JP2002501932A - Liquid and Solid Sulfoxide Glycosylation: Synthesis of β-Linked Oligosaccharides Using a 2-Deoxy-2-N-trifluoroacetamide-Glycopyranosyl Donor - Google Patents

Abstract

  (57) [Summary] The present invention relates to a process for synthesizing β-oligosaccharides. Beta-oligosaccharides can be converted to liquid and solid phases via sulfoxide glycosylation using alkylsulfenyl- or arylsulfenyl-2-deoxy-2-N-trifluoroacetamidoglycopyranose as the glycosyl donor. Synthesized in both. Once activated under glycosylation conditions, these donors produce β-glycosides exclusively and in high yield. Since the trifluoroacetamide group is easily removed under mild conditions, the corresponding amino group can be appropriately derivatized even in the presence of an unprotected hydroxyl group. A disaccharide library is designed, constructed and analyzed. The present invention also relates to a process for synthesizing a glycosyl donor.

Description

DETAILED DESCRIPTION OF THE INVENTION

TECHNICAL FIELD The present invention relates generally to β-oligosaccharides and processes for synthesizing β-oligosaccharides, including reacting a glycosyl donor with a glycosyl acceptor. More specifically, the present invention relates to a process for the synthesis of β-oligosaccharides using 2-deoxy-2-N-trifluoroacetamidoglycopyranosylsulfoxide as a glycosyl donor in liquid and solid phase sulfoxide glycosylation. About. The invention also relates to the design, construction and analysis of a disaccharide combinatorial library.

BACKGROUND OF THE INVENTION The biological relevance of oligosaccharide chains of glycoproteins and glycolipids is becoming increasingly clear. As both are found on cell surface membranes and circulate in biological fluids, these glycoside residues act as recognition signals that mediate key events in normal cell development and function I do. They are involved in embryogenesis, hormonal activity, neuronal development, inflammation, cell proliferation, fertilization, and organization of various cell types into specific tissues. They can regulate the intercellular transport of proteins and should therefore be regarded as metabolic signalers. They are also involved in the intracellular localization and secretion of glycoproteins, and the elimination of plasma glycoproteins from the circulation, and many oligosaccharides are involved in cell-cell recognition, cellular immune responses, cell carcinogenesis. It is involved in transformation and inflammation, and other cell biological phenomena. Sharon et al. , Sci. Am. 268: 82-89 (1993)
); Karlsson, Trends Pharm. Sci. , 12 (7): 2
65-272 (1991); Drickamer et al. Curr. O
pn. Struct. Biol. 2 (5): 653-654 (1992).

[0003] Oligosaccharides are also involved in the prevention and treatment of diseases. This class of compounds includes glucosamine and macrolide antibiotics, anthracyclines and enediyne anticancer antibiotics. For example, oligosaccharides on the cell surface function as receptors for viruses, toxins, infectious bacteria, hormones, pathogens, enzymes, proteins, as well as better ligands. Modified cell surface carbohydrates have been implicated in tumorigenesis and metastasis. Oligosaccharide structures mediate cell migration, the process of infection, and other phenomena during embryonic development. Rademacher
et al. Annu. Rev .. Biochem. 57: 785 (1988).
); Feizi et al. , TIBS, 24 (1985); Hakomor.
i, TIBS, 45 (1984); Feizi TIBS, 84 (1991);
Dennis and Laferte Cancer Res. , 45: 6034
(1985); Markwell et al. Proc. Natl. Acad
. Sci. USA, 78: 5406 (1981); Wiley and Skehe.
l. Annu. Rev .. Biochem. 56: 365 (1987); W.
alz et al. , Science, 250 (1990); Kleinma.
net et al. Proc. Natl. Acad. Sci. USA, 76:33
67 (1979).

[0004] Efforts to develop products related to oligosaccharides continue, including drug delivery vehicles that recognize carbohydrate receptors, vaccines that block infection by viruses that recognize cell surface carbohydrates, and various diseases. Diagnostic kits for identifying, detecting and analyzing isolated carbohydrates, and monoclonal antibodies recognizing abnormal carbohydrates for use as drugs. The development of biomedical products based on these and other carbohydrates has led to the technology to produce oligosaccharides and other glycoconjugates quickly, efficiently, and in practical quantities for basic and developmental research. Depends on availability.

[0005] More specifically, there is a need for a method that allows for the rapid preparation of glycoside libraries containing a mixture of various oligosaccharides and other glycoconjugates, which is then followed by a specific biological activity Can be screened for For example, screening of peptide mixtures has been shown to be an efficient method of identifying active compounds and elucidating structure-activity relationships. There are a number of methods for the production of chemically distinct peptide mixtures and for determining the active compound. For example, Zucker
mann et al. Proc. Natl. Acad. Sci. USA, 8
9: 4505 (1992); Lam et al. , Nature, 354: 8
2 (1991); Houghten, Nature, 354: 84 (1991).
Petritory, Proc. Natl. Acad. Sci. USA, 88
: 11510 (1991); Geyse, Proc. Natl. Acad. Sc
i. ScL USA, 81: 3998 (1984); Houghten, Proc. Na
tl. Acad. Sci. USA, 82: 5131 (1985); Fodor,
Science, 251: 767 (1991).

[0006] Oligosaccharide synthesis is by far one of the most challenging areas of modern organic chemistry. Efficient production of oligosaccharides or glycoconjugates involves attaching glycosyl moieties to specific sites on glycosyl receptors with high yields as well as high steric control. Based on this challenge, several glycosylation methods have been developed.

[0007] The classic donor of glycosylation is a glycosyl halide. Nicol
aou et al. Preparative carbohydrate chemistry, Hanesian, S .; Hen,
Marcel Dekker, Inc. (1997) reviewed oligosaccharide synthesis from glycosyl fluoride. One of the oldest glycosylation methods is the König-Knoll method, which is carried out in the presence of heavy metal salts such as silver or mercury.
Includes activation of the anomeric center by decomposition of a glycoside halide such as bromide or chloride. However, problems can occur in the conversion of many oligosaccharides, such as glycosides or glycosyl esters, to glycosyl halides, resulting in low yield steps far from the synthesis sequence. In addition, the stringent requirements for the production of glycosyl halides, their low thermal stability, and their high susceptibility to hydrolysis make it possible, for example, to use trichloroacetamide as a glycosyl donor under Lewis or protonic acid conditions. Other methods had to be developed, such as using dating. Glycosylation proceeds with an inversion of the configuration if the participating group is not present at C-2, in which case a β-glycoside is obtained.
Schmidt et al. , Tetrahedron Lett. , 25:
821 (1984).

[0008] Several types of glycal glycosidation methods are described in Lemieux et al.
. , Can. J. Chem. 43: 2190 (1965); Thiem et.
al. , Synthesis, 696 (1978); Sinay et al.
. J. Chem. Soc. Chem. Commun. , P. 572 (1981)
Ogawa et al.); , Tetrahedron Lett. , 28:
2723 (1987) and Danishefsky et al. J. Am
. Chem. Soc. , 111: 6661 (1989). In this method, an electrophile (E ⁺ ) is used as a means to activate the anomeric center,
Attacks the electron-rich glycal, and then reacts with the carbohydrate receptor to give the glycoside. Thereafter, the C-2 substituent is removed to give 2-deoxyglycoside.

The n-pentenyl glycoside method is described in Fraser-Reid et al. , J
. Chem. Soc. Chem. Commun. , P. 823 (1988). This method relies on electrophilic addition to olefins, followed by intramolecular displacement by ring oxygen and eventual removal of the pentenyl chain to form an oxonium species. Trapping (capture) at the glycoside receptor gives the desired glycoside.

[0010] Schmidt et al. , Tetrahedron Lett. , 33
: 6123 (1992) described a relatively new method involving glycosyl phosphite as a glycosyl donor. This method finds important application in the synthesis of sial derivatives.

[0011] Thioglycosides have received considerable attention as glycosyl donors. Thioalkyl or aryl groups can efficiently and temporarily protect the anomeric center, while being selectively activated under a variety of conditions. Many glycosylation reactions using thioglycosides have been reported. Early attempts to use thioglycosides as glycosyl donors were made by mercury (II) sulfate, Ferrier et al. J.
Glycoconjugate, 4: 97-108 (1987); mercury chloride (I
I), Tsai et al. , Can. J. Chem. , 62: 1403-5.
(1984); phenylmercury trifluoromethanesulfonate, Garegge
t al. , Carbohyd. Res. , 116: 162-65 (1983).
Mercury (II) benzoate, van Cleve, Carbohyd. Res. ,
116: 162-65 (1983); mercuric nitrate, Hanesian et a.
l. , Carbohyd. Res. , 80: C17-22 (1980); silver triflate, Hanesian et al. , Carbohyd. Res. ,
53: C13 (1977); copper (II) trifluoromethanesulfonate, Muk
aiyama et al. Chem. Lett. 487-90 (1979).
N-bromosuccinimide (NBS), Hanesian, supra and N
icolaou et al. J. Am. Chem. Soc. , 105: 24
30-34 (1982); Lead (II) perchlorate, Woodward et al.
. J. Am. Chem. Soc. , 103: 3215-17 (1981); benzeneselenenyl triflate, Ito et al. Tetrahed. L
ett. , 1061-64 (1988), methyl triflate, Lonn, Ca.
rbhyd. Res. 39: 105-113 (1985); dimethyl (methylthio) sulfonium triflate (DMTST), Andersson et al.
al. Tetrahed. Lett. 3919-3922 (1986); benzenesulfonyl derivatives, Brown et al. Tetrahed. Le
tt. , 29/38: 4873-4876 (1988); Alkylsulfonyl triflate, Dasgupta et al. , Carbohyd. Resea
rch. 177: c13-c17 (1988). These various accelerators did not result in widespread use in oligosaccharide synthesis. Heavy metals are not reactive enough for general applications. This problem was circumvented by the use of more reactive heterocyclic thioglycosides. Hanesian
et al. And Mukaiyama et al. the above.

Glycosidation methods using thioglycosides are described in Kahne et al. , J
. Am. Chem. Soc. , 111: 6881-2 (1989), wherein the anomeric sulfide is first oxidized to the corresponding sulfoxide and then activated by the addition of anhydrous triflate in the presence of a glycosyl receptor to produce a glycoside I do. Glycosyl donors activated with anhydrous triflate have been found to be very reactive in solution and can be used to glycosylate extremely unreactive substrates under mild conditions.

[0013] Kahne et al. Nos. 5,635,612 and 5,638,866, the contents of which are incorporated herein by reference.
Methods are described for forming multiple glycoside chains in solution in one step using anomeric sugar sulfoxides as glycosyl donors and for constructing continuous glycoside chains in solid and liquid phases. Activators include strong acids such as trifluoromethanesulfonic acid or triflate acid (TfOH), p-toluenesulfonic acid or methanesulfonic acid.

[0014] Recent studies have shown that thioglycosides can be used in at least two other different technologies,
It has been shown that it can be conveniently and regeneratively activated. The first is a "two-step" activation that involves first forming a glycosyl halide and then further activating it with a halophilic reagent. Sato et al. , Carbohyd. R
es. 155: C6-10 (1986).

A two-step activation method using a mixture of glycosyl fluoride and sulfide for oligosaccharide synthesis is described in Nicolaou et al. J. Am. Chem. S
oc. , 106: 4189 (1984). Glycosyl fluoride is described in Mukaiyama et al. Chem. Lett. , P. 43
1 (1981) as a glycosyl donor. This method involved the activation of a carbohydrate intermediate and coupling to a glycosyl receptor in the presence of silver perchlorate and tin dichloride. The mechanism of activation is similar to the Königs-Knoll process, as is the selectivity of the reaction. If an equatorial participation group is present at C2, a β glycoside is formed. Without such moieties, the alpha anomer is the preferred product. Thioglycosides are described in Ferrier et al. Using an ethylthio group at the anomeric position. , Carbohyd. Res. , 127
157 (1984) as glycosyl donors. Hanes
Sian et al. , Carbohyd. Res. , 80: C17 (198
0) demonstrated the use of 2-pyridylthioglycoside. Nicolaou e
t al. J. Am. Chem. Soc. , 106: 4189 (1984) uses stable phenylthioglycosides as key building blocks. Phenylthioglycoside can be obtained from NBS and diethylaminosulfur trifluoride (
DAST) converts it to more reactive glycosyl fluoride. In the second activation, in the preparation of the oligosaccharide chain, glycosyl fluoride is coupled to a glycosyl receptor having phenylthio at the anomeric position. The two-step activation method is particularly suitable for solid-phase oligosaccharide synthesis.

[0016] The second technique involves one-step activation with a sulfurizing reagent such as methyl triflate or dimethyl (methylthio) sulfonium trifluoromethanesulfonate (DMTST). Full stereospecificity is achieved only at the 1,2 trans linkage, due to the use of adjacent group participation from the 2-acyl substituent, which controls the steric outcome of the reaction. DMTST is a very lipophilic promoter in the synthesis of 1,2 transglycosides using various thioglycosides with 2-substituent participation as glycosyl donors. Fugedi et al. J. Glycoconjug
ate, 4: 97-108 (1987). It has also been used in the synthesis of 1,2 cis glycosides, Andersson et al. Tetrahed.
Lett. , 27: 3919-3922 (1986).

Β-selective glycosylation without the involvement of neighboring groups has been performed using an insoluble silver catalyst. Garegg et al. , Carbohyd. Res. , 70
: A solvent capable of cationic interaction such as C13 (1979) or acetonitrile; Hashimoto et al. Tetrahed. Lett. , 25
: 1379 (1984).

A sulfonate ester, which is a glycosyl acceptor, and a benzylated methyl or phenyl ester at −35 ° C. in the presence of a Lewis acid such as CF ₃ SO ₃ SiMe ₃ , TrBF ₄ or BF ₃ .Et ₂ O The reaction of 1-thio-β-glucopyranoside generally produces an α-β mixture. Ito et al. , Tetrah
ed. Lett. , 28: 4701-4704 (1987). Stereoselectivity is very dependent on the solvent.

[0019] Sinay et al. Pure Appl. Chem. 63: 519 (
Another thioglycoside activation method developed by 1991) involves the electron transfer from sulfur to the activator tris ([alpha] -bromophenyl) ammonium hexachloroantimonate (TBPA). The glycosyl radical cation produced undergoes cleavage to a thiyl radical, leaving an oxonium species, which subsequently undergoes glycosidation.

Garegg et al. , Carbohyd. Res. , 116: 162
-65 (1983) described the use of benzylated or acylated phenyl-1-thio-β-D-glucopyranoside or 1-thio-β-D-galactopyranoside as a glycosyl donor. Similarly, Van Cleve, Carbohy
d. Res. , 70: 161-164 (1979) described the use of phenyl-1-thio-β-D-glucopyranoside as a glycosyl donor. Ferr
ier et al. J. Glycoconjugate: 4: 97-108
(1987) described the use of benzylated phenylthioglycosides as glycosyl donors. The previous method produces both α- and β-anomers.

As a result, there is a need to improve the stereoselectivity of the glycosyl donor and the stability and yield of the glycosylated product. The sulfoxide glycosylation method of the present invention comprises:
It has been shown to be successful in both liquid and solid phases. It allows glycosylation of sensitive and / or non-reactive substrates, generally at high stereoselectivity, under low temperature and substantially neutral conditions.

SUMMARY OF THE INVENTION The present invention generally relates to a process for synthesizing β-oligosaccharides, including β-disaccharides and conjugates thereof, wherein the process comprises reacting a glycosyl donor with a glycosyl acceptor. Including. In particular, the present invention relates to the use of β-alkylsulfenyl- or arylsulfenyl-2-deoxy-2-N-trifluoroacetamidoglycopyranoses as glycosyl donors in sulfoxide-mediated glycosylation reactions in solution or solid phase. The present invention relates to an oligosaccharide synthesis process. Suitable te
rt-butylsulfenyl- or phenylsulfenyl-2-deoxy-2-
A glycosyl donor, including N-trifluoroacetamidoglycopyranose,
It is described together with the preparation method of the same.

[0023] A further aspect of the present invention is that the glycosyl donor is arylsulfenyl-
Or using alkylsulfenyl-2-deoxy-2-N-trifluoroacetamidoglycopyranose to provide a process for building glycoside linkages. Using selected conditions and starting materials such as galactosamine hydrochloride and glucosamine hydrochloride, produce β-anomers (ie, β-glycoside chains) on a solid phase using the anomeric sugar sulfoxide as a glycosyl donor. it can. The process of the present invention may be used for the preparation of specific oligosaccharides or glycoconjugates, or for the creation of glycoside chemical libraries, which can subsequently be screened to detect compounds with the desired biological activity. Can be applied to the preparation of mixtures of various oligosaccharides or glycoconjugates.

In another embodiment, the present invention provides a method for preparing β-isomers in high yield in the substantial absence of
A process for synthesizing glycosides;

In yet another aspect, the present invention is broadly directed to β-selection in glycosylation using a glycosyl donor with a C-2 protecting group that can involve neighboring groups such as amides, esters, imides, or carbamates. About sex.

Another aspect of the invention relates to the design, construction and analysis of combinatorial libraries comprising a plurality of novel compounds of the invention, including salts and conjugates thereof, preferably those attached to a solid phase.

[0027] Other embodiments of the invention will be apparent to those skilled in the art from consideration of the present disclosure.

DETAILED DESCRIPTION OF THE INVENTION The following glossary is provided as an aid to understanding the use of terms used herein. The definitions provided are for explanation and illustration purposes only. They should not be construed narrowly or to unduly limit the scope of the invention.
The present invention is limited only by the disclosure of the prior art.

Oligosaccharide: One or more monosaccharides (or sugars) linked via glycosidic bonds
Oligomeric sugars (or carbohydrates) containing units. Oligosaccharides can be referred to as disaccharides, trisaccharides, tetrasaccharides, etc., depending on the number of monosaccharides generated during hydrolysis. As used herein, an oligosaccharide may include its conjugate (ie, another non-sugar chemical moiety, including anticancer agents, macrolides, other natural products, amino acids, peptides, nucleosides, oligonucleotides, etc.). Compounds containing one, two, three, etc. sugars covalently bonded to

Glycoside: Any sugar containing at least one pentose or hexose residue, wherein the anomeric carbon has a hydrogen-free substituent. Typically, the hydrogen-free substituent is a heteroatom, such as nitrogen, oxygen, phosphorus, silicon or sulfur.

Glycoside linkage (also referred to as a glycosidic linkage or linkage): A linkage, linkage, or linkage formed by the reaction of a sugar, such as a cyclic aldose or ketose, with an alcohol ROH to form a mixed acetal or glycoside. . If the alcohol is itself a hydroxyl group of a monosaccharide, the product is a disaccharide. The disaccharide can react in a similar manner to form higher oligosaccharides and ultimately polysaccharides. Thus, the monosaccharide units of the polysaccharide are linked via glycosidic bonds. The linkage may be formed by the reaction of C-1 or C-2 hemiacetal hydroxyl with any of the hydroxyl groups of other monosaccharides. The bond can be formed such that the anomeric carbon has either configuration. By convention, in the standard ring orientation of the β-glycoside linkage (the hemiacetal carbon atom on the extreme right hand side of the structure), the linkage is pointing up the ring plane, and in the α-glycoside linkage, the linkage is Point below the plane of the ring.

Activator: A chemical reagent that, when added to glycosyl sulfoxide, reacts with the anomeric sulfoxide group to render the anomeric carbon susceptible to nucleophilic attack. In the case of a bifunctional sugar or glycoside residue, the activator can also deprotect a blocked nucleophile under the same conditions used to activate the anomeric sulfoxide group.

Glycosyl acceptor: Any compound containing at least one nucleophilic group capable of forming a covalent bond with the anomeric carbon of a glycosyl donor under the conditions of the process of the invention. As described herein, a glycosyl receptor includes an unprotected hydroxyl, amino, or mercapto group, or a group blocked by a protecting group that can be removed in situ, ie, under the reaction conditions of the invention. , Any organic molecule, including sugars.

Glycosyl Donor: A sugar or glycoside residue having a sulfoxide group at the anomeric carbon, which group upon activation activates the anomeric carbon to be attacked by the nucleophilic group of the glycosyl acceptor, resulting in a glycosidic linkage. Is formed. A preferred glycosyl donor is phenylsulfenyl-2-deoxy-2-N-trifluoroacetamidoglycopyranose. It is important to point out that those of ordinary skill in the art will understand that donor glycosides can be monosaccharides, disaccharides, trisaccharides, and the like. Thus, chemical libraries that can possibly be produced using the methods and compounds of the present invention include all forms of oligosaccharides or conjugates or glycoconjugates thereof.

Glycoside library: An oligosaccharide of a mixture, collection, or a plurality of different sequences that can be subjected to screening methods to identify compounds or molecules that exhibit biological activity. The chemical library also contains various conjugates or glycoconjugates.

Glycoconjugate: Any compound or molecule that contains a sugar-free moiety covalently linked to a glycoside residue. See also the definition of oligosaccharides above.

Protecting group: A blocking or protecting group that can be removed in situ, preferably, but not necessarily, under the same conditions used to activate the anomeric sulfoxide group. It means a moiety commonly used in oligosaccharide synthesis to prevent hydroxyl or amino group reactions during the reaction. A preferred protecting group is a trifluoroacetyl moiety.

Blocking Group: Like protecting groups, blocking groups are used to prevent inappropriate reaction of a functional group of interest. The terms protecting group or blocking group can be used as synonyms.

The term “substantially free” refers, in a preferred embodiment of the invention, to a glycosylation product formed under the conditions of the invention, in which the presence of significant amounts of α-anomers has been excluded. Could mean.

In the present sulfoxide glycosylation method, the use of a glycosyl sulfoxide donor based on galactosamine and glucosamine is investigated. More specifically,
Develop sulfoxide donors that provide high β-selectivity. Generally, the β-selectivity of the glycosylation reaction is achieved by the use of a glycosyl donor with a C-2 protecting group that can participate in adjacent groups, including but not limited to amides, esters, imides, or carbamates. In the case of sulphoxides based on glucosamine and galactosamine, the nature of this protecting group at C-2 is important. This means that the sulfoxide should not only be reactive enough to glycosylate the relatively non-reactive nucleophiles, but the group should be easy enough to allow subsequent derivatization of the amino functionality. Because it should be removed.

In one embodiment of the process for synthesizing β-oligosaccharides, the glycosyl acceptor is converted to a glycosyl donor (e.g., phenylsulfenyl-
2-deoxy-2-N-trifluoroacetamide-3,4,6-tri-O-acetyl-β-D-glycopyranose).

Another embodiment of the present invention is a glycosyl donor, which is a phenylsulfenyl-2-deoxy-2-N-trifluoroacetamide-3,4,6-tri-O-
Acetyl-β-D-glycopyranose). These donors have been found to give exclusively β-glycosides in high yield.
Thereafter, the trifluoroacetamide is removed under mild conditions and the resulting 2-amino group is selectively derivatized as an amide. Thus, this protecting group, which is newly used in this chemistry, provides flexibility to the sulfoxide glycosylation process.

In yet another embodiment of the present invention: a) reacting glycosamine hydrochloride with p-methoxybenzaldehyde in the presence of an alkali to form 2-Np-methoxy-benzylideneglycosamine. B) acetylating 2-Np-methoxybenzylideneglycosamine with acetic anhydride in the presence of pyridine and dimethylaminopyridine (DMAP);
O-acetylated 2-N-p-methoxybenzylidene glycosamine is formed; c) The p-methoxybenzaldehyde group is removed using hydrochloric acid in acetone and O
To form an acetylated glycosamine hydrochloride; d) protecting the O-acetylated glycosamine hydrochloride with trifluoroacetic anhydride in the presence of pyridine and methylene chloride; E) subjecting the O-acetylated trifluoroacetamidoglycopyranose to thiophenol, boron trifluoride etherate and methylene chloride to give phenyl-1
-Thio-O-acetylated 2-N-trifluoroacetamido glycopyranose; f) reacting phenyl-1-thio-O-acetylated 2-N-trifluoroacetamide with m-chloroperoxybenzoic acid; The corresponding phenylsulfenyl-2-deoxy-2-N-trifluoroacetamido-3,4,6-tri-O-
G) phenylsulfenyl-2-deoxy-2-N-trifluoroacetamide-3,4,6-tri-O-, which comprises the step of producing acetyl-β-D-glycopyranose; A process for the synthesis of acetyl-β-D-glycopyranose is provided.

The preparation of the tert-butylsulphenyl equivalent can be achieved in the same manner by using a thiotert-butanol reagent instead of a thiophenol reagent. The reaction solvent plays a role in the stereoselectivity of glycosylation in the absence of neighboring group participation. When a non-polar aprotic solvent is used, the selectivity of α-glycosidic bond formation increases, while the use of a polar aprotic solvent such as propionitrile increases β
-Increase the selectivity of glycosidic bond formation.

Protecting groups on glycosyl donors also influence the stereochemical course of the glycosylation reaction. If the protecting group at the equatorial position at the C-2 center of the glycosyl donor is trifluoroacetamide, the β-glycosidic bond will be involved in the glycosylation process, regardless of whether an aprotic non-polar or aprotic polar solvent is used in the reaction. Only are formed. Many functionalities suitable for use as protecting groups for amino groups include
T. W. Greene, Protecting Group for Organic Synthesis, John Wiley & Son
s. A suitable protecting group is 9-fluorenylmethoxycarbonyl (
Carbamates such as Fmoc); and allyloxycarbonyl (Alloc)
; Including amides such or trifluoroacetamide (TFA); phthalimide (Phth) and imide such tetrachlorophthalimides (PhthCl _4). Among the many possible protecting groups used for glycosyl donors, the inventors have found that TF
A was found to provide unexpected benefits. First, the corresponding glycosyl sulfoxides were shown to be very reactive, giving high glycosylation yields in both liquid and solid phases. Second, the analysis of the reaction products gives β
The isomer was formed, indicating that there was no obvious trace of the alpha isomer. Third, the trifluoroacetamide group was deprotected under mild conditions and the corresponding amino group was derivatized as an amide.

The TFA group is easily removed by treatment with LiOH in an anhydrous mixture of 50:50 MeOH-THF. This method is compatible with the solid phase method. Under the conditions of the present invention, all common ester groups are also removed. This is not a disadvantage since the amine can be selectively converted to an amide in the presence of an unprotected alcohol.

Solution Methods for Obtaining Oligosaccharides Two general methods for obtaining oligosaccharides are: a) isolation from natural sources (this approach involves the production of large quantities of naturally occurring oligosaccharides). Sugars); and b
2.) Enzymes or chemical synthesis. Enzyme synthesis is limited because enzymes are highly specific and can only accept certain substrates. In contrast, chemical synthesis is more versatile than enzyme synthesis and has the potential to produce a vast variety of oligosaccharides. The problem with chemical synthesis is that it is extremely expensive in terms of time and labor.

[0048] Oligosaccharides are formed from monosaccharides connected by glycosidic linkages. In a typical liquid-phase chemical synthesis of oligosaccharides, a fully protected glycosyl donor is activated and reacted in solution with a glycosyl acceptor, typically another monosaccharide having an unprotected hydroxyl group. . The glycosylation reaction itself depends on the method used,
It can be done anywhere, in minutes to days. Thereafter, the coupling product is purified and chemically modified to convert it to a glycosyl donor. Each purification is time consuming and can lead to considerable material loss. The new glycosyl donor, disaccharide, is then coupled to another glycosyl acceptor. Thereafter, the product is isolated and chemically modified as described above. It is not uncommon for the synthesis of trisaccharides to require 10 or more steps from the component monosaccharide. Thus, the time and expense involved in oligosaccharide synthesis has been a serious obstacle to the development of carbohydrate drugs and other biomedical products.

(Solid Phase Synthesis of Oligosaccharides) The solid phase synthesis of oligosaccharides does not require isolation and purification.

[0050] Frechet and Schuerch, J. et al. Am. Chem. Soc. , 9
3: 492-496 (1971), solid phase synthesis of oligosaccharides comprises: a) the use of a sugar derivative having a reactive leaving group at C1; A resin that can separate the hydroxyl group; c) the remaining hydroxyl protected by a stable blocking group; and d) the formed oligosaccharide derivative without degradation of the product degradation.

Previous attempts to synthesize oligosaccharides on insoluble resins have resulted in low coupling yields and stereochemical control was inadequate, especially for the creation of β-glycoside linkages.
This is clearly because the reaction kinetics on the solid phase is slower than in the liquid phase. E
by and Schuerch, Carbohyd. Res. , 39: 151-1
55 (1975). As a result, factors such as stereochemical control and yield are affected. Frechet and Schuerch describe two glycosylation reactions (
It is found that both involve the displacement of anomeric halide in the presence of a catalyst), but preferentially give β-anomers in solution, but give mixtures on solid phase. It was concluded that the formation of β-glycoside linkages on the solid phase would require the use of adjacent group participation.

However, it has been found that existing glycosylation methods cannot be applied to solid phases. This is because adjacent participating groups (NPGs) often inactivate glycosyl donors. Often, the glycosyl donor is broken down in the resin mixture before glycosylation occurs. See Eby and Schuerch, supra. In some cases, resins are also known to degrade due to the harsh conditions required for glycosylation. In addition, many ester-type NPGs have significant problems with acyl transfer from a glycosyl donor to a glycosyl acceptor on the resin. This side reaction covers the resin and prevents further reaction.

Frechet reviewed the issues that arise in attempting to implement a strategy for solid phase oligosaccharide synthesis. Frechet, Polymer Supported Reactions in Organic Synthesis, p. 407, p. Hodge and D.M. C. Sherrington edition, Jo
hn Wiley & Sons, 1980.

[0054] Soluble resin was used to overcome the undesirable reaction kinetics associated with the solid phase reaction. Douglas et al. J. Am. Chem. Soc. , 1
13: 5095 (1991) use a soluble polyethylene glycol resin with a succinic acid linker and use the Koenigs-Knoll reaction to achieve 85-95% coupling yields with excellent control of anomeric stereochemistry. Achieved. Certain glycosylation reactions may be advantageous because soluble resins provide a more "solution-like" environment. However, stepwise synthesis on a soluble polymer requires that the intermediate be precipitated after each step and crystallized before another sugar residue can be coupled.

Thus, there is a need for a glycosylation method that provides for rapid, efficient, and high yield solid phase synthesis of oligosaccharides.

Glycoside Linkage Formation in Solution Glycosyl donors with alkyl or aryl sulfoxides at the anomeric position, and one or more free hydroxyl and / or other nucleophilic groups (eg, amines) and / or silyl A glycosyl acceptor having an ether-protected hydroxyl is combined in the reaction vessel.

The glycosyl donor is blocked at the C-2 position by a suitable protecting group such as TFA,
A 1,2-transglycoside linkage is obtained.

The mixture of glycosyl donor and acceptor can be prepared under anhydrous conditions using a non-nucleophilic solvent, including toluene, ether, tetrahydrofuran (THF), methylene chloride, chloroform, propionitrile, ethyl acetate or a mixture thereof. Is not limited to this). The choice of solvent was found to affect the stereochemical consequences of glycosylation in reactions without neighboring group involvement. In general, for a given donor / acceptor pair, the use of a non-polar solvent, such as toluene, results in the formation of a higher proportion of α-isomers, while the use of a more polar solvent, such as propionitrile. By
The β-anomer is formed in a higher proportion.

Further, in another embodiment of the present invention, a mixture of chemically diverse oligosaccharide or glycoconjugate products is produced for the creation of a library, which can be screened for biological activity. In order to do so, it may be desirable to include several different types of sugars in the reaction mixture.

Glycosidic Linkage Formation on Solid Phase Glycosyl receptors can be readily cleaved at the end of synthesis using conditions that do not damage the glycosidic linkage on the insoluble support (hereinafter referred to as resin). Attach through the chain. The resin can be any insoluble polymer that swells in an organic solvent and has a glycosyl receptor attachment site. Suitable resins are polystyrene resins, such as Merrifield resin, and PEG-derivatized polystyrene resins,
Examples include, but are not limited to, TentaGel® resin. The type of linkage depends on the type of functional group available on the polymer phase and on the glycosyl acceptor.

A glycosyl receptor can be any molecule having one or more reactive nucleophiles, including reactive hydroxyls, amines and / or thiols, provided that it also has suitable sites for resin attachment . Reactive nucleophiles are free nucleophiles or nucleophiles with temporary protecting groups that can be easily removed once the glycosyl receptor is attached to the resin. Glycosyl receptors may have permanent protected nucleophiles, which are nucleophiles that cannot be deprotected under the conditions used to remove the temporary protecting groups. Glycosyl receptors can also be sugars or molecules with some other nucleophiles, including but not limited to steroids, amino acids or peptides, polar lipids, polycyclic aromatic compounds, macrolides, natural products, etc. May be. Suitable receptors (13-15) for immobilization on link amide polystyrene are shown in FIG. Sugar protecting group schemes that allow selective protection and deprotection at any position are known. See, for example, Brinkley, Modern Carbohydrate Chemistry, Marcel Dekker, Inc. : New York (198
8).

After attaching the potentially reactive nucleophile to the resin, it is optionally deprotected, if necessary.

[0063] All cited patents, publications and literature are incorporated herein by reference. The following specific examples are provided to assist the reader in various aspects of practicing the invention. These specific examples are merely illustrative, and nothing in the following should be construed as limiting the invention in any way.

Example Synthesis of Glycosyl Sulfoxide Donor Phenylsulfenyl 2-deoxy-2-N-trifluoroacetamidoglycopyranose was obtained from commercially available glucosamine and galactosamine hydrochlorides, as shown in FIGS. Synthesized from Both syntheses are performed on a multigram scale. The corresponding sulfoxide can be easily synthesized from commercially available sugars and is stable for several weeks at room temperature.

Example 1 Synthesis of phenylsulfenyl-2-deoxy-2-N-trifluoroacetamido-3,4,6-tri-O-acetyl-β-D-glucopyranose (6) 2-Np- Methoxybenzylidene-β-D-glucosamine (1) The method is described in Bergman, M, and Zervas, L .; Chem. Ber
. Adopted from 1931 and 975. Glucosamine hydrochloride (50 g;
(0.232 mol) is dissolved in 240 mL of 1 M aqueous sodium hydroxide to form a colorless solution. Anisaldehyde (28.5 mL; 0.235 mol) is added via syringe with vigorous stirring to form a cloudy solution. After stirring for a few minutes, a white precipitate forms. The system is left in an ice bath for 1 hour to completely settle. The solid is then collected by filtration and washed successively with water (2 × 200 mL) and a 1: 1 mixture of methanol and ether (2 × 200 mL). The precipitate is dried under vacuum overnight to give 1 (50 g; 72% yield). ¹ H NMR (3
00 MHz, DMSO-d ₆ ): δ 8.17 (s, 1H), 7.74 (d, 2H)
, J = 8.1 Hz), 7.04 (d, 2H, J = 7.8 Hz), 6.60 (d,
1H, J = 6.6 Hz), 4.99 (d, 1H, J = 4.5 Hz), 4.88 (
d, 1H, J = 4.8 Hz), 4.75 (d, 1H, J = 7.2 Hz), 4.6
2 (t, 1H, J = 5.4 Hz), 3.85 (s, 3H), 3.69 (dd, 1
H, J = 5.4, 11.1 Hz), 3.58-3.42 (m, 2H), 3.32
-3.16 (m, 2H), 2.85 (t, 1H, J = 8.7 Hz). ¹³ CN
MR (75.4 MHz, DMSO-d ₆ ): δ161.24, 161.06, 1
29.65, 129.11, 113.91, 95.64, 78.21, 76.8
8, 74.61, 70.36, 61.27, 55.29. C ₁₄ H ₁₉ NO ₆ (
297.31) Elemental analysis: C, 56.56%; H, 6.44%; N, 4.71.
%. Found: C, 55.97%; H, 6.38%; N, 4.56%. Melting point 148
１５０150 ° C. (decomposition).

2-deoxy-2-Np-methoxybenzylidene-1,3,4,6-tetra-
O-acetyl-β-D-glucopyranose (2) 1 (50 g; 0.168 mol) was added to acetic anhydride (150 mL;
59 mol), pyridine (270 mL; 3.34 mol) and DMAP (0.5 g)
). The solid slowly goes into solution and the reaction mixture is left at room temperature overnight
. When the solution is poured onto 1.5 L of ice, a white crystalline solid is formed. Crystals for filtration
Collected and washed with water (2 × 100 mL) and ether (2 × 100 mL),
Drying under vacuum gives 2 (60 g; 77% yield). TLCR_f=
0.45 (50% ethyl acetate-hexane).¹1 H NMR (300 MHz, CD
Cl₃): Δ 8.15 (s, 1H), 7.64 (d, 2H, J = 8.4 Hz),
6.91 (d, 2H, J = 8.7 Hz), 5.94 (d, 1H, J = 8.1 Hz)
), 5.42 (t, 1H, J = 9.3 Hz), 5.14 (t, 1H, J = 9.6)
Hz), 4.37 (dd, 1H, J = 4.5, 12.3 Hz), 4.12 (dd
, 1H, J = 2.1, 12.6 Hz), 3.97 (ddd, 1H, J = 2.4,
4.8, 9.6 Hz), 3.84 (s, 3H), 3.44 (t, 1H, J = 9.
6 Hz), 2.10 (s, 3H), 2.03 (s, 3H), 2.01 (s, 3H)
), 1.88 (s, 3H).¹³C NMR (75.4 MHz, CDCl₃):
δ 170.69, 169.89, 169.54, 168.77, 164.27,
162.26, 130.22, 128.24, 114.02, 93.12, 73
. 20, 72.91, 72.72, 67.98, 61.78, 55.39, 20
. 79, 20.67, 20.49 (br). IR (neat, cm^-1): 294
8, 2869, 1752, 1640, 1606, 1508, 1372, 1223
, 1039. C₂₂H₂₇NO₁₀FAB + of [MH] calculated m / z 466;
Found m / z 466; [MNa] Calculated m / z 488, Found m / z 488. C ₂₂ H₂₇NO₁₀Elemental analysis for (465.46): C, 56.77%;
85%; N, 3.01%. Found: C, 56.56%; H, 5.90%; N, 2
. 99%. Melting point 168-172 [deg.] C (decomposition).

2-Deoxy-2-amino-1,3,4,6-tetra-O-acetyl-β-D-
Glucopyranosyl hydrochloride (3) 2 (50 g; 0.108 mol) is dissolved in 250 mL of refluxing acetone and 25 mL of 5N HCl is added dropwise to this solution. After 5 minutes, a thick white precipitate forms and the system is cooled to room temperature. Filter the precipitate and wash with acetone (100 mL) and ether (2 × 250 mL). Drying the crude product 3 under vacuum overnight gives 41.8 g (quant.). ¹ H NMR (300 MHz, DMS
_{O-d 6): δ8.93 (} s, br, 2.6H), 5.97 (d, 1H, J = 8
. 7 Hz), 5.42 (t, 1H, J = 9.9 Hz), 4.99 (t, 1H, J
= 9.3 Hz), 4.25 (dd, 1H, J = 3.9, 12 Hz), 4.11-
4.03 (m, 2H), 3.62 (t, 1H, J = 9.3 Hz), 2.23 (s
, 3H), 2.09 (s, 3H), 2.05 (s, 3H), 2.03 (s, 3H)
). ¹³ C NMR (75.4 MHz, DMSO-d ₆ ): δ 169.98, 1
69.78, 169.32, 168.67, 90.08, 71.59, 70.3
1, 67.76, 61.25, 52.11, 20.97, 20.88, 20.5
1, 20.37. IR (neat, cm ^-1 ): 2805, 2745, 2683,
1757, 1595, 1519, 1366, 1247, 1208, 1084, 1
060, 1040. C ₁₄ H ₂₇ NO ₉ Cl in FAB +: [M-Cl] calcd m
/ Z 348, found m / z 348; [MNa-HCl] calculated m / z 370, found m / z 370. C ₁₄ H ₂₂ NO ₉ elemental analysis of Cl (383.78): C,
43.81%; H, 5.78%: N, 3.65%; Cl, 9.24%. Measured value:
C, 43.80%; H, 5.80%; N, 3.57%; Cl, 9,15%. Melting point> 200 ° C.

2-deoxy-2-N-trifluoroacetamide-1,3,4,6-tetra-
O-acetyl-β-D-glucopyranose (4) 3 (41.6 g; 0.108 mol) was added to pyridine (90 mL; 1.11 mol).
And suspended in methylene chloride (90 mL). Trifluoroacetic anhydride (18.5)
mL; 0.131 mol) is added slowly via syringe. The solid slowly goes into solution with a slight increase in temperature. The reaction mixture is concentrated in vacuo and dried. The residue was dissolved in 100 mL of methylene chloride and 2N HCl (1 × 100 mL)
), Washed with aqueous NaHCO ₃ (2 × 100 mL) and brine (1 × 50 mL), dried over anhydrous _Na 2 SO _4. The clear solution is concentrated and dried to give off-yellow solid 4 (48.5 g; quantitative). TLC R _f = 0.47 (
50% ethyl acetate-hexane). ¹ H NMR (300 MHz, CDCl ₃ ):
δ 7.24 (d, 1H, J = 9.0 Hz), 5.75 (d, 1H, J = 9.0 H)
z), 5.31 (t, 1H, J = 10.0 Hz), 5.13 (t, 1H, J = 9)
. 6 Hz), 4.35 (q, 1H, J = 9.9 Hz), 4.27 (dd, 1H,
J = 4.8, 12.6 Hz), 4.15 (dd, 1H, J = 2.1, 12.6H)
z), 3.90 (ddd, 1H, J = 2.1, 4.8, 9.9 Hz), 2.12
(S, 3H), 2.10 (s, 3H), 2.06 (s, 3H), 2.05 (s,
3H). ¹³ C NMR (75.4 MHz, CDCl ₃ ): δ 171.67, 1
70.67, 169.43, 169.34, 157.50 (J = 38 Hz), 1
15.51 (J = 286 Hz), 91.78, 72.95, 72.00, 67.
76, 61.54, 53.15, 20.67-20.37 (br). IR (neat, cm- ¹ ): 3326, 3100, 2952; 1748, 1560, 137
4, 1219, 1079, 1042. FAB + for C ₁₆ H ₂₀ NO ₁₀ F ₃ :
MNa] Calculated m / z 466, found m / z 466. C ₁₆ H ₂₀ NO ₁₀ F elemental analysis of _{3 (443.33): C, 43.35} %; H, 4.55%; N, 3.1
6%. Found: C, 43.16%; H, 4.51%; N, 3.15%.

Phenyl-1-thio-2-deoxy-2-N-trifluoroacetamido-3,
4,6-Tri-O-acetyl-β-D-glucopyranose (5) 4 (48.25 g; 0.109 mol) was dissolved in 400 mL of anhydrous methylene chloride.
Sulfur, thiophenol (17 mL; 0.166 mol) and boron trifluoride
Treat with rate (42 mL; 0.331 mol). Leave the reaction mixture at room temperature overnight
And then 100 mL of saturated NaHCO₃Water, 100 mL of Na₂CO₃water,
And pour into 50 mL of saline solution. The organic layer was further washed with 50 mL of saturated NaHC
O₃Water and 50 mL of Na₂CO₃Wash with a mixture of water. The organic layer was dried over anhydrous Na ₂ SO₄And concentrated under vacuum. The obtained solid is quenched with 300 mL boiling water.
Wash with sun and filter. The filtrate was further washed with 300 mL of ice water hexane,
Drying in the air gives 5 (49.2 g; 92% yield). TLCR_f=
0.54 (50% ethyl acetate-hexane).¹1 H NMR (300 MHz, CD
Cl₃): Δ 7.51-7.48 (m, 2H), 7.33-7.26 (m, 3H)
), 7.02 (d, 1H, J = 9.3 Hz), 5.28 (t, 1H, J = 9.9)
Hz), 5.03 (t, 1H, J = 9.6 Hz), 4.78 (d, 1H, J = 1
0.2Hz), 4.22-4.13 (m, 2H), 4.08 (q, 1H, J = 1
0.2Hz), 3.78 (m, 1H), 2.08 (s, 3H), 2.00 (s,
3H), 1.93 (s, 3H).¹³C NMR (75.4 MHz, CDCl₃ ): Δ 168.32, 167.55, 166.15, 154.03 (q, J = 3
8 Hz), 130.38, 128.02, 125.95, 125.68, 112
. 46 (q, J = 288 Hz), 82.87, 72.82, 70.23, 65.
13, 59.12, 50.12, 17.64, 17.34, 17.29. IR (
Neat, cm^-1): 3302, 3103, 2951, 2879, 1748, 1
706, 1557, 1371, 1217, 1178, 1077, 1037. C_{2 0} H₂₂NO₈F₃FAB +: [MNa] calculated m / z 516, measured m / z
516. Melting point 178-180 [deg.] C (decomposition).

Phenylsulfenyl-2-deoxy-2-N-trifluoroacetamido-3
, 4,6-Tri-O-acetyl-β-D-glucopyranose (6) 4 (49 g; 0.099 mol) in 500 mL of CH₂Cl₂Melted into -78
Cool to ° C. Sodium bicarbonate (0.5 g) and mCPBA (26.3 g;
68.7% pure: 0.104 mol) and slowly raise the temperature to -25 ° C.
Let As the reaction proceeds, the product precipitates out of solution. Reaction by TLC
Quenched with 1 mL dimethyl sulfide and allowed to reach room temperature
You. The reaction mixture was washed with 200 mL of water, 200 mL of NaHCO₃Water and 100m
L CH₂Cl₂Dilute with. The organic layer was washed with 100 mL of Na₂CO₃Water and 10
Wash with a mixture of 0 mL saline and dry₂SO₄And concentrated under vacuum
I do. Wash the solid with 300 mL hot ether, filter and add another 300 mL ice
Wash with cold ether and dry under vacuum to give 5 (48 g; 95% yield).
It is. TLCR_f= 0.15-0.26 (50% ethyl acetate-hexane).¹ 1 H NMR (300 MHz, CDCl₃5: 1 ratio of diastereoisomers): δ
8.37 (d, 0.16H, J = 7.8 Hz), 7.83 (d, 0.84H, J
= 8.7 Hz), 7.68-7.41 (m, 5H), 5.65 (t, 0.16H)
, J = 9.9 Hz), 5.45 (t, 0.84 H, J = 9.6 Hz), 5.02
(T, 0.16H, J = 9.9Hz), 4.96-4.88 (m, 1.84Hz
), 4.21-3.94 (m, 3H), 3.81 (ddd, 0.84H, J = 2)
. 7, 3.9, 10.2 Hz), 3.68 (ddd, 0.16H, J = 2.4,
6.0, 9.6 Hz), 2.22-1.93 (m, 9H).¹³C NMR (7
5.4 MHz, CDCl₃5: 1 ratio of diastereoisomers): δ 170.76
, 170.42, 169.33, 157.27 (q, J = 38 Hz), 137.
42, 137.00, 131.970, 131.88, 129.08, 125.
60, 125.40, 115.23 (q, J = 288 Hz), 92.03, 88
. 52, 76.75, 76.28, 72.30, 71.53, 68.23, 67
. 58, 61.81, 61.33, 51.51, 50.17, 20.60, 20
. 49, 20.37. IR (neat, cm^-1): 3232, 3070, 295
5, 1750, 1372, 1222, 1182, 1110, 1037. C₂₀H ₂₇ NSO₉F₃FAB +: [MNa] calculated value m / z 532, measured value m / z 5
32. C₂₀H₂₂NSO₉F₃Elemental analysis for (509.45): C, 47.15
%; H, 4.35%; N, 2.75%; S, 6.29%. Found: C, 47.0
H, 4.34%; N, 2.70%; S, 6.21%. 134-140
° C (decomposition).

FIG. 1 shows a synthetic scheme leading to compound (6).

Example 2 Phenylsulfenyl-2-deoxy-2-N-trifluoroacetoa
Mido-3,4,6-tri-O-acetyl-β-D-galactopyranose (12)
2-Np-methoxybenzylidene-β-D-galactosamine (7) The method was described by Bergman, M, and Zervas, L. Chem. Ber
. Adopted from 1931 and 975. Galactosamine hydrochloride (5.0
g; 215.64 g / mol; 23.2 mmol) in 24 mL of 1 M sodium hydroxide.
When dissolved in water (24 mmol), a colorless solution is formed. Anisaldehyde
(2.85 mL; 3.2 g; 136 g / mol; ρ1.119; 23.5 mol)
Upon addition via syringe with vigorous stirring, a cloudy solution is formed. For a few minutes
After stirring, a white precipitate forms. Place the system in an ice bath for 1 hour to completely precipitate
Let it. Thereafter, the solid was collected by filtration and combined with water (100 mL) and methanol.
Wash with a 1: 1 mixture of ethers (2 × 25 mL). Dry the precipitate overnight under vacuum
Drying gives 7 (4.5 g; 297 g / mol).¹1 H NMR (30
0 MHz, DMSO-d₆): Δ 8.18 (s, 1H), 7.73 (d, 2H,
J = 8.7 Hz), 7.04 (d, 2H, J = 8.7 Hz), 6.52 (d, 1
H, J = 6.9 Hz), 4.72-4.67 (m, 2H), 4.61 (d, 1H)
, J = 6.9 Hz), 4.53 (d, 1H, J = 4.5 Hz), 3.85 (s,
3H), 3.73 (t, 1H, J = 3.9 Hz), 3.67-3.49 (m, 4
H), 3.15 (t, 1H, J = 7.8 Hz).¹³C NMR (75.4 MH
z, DMSO-d₆): 161.21, 160.97, 129.54, 129.
20, 113.88, 96.15, 75.15, 74.51, 71.62, 67
. 18, 60.79, 55.28. C₁₄H₁₉NO₆ES-MS of [MH] ⁺ M / z 298, m / z 298; [M-H]⁻Calculated value m / z 296
, Found 296. C₁₄H₁₉NO₆Elemental analysis for (297.31): C, 56.
56%; H, 6.44%; N, 4.71%. Found: C, 55.46%; H, 6
. 42%; N, 4.58%.

2-Deoxy-2-Np-methoxybenzylidene-1,3,4,6-tetra-
O-acetyl-β-D-galactopyranose (8) 7 (4.5 g; 83% pure; 297 g / mol; 13 mmol) was treated with acetic anhydride (15 mL; 102.09 g / mol; 082; 159 mmol)
, Pyridine (79.10 g / mol; ρ0.978; 27 mL; 334 mmol)
And DMAP (0.05 g). The solid slowly goes into solution and the reaction mixture is left at room temperature for 3 hours. Upon pouring the solution into 200 mL of ice, a white crystalline solid is formed. The crystals were collected by filtration, and water (100 mL) and ether (
100 mL) and dried under vacuum to give 8 (3.8 g; 465 g / mol; 63% yield). TLC _Rf = 0.71 (60% ethyl acetate-hexane). ¹ H NMR (300 MHz, CDCl ₃ ): δ 8.21 (s, 1H)
, 7.65 (d, 2H, J = 9.0 Hz), 6.92 (d, 2H, J = 8.4 H)
z), 5.92 (d, 1H, J = 8.1 Hz), 5.45 (d, 1H, J = 3.
3 Hz), 5.25 (dd, 1H, J = 3.3, 10.5 Hz), 4.22-4
. 14 (m, 3H), 3.84 (s, 3H), 3.61 (dd, 1H, J = 8.
4, 10.5 Hz), 2.17 (s, 3H), 2.06 (s, 3H), 2.02
(S, 3H), 1.88 (s, 3H). ¹³ C NMR (75.4 MHz, CD
Cl ₃ ): δ 170.45, 170.12, 169.69, 168.75, 16
4.47, 162.17, 130.16, 128.36, 11.97, 93.4
5, 93.45, 71.71, 71.52, 68.78, 65.90, 61.2
9, 55.41, 20.71-20.54 (m). ES of C ₂₂ H ₂₇ NO ₁₀
-MS: [M-H] ⁺ calculated m / z 466, found m / z 466; [M + OAc
^- Calculated m / z524, Found m / z524.

2-Deoxy-2-amino-1,3,4,6-tetra-O-acetyl-β-D-
Galactopyranosyl hydrochloride (9) 8 (7.3 g; 465 g / mol; 15.7 mmol) is dissolved in 37 mL of refluxing acetone, and to this solution 3.65 mL of 5N HCl is added dropwise. After 5 minutes, a thick white precipitate forms and the system is cooled to room temperature. Filter the precipitate and wash with acetone (30 mL) and ether (60 mL). If more solids are separated from this filtrate, it will integrate with the solids obtained from the first filtrate. The combined products are dried under vacuum overnight to give 5.8 g of crude 9 (95% yield). ¹ H NMR (300 MHz, DMSO-d ₆ ): δ 8.81 (s, br, 2
. 46H), 5.95 (d, 1H, J = 8.7 Hz), 5.36-5.33 (m
, 2H), 4.35 (t, 1H, J = 6.3 Hz), 4.12-4.073 (m
, 2H), 3.48 (br, 1H), 2.22 (s, 3H), 2.18 (s, 3H)
H), 2.06 (s, 3H), 2.05 (s, 3H). ¹³ C NMR (75.
_{4MHz, DMSO-d 6):} 169.98,169.89,169.33,1
68.65, 90.258, 71.04, 68.80, 65.76, 61.18
, 49.33, 20.85, 20.88, 20.70, 20.53, 20.36
. C ₁₄ H ₂₇ NO ₉ Cl of ES-MS: [M-Cl ] + calcd m / z348,
Found m / z348; [M-H ] - calcd m / z 382 and 384, found m
/ Z 382 and 384.

2-Deoxy-2-trifluoroacetamide-1,3,4,6-tetra-O
-Acetyl-β-D-galactopyranoose (10) 9 (5.6 g; 383.5 g / mol; 14.6 mmol) was converted to pyridine (15
mL; 14.7 g; 79.1 g / mol; ρ 0.978; 185 mmol) and
Suspend in methylene chloride (15 mL). Trifluoroacetic anhydride (2.5 mL; 3
. 7g; 210.3 g / mol; ρ 1.487; 17.7 mmol) slowly
Add via syringe. Solids slowly go into solution with a slight increase in temperature
. The reaction mixture is concentrated in vacuo and dried. Residue in methylene chloride (50 mL)
Dissolve, 2N HCl (2 × 50 mL), NaHCO₃Water (50 mL) and food
Wash with brine (20 mL) and dry with anhydrous Na₂SO₄And dry. Concentrate clear solution
To dry off-yellow solid 10 (5.6 g; 443 g / mol; 12
. 6 mmol; 87% yield). TLCR_f= 0.36 (40% acetic acid
Ethyl-hexane).¹1 H NMR (300 MHz, CDCl₃): Δ7.15
(D, 1H, J = 9.6 Hz), 5.77 (d, 1H, J = 8.7 Hz), 5.77.
40 (d, 1H, J = 2.7 Hz), 5.17 (dd, 1H, J = 3.3, 11
. 1 Hz), 4.48 (q, 1H, J = 9.3 Hz), 4.19-4.14 (m
, 2H), 4.07 (q, 1H, J = 6.6 Hz), 2.19 (s, 3H), 2
. 13 (s, 3H), 2.05 (s, 3H), 2.01 (s, 3H).¹³C
NMR (75.4 MHz, CDCl₃): Δ 170.81, 170.64, 17
0.14, 169.61, 157.66 (q, J = 38 Hz), 115.52 (
J = 286 Hz), 92.22, 71.97, 69.84, 67.92, 66.
10, 61.33, 50.152, 20.63-20.41 (m). C₁₆H_{2 0} NO₁₀F₃ES-MS: [M + NH₄]⁺Calculated value m / z 461, measured value m
/ Z461; [M-H]⁻Calculated value m / z 442, measured value m / z 442. C₁₆H ₂₀ NO₁₀F₃Elemental analysis for (443.33): C, 43.35%; H, 4.5
5%; N, 3.16%. Found: C, 43.34%; H, 4.60%;
11%.

Phenyl-1-thio-2-deoxy-2-N-trifluoroacetamido-3
, 4,6-Tetra-O-acetyl-β-D-galactopyranoose (11) 10 (5.55 g; 443 g / mol; 12.5 mmol) was added to anhydrous methyl chloride
Thiophenol (2.0 mL; 110.15 g / mol)
1.073; 19.5 mmol) and boron trifluoride etherate (4.8).
mL; 141.93 g / mol; ρ1.120; 37.9 mmol).
The reaction mixture was left at room temperature overnight, after which 25 mL of methylene chloride, 15 mL of
Saturated NaHCO₃Water, 15 mL of Na₂CO₃To a mixture of water and 15 mL of saline
pour it up. The organic layer was washed with an additional 15 mL of saturated NaHCO₃Water and 15 mL of Na₂C
O₃Wash with a mixture of water. The organic layer was dried over anhydrous Na₂SO₄And concentrated under vacuum
Shrink. Wash the resulting solid with 50 mL boiling hexane and filter. Remove the filtrate
After washing with 50 mL of ice-water hexane and drying in vacuo, 11 (5.75 g) was obtained.
493 g / mol; 11.7 mmol; 93% yield). TLCR_f = 0.35 (40% ethyl acetate-hexane).¹1 H NMR (300 MHz, C
DCI₃): Δ 7.53-7.50 (m, 2H), 7.34-7.31 (m, 3
H), 6.63 (d, 1H, J = 9.0 Hz), 5.39 (d, 1H, J = 3.0 Hz).
0 Hz), 5.20 (dd, 1H, J = 3.3, 10.8 Hz), 4.87 (d
, 1H, J = 10.2 Hz), 4.30-4.10 (m, 3H), 3.96 (t
, 1H, J = 6.3 Hz), 2.13 (s, 3H), 2.040 (s, 3H),
1.97 (s, 3H).¹³C NMR (75.4 MHz, CDCl₃): Δ1
70.63, 170.50, 170.10, 157.24 (q, J = 37 Hz)
, 132.87, 131.71, 129.00, 128.48, 115.48 (
q, J = 286 Hz), 86.43, 74.60, 70.76, 66.60, 6
1.69, 50.05, 20.65-20.41 (br). C₂₀H₂₂NSO ₈ F₃ES-MS: [M + NH₄]⁺Calculated value m / z 511, measured value m / z 51
1; [MH]⁻Calculated value m / z 492, measured value m / z 492. C₂₀H₂₂NS
O₈F₃Elemental analysis for (493.45): C, 48.68%; H, 4.49%; N
, 2.84%; S, 6.50%. Found: C, 48.43%; H, 4.37%;
N, 2.75%; S, 6.40%.

Phenylsulfenyl-2-deoxy-2-N-trifluoroacetamide-3
, 4,6-Tri-O-acetyl-β-D-galactopyranoose (12) 11 (5.85 g; 493 g / mol; 11.9 mmol) in 50 mL of CH ₂ Cl₂And cooled to -78 ° C. Sodium bicarbonate (0.1 g) and
mCPBA (3.13 g; 68.7% pure: 172.57 g / mol; 12.5 mi
) And the temperature is slowly raised to -25 ° C. As the reaction progresses
The product precipitates out of solution. If the reaction is determined to be complete by TLC
Quench with 1 mL dimethyl sulfide and allow to reach room temperature. The reaction mixture is
mL NaHCO₃Water and 50 mL CH₂Cl₂Dilute with. 50 organic layers
mL of Na₂CO₃Wash with a mixture of water and 10 mL of brine and dry with anhydrous Na₂S
O₄And concentrated under vacuum. Wash the solid with 75 mL hot ether and filter
Wash with an additional 100 mL of ice-cold ether and dry under vacuum to give a single
12 as diastereoisomer (4.75 g; 509 g / mol; 9.3 mmol
; 78% yield). TLCR_f= 0.31 (60% ethyl acetate-hex
Sun).¹1 H NMR (300 MHz, CDCl₃): Δ 7.72-7.53 (
m, 6H), 5.40 (dd, 1H, J = 3.3, 10.8 Hz), 5.34 (
d, 1H, J = 3.3 Hz), 4.84 (d, 1H, J = 10.5 Hz);
34 (q, 1H, J = 10.2 Hz), 4.09-3.94 (m, 3H), 2.
00-1.96 (m, 9H).¹³C NMR (75.4 MHz, CDCl₃)
: Δ170.33, 170.27, 169.83, 157.46 (J = 38 Hz
), 138.01, 131.85, 128.95, 125.57, 115.23.
(J = 286 Hz), 92.33, 74.95, 69.74, 66.21, 61
. 09, 46.80, 20.60-20.39 (br). C₂₀H₂₇NSO₉ F₃ES-MS of [M + H]⁺Calculated value m / z 510, measured value m / z 510;
M + NH₄]⁺Calculated value m / z 527, measured value m / z 527; [M-H]⁻Calculated value
m / z 508, found m / z 508. C₂₀H₂₂NSO₉F₃(509.45
)): C, 47.15%; H, 4.35%; N, 2.75%; S, 6.
29% found: C, 47.35%; H, 4.50%; N, 2.63%; S, 6.
10%. Melting point 116-118 [deg.] C.

FIG. 2 shows a synthetic scheme leading to compound (12).

Example 3 Synthesis of β-Disaccharide or Glycoconjugate on Solid Phase In a typical glycosylation method using a phenylsulfenyl-2-deoxy-2-N-trifluoroacetamidoglycopyranosyl donor, The glycosyl receptor immobilized on Linkamide resin and containing free hydroxyl groups is dried under high vacuum and then placed under argon. The resin is combined with a glycosyl donor (4 eq) and 2,6-di-t-butyl in a solvent system compatible with sulfoxide glycosylation reaction conditions (generally a 9: 1 mixture of anhydrous methylene chloride and anhydrous ethyl acetate). Add a solution of -4-methyl-pyridine (2 equivalents). The mixture is stirred at room temperature for 5 minutes and then cooled to -78C. Trifluoromethanesulfonic anhydride (4
Eq) is added slowly and the system is kept at -70 ° C for 1 hour. Thereafter, the reaction mixture is
Maintain at -45 ° C to -40 ° C for 3-16 hours and quench with a mixture of methanol and diisopropylethylamine. The reaction mixture was warmed to room temperature and the resin was added to D
Wash with MF (3 ×), tetrahydrofuran (2 ×), methanol (2 ×), and methylene chloride (2 ×). For analytical purposes, a sample of the resin was then removed using a 30% reaction mixture of trifluoroacetic acid and methylene chloride for 30 minutes. The supernatant is evaporated to dryness, the residue is dissolved and analyzed by HPLC and LC-MS.

Glycosyl donor (6) is converted to glycosyl acceptors (13), (14) and (1)
When coupled with 5), the corresponding β-linked disaccharides (16), (17) and (17)
18) are obtained respectively. The trifluoroacetamide group provides a high degree of flexibility for sulfoxide glycosylation. Once activated, sulfoxides are highly reactive and glycosylate non-reactive nucleophiles such as the glycosyl receptor (13). The high reactivity of glycosyl donors such as (6) and (12) can be evaluated by comparing them with other studied sulfoxides.

Glycosylation with> 90% conversion of the glycosyl acceptor (13) can be achieved with 4 equivalents of the glycosyl donor (6) or (12). When using the corresponding phthalimide protected sulfoxide, up to 8 equivalents of sulfoxide must be used to achieve the same level of conversion. The use of 4 equivalents of the relevant sulfoxide containing Alloc and Fmoc protecting groups resulted in up to 10% conversion to the corresponding disaccharide. Glycosyl donors (6) have been used to successfully glycosylate acceptors (13), (14) and (15), as shown in FIG. The glycosylated products (16), (17) and (18) (FIG. 4) have>
Obtained in 90% yield (with _{TFA-CH 2} Cl 2 the reaction mixture to cleave the product from the resin, determined by analyzing it by HPLC). In each product, the stereochemistry of the glycosidic linkage is β by ¹ H NMR, indicating no trace of the α-anomer (FIG. 5). The glycosyl donor (12) also
It is shown to glycosylate receptor (13) in> 90% yield (determined by HPLC).

In the β-disaccharide products (16) and (17), the trifluoroacetamide group is shown to be completely removed by treating the resin with LiOH in a 1: 1 MeOH-THF solution. Under these conditions, benzoate and acetate groups are also removed. Selective derivatization of amino groups with carboxylic acids in the presence of unprotected alcohol is possible using HATU-DIPEA coupling conditions. The β-disaccharide product (16) undergoes a reaction sequence as shown in FIG. The identity and purity of each reaction product is determined by HPLC and LC-MS (FIG. 7)
.

Liquid-Phase Glycosylation So far, studies in the liquid-phase glycosylation chemistry of these glycosyl donors have been limited to the glycosylation of isopropanol with the glycosyl donor (6). Under standard glycosylation conditions, β-isopropoxy glycoside has 60%
Isolated in yield and characterized by mass spectroscopy and ¹ H NMR. No trace of the corresponding α-isomer is observed.

Solid Phase Library Design combinatorial libraries around 16 and 17 according to the chemistry set forth above. In the case of 16, the disaccharide core is derivatized with 8 different isocyanates and 12 different carboxylic acids (FIG. 8). In this library, the anomeric group of the acceptor sugar is either a β-thiophenyl group or an α, β-hydroxy group (lactol). These three diverse elements give rise to a combinatorial library of 192 members. In the case of 17, the disaccharide core is 6
Derivatised with two different isocyanates and eight different carboxylic acids,
A combinatorial library of 48 members is obtained (FIG. 9).

Example 4 Library Construction For each disaccharide library, the corresponding disaccharide (16
Or 17) is completely deprotected by treatment with LiOH in 1: 1 THF-MeOH. The deprotected resin is then suspended in a 4: 1 mixture of methylene chloride and tetrahydrofuran, and an aliquot of this suspension containing the RF microtag,
Dispense into Irori MicroKan®. Aliquots are calculated so that each Microcan® contains 15 mg resin (for 16) or 20 mg resin (for 17). Each Microcan® and its RF tag are scanned by Irori synthesis software and assigned a confirmation number. The library is then synthesized according to the reaction scheme shown in FIGS. The results of the LC-MS analysis are consistent with the production of the desired library compound based on its molecular weight (see FIGS. 14 and 15 below).

Once the synthesis step is complete, place the Microcan® container containing the derivatized resin in a separate test tube and treat with a 30% TFA-CH ₂ Cl ₂ reaction mixture for 30 minutes. The supernatant is then transferred to the wells of a microtiter plate and concentrated under vacuum using a Savant evaporator. The residue obtained is then reconstituted in 1 ml of DMSO and the solution is dispensed for control by LC-MS analysis, antimicrobial screen and compound storage.

An LC-MS trace of a representative product for the scaffold 16 based library is shown in FIG. The analytical results obtained from LC-MS analysis of this library are summarized in FIG.

An LC-MS trace of a representative product for the scaffold 17 based library is shown in FIG. The analytical results obtained from LC-MS analysis of this library are summarized in FIG.

The above examples are provided for illustrative purposes only and do not limit the scope or embodiments of the present invention. The invention is further described with reference to the claims that follow this specification.

[Brief description of the drawings]

FIG. 1 shows a process for synthesizing phenylsulfenyl-2-deoxy-2-N-trifluoroacetamide-3,4,6-tri-O-acetyl-β-D-glucopyranose (6). .

FIG. 2 shows phenylsulfenyl-2-deoxy-2-N-trifluoroacetamide-3,4,6-tri-O-acetyl-β-D-galactopyranose (12
2) shows the synthesis process.

FIG. 3 shows receptors 13-15 immobilized on Rink amide polystyrene resin.

FIG. 4 shows the characterized products 16-18 of the solid-phase glycosylation reaction.

FIG. 5 shows a ¹ H NMR spectrum of the reaction product showing that only β-disaccharide was formed (no trace of the corresponding α-isomer is observed).

FIG. 6 shows solid phase derivatization of 16.

FIG. 7 shows the analytical characterization of intermediates and end products in derivatization of 16 (
HPLC and LC-MS).

FIG. 8 shows the general structure and building blocks used for a combinatorial library based on 16.

FIG. 9 shows the general structure and building blocks used for a combinatorial library based on 17.

FIG. 10 shows the reaction sequence used for derivatization of 16 to generate a library.

FIG. 11 shows the reaction sequence used for derivatization of 17 to generate a library.

FIG. 12 shows LC-MS spectra of representative members of a combinatorial library generated around 16.

FIG. 13 shows analytical data obtained for a library created around 16.

FIG. 14 shows LC-MS spectra of representative members of a combinatorial library generated around 17.

FIG. 15 shows analytical data obtained for a library created around 17.

──────────────────────────────────────────────────続 き Continued on the front page (51) Int.Cl. ⁷ Identification symbol FI Theme coat ゛ (Reference) C07H 15/203 C07H 15/203 (81) Designated country EP (AT, BE, CH, CY, DE, DK, ES, FI, FR, GB, GR, IE, IT, LU, MC, NL, PT, SE), OA (BF, BJ, CF, CG, CI, CM, GA, GN, GW, ML, MR, NE, SN, TD, TG), AP (GH, GM, KE, LS, MW, SD, SZ, UG, ZW), EA (AM, AZ, BY, KG, KZ, MD, RU, TJ, TM ), AL, AM, AT, AU, AZ, BA, BB, BG, BR, BY, CA, CH, CN, CZ, DE, DK, EE, ES, FI, GB, GE, H, GM, HR, HU, ID, IL, IN, IS, JP, KE, KG, KR, KZ, LC, LK, LR, LS, LT, LU, LV, MD, MG, MK, MN, MW , MX, NO, NZ, PL, PT, RO, RU, SD, SE, SG, SI, SK, SL, TJ, TM, TR, TT, UA, UG, UZ, VN, YU, ZW (72) Inventors Sofia, Michael Jay. United States 08648 Lawrenceville, New Jersey Holly Lane 3F Term (Reference) 4C057 BB03 BB04 CC03 JJ03 JJ13 JJ24 4H055 AA01 AA02 AB29 AB81 AC01 AD30 BA30

Claims (26)

[Claims] 1. An alkylsulfenyl- or arylsulfenyl-2-amine.
A compound which is deoxy-2-N-trifluoroacetamidoglycopyranose. 2. The compound according to claim 1, which is phenylsulfenyl-2-deoxy-2-N-trifluoroacetamidoglucopyranose. 3. The compound according to claim 2, which is phenylsulfenyl-2-deoxy-2-N-trifluoroacetamido-3,4,6-tri-O-acetyl-β-D-glucopyranose. 4. The compound according to claim 1, which is phenylsulfenyl-2-deoxy-2-N-trifluoroacetamidogalactopyranose. 5. The compound according to claim 4, which is phenylsulfenyl-2-deoxy-2-N-trifluoroacetamido-3,4,6-tri-O-acetyl-β-D-galactopyranose. 6. A glycoside β-glycosidic chain comprising reacting a glycosyl acceptor with a glycosyl donor that is an alkylsulfenyl- or arylsulfenyl-2-deoxy-2-N-trifluoroacetamido glycopyranose. Process to do. 7. The process of claim 6, wherein the glycosyl donor is phenylsulfenyl-2-deoxy-2-N-trifluoroacetamidoglucopyranose. 8. The glycosyl donor is phenylsulfenyl-2-deoxy-2-N-trifluoroacetamide-3,4,6-tri-O-acetyl-β-
The process of claim 7, wherein the process is D-glucopyranose. 9. The method according to claim 6, wherein the glycosyl donor is 2-deoxy-2-N-trifluoroacetamide-3,4,6-tri-O-acetyl-β-D-galactopyranose sulfoxide. process. 10. The glycosyl donor is phenylsulfenyl-2-deoxy-2-N-trifluoroacetamide-3,4,6-tri-O-acetyl-β
The process according to claim 7, which is -D-galactopyranose. 11. The process of claim 6, wherein the linkage is substantially free of α-glycoside linkages. 12. The process according to claim 6, wherein the process is performed by a solid-state reaction. 13. The process of claim 12, wherein the glycosyl receptor is bound to a solid support. 14. The process of claim 12, wherein the glycosyl receptor is attached to the solid support via a linker arm. 15. The process of claim 12, wherein the solid support is selected from the group consisting of a polystyrene resin and a PEG-derived polystyrene resin. 16. The process according to claim 6, wherein the process is performed by a liquid phase reaction. 17. The process of claim 6, wherein the glycosyl donor has an adjacent participating group (NPG) at the C-2 position. 18. The process of claim 17, wherein said NPG is selected from the group consisting of amides, esters, imides and carbamates. 19. The glycosyl donor is activated with an effective amount of an activating agent.
The process of claim 11. 20. The process of claim 19, wherein the activator is trifluoromethanesulfonic anhydride or an acid thereof. 21. A process for the synthesis of β-oligosaccharides, comprising: a) converting a glycosyl acceptor bound to a solid support into an alkylsulfenyl- or arylsulfenyl-2-deoxy-2-N as glycosyl donor. Treating with trifluoroacetamidoglycopyranose; b) reacting a glycosyl donor with a glycosyl acceptor to yield the corresponding β-glycosylation product. 22. A process for the synthesis of phenylsulfenyl-2-deoxy-2-N-trifluoroacetamido-3,4,6-tri-O-acetyl-β-D-glycopyranose, comprising: a) glycosamine Reacting hydrochloride with p-methoxybenzaldehyde in the presence of an alkali to form 2-Np-methoxy-benzylidene glycosamine; b) converting 2-Np-methoxybenzylidene glycosamine to pyridine and Acetylation with acetic anhydride in the presence of dimethylaminopyridine (DMAP)
O-acetylated 2-N-p-methoxybenzylideneglycosamine is formed; c) p-methoxybenzaldehyde is removed using hydrochloric acid in acetone and O-
Forming an acetylated glycosamine hydrochloride; d) protecting the O-acetylated glycosamine hydrochloride with trifluoroacetic anhydride in the presence of pyridine and methylene chloride to form O-acetylated 2-N- E) subjecting the O-acetylated trifluoroacetamidoglycopyranose to thiophenol, boron trifluoride etherate and methylene chloride to give phenyl-1
-Thio-O-acetylated 2-N-trifluoroacetamido glycopyranose; f) reacting phenyl-1-thio-O-acetylated 2-N-trifluoroacetamide with m-chloroperoxybenzoic acid; The corresponding phenylsulfenyl-2-deoxy-2-N-trifluoroacetamido-3,4,6-tri-O-
Producing acetyl-β-D-glycopyranose; and g) recovering said glycopyranose. 23. The formula [Wherein, Wherein R ₃ is β-SPh or α, β-OH], a salt thereof, or a conjugate thereof. 24. The formula [Wherein, Or a salt thereof, or a conjugate thereof. 25. The formula [Wherein, Wherein R ₃ is β-SPh or α, β-OH], a salt thereof, and a conjugate thereof. 26. The compound of the formula [Wherein, The library according to claim 25, which is a compound corresponding to the above, a salt thereof, or a conjugate thereof.