Sugar Chain and Compositions Thereof and Use Thereof in Prevention and/or Treatment of Coronavirus Infection

Abstract

The present application belongs to the field of life health, and discloses a sugar chain and a composition thereof, and use in the prevention and/or treatment of coronavirus infection. The sugar chain contains any one or more of Neu5Acα2-N.sub.1Gal building blocks, and/or any one or more of xFuc-N.sub.1Gal-N.sub.1(xFuc-N.sub.1)GlcNAc building blocks, at the non-reducing end, where, x=0 or 1, and N.sub.1=1, 2, 3, 4 or 6. A glycosidic bond formed between Neu5Ac and Gal is an α2 glycosidic bond. In the xFuc-N.sub.1Gal-N.sub.1(xFuc-N.sub.1)GlcNAc building blocks, a glycosidic bond formed between any two adjacent monosaccharides is an α1 or β1 glycosidic bond. The specific building block contained at the non-reducing end of the sugar chain blocks the binding of the virus to the host, thereby blocking virus invasion and infection of the respiratory tract/lung, and achieving the specific prevention and treatment.

Claims

1. A sugar chain comprising one or more of Neu5Acα2-N1Gal building blocks, and/or one or more of xFuc-N1Gal-N1(xFuc-N1)GlcNAc building blocks, at a non-reducing end, where, x=0 or 1, and N1=1, 2, 3, 4 or 6.

2. The sugar chain according to claim 1 wherein in the xFuc-N1Gal-N1(xFuc-N1)GlcNAc building blocks, a glycosidic bond formed between any two adjacent monosaccharides is an α1 or β1 glycosidic bond.

3. The sugar chain according to claim 1 wherein in the xFuc-N1Gal-N1(xFuc-N1)GlcNAc building blocks, a glycosidic bond formed between Fuc and Gal is an α1 or β1 glycosidic bond, a glycosidic bond formed between Gal and GlcNAc is an α1 or β1 glycosidic bond, and a glycosidic bond formed between Fuc and GlcNAc is an α1 or β1 glycosidic bond.

4. The sugar chain according to any one of claim 1, wherein the xFuc-N1Gal-N1(xFuc-N1)GlcNAc building blocks comprise xFuc-N1Gal-(xFuc-N1)GlcNAcα1 or xFuc-N1Gal-(xFuc-N1)GlcNAcβ1, where, x=0 or 1, and N1=1, 2, 3, 4 or 6.

5. The sugar chain according to claim 1, wherein the xFuc-N1Gal-N1(xFuc-N1)GlcNAc building blocks comprise xFucα1-N1Galβ1-(xFucα1-N1)GlcNAcβ1, where, x=0 or 1, and N1=1, 2, 3, 4 or 6.

6. The sugar chain claim 1, wherein the Neu5Ac-Gal building blocks comprise the Neu5Acα2-6Gal or Neu5Acα2-3Gal building block, and the xFuc-N1Gal-N1(xFuc-N1)GlcNAc building blocks comprise the Lewis a, Lewis x, Lewis y or Blood Group H building block.

7. The sugar chain according to claim 1, wherein the sugar chain contains A and B building blocks at the non-reducing end, the A building block is one or more of Lewis a, Lewis x, Lewis y, and Blood Group H, and the B building block is the Neu5Acα2-6Gal and/or Neu5Acα2-3Gal building blocks.

8. (canceled)

9. The sugar chain according to claim 1, wherein the sugar chain is any one of Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-3Galβ1-3GalNAcα, Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-3GalNAcα, Neu5Acα2-3Galβ1-4GlcNAcβ1-3GalNAcα, Neu5Acα2-6Galβ1-4GlcNAcβ1-3(Neu5Acα2-6Galβ1-3GlcNAcβ1-6)GalNAcα, Galβ1-4(Fucα1-3)GlcNAcβ1-3Galβ1-3GalNAcα, Galβ1-4(Fucα1-3)GlcNAcβ1-3GalNAcα, Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-6GalNAcα, 3′-sialyllactose, 3′-sialyllactosamine, 6′-sialyllactose, 6′-N-acetyl sialyllactosamine, Galβ1-3(Fucα1-4)GlcNAcβ1-3Galβ1-4Glc, Galβ1-4(Fucα1-3)GlcNAcβ1-3Galβ1-4Glc, Fucα1-2Galβ1-4GlcNAcβ1-3Galβ1-4Glc, Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-3Galβ1-4Glc, Galβ1-4(Fucα1-3)GlcNAcβ1-6(GlcNAcβ1-3)Galβ1-4Glc, Galβ1-4(Fucα1-3)GlcNAcβ1-6(Galβ1-4GlcNAcβ1-3)Galβ1-4Glc, Fucα1-2Galβ1-4GlcNAcβ1-6(Fucα1-2Galβ1-4GlcNAcβ1-3)Galβ1-4Glc, and Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-6(Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-3)Galβ1-4Glc.

10. A complex sugar chain, wherein the complex sugar chain is a sugar chain composition containing at least two different sugar chains, and at least one sugar chain in the sugar chain composition is the sugar chain according to claim 1.

11. (canceled)

12. The complex sugar chain according to claim 10, wherein at least one sugar chain in the sugar chain composition is selected from the group consisting of sulfated fucoidan, HMOs, ginsenosides, chitosan oligosaccharide, heparin sulfate, heparin sulfate oligosaccharide, chondroitin sulfate, chondroitin sulfate oligosaccharide, dermatan sulfate, keratan sulfate, hyaluronic acid, hyaluronic acid oligosaccharide, fucoidan, fucose oligosaccharide, protein N/O-sugar chains, and exopolysaccharide from Haloarcula hispanica.

13. The complex sugar chain according to claim 10, wherein at least one sugar chain in the sugar chain composition is any one of GlcNAcβ1-6(Galβ1-3)GalNAcα, GlcNAcβ1-6[Galβ1-4(Fucα1-3)GlcNAcβ1-3]Galβ1-4Glc, GlcNAcβ1-6[Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-3]Galβ1-4Glc, Galβ1-4GlcNAcβ1-6(Fucα1-2Galβ1-4GlcNAcβ1-3)Galβ1-4Glc, Galβ1-4(Fucα1-3)Glc, Fucα1-2Galβ1-4Glc, Galα1-3(Fucα1-2)Galβ1-4GlcNAcβ1-3NAcα, GlcNAcβ1-6(GlcNAcβ1-3)GalNAcα, and Galβ1-4(Fucα1-6)GlcNAcβ1-3GalNAcα, GlcNAcβ1-6(GlcNAcβ1-3)GalNAcα.

14. The complex sugar chain according to claim 10, wherein each sugar chain in the sugar chain composition contains any one or more of the following building blocks at the non-reducing end: a) Neu5Acα2-6Gal; b) Neu5Acα2-3Gal; c) Lewis a; d) Lewis x; e) Lewis y; and f) Blood Group H.

15. The complex sugar chain according to claim 10, wherein at least one sugar chain in the sugar chain composition is exopolysaccharide from Haloarcula hispanica.

16. The complex sugar chain according to claim 10, wherein the sugar chain composition contains at least one of Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-3Galβ1-3GalNAcα, and/or Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-3GalNAcα, and/or Neu5Acα2-3Galβ1-4GlcNAcβ1-3GalNAcα, and/or Neu5Acα2-6Galβ1-4GlcNAcβ1-3(Neu5Acα2-6Galβ1-3GlcNAcβ1-6)GalNAcα, and/or Galβ1-4(Fucα1-3)GlcNAcβ1-3Galβ1-3GalNAcα, and/or Galβ1-4(Fucα1-3)GlcNAcβ1-3GalNAcα, and/or Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-6GalNAcα, and/or 3′-sialyllactose, and/or 3′-sialyllactosamine, and/or 6′-sialyllactose, and/or 6′-N-acetyl sialyllactosamine, and/or Galβ1-3(Fucα1-4)GlcNAcβ1-3Galβ1-4Glc, and/or Galβ1-4(Fucα1-3)GlcNAcβ1-3Galβ1-4Glc, and/or Fucα1-2Galβ1-4GlcNAcβ1-3Galβ1-4Glc, and/or Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-3Galβ1-4Glc, and/or Galβ1-4(Fucα1-3)GlcNAcβ1-6(GlcNAcβ1-3)Galβ1-4Glc, and/or Galβ1-4(Fucα1-3)GlcNAcβ1-6(Galβ1-4GlcNAcβ1-3)Galβ1-4Glc, and/or Fucα1-2Galβ1-4GlcNAcβ1-6(Fucα1-2Galβ1-4GlcNAcβ1-3)Galβ1-4Glc, and/or Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-6(Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-3)Galβ1-4Glc.

17. A method of preventing and/or treating coronavirus infection comprising administering to a mammal in need the sugar chain according to claim 1.

18. (canceled)

19. (canceled)

20. (canceled)

21. (canceled)

22. (canceled)

23. (canceled)

24. A composition for preventing and/or treating coronavirus infection comprising the sugar chain according to claim 1.

25. (canceled)

26. (canceled)

27. (canceled)

28. (canceled)

29. (canceled)

30. (canceled)

31. A saccharide chip for preventing and/or treating coronavirus infection, comprising the sugar chain according to claim 1.

32. A saccharide chip for preventing and/or treating coronavirus infection, comprising the complex sugar chain according to claim 10.

33. A composition for preventing and/or treating coronavirus infection comprising the complex sugar chain according to claim 10.

34. A method of preventing and/or treating coronavirus infection comprising administering to a mammal in need the complex sugar chain according to claim 10.

Description

BRIEF DESCRIPTION OF THE DRAWINGS

[0104] FIG. 1 is a diagram of structural formulas of Neu5Acα2-6Gal, Neu5Acα2-3Gal, Lewis a, Lewis x, Lewis y, and Blood Group H type II, and in the figure, 1A shows Neu5Acα2-6Gal; 1B shows Neu5Acα2-3Gal; 1C shows Lewis a; 1D shows Lewis x; 1E shows Lewis y; and 1F shows Blood Group H type II;

[0105] FIG. 2 is a .sup.1H NMR image of 6′-sialyllactose obtained by enzymatic synthesis of Example 1;

[0106] FIG. 3 is a .sup.13C NMR image of 6′-sialyllactose obtained by enzymatic synthesis of Example 1;

[0107] FIG. 4 is a mass spectrogram of 6′-sialyllactose obtained by enzymatic synthesis of Example 1;

[0108] FIG. 5 is a diagram of the toxicity of sialic acid, sialyloligosaccharides, chitosan oligosaccharide, and chondroitin sulfate of Example 2 to lung epithelial cells (BEAS-2B cells), and the in the figure, A shows sialic acid (Sia); B shows 3′-sialyllactose (3′-SL); C shows 6′-sialyllactose (6′-SL); D shows 6′-N-acetyl sialyllactosamine (6′-SLN); E shows chitosan oligosaccharide (COS); F shows chondroitin sulfate (CS); G shows fucoidan (Fucoidan); H shows sodium heparin sulfate (Heparin-Na); I shows exopolysaccharide from Haloarcula hispanica (HhEPS); and J shows nadroparin calcium (Nadroparin);

[0109] FIG. 6 is a diagram of the toxicity of sialic acid, sialyloligosaccharides, chitosan oligosaccharide, and chondroitin sulfate of Example 2 to bronchial epithelial cells (16HBE cells), and the in the figure, A shows sialic acid (Sia); B shows 3′-sialyllactose (3′-SL); C shows 6′-sialyllactose (6′-SL); D shows 6′-N-acetyl sialyllactosamine (6′-SLN); E shows chitosan oligosaccharide (COS); and F shows chondroitin sulfate (CS);

[0110] FIG. 7 is a diagram of the toxicity of fucoidan, nadroparin calcium, chitosan oligosaccharide, and exopolysaccharide from Haloarcula hispanica of Example 2 to African green monkey kidney cells (Vero E6 cells), and the in the figure, A shows fucoidan (Fucoidan); B shows nadroparin calcium (Nadroparin); and C shows exopolysaccharide from Haloarcula hispanica (HhEPS);

[0111] FIG. 8 is a diagram of the binding of the S1 subunit of a virus to an O-sugar chain chip in an experiment of Example 3;

[0112] FIG. 9 is a diagram of the binding of the DC-SIGN receptor to an O-sugar chain chip in an experiment of Example 3;

[0113] FIG. 10 is a diagram of the binding of the DC-SIGN receptor to an HMO chip in an experiment of Example 3;

[0114] FIG. 11 is a diagram of the binding of the S1 subunit of a virus to heparin sulfate in an experiment of Example 4;

[0115] FIG. 12 is a diagram of the binding of the S1 subunit of a virus to chondroitin sulfate in an experiment of Example 4;

[0116] FIG. 13 is a diagram of the binding of the S1 subunit of a virus to exopolysaccharide from Haloarcula hispanica in an experiment of Example 4;

[0117] FIG. 14 is a diagram of the binding of the S1 subunit of a virus to fucoidan in an experiment of Example 4;

[0118] FIG. 15 is a diagram of the binding of the S1 subunit of a virus to 3-fucosyllactose in an experiment of Example 4;

[0119] FIG. 16 is a diagram of the binding of the S1 subunit of a virus to 2′-fucosyllactose in an experiment of Example 4;

[0120] FIG. 17 is a diagram of the binding of the S1 subunit of a virus to nadroparin calcium in an experiment of Example 4;

[0121] FIG. 18 is a diagram of an observation result of a control without any sugar chain of Example 5;

[0122] FIG. 19 is a diagram of blocking the binding of the S1 subunit to 16HBE cells (bronchial epithelial cells) by chondroitin sulfate;

[0123] FIG. 20 is a diagram of blocking the binding of the S1 subunit to 16HBE cells (bronchial epithelial cells) by fucoidan;

[0124] FIG. 21 is a diagram of blocking the S1 subunit (the S1 protein) of SARS-CoV-2 to lung epithelial cells (BEAS-2B cells) by sugar chains of Example 6, and in the figure, A shows BEAS-2B cells; B shows the binding of the S1 protein to BEAS-2B cells; C shows an inhibitory effect of exopolysaccharide from Haloarcula hispanica (HhEPS) on the binding of the S1 protein to BEAS-2B cells; D shows an inhibitory effect of fucoidan (Fucoidan) on the binding of the S1 protein to BEAS-2B cells; E shows an inhibitory effect of sodium heparin sulfate (Heparin-Na) on the binding of the S1 protein to BEAS-2B cells; and F shows an inhibitory effect of nadroparin calcium (Nadroparin) on the binding of the S1 protein to BEAS-2B cells;

[0125] FIG. 22 is a diagram of blocking the S1 subunit (the S1 protein) of SARS-CoV-2 to African green monkey kidney cells (Vero-E6 cells) by sugar chains of Example 6, and in the figure, A shows Vero-E6 cells; B shows the binding of the S1 protein to Vero-E6 cells; C shows an inhibitory effect of exopolysaccharide from Haloarcula hispanica (HhEPS) on the binding of the S1 protein to Vero-E6 cells; D shows an inhibitory effect of fucoidan (Fucoidan) on the binding of the S1 protein to Vero-E6 cells; E shows an inhibitory effect of sodium heparin sulfate (Heparin-Na) on the binding of the S1 protein to Vero-E6 cells; and F shows an inhibitory effect of nadroparin calcium (Nadroparin) on the binding of the S1 protein to Vero-E6 cells; and

[0126] FIG. 23 is a diagram of inhibitory effects of sugar chains of Example 7 on pseudovirus invasion and infection of African green monkey kidney cells (Vero-E6 cells), and in the figure, A shows fucoidan (Fucoidan); B shows exopolysaccharide from Haloarcula hispanica (HhEPS); and C shows nadroparin calcium (Nadroparin).

DETAILED DESCRIPTION OF THE INVENTION

[0127] The present application will be further described below with reference to specific embodiments.

[0128] It should be noted that the terms “above”, “below”, “left”, “right”, “middle” and the like used herein are only for the convenience of description and understanding, and are not intended to limit the implementable scope. Changes or adjustments made to a relative relationship of technical features without substantially changing the technical content shall fall within the implementable scope of the present application.

[0129] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art to which the present application belongs. The term “and/or” used herein includes any and all combinations of one or more of associated items listed.

[0130] Experiments described without introducing the conditions are carried out in accordance with the conventional conditions or the conditions suggested by manufacturers. Reagents or instruments described without introducing manufacturers are commercially available common products.

[0131] The term “about” used herein is for providing flexibility in association with a given term, measurement or value. Those skilled in the art can readily determine the degree of flexibility of a specific variate.

[0132] The term “at least one of” used herein is intended to be synonymous with “one or more of”. For example, “at least one of A, B, C” expressly includes only A, only B, only C, and combinations of any two or more of A, B, and C.

[0133] Concentration, quantity, and other numerical data may be presented herein in range format. It should be understood that such range format is only for convenience and brevity, and should be flexibly construed to include not only values expressly stated as the limits of the range, but also all individual values or subranges subsumed within the range, as if each numerical value and subrange were expressly stated. For example, a numerical range of about 1 to about 4.5 should be construed to include not only the expressly stated limits of about 1 and about 4.5, but also individual numbers (e.g. 2, 3, and 4) and subranges (e.g. 1 to 3 and 2 to 4). The same principle is applicable to ranges with only one stated numerical value, such as “less than about 4.5”, which should be construed to include all values and subranges that are less than about 4.5. In addition, this interpretation should applicable regardless of the range of the scope or features described.

[0134] Steps in any method or process claimed may be performed in any order and are not limited to the order defined in the claims.

[0135] The present application will be further described below with reference to specific examples.

Example 1

[0136] Experiment on Enzymatic Synthesis of Functional Oligosaccharides Such as 6′-Sialyllactose (6′-SL)

[0137] Related enzymes required by the synthesis of functional oligosaccharides were expressed and purified, and respectively subjected to activity analysis. Functional oligosaccharides were efficiently obtained by in vitro enzymatic synthesis, and then subjected to HMR and MS characterization.

[0138] The synthesis of 6′-sialyllactose (6′-SL) was taken as an example. A reaction system (10 mL) was composed of 3.6 mM sialic acid, 3 mM lactose, 3.6 mM CTP, Tris-HCl (pH=8.5, 100 mM), 20 mM MgCl.sub.2, 2 μM NmCSS (CMP-sialic acid synthetase, Yu H, et al., Bioorganic & Medicinal Chemistry, 2004, 12, 6427-6435), 2 μM Pd26ST (α-2,6-sialyltransferase, Sun M, et al., Biotechnol Lett. 2008, 30, 671-676), and 2 μM PPA (pyrophosphatase, Li L, et al., Org. Lett., 2013, 15, 5528-5530). The conditions included that: the reaction system reacted at 140 rpm and 37° C. for 48 h. After the reaction was completed, anhydrous ethanol with the same volume was added, and the mixture was concentrated, purified on Sepahadex G15, and further separated and purified by HPLC. Finally, a purified product, i.e. 6′-sialyllactose (6′-SL), was subjected NMR and mass spectrometry characterization.

[0139] The purified product, i.e. 6′-SL, was subjected to .sup.1H NMR (see FIG. 1), .sup.13C NMR (see FIG. 2), and mass spectrometry (see FIG. 3) characterization. .sup.1HNMR (600 MHz, D.sub.2O) δ 5.14 (d, J=6.0 Hz, H-1α), 4.59 (d, J=12.0 Hz, H-1β), 4.35 (d, J=12.0 Hz, 1H), 3.89-3.48 (m, 18H), 3.23 (t, J=6.0 Hz, 1H), 2.63 (dd, J=6.0, 12.0 Hz, 1H), 1.95 (s, 3H), 1.65 (t, J=12.0 Hz, 1H) (see FIG. 1).

[0140] .sup.13C NMR (600 MHz, D.sub.2O) δ 175.7, 174.9, 103.4, 100.4, 95.6, 91.8, 79.6, 74.6, 73.8, 72.5, 72.4, 71.8, 70.8, 68.5, 68.4 63.6, 62.7, 61.4, 60.3, 60.1, 51.8, 40.1, 22.1 (see FIG. 2).

[0141] MALDI-TOF, the calculated molecular weight was 633.55 (the molecular formula of 6′-SL: C.sub.23H.sub.39NO.sub.19), and the actual molecular weight was 632.234 (see FIG. 3).

[0142] 6′-sialyllactose (6′-SL) was a sugar chain containing the Neu5Acα2-6Gal building block at the non-reducing end.

Example 2

[0143] Cytotoxicity Tests of a Series of Functional Oligosaccharides

[0144] Effects of oligosaccharides (polysaccharides), such as sialic acid (Sia), 3′-sialyllactose (3′-SL), 6′-sialyllactose (6′-SL), 6′-N-acetyl sialyllactosamine (6′-SLN), chitosan oligosaccharide (COS), chondroitin sulfate (CS), sodium heparin sulfate (Heparin-Na), nadroparin calcium (Nadroparin), fucoidan (Fucoidan), and exopolysaccharide from Haloarcula hispanica (HhEPS), on 16HBE cells (bronchial epithelial cells), BEAS-2B cells (lung epithelial cells), and Vero-E6 (African green monkey kidney cells) were determined by MTT assays.

[0145] Specific experimental steps were as follows.

[0146] 1) The above cells were respectively prepared into 20 mg/mL mother solutions.

[0147] 2) Concentration gradients were set according to Table 1.

TABLE-US-00001 TABLE 1 Concentration gradients Volume of 20 mg/mL 1640 mother culture Configured solution solution volume Concentration (μL) (μL) (μL) Ctrl (Control) 0 2000 2000 10 μg/mL 1 1999 2000 50 μg/mL 5 1995 2000 100 μg/mL 10 1990 2000 200 μg/mL 20 1980 2000 500 μg/mL 50 1950 2000 1000 μg/mL 100 1900 2000

[0148] 2) The cell concentration was set to be 2×10.sup.5 cells/mL, and 150 μL of each prepared solution was inoculated into each well of a 96-well plate.

[0149] 3) After the cells adhered to wells for 12 h, the culture solution was removed by suction, and 200 μL of oligosaccharide culture solutions at different concentrations were added to each well, and the same oligosaccharide culture solution was added to 6 wells.

[0150] 4) 24 h after the oligosaccharide solutions at different concentrations were added, 20 μL of MTT solution at a concentration of 5 mg/mL was added to each well.

[0151] 5) After the MTT solution was added, the cells were incubated for 3.5 h, and 150 μL of DMSO was added to each well.

[0152] 6) After the cells were uniformly mixed with DMSO for 10 min, the 96-well plates were placed into a microplate reader and detected at a detection wavelength of 490 nm. 7) The relative cell viability was determined by comparing with values of the control at 490 nm.

[0153] It can be known from results shown in FIG. 5, FIG. 6, and FIG. 7 that most of the sugar chains have weak toxicity to lung epithelial cells (BEAS-2B cells), bronchial epithelial cells (16HBE cells), and African green monkey kidney cells (Vero-E6).

[0154] Among the sugar chains, 6′-N-acetyl sialyllactosamine (Neu5Acα2-6Galβ1-4GlcNAc, 6′-SLN) was an oligosaccharide containing the Neu5Acα2-6Gal building block.

Example 3

[0155] Experiments on the binding of the S1 subunit of SARS-CoV-2 and DC-SIGN to an O-sugar chain chip were carried out in the present example. The O-sugar chain chip and HMO chip used in the present application were purchased from Nanjing Creative Biochip Co., Ltd.

[0156] 1) Fluorescent labeling of S1 subunit and DC-SIGN sample solutions

[0157] 1 mg/mL S1 subunit sample and 1 mg/mL DC-SIGN sample were respectively prepared. 100 μL of sample was added to 25 μL of labeling reaction solution at 5-fold diluted concentration (containing 130 mM NaHCO.sub.3 and 50 mM NaCl, pH=8.2-8.3), an appropriate volume of Cy3-Se (cyanine 3 succinimidyl ester, an N-hydroxysuccinimide activated anthocyanidin 3 fluorescent dye) fluorescent dye (10 mg of Cy3-Se/1 mL of dimethyl sulfoxide), 0.5 μL of fluorescent dye was correspondingly added to the S1 subunit sample, 0.9 μL of fluorescent dye was correspondingly added to the DC-SIGN sample, and the mixtures were slightly shaken to form uniform reaction solutions.

TABLE-US-00002 TABLE 2 Corresponding addition volume of fluorescent dye Item 1 (S1 subunit) 2 (DC-SIGN) Concentration (mg/mL) 1 mg/mL 1 mg/mL CY3-Se (μL) 0.5 0.9

[0158] 2) The reaction solutions were shaken on a shaker for 1 h.

[0159] 3) Dialysis after labeling: 125 μL of each reaction solution was injected into a mini dialysis tubing.

[0160] 4) The reaction solutions were respectively stirred 3 times in a dialysis buffer solution (300 mL×3) in the dark, and each stirring was performed for 1 h (after the last stirring, the reaction solutions were placed overnight).

[0161] 5) The samples were taken out and respectively transferred into a new centrifuge tube.

3.2 Preparation of Samples to be Analyzed

[0162] Concentration of samples to be analyzed: the 2 samples were denoted as No. 1 and No. 2 samples, respectively. The No. 1 sample referred to the S1 subunit, and the No. 2 sample referred to DC-SIGN. The above No. 1 and No. 2 samples were subjected to serial dilution to form samples at different test concentrations.

[0163] Specific dilutions were as follows: at the first dilution, the samples were diluted to a concentration of 10 μg/mL (the volume of each sample was 250 μL); at the second dilution, the samples were subjected to 1-fold dilution to a concentration of 5 μg/mL (the volume of each sample was 250 μL); at the third dilution, the samples were subjected to 1-fold dilution to a concentration of 2.5 μg/mL (the volume of each sample was 250 μL); and at the forth dilution, the samples were subjected to 1-fold dilution to a concentration of 1.25 μg/mL (the volume of each sample was 250 μL).

[0164] The corresponding arrangement of the samples at different test concentrations on an 8-subarray microarray chip is shown in Table 3.

TABLE-US-00003 TABLE 3 Arrangement of samples on an 8-subarray microarray chip Sugar chain Serial No. of samples concentration Volume No. 1 sample (S1 subunit) 10 μg/mL 250 μL No. 1 sample (S1 subunit) 5 μg/mL 250 μL No. 1 sample (S1 subunit) 2.5 μg/mL 250 μL No. 1 sample (S1 subunit) 1.25 μg/mL 250 μL No. 2 sample (DC-SIGN) 10 μg/mL 250 μL No. 2 sample (DC-SIGN) 5 μg/mL 250 μL No. 2 sample (DC-SIGN) 2.5 μg/mL 250 μL No. 2 sample (DC-SIGN) 1.25 μg/mL 250 μL

3.3. Chip Analysis

[0165] Part I. Blocking

[0166] The microarray chip was treated in a clean and dry environment. Gloves were required when touching the surface of the chip.

[0167] Before being opened, the chip package was placed at room temperature for about 20 min. Then, the package was opened, and a chip to be used was taken out, encapsulated into a reusable pouch with a desiccant inside, and stored in a refrigerator at −20° C.

[0168] Fences for analysis were mounted on the chip. A blocking buffer solution was added to each subarray. For an 8-subarray chip (referring to a glass substrate with 8 printed sugar chain areas, which contain the same sugar chain and are used to detect 8 different samples at the same time), 200 μL of buffer solution added to each subarray.

[0169] The subarrays were sealed with a plastic film to prevent evaporation of the blocking buffer solution, and incubated on a rotary shaker (at 60 rpm) for 30 min.

[0170] Part II. Binding Experiment

[0171] The samples were centrifuged on a centrifuge to avoid increasing particles on the chip.

[0172] A pipette was carefully put at the corner of each subarray to remove the blocking buffer solution from each subarray without touching the surface of the microarray chip.

[0173] The samples to be analyzed were added to each subarray immediately. For the 8-subarray chip, 200 μL of sample was added to each subarray. The sample solution should cover the entire area of each subarray without producing air bubbles in the sample solution.

[0174] The microarray chip was sealed with a plastic film to prevent evaporation of the sample solution, and covered with aluminum foil and incubated in the dark for 1 h (on a shaker at 80 rpm).

[0175] 5) During the experiment, especially during the long incubation, the surface of the chip was prevented from being dried. A sealing effect of the plastic film should be ensured.

[0176] Part III: Final Washing and Drying

[0177] A pipette was carefully put at the corner of each subarray to take the sample from each subarray without touching the surface of the microarray chip.

[0178] A washing buffer solution was added to each subarray. For the 8-subarray chip, 200 μL of buffer solution was added to each subarray. The microarray chip was sealed with a plastic film, and incubated on a shaker (at 80 rpm) for 5 min. The washing buffer solution was removed immediately by using a pipette, and the above steps were repeated.

[0179] After the washing buffer solution was completely removed, the fences for analysis were unmounted from the chip, the glass slide was immersed in a washing buffer solution, and the chip was incubated on a shaker (at 80 rpm) for 10 min.

[0180] The glass slide was immersed in deionized water, and the chip was incubated on a shaker (at 80 rpm) for 2 min.

[0181] Before data scanning, the chip was dried in a clean and dust-free environment.

[0182] Part IV: Data Reading and Analysis

[0183] FIG. 8 is a diagram of the binding of the S1 subunit of SARS-CoV-2 to the O-sugar chain chip. It can be seen from the results shown in FIG. 8 that when the concentration of the S1 subunit is 1.25 μg/mL, some O-sugar chains bind to the S1 subunit of SARS-CoV-2 to a certain extent.

[0184] The sugar chains include O20 (Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-3Galβ1-3GalNAcα), O33 (Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-3GalNAcα), O23 (GlcNAcβ1-6(Galβ1-3)GalNAcα), O29 (Neu5Acα2-3Galβ1-4GlcNAcβ1-3GalNAcα), O36 (Galα1-3(Fucα1-2)Galβ1-4GlcNAcβ1-3NAcα), O39 (GlcNAcβ1-6(GlcNAcβ1-3)GalNAcα), and O83 (Neu5Acα2-6Galβ1-4GlcNAcβ1-3(Neu5Acα2-6Galβ1-3GlcNAcβ1-6)GalNAcα).

TABLE-US-00004 TABLE 4 Binding of the S1 subunit of a virus to an O-sugar chain chip Sugar chain RFU O20 304.5 O23 69.84 O29 2866.34 O33 6320.17 O36 320.5 O39 136.5 O83 6012.67

[0185] Among the sugar chains, O20 and O33 are sugar chains containing the Lewis y building block at the non-reducing end, O83 is a sugar chain containing the Neu5Acα2-6Gal building block at the non-reducing end, O29 is a sugar chain containing the Neu5Acα2-3Gal building block at the non-reducing end, and O23 and O39 are sugar chains containing the GlcNAcβ1-6 building block at the non-reducing end. Therefore, it can be determined from the above results that HMNOs, such as 3′-sialyllactose containing Neu5Acα2-6Gal at the non-reducing end and 3′-sialyllactosamine containing Neu5Acα2-3Gal at the non-reducing end, can also bind to the S1 subunit.

[0186] FIG. 9 is a diagram of the binding of the DC-SIGN protein to the O-sugar chain chip. It can be seen from the results shown in FIG. 9 that some O-sugar chains bind to the DC-SIGN protein of a receptor to a certain extent.

TABLE-US-00005 TABLE 5 Binding of DC-SIGN to an O-sugar chain chip Sugar chain RFU O19 83.33 O20 135 O32 68.83 O33 124.5 O44 69 O46 65.83

[0187] The sugar chain include O19 (Galβ1-4(Fucα1-3)GlcNAcβ1-3Galβ1-3GalNAcα), O20 (Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-3Galβ1-3 GalNAcα), O32 (Galβ1-4(Fuca 1-3)GlcNAcβ1-3 GalNAcα), O33 (Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-3GalNAcα), O44 (Galβ1-4(Fuca 1-3)GlcNAcβ1-3 GalNAcα), and O46 (Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-6GalNAcα.

[0188] Among the sugar chains, O19, O32, and O44 are sugar chains containing the Lewis x building block at the non-reducing end, and O20, O33, and O46 are sugar chains containing the Lewis y building block at the non-reducing end.

[0189] It can be known from the above results that O20 (Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-3Galβ1-3GalNAcα) and 033 (Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-3GalNAcα) are sugar chains that can bind to not only the S1 subunit, but also DC-SIGN.

[0190] FIG. 10 is a diagram of the binding of DC-SIGN of a receptor to an HMO chip. It can be seen from the results shown in FIG. 10 that some HMOs bind to DC-SIGN to a certain extent.

[0191] The HMOs contain the Lewis a, Lewis x, Lewis y or Blood Group H type II building block at the non-reducing end, such as HMO13 (Galβ1-3(Fucα1-4)GlcNAcβ1-3Galβ1-4Glc), HMO14 (Galβ1-4(Fucα1-3)GlcNAcβ1-3Galβ1-4Glc), HMO15 (Fucα1-2Gal 1-4GlcNAcβ1-3Galβ1-4Glc), HMO16 (Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-3Gal 1-4Glc), HMO31 (GlcNAcβ1-6[Galβ1-4(Fucα1-3)GlcNAcβ1-3]Galβ1-4Glc), HMO33 (GlcNAcβ1-6[Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-3]Galβ1-4Glc), HMO36 (Galβ1-4(Fucα1-3)GlcNAcβ1-6(GlcNAcβ1-3)Gal 1-4Glc), HMO40 (Galβ1-4GlcNAcβ1-6(Fucα1-2Gal 1-4GlcNAcβ1-3)Gal 1-4Glc), HMO43 (Galβ1-4(Fucα1-3)GlcNAcβ1-6(Galβ1-4GlcNAcβ1-3)Gal 1-4Glc), HMO45 (Fucα1-2Galβ1-4GlcNAcβ1-6(Fucα1-2Galβ1-4GlcNAcβ1-3)Galβ1-4Glc), HMO46 (Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-6(Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-3)Galβ1-4Glc), HMO2 (Galβ1-4(Fucα1-3)Glc), and HMO3 (Fucα1-2Galβ1-4Glc).

TABLE-US-00006 TABLE 6 Binding of DC-SIGN to an HMO chip Sugar chain RFU HMO2 2329.09 HMO3 2930.75 HMO13 1896.5 HMO14 2125.34 HMO15 1683.84 HMO16 2537.92 HMO31 2175.59 HMO33 1913.59 HMO36 1707.42 HMO40 801.5 HMO43 1196.59 HMO45 838.42 HMO46 2425.92

[0192] FIG. 1 is a diagram of structural formulas of Neu5Acα2-6Gal, Neu5Acα2-3Gal, Lewis a, Lewis x, Lewis y, and Blood Group H type II, and in the figure, 1A shows Neu5Acα2-6Gal, 1B shows Neu5Acα2-3Gal, 1C shows Lewis a, 1D shows Lewis x, 1E shows Lewis y, and 1F shows Blood Group H type II.

Example 4

[0193] Experiments on the binding of the S1 protein (or the S1 subunit) of SARS-CoV-2 to various oligosaccharides were carried out in the present example. An MO NT. 115 microscale thermophoresis measurement instrument purchased from Nano Temper was used for detection, and experimental steps were as follows.

[0194] 1) First, the S1 protein was subjected to fluorescent labeling. In the present experiment, a Monolith™ RED-NHS 2.sup.nd generation protein labeling kit was used for labeling.

[0195] 2) A buffer solution for the S1 protein was replaced. 100 μg of S1 protein (1 mg/mL) expressed in mammalian cells was taken, the column A of the kit was used to replace the protein dissolution buffer solution with a labeling buffer solution (containing 130 mM NaHCO.sub.3 and 50 mM NaCl, pH=8.2-8.3).

[0196] 3) 25 μL of dimethyl sulfoxide (DMSO) was used to dissolve a RED-NHS dye of the kit.

[0197] 4) 5 μL of RED-NHS dye was diluted and uniformly mixed with 5 μL of labeling buffer solution.

[0198] 5) 90 μL of S1 protein was added and uniformly mixed with the fluorescent dye, and labelled in the dark at 37° C. for 30 min.

[0199] 6) The labelled protein was purified by using the column B, and 450 μL of purified protein was obtained.

[0200] 7) Oligosaccharides used in the experiment were subjected to serial dilutions with ddH.sub.2O, each oligosaccharide was diluted at 16 concentration gradients, and the initial concentration of each oligosaccharide was as follows:

[0201] sodium heparin sulfate (Heparin-Na): 83.33 μM;

[0202] sodium chondroitin sulfate (Chondroitin sulfate-Na): 40 mg/mL, 285.75 μM;

[0203] exopolysaccharide from Haloarcula hispanica (HhEPS, EPS from Haloarculahispanica): 22 mg/mL, 10 μM;

[0204] fucoidan (Fucoidan): 30 mg/mL, 60 μM;

[0205] 3-fucosyllactose (Galβ1-4(Fucα1-2)Glc, 3-FL): 1 mg/mL, 2049 μM;

[0206] 2′-fucosyllactose (Fucα1-2Galβ1-4Glc, 2′-FL): 5 mg/mL, 10.24 mM; and

[0207] nadroparin calcium (Nadroparin): 444.33 μM.

[0208] The labelled protein was mixed with the oligosaccharide solution with the same volume. Generally, 10 μL of labelled protein was mixed with 10 μL of oligosaccharide solution, and 20 μL of reaction system was obtained.

[0209] The oligosaccharide-protein mixed solutions at 16 concentration gradients were sampled by using capillaries. 7 μL of sample was taken each time, and 20 μL of reaction solution was used in three repeated experiments.

[0210] The 16 capillaries with samples were placed on the lanes of the MO NT. 115 microscale thermophoresis measurement instrument in order to be subjected to a protein-oligosaccharide binding experiment.

[0211] Dissociation constants (K.sub.d) of the binding of the S1 protein to the sugar chains were calculated from obtained data. The dissociation constants (K.sub.d) of the binding of the S1 protein to the sugar chains are shown in Table 7.

TABLE-US-00007 TABLE 7 Dissociation constants (K.sub.d) of the binding of the S1 protein to various sugar chains Sugar chain Kd (μM) Sodium heparin sulfate (Heparin-Na) 1.25 Sodium chondroitin sulfate (Chondroitin sulfate-Na) 19 Exopolysaccharide from Haloarcula hispanica (HhEPS, 3.77 EPS from Haloarculahispanica) Fucoidan (Fucoidan) 1.61 3-fucosyllactose (Galβ1-4(Fucα1-2)Glc, 3-FL) 7.1 2′-fucosyllactose (Fucα1-2Galβ1-4Glc, 2′-FL) 71 Nadroparin calcium (Nadroparin) 6.2

[0212] It can be known from the data in Table 7 that the affinity of sugar chains, such as sodium heparin sulfate, fucoidan, exopolysaccharide of Haloarcula hispanica, and 3-fucosyllactose, with the S1 protein is relatively strong, and the affinity of sodium chondroitin sulfate and 2′-fucosyllactose with the S1 protein is relatively weak.

[0213] FIG. 11 is a diagram of the binding of the S1 subunit of the virus to heparin sulfate in the experiment; FIG. 12 is a diagram of the binding of the S1 subunit of the virus to chondroitin sulfate; FIG. 13 is a diagram of the binding of the S1 subunit of the virus to exopolysaccharide from Haloarcula hispanica; FIG. 14 is a diagram of the binding of the S1 subunit to fucoidan; FIG. 15 is a diagram of the binding of the S1 subunit to 3-fucosyllactose; FIG. 16 is a diagram of the binding of the S1 subunit to 2′-fucosyllactose; and FIG. 17 is a diagram of the binding of the S1 subunit to nadroparin calcium.

[0214] In FIG. 11 to FIG. 17, the left diagrams show thermophoresis curves of the binding of the sugar chains at different concentrations to the S1 subunit, and the right diagrams show curves of the binding of the sugar chains at different concentrations to the S1 subunits. In the left diagrams, the x-axis refers to time, and y-axis refers to relative fluorescence intensities; and in the right diagrams, the x-axis refers to concentrations of the sugar chains, and the y-axis refers to one thousandth of differences between fluorescence intensities of the binding of the sugar chains at different concentrations to the S1 subunit and a fluorescence intensity of the binding of the sugar chains at the lowest concentration to the S1 subunit.

[0215] It can be known from FIG. 11 to FIG. 17 that the 6 sugar chains in the above example can all specifically bind to the S1 subunit of the virus in the experiment, but exhibits different concentration dependencies upon binding.

Example 5

[0216] Experiments on blocking of the binding of the S1 subunit (the S1 protein) of SARS-CoV-2 to human bronchial epithelial cells by sugar chains were carried out in the present example. Experimental steps were as follows.

[0217] 1) 16HBE cells (bronchial epithelial cells) were inoculated into a 24-well cell culture plate with a glass bottom (Cellvis), 1 mL of culture medium (containing 90% 1640 culture medium, 10% fetal calf serum, and 1% double antibody (Pen Strep: 100 U/mL penicillin and 10 μg/mL streptomycin)) was added to each well, after being cultured for 24-48 h, the cells covered the bottom of each well, and next step was performed.

[0218] 2) The cell culture solution in the 24-well plate was removed by suction, and each well was washed 3 times with PBS (pH=7.4, 1 mL×3).

[0219] 3) 200 μL of each sugar chain solution, i.e. 40 μM chondroitin sulfate and 20 μM fucoidan, was added to each well to incubate the cells, 200 μL of PBS was added to a control (without any saccharide), and the cells were incubated for 1 h.

[0220] 4) After 1 h, 100 μL of fluorescently labeled (using a Monolith™ RED-NHS 2.sup.nd generation protein labeling kit that was purchased from Nano Temper) S1 protein (10 g/mL) was added to each well, and the cells were incubated for another 1 h.

[0221] 5) Solution in each well was removed by suction, and each well was washed 3 times with PBS (500 μL×3).

[0222] 6) 200 μL of paraformaldehyde (4%) tissue fixation solution was added to each well to fix the cells for 30 min.

[0223] 7) Solution in each well was removed by suction, and each well was washed 3 times with PBS (500 μL×3).

[0224] 8) 1 μg/mL of DAPI (4′,6-diamidino-2-phenylindole) dye was added to each well to stain cell nucleus for 10 min.

[0225] 9) Solution in each well was removed by suction, and each well was washed 3 times with PBS (500 μL×3).

[0226] 10) 300 μL of PBS was added to each well, and the binding of the S1 protein (red light) to the cells (blue light) was observed under a laser scanning confocal microscope.

[0227] FIG. 18 is a diagram of the control without any sugar chain; FIG. 19 is a diagram of blocking the binding of the S1 subunit to 16HBE cells (bronchial epithelial cells) by chondroitin sulfate; and FIG. 20 is a diagram of blocking the binding of the S1 subunit to 16HBE cells (bronchial epithelial cells) by fucoidan.

[0228] In FIG. 18 to FIG. 20, the left diagrams show observation results of RED staining, and the right diagrams show observations results of DAPI staining. As shown in FIG. 18 to FIG. 20, the S1 protein directly binds to the cells, and RED and DAPI staining conditions are respectively observed by using red light and blue light. The results show that clear cell morphologies can be observed at both wavelengths, and the two cell morphologies coincide, which indicates that the S1 protein successfully binds to 16HBE cells (bronchial epithelial cells). On the contrary, after chondroitin sulfate or fucoidan is added, the clear cell morphology can be observed from the DAPI staining condition, and the cellular outline cannot be seen under red light, which indicates that the S1 protein does not bind to the cells, and also proves that chondroitin sulfate and fucoidan successfully block the binding of the S1 protein to bronchial epithelial cells.

Example 6

[0229] Experiments on blocking the binding of the S1 subunit (the S1 protein) of SARS-CoV-2 to lung epithelial cells (BEAS-2B cells) and African green monkey kidney cells (Vero-E6 cells) by sugar chains were carried out in the present example. Experimental steps were as follows.

[0230] (1) The cells were inoculated into a 24-well cell culture plate with a glass bottom (1×10.sup.5 cells/well), 1 mL of culture medium was added to each well, after being cultured for 24-48 h, the cells covered the bottom of each well, and next step was performed.

[0231] (2) The cell culture solution in the 24-well plate was removed by suction, and each well was washed 3 times with PBS (1 mL×3).

[0232] (3) 300 μL of 4% paraformaldehyde (PFA) solution was added to each well to fix the cells at room temperature for 1 h.

[0233] (4) The fixation solution was removed by suction, and each well was washed 3 times with PBS (1 mL×3).

[0234] (5) Sugar chains were added to the cells to pre-protect the cells, and the cells were incubated at 37° C. for 1 h.

[0235] (6) After 1 h, the S1 subunit (1 μM) was added to each well, and the cells were incubated at 37° C. for 1 h.

[0236] (7) The solution in each well was removed by suction, and each well was washed 3 times with PBS (1 mL×3).

[0237] (8) 200 μL of Mouse anti-SARS-CoV-2 mAb (Sino Biological, 40592-MM57) diluted with an HBSSA solution in a ratio of 1:1000 was added to each well, and the cells were incubated at 37° C. for 1 h.

[0238] (9) The primary antibody solution was removed by suction, and each well was washed 3 times with PBS (1 mL×3).

[0239] (10) 200 μL of Goat anti-Mouse IgG (H+L) Cross-Adsorbed Secondary Ab (Alexa Fluor 488) diluted with an HBSSA solution in a ratio of 1:1000 was added to each well, and the cells were incubated at 37° C. for 1 h.

[0240] (11) The solution in each well was removed by suction, and each well was washed 3 times with PBS (500 μL×3).

[0241] (12) 1 μg/mL of DAPI dye was added to each well to stain the cells for 10 min.

[0242] (13) The solution in each well was removed by suction, and each well was washed 3 times with PBS (500 μL×3).

[0243] (14) The cells were observed under a laser scanning confocal microscope at 405 nm (DAPI) and 490 nm (FITC), respectively, and inhibitory effects of the sugar chains on the binding of the S1 protein to the cells were evaluated.

[0244] FIG. 21 is a diagram of blocking the binding of the S1 subunit to BEAS-2B cells (lung epithelial cells) by the sugar chains; and FIG. 22 is a diagram of blocking the binding of the S1 subunit to Vero-E6 cells (African green monkey kidney cells) by the sugar chains.

[0245] It can be seen from FIG. 20 and FIG. 21 that sugar chains, such as exopolysaccharide from Haloarcula hispanica (HhEPS), fucoidan (Fucoidan), sodium heparin sulfate (Heparin-Na), and nadroparin (Nadroparin), can block the binding of the S1 subunit to BEAS-2B cells and Vero-E6 cells.

Example 7

[0246] Experiments on Inhibitory Effects of Oligosaccharides on Pseudovirus Invasion and Infection of Vero-E6 Cells

[0247] (1) Vero-E6 cells were inoculated into a 48-well plate (5×10.sup.4 cells/well), 1 mL of culture medium was added to each well, after being cultured for 24 h, the cells covered the bottom of each well, and the next step was performed.

[0248] (2) Viroid of SARS-CoV-2 was amplified in 293T cells and subjected to TCID50 titration.

[0249] (3) The culture medium in the 48-well plate was removed by suction, and the 48-well plate was washed twice with DMEM (without serum).

[0250] (4) Sugar chains were subjected to gradient dilution with fresh DMEM (containing 10% FBS), and 200 μL of diluted sugar chain was added to each well of the 48-well plate, and the cells were incubated at 37° C. for 2 h.

[0251] (5) A pseudovirus solution was added to the 48-well plate, and the cells were incubated at 37° C. for 24 h.

[0252] (6) A supernate was taken, and each well was washed twice with PBS (500 μL×2).

[0253] (7) 100 μL of pancrelipase solution was added to each well, and the cells were digested for 3-5 min.

[0254] (8) 100 μL of DMEM containing 2% FBS was added to each well, and the cells were suspended by blowing and beating and then transferred into a flow tube.

[0255] (9) Proliferation of the pseudovirus was analyzed by using a flow cytometer, and inhibitory effects of the sugar chains on the pseudovirus were evaluated.

[0256] FIG. 23 is a diagram of inhibitory effects of the sugar chains on pseudovirus invasion and infection of Vero-E6 cells.

[0257] It can be seen from FIG. 23 that sugar chains, such as fucoidan (Fucoidan), exopolysaccharide from Haloarcula hispanica (HhEPS), and nadroparin (Nadroparin), exhibit an inhibition activity to pseudovirus invasion and infection of the cells, and the highest inhibition rate is.

Example 8

[0258] A mouthwash for preventing and/or treating 2019-nCoV infection was prepared, in which the concentration of an active ingredient was 0.01%-1% (W/V), and the active ingredient contained the following sugar chain components:

[0259] Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-3Galβ1-3GalNAcα, Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-3GalNAcα, sulfated fucoidan, HMOs, ginsenosides, chitosan oligosaccharide, heparin sulfate, heparan sulfate oligosaccharide, chondroitin sulfate, chondroitin sulfate oligosaccharide, dermatan sulfate, keratan sulfate, hyaluronic acid, hyaluronic acid oligosaccharide, fucoidan, fucose oligosaccharide, protein N/O-sugar chains, and exopolysaccharide from Haloarcula hispanica.

[0260] Other ingredients of the mouthwash were water or ingredients applicable to mouthwash.

Example 9

[0261] A mouthwash for preventing and/or treating 2019-nCoV infection was prepared, in which the concentration of an active ingredient was 0.01%-1% (W/V), and the active ingredient contained the following sugar chain components:

[0262] Fucα1-2Galβ1-4GlcNAcβ1-3Galβ1-4Glc, Fucα1-2Galβ1-4GlcNAcβ1-6(Fucα1-2Galβ1-4GlcNAcβ1-3)Galβ1-4Glc, sodium heparin sulfate, fucoidan, exopolysaccharide from Haloarcula hispanica, and 3-fucosyllactose.

[0263] Other ingredients of the mouthwash were glycerol or ingredients applicable to mouthwash.

Example 10

[0264] A nasal wash for preventing and/or treating coronavirus infection was prepared, in which the concentration of an active ingredient was 0.01%-1% (W/V), and the active ingredient contained the following sugar chain components:

[0265] Neu5Acα2-6Galβ1-4GlcNAcβ1-3(Neu5Acα2-6Galβ1-3GlcNAcβ1-6)GalNAc, 6′-sialyllactose (Neu5Acα2-6Galβ1-4Glc), 6′-N-acetyl sialyllactosamine (Neu5Acα2-6Galβ1-4GlcNAc), Fucα1-2Galβ1-4(Fucα1-3)Galβ1-4GlcNAcβ1-3Galβ1-3GalNAcα, Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-3Galβ1-3GalNAcα, Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-6GalNAcα, Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-3Galβ1-4Glc, and Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-6(Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-3)Galβ1-4Glc.

[0266] Other ingredients of the nasal wash were glycerol or ingredients applicable to nasal wash.

Example 11

[0267] A pharmaceutical composition for preventing and/or treating 2019-nCoV infection, in which the concentration of an active ingredient was 0.01%-1% (W/V), and the active ingredient contained the following sugar chain components:

[0268] Fucα1-2Galβ1-4(Fucα1-3)Galβ1-4GlcNAcβ1-3Galβ1-3GalNAcα, Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-3Galβ1-3GalNAcα, Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-6GalNAcα, Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-3Galβ1-4Glc, Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-6(Fucα1-2Galβ1-4(Fucα1-3)GlcNAcβ1-3)Galβ1-4Glc, Fucα1-2Galβ1-4GlcNAcβ1-3Galβ1-4Glc, and Fucα1-2Galβ1-4GlcNAcβ1-6(Fucα1-2Galβ1-4GlcNAcβ1-3)Galβ1-4Glc.

[0269] Other ingredients of the pharmaceutical composition are preferably water, and the pharmaceutical composition was delivered to the lung with the assistance of a gasifying agent.

[0270] The above content is a schematic description of the present application and its implementation modes, and the description is not restrictive. Implementation modes shown in the embodiments are only some of the implementation modes of the present application, and the actual implementation modes are not limited thereto. Therefore, any structural modes and embodiments similar to the technical solutions designed by those of ordinary skill in the art inspired by the implementation modes without creativity and without departing from the purpose of the present invention shall fall within the scope of protection of the present application.