] Step S3:

[0373] ,

[0374] M2 (150 mg, 1.0 eq.) and anhydrous tetrahydrofuran (1.5 mL, 10 V) were added to the reaction flask in sequence. The mixture was cooled to 4°C in an ice bath. 2-Chloro-2-oxo-1,3,2-dioxaphospholane (51.35 mg, 2.0 eq.) was added dropwise. After stirring for 5 minutes, 1-methylpyrrolidine (38.36 mg, 2.5 eq.) was slowly added dropwise. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material 1 hour after the addition, indicating the reaction was complete. The reaction solution was used directly in the next step without further purification. The estimated crude product weight was 160 mg.

[0375] Step S4:

[0376] ,

[0377] Under nitrogen, the crude product solution M3 (160 mg, 1.0 eq.) was stirred at room temperature in a reaction flask. 1-Methylpyrrolidine (21.78 mg, 1.5 eq.) was added with stirring. The temperature was slowly raised to 50°C and stirred for 30 h. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly separated and purified by silica gel column chromatography (DCM / MeOH = 1 / 0-5 / 1). The desired product was obtained as a white solid (42 mg) in a 24.07% yield.

[0378] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0379] 1H NMR (400 MHz, Chloroform-d) δ 4.31 (t, J = 5.1 Hz, 2H), 4.07 (q, J= 5.8, 5.2 Hz, 2H), 3.97 (d, J = 7.7 Hz, 2H), 3.83 – 3.64 (m, 3H), 2.88 (dp, J= 14.9, 7.4, 6.9 Hz, 9H), 2.79 – 2.57 (m, 9H), 1.65 – 1.49 (m, 4H), 1.26 (s, 60H), 0.88 (t, J = 6.7 Hz, 6H).

[0380] The synthesized product was subjected to high-resolution mass spectrometry analysis, and the characterization data are as follows:

[0381] MS (m / z) : [M+H] + = 1023.61454.

[0382] Example 31 Synthesis of a novel phosphatidylcholine compound (31)

[0383] The structural formula of the novel phosphatidylcholine compound (31) is as follows:

[0384] ,

[0385] Step S3:

[0386] ,

[0387] M2 (150 mg, 1.0 eq.) and anhydrous tetrahydrofuran (1.5 mL, 10 V) were added to the reaction flask in sequence. The mixture was cooled to 4°C in an ice bath and 2-chloro-2-oxo-1,3,2-dioxaphospholane (51.35 mg, 2.0 eq.) was added. After stirring for 5 minutes, N-methylpiperidine (44.68 mg, 2.5 eq.) was slowly added dropwise. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material 1 hour after the addition, indicating the reaction was complete. The reaction mixture was used directly in the next step without further purification. The estimated crude product weight was 160 mg.

[0388] Step S4:

[0389] ,

[0390] Under nitrogen, the crude product solution M3 (160 mg, 1.0 eq.) was stirred in a reaction flask at room temperature. N-methylpiperidine (25.36 mg, 1.5 eq.) was added with stirring. The temperature was slowly raised to 50°C and stirred for 30 h. Samples were taken and checked by TLC (PE / EA = 1 / 1), confirming complete consumption of the starting material. The reaction solution was directly separated and purified by silica gel column chromatography (DCM / MeOH = 1 / 0-5 / 1). The desired product was obtained as a white solid (50.30 mg) in a 28.43% yield.

[0391] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0392] 1H NMR (400 MHz, Chloroform-d) δ 4.35 – 4.26 (m, 2H), 4.06 (q, J =5.6 Hz, 2H), 3.95 (q, J = 6.9, 5.0 Hz, 2H), 3.73 (t, J = 4.3 Hz, 2H), 2.87 (q,J = 7.4, 7.0 Hz, 9H), 2.77 – 2.57 (m, 10H), 1.63 – 1.49 (m, 4H), 1.26 (s,61H), 0.88 (t, J = 6.8 Hz, 6H).

[0393] The synthesized product was subjected to high-resolution mass spectrometry analysis, and the characterization data are as follows:

[0394] MS (m / z) : [M+H] + = 1037.68125.

[0395] Example 32 Synthesis of a novel phosphatidylcholine compound (32)

[0396] The structural formula of the novel phosphatidylcholine compound (32) is as follows:

[0397] ,

[0398] Step S3:

[0399] ,

[0400] M2 (150 mg, 1.0 eq.) and anhydrous tetrahydrofuran (1.5 mL, 10 V) were added to the reaction flask in sequence. The mixture was cooled to 4°C in an ice bath. 2-Chloro-2-oxo-1,3,2-dioxaphospholane (51.35 mg, 2.0 eq.) was added dropwise. After the addition was complete, the mixture was stirred for 5 minutes. N-ethylpiperidine (51.00 mg, 2.5 eq.) was slowly added dropwise. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material 1 hour after the addition, indicating the reaction was complete. The reaction solution was used directly in the next step without further purification. The crude product weighed an estimated 160 mg.

[0401] Step S4:

[0402] ,

[0403] Under nitrogen, the crude product solution M3 (160 mg, 1.0 eq.) was stirred in a reaction flask at room temperature. N-ethylpiperidine (28.95 mg, 1.5 eq.) was added with stirring. The temperature was slowly raised to 50°C and stirred for 30 h. Samples were taken and checked by TLC (PE / EA = 1 / 1), confirming complete consumption of the starting material. The reaction solution was directly separated and purified by silica gel column chromatography (DCM / MeOH = 1 / 0-5 / 1). The desired product was obtained as a white solid (64 mg) in a 35.69% yield.

[0404] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0405] 1H NMR (400 MHz, Chloroform-d) δ 4.37 – 4.19 (m, 2H), 4.13 – 4.02 (m,2H), 3.96 (d, J = 7.3 Hz, 2H), 3.74 (d, J = 4.9Hz, 2H), 2.93 – 2.81 (m, 8H), 2.76 – 2.57 (m, 9H), 1.65 – 1.49 (m, 4H), 1.26 (s, 61H), 0.88 (t, J = 6.8 Hz, 6H).

[0406] Example 33 Synthesis of a novel phosphatidylcholine compound (33)

[0407] The structural formula of the novel phosphatidylcholine compound (33) is as follows:

[0408] ,

[0409] Step S3:

[0410] ,

[0411] M2 (150 mg, 1.0 eq.) and anhydrous tetrahydrofuran (1.5 mL, 10 V) were added to the reaction flask in sequence. The mixture was cooled to 4°C in an ice bath. 2-Chloro-2-oxo-1,3,2-dioxaphospholane (51.35 mg, 2.0 eq.) was added dropwise. After stirring for 5 minutes, N,N-dimethylcyclohexylamine (57.32 mg, 2.5 eq.) was slowly added dropwise. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material 1 hour after the addition, indicating the reaction was complete. The reaction solution was used directly in the next step without further purification. The estimated crude product weight was 160 mg.

[0412] Step S4:

[0413] ,

[0414] Under nitrogen, the crude product solution M3 (160 mg, 1.0 eq.) was stirred at room temperature in a reaction flask. N,N-dimethylcyclohexylamine (32.54 mg, 1.5 eq.) was added. The temperature was slowly raised to 50°C and stirred for 30 hours. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material. The reaction solution was directly separated and purified by silica gel column chromatography (DCM / MeOH = 1 / 0-5 / 1). The desired product was obtained as a white solid (71 mg) in a 39.08% yield.

[0415] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0416] 1H NMR (400 MHz, Chloroform-d) δ 4.30 (d, J = 6.3 Hz, 2H), 4.06 (dd,J = 7.0, 4.6 Hz, 2H), 3.94 (q, J = 7.0, 5.1 Hz,2H), 3.73 (t, J = 4.1 Hz, 2H), 2.86 (t, J = 7.4 Hz, 9H), 2.74 – 2.55 (m, 10H), 1.63 – 1.49 (m, 4H), 1.26 (s, 65H), 0.90 – 0.85 (m, 6H).

[0417] The synthesized product was subjected to high-resolution mass spectrometry analysis, and the characterization data are as follows:

[0418] MS (m / z) : [M+H] + = 1065.72139.

[0419] Example 34 Synthesis of a novel phosphatidylcholine compound (34)

[0420] The structural formula of the novel phosphatidylcholine compound (34) is as follows:

[0421] ,

[0422] Step S2:

[0423] ,

[0424] M1 (1.5 g, 1.0 eq.), SM3 (containing 0.05% BHT stabilizer) (3.83 g, 2.2 eq.), and acetic acid (21.73 mg, 0.05 eq.) were added sequentially to a reaction flask at room temperature. The mixture was heated to 70°C with stirring and reacted for 40 h. TLC (DCM / MeOH = 20 / 1) was performed to confirm that the starting materials were essentially consumed. The reaction solution was directly purified by silica gel column chromatography (DCM / MeOH = 1 / 0-100 / 1). 2.4 g of the desired product was obtained as a light yellow oil in a 48.20% yield.

[0425] The synthesized intermediate compound M2 was subjected to nuclear magnetic resonance analysis, and its characterization data are as follows:

[0426] 1H NMR (400 MHz, Chloroform-d) δ 4.36 – 4.27 (m, 1H), 4.24 – 4.08 (m,2H), 3.97 (d, J = 5.8 Hz, 4H), 3.83 (t, J = 4.6Hz, 2H), 2.87 – 2.69 (m, 6H), 2.69 – 2.56 (m, 6H), 2.45 (t, J = 7.2 Hz, 4H), 1.78 – 1.54 (m, 3H), 1.27 (tt,J = 7.3, 4.4 Hz, 39H), 0.89 (td, J = 6.6, 4.2 Hz, 12H).

[0427] Step S3:

[0428] ,

[0429] M2 (150 mg, 1.0 eq.) and anhydrous tetrahydrofuran (1.5 mL, 10 V) were added to the reaction flask in sequence. The mixture was cooled to 4°C in an ice bath. 2-Chloro-2-oxo-1,3,2-dioxaphospholane (62.12 mg, 2.0 eq.) was added. After the addition was complete, the mixture was stirred for 5 minutes. N,N-diethylmethylamine (47.51 mg, 2.5 eq.) was slowly added dropwise. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material 1 hour after the addition, indicating the reaction was complete. The reaction mixture was used directly in the next step without further purification. The estimated crude product weight was 160 mg.

[0430] Step S4:

[0431] ,

[0432] Under nitrogen, the crude product solution M3 (160 mg, 1.0 eq.) was stirred at room temperature in a reaction flask. N,N-diethylmethylamine (26.34 mg, 1.5 eq.) was added with stirring. The temperature was slowly raised to 50°C and stirred for 30 h. Samples were taken and analyzed by TLC (PE / EA = 1 / 1), confirming complete consumption of the starting material. The reaction solution was directly separated and purified by silica gel column chromatography (DCM / MeOH = 1 / 0-5 / 1). The desired product was obtained as a colorless oil (65.70 mg) in a 25.74% yield.

[0433] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0434] 1H NMR (400 MHz, Chloroform-d) δ 4.32 (ddd, J = 38.0, 12.4, 7.0 Hz, 2H), 4.14 – 4.05(m, 4H), 3.98 (d, J = 5.8 Hz, 4H), 3.70 (t, J = 5.8 Hz, 2H), 3.02 – 2.44 (m, 16H), 1.61 (q, J = 5.7 Hz, 2H), 1.28 (hept, J = 7.1 Hz, 43H), 0.89 (td, J = 6.6, 4.2 Hz, 12H).

[0435] Example 35 Synthesis of a novel phosphatidylcholine compound (35)

[0436] The structural formula of the novel phosphatidylcholine compound (35) is as follows:

[0437] ,

[0438] Step S3:

[0439] ,

[0440] M2 (150 mg, 1.0 eq.) and anhydrous tetrahydrofuran (1.5 mL, 10 V) were added to the reaction flask in sequence. The mixture was cooled to 4°C in an ice bath. 2-Chloro-2-oxo-1,3,2-dioxaphospholane (62.12 mg, 2.0 eq.) was added dropwise. After stirring for 5 minutes, N,N-dimethylbutylamine (55.15 mg, 2.5 eq.) was slowly added dropwise. TLC (PE / EA = 1 / 1) confirmed complete consumption of the starting material 1 hour after the addition, indicating the reaction was complete. The reaction mixture was used directly in the next step without further purification. The estimated crude product weight was 160 mg.

[0441] Step S4:

[0442] ,

[0443] Under nitrogen, the crude product solution M3 (160 mg, 1.0 eq.) was stirred at room temperature in a reaction flask. N,N-dimethylbutylamine (30.08 mg, 1.5 eq.) was added. The temperature was slowly raised to 50°C and stirred for 30 h. Samples were taken and analyzed by TLC (PE / EA = 1 / 1), confirming complete consumption of the starting material. The reaction solution was directly separated and purified by silica gel column chromatography (DCM / MeOH = 1 / 0 to 5 / 1). The desired product (100 mg) was obtained as a colorless oil in a 55.44% yield.

[0444] The synthesized product was subjected to NMR analysis, and its characterization data are as follows:

[0445] 1H NMR (400 MHz, Chloroform-d) δ 4.40 – 4.24 (m, 2H), 4.14 – 4.04 (m,4H), 3.98 (d, J = 5.8 Hz, 4H), 3.70 (t, J = 5.7Hz, 2H), 3.00 – 2.46 (m, 15H), 1.62 (t, J = 6.0 Hz, 2H), 1.37 – 1.12 (m, 37H), 0.89 (td, J = 7.0, 6.6, 4.2Hz, 12H).

[0446] Preparation and screening of LNPs

[0447] mRNA: All mRNA used in these studies was synthesized and purified by Xingrui Pharmaceuticals (Suzhou) Co., Ltd. through in vitro transcription. Fluci mRNA encodes luciferase, which can express luciferase protein after transfection into cells or administration to animals. When luciferin is added, it can emit light. The intensity of the luminescence is positively correlated with the expression of luciferase and is used to evaluate the delivery capacity of the delivery vector.

[0448] Formula 1 LNP: Formula 1 LNP contains four lipids, namely SM-102, the lipid of the present invention, cholesterol, and DMG-PEG2000; the molar percentages of the four lipids are 46% SM-102, 8% lipid of the present invention, 43.95% cholesterol, and 2.05% DMG-PEG2000; the mass ratio of total lipids to Fluci mRNA is 18:1;

[0449] SM-102 LNP: SM-102 LNP contains four lipids, with the components and molar percentages being 50% SM-102, 10% DSPC, 38.5% cholesterol, and 1.5% DMG-PEG2000; the mass ratio of total lipids to Fluci mRNA is 19:1;

[0450] LNP preparation: The four lipids were dissolved in ethanol according to the corresponding formula ratio, and Fluci mRNA was dissolved in 10mM sodium citrate solution (pH=4.0). The ethanol phase (four lipids) and the aqueous phase (mRNA) were mixed in a volume ratio of 1:3 to prepare a crude LNP product. The crude LNP product was diluted three times with PBS and purified using a 100kD ultrafiltration tube to replace the ethanol with PBS. The solution was then filtered through a 0.22μm sterile filter to obtain LNP encapsulated with Fluci mRNA.

[0451] LNP physicochemical parameter testing: particle size, polydispersity coefficient, and zeta potential were determined using a Zetasizer Pro (Malvern Panalytical); LNP encapsulation efficiency was determined using the Quant-it RiboGreen assay (Thermo Fisher Scientific);

[0452] The lipid structure of SM-102 is as follows:

[0453]

[0454] Example 37 In vivo delivery and expression of LNPs using novel lipid formulation 1

[0455] Formula 1 LNP is prepared using Fluci mRNA as the nucleic acid component; Fluci is luciferase mRNA, a common reporter gene expressed in cells, using luciferin as a substrate to catalyze bioluminescence. The abundance of luciferase expression is assessed by collecting fluorescence intensity. The components and physicochemical properties of the LNP preparation are shown in Table 1:

[0456] Table 1 Representative physicochemical properties of LNP preparations

[0457] Serial number Lipid name LNP formulations Particle size (nm) polydispersity coefficient Zeta potential (mV) 1 Compound (1) Recipe 1 124.7 0.117 -10.24 2 Compound (2) Recipe 1 130.4 0.092 -12.05 3 Compound (3) Recipe 1 133.5 0.070 -10.59 4 Compound (4) Recipe 1 130.9 0.117 -13.03 5 Compound (5) Recipe 1 126.7 0.142 -11.15 6 Compound (6) Recipe 1 127.3 0.006 -12.34 7 Compound (7) Recipe 1 128.6 0.049 -12.00 8 Compound (8) Recipe 1 134.1 0.065 -12.36 9 Compound (9) Recipe 1 144.9 0.114 -10.52 10 Compound (10) Recipe 1 132.7 0.062 -13.57 11 Compound (11) Recipe 1 129.0 0.021 -11.03 12 Compound (12) Recipe 1 129.9 0.089 -13.76 13 Compound (13) Recipe 1 133.2 0.035 -11.41 14 Compound (14) Recipe 1 127.2 0.078 -14.68 15 Compound (15) Recipe 1 130.7 0.158 -10.48 16 Compound (16) Recipe 1 130.7 0.089 -14.00 17 Compound (17) Recipe 1 139.8 0.155 -12.79 18 Compound (18) Recipe 1 131.1 0.015 -13.84 19 Compound (19) Recipe 1 127.1 0.090 -14.00 20 Compound (20) Recipe 1 130.4 0.014 -12.62 21 Compound (21) Recipe 1 130.9 0.156 -10.65 22 Compound (22) Recipe 1 125.0 0.121 -12.61 23 Compound (23) Recipe 1 135.8 0.144 -8.09 24 Compound (24) Recipe 1 137.1 0.127 -14.39 25 Compound (25) Recipe 1 129.4 0.129 -7.50 26 Compound (26) Recipe 1 134.2 0.086 -11.88 27 Compound (27) Recipe 1 136.4 0.123 -11.49 28 Compound (28) Recipe 1 128.1 0.072 -10.02 29 Compound (29) Recipe 1 146.3 0.044 -8.96 30 Compound (30) Recipe 1 131.2 0.120 -12.12 31 Compound (31) Recipe 1 135.9 0.077 -10.49 32 Compound (32) Recipe 1 129.0 0.084 -12.64 33 Compound (33) Recipe 1 128.7 0.156 -10.49 .

[0458] Example 38 In vitro delivery and expression activity of novel lipid formulation 1 LNP

[0459] According to Example 37, LNPs were prepared using Fluci mRNA as the nucleic acid component and transfected into HEK293T and Huh7 cell lines cultured in vitro. Post-transfection cell viability and Fluci expression levels were determined using the commercial reagent ONE GLO Kit. Fluci expression levels were normalized using cell viability. Both cell lines were plated in 96-well plates at approximately 2E4 cells / well and transfected with 25 ng / well of LNPs.

[0460] Figure 2 Figure 3 The results showed that the LNP formulation of compound (1) to compound (22) had higher in vitro delivery expression activity in HEK293T and Huh7 cells.

[0461] Example 39 Comparison of Targeted Delivery Expression Activity of Compound (1) and Compound (4) Formula 1 LNP and SM-102 LNP

[0462] According to Example 37, the lipid formulations 1 LNPs of compound (1) and compound (4) and SM-102 LNPs were prepared using Fluci mRNA as the nucleic acid component; the abundance of luciferase expression was assessed by collecting fluorescence intensity;

[0463] C57 mice (SPF grade, female, 6-8 weeks old) were administered once via the tail vein on day 0, with a dose of 0.25 mpk in a volume of 200 μL. 6 hours after administration, 30 mg of luciferase substrate was injected intraperitoneally into the experimental mice. Organs were dissected within 10-15 minutes and imaged using the PerkinElmer IVIS small animal imaging system to assess the expression level of Fluci by fluorescence intensity.

[0464] Figure 4 The results showed that the Fluci of the lipid formulation 1 LNP of compound (1) and compound (4) had good spleen targeting specificity, and the Fluci of the SM-102 LNP formulation had good liver targeting specificity and high expression efficiency. Among them, the Fluci of the lipid formulation 1 LNP of compound (1) and compound (4) had the highest expression level in the spleen, which could reach more than 90%, while the SM-102 LNP had the highest expression level in the liver, which could reach more than 80%. Figure 5 shown.

[0465] Example 40 Delivery and Expression Activity of Novel Lipid Formulation 1 LNPs of Compound (2), Compound (13), Compound (24), Compound (5), Compound (16) and Compound (27)

[0466] Formula 1 of the novel lipid LNPs of compound (2), compound (13), compound (24), compound (5), compound (16) and compound (27) was prepared using Fluci mRNA as the nucleic acid component; the abundance of luciferase expression was evaluated by collecting fluorescence intensity;

[0467] The components and physicochemical properties of the LNP preparation are shown in Table 2:

[0468] Table 2 Physicochemical properties of LNP preparations

[0469] Serial number Lipid name LNP formulations Particle size (nm) polydispersity coefficient Zeta potential (mV) Encapsulation rate% 1 Compound (2) Recipe 1 105.0 0.071 -7.39 88.8% 2 Compound (13) Recipe 1 106.4 0.103 -6.94 89.5% 3 Compound (24) Recipe 1 104.8 0.081 -8.29 89.1% 4 Compound (5) Recipe 1 108.4 0.092 -7.64 88.3% 5 Compound (16) Recipe 1 106.3 0.103 -8.95 87.2% 6 Compound (27) Recipe 1 105.8 0.113 -7.61 87.3%

[0470] C57 mice (SPF grade, female, 6-8 weeks old) were administered once via the tail vein on day 0, with a dose of 0.25 mpk in a volume of 200 μL. 6 hours after administration, 30 mg of luciferase substrate was injected intraperitoneally into the experimental mice. Organs were dissected within 10-15 minutes and imaged using the PerkinElmer IVIS small animal imaging system to assess the expression level of Fluci by fluorescence intensity.

[0471] Figure 6 The results showed that the Fluci of the LNP formulation of the novel lipid formula 1 of compound (2), compound (13), compound (24), compound (5), compound (16) and compound (27) were significantly expressed in the liver and spleen, and the expression efficiency in the spleen was higher; among them, the Fluci of the LNP formulation of the lipid formula 1 of compound (2) was the highest in the spleen, reaching more than 80%, as shown in FIG. Figure 7 shown.

[0472] The basic principles, main features, and advantages of the present invention are shown and described above. Those skilled in the art should understand that the present invention is not limited to the above embodiments. The above embodiments and descriptions are merely illustrative of the principles of the present invention. Various changes and modifications may be made to the present invention without departing from the spirit and scope of the present invention. Such changes and modifications are intended to fall within the scope of the present invention. The scope of protection claimed in the present invention is defined by the appended claims and their equivalents.

Claims

1. A phosphatidylcholine compound of the following structure, a pharmaceutically acceptable salt, a tautomer or a stereoisomer of the compound, characterized in that The phosphatidylcholine compound has the structure shown below: 、 、 。 2. Use of the phosphatidylcholine compound, or a pharmaceutically acceptable salt, tautomer or stereoisomer of the compound according to claim 1, in the preparation of a drug carrier.

3. A composition characterized in that The composition comprises the phosphatidylcholine compound according to claim 1, or a pharmaceutically acceptable salt, tautomer or stereoisomer of the compound.

4. The composition according to claim 3, characterized in that The composition comprises the phosphatidylcholine compound according to claim 1, a pharmaceutically acceptable salt, tautomer or stereoisomer of the compound, and other lipids.

5. The composition according to claim 4, characterized in that The other lipids are selected from , cholesterol, DMG-PEG2000 or one or more.

6. The composition according to claim 4 or 5, characterized in that The other lipids are selected from .

7. The composition according to claim 4 or 5, characterized in that The other lipids are selected from cholesterol.

8. The composition according to claim 4 or 5, characterized in that The other lipids are selected from DMG-PEG2000.

9. The composition according to claim 4 or 5, characterized in that The other lipids are selected from , cholesterol and DMG-PEG2000.

10. The composition according to claim 9, characterized in that The molar percentages of the other lipids, the phosphatidylcholine compounds according to claim 1, cholesterol, and DMG-PEG2000 are (40%~60%): (5%~15%): (22%~54.5%): (0.5%~1.5%).

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