Chemical synthesis method for serotype o5 o-antigen oligosaccharide of pseudomonas aeruginosa

Abstract

The present invention relates to the field of chemical synthesis, and disclosed is a chemical synthesis method for a serotype O5 O-antigen oligosaccharide of pseudomonas aeruginosa. In the present invention, a D-glucuronic acid building block and a D-fucosamine building block are used to construct an O-antigen trisaccharide, wherein the stereoselective synthesis of a D-fucosamine 1,2-α-cis-glycosidic bond relies on remote acyl participation and reagent regulation, and the synthesis of two 2,3-diaminomannuronic acid 1,2-β-trans glycosidic bonds is realized by means of SN2 nucleophilic substitution of a C2 azido group; and by means of selective assembly of protecting groups, orthogonal modification of modifying groups, and regulation of the reactivity of glycosyl donors and acceptors, the preparation of a multifunctionally modified O-antigen target trisaccharide is successfully completed. The method of the present invention is characterized in that raw materials are cheap and easily available and the preparation method is simple and easy to repeat, and has good prospects of application in the development of vaccines against pseudomonas aeruginosa.

Description

A chemical synthesis method for oligosaccharides of O5 serotype O antigen from Pseudomonas aeruginosa. Technical Field

[0001] This invention relates to a chemical synthesis method for oligosaccharides of O5 serotype O-antigen from Pseudomonas aeruginosa, belonging to the field of chemical synthesis. Background Technology

[0002] Pseudomonas aeruginosa, also known as Pseudomonas aeruginosa, is a highly adaptable opportunistic pathogen and one of the main causes of hospital-acquired infections. Pseudomonas aeruginosa infection can cause a series of complications such as sepsis and septicemia, posing a potentially fatal risk to immunocompromised individuals, including those with cystic fibrosis, HIV / AIDS, and those undergoing chemotherapy (K. Grimwood et al., Hum Vaccin Immunother 2015, 11, 14-20). According to the US Centers for Disease Control and Prevention (CDC), approximately 51,000 patients are infected with Pseudomonas aeruginosa annually, with a mortality rate of about 5.3% (ME Chirgwin et al., Materials 2019, 12, 4093). Pseudomonas aeruginosa exhibits strong resistance to multiple antibiotics (CKElogne et al., Afr J Microbiol Res, 2018, 12, 62-67). In 2017, the World Health Organization (WHO) listed Pseudomonas aeruginosa as a "highest priority" pathogen in its first list of antibiotic-resistant pathogens, highlighting its resistance as one of the greatest threats to human health today (E. Tacconelli et al., World Health Organization, 2017). Since immunoprophylaxis strategies are not affected by antibiotic resistance mechanisms, the development of highly effective and safe vaccines for treating drug-resistant pathogens is urgently needed. Researchers have conducted extensive studies on Pseudomonas aeruginosa vaccines, primarily including whole-cell vaccines and various subunit vaccines, but no vaccines currently approved for human use have been approved for marketing (GPPriebe et al., Expert Rev Vaccines, 2014, 13, 507-519). Compared to whole-cell vaccines containing many ineffective antigens, subunit vaccines containing Pseudomonas aeruginosa cell surface polysaccharides and membrane proteins offer more effective immune protection. Studies have shown that using Pseudomonas aeruginosa lipopolysaccharide as an immunogen in vaccine research has demonstrated good protective effects. The most effective antigenic target for immunity is the O-antigen polysaccharide portion of bacterial surface lipopolysaccharide (LPS) (SJCryz et al., Infect Immun, 1984, 44, 508-513). Therefore, lipopolysaccharide O-antigen has been widely used in the development of Pseudomonas aeruginosa vaccines (NFAbu-Baker et al., Advances in Microbiology, 2016, 6, 332-342).

[0003] Based on the different O-antigens of Pseudomonas aeruginosa lipopolysaccharides, 20 Pseudomonas aeruginosa serotypes with different immunological specificities have been identified (PVLiu et al., J Clin Microbiol, 1990, 28, 922-925). Among the 20 Pseudomonas aeruginosa serotype O-antigens with well-defined structures, the O5 serotype O-antigen shows high similarity to the O2, O16, O18, and O20 serotype O-antigens in their trisaccharide repeat fragment structures. The O5 serotype O-antigen polysaccharide structure has attracted attention due to its high structural specificity. This polysaccharide structure consists of a complex modified polyaminotrisaccharide repeat unit: [→4)-β-D-ManpNAc3AmA-(1→4)-β-D-ManpNAc3AcA-(1→3)-α-D-FucpNAc-(1→](I.Sadovskaya et al. (al., Eur. J. Biochem. 1997, 255, 673-684). It is noteworthy that polysaccharides with two consecutive β-D-ManpN3NA segments are uncommon in nature and had not been synthesized before. Furthermore, most of the hydroxyl groups on the sugar ring of the trisaccharide repeating segments are replaced by amino groups, with only one hydroxyl group exposed, resulting in a high-density functionalized aminoglycoside. Another distinctive feature is that the trisaccharide repeating segments are all linked by 1,2-cis-glycosidic bonds. A unique functional group, acetamidine (Am), is predominantly present in the bacterial O-antigen, specifically attached to the O-antigen trisaccharide 2,3-diamino-D-mannoside. The amino group at the C3 position of the aldehyde acid. Despite significant progress in recent years in stereoselective glycosylation synthesis and strategies for assembling complex oligosaccharides, the chemical synthesis of high-density functionalized aminoglycosides remains challenging. These challenges include: difficulty in synthesizing stereospecific trisaccharides due to all linkages being 1,2-cis-glycosidic bonds; the poor reactivity of the rare sugar ManpNAc3NAcA during glycosylation; and the need for orthogonal assembly of acetamidine and acetyl groups at the amino group. Therefore, the total synthesis of this structure requires comprehensive route design, including the selection of protecting groups, the timing of introducing modifying groups, and key aspects such as the efficiency and selectivity of the glycosylation reaction.

[0004] Summary of the Invention

[0005] This invention relates to the synthesis of a trisaccharide fragment of Pseudomonas aeruginosa O5 serotype O-antigen using three glycoblock chemical methods. The fragment mainly comprises 3-amino-D-glucose blocks, D-glucuronic acid blocks, D-fucoglycosamine blocks, and orthogonal modifications of 1,2-α-cis-glycosidic bonds, 1,2-β-cis-glycosidic bonds, and amino functional groups. The construction of the 1,2-cis-glycosidic bond and the orthogonal modification of the amino functional groups are key steps in the formation of this target trisaccharide. This invention successfully solves the stereoselectivity problem in constructing the 1,2-α-cis-glycosidic bond through the synergistic effect of acyl long-range participation and additive effects. The stereoconstruction of two 1,2-β-cis-glycosidic bonds is achieved through SN2 nucleophilic substitution of the C2-position azido group. Using the synthesized trisaccharide precursor, the target trisaccharide, as shown in Formula VI, is obtained through C6-position hydroxyl oxidation and orthogonal modification of different amino functional groups, followed by deprotection. This invention uses amino linkers to immobilize synthetic oligosaccharides on the chip surface and uses Pseudomonas aeruginosa patient serum to screen oligosaccharide antigens for antigenicity studies of synthetic oligosaccharides.

[0006] The first objective of this invention is to provide a method for synthesizing the O-antigen trisaccharide of O5 serotype Pseudomonas aeruginosa. The method includes: constructing the O-antigen trisaccharide using 3-amino-D-glucose building blocks, D-glucuronic acid building blocks, and D-fucosamine building blocks; constructing a 1,2-α-cis-D-fucosamine glycosidic bond using the synergistic effect of acyl long-range participation and additive effects; constructing a 1,2-β-trans-glycosidic bond of two 2,3-di-amino-D-mannuronic acids using an SN2 substitution reaction of the C2-position azide group; and completing the orthogonal assembly of the five amino functional modifying groups in the O-antigen trisaccharide.

[0007] This invention relates to an oligosaccharide fragment of Pseudomonas aeruginosa O5 serotype O-antigen trisaccharide assembled with a linker arm, the chemical structure of which can be represented by general formula (VI):

[0008] Linker is -L-NH2, where L represents the connecting arm;

[0009] In this invention, the connecting arm L can be a chain structure containing 2-40 carbon atoms (including the carbon atoms of the side chain) with 0-6 heteroatoms.

[0010] In this invention, when the main chain length of the connecting arm is 4-8 atoms, the chain may contain 1, 2 or 3 heteroatoms (O, N and S). When the main chain length of the connecting arm is 9-14 atoms, the chain may contain 1, 2, 3, 4, 5 or 6 heteroatoms (O, N and S).

[0011] In this invention, the linker arm -L- can be a cyclic structure that is wholly or partially fluorinated. The linker arm -L- can contain a three-, four-, five-, or six-membered saturated carbon ring; it can also contain a five-membered unsaturated carbon ring (non-aromatic ring); it can also contain a four-, five-, or six-membered saturated oxygen heterocycle; it can also contain a four-, five-, or six-membered saturated nitrogen heterocycle; or it can contain a six-membered aromatic carbon ring.

[0012] In this invention, the connecting arm -L- may also contain an amide bond and / or a urea group.

[0013] In this invention, the aforementioned linking arms -L- may further contain one or more substituents, which may include: -F, -Cl, -CH3, -C2H5, -C3H7, -C5H9, -C6H 13 , -OCH3, -OC2H5, -CH2F, -CHF2, -CF3, -C(O)-NH2, -SCH3, -SC2H5, -NHC(O)CH3, -N(CH3)2 and -N(C2H5)2.

[0014] The sugar chain structure synthesized in this invention contains basic (ethyl) and acidic (carboxyl) groups, which can form corresponding salts with organic or inorganic acids or bases.

[0015] The sugar chain structure synthesized in this invention contains both basic (ethyl) and acidic (carboxyl) groups, and can also be transferred intramolecularly via proton migration, i.e., the proton of the acidic group is transferred to the basic group. The general formula may contain -O. - and -NH3 + Amphoteric molecules.

[0016] This invention provides a chemical synthesis method for modifying the linker arm of a polysaccharide linker arm of Pseudomonas aeruginosa O5 serotype O-antigen (as shown in general formula VI), using three monosaccharide building blocks A, B, and C as raw materials, as shown in formulas (I) to (III), respectively:

[0017] in:

[0018] PG1 is H;

[0019] PG2, PG3, PG4 and PG6 are temporary hydroxyl protecting groups, each independently selected from: acetyl (Ac), or benzoyl (Bz), or neopentyl (Piv), or chloroacetyl (ClAc), or acetylpropionyl (Lev), or 9-fluorenylmethoxycarbonyl (Fmoc), or allyloxycarbonyl (Alloc), or 2-naphthylmethyl (Nap), or p-methoxybenzyl (pMB), or allyl (All), or tert-butyldimethylsilyl (TBS), or triethylsilyl (TES);

[0020] PG5 is a carboxyl protecting group, which can be benzyl (Bn), methyl (Me), ethyl (Et), tert-butyl (tBu), or allyl (All);

[0021] PG7 is a temporary protecting group for the amino group, which can be trichloroethoxycarbonyl (Troc), phthaloyl (Phth), 9-fluorenylmethoxycarbonyl (Fmoc), tert-butyloxycarbonyl (Boc), etc.

[0022] PG8 and PG9 are hydroxyl protecting groups, which can be benzene methylene (PhCH), naphthyl methylene (NapCH), isopropyl ketal ((CH3)2CH), etc.

[0023] PG 10 PG 11 It is an amino protecting group, which can be benzyl (Bn) or benzyloxycarbonyl (Cbz);

[0024] LG is a leaving group used in glycosylation reactions, such as fluorine (F), chlorine (Cl), bromine (Br), iodine (I), trichloroacetylimide ester (CCl3C(=NH)O-), N-phenyltrifluoroacetylimide ester glycoside (CF3C(=NPh)O-), ethylthio (SEt), phenylthio (SPh), p-toluenethio (STol), ethylthio (SEt), or dibutylphosphonic acid (-P(=O)-(OBu)2), etc.

[0025] The process includes the following:

[0026] (1) By glycosylation reaction of monosaccharide building block B (D-glucuronic acid building block) and monosaccharide building block A (D-fucoglycosamine building block), the disaccharide fragment shown in Formula IV was synthesized.

[0027] (2) The obtained disaccharide fragment was first deprotected and the protecting group PG4 was removed. Then, it was glycosylated with the monosaccharide building block C (3-amino-D-glucose building block) to synthesize the trisaccharide intermediate fragment as shown in Formula V.

[0028] (3) The obtained trisaccharide intermediate fragment was reduced, and PG3 and PG6 were deprotected. The deprotected position 2 in monosaccharide building block B and the deprotected position 2 in monosaccharide building block C were azidified. Then, the positions 4 and 6 of monosaccharide building block C in the trisaccharide intermediate fragment were reduced, PG8 and PG9 were removed, and the position 6 was oxidized to obtain a carboxylic acid group. The deprotected position 2 in monosaccharide building block B and the azid group at position 2 in monosaccharide building block C were reduced to obtain an acetamido group. Then, an imine reagent was used to construct an acetamidine structure at position 3 in monosaccharide building block C. Finally, the amino group in the linker was reduced to obtain the target product shown in Formula VI.

[0029] In one embodiment of the present invention, the synthetic route of the method is as follows:

[0030] This invention utilizes D-glucuronic acid building blocks and D-fucosamine building blocks to construct O-antigen trisaccharides. The stereoselective synthesis of the D-fucosamine 1,2-α-cis-glycosidic bond depends on the participation of a long-range acyl group and reagent regulation. The synthesis of the two 2,3-diaminomannuronic acid 1,2-β-trans-glycosidic bonds is achieved through SN2 nucleophilic substitution of the C2-position azide group. Through selective assembly of protecting groups, orthogonal modification of modifying groups, and regulation of glycosyl donor and acceptor reactivity, the preparation of multifunctional modified O-antigen target trisaccharides and their oligosaccharide mimics was successfully completed. The synthesized O-antigen oligosaccharide fragments were immobilized on a glass slide to prepare a sugar chip. Effective antigens were screened using patient serum to determine the O-antigen oligosaccharide epitopes. The raw materials are inexpensive and readily available, and the preparation method is simple and reproducible, showing great promise for the development of vaccines against Pseudomonas aeruginosa.

[0031] In one embodiment of the present invention, the method specifically includes: using 3,4-O-diacetylfucose as a raw material, reacting it with trimethyl azidosilane (TMS-N3) and diphenyldiselenose (Ph2Se2) under the action of diacetate iodobenzene (PhI(OAc)2) to generate 1-selenobenzene-2-azidofucose, then removing the acetyl group under alkaline conditions, selectively protecting the C3 hydroxyl group to 3-ONap, and protecting the C4 hydroxyl group with benzoyl group, and finally obtaining a highly stereoselective 1,2-α-cis-D-fucosamine glycosidic bond under the synergistic effect of additive effect (Ph3PO) and acyl group long-range participation effect (Bz).

[0032] In one embodiment of the present invention, the method further includes: using glucose as a raw material, obtaining a C3-amino precursor by nucleophilic substitution of the C3-position azido group mediated by trifluorosulfonyl group, and similarly using SN2 nucleophilic substitution of the C2-position azido group for the C2-position amino precursor, thereby completing the efficient construction of two 2,3-diaminomannuronic acid 1,2-β-trans-glycosidic bonds.

[0033] In one embodiment of the present invention, the Lewis acid includes boron trifluoride ether (BF3·Et2O), trifluoromethanesulfonic acid (TfOH), trimethylsilyl trifluoromethanesulfonate (TMSOTf), or silver salt silver trifluoromethanesulfonate (AgOTf), etc., and the thioglycoside is reacted by adding NIS together with the Lewis acid as a promoter.

[0034] In one embodiment of the present invention, the glycosylation reaction in step (1) is carried out in an environment of activating reagent Lewis acid, solvent, and molecular sieve.

[0035] In one embodiment of the present invention, the Lewis acid used as the activating agent for the glycosylation reaction in step (1) is any one or more of boron trifluoride diethyl ether (BF3·Et2O), trifluoromethanesulfonic acid (TfOH), trimethylsilyl trifluoromethanesulfonate (TMSOTf), or silver trifluoromethanesulfonate silver (AgOTf).

[0036] In one embodiment of the present invention, the glycosylation reaction in step (2) is carried out in an environment containing Lewis acid as an activating reagent, NIS (iodosuccinimide), solvent, and molecular sieve.

[0037] In one embodiment of the present invention, the amount of Lewis acid used as the activating agent for the glycosylation reaction in step (2) is 0.1 to 1 eq, and the amount of NIS is 1.2 to 1.5 eq.

[0038] In one embodiment of the present invention, the Lewis acid used as the activating agent for the glycosylation reaction in step (2) is any one or more of boron trifluoride diethyl ether (BF3·Et2O), trifluoromethanesulfonic acid (TfOH), trimethylsilyl trifluoromethanesulfonate (TMSOTf), or silver trifluoromethanesulfonate silver (AgOTf).

[0039] The purpose of this invention is to provide a method for synthesizing a Pseudomonas aeruginosa O5 serotype O-antigen trisaccharide assembled with an amino linker arm. The O5 serotype O-antigen oligosaccharide fragment prepared by the above method may be used in the development of Pseudomonas aeruginosa vaccines or Pseudomonas aeruginosa infectious drugs, etc. Beneficial effects:

[0040] This invention presents, for the first time, a chemical synthesis method for the O-antigen trisaccharide of Pseudomonas aeruginosa O5 serotype. The method utilizes D-glucuronic acid and D-fucosamine building blocks to construct the O-antigen trisaccharide. The stereoselective synthesis of the D-fucosamine 1,2-α-cis-glycosidic bond depends on the participation of a long-range acyl group and reagent regulation. The synthesis of the two 2,3-diaminomannuronic acid 1,2-β-trans-glycosidic bonds is achieved through SN2 nucleophilic substitution of the C2-position azido group. Through selective assembly of protecting groups, orthogonal modification of modifying groups, and regulation of the reactivity of glycosyl donors and acceptors, the preparation of the multifunctional modified O-antigen target trisaccharide and its oligosaccharide mimics was successfully completed. The synthesized O-antigen oligosaccharide fragments were immobilized on a glass slide to prepare a sugar chip. Effective antigens were screened using patient serum to determine the O-antigen oligosaccharide epitopes. The raw materials are inexpensive and readily available, and the preparation method is simple and reproducible, showing great promise for the development of vaccines against Pseudomonas aeruginosa. Attached Figure Description

[0041] Figure 1 shows the trisaccharide repeat unit of the O-antigen of Pseudomonas aeruginosa O5 serotype.

[0042] Figure 2 shows compounds A, B, and C as represented by general formulas I, II, and III.

[0043] Figure 3 shows compound F as shown in general formula VI.

[0044] Figure 4 shows the synthesis of sugar block 9.

[0045] Figure 5 shows the synthesis of sugar block 16.

[0046] Figure 6 shows the synthesis of sugar block 19.

[0047] Figure 7 shows the synthesis of trisaccharide 20.

[0048] Figure 8 shows the synthesis of the target trisaccharide 30.

[0049] Figure 9 shows an attempt to synthesize trisaccharide 35.

[0050] Figure 10 shows an attempt to synthesize trisaccharide 38.

[0051] Figure 11 shows an attempt to synthesize trisaccharides 43 and 44. Detailed Implementation

[0052] The embodiments of the present invention will be described in detail below with reference to examples. However, those skilled in the art will understand that the following examples are for illustrative purposes only and should not be considered as limiting the scope of the invention. Unless otherwise specified in the examples, conventional conditions or conditions recommended by the manufacturer are followed. Reagents or instruments used without specified manufacturers are all commercially available conventional products.

[0053] The yield calculation method of this invention is "product (mol) / reaction substrate (mol) * 100%". The methods for identifying the compound structures in this invention include nuclear magnetic resonance (NMR) spectroscopy (400MHz, 600MHz), high-resolution mass spectrometry, optical rotation determination, and infrared spectroscopy. The results are listed in the specific synthesis of each compound.

[0054] Example 1

[0055] Crafting Sugar Block 9:

[0056] As shown in Figure 4, starting with allyl α-D-aloose 1, the benzoyl group (Bz) was removed using sodium methoxide to obtain the corresponding diol. Then, a naphthyl methylene group (Nap) was assembled at the 2-OH position via a tin-mediated regioselective 2-naphthylmethyleneization reaction to yield compound 2. Subsequently, the O3 in compound 2 was trifluoroacetylated, followed by azido substitution to obtain compound 3 with an azide group at the C3 position. The 2-O-Nap group in compound 13 was removed using 2,3-dichloro-5,6-dicyano-1,4-benzoquinone (DDQ), and its acetylpropionyl group was protected to obtain compound 5. Subsequently, the 4,6-O-benzyl group was hydrolyzed using 80% acetic acid solution to obtain diol 6. Then, the C6 hydroxyl group was oxidized using TEMPO / BAIB, followed by benzyl esterification of the carboxylic acid to obtain compound 7. The temporary protecting group tert-butyldimethylsilyl (TBS) was assembled at the C4 hydroxyl group to obtain compound 8. Finally, the terminal allyl group is hydrolyzed and converted into trifluoroacetylimine ester donor 9.

[0057] Specific experimental procedures and steps:

[0058] Compound 2: Compound 1 (6.6 g, 16.01 mmol) was dissolved in methanol (40 mL) solution by stirring, and NaOMe (0.43 g, 8.01 mmol) was added. The reaction was stirred at room temperature for 5 h, and then analyzed using Amberlite IR 120 (H2O) reagent. + The solution was neutralized with ion exchange resin and filtered. The filtrate was concentrated under reduced pressure to obtain the crude product. The crude product was purified by silica gel column chromatography (PE / EA v / v 2:1) to obtain the corresponding diol compound. The above compound was azeotropically reacted with anhydrous toluene and dried under high vacuum for 30 min. Then, under nitrogen protection, anhydrous toluene (80 mL) was added, followed by Bu₂SnO (5.98 g, 24.02 mmol) and... Molecular sieves (flame-dried). The reaction was refluxed at 110 °C and stirred thoroughly for 3 h. The reaction was cooled to room temperature, and 2-bromomethylnaphthalene (5.32 g, 24.02 mmol) and TBAB (7.74 g, 24.02 mmol) were added. The mixture was stirred at 60 °C for 5 h. The reaction progress was monitored by TLC. After the reaction was complete, the reaction mixture was filtered and concentrated under reduced pressure to obtain a crude product. The crude product was purified by silica gel column chromatography (PE / EA v / v 6:1) to give syrupy compound 2 (5.88 g, 13.13 mmol, 82%). [α] 25 D =+105.1°(c=1.00,CHCl3); 1H NMR(400MHz,Chloroform-d)δ7.90–7.30(m,12H,Ar-H),5.95(dddd,J=17.0,10.4,6.3,5.2Hz,1H,All-CH),5.50(s,1H,ArCH),5.37(dq,J=17.2,1.5Hz,1H,All-CH2),5.25(dq,J=10.4,1.3Hz,1H,All-CH2),4.97–4.88(m,2H,1-H,ArCH),4.77(d,J=12.5Hz,1H,ArCH),4.47(dt,J=6.3,2.8Hz,1H,3-H),4.32(dd,J=10.2,5.1Hz,1H,6’-H),4.29–4.16(m,2H,All-CH2,5-H),4.07(ddt,J=12.9,6.3,1.3Hz,1H,All-CH2),3.67(t,J=10.2Hz,1H,6-H),3.53(t,J=3.4Hz,1H,2-H),3.39(dd,J=9.7,2.6Hz,1H,4-H),3.31(d,J=7.0Hz,1H,3-OH); 13 C NMR(101MHz,Chloroform-d)δ137.13,134.75,133.26,133.15,129.05,128.50,128.20,127.86,127.74,126.90,126.32,126.17,125.71,118.39,102.00,97.44,78.93,73.68,70.47,69.14,69.11,67.14,58.09;IR ν max (film)3510,2929,1601,1451,1316,1270,1177,1106,1070,1028,862,756,713cm -1 ;HR-ESI-MS(m / z):calcdfor C 27 H 28 O6Na + (M+Na + ):471.1778,found:471.1783.

[0059] Compound 3: Compound 2 (5.80 g, 12.94 mmol) was dissolved in anhydrous DCM (65 mL) under nitrogen atmosphere with stirring. Pyridine (10.6 mL, 129.4 mmol, 10 eq) was added at -20 °C. Tf₂O (4.35 mL, 25.88 mmol) was added dropwise, and the solvent was stirred to allow the temperature to rise from -20 °C to 0 °C over 4 h. The organic phase was dried over Na₂SO₄ and concentrated under vacuum at 30 °C to give a yellow syrupy trichloroate product. The trichloroate was dissolved in anhydrous DMF (65 mL) under nitrogen atmosphere, and TBAN₃ (11.04 g, 38.82 mmol) was added at 0 °C. The mixture was stirred thoroughly overnight at room temperature. The reaction progress was monitored by TLC. After the reaction was complete, the reaction solution was diluted with EA (150 mL) and washed with water and brine. The organic phase was dried over anhydrous Na₂SO₄ and concentrated under reduced pressure to give the crude product. The crude product was purified by silica gel column chromatography (PE / EA v / v 10:1) to give product 3 (4.65 g, 9.83 mmol, two-step yield 76%). [α] 25 D =+173.0°(c=1.00,CHCl3); 1 H NMR(400MHz,Chloroform-d)δ7.89–7.34(m,12H,Ar-H),5.92(dddd,J=17.1,10.3,6.6,5.2Hz,1H,All-CH),5.52(s,1H,ArCH),5.33(dd,J=17. 2,1.6Hz,1H,All-CH2),5.23(dd,J=10.3,1.4Hz,1H,All-CH2),4.96(d,J=12.2Hz,1H,ArCH),4.82(d,J=12.2Hz,1H,ArCH),4.77(d,J=3.6Hz,1H ,1-H),4.24(dd,J=10.3,4.9Hz,1H,6'-H),4.18(ddt,J=12.9,5.2,1.5Hz,1H,All-CH2),4.09(t,J=9.9Hz,1H,3-H),3.98(ddt,J=12.8,6.6,1. 3Hz,1H,All-CH2),3.89(td,J=10.0,4.9Hz,1H,5-H),3.65(t,J=10.3Hz,1H,6-H),3.47(dd,J=9.9,3.6Hz,1H,2-H),3.39(t,J=9.8Hz,1H,4-H); 13C NMR(101MHz,Chloroform-d)δ136.81,134.89,133.28,133.18,129.13,128.44,128.32,127.93,127.74,127.1 1,126.29,126.19,126.07,125.92,118.59,101.61,95.80,79.89,78.16,73.34,68.89,68.60,62.68,61.74; IR ν max (film)2868,2109,1451,1374,1268,1094,1071,1044,1028,995,929,751,713,699cm -1 HR-ESI-MS(m / z):calcd for C 27 H 27 N3O5Na + (M+Na + ):496.1843,found:496.1848.

[0060] Compound 4: Compound 3 (4.62 g, 9.76 mmol) was dissolved in DCM (90 mL) and H2O (10 mL) solution, and DDQ (3.33 g, 14.64 mmol) was added. The mixture was stirred at room temperature for 6.5 h, and the reaction progress was monitored by TLC. The reaction was quenched dropwise with 5% (w / v) Na2S2O3 solution, and the organic phase was washed with saturated NaHCO3 solution and brine. DCM was back-extracted. The organic phase was then dried over anhydrous Na2SO4 and concentrated under vacuum. The crude product was purified by silica gel column chromatography (PE / EA v / v 4:1) to give compound 4 (2.79 g, 8.39 mmol, 86%). [α] 25 D = +162.1° (c 1.00, CH3Cl). 1H NMR(400MHz,Chloroform-d)δ7.55–7.32(m,5H,Ar-H),5.93(dddd,J=16.9,10.3,6.4,5.4Hz,1H,All-CH),5.56(s,1H,ArCH),5.34(dq,J=17.2,1.5Hz,1H,All-CH2),5.27(dq,J=10.3,1.2Hz,1H,All-CH2),4.94(d,J=3.9Hz,1H,1-H),4.30(dd,J=10.3,4.9Hz,1H,6’-H),4.29–4.23(m,1H,All-CH2),4.08(ddt,J=12.6,6.3,1.3Hz,1H,All-CH2),3.89(td,J=9.9,4.9Hz,1H,5-H),3.84(t,J=9.8Hz,1H,3-H),3.73(t,J=10.3Hz,1H,6-H),3.61(td,J=10.0,3.9Hz,1H,2-H),3.48(t,J=9.7Hz,1H,4-H),2.26(d,J=10.4Hz,1H,2-OH); 13 C NMR(101MHz,Chloroform-d)δ136.78,132.98,129.12,128.32,126.01,118.68,101.53,97.37,79.63,77.22,71.78,68.97,68.85,63.74,63.01;IR ν max (film)3345,2916,1870,2105,1368,1260,1152,1077,1058,1013,994,931,746,698,653cm -1 ;HR-ESI-MS(m / z):calcd for C 16 H 19 N3O5Na + (M+Na + ):356.1217,found:356.1218.

[0061] Compound 5: Compound 4 (2.77 g, 8.32 mmol) was dissolved in a dry DCM (40 mL) solution, and levulinic acid (LevOH) (1.45 g, 12.48 mmol), N,N-dicyclohexylcarbodiimide (DCC) (2.57 g, 12.48 mmol), and DMAP (1.52 g, 12.48 mmol) were added. The mixture was stirred at room temperature for 4 h until the starting material was completely consumed, and the reaction solution was diluted with DCM. The organic phase was washed with saturated NaHCO3 and saturated NaCl solutions, and back-extracted with DCM. The organic phase was then dried over anhydrous Na2SO4 and concentrated under vacuum. The crude product was purified by silica gel column chromatography (PE / EA v / v 8:1) to give compound 5 (3.48 g, 8.07 mmol, 97%). [α] 25 D = +110.8° (c 1.00, CH3Cl); 1 H NMR(400MHz,Chloroform-d)δ7.54–7.33(m,5H,Ar-H),5.91(dddd,J=16.9,10.3,6.2,5.3Hz,1H,All-CH),5.58(s,1H,ArCH),5.32(dq,J=17 .1,1.6Hz,1H,All-CH2),5.24(dq,J=10.4,1.4Hz,1H,All-CH2),5.05(d,J=3.7Hz,1H,1-H),4.74(dd,J=10.3,3.7Hz,1H,2-H),4.29(dd,J=1 0.4,4.9Hz,1H,6'-H),4.21(ddt,J=12.9,5.3,1.5Hz,1H,All-CH2),4.12(t,J=10.1Hz,1H,3-H),4.04(ddt,J=12.9,6.2,1.4Hz,1H,All-CH2 ),3.95(td,J=9.9,4.9Hz,1H,5-H),3.75(t,J=10.3Hz,1H,6-H),3.54(t,J=9.7Hz,1H,4-H),2.92–2.56(m,4H,CH2-Lev),2.20(s,3H,CH3CO); 13 C NMR(101MHz,Chloroform-d)δ206.02,171.88,136.72,133.23,129.15,128.32,126.03,1 18.19,101.63,95.20,79.83,72.15,68.97,68.82,62.79,60.04,37.79,29.79,27.88;IR ν max(film)2923,2868,2109,1743,1718,1454,1368,1260,1208,1151,1123,1096,1072,1045,996,933,753,700,653cm -1 HR-ESI-MS(m / z):calcd for C 21 H 25 N3O7Na + (M+Na + ):454.1585,found:454.1581.

[0062] Compound 6: Compound 5 (3.46 g, 8.02 mmol) was dissolved in 80% AcOH (40 mL) solution. The system was heated to 55 °C using an oil bath and stirred. The reaction progress was monitored by TLC until the starting material was completely consumed. After removing most of the solvent by vacuum concentration, the product was purified by silica gel column chromatography (DCM / MeOH v / v 30:1) to give product 6 (2.64 g, 7.70 mmol, 96%). [α] 25 D = +96.6° (c 1.00, CH3Cl); 1 H NMR(400MHz,Chloroform-d)δ5.90(dddd,J=17.2,10.4,6.1,5.3Hz,1H,All-CH),5.31(dq,J=17.2,1.6Hz,1H,All-CH2),5.23(dq ,J=10.4,1.4Hz,1H,All-CH2),5.03(d,J=3.6Hz,1H,1-H),4.69(dd,J=10.5,3.6Hz,1H,2-H),4.19(ddt,J=13.0,5.3,1.5Hz,1H,A ll-CH2),4.02(ddt,J=13.0,6.1,1.4Hz,1H,All-CH2),3.94(dd,J=10.6,9.5Hz,1H,3-H),3.88–3.80(m,2H,6-H),3.73(dt,J=9.8 ,3.5Hz,1H,5-H),3.62–3.52(m,1H,4-H),3.34(s,1H,4-OH),2.84–2.62(m,4H,CH2-Lev),2.37(br,1H,6-OH),2.20(s,3H,CH3CO); 13C NMR (101MHz, Chloroform-d) δ206.30,172.03,133.35,118.03,94.57,72.29,70.98,69.06,68.76,63.65,61.78,37.80,29.79,27.90; IR ν max (film)3395,2925,2108,1743,1716,1417,1362,1258,1207,1155,1040,929,840,770,607cm -1 HR-ESI-MS(m / z):calcd for C 14 H 21 N3O7Na + (M+Na + ):366.1272,found:366.1275.

[0063] Compound 7: Compound 6 (2.64 g, 7.70 mmol) was dissolved in DCM (77 mL) and H2O (15 mL) solution under argon protection. The system was cooled to 0 °C using an ice bath, and 2,2,6,6-tetramethylpiperidine oxide (TEMPO) (240 mg, 1.54 mmol) and diacetoxyiodobenzene (BAIB) (4.96 g, 15.4 mmol) were added. The mixture was stirred at room temperature for 4 h. The reaction progress was monitored by TLC until the starting material was exhausted. The reaction was quenched dropwise with 5% (w / v) Na2S2O3 solution, and the organic phase was washed with saturated NaHCO3 and saturated NaCl solutions, followed by DCM back-extraction. The organic phase was then dried over anhydrous Na2SO4 and concentrated under vacuum. The crude product was dissolved in anhydrous DMF (77 mL) under nitrogen protection. NaHCO3 (3.23 g, 38.5 mmol) and BnBr (1.83 mL, 15.4 mmol) were added at room temperature. When TLC showed complete consumption of the starting material, the solvent was removed by vacuum concentration. Purification was then performed by silica gel column chromatography (PE / EA v / v 5:1). The two-step reaction yielded glucuronic acid syrup 7 (2.75 g, 6.16 mmol, 80%). [α] 25 D = +74.6° (c 1.00, CH3Cl); 1H NMR(400MHz,Chloroform-d)δ7.41–7.32(m,5H,Ar-H),5.90(dddd,J=17.0,10.4,6.2,5.3Hz,1H,All-CH),5.31(dq,J=17.2,1.6Hz,1H,All-CH2),5.29(d,J=12.3Hz,1H,ArCH),5.24(d,J=12.2Hz,1H,ArCH),5.22(dq,J=10.4,1.3Hz,1H,All-CH2),5.10(d,J=3.6Hz,1H,1-H),4.68(dd,J=10.6,3.6Hz,1H,2-H),4.27(d,J=9.8Hz,1H,5-H),4.25–4.19(m,1H,All-CH2),4.07(ddt,J=12.9,6.2,1.4Hz,1H,All-CH2),3.95(dd,J=10.6,9.5Hz,1H,3-H),3.77(td,J=9.7,2.8Hz,1H,4-H),3.17(s,1H,4-OH),2.87–2.55(m,4H,CH2-Lev),2.19(s,3H,CH3CO). 13 C NMR(400MHz,Chloroform-d)δ206.00,171.80,169.73,134.85,133.03,128.74,128.67,128.24,118.41,94.86,71.26,70.78,70.17,69.35,67.61,62.45,37.79,29.76,27.84;IR ν max (film)3480,2935,2110,1745,1717,1361,1259,1181,1155,1051,939,754,698,609cm -1 ;HR-ESI-MS(m / z):calcd for C 21 H 25 N3O8Na + (M+Na + ):470.1534,found:470.1537.

[0064] Compound 8: Anhydrous DMF (3.5 mL) was added to glucuronic acid 7 (1.55 g, 3.47 mmol), and the reaction mixture was stirred thoroughly under argon protection. Then, tert-butyldimethylsilyl chloride (1.05 g, 6.94 mmol), imidazole (472 mg, 6.94 mmol), and 4-dimethylaminopyridine (43 mg, 0.35 mmol) were added to the reaction flask. The mixture was stirred overnight at 80 °C. The reaction progress was monitored by TLC until the starting material was completely consumed. The reaction was then quenched with methanol (3 mL). The reaction mixture was diluted with ethyl acetate (50 mL), and the organic phase was washed with saturated NaHCO3 and NaCl solutions, followed by DCM back-extraction. The organic phase was then dried over anhydrous Na2SO4 and concentrated under vacuum. The crude product was purified by silica gel column chromatography (PE / EA v / v 10:1) to give colorless syrup 8 (1.73 g, 3.09 mmol, 89%). [α] 25 D = +74.5° (c 1.00, CH3Cl); 1 H NMR(600MHz,Chloroform-d)δ7.40–7.32(m,5H,Ar-H),5.90(dddd,J=16.8,10.2,6.3,5.3Hz,1H,All-CH),5.30(dd,J=17.2,1.6Hz,1H,All-CH2),5. 23(d,J=12.3Hz,1H,ArCH),5.25–5.20(m,1H,All-CH2),5.13(d,J=12.4Hz,1H,ArCH),5.07(d,J=3.6Hz,1H,1-H),4.75(dd,J=10.3,3.6Hz,1H,2-H),4 .23–4.20(m,1H,All-CH2),4.20(d,J=9.4Hz,1H,5-H),4.03(ddt,J=13.0,6.3,1.3Hz,1H,All-CH2),3.82(t,J=9.8Hz,1H,3-H),3.76(t,J=9.3Hz,1H, 4-H),2.84–2.73(m,2H,CH2-Lev),2.71–2.60(m,2H,CH2-Lev),2.19(s,3H,CH3CO),0.85(s,9H,SiC(CH3)3),0.15(s,3H,SiCH3),0.01(s,3H,SiCH3); 13C NMR (151MHz, CDCl3) δ205.90,171.79,168.70,134.98,133.17,128.63,128.49,128.26,118.43,94 .88,72.21,72.18,70.94,69.27,67.38,64.49,37.80,29.74,27.88,25.72,18.05,-4.43,-5.02; IR ν max (film)=2930,2857,2109,1748,1721,1456,1404,1361,1252,1182,1136,1052,939,838,781,749,697,669cm -1 HR-ESI-MS(m / z):calcd for C 27 H 39 N3O8SiNa + (M+Na + ):584.2399,found:584.2401.

[0065] Compound 9: PdCl2 (485 mg, 2.74 mmol) was added to a MeOH / DCM (v / v, 2 / 1, 30 mL) solution of Compound 8 (1.54 g, 2.74 mmol), and the mixture was stirred at 40 °C for 6 h. The mixture was filtered and concentrated under vacuum, and purified by silica gel column chromatography (PE / EA v / v 2:1) to obtain the corresponding hemiacetal, a pale yellow syrup. This pale yellow syrup was dissolved in DCM (30 mL). 2,2,2-trifluoro-N-phenylacetylimine chloride (2.05 mL, 13.7 mmol) and DBU (1.23 mL, 8.22 mmol) were added to the reaction mixture at 0 °C. The reaction system was heated to room temperature and stirred for 3 h. The reaction progress was monitored by TLC until the starting materials were completely consumed. The reaction solution was concentrated under vacuum, and the crude product was purified by silica gel column chromatography (PE / EA v / v 20:1→10:1) to obtain a pale yellow syrup 9 (1.43 g, 2.06 mmol, 75%). [α] 25 D = +29.9° (c 1.00, CH3Cl); 1H NMR(600MHz,Chloroform-d)δ7.39–6.77(m,10H,Ar-H),5.28(d,J=12.3Hz,1H,ArCH ),5.20–5.15(m,1H,2-H),5.13(d,J=12.3Hz,1H,ArCH),4.10–3.78(m,2H,3-H,5-H) ,3.62–3.44(m,1H,4-H),2.85–2.79(m,2H,CH2-Lev),2.70–2.59(m,2H,CH2-Lev),2 .18(s,3H,CH3CO),0.84(s,9H,SiC(CH3)3),0.16(s,3H,SiCH3),0.01(s,3H,SiCH3); 13 C NMR(151MHz,Chloroform-d)δ205.73,171.02,166.98,142.94,134.69,128.78,128.67,128.63,128.34,124 .56,119.26,94.44,77.40,70.86,70.45,67.63,67.48,37.86,29.66,27.72,25.64,17.97,-4.53,-5.09; IR ν max (film)=2930,2857,2109,1753,1720,1597,1489,1405,1363,1327,1257,1212,1162,1137,1090,910,839,779,754,696,584cm -1 HR-ESI-MS(m / z):calcd for C 32 H 39 F3N4O8Na + (M+Na + ):715.2381,found:715.2377.

[0066] Example 2

[0067] Crafting Sugar Block 16:

[0068] As shown in Figure 5, using D-fucose as a starting material, dilute sugar 10 was obtained through acetylation, bromination, and elimination reactions. Subsequently, an azidization reaction was carried out under the action of diphenyldiselenyl (Ph₂Se₂) and trimethylsilyl azide (TMSN₃) to give compound 11 with an azide group at the C2 position. Then, deacetylation was performed under the action of sodium methoxide, followed by selective 2-naphthylmethyleneization on O₃ to give compound 13. Benzyl bromide and sodium hydride (NaH) were used in N,N-dimethylformamide (DMF) solvent for benzylation on O₄ to give compound 14. Subsequently, donor 14 was glycosylated with N-benzyl-N-benzyloxycarbonyl-3-aminopropanol in diethyl ether / dichloromethane to give compound 15. Finally, the 2-naphthylmethylene group (Nap) was removed under the action of DDQ to give glycosyl acceptor 16.

[0069] Specific experimental procedures and steps:

[0070] Compound 10: D-fucose (5.0 g, 30.5 mmol) was dissolved in Ac₂O (25.0 mL, 0.266 mol) under nitrogen atmosphere. The reaction system was cooled to 0 °C, and HClO₄ (50 μL, 0.83 mmol) was added. After stirring at room temperature for 30 min, 100 mL of ice water was added to the reaction solution. The organic phase was then washed with saturated NaHCO₃ solution (3 × 100 mL) and saturated NaCl solution (100 mL), followed by back-extraction with DCM (3 × 50 mL). The organic phase was then dried over anhydrous Na₂SO₄ and concentrated under vacuum. The organic layer was concentrated to 15 mL and used directly in the next step.

[0071] Under nitrogen atmosphere, peracetylglucose was dissolved in dry DCM (15 mL). The reaction system was cooled to 0 °C, and HBr-AcOH (33% w / w, 10 mL, 57.8 mmol) was added dropwise over 30 min. The system temperature was then raised to room temperature, and after stirring for 7 h, the reaction solution was diluted with DCM (50 mL), washed with ice water (3 × 100 mL), saturated NaHCO3 (3 × 100 mL), and brine (100 mL), and dried over Na2SO4. After concentration under high vacuum, the crude product was directly used for the next step without further purification.

[0072] Under nitrogen atmosphere, the crude product was dissolved in ethyl acetate (100 mL), and saturated NaH₂PO₄ (50 mL) was added. After the reaction was complete as monitored by TLC, the solution was filtered through diatomaceous earth, and the filtrate was washed with saturated NaHCO₃ (3 × 100 mL) and back-extracted with ethyl acetate (3 × 50 mL). The organic phase was then dried over Na₂SO₄ and concentrated. The concentrated crude product was purified by silica gel column chromatography (PE / EA v / v 20:1) to obtain 10 (4.11 g, 19.2 mmol, 63%) of colorless syrup. [α]25 D = +22.1° (c = 1.00, CHCl3); 1 H NMR(400MHz,Chloroform-d)δ6.46(dd,J=6.3,1.9Hz,1H,1-H),5.58(ddd,J=4.1,2.1,1.0Hz,1H,4-H),5.29(dt,J=4.7,1.6Hz,1H, 2-H),4.64(dt,J=6.4,2.0Hz,1H,3-H),4.30–4.12(m,1H,5-H),2.16(s,3H,CH3CO),2.02(s,3H,CH3CO),1.28(d,J=6.6Hz,3H,6-H); 13 C NMR (101MHz, Chloroform-d) δ170.72,170.42,146.12,98.27,71.54,66.30,65.07,20.87,20.71,16.53; IR ν max (film)1747,1650,1372,1243,1163,1091,1073,1028,989,924,892,851,761cm -1 ;HR-ESI-MS(m / z):calcdfor C 10 H 14 O5Na + (M+Na + ):237.0733,found:237.0734.

[0073] Compound 11: Compound 10 (3.7 g, 17.3 mmol) was dissolved in dry DCM (86 mL), followed by the addition of diphenyldiselenoether (5.4 g, 17.3 mmol). The solution was cooled to -30 °C under argon atmosphere, and then BAIB (5.6 g, 17.3 mmol) and azidotrimethylsilane (3.98 g, 34.6 mmol) were added. The reaction progress was monitored by TLC until the starting material was exhausted. The mixture was washed with saturated NaHCO3 (3 × 100 mL) and back-extracted with DCM (3 × 50 mL). The organic phase was then dried over Na2SO4 and concentrated under vacuum. The crude product was purified by silica gel column chromatography (PE / EA v / v 5:1) to give a pale yellow syrupy product 11 (5.0 g, 12.1 mmol, 70%). [α] 25 D = +204.3° (c 1.00, CH3Cl); 1H NMR(400MHz,Chloroform-d)δ7.68–7.49(m,2H,Ar-H),7.41–7.13(m,3H,Ar-H),5.95(d,J=5.4Hz,1H,1-H),5.32(dd,J=3.3,1.2Hz,1H,4-H),5.13(dd, J=10.8,3.2Hz,1H,3-H),4.57–4.44(m,1H,5-H),4.24(dd,J=10.8,5.4Hz,1 H,2-H),2.17(s,3H,CH3CO),2.07(s,3H,CH3CO),1.09(d,J=6.5Hz,3H,6-H); 13 C NMR(101MHz,Chloroform-d)δ170.44,169.83,134.77,129.29,128.15,84.54,71.74,70.27,67 .54,58.88,20.78,15.93; IR(film):ν=2110,1747,1368,1233,1084,1020,954,908,741,692cm -1 HR-ESI-MS(m / z):calcd for C 16 H 19 N3O5SeNa[M+Na] + 436.0382, found 436.0385.

[0074] Compound 12: Compound 11 (4.6 g, 11.1 mmol) was dissolved in methanol (55 mL), and NaOMe (0.30 g, 5.55 mmol) was added. The solution was stirred at room temperature for 5 h. The solution was purified using Amberlite IR 120 (H) + The sample was neutralized with ion exchange resin and then filtered through a cotton plug. The filtrate was concentrated under vacuum, and the crude product was purified by silica gel column chromatography (dichloromethane / methanol v / v 100:1) to give product 12 (3.32 g, 10.1 mmol, 91%). [α] D 25 =+247.6°(c=1.00,CHCl3); 1 HNMR (400MHz, Methanol-d4)δ=7.66–7.21(m,5H,Ar-H),5.94(d,J=5.3Hz,1H,1-H),4.32(q,J=6.5Hz ,1H,5-H),4.04(dd,J=10.0,5.2Hz,1H,2-H),3.79–3.71(m,2H,3-H,4-H),1.18(d,J=6.5Hz,3H,6-H);13 C NMR (100MHz, Methanol-d4) δ135.88,130.02,128.70,86.91,72.91,72.66,70.62,62.92,16.41; IR ν max (film)2106,1093,1058,988,827,761,689,670,633cm -1 HR-ESI-MS(m / z):calcd for C 12 H 15 N3O3SeNa + (M+Na + ):352.0171,found:352.0176.

[0075] Compound 13: Diol 12 (1.76 g, 5.35 mmol) was azeotropically reacted with anhydrous toluene and dried under high vacuum for 30 min. Then, under nitrogen protection, anhydrous toluene (50 mL) was added, followed by Bu₂SnO (2.0 g, 8.03 mmol) and... Molecular sieves (flame-dried). The reaction was stirred under reflux for 2 h. The reaction mixture was cooled to room temperature, and 2-bromomethylnaphthalene (1.78 g, 8.03 mmol) and TBAB (2.59 g, 8.03 mmol) were added. The mixture was stirred at 60 °C for 4 h. The reaction mixture was filtered, and the solvent was removed by concentration under reduced pressure. The crude product was purified by silica gel column chromatography (PE / EA v / v 6:1) to give a white solid compound 13 (1.95 g, 4.16 mmol, 78%). [α] D 25 =+125.9°(c=1.00,CHCl3); 1 H NMR(400MHz,Chloroform-d)δ7.91–7.26(m,12H,Ar-H),5.90(d,J=5.3Hz,1H,1-H),4.89(t,J=11.9Hz,2H,ArCH),4.29(q,J=6.6Hz,1H, 5-H),4.20(dd,J=10.1,5.3Hz,1H,2-H),3.90(dd,J=3.3,1.4Hz,1H,4-H),3.75(dd,J=10.2,3.1Hz,1H,3-H),1.25(d,J=6.5Hz,3H,6-H); 13C NMR(101MHz,Chloroform-d)δ134.46,133.27,133.24,129.11,128.65,128.55,128.01,127.80 ,127.79,127.03,126.40,126.32,125.69,85.22,79.24,72.32,68.64,60.34,29.71,16.05;IR ν max (film)3500,3054,2975,2925,2111,1578,1509,1476,1438,1346,1271, 1215,1163,1091,1065,1021,999,936,857,817,759,670,692,630,573cm -1 HR-ESI-MS(m / z):calcd for C 23 H 23 N3O3SeNa + (M+Na + ):492.0797,found:492.0799.

[0076] Compound 14: Compound 13 (127 mg, 0.27 mmol) was dissolved in anhydrous DMF (3 mL) and NaH (22 mg, 0.54 mmol, 60% dispersible in mineral oil) was added. The mixture was stirred at 0 °C for 10 min. Then, BnBr (50 μL, 0.41 mmol) was added to the reaction mixture, and the mixture was stirred at room temperature for 4 h. After the reaction was complete, the mixture was cooled to 0 °C, and methanol was added dropwise. The reaction mixture was then washed with water and brine, and back-extracted using DCM. The organic phase was dried over anhydrous Na₂SO₄, concentrated under vacuum, and the crude product was purified by silica gel column chromatography (PE / EA v / v 20:1) to obtain compound 14 (135 mg, 0.24 mmol, 89%). [α] D 25 =+202.8°(c=1.00,CHCl3); 1H NMR(400MHz,Chloroform-d)δ7.91–7.24(m,17H,Ar-H),5.94(d,J=5.3Hz,1H,1-H),4.93(m,3H,ArCH),4.64(d,J=11.5Hz,1H,ArCH),4.39(dd, J=10.3,5.3Hz,1H,2-H),4.22(q,J=6.5Hz,1H,5-H),3.78(dd,J=10.4,2.7Hz,1H,3-H),3.73(d,J=2.8Hz,1H,4-H),1.13(d,J=6.5Hz,3H,6-H); 13 C NMR(101MHz,Chloroform-d)δ138.14, 134.98,134.38,133.31,133.11,129.01,128.72,128.38,128.30,128.12,127.98,127.75,127.64, 126.62,126.25,126.10,125.67,85.56,80.65,75.97,75.04,72.67,69.45,61.09,29.70,16.54; IR ν max (film)3056,2976,2882,2108,1718,1602,1578,1509,1496,1476,1454,1438,1346,1297,12 69,1213,1158,1102,1080,1065,1022,999,970,895,856,816,739,691,671,631,596,566cm -1 HR-ESI-MS(m / z):calcd for C 30 H 29 N3O3SeNa + (M+Na + ):582.1266,found:582.1268.

[0077] Compound 15: Compound 14 (20 mg, 0.034 mmol) was added to anhydrous DCM / Et2O (v / v 3:1, 1 mL) at -20 °C under argon protection with stirring. Then N-benzyl-N-benzyloxycarbonyl-3-aminopropanol (15 mg, 0.051 mmol) was added and Molecular sieves (flame-dried). The reaction system temperature was lowered to 0°C, and NIS (9.2 mg, 0.041 mmol) and TMSOTf (1.27 μL, 0.007 mmol) were added. The reaction was stirred at 0°C for 3 h until TLC showed complete conversion of the starting material. The reaction solution was diluted with DCM and filtered. The organic phase was treated with 10% (w / v) Na₂S₂O₂. 3、 The product was washed with saturated NaHCO3 and NaCl solutions and back-extracted using DCM. The organic phase was then dried over anhydrous Na2SO4 and concentrated under vacuum. The crude product was purified by silica gel column chromatography (PE / EA v / v 10:1→5:1) to give product 15 (20 mg, 0.029 mmol, 84%, α:β = 1:1.1).

[0078] Compound 16: Compound 26 (1.45 g, 2.07 mmol) was dissolved in DCM / H2O (v / v 9:1, 40 mL), and DDQ (0.71 g, 3.11 mmol) was added. After stirring at room temperature for 6 h, the mixture was quenched with 5% (w / v) Na2S2O3 solution, washed with saturated NaHCO3 and brine, and back-extracted with DCM. The organic phase was dried over anhydrous Na2SO4 and concentrated under vacuum. The crude product was purified by silica gel column chromatography (PE / EA v / v 5:1→4:1) to give compound 11 (0.92 g, 1.64 mmol, 79%). [α] 25 D = +108.1° (c 1.00, CH3Cl); 1 H NMR(400MHz,Chloroform-d)δ7.62–7.04(m,15H,Ar-H),5.17(d,J=5.8Hz,2H,ArCH),4.79(d,J= 11.7Hz,1H,ArCH),4.82–4.72(m,1H,1-H),4.66(d,J=11.5Hz,1H,ArCH),4.56(d,J=15.5Hz,1H,A rCH),4.51–4.40(m,1H,ArCH),4.08–3.73(m,2H,3-H,5-H),3.71–3.52(m,2H,CH2-Linker,4-H), 3.51–3.21(m,4H,2-H,CH2-Linker),1.94–1.68(m,2H,CH2-Linker),1.22(d,J=6.1Hz,3H,6-H); 13C NMR(101MHz,Chloroform-d)δ156.72,156.25,137.82,136.73,128.71,128.58,128.24,128.15,128.01,127.90,1 27.36,98.24(C-1),80.17,76.20,68.59,67.30,66.48,65.81,60.91,50.87,44.68,43.83,28.37,27.87,16.79; IR ν max (film)3444,2925,2106,1695,1496,1496,1454,1421,1359,1221,1165,1127,1103,1042,968,914,735,698cm -1 HR-ESI-MS(m / z):calcd for C 31 H 36 N4O6Na + (M+Na + ):583.2527,found:583.2524.

[0079] Example 3

[0080] Synthesis optimization of 1,2-cis-α-D-fucoglycoside:

[0081] As shown in Table 1, a series of glycosyl donors were tested under different conditions to improve the α-glycosidic bond selectivity of D-fucoside. The reaction efficiency of trifluoroacetylimine donor 14a was investigated using TMSOTf as an activator in diethyl ether / dichloromethane. The resulting product 15 showed a slight improvement in glycosidic bond selectivity, reaching α:β = 1.5:1. Subsequently, donor 14a was glycosylated with N-benzyl-N-benzyloxycarbonyl-3-aminopropanol using TMSI-Ph3PO reagent, yielding product 15 with significantly improved α-configuration selectivity (α / β = 4.2:1). Given the synergistic strategy combining the long-range participation effect of the acyl group and the reagent effect, the stereoselectivity of 1,2-cis-α-aminoglycosides can be enhanced. Therefore, Ac, Bz, and Lev were assembled at the O4 position of compound 13 to obtain donors 14b, 14c, and 14d, respectively, to utilize their long-range participation effect of the acyl group to generate α-D-fucoside. Finally, with the synergistic effect of remote involvement and solvent effect, target glycosides 15a, 15b, and 15c were obtained, all with high yields and excellent stereoselectivity, with α / β selectivities of 7.5:1, 13.5:1, and 12.1:1, respectively. Ultimately, the benzoyl group was chosen as the temporary protecting group at the O4 position (Bz) because the α-configuration isomer of compound 15c was easier to isolate and had a higher yield (91%). Finally, the benzoyl group of compound 15c was removed using sodium methoxide, followed by benzylation to obtain compound 15.

[0082] Table 1. Optimization of the synthesis of 1,2-cis-α-D-fucosamine glycosides

[0083] Specific experimental procedures and steps:

[0084] Compound 14a: At room temperature, NBS (166 mg, 0.93 mmol) was added to a mixture of Compound 14 (130 mg, 0.31 mmol) stirred in THF / H₂O (v / v 4:1, 3 mL), and the mixture was stirred for 2.5 h. When TLC showed complete conversion of the starting material, the mixture was diluted with DCM and washed with 10% (w / v) Na₂S₂O₃ and brine. The organic phase was dried over anhydrous Na₂SO₄, filtered, and concentrated under reduced pressure. The crude product was purified by silica gel column chromatography (PE / EA v / v 4:1→2:1) to obtain the corresponding hemiacetal as a colorless syrup. This hemiacetal was dissolved in DCM (3 mL). The reaction system temperature was lowered to 0 °C, and 2,2,2-trifluoro-N-phenylacetylimine chloride (230 μL, 1.55 mmol) and DBU (140 μL, 0.93 mmol) were added. The reaction system temperature was then raised to room temperature and stirred for 3 h. The reaction progress was monitored by TLC until the reactants were completely consumed. The reaction solution was concentrated under reduced pressure at room temperature to obtain a crude product, which was then purified by silica gel column chromatography (PE / EA v / v 20:1→10:1) to obtain a pale yellow syrup 14a (153 mg, 0.26 mmol, 83%). [α] 25 D = +33.7° (c 1.00, CH3Cl); 1 H NMR(400MHz,Chloroform-d)δ8.13–6.41(m,17H,Ar-H),4.98(d,J=11.6Hz,1H,ArCH),4.89(s,2H,ArCH),4.70(d,J= 11.5Hz,1H,ArCH),4.17–3.99(m,1H,5-H),3.58(s,1H,4-H),3.56–3.28(m,2H,2-H,3-H),1.20(d,J=6.1Hz,3H,6-H); 13 CNMR(100MHz,Chloroform-d)δ143.43,137.97,135.09,133.25,133.16,129.43,128.70,128.47,128.42,128.36,127.96,127.91,127. 79,126.78,126.41,126.36,126.21,125.71,124.29,120.47,119.33,95.97,80.88,74.94,74.68,72.90,71.90,62.17,29.72,16.70; IR ν max(film)2931,2114,1720,1598,1549,1489,1452,1318,1285,1211,1163,1107,1082,1028,910,856,818,777,753,732,695cm -1 HR-ESI-MS(m / z):calcd for C 32 H 29 F3N4O4Na + (M+Na + ):613.2033,found:613.2037.

[0085] Compound 15: Donor 14a (20 mg, 0.034 mmol) and the linker arm N-benzyl-N-benzyloxycarbonyl-3-aminopropanol (15 mg, 0.051 mmol) were azeotropically reacted with toluene and dried under vacuum overnight. These substances were then dissolved in dry DCM / Et2O (v / v 3:1, 1 mL) and added... Molecular sieve (flame-dried). Cool the reaction system to 0°C. Under argon protection, add TMSOTf (1.27 μL, 0.007 mmol) to the reaction solution. Stir the reaction solution at 0°C until the reaction is complete as monitored by TLC. Then quench the reaction with triethylamine dropwise and filter. Molecular sieves were used and diluted with DCM. The organic phase was washed with saturated NaHCO3, then dried over anhydrous Na2SO4 and concentrated under vacuum. The crude product was purified by silica gel column chromatography (PE / EA v / v 10:1→5:1) to give compound 15 (21 mg, 0.03 mmol, 87%, α:β = 1.5:1).

[0086] Donor 14a (20 mg, 0.034 mmol) and the linker N-benzyl-N-benzyloxycarbonyl-3-aminopropanol (15 mg, 0.051 mmol) were azeotropically dissolved in anhydrous toluene, and then dissolved in dry DCM (1 mL) under argon protection. Add [the following to the reaction flask]... Molecular sieves (flame-dried) and Ph3OP (67 mg, 0.24 mmol) were added. Then, TMSI (5 μL, 0.034 mmol, 1.0 eq) was slowly added dropwise to the reaction flask. The reaction was stirred at room temperature until TLC monitoring showed completion. The reaction was quenched with saturated Na2S2O3, and the solid was filtered off. Molecular sieves were used and washed with DCM. The organic phase was washed with water and brine and dried over anhydrous Na2SO4. The crude product was then concentrated under vacuum. The crude product was purified by silica gel column chromatography (PE / EA v / v 10:1→5:1) to give compound 15 (22 mg, 0.031 mmol, 91%, α:β = 4.2:1).

[0087] 15α:[α] 25 D = +74.6° (c 1.00, CH3Cl). 1 H NMR(600MHz,Chloroform-d)δ8.05–7.05(m,22H,Ar-H),5.17(d,J=9.7Hz,2H,ArCH),4.93(d,J =11.5Hz,1H,ArCH),4.89–4.75(m,3H,ArCH,1-H),4.63(d,J=11.5Hz,1H,ArCH),4.58–4.39(m,2 H,ArCH),4.00–3.88(m,1H,4-H),3.88–3.73(m,2H,2-H,5-H),3.72–3.54(m,2H,3-H,CH2-Linke r),3.51–3.26(m,3H,CH2-Linker),1.97–1.69(m,2H,CH2-Linker),1.13(d,J=7.5Hz,3H,6-H); 13 C NMR(151MHz,Chloroform-d)δ156.70,156.22,138.24,137.87,136.81,135.19, 133.32,133.09,128.55,128.49,128.34,128.29,128.26,127.98,127.96,127. 85,127.75,127.73,127.31,126.53,126.21,126.03,125.69,98.26(C-1),77.7 0,76.25,74.94,72.39,67.25,66.69,65.74,59.69,50.85,50.55,44.63,43.81, 28.34,27.85,16.75;IR ν max (film)2920,2107,1697,1496,1454,1421,1359,1218,1169,1124,1044,965,914,857,818,749,699cm -1 HR-ESI-MS(m / z):calcd for C 42 H44 N4O6Na + (M+Na + ):723.3153,found:723.3157.

[0088] 15β:[α] 25 D =-30.8°(c 1.00,CH3Cl). 1 H NMR(400MHz,Chloroform-d)δ7.89–7.15(m,22H,Ar-H),5.21–5.10(m,2H,ArCH),4.96(d,J=11.7Hz,1H,ArCH),4.86(s,2H,ArCH),4.70(d,J=11.6Hz,1H,ArCH),4.59–4.45(m,2H,ArCH),4.17–4.02(d,J=8.0Hz,1H,1-H),3.98–3.84(m,1H,CH2-Linker),3.83–3.75(m,1H,2-H),3.54(dd,J=2.9,1.0Hz,1H,4-H),3.52–3.41(m,1H,CH2-Linker),3.41–3.27(m,4H,3-H,5-H,CH2-Linker),1.96–1.77(m,2H,CH2-Linker),1.17(d,J=6.3Hz,3H,6-H); 13 C NMR(101MHz,Chloroform-d)δ156.79,138.21,137.96,135.17,133.22,133.08,128.50,128.44,128.35,128.24,127.92,127.88,127.83,127.75,127.72,127.37,127.23,126.64,126.26,126.08,125.75,102.08(C-1),80.68,75.00,74.75,72.69,70.55,67.15,63.20,50.88,44.65,43.60,28.49,27.99,16.84.IR ν max (film)2935,2110,1697,1603,1496,1474,1454,1420,1362,1271,1218,1172,1111,1067,1028,988,893,857,818,754,698,690,664,604cm -1 ;HR-ESI-MS(m / z):calcd for C42 H 44 N4O6Na + (M+Na + ):723.3153,found:723.3156.

[0089] Compound 15a: Donor 14b (49 mg, 0.09 mmol) and linker HO(CH2)3NBnCbz (40 mg, 0.136 mmol) were azeotropically reacted with toluene, and then dissolved in dry DCM (2 mL) under argon atmosphere. The following were added to the reaction flask: Molecular sieves (flame-dried) and Ph3OP (200 mg, 0.72 mmol) were added. Then, TMSI (13 μL, 0.09 mmol, 1.0 eq) was slowly added dropwise to the reaction flask. The reaction was stirred at room temperature until TLC monitoring showed completion. The reaction was quenched with saturated Na2S2O3, and the solid was filtered off. Molecular sieves were used and washed with DCM. The organic phase was washed with water and brine and dried over anhydrous Na2SO4. The crude product was then concentrated under vacuum. The crude product was purified by silica gel column chromatography (PE / EA v / v 10:1→5:1) to give compound 15a (52 mg, 0.079 mmol, 88%, α:β = 7.5:1).

[0090] Compound 15b: Donor 14c (65 mg, 0.10 mmol) and linker HO(CH2)3NBnCbz (45 mg, 0.15 mmol) were azeotropically reacted with toluene, and then dissolved in dry DCM (2 mL) under argon atmosphere. The following were added to the reaction flask: Molecular sieves (flame-dried) and Ph3OP (222 mg, 0.8 mmol) were added. Then, TMSI (15 μL, 0.10 mmol, 1.0 eq) was slowly added dropwise to the reaction flask. The reaction was stirred at room temperature until TLC monitoring showed completion. The reaction was quenched with saturated Na2S2O3, and the solid was filtered off. Molecular sieves were used and washed with DCM. The organic phase was washed with water and brine and dried over anhydrous Na2SO4. The crude product was then concentrated under vacuum. The crude product was purified by silica gel column chromatography (PE / EA v / v 10:1→5:1) to give compound 15b (60 mg, 0.085 mmol, 85%, α:β = 13.5:1).

[0091] Compound 15c: Donor 14d (1.70 g, 2.81 mmol) and linker HO(CH2)3NBnCbz (1.26 g, 4.22 mmol) were azeotropically reacted with toluene, and then dissolved in dry DCM (28 mL) under argon atmosphere. The following were added to the reaction flask: Molecular sieves (flame-dried) and Ph3OP (6.26 g, 22.5 mmol) were used. Then, TMSI (0.42 mL, 2.81 mmol, 1.0 eq) was slowly added dropwise to the reaction flask. The reaction was stirred at room temperature until TLC monitoring showed completion. The reaction was quenched with saturated Na2S2O3, and the solid was filtered off. Molecular sieves were used and washed with DCM. The organic phase was washed with water and brine and dried over anhydrous Na₂SO₄. The mixture was then concentrated under vacuum to obtain a crude product. The crude product was purified by silica gel column chromatography (PE / EA v / v 10:1→5:1) to give compound 15c (1.83 g, 2.56 mmol, 91%, α:β = 12.1:1). [α] 25 D = +159.9° (c 1.00, CH3Cl); 1 H NMR(400MHz,Chloroform-d)δ8.21–7.08(m,22H,Ar-H),5.73–5.59(m,1H,4-H),5. 18(s,2H,ArCH),5.00–4.81(m,2H,ArCH,1-H),4.73–4.61(m,1H,ArCH),4.59–4.37 (m,2H,ArCH),4.16–3.94(m,2H,3-H,5-H),3.78–3.58(m,2H,2-H,CH2-Linker),3. 53–3.27(m,3H,CH2-Linker),1.85(s,2H,CH2-Linker),1.18(d,J=6.1Hz,3H,6-H); 13 C NMR(101MHz,Chloroform-d)δ166.14,156.65,156.23,137.77,136.61,134.65,133.2 9,133.19,133.02,131.46,129.86,129.61,128.55,128.47,128.16,128.00,127.92,1 27.87,127.60,127.33,127.25,127.01,125.95,125.86,98.19(C-1),74.18,71.40,6 9.93,67.27,65.97,65.21,59.33,50.84,50.51,44.52,43.71,28.23,27.78,16.33;IR ν max(film)2936,2108,1719,1698,1601,1496,1471,1452,1421,1358,1314,1269,1219,1169,1112,1052,1026,968,857,819,752,712,670cm -1 HR-ESI-MS(m / z):calcd for C 42 H 42 N4O7Na + (M+Na + ):737.2946,found:737.2951.

[0092] Compound 15: A solution of compound 15c (1.82 g, 2.55 mmol) dissolved in 25 mL of methanol was added to a solution containing NaOMe (70 mg, 1.28 mmol). The solution was stirred at 40 °C for 6 h. The solution was then purified using an Amberlite IR 120 (H) reagent. + The mixture was neutralized with ion exchange resin and filtered through a cotton plug. The crude product was concentrated under reduced pressure and then purified by silica gel column chromatography (PE / EA v / v 5:1→2:1) to obtain the corresponding product. The product was dissolved in DMF (13 mL), and NaH (60% dispersion in mineral oil) (205 mg, 5.10 mmol) was added. Then, BnBr (0.45 mL, 3.82 mmol) was added at 0 °C. The system was heated to room temperature and stirred thoroughly for 3 h. After the reaction was completed by TLC monitoring, the mixture was quenched with methanol and washed with water and brine, followed by DCM back-extraction. The organic phase was dried over anhydrous Na₂SO₄ and concentrated under vacuum to obtain the crude product. The crude product was purified by silica gel column chromatography (PE / EA v / v 10:1→5:1), and compound 15 (1.47 g, 2.1 mmol, 82%) was obtained through a two-step reaction. [α] 25 D = +74.6° (c 1.00, CH3Cl); 1H NMR(600MHz,Chloroform-d)δ8.05–7.05(m,22H,Ar-H),5.17(d,J=9.7Hz,2H,ArCH),4.93(d,J =11.5Hz,1H,ArCH),4.89–4.75(m,3H,ArCH,1-H),4.63(d,J=11.5Hz,1H,ArCH),4.58–4.39(m,2 H,ArCH),4.00–3.88(m,1H,4-H),3.88–3.73(m,2H,2-H,5-H),3.72–3.54(m,2H,3-H,CH2-Linke r),3.51–3.26(m,3H,CH2-Linker),1.97–1.69(m,2H,CH2-Linker),1.13(d,J=7.5Hz,3H,6-H); 13 C NMR(151MHz,Chloroform-d)δ156.70,156.22,138.24,137.87,136.81,135.19,133.3 2,133.09,128.55,128.49,128.34,128.29,128.26,127.98,127.96,127.85,127.75, 127.73,127.31,126.53,126.21,126.03,125.69,98.26(C-1),77.70,76.25,74.94,7 2.39,67.25,66.69,65.74,59.69,50.85,50.55,44.63,43.81,28.34,27.85,16.75;IR ν max (film)2920,2107,1697,1496,1454,1421,1359,1218,1169,1124,1044,965,914,857,818,749,699cm -1 HR-ESI-MS(m / z):calcd for C 42 H 44 N4O6Na + (M+Na + ):723.3153,found:723.3157.

[0093] Example 4

[0094] Crafting Sugar Block 19:

[0095] As shown in Figure 6, using p-toluenethio-β-D-glucose 17 as a starting material, the C3-position azide group was reduced with lithium aluminum hydride to obtain the corresponding amino sugar. Then, the C3-position amino group was selectively protected with a trichloroethoxycarbonyl (Troc) group to obtain compound 18. Subsequently, the C2-position hydroxyl group in compound 18 was protected with an acetylpropionyl group to obtain compound 19.

[0096] Specific experimental procedures and steps:

[0097] Compound 18: Compound 17 (1.6 g, 4.01 mmol) was placed in a 20 mL THF solution at 0 °C, and LiAlH4 (460 mg, 12.03 mmol) was added. The reaction mixture was stirred at room temperature for 1.5 h until TLC showed complete conversion of the starting material. Then, the reaction was quenched dropwise with H2O at 0 °C and diluted with DCM. The reaction mixture was washed with H2O, saturated NaHCO3 solution, and brine. The organic phase was dried over anhydrous Na2SO4, filtered, and concentrated under vacuum. The crude product was purified by silica gel column chromatography (DCM / MeOH v / v 50:1 → 20:1) to obtain amino sugars. The amino sugars were dissolved in 35 mL THF and pyridine (0.97 mL, 12.03 mmol, 3 eq). Then, a mixture of 5 mL THF containing TrocCl (0.55 mL, 4.01 mmol) was added dropwise at 0 °C. The reaction was stirred at 0°C for 2 h until TLC showed complete conversion of the starting material. The reaction was then quenched with methanol at 0°C. The reaction solution was concentrated under vacuum to obtain a crude product, which was purified by silica gel column chromatography (PE / EA v / v 5:1→2:1) to give a colorless, syrupy product 18 (1.86 g, 3.41 mmol, 85%). [α] 25 D = +57.3° (c 1.00, CH3Cl); 1 H NMR(400MHz,Chloroform-d)δ7.51–7.10(m,9H,Ar-H),5.48(s,1H,ArCH),5.35–5.14(m,1H,NH),4.72(s,2H,CH2-Troc),4.61(d,J=9.4Hz,1H,1-H),4.37(d d,J=10.6,4.5Hz,1H,6'-H),3.85–3.79(m,1H,3-H),3.75(t,J=10.2Hz,1H,6- H),3.63–3.49(m,3H,2-H,4-H,5-H),3.38(br,1H,2-OH),2.36(s,3H,ArCH3); 13C NMR(101MHz,Chloroform-d)δ155.26,138.84,136.69,133.71,129.89,129.28,128.3 2,127.42,126.15,101.75,95.27,89.82,74.69,71.81,71.53,68.58,58.35,21.20;IR ν max (film)3328,2881,1715,1542,1493,1451,1372,1318,1245,1168,1109,1071,1031,1006,810,749,733,698cm -1 HR-ESI-MS(m / z):calcd for C 23 H 24 Cl3NO6SNa + (M+Na + ):570.0282,found:570.0279.

[0098] Compound 19: Compound 18 (1.84 g, 3.36 mmol) was placed in a dry DCM (33 mL) and stirred. Levylpropionic acid (LevOH) (585 mg, 5.04 mmol), N,N-dicyclohexylcarboimide (DCC) (1.04 g, 5.04 mmol), and DMAP (615 mg, 5.04 mmol) were added. The reaction mixture was stirred at room temperature for 4 h. The reaction mixture was diluted with DCM until the starting material was completely consumed. The organic phase was washed with saturated NaHCO3 solution and brine, dried over anhydrous Na2SO4, and filtered. The filtrate was concentrated under vacuum to obtain the crude product. Purification was achieved by silica gel column chromatography (PE / EA v / v 10:1→4:1) to give compound 19 (2.06 g, 3.19 mmol, 95%). [α] 25 D = +41.3° (c 1.00, CH3Cl); 1H NMR(600MHz,Chloroform-d)δ7.46–7.10(m,9H,Ar-H),5.47(s,1H,ArCH),5.12(d,J=9.0Hz,1H,NH),4.90(t,J =9.8Hz,1H,2-H),4.76(d,J=12.1Hz,1H,CH2-Troc),4.73(d,J=9.8Hz,1H,1-H),4.68(d,J=12.0Hz,1H,CH2-Tro c),4.37(dd,J=10.7,4.4Hz,1H,6'-H),4.02(q,J=9.4Hz,1H,3-H),3.75(t,J=10.0Hz,1H,6-H),3.62–3.48(m,2 H,4-H,5-H),2.83–2.73(m,2H,CH2-Lev),2.69–2.60(m,2H,CH2-Lev),2.35(s,3H,CH3CO),2.19(s,3H,ArCH3); 13 C NMR(151MHz,Chloroform-d)δ205.95,171.82,154.35,138.83,136.74,133.87,129.77,129.14,128.23,127 .59,126.13,101.56,95.56,87.07,77.92,74.49,72.00,70.54,68.48,56.75,37.89,29.79,28.01,21.20; IR ν max (film)3326,2880,1742,1721,1551,1375,1318,1262,1245,1156,1103,1069,1030,811,751,734,697cm -1 HR-ESI-MS(m / z):calcd for C 28 H 30 Cl3NO8SNa + (M+Na + ):668.0650,found:668.0652.

[0099] Example 5

[0100] Synthesis of trisaccharide intermediate compound 22:

[0101] As shown in Figure 7, glycosylation was performed using glycosyl donor 9 and acceptor 16 as raw materials under the action of the activating reagent TMSOTf to obtain disaccharide compound 20. Subsequently, the TBS at the C4 position of the disaccharide was removed using tetrabutylamine fluoride (TBAF) to obtain disaccharide acceptor 21. Finally, monosaccharide donor 19 and disaccharide acceptor 21 were glycosylated under the catalysis of TMSOTf and NIS to obtain trisaccharide intermediate compound 22.

[0102] Specific experimental procedures and steps:

[0103] Compound 20: At -10°C, donor 9 (1.43 g, 2.06 mmol), acceptor 16 (0.77 g, 1.37 mmol) and... The molecular sieve was dissolved in dry DCM (20 mL). TMSOTf (75 μL, 0.412 mmol) was added dropwise under argon protection. When TLC showed complete conversion of the starting material, the mixture was quenched with Et3N, then filtered and concentrated under vacuum. The crude product was purified by silica gel column chromatography (PE / EA v / v 5:1→2:1) to give 20 g of colorless syrup (1.71 g, 1.61 mmol, 78%). [α] 25 D = +24.5° (c 1.00, CH3Cl); 1H NMR(400MHz,Chloroform-d)δ7.41–7.13(m,20H,Ar-H),5.21–5.12(m,4H,ArCH),5.03–4.97(m,1H,2’-H),4.83(d,J=11.4Hz,1H,ArCH),4.81–4.73(m,1H,1-H),4.72–4.66(m,1H,1’-H),4.54(d,J=15.5Hz,1H,ArCH),4.51–4.44(m,1H,ArCH),4.40(d,J=11.4Hz,1H,ArCH),3.99–3.88(m,2H,5’-H,3-H),3.84(td,J=9.2,1.3Hz,1H,4’-H),3.78–3.63(m,1H,5-H),3.65(dd,J=10.6,3.7Hz,1H,2-H),3.57–3.52(m,1H,4-H),3.53–3.46(m,1H,3’-H),3.46–3.36(m,1H,CH2-Linker),3.38–3.26(m,1H,CH2-Linker),2.89–2.67(m,4H,CH2-Lev),2.16(s,3H,CH3CO),1.93–1.71(m,2H,CH2-Linker),1.06–0.96(m,3H,6-H),0.86(s,9H,SiC(CH3)3),0.18(s,3H,SiCH3),0.03(s,3H,SiCH3); 13 C NMR(151MHz,Chloroform-d)δ205.95,171.33,167.26,156.65,156.26,138.49,137.87,136.77,134.83,102.48(C-1’),98.26(C-1),78.96,78.50,77.10,75.11,71.77,70.89,67.75,67.39,67.27,66.46,65.80,59.50,50.89,50.62,44.66,43.89,37.89,31.94,29.71,29.69,29.69,29.68,27.75,25.70,24.76,22.69,18.02,16.34,14.10,-4.45,-5.01;IR ν max (film)2917,2850,2107,1750,1697,1455,1418,1362,1255,1217,1134,1041,838,780,752,698cm -1HR-ESI-MS(m / z):calcd for C 55 H 69 N7O 13 SiNa + (M+Na + ):1086.4615,found:1086.4619.

[0104] Compound 21: Compound 20 (1.70 g, 1.60 mmol) was dissolved in THF (16 mL) at room temperature, followed by the addition of AcOH (91 μL, 1.60 mmol) and TBAF (2.4 mL, 2.4 mmol), and stirred for 5 h. The organic phase was dried over Na₂SO₄ and concentrated under vacuum. The crude product was purified by silica gel column chromatography (PE / EA v / v 4:1→3:2) to obtain colorless syrup 21 (1.40 g, 1.47 mmol, 92%). [α] 25 D = +21.6° (c 1.00, CH3Cl). 1 H NMR(600MHz,Chloroform-d)δ7.40–7.13(m,20H,Ar-H),5.26–5.11(m,4H,ArCH),4.87(dd,J=10.3,7.9Hz,1H,2'-H),4.80(d,J=11.4Hz,1H,A rCH),4.79–4.72(m,1H,1-H),4.72–4.65(m,1H,1'-H),4.57–4.42(m,2H,ArCH),4.38(d,J=11.4Hz,1H,ArCH),4.01–3.88(m,2H,5'-H,3-H),3 .85(t,J=9.5Hz,1H,4'-H),3.77–3.64(m,1H,5-H),3.69–3.63(m,2H,2-H,4-H),3.65–3.54(m,1H,CH2-Linker),3.59(t,J=9.9Hz,1H,3'-H), 3.51–3.25(m,3H,CH2-Linker),2.87–2.64(m,4H,CH2-Lev),2.17(s,3H,CH3CO),1.91–1.71(m,2H,CH2-Linker),0.99(d,J=6.3Hz,3H,6-H); 13C NMR(151MHz,Chloroform-d)δ206.05,171.31,156.23,138.53,137.83,134.45,12 8.97,128.82,128.58,128.50,128.17,128.02,127.82,127.57,127.38,127.22,1 02.61(C-1”),98.24(C-1),79.06,75.14,74.35,70.90,70.81,67.96,67.25,66.4 7,65.80,65.59,59.30,50.85,50.56,44.67,43.84,37.91,29.75,27.75,16.38;IR ν max (film)3445,2919,2108,1750,1697,1454,1422,1362,1314,1218,1161,1042,914,752,698cm -1 HR-ESI-MS(m / z):calcd for C 49 H 55 N7O 13 Na + (M+Na + ):972.3750,found:972.3756.

[0105] Compound 22: At -10°C, glucosinolate donor 19 (0.56 g, 0.87 mmol), acceptor 21 (0.55 g, 0.58 mmol) and... Molecular sieves were placed in dry DCM (6 mL) and stirred. NIS (235 mg, 1.04 mmol) and TMSOTf (30 μL, 0.174 mmol) were added. The reaction was stirred at 0 °C for 5 h until TLC showed complete conversion of the glycosyl donor. The reaction solution was diluted with DCM and filtered. The filtrate was washed with 10% (w / v) Na₂S₂O₃, saturated NaHCO₃ solution, and brine. The organic phase was dried over anhydrous Na₂SO₄ and filtered. The filtrate was concentrated under vacuum. The crude product was purified by silica gel column chromatography (PE / EA v / v 5:1→2:1) to obtain colorless syrup 22 (648 mg, 0.44 mmol, 76%). [α] 25 D = +10.2° (c 1.00, CH3Cl); 1H NMR(600MHz,Chloroform-d)δ7.46–7.16(m,25H,Ar-H),5.47(s,1H,ArCH),5.41(d,J=11.8Hz,1H,ArCH),5.21–5.14(m,2H,ArCH),5.09(d,J=11.8Hz,1H,ArCH),4.98(d,J=9.2Hz,1H,NH),4.87–4.81(m,2H,2’-H,2”-H),4.82(d,J=11.4Hz,1H,ArCH),4.80–4.74(m,1H,1-H),4.78(d,J=12.3Hz,1H,CH2-Troc),4.74–4.67(m,2H,1’-H,CH2-Troc),4.54(d,J=15.5Hz,1H,ArCH),4.52–4.47(m,1H,ArCH),4.46(d,J=8.1Hz,1H,1”-H),4.43(d,J=11.4Hz,1H,ArCH),4.34(dd,J=10.7,5.0Hz,1H,6”-H),4.10–4.03(m,2H,5’-H,6’-H),4.00–3.91(m,1H,3-H),3.88(q,J=10.3Hz,1H,3”-H),3.77-3.64(m,1H,5-H),3.72(t,J=10.4Hz,1H,6”-H),3.65(dd,J=10.8,3.5Hz,1H,4-H),3.63–3.51(m,4H,3’-H,CH2-Linker,2-H,4”-H),3.50–3.29(m,3H,CH2-Linker),3.18(td,J=9.6,5.0Hz,1H,5”-H),2.86–2.49(m, 8H,CH2-Lev),2.18(s,3H,CH3CO),2.17(s,3H,CH3CO)1.92–1.71(m,2H,CH2-Linker),1.05–0.95(m,3H,6-H); 13C NMR(151MHz,Chloroform-d)δ206.08,206.02,172.10,171.31,166.67,154.37,138.40,137.83,136.75,134.57,129.34,129. 17,129.02,128.66,128.65,128.57,128.51,128.26,128.22,128.01,127.83,127.67,127.38,127.24,126.13,102.51(C-1') ,101.60(ArCH),100.52(C-1”),98.23(C-1),95.54(CCl3-Troc),78.81,78.03,76.23,74.98,74.48,71.83,70.46,68.39,67. 82,67.59,67.25,66.38,65.79,64.61,59.40,54.95,50.56,44.61,37.91,37.79,29.83,29.75,27.84,27.73,27.63,16.37; IR ν max (film)3323,2924,2111,1719,1543,1455,1420,1364,1314,1274,1239,1152,1081,1029,917,820,752,699cm -1 HR-ESI-MS(m / z):calcd for C 70 H 77 Cl3N8O 21 Na + (M+Na + ):1493.4161,found:1493.4168.

[0106] Example 6

[0107] Synthesis of target trisaccharide 30:

[0108] As shown in Figure 8, using trisaccharide intermediate compound 22 as a starting material, two acetylpropionyl groups in compound 22 were simultaneously removed under the action of hydrazine acetate to obtain diol 23. Subsequently, trifluoro anhydride (Tf2O) and pyridine were used to convert the two hydroxyl groups in compound 23 into trifluoromethanesulfonyl groups, and then tetrabutylammonium azide (TBAN3) was used for axial substitution to obtain compound 24. Then, after hydrolyzing the 4,6-O-benzyl group in trifluoroacetic acid (TFA)-mediated reaction, the C6” hydroxyl group of the resulting diol 25 was selectively oxidized to generate the corresponding carboxylic acid. This was then selectively benzyl esterified using benzyl bromide and sodium bicarbonate (NaHCO3) to give compound 26. The azide group was reduced in pyridine using thioacetic acid (AcSH) to give acetaminosaccharide 27. Subsequently, the Troc group on trisaccharide 27 was removed in acetic acid at 55°C using excess zinc powder. Then, it was reacted in pyridine using S-benzylthioacetylimine hydrochloride to give trisaccharide 28 containing an acetamidine group at the C3” position. Due to the increased polarity of trisaccharide 26, it was difficult to dissolve in dichloromethane but readily soluble in methanol. It was found that during dissolution, transfer, and storage in methanol, due to the effect of the basic acetamidine group, part of product 28 underwent an transesterification reaction from benzyl carboxylate to methyl carboxylate, yielding trisaccharide 29. Finally, the methyl ester on trisaccharide 29 was hydrolyzed at 0°C using lithium hydroxide, followed by complete deprotection using a Pd / C hydrogenation reaction to obtain the target trisaccharide 30.

[0109] Specific experimental procedures and steps:

[0110] Compound 23: Compound 22 (620 mg, 0.422 mmol) was placed in a DCM / MeOH solution (v / v 20:1, 4.2 mL) and stirred. Hydrazine acetate (117 mg, 1.27 mmol) was added. The reaction mixture was stirred at room temperature for 4.5 h until TLC showed complete conversion of the starting material. The reaction mixture was diluted with DCM and washed with saturated NaHCO3 solution and brine. The organic phase was dried over anhydrous Na2SO4 and concentrated under vacuum. The crude product was purified by silica gel column chromatography (PE / EA v / v 5:1→2:1) to give colorless syrup product 23 (457 mg, 0.359 mmol, 85%). [α] 25 D = +35.8° (c 1.00, CH3Cl); 1H NMR(600MHz,Chloroform-d)δ7.49–7.14(m,25H,Ar-H),5.48(s,1H,ArCH),5.29(d,J=11.9Hz,1H,ArCH),5.23(s,1H,NH),5.19–5.11(m,3H,ArCH),4.86–4.78(m,1H,1-H),4.77(d,J=11.7Hz,1H,ArCH),4.77–4.72(m,2H,CH2-Troc),4.58–4.49(m,1H,1’-H),4.50(d,J=7.4Hz,1H,1”-H),4.43(d,J=11.6Hz,1H,ArCH),4.40(dd,J=10.7,5.0Hz,1H,6”-H),4.08–4.02(m,1H,5’-H),4.03–3.94(m,1H,3-H),3.91(t,J=9.5Hz,1H,4’-H),3.80–3.65(m,3H,6”-H,2-H,5-H),3.72(dd,J=10.1,7.6Hz,1H,3”-H),3.65–3.56(m,1H,CH2-Linker),3.65–3.60(m,2H,4”-H,4-H),3.59–3.51(m,2H,2”-H,3’-H),3.43–3.39(m,1H,5”-H),3.48–3.28(m,3H,CH2-Linker),3.38(dd,J=9.9,7.7Hz,1H,2’-H),2.90(s,1H,2’-OH),1.91–1.70(m,2H,CH2-Linker),1.04–0.94(m,3H,6-H); 13C NMR(151MHz,Chloroform-d)δ167.89,155.05,138.35,137.79,136.82,134.56,129.24,129.08,128.85,128.59,128.54,128.51,128.40,128.31,128.24,128.03,127.85,127.74,127.41,127.25,126.17,105.07(C-1’),104.34(C-1”),101.78(ArCH),97.87(C-1),95.39(CCl3-Troc),80.27,75.02,74.67,74.46,73.11,72.47,68.50,68.16,67.86,67.28,66.88,66.36,65.89,59.35,56.90,27.82,27.23,16.35.IR ν max (film)3390,2925,2112,1743,1683,1537,1497,1454,1423,1363,1239,1220,1176,1082,1029,916,819,752,698cm -1 ;HR-ESI-MS(m / z):calcd for C 60 H 65 Cl3N8O 17 Na + (M+Na + ):1297.3425,found:1297.3431.

[0111] Compound 24: Under nitrogen protection, compound 23 (440 mg, 0.345 mmol) was placed in a mixed solution of dry DCM (6.0 mL) and pyridine (0.56 mL, 6.9 mmol, 20 eq) and stirred. The reaction system was cooled to -20 °C, and a mixture of dry DCM (1 mL) containing Tf₂O (0.35 mL, 2.07 mmol) was added dropwise. The reaction mixture was stirred and heated from -20 °C to 0 °C over 4 h. The organic phase was dried over anhydrous sodium sulfate and concentrated under vacuum at 30 °C to give a yellow syrupy trichlorate product. The above trichlorate was dissolved in anhydrous DMF (7 mL) under nitrogen. TBAN₃ (0.98 g, 3.45 mmol) was added at 0 °C. The reaction was stirred overnight at room temperature. The reaction progress was monitored by TLC. The reaction solution was diluted with ethyl acetate (20 mL), washed with water and brine, and the organic phase was dried over anhydrous Na₂SO₄ and concentrated under vacuum. The crude product was purified by silica gel column chromatography (PE / EA v / v 2:1). Product 24 (334 mg, 0.252 mmol, 73%) was obtained through a two-step reaction. [α] 25 D = -8.4° (c = 1.00, CHCl3); 1H NMR(600MHz,Chloroform-d)δ7.60–7.05(m,25H,Ar-H),5.46(s,1H,ArCH),5.29(d,J=11.6Hz,1H,ArCH),5.25(d,J=8.5Hz,1H,NH),5.22–5.12(m,3H,ArCH),4.99(d,J=11.4Hz,1H,ArCH),4.92–4.81(m,1H,1’-H),4.81–4.72(m,3H,CH2-Troc,1-H),4.66(s,1H,1”-H),4.59–4.43(m,3H,ArCH),4.35(dd,J=11.1,4.9Hz,1H,6”-H),4.22(t,J=9.6Hz,1H,4’-H),4.14–4.08(m,1H,5’-H),4.08–3.97(m,1H,3-H),3.96–3.85(m,3H,3”-H,2-H,2’-H),3.83(s,1H,2”-H),3.78(t,J=10.4Hz,1H,6”-H),3.75–3.68(m,1H,5-H),3.68–3.53(m,2H,4-H,CH2-Linker),3.56(t,J=9.7Hz,1H,4”-H),3.52–3.26(m,4H,CH2-Linker,3’-H),3.22(td,J=9.9,4.9Hz,1H,5”-H),1.90–1.70(m,2H,CH2-Linker),1.01–0.92(m,3H,6-H); 13C NMR(151MHz,Chloroform-d)δ167.18,154.03,138.34,137.84,136.85,134.67,129.40,129.28,129.18,129.04,128.75,12 8.70,128.59,128.52,128.27,128.24,128.13,128.02,127.89,127.81,127.75,127.39,127.20,126.14,102.01(C-1'),101 .95(C-1”),101.40(ArCH),97.52(C-1),95.38(CCl3-Troc),80.80,78.25,75.77,75.02,74.66,74.57,74.25,68.36,68.17 ,68.01,67.23,66.41,65.75,63.57,62.68,62.23,60.40,59.43,52.08,50.77,50.52,44.57,43.72,28.20,27.83,16.39; IR ν max (film)2920,2108,1743,1693,1455,1275,1234,1178,1090,1045,1000,905,822,753,699cm -1 HR-ESI-MS(m / z):calcd for C 60 H 63 Cl3N 14 O 15 Na + (M+Na + ):1347.3555,found:1347.3559.

[0112] Compound 25: Compound 24 (240 mg, 0.181 mmol) was placed in DCM (4.0 mL) solution and stirred. Trifluoroacetic acid (0.5 mL, 6.72 mmol) and H2O (33 μL, 1.81 mmol, 10 eq) were added. The reaction was stirred at room temperature for 4 h until TLC showed complete conversion of the starting material. The reaction solution was diluted with DCM, washed with water, saturated NaHCO3 solution, and brine. The organic phase was dried over anhydrous Na2SO4 and concentrated under vacuum. The crude product was purified by silica gel column chromatography (DCM / MeOH v / v 50:1→20:1) to give colorless syrup 25 (199 mg, 0.161 mmol, 89%). [α] 25 D = -5.5° (c 1.00, CH3Cl); 1H NMR(600MHz,Chloroform-d)δ7.42–7.14(m,20H,Ar-H),5.53(d,J=7.8Hz,1H,NH),5.28–5.11(m,4H,ArCH),4.99(d,J=11.5Hz,1H,ArCH),4.93–4.79(m,1H,1’-H),4.82(d,J=11.9Hz,1H,CH2-Troc),4.82–4.70(m,1H,1-H),4.77(d,J=12.0Hz,1H,CH2-Troc),4.62(s,1H,1”-H),4.56(d,J=11.5Hz,1H,ArCH),4.55–4.42(m,2H,ArCH),4.21(t,J=9.6Hz,1H,4’-H),4.09(d,J=8.8Hz,1H,5’-H),4.07–3.97(m,1H,3-H),3.97–3.86(m,1H,2-H),3.93–3.86(m,1H,2’-H),3.92(dd,J=12.2,3.1Hz,1H,6”-H),3.83(dd,J=12.3,3.9Hz,1H,6”-H),3.79–3.66(m,1H,5-H),3.66–3.55(m,5H,4”-H,4-H,CH2-Linker,2”-H,3”-H),3.53–3.26(m,4H,3’-H,CH2-Linker),3.15(dt,J=8.1,3.5Hz,1H,5”-H),2.84(br,1H,4”-OH),2.28(br,1H,6”-OH),1.93–1.70(m,2H,CH2-Linker),1.02–0.91(m,3H,6-H); 13CNMR(151MHz,Chloroform-d)δ167.20,156.61,156.25,155.16,138.28,137.79,136.57,134.67,129.34,129.25,128.98,128.78,128.59,128.50,128.23,128.02,127.81,127.75,127.42,127.37,127.17,101.86(C-1’),101.25(C-1”),97.56(C-1),95.24(CCl3-Troc),80.71,78.19,75.02,74.92,74.55,74.47,67.97,67.24,66.46,66.38,65.73,63.32,62.42,62.06,61.88,59.40,54.55,50.74,50.49,44.54,43.74,28.16,27.75,16.37.IR ν max (film)3411,2926,2109,1738,1697,1516,1455,1423,1362,1273,1168,1076,1045,820,737,699cm -1 ;HR-ESI-MS(m / z):calcd for C 53 H 59 Cl3N 14 O 15 Na + (M+Na + ):1259.3242,found:1259.3247.

[0113] Compound 26: Compound 25 (190 mg, 0.154 mmol) was dissolved in DCM (4 mL) and H2O (1 mL) solution under argon protection. The system temperature was lowered to 0 °C using an ice bath, and TEMPO (5 mg, 0.031 mmol) and BAIB (99 gm, 0.308 mmol) were added. The mixture was stirred at room temperature for 4 h. The reaction progress was monitored by TLC until the starting material was completely consumed. The reaction was quenched dropwise with 10% (w / v) Na2S2O3 solution, and the organic phase was washed with saturated NaHCO3 and saturated NaCl solutions, followed by DCM back-extraction. The organic phase was then dried with anhydrous Na2SO4 and concentrated under vacuum. The crude product was dissolved in anhydrous DMF (3 mL) under nitrogen protection. NaHCO3 (65 mg, 0.77 mmol) and BnBr (55 μL, 0.462 mmol) were added at room temperature. When TLC showed that the starting material was completely consumed, the solvent was removed by vacuum concentration. Purified by silica gel column chromatography (PE / EA v / v 2:1). The two-step reaction yielded syrup 26 (2.75 g, 6.16 mmol, 80%). [α] 25 D = -12.5° (c 1.00, CH3Cl); 1H NMR(600MHz,Chloroform-d)δ7.43–7.13(m,25H,Ar-H),5.36(d,J=8.4Hz,1H,NH),5.27(d,J=12.2Hz,1H,ArCH),5.26–5.22(m,1H,ArCH),5.21(d,J=12.2Hz,1H,ArCH),5.19–5.12(m,3H,ArCH),4.98(d,J=11.5Hz,1H,ArCH),4.90–4.77(m,1H,1’-H),4.84(d,J=12.0Hz,1H,CH2-Troc),4.80–4.72(m,1H,1-H),4.72(d,J=12.0Hz,1H,CH2-Troc),4.64(s,1H,1”-H),4.56(d,J=11.5Hz,1H,ArCH),4.56–4.43(m,2H,ArCH),4.22(t,J=9.6Hz,1H,4’-H),4.11–3.96(m,2H,5’-H,3-H),3.95–3.83(m,2H,2-H,2’-H),3.77–3.61(m,6H,4”-H,5-H,4-H,2”-H,3”-H,5”-H),3.63–3.53(m,1H,CH2-Linker),3.53–3.27(m,4H,3’-H, CH2-Linker),3.17(br,1H,4”-OH),1.89–1.70(m,2H,CH2-Linker),0.96(t,J=8.5Hz,3H,6”-H); 13C NMR(151MHz,Chloroform-d)δ168.67,167.18,156.60,156.22,154.55,138.32,137.80,136.83,136.59,134.71,129 .33,129.23,128.97,128.78,128.63,128.58,128.49,128.43,128.21,128.01,127.80,127.72,127.38,127.16,101 .73(C-1'),101.39(C-1”),97.56(C-1),95.24(CCl3-Troc),80.65,78.12,75.34,74.96,74.89,74.87,74.48,67.95 ,67.79,67.22,66.37,65.71,63.35,62.62,61.66,59.39,53.75,50.77,50.49,44.54,43.72,28.17,27.74,16.38.IR ν max (film)3367,2925,2110,1743,1697,1517,1455,1362,1273,1176,1127,1075,1044,1001,823,753,698cm -1 HR-ESI-MS(m / z):calcd for C 60 H 63 Cl3N 14 O 16 Na + (M+Na + ):1363.3504,found:1259.3501.

[0114] Compound 27: Compound 26 (56 mg, 0.042 mmol) was placed in a pyridine (2.1 mL) solution and stirred under nitrogen protection. The reaction system was cooled to 0 °C, and AcSH (2.1 mL) was added. The reaction was stirred at room temperature until TLC showed complete conversion of the starting material. The reaction solution was diluted with DCM, washed with water, saturated NaHCO3 solution, and brine, and the organic phase was dried over anhydrous Na2SO4 and concentrated under vacuum. The crude product was purified by silica gel column chromatography (DCM / MeOH v / v 50:1→20:1) to give colorless syrup product 27 (48 mg, 0.034 mmol, 81%). [α] 25 D = +16.2° (c 1.00, CH3Cl); 1H NMR(600MHz,Chloroform-d)δ7.91–7.74(m,1H,NH),7.45–7.16(m,25H,Ar-H),6.96–6.86(m,1H,NH),6.86–6.79(m,1H,NH),6.80–6.72(m,1H,NH),6.08–5.86(m,1H,NH),5.29(d,J=11.9Hz,1H,ArCH),5.23–5.04(m,5H,ArCH),4.91–4.83(m,2H,1’-H,ArCH),4.80(d,J=12.0Hz,1H,CH2-Troc),4.72(d,J=12.1Hz,1H,CH2-Troc),4.71–4.66(m,1H,3’-H),4.65–4.59(m,2H,1-H,ArCH),4.54(d,J=11.8Hz,1H,ArCH),4.38(d,J=15.9Hz,1H,ArCH),4.31–4.19(m,3H,3”-H,-H,1”-H),4.09(d,J=9.8Hz,1H,5”-H),3.98(q,J=10.6,9.5Hz,1H,4”-H),3.90–3.82(m,2H,4’-H,5-H),3.79–3.71(m,3H,3-H,4-H,CH2-Linker),3.68(s,1H,2’-H),3.65–3.61(m,1H,CH2-Linker),3.55(d,J=9.5Hz,1H,5’-H),3.33–3.23(m,2H,CH2-Linker,2-H),3.23–3.15(m,1H,CH2-Linker),2.13(s,3H,CH3CO),2.02(s,3H,CH3CO),1.89(s,3H,CH3CO),1.78(s,3H,CH3CO),1.08(d,J=6.4Hz,3H,6-H); 13C NMR(151MHz,Chloroform-d)δ175.35,172.52,172.22,171.43,168.43,167.65,157.71,156.53,138.68,137.33,136.4 0,135.02,134.75,129.51,129.38,129.00,128.71,128.65,128.55,128.47,128.24,128.15,127.83,127.70,127.61, 127.22,101.44(C-1'),98.02(C-1”),97.82(C-1),95.26(CCl3-Troc),80.59,79.40,74.96,74.21,69.58,67.79,67.4 8,67.33,66.76,63.43,57.96,52.78,51.84,51.22,49.65,48.50,42.66,27.20,23.50,23.22,23.15,22.78,16.64; IRν max (film)3306,2926,1743,1668,1537,1455,1423,1373,1240,1180,1093,1050,819,736,698cm -1 HR-ESI-MS(m / z):calcd for C 68 H 79 Cl3N6O 20 Na + (M+Na + ):1427.4307,found:1427.4311.

[0115] Compound 28: Compound 27 (30 mg, 21.4 μL) was placed in acetic acid (2.0 mL) solution and stirred, and activated zinc powder (50 mg, 0.78 mmol) was added. After stirring at room temperature for 12 h, the reaction mixture was filtered through diatomaceous earth, and the filtrate was concentrated under vacuum. The crude product was purified by silica gel column chromatography (DCM / MeOH v / v 20:1→10:1) to obtain the corresponding amino sugar. The above amino sugar was placed in anhydrous pyridine (2 mL) solution and stirred at 0 °C, and benzylthioacetylimine hydrochloride (8.6 mg, 42.8 μmol) was added. The reaction was stirred at 0 °C for 5 h. The reaction solution was then concentrated under vacuum, and the crude product was purified by silica gel column chromatography (DCM / MeOH v / v 10:1). The two-step reaction yielded colorless syrup product 28 (19 mg, 15.6 μmol, 73%). [α] 25 D=-2.7°(c 1.00,CH3Cl); 1 H NMR(600MHz,Methanol-d4)δ7.43–7.17(m,25H,Ar-H),5.37(d,J=12.3Hz,1H,ArCH),5.26(s,2H,ArCH),5.19–5.10(m,3H,ArCH),5.08(s,1H,1’-H),4.99–4.90(m,1H,1-H),4.88(d,J=11.4Hz,1H,ArCH),4.65–4.58(m,1H,1”-H),4.57–4.53(m,1H,2-H),4.53–4.50(m,2H,),4.47–4.45(m,1H,2’-H),4.45–4.39(m,1H,2”-H),4.35(d,J=11.3Hz,1H,ArCH),4.26–4.15(m,2H,3-H,5-H),4.13–4.06(m,2H,5’-H,4-H),4.01–3.89(m,2H,3”-H,3’-H),3.83–3.76(m,2H,5-H,4’-H),3.66–3.59(m,1H,4”-H),3.59–3.48(m,1H,CH2-Linker),3.46–3.33(m,3H,CH2-Linker),2.20(s,3H,CH3CO),2.02(s,3H,CH3CO),1.95–1.90(m,3H,CH3CO),1.85(s,3H,CH3CO),1.81(s,3H,CH3CO),1.05–0.95(m,3H,6-H); 13 C NMR(151MHz,Methanol-d4)δ175.03,174.27,173.88,173.71,170.65,169.77,167.49,140.34,139.29,138.10,136.71,136.56,129.77,129.66,129.21,129.17,128.93,128.53,128.33,127.99,103.49(C-1),100.69(C-1’),99.03(C”-1),80.53,78.19,76.46,76.14,68.81,68.47,67.73,67.58,66.05,58.19,53.03,52.26,51.35,50.21,45.66,45.49,29.43,28.80,23.04,22.87,22.78,19.24,16.97;IR ν max(film)3291,2931,1743,1657,1550,1454,1374,1293,1244,1177,1112,1051,739,699,599cm -1 HR-ESI-MS(m / z):calcd for C 67 H 82 N7O 18 + (M+H + ):1272.5711,found:1272.5714.

[0116] Compound 30: Compound 29 (11 mg, 9.20 μmol) was placed in a 1.0 mL THF solution at 0 °C, and 20 μL of 1 M LiOH aqueous solution was added. After complete conversion of the starting material by TLC, the mixture was diluted with DCM and washed with water and brine. After filtration, the filtrate was concentrated under reduced pressure to obtain the crude product. The crude product was purified by silica gel column chromatography (DCM / MeOH v / v 10:1) to obtain a partially deprotected product. The partially protected product was placed in a THF / MeOH / H2O / AcOH solution (v / v / v / v 10:5:4:1, 2 mL), and 10% Pd / C (50 mg) was added. The reaction was stirred in H2 for 24 h. The mixture was then filtered and concentrated under vacuum, and eluted using a Sephadex LH-20 column (H2O). The two-step reaction yielded a white solid 30 (4.5 mg, 5.88 μmol, 64%). 1H NMR(600MHz,Deuterium Oxide)δ4.98(s,1H,1'-H),4.84(s,1H,1-H),4.77(s,1H,1”-H),4.51(d, J=3.3Hz,1H,2”-H),4.41–4.38(m,1H,2’-H),4.24(dd,J=11.7,3.4Hz,1H, 2-H),4.16–4.12(m,1H,3'-H),4.05(q,J=6.4Hz,1H,5-H),4.03–3.98(m, 3H,3-H,4-H,5"-H),3.97–3.92(m,2H,5'-H,3"-H),3.85–3.81(m,2H,4'-H ,4”-H),3.78(dt,J=11.3,6.4Hz,1H,CH2-Linker),3.54(dt,J=11.3,6.1Hz,1H,CH2-Linker),3.11(t,J=7.5Hz,2H,CH2-Linker),2.20(s,3H,CH3C (N)),2.05(s,3H,CH3CO),2.05(s,3H,CH3CO),2.02–1.97(m,2H,CH2-Linker),1.99(s,3H,CH3CO),1.97(s,3H,CH3CO),1.23(d,J=6.5Hz,3H,6-H); 13 C NMR (151MHz, Deuterium Oxide)δ174.82,174.26,174.20,173.93,166.06,163.13,162.89,99.90(C-1'),98.56(C-1”),97.23(C-1),78.23,77.39,76.93,75.13,70. 00,66.54,65.70,64.94,56.20,51.95,51.35,49.49,47.75,37.14,26.74,21.92,21.91,21.80,21.75,18.53,15.35; HR-ESI-MS(m / z):calcd for C 31 H 52 N7O 16 + (M+H + ):778.3465,found:778.3466.

[0117] Comparative Example 1

[0118] An attempt to synthesize trisaccharide 35:

[0119] As shown in Figure 9, intermediate compound 7 was used as a starting material. The C4 hydroxyl group of compound 7 was benzyl-protected with benzyl bromide and silver oxide to obtain compound 31. Subsequently, the allyl group at the reducing end of compound 31 was removed using palladium chloride (PdCl2) to obtain the corresponding hemiacetal derivative, which was then converted into N-phenyltrifluoroimine ester donor 32. Under the catalysis of TMSOTf, compound 32 reacted with p-toluenethiophenol to obtain thioglycoside donor 33. Intermediate disaccharide 21 underwent reduction of the azide groups at the C2 and C3' positions to aminoacetyl groups in the presence of zinc powder and acetic anhydride to obtain compound 34. Subsequently, using TMSOTf as an activator, disaccharide acceptor 34 was glycosylated with trifluoroacetylimine ester donor 32 and thioglycoside donor 33, respectively, in an attempt to assemble and synthesize trisaccharide 35.

[0120] Specific experimental procedures and steps:

[0121] Compound 31: Under nitrogen protection, benzyl glucuronide 7 (1.2 g, 2.68 mmol) was dissolved in anhydrous DCM (27 mL), and BnBr (1.59 mL, 13.4 mmol) and Ag₂O (1.86 g, 8.04 mmol) were added at 0 °C. The reaction mixture was heated to 30 °C and stirred for 24 hours. After filtration through diatomaceous earth and concentration, the residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate 10:1 v / v) to give product 31 as a white solid (1.14 g, 2.12 mmol, 79%). [α] 25 D = +68.4° (c 1.00, CH3Cl); 1 H NMR(400MHz,Chloroform-d)δ7.40–7.16(m,10H,Ar-H),5.94–5.81(m,1H,All-CH),5.28(dq,J=17.2,1.6Hz, 1H,All-CH2),5.20(s,2H,ArCH),5.07(d,J=3.6Hz,1H,1-H),4.75–4.67(m,2H,2-H,ArCH),4.47(d,J=10.5Hz ,1H,ArCH),4.30(d,J=9.8Hz,1H,5-H),4.20(ddt,J=13.0,5.3,1.5Hz,1H,All-CH2),4.07–4.01(m,1H,All-C H2),4.00(t,J=10.1Hz,1H,3-H),3.66(t,J=9.7Hz,1H,4-H),2.88–2.57(m,4H,CH2-Lev),2.19(s,3H,CH3CO); 13C NMR(101MHz,Chloroform-d)δ206.06,171.85,168.82,140.90,137.13,134.96,133.01,128.66,128.59,128.51,128.44,128. 42,128.17,128.05,127.68,127.00,118.48,77.99,75.07,71.51,70.33,69.14,67.51,65.40,63.04,37.80,29.82,27.86; IR ν max (film)=2926,2108,1747,1716,1455,1362,1181,1154,1053,739,697cm -1 HR-ESI-MS(m / z):calcd for C 28 H 31 N3O8Na + (M+Na + ):560.2003,found:560.2005.

[0122] Compound 32: Compound 31 (0.64 g, 1.19 mmol) was dissolved in MeOH / DCM (2 / 1, v / v, 24 mL), PdCl2 (210 mg, 1.19 mmol) was added, and the reaction was stirred at 35 °C for 4.5 h. The mixture was filtered and concentrated under vacuum, and purified by silica gel column chromatography (petroleum ether / ethyl acetate 2:1 v / v) to give the corresponding hemiacetal. At 0 °C, 2,2,2-trifluoro-N-phenyliminoacetyl chloride (0.89 mL, 5.95 mmol) and DBU (0.53 mL, 3.57 mmol) were added to the above hemiacetal in DCM (12 mL). The reaction was heated to room temperature and stirred for 3 h. The solution was concentrated under vacuum to the residue, which was purified by silica gel column chromatography (petroleum ether / ethyl acetate 20:1 → 10:1 v / v) to give pale yellow syrup 32 (0.62 g, 0.93 mmol, 78%, two-step reaction). [α] 25 D = +31.1° (c 1.00, CH3Cl); 1H NMR(600MHz,Chloroform-d)δ7.43–6.76(m,15H,Ar-H),5.23(d,J=12.2Hz,1H,ArCH),5 .19(d,J=12.2Hz,1H,ArCH),5.13–5.04(m,1H,2-H),4.68(d,J=10.5Hz,1H,ArCH),4.49( d,J=10.5Hz,1H,ArCH),4.17–3.89(m,1H,5-H),3.83(t,J=9.5Hz,1H,4-H),3.75–3.63(m ,1H,3-H),2.84–2.78(m,2H,CH2-Lev),2.67–2.62(m,2H,CH2-Lev),2.18(s,3H,CH3CO); 13 C NMR(151MHz,Chloroform-d)δ205.76,171.05,167.20,142.92,136.71,134.69,128.78,128.73,128.70,128.64,128.48,128.25,128.22, 124.57,119.25,94.49,77.29,75.54,75.14,70.60,67.81,65.67,37.88,29.67,27.74; IR ν max (film)=2919,2109,1749,1719,1597,1489,1455,1405,1365,1327,1212,1161,1087,1028,910,778,755,696cm -1 .

[0123] Compound 33: At -20°C, N-phenyltrifluoroacetylimine donor 32 (0.45 g, 0.67 mmol) was placed in anhydrous DCM (7 mL), and TolSH (83 mg, 0.67 mmol) was added. Molecular sieve (flame dried). TMSOTf (23 μL, 0.134 mmol) was added dropwise under argon atmosphere. When TLC showed complete conversion of the starting material, the reactants were quenched with Et3N, then filtered and concentrated under vacuum. The crude product was purified by silica gel column chromatography (petroleum ether / ethyl acetate v / v 5:1) to give a white solid 33 (343 mg, 0.57 mmol, 85%). [α] 25 D = -14.32 (c 1.0, CH3Cl); 1H NMR(600MHz,Chloroform-d)δ7.36(d,J=6.5Hz,5H,Ar-H),7.30–7.24(m,5H,Ar-H),7.18(d,J=7.8Hz,2H,Ar-H),7.05(d,J=7.7Hz,2H,Ar-H),5.23–5.16(m,2H,ArCH),4.81(dd,J=10.8,8.9Hz,1H,2-H),4.67(d,J=10.5Hz,1H,ArCH),4.56(d,J=9.9Hz,1H,1-H),4.49(d,J=10.5Hz,1H,ArCH),3.95(d,J=9.5Hz,1H,5-H),3.70(t,J=9.6Hz,1H,4-H),3.62(t,J=9.7Hz,1H,3-H),2.83(td,J=6.6,2.1Hz,2H,Lev-H),2.69(q,J=6.9Hz,2H,Lev-H),2.32(s,3H,Lev-CH3),2.21(d,J=1.8Hz,3H,Tol(CH3)); 13 C NMR(151MHz,Chloroform-d)δ205.94,171.15,167.25,138.74,136.97,134.92,133.76,129.72,128.66,128.62,128.53,128.43,128.21,128.16,128.10,127.64,87.00,78.55,77.55,75.06,70.04,67.87,67.56,37.83,29.84,27.99,21.18;IR ν max (film)=2919,2107,1748,1718,1494,1455,1401,1363,1262,1201,1174,1150,1072,1020,910,810,752,698,580cm -1 ;HR-ESI-MS(m / z):calcd for C 32 H 33 N3O7SNa + (M+Na + ):626.1932,found:626.1929.

[0124] Compound 34: Compound 21 (1.70 g, 1.60 mmol) was placed in 16 mL of THF solution and stirred at room temperature. AcOH (91 μL, 1.60 mmol) and TBAF (2.4 mL, 2.4 mmol) were added, and the mixture was stirred for 5 h. The organic phase was dried over anhydrous Na₂SO₄ and concentrated under vacuum. The crude product was purified by silica gel column chromatography (PE / EA v / v 4:1→3:2) to obtain colorless syrup 34 (1.40 g, 1.47 mmol, 92%). [α] 25 D = +37.8° (c 1.00, CH3Cl); 1 H NMR(600MHz,Chloroform-d)δ7.40–7.14(m,20H,Ar-H),6.71–6.57(m,2H,NH),5.3 1–5.14(m,4H,ArCH),4.88(d,J=11.6Hz,1H,ArCH),4.78(t,J=7.1Hz,1H,2'-H),4.7 5(d,J=6.4Hz,1H,1'-H),4.67–4.56(m,3H,2-H,1-H,ArCH),4.53(d,J=11.6Hz,1H,A rCH),4.40(d,J=15.9Hz,1H,ArCH),4.07(d,J=9.2Hz,1H,5'-H),4.01–3.90(m,2H,3 '-H,4'-H),3.83–3.71(m,2H,5-H,3-H),3.69–3.57(m,3H,4-H,CH2-Linker),3.39 –3.22(m,2H,CH2-Linker),2.90(ddd,J=18.4,9.8,4.4Hz,1H,CH2-Lev),2.81–2.71 (m,1H,CH2-Lev),2.69–2.55(m,2H,CH2-Lev),2.15(s,3H,CH3CO),2.04(s,3H,CH3C O),2.01(s,3H,CH3CO),1.81–1.58(m,2H,CH2-Linker),1.08(d,J=6.4Hz,3H,6-H); 13C NMR(151MHz,Chloroform-d)δ206.81,174.23,173.56,170.80,168.55,156.43,138.80,137.4 2,136.46,135.27,128.69,128.58,128.41,128.28,128.14,128.13,127.88,127.69,127.52,1 27.25,101.48(C-1'),97.88(C-1),79.69,78.94,75.78,74.92,73.02,70.37,67.43,67.20,6 6.74,63.58,57.57,49.72,48.34,42.92,38.04,29.68,27.69,27.25,23.38,22.72,16.58;IRν max (film)3305,2924,1747,1661,1549,1454,1423,1372,1218,1177,1127,1072,1042,751,699,593cm -1 ;HR-ESI-MS(m / z):calcdfor C 53 H 63 N3O 15 Na + (M+Na + ):1004.4151,found:1004.4157.

[0125] Compound 35:

[0126] Condition 1: At -10℃, donor 32 (21 mg, 0.0306 mmol), recipient 34 (20 mg, 0.0204 mmol), and Molecular sieves (flame-dried) were placed in a dry DCM (1.5 mL) solution and stirred. Under argon protection, TMSOTf (1.1 μL, 0.006 mmol) was added dropwise. When TLC showed complete conversion of the glycosyl donor, Et3N was added to quench the reaction, followed by filtration and vacuum concentration. The crude product was purified by silica gel column chromatography and detected by MALDI-TOF and NMR. Unreacted acceptor and donor degradation products indicated glycosylation failure.

[0127] Condition 2: At -20℃, glucosinolate donor 33 (18 mg, 0.0306 mmol), receptor 34 (20 mg, 0.0204 mmol), and Molecular sieves were placed in dry DCM (1.5 mL) and stirred. NIS (8.3 mg, 0.0367 mmol) and TMSOTf (1.1 μL, 0.006 mmol) were added. The reaction was stirred at 0 °C for 5 h until TLC showed complete conversion of the glycosyl donor. The reaction solution was diluted with DCM and filtered. The filtrate was washed with 10% (w / v) Na₂S₂O₃, saturated NaHCO₃ solution, and brine. The organic phase was dried over anhydrous Na₂SO₄ and concentrated under vacuum. The crude product was purified by silica gel column chromatography, yielding only trace amounts. The large amount of unreacted receptor residue indicated glycosylation failure.

[0128] Comparative Example 2

[0129] An attempt to synthesize trisaccharide 38:

[0130] As shown in Figure 10, to improve the reactivity of the disaccharide receptor, we attempted to use unreduced azide disaccharide 8 as the receptor for trisaccharide assembly. First, we tried glycosylation of disaccharide receptor 8 with trifluoroacetylimine 32 and thioglycoside 33 as glycosyl donors, respectively, to obtain trisaccharide 27 with only the β-configuration, with yields of 15% and 21%, respectively. Although the yields were not ideal, the reactivity of the disaccharide receptor was improved, and the thioglycoside donor was more reactive than the trifluoroacetylimine donor. To achieve orthogonal modification of the C3” amino group, we attempted to synthesize trisaccharide 38 by glycosylation of thioglycoside donor 37 with disaccharide receptor 21 at -10℃ to 0℃ using TMSOTf and NIS as activators. However, the yield of the target trisaccharide 29 was not ideal, only 3%, and the reaction yielded byproducts of donor decomposition and glycosyl receptor 21.

[0131] Specific experimental procedures and steps:

[0132] Compound 36: Condition 1: At -10°C, donor 32 (21 mg, 0.0316 mmol), acceptor 21 (20 mg, 0.0211 mmol) and Molecular sieves (flame-dried) were placed in a dry DCM solution (1.6 mL) and stirred. Under argon protection, TMSOTf (1.1 μL, 6.32 μmol) was added dropwise. When TLC showed complete conversion of the glycosyl donor, Et3N was added dropwise to quench the reaction, followed by filtration and vacuum concentration. The crude product was purified by silica gel column chromatography (PE / EA v / v 5:1→2:1) to obtain colorless syrup 36 (4.5 mg, 4.74 μmol, 15%).

[0133] Condition 2: At -20℃, glucosinolate donor 33 (19 mg, 0.0316 mmol), receptor 21 (20 mg, 0.0211 mmol), and Molecular sieves were placed in dry DCM (1.5 mL) and stirred. NIS (8.5 mg, 0.038 mmol) and TMSOTf (1.1 μL, 6.62 μmol) were added. The reaction was stirred at 0 °C for 5 h until TLC showed complete conversion of the glycosyl donor. The reaction solution was diluted with DCM and filtered. The filtrate was washed with 10% (w / v) Na₂S₂O₃, saturated NaHCO₃ solution, and brine. The organic phase was dried over anhydrous Na₂SO₄ and concentrated under vacuum. The crude product was purified by silica gel column chromatography (PE / EA v / v 5:1→2:1) to obtain colorless syrup 36 (9.5 mg, 6.64 μmol, 21%). [α] 25 D = +18.9° (c 0.50, CH3Cl); 1H NMR(600MHz,Chloroform-d)δ7.38–7.14(m,30H,Ar-H),5.32(d,J=11.8Hz,1H,ArCH),5.20–5.13(m,4H,ArCH),5.11(d,J=11.9Hz,1H,ArCH),4.85(dd,J=10.4,7.9Hz,1H,2’-H),4.80(d,J=11.4Hz,1H,ArCH),4.82–4.74(m,1H,1-H),4.75(dd,J=10.3,7.9Hz,1H,2”-H),4.73–4.66(m,1H,1’-H),4.62(d,J=10.7Hz,1H,ArCH),4.55–4.46(m,2H,ArCH),4.44(J=10.4Hz,1H,ArCH),4.44(d,J=8.3Hz,1H,1”-H),4.40(d,J=11.5Hz,1H,ArCH),4.14–4.07(m,1H,5’-H),3.98(t,J=9.5Hz,1H,4’-H),3.96–3.88(m,1H,3-H),3.77(d,J=9.6Hz,1H,5”-H),3.71–3.75(m,1H,5-H),3.69(t,J=9.6Hz,1H,4”-H),3.67–3.64(m,2H,2-H),3.64–3.58(m,1H,CH2-Linker),3.59–3.54(m,2H,3’-H,4-H),3.48–3.35(m,2H,CH2-Linker),3.31(dd,J=11.6,8.2Hz,2H,3”-H,CH2-Linker),2.87–2.50(m,8H,CH2-Lev),2.19(s,3H,CH3CO),2.16(s,3H,CH3CO),1.89–1.73(m,2H,CH2-Linker),1.04–0.94(m,3H,6-H); 13C NMR(151MHz,Chloroform-d)δ206.17,205.93,171.23,167.06,166.88,138.43,137.83,137.04,134.82,134.71,130.04,129 .74,129.26,129.19,128.90,128.71,128.65,128.64,128.58,128.40,128.19,128.10,128.04,127.84,127.62,127.42,127 .25,102.54(C-1'),100.31(C-1”),98.15(C-1),78.85,77.65,76.66,75.21,74.93,74.91,74.69,70.96,70.63,67.72,67.2 6,66.39,65.83,65.80,64.61,59.38,50.59,37.95,37.70,29.87,29.86,27.77,27.60,27.23,27.17,24.74,22.70,16.38; IR ν max (film)2918,2109,1749,1715,1372,1218,1146,1042,751,699cm -1 HR-ESI-MS(m / z):calcd for C 74 H 80 N 10 O 20 Na + (M+Na + ):1451.5443,found:1451.5451.

[0134] Compound 37: Under nitrogen protection, zinc powder (106 mg, 1.66 mmol) was added to a solution of compound 33 (50 mg, 0.083 mmol) in AcOH / THF (1:2 v / v, 3 mL). The mixture was stirred at 37 °C for 12 hours. The solution was filtered and concentrated under vacuum to obtain a crude product. The crude product was dissolved in THF (7 mL) and pyridine (20 μL, 0.249 mmol, 3 equivalents), and then a solution of TrocCl (11.4 μL, 0.083 mmol) in THF (1 mL) was added dropwise at 0 °C. The reaction mixture was stirred at 0 °C for 2 hours until TLC showed complete conversion of the starting material, and then quenched by the addition of MeOH at 0 °C. The mixture was concentrated under vacuum to the residue and purified by silica gel column chromatography (petroleum ether / ethyl acetate 5:1 → 2:1 v / v) to give product 37 (19 mg, 0.026 mmol, 31%) as a colorless syrup. [α]25 D =-25.3°(c 1.00,CH3Cl); 1 H NMR(600MHz,Chloroform-d)δ7.47–6.94(m,14H,Ar-H),5.21(s,2H,CH2-Troc),4.88(d,J=9.4Hz,1H,NH),4.76(d,J=12.1Hz,1H,CH2-Troc),4.75(t,J=9.7Hz,1H,2-H),4.70(d,J=12.1Hz,1H,CH2-Troc),4.64(d,J=9.8Hz,1H,1-H),4.48(d,J=11.0Hz,1H,ArCH),4.44(d,J=11.1Hz,1H,ArCH),4.03(d,J=9.4Hz,1H,5-H),3.94(q,J=9.8Hz,1H,3-H),3.74(t,J=9.7Hz,1H,4-H),2.81–2.68(m,2H,CH2-Lev),2.64–2.57(m,2H,CH2-Lev),2.33(s,3H,CH3CO),2.18(s,3H,ArCH3); 13 C NMR(151MHz,Chloroform-d)δ205.86,171.78,167.67,154.25,138.82,137.05,134.95,134.03,129.71,128.66,128.60,128.53,128.41,128.11,128.03,127.22,95.47,86.74,79.10,76.52,74.64,74.26,69.82,67.53,58.36,37.80,29.78,27.97,21.21;IR ν max (film)3334,2923,1742,1719,1544,1495,1455,1401,1364,1317,1288,1257,1205,1157,1074,1029,814,735,698cm -1 ;HR-ESI-MS(m / z):calcd for C 35 H 36 Cl3NO9SNa + (M+Na + ):774.1069,found:774.1072.

[0135] Compound 38: At -10°C, glucosinolate donor 37 (24 mg, 0.0316 mmol), receptor 21 (20 mg, 0.0211 mmol) and... Molecular sieves were placed in dry DCM (1.6 mL) and stirred. NIS (9 mg, 0.038 mmol) and TMSOTf (1.1 μL, 6.32 μmol) were added. The reaction was stirred at 0 °C for 5 h until TLC showed complete conversion of the glycosyl donor. The reaction solution was diluted with DCM and filtered. The filtrate was washed with 10% (w / v) Na₂S₂O₃, saturated NaHCO₃ solution, and brine. The organic phase was dried over anhydrous Na₂SO₄ and concentrated under vacuum. The crude product was purified by silica gel column chromatography (PE / EA v / v 3:1→1:1) to obtain colorless syrup 38 (1.5 mg, 0.95 μmol, 3%). 1 H NMR(600MHz,Chloroform-d)δ7.38–7.10(m,30H,Ar-H),5.35(d,J=11.8Hz,1H,ArCH), 5.21–5.12(m,4H,ArCH),5.07(d,J=11.8Hz,1H,ArCH),4.86(dd,J=10.3,7.9Hz,1H,2' -H),4.83–4.74(m,3H,1-H,CH2-Troc),4.76–4.70(m,2H,2”-H,ArCH),4.70–4.65(m,1 H,1'-H),4.55–4.48(m,2H,ArCH),4.46–4.39(m,4H,1”-H,ArCH),4.06–3.97(m,2H),3. 97–3.87(m,1H),3.81(q,J=9.8Hz,1H),3.79–3.70(m,3H),3.66(dd,J=10.9,3.7Hz,1H ),3.66–3.61(m,1H,CH2-Linker),3.59–3.54(m,2H),3.48–3.29(m,3H,CH2-Linker),2 .85–2.65(m,6H,CH2-Lev),2.58–2.46(m,2H,CH2-Lev),2.17(s,3H,CH3CO),2.16(s,3 H,CH3CO),1.90–1.71(m,2H,CH2-Lev),1.03–0.95(m,3H,6-H); HR-ESI-MS(m / z):calcd for C 77 H 83 Cl3N8O 22 Na + (M+Na +):1599.4580,found:1599.4586.

[0136] Comparative Example 3

[0137] An attempt to synthesize trisaccharides 43 and 44:

[0138] As shown in Figure 11, under the action of TMSOTf and N-iodosuccinimide (NIS), acceptor 16 and thioglycoside donor 39 underwent glycosylation at temperatures ranging from -10°C to 0°C to obtain disaccharide 39 with complete β-selectivity in a yield of 79%. Hydrolysis of the 4,6-O-benzyl group of compound 40 with trifluoroacetic acid (TFA) yielded diol 41 in a yield of 82%. Subsequently, selective protection of the C6' hydroxyl group with tert-butyldimethylsilyl (TBDPS) failed to yield the desired product despite the reaction being carried out at 80°C. Considering that selective protection of the C6' hydroxyl group with tert-butyldimethylsilyl (TBS) could reduce the influence of steric hindrance, the reaction was subsequently carried out at 50°C, successfully yielding product 42 in a yield of 69%. However, unfortunately, even with the equivalent amount of catalyst TMSOTf increased to 0.6 equivalents, the glycosylation reaction between glycosyl donor 37 and disaccharide acceptor 42 still failed to proceed successfully. Furthermore, the glycosylation reaction of the more reactive donor 19 with the disaccharide acceptor 42 also failed to yield the desired trisaccharide. Analysis of the decomposed donor byproducts and unreacted acceptor suggests that the failure of the above glycosylation reaction may be due to steric hindrance of the TBS at the O6 position.

[0139] Specific experimental procedures and steps:

[0140] Compound 40: Under argon protection and at -10°C, the solution containing sulfur donor 39 (610 mg, 1.22 mmol), acceptor 16 (380 mg, 0.68 mmol), and newly activated sulfur were reacted. NIS (330 mg, 1.46 mmol) and TMSOTf (44 μL, 0.244 mmol) were added to a mixture of anhydrous DCM (6.8 mL) and MS. The reaction mixture was stirred at 0 °C for 4.5 h until TLC showed complete conversion. The mixture was diluted with DCM and filtered. The filtrate was washed successively with 10% (w / v) Na₂S₂O₃ solution, saturated NaHCO₃, and brine. The organic layer was dried over anhydrous Na₂SO₄, filtered, and concentrated under vacuum. The residue was purified by silica gel column chromatography (petroleum ether / ethyl acetate 5:1 → 2:1 v / v) to give product 40 (501 mg, 0.54 mmol, 79%). [α] 25 D = +28.0° (c 1.00, CH3Cl); 1H NMR(400MHz,Chloroform-d)δ7.56–7.09(m,20H,Ar-H),5.59(s,1H,ArCH),5.23–5.12(m,1H,ArCH),4.96(dd,J=9.9,7.8Hz,1H,2’-H),4.88(d,J=11.5Hz,1H,ArCH),4.86–4.80(m,1H,1-H),4.80–4.76(m,1H,1’-H),4.62–4.31(m,4H,ArCH,6’-H),4.06–3.93(m,1H,CH2-Linker),3.88–3.74(m,3H,3’-H,6’-H,5-H),3.57–3.26(m,5H,4’-H,5’-H,CH2-Linker,3-H,4-H),3.56–3.41(m,2H,2-H,CH2-Linker),3.37–3.30(m,1H,CH2-Linker),2.91–2.63(m,4H,CH2-Lev),2.16(s,3H,CH3CO),1.90–1.82(m,2H,CH2-Linker),1.14–1.05(m,3H,6-H); 13 C NMR(101MHz,Chloroform-d)δ206.01,171.40,138.40,137.82,136.50,129.28,128.69,128.60,128.51,128.39,128.35,128.22,128.03,127.83,127.72,127.42,127.23,126.02,102.63(C-1’),101.60,98.34(C-1),79.41,77.35,77.35,77.03,76.72,74.95,71.90,68.52,67.27,66.52,65.89,63.11,59.54,50.49,37.92,29.77,27.75,16.44;IR ν max (film)2933,2108,1753,1698,1497,1454,1421,1360,1315,1218,1125,1092,1043,1002cm -1 ;HR-ESI-MS(m / z):calcd for C 49 H 55 N7O 12 Na + (M+Na + ):956.3801,found:956.3805.

[0141] Compound 41: Trifluoroacetic acid (0.5 mL, 6.72 mmol) and H₂O (88 μL, 4.90 mmol, 10 eq.) were added to a DCM (5.0 mL) solution containing compound 40 (460 mg, 0.49 mmol). The reaction mixture was stirred at room temperature for 4 hours until TLC showed complete conversion of the starting material. The mixture was diluted with DCM and washed with water, saturated NaHCO₃ solution, and saturated brine, dried over Na₂SO₄, and concentrated under vacuum. The residue was purified by silica gel column chromatography (DCM / MeOH 50:1 → 20:1 v / v) to give compound 41 (340 mg, 0.402 mmol, 82%) as a colorless syrup. [α] 25 D = +27.2° (c 1.00, CH3Cl); 1 H NMR(600MHz,Chloroform-d)δ7.46–7.13(m,15H,Ar-H),5.22–5.10(m,2H,ArCH),4.91(dd,J=9.7,7.8Hz,1H,2'-H),4.88(d,J=11.6Hz,1H,Ar CH),4.86–4.77(m,1H,1-H),4.77–4.69(m,1H,1'-H),4.59(d,J=11.6Hz,1H,ArCH),4.57–4.41(m,2H,ArCH),4.12–3.94(m,1H,3-H),3.90–3.7 4(m,3H,5-H,6'-H),3.72(dd,J=10.8,3.6Hz,1H,2-H),3.67–3.53(m,4H,4-H,CH2-Linker,3'-H,4'-H),3.49–3.38(m,2H,5'-H,CH2-Linker) ,3.38–3.27(m,1H,CH2-Linker),2.84–2.66(m,4H,CH2-Lev),2.15(s,3H,CH3CO),1.90–1.74(m,2H,CH2-Linker),1.10(d,J=6.4Hz,3H,6-H); 13C NMR(151MHz,Chloroform-d)δ206.31,171.52,156.38,138.44,137.68,136.49 ,128.60,128.56,128.50,128.20,128.05,127.80,127.69,127.42,127.20,10 1.86(C-1),98.27(C-1'),78.86,77.46,76.10,74.80,71.83,69.75,67.35,66 .83,66.60,65.38,62.29,59.77,50.31,44.34,37.87,29.74,28.22,16.47; IR ν max (film)3462,2935,2107,1750,1696,1497,1454,1423,1362,1314,1220,1161,1126,1044cm -1 HR-ESI-MS(m / z):calcd for C 42 H 51 N7O 12 Na + (M+Na + ):868.3488,found:868.3493.

[0142] Compound 42: To a solution of 41 (140 mg, 0.166 mmol) in anhydrous DMF (0.3 mL), tert-butyldimethylchlorosilane (30 mg, 0.20 mmol), imidazole (23 mg, 0.33 mmol), and 4-dimethylaminopyridine (3 mg, 0.02 mmol) were added. The reaction mixture was stirred overnight at 50 °C, and then quenched by adding methanol (3 mL). The mixture was diluted with ethyl acetate (30 mL) and washed with water, saturated NaHCO3 solution, and saturated brine. The combined organic phases were dried over Na2SO4, filtered, and concentrated. The crude product was purified by rapid silica column chromatography (petroleum ether / ethyl acetate 10:1 v / v) to give 42 (110 mg, 0.115 mmol, 69%) as a colorless oil. [α] 25 D = +19.7° (c 1.00, CH3Cl); 1H NMR(600MHz,Chloroform-d)δ7.40–7.15(m,15H,Ar-H),5.22–5.14(m,2H,ArCH),4.92–4.88(m,2H,ArCH,2’-H),4.84–4.76(m,1H,1-H),4.73–4.65(m,1H,1’-H),4.57–4.43(m,3H,ArCH),4.02–3.93(m,1H,3-H),3.93–3.86(m,1H,6’-H),3.86–3.73(m,1H,5-H),3.81(dd,J=10.3,6.0Hz,1H,6’-H),3.69(dd,J=10.8,3.6Hz,1H,2-H),3.68–3.65(m,1H,4-H),3.66–3.54(m,3H,CH2-Linker,3’-H,4’-H),3.51–3.30(m,4H,5’-H,CH2-Linker),2.86–2.68(m,4H,CH2-Lev),2.16(s,3H,CH3CO),1.91–1.74(m,2H,CH2-Linker),1.10(d,J=6.6Hz,3H,6-H),0.89(s,9H,SiC(CH3)3),0.12–0.06(m,6H,Si(CH3)2); 13 C NMR(151MHz,Chloroform-d)δ206.09,171.50,156.23,138.53,137.81,136.60,128.58,128.50,128.15,128.02,127.82,127.59,127.38,127.21,102.01(C-1’),98.33(C-1),79.05,77.84,75.03,74.71,72.15,71.59,67.27,66.60,66.52,65.82,64.41,59.61,50.55,44.67,37.92,29.75,27.80,25.78,18.17,16.42,-5.49;IR ν max (film)3419,2929,2108,1754,1704,1472,1455,1423,1361,1317,1253,1153,1130,1047,970cm -1 ;HR-ESI-MS(m / z):calcd for C 48 H 65 N7O 12 SiNa + (M+Na+ ):982.4353,found:982.4355.

[0143] Compound 43: Under argon protection at -10°C, the reaction mixture was subjected to an atmosphere containing donor 37 (32 mg, 0.042 mmol), disaccharide acceptor 42 (20 mg, 0.021 mmol), and newly activated [agent / acceptor]. NIS (11 mg, 0.05 mmol) and TMSOTf (1.5 μL, 8.4 μmol) were added to anhydrous DCM (0.5 mL) mixture in MS. The reaction mixture was stirred at 0 °C for 10 h until TLC showed complete conversion of the glycosyl donor. The reaction was quenched with Et3N, then filtered and concentrated under vacuum. The residue was purified by silica gel column chromatography and analyzed using MALDI-TOF and NMR. Unreacted acceptor and donor degradation products indicated failure of the glycosylation reaction.

[0144] Compound 44: Under argon protection at -10°C, the reaction mixture was subjected to an atmosphere containing donor 19 (21 mg, 0.042 mmol), disaccharide acceptor 42 (20 mg, 0.021 mmol), and newly activated [agent / acceptor]. NIS (11 mg, 0.05 mmol) and TMSOTf (1.5 μL, 8.4 μmol) were added to anhydrous DCM (0.5 mL) mixture in MS. The reaction mixture was stirred at 0 °C for 8 hours until TLC showed complete conversion of the glycosyl donor. The reaction was quenched with Et3N, then filtered and concentrated under vacuum. The residue was purified by silica gel column chromatography and analyzed using MALDI-TOF and NMR. Unreacted acceptor and donor degradation products indicated failure of the glycosylation reaction.

[0145] The present invention has been described above with reference to specific embodiments. Obviously, the implementation of the present invention is not limited to the above-described manner. Any improvements made using the inventive concept and technical solution of the present invention, or the direct application of the inventive concept and technical solution to other situations without modification, are all within the protection scope of the present invention.

Claims

1. A chemical synthesis method for an oligosaccharide fragment with a linker arm assembled from the O-antigen trisaccharide of Pseudomonas aeruginosa O5 serotype O-antigen, characterized in that, The structure of the oligosaccharide fragment with a linker arm assembled from the O-antigen trisaccharide of Pseudomonas aeruginosa O5 serotype is shown in general formula VI: Wherein, Linker is -L-NH2, where L represents the linker arm; the linker arm L is a chain structure with 2-40 carbon atoms containing 0-6 heteroatoms, a substituted or unsubstituted three- or six-membered ring structure, an amide bond, or a urea group; The three monosaccharide building blocks A, B, and C are used as raw materials, as shown in formulas (I) to (III) respectively: in: PG1 is H; PG2, PG3, PG4 and PG6 are temporary hydroxyl protecting groups, each independently selected from: acetyl, or benzoyl, or neopentyl, or chloroacetyl, or acetylpropionyl, or 9-fluorenylmethoxycarbonyl, or allyloxycarbonyl, or 2-naphthylmethyl, or p-methoxybenzyl, or allyl, or tert-butyldimethylsilyl, or triethylsilyl; PG5 is a carboxyl protecting group, selected from benzyl, methyl, ethyl, tert-butyl, and allyl. PG7 is a temporary amino protecting group selected from trichloroethoxycarbonyl, phthaloyl, 9-fluorenylmethoxycarbonyl, and tert-butyloxycarbonyl. PG8 and PG9 are hydroxyl protecting groups selected from benzenemethylene, naphthenemethylene, and isopropyl ketal; PG 10 PG 11 It is an amino protecting group, selected from benzyl or benzyloxycarbonyl; LG is a leaving group used in glycosylation reactions, selected from fluorine, chlorine, bromine, iodine, trichloroacetylimine ester, N-phenyltrifluoroacetylimine ester glycoside, ethylthio, phenylthio, p-toluenethio, ethylthio, or dibutylphosphonic acid group; The process includes the following: (1) By glycosylation reaction of monosaccharide building block B and monosaccharide building block A, the disaccharide fragment shown in Formula IV was synthesized. (2) The obtained disaccharide fragment was first deprotected and the protecting group PG4 was removed. Then, it was glycosylated with the monosaccharide building block C to synthesize the trisaccharide intermediate fragment as shown in Formula V. (3) The obtained trisaccharide intermediate fragment was reduced, and PG3 and PG6 were deprotected. The deprotected position 2 in monosaccharide building block B and the deprotected position 2 in monosaccharide building block C were azidized. Subsequently, positions 4 and 6 of monosaccharide building block C in the trisaccharide intermediate fragment were reduced, PG8 and PG9 were removed, and position 6 was oxidized to obtain a carboxylic acid group. Then, the deprotected position 2 in monosaccharide building block B and the azid group at position 2 in monosaccharide building block C were reduced to obtain acetyl. The amino group was then used to construct an acetamidine structure at position 3 of the monosaccharide building block C using an imine reagent. Finally, the amino group in the linker was reduced to obtain the target product shown in Formula VI.

2. The chemical synthesis method according to claim 1, characterized in that, The synthesis route is shown below:

3. The chemical synthesis method according to claim 1, characterized in that, The glycosylation reaction in step (1) is carried out in an environment with Lewis acid as the activating reagent, solvent, and molecular sieve.

4. The chemical synthesis method according to claim 3, characterized in that, The Lewis acid used as the activating agent for the glycosylation reaction in step (1) is selected from any one or more of the following: boron trifluoride ether, trifluoromethanesulfonic acid, trimethylsilyl trifluoromethanesulfonate, and silver salt silver trifluoromethanesulfonate.

5. The chemical synthesis method according to claim 1, characterized in that, The glycosylation reaction in step (2) is carried out in an environment with Lewis acid as the activating reagent, NIS, solvent, and molecular sieve.

6. The chemical synthesis method according to claim 5, characterized in that, In step (2), the amount of Lewis acid used as the activating agent for the glycosylation reaction is 0.1–1 eq, and the amount of NIS is 1.2–1.5 eq.

7. The chemical synthesis method according to claim 5, characterized in that, The Lewis acid used as the activating agent for the glycosylation reaction in step (2) is selected from any one or more of the following: boron trifluoride ether, trifluoromethanesulfonic acid, trimethylsilyl trifluoromethanesulfonate, and silver salt silver trifluoromethanesulfonate.

8. A Pseudomonas aeruginosa O5 serotype O-antigen trisaccharide compound assembled with an amino linker arm, characterized in that, The compound is prepared using the method described in any one of claims 1-7, and has the following structure:

9. The use of the Pseudomonas aeruginosa O5 serotype O-antigen trisaccharide compound assembled with amino linker arms as described in claim 8 in the development or preparation of Pseudomonas aeruginosa vaccines.

10. The use of the Pseudomonas aeruginosa O5 serotype O-antigen trisaccharide compound assembled with an amino linker arm as described in claim 8 in the development or preparation of a therapeutic drug for Pseudomonas aeruginosa infection.

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