WO1995005182A1

WO1995005182A1 - Bridged oligosaccharides and sulfated derivatives thereof

Info

Publication number: WO1995005182A1
Application number: PCT/US1994/008818
Authority: WO
Inventors: Peter Fugedi; Csaba Janos Peto; John Henry Musser
Original assignee: Glycomed Incorporated
Priority date: 1993-08-13
Filing date: 1994-08-04
Publication date: 1995-02-23
Also published as: AU7520394A

Abstract

The present invention relates to bridged oligosaccharide(s) compounds and sulfated derivatives thereof having the structure R?1-[X1-R2]¿n-X2-R3 wherein R?1, R2 and R3¿ are independently one or more saccharide(s); X?1 and X2¿ are independently difunctional or polyfunctional alkyl, aryl or aralkyl compounds capable of covalently joining together said saccharides; and n is an integer of zero to ten.

Description

BRIDGED OLIGOSACCHARIDES AND SULFATED DERIVATIVES THEREOF

Field of the Invention

The present invention relates to a novel class of bridged mono- and oligosaccharides and sulfated derivatives thereof. The present invention also relates to a method for using this novel class of sulfated derivatives for use as antithrombotic, anticoagulant, antiangiogenic and anti-inflammatory agents.

Background of the Invention

Heparin and other naturally occurring mucopolysaccharides, such as dermatan sulfate or heparin sulfate, possess antithrombotic properties. Recently, several low molecular weight heparins from various chemical or enzymatic depolymerization processes have been developed for clinical use. Thomas et al., Thrombos. Res. (1982) 28:343-350; Walenga et al., Thrombos. Res. (1986) 43:243-248; Roller et al., Thrombos Haemostas. (1986) 56:243-246. Heparins and low molecular weight heparins have the disadvantage that they are of natural origin, generally being derived from animals. As such, small amounts of tissue present from the animal tissues can be present to cause anaphylactic reactions such as a decrease in the number of thrombocytes, thrombosis and embolism. Therefore, the preparation of synthetic agents possessing comparable antithrombotic, anticoagulant, antiangiogenic and anti-inflammatory properties as heparin are desired.

The present invention relates to a new class of oligosaccharides useful as antithrombotic, anticoagulant, antiangiogenic and anti-inflammatory agents.

Summary of the Invention

The present invention relates to bridged saccharide(s) compounds and sulfated derivatives thereof having the structure

R'-tX'-R X²-!*³ wherein R¹, R² and R³ are independently one or more saccharide(s);

X¹ and X² are independently difunctional or polyfunctional alkyl, aryl or aralkyl compounds capable of covalently joining together said saccharides; and n is an integer of zero to ten.

These and other objects of the invention will become apparent upon a full consideration of the following disclosure.

A second object of the invention is a description of bridged saccharide(s) compounds as described above wherein R¹, R², and/or R³ have sulfateable hydroxyl or amine groups, and at least certain of these groups are preferably sulfated.

A third object of the invention is a description of methods of treating disease with the bridged saccharide(s) compounds of the invention, preferably diseases that are benefitted by treatment with heparin.

Detailed Description of the Invention

The present invention relates to the synthesis of a novel class of bridged mono- and oligosaccharides and their sulfated derivatives. The compounds of the present invention have the general structure R¹-[X¹-R ]_n-X²-R³ wherein R¹, R² and R³ are one or more saccharide(s) having sulfateable hydroxyl or amine groups and certain of these groups are preferably sulfated. X corresponds to a bridging group between the saccharide units and encompasses difunctional and polyfunctional compounds, preferably alkyl, aryl and aralkyl compounds wherein the bridging group joins together two saccharide compounds by, preferably, an ether, thioether, glycosidic or thioglycosidic bond, n is an integer, equal to or greater than zero preferably n = 1 - 10, most preferably n = 1 - 5.

With regard to the monosaccharide or oligosaccharide group, R¹, R² and R³ can be in the pyranose or furanose ring form and have either an α or β configuration at the anomeric center. R¹, R² and R³ can be the same or different monosaccharide or oligosaccharide. Examples of monosaccharides useful in the present invention include, but are not limited to, D-glucose, D-galactose, D-mannose, D-xylose, D- and L-arabinose, D- ribose, L-rhamnose, L-fucose, D-glucuronic acid, D-galacturonic acid, L-iduronic acid, D- glucosamine, D-galactosamine and sialic acid. Examples of disaccharides useful in the present invention include, but are not limited to, maltose, lactose, cellobiose, melibiose and 3-O-β-D-galactopyranosyl-D-arabinose. Examples of trisaccharides and higher oligosaccharides useful in the present invention include, but are not limited to, maltotriose, and maltotetraose.

With regard to the bridging group, X can be a difunctional or polyfunctional alkyl, aryl or aralkyl group capable of linking two saccharide units together through preferably either an ether, thioether, glycosidic or thioglycosidic bond. Examples of bridging groups include but are not limited to diols, oligomers of diols, aromatic diols such as hydroquinone and dihydroxynapthalenes, aralkyl diols such as benzenedimethanol, dithiols, oligomers of dithiols and thiohydroxy compounds. Optionally, the bridging group may possess additional functional groups such as hydroxyls, thiols, amines, carboxylic acids, amides or sulfonic acids wherein these groups do not form bonds with the saccharide units.

In the case where n > 0, the compound contains more than one bridging group. When a compound contains more than one bridging group, the same bridging group need not be used throughout the compound. Further, when more than one bridging group is present in a compound, each bridging group - saccharide linkage can be either an ether, thioether, glycosidic or thioglycosidic bond.

Attachment of the bridging group to the anomeric center of the saccharide unit to form either the glycosidic or thioglycosidic bond can be achieved by the reaction of an activated saccharide derivative, e.g. glycosyl halides, thioglycosides, glycosyl imidates, or n-pentenyl glycosides with the hydroxyl or thiol of the bridging group. Attachment of the bridging group by an ether or thioether linkage can be achieved using classical methods of ether preparation.

If so desired, the bridged non-sulfated compounds can be functionalized to enable the attachment of additional mono- or oligosaccharide units. An example of a chain extension is provided in Example 7. An important aspect of the present invention is the sulfation of the saccharide groups preferably via their hydroxyl or amine groups. The hydroxyl and amine groups can be either partially or completely sulfated.

After the saccharide groups have been joined together by the bridging group(s), the saccharide groups are deprotected to yield free hydroxyls and amines. The free hydroxyls and amines are then sulfated using an appropriate an appropriate sulfating agent such as but not limited to chlorosulfonic acid or complexes of sulfur trioxide with organic bases in an inert solvent such as N,N-dimethylformamide, hexamethylphosphoric triamide, dimethyl sulfoxide or pyridine. Using techniques known in the art, selective sulfation of either the hydroxyl or amine groups can be obtained. In the case of sulfating amines, water can be used as a solvent. After sulfation, the sulfate groups can be modified to possess biologically acceptable cations, including but not limited to Na, K, Li, Ca, Mg, NH₄, aluminum, ethanolamine, triethanolamine, morpholine, pyridine and piperidine.

Labeled Forms of the Invention Non-Anticoagulant Compositions

The compositions of the invention can be provided with fluorescent, radioisotope, or enzyme labels as desired. Conventional techniques for coupling of label to carbohydrates or related moieties can be used. Such techniques are well established in the art. See, for example, U.S. Patent No. 4,613,665. The labeled mixtures of the invention may be used to identify sites of disease as well as in competitive immunoassays, and as a means to trace the pharmacokinetics of the compositions in vivo. Suitable radioisotope labels for this purpose include hydrogen³, iodine¹³¹, indium¹¹¹, technetium", and phosphorus³². Suitable enzymic labels include alkaline phosphatase, glucose-6-phosphate-dehydrogenase, and horseradish peroxidase. Particularly preferred fluorescent labels include fluorescein and dansyl. A wide variety of labels of all three types is known in the art.

Administration and Use

The non-anticoagulant heparin compositions of the instant invention are useful in therapeutic applications for treating or preventing a variety of diseases including cancer, inflammation, and diseases caused or exacerbated by platelet aggregation, heparanase or angiogenic activity. The instant compositions, because of their anti-angiogenic activity, will be preferably applied for the beneficial treatment of angiogenic based diseases. One such class of diseases is retinopathies. A member of this class is diabetic retinopathy that will be favorably treated by the compositions of the instant invention.

Administration of the bridged saccharide(s) compounds of the invention is typically by routes appropriate for glycosaminoglycan compositions, and generally includes systemic administration, such as by injection.

Particularly preferred is intravenous injection, as continuous injection over long time periods can be easily continued. Also preferred are introduction into the vascular system through intraluminal administration or by adventitial administration using osmotic pumps or implants. Typical implants contain biodegradable materials such as collagen, polylactate, polylactate/polyglycoside mixtures, and the like. These may be formulated as patches or beads. Typical dosage ranges are in the range of 0.1-10 mg/kg/hr on a constant basis over a period of 5-30, preferably 7-14, days. Particularly preferred dosage is about 0.3 mg/kg/hr, or, for a 70 kg adult, 21 mg/hr or about 500 mg/day.

Other modes of administration are less preferred but may be more convenient. Injection subcutaneously at a lower dose or administered orally at a slightly higher dose than intravenous injection, or by transmembrane or transdermal or other topical administration for localized injury may also be effective. Localized administration through a continuous release device, such as a supporting matrix, perhaps included in a vascular graft material, is particularly useful where the location of the trauma is accessible.

Formulations suitable for the foregoing modes of administration are known in the art, and a suitable compendium of formulations is found in Remington's Pharmaceutical Sciences. Mack Publishing Company, Easton, PA, latest edition.

The compositions of the invention may also be labeled using typical methods such as radiolabeling, fluorescent labeling, chromophores or enzymes, and used to assay the amount of such compositions in a biological sample following its administration. Suitable protocols for competitive assays of analytes in biological samples are well known in the art, and generally involve treatment of the sample, in admixture with the labeled competitor, with a specific binding partner which is reactive with the analyte such as, typically, an immunoglobulin or fragment thereof. The antibodies prepared according to the invention, as described below, are useful for this purpose. The binding of analyte and competitor to the antibody can be measured by removing the bound complex and assaying either the complex or the supernatant for the label. The separation can be made more facile by preliminary conjugation of the specific binding partner to a solid support. Such techniques are well known in the art, and the protocols available for such competitive assays are too numerous and too well known to be set forth in detail here.

The synthesis and biological activity of the compounds of the present invention are illustrated in the following examples. Further objectives and advantages other than those set forth above will become apparent from the examples and accompanying figures. Examples

Example 1 Ethane- 1.2-diyl bis(β-D-glucopyranoside) sulfate (PF#1) A mixture of ethylene glycol (0.31g), mercuric cyanide (3.79 g), mercuric bromide (0.36 g), and 4 A molecular sieves powder (2 g) in a mixture of nitromethane (150 mL) and toluene (150 mL) was distilled at atmospheric pressure until about half of the solvent was distilled off. The mixture was cooled to 45 °C and 2,3,4,6-tetra-O-acetyl-α-D- glucopyranosyl bromide (6.17 g) was added. After stirring the mixture at this temperature for 10 hours, additional 2,3,4,6-tetra-O-acetyl-α-D-glucopyranosyl bromide (4.11 g) and mercuric cyanide (1.89 g) were added, and stirring was continued overnight. The mixture was filtered, the filtrate was evaporated, the residue was taken up in chloroform (300 mL), washed with 10% aqueous KI, saturated aqueous NaHCO₃ and then water, dried and evaporated. Column chromatography (toluene-ethyl acetate 3:2 — » 6:5) gave ethane- 1,2-diyl bis(2,3,4,6-tetra-O-acetyl-β-D-glucopyranoside) (2.12 g, 58.6%). [α]_D -29.2° (c 0.96, chloroform). Η-NMR data (CDC1₃): δ 2.00, 2.02, 2.08, 2.09 (4s, 4 X 3H, COCH₅); 3.72 (ddd, 1Η, Η-5); 3.86 (dd, 2H, OCH₂); 4.14 (dd, 1Η, Η-6b); 4.29 (dd, 1H, H-6a); 4.58 (d, 1H, H-l 7^=8.0 Hz); 4.99 (dd, 1H, H-2); 5.09 (dd t, 1H, H-3); 5.23 (dd t, 1H, H-4). ¹³C- NMR data (CDC1₃): 5 20.62, 20.73, 20.76 (4C, COCH₃); 61.79 (C-6); 68.33 (C-4); 68.56 (OCH₂); 71.37 (C-2); 71.72 (C-5); 73.72 (C-3); 100.45 (C-l) 169.47, 169.55, 170.22, 170.68 (4C, COCH₃)

To a solution of this compound (1.82 g) in methanol (20 mL) methanolic sodium methoxide was added to adjust the pH to about 9. The mixture was stirred at room temperature for 5 h and was then neutralized with Amberlite IR-120 (H⁺) resin. The resin was filtered off, the filtrate was evaporated, and the residue was dried in vacuo to leave ethane- 1,2-diyl bis(β-D-glucopyranoside) (0.97 g, 100%) as a syrup. [α]_D -25.5° (c 0.93, methanol). Η-NMR data (D₂O): δ 3.84 (dd, 2H, OCH₂); 4.36 (d, 1Η, Η-l 7^=7.8 Hz). ^I3C-NMR data (D₂O): δ 62.73 (C-6); 70.11 (OCH₂); 71.57, 75.12, 77.96 (C-2,3,4,5); 104.50 (C-l).

To a solution of the above glucoside (0.193 g) in N,N-dimethylformamide (20 mL), sulfur trioxide pyridine complex (1.27 g) was added and the mixture was stirred at room temperature for 2 days. The pH was adjusted to 9 by the addition of M NaOH, and the mixture was evaporated to dryness. The residue was desalted on a Biogel P-2 column with 0.5 M NH₄HCO₃, and the desalted product was passed through an SP Sephadex C-25 (Na⁺) column with water to give 0.511 g ethane- 1,2-diyl bis(β-D-glucopyranoside) sulfate (PF#1). ¹³C-NMR-data (D₂O): δ 67.79 (C-6); 69.19 (OCH₂); 72.47, 76.88, 79.92, 83.91 (C-2,3,4,5); 100.70 (C-l). Anal. Calc. for C₁₄H₁₉O₃₃S₇Na₇ x 5H₂O: C, 14.12; H, 2.45; S, 18.85. Found: C, 14.77; H, 2.43; S, 18.77.

Example 2 3.6.9.12.15-Pentaoxa-heptadecane- 1 , 17-diyl bis(β-D-glucopyranoside) sulfate (PF#2)

To a mixture of 2,3,4,6-tetra-O-benzoyl-α-D-glucopyranosyl bromide (3.957 g), hexaethyleneglycol (0.565 g), and 4 A molecular sieves powder (2.5 g) in dichloromethane (20 mL) a solution of silver trifluoromethanesulfonate (1. 925 g) and 2,4,6-collidine (1 mL) in dichloromethane-toluene (1:1, 20 mL) was added dropwise with stirring at -40 °C. Additional silver trifluoromethanesulfonate (0.963 g) and 2,4,6-collidine (0.4 mL) in dichloromethane-toluene (1:1, 10 mL) were added after 2 h. The reaction was quenched by the addition of pyridine. The mixture was then diluted with dichloromethane and filtered through a pad of Celite. The filtrate was subsequently washed with water, 10% aqueous Na₂S₂O₃, M HC1, water, aqueous NaHCO₃, and water, dried and evaporated. Column chromatography (toluene-ethyl acetate 6:5) gave 3,6,9,12,15-pentaoxa-heptadecane-l,2-diyl bis(2,3,4,6-tetra-O-benzoyl-β-D-glucoρyranoside) (1.51 g, 52%). [α]_D +18.3° (c 1.05, chloroform). Η-NMR data (CDC1₃): δ 3.38, 3.45, 3.51, 3.56 (m, 9H, OCH₂); 3.60 (ddd, 1Η, Η-5); 3.80, 3.98, 4.16 (m, 3H, OCH₂); 4.49 (dd, 1Η, Η-6a); 4.64 (dd, 1H, H-6b); 4.97 (d, 1H, H-l 7^=7.9 Hz); 5.53, 5.69, 5.91 (dd, 3H, H-2,3,4); 7.25-7.57, 7.80-8.05 (m, 20H, 4 Ph). ¹ C-NMR data (CDC1₃): δ 63.11 (C-6); 69.36, 70.29 70.40, 70.60 (OCH₂); 69.70, 71.85, 72.15, 72.87 (C-2,3,4,5); 101.34 (C-l); 128.28, 128.36, 128.73, 129.78, 133.12, 133.22, 133.42 (Ph); 165.05, 165.17, 165.78, 166.12 (4 COCH₃).

A solution of the benzoate (1.08 g) in a mixture of methanol (20 mL) and tetrahydrofuran (10 mL) was treated with methanolic sodium methoxide to adjust the pH to 9. After one day, the mixture was neutralized with AG 50W-X8 (H⁺) resin. The resin was then filtered off, and the filtrate evaporated. The residue was purified by column chromatography (chloroform-90% aqueous methanol 9:1) to give 3,6,9,12, 15-pentaoxa- heptadecane-l,17-diyl bis(β-D-glucopyranoside) (0.45 g, 100%). [α]_D -16.5° (c 0.99, methanol).

Sulfation of this product (54.9 mg) with sulfur trioxide pyridine complex (229 mg) in DMF (5 mL) was performed as described in Example 1 and gave 3,6,9, 12,15-pentaoxa- heptadecane-l,17-diyl bis(β-D-glucopyranoside) sulfate (PF#2) (39.6 mg). ¹³C-NMR-data (D₂O 300MHz): δ 68.08 (C-6); 69.18, 69.69,69.72, 69.83 (OCH₂); 72.74, 73.65, 75.94, 76.34 (C-2,3,4,5); 100.39 (C-l).

Example 3

Ethane- 1,2-diyl bis(O- -D-glucopyranosyl-(l→4)-β-D-glucopyranoside) sulfate (PF#3). The glycosylation was performed as described in Example 2 using ethylene glycol (0.21 g), 2,3,6-tri-O-benzoyl-4-O-(2,3,4,6-tetra-O-benzoyl-α-D-glucopyranosyl)-α-D- glucopyranosyl bromide (11.5 g), 4 A molecular sieves powder (7 g), silver trifluoromethanesulfonate (3.21 g), 2,4,6-collidine (0.34 mL). The crude product was purified by column chromatography (toluene-ethyl acetate 95:5— »9:1) to give ethane-1,2- diyl bis[O-(2,3,4,6-tetra-O-benzoyl-α-D-glucopyranosyl)-(l→4)-2,3,6-tri-O-benzoyl-β-D- glucopyranoside] (5.28g 73%). [oc]_D +77.1° (c 1.21, chloroform). Η-NMR data (CDC1₃): δ 3.72 (dd, 2H, OCH₂); 4.45 (d, 1Η, Η-l 7^=7.7 Hz); 5.69 (d, IH, H-l' 7^=3.6 Hz). ¹³C- NMR data (CDC1₃): δ 62.49, 63.31 (C-6, C-6'); 68.82 (OCH₂); 69.06, 69.07, 69.77, 71.10, 72.33, 72.52, 73.08, 74.63 (C-2,3,4,5,2',3',4',5'); 96.32 (C-l'); 100.63 (C-l); 128.16-133.53 (Ph); 164.97, 165.03, 165.15, 165.53, 165.75, 165.82, 166.04 (7 COPh).

The benzoate (6.06 g) was debenzoylated as described in Example 2, the crude product was purified by column chromatography (chloroform -90% aqueous methanol 1:1) to give 1.89 g (95 %) ethane- 1,2-diyl bis(O-α-D-glucopyranosyl-(l→4)-β-D- glucopyranoside). [α]_D +62.2° (c 1.0, methanol). Η-NMR data (CD₃OD): δ 4.45 (d, IH, H-l 7^=7.8 Hz); 5.25 (d, IH, H-l' 7^=3.7 Hz). ¹³C-NMR data (CD₃OD): δ 61.93, 62.22 (C-6, C-6'); 70.11 (OCH₂); 70.99, 73.58, 74.35, 74.63, 76.22, 77.45, 80.16); 102.13 (C-l'); 103.97 (C-l).

The compound was sulfated as described in Example 1 to give ethane- 1,2-diyl bis[O-α-D-glucopyranosyl-(l→4)-β-D-glucopyranoside] sulfate (PF#3). Η-NMR data (D,O): δ 5.06 (d, IH, H-l 7^=5.2 Hz); 5.63 (d, IH, H-l' 7^=3.3 Hz). ¹ C-NMR data (D₂O): δ 66.32, 67.67 (C-6, C-6'); 68.92 (OCH₂); 70.27, 72.28, 72.57, 73.37, 73.53, 75.02, 76.53, 77.79 (C-2,3,4,5,2',3',4',5'); 94.40 (C-l'); 100.70 (C-l). Anal. Calc. for Q_tfH^O^S^Na^ x 12H₂O: C, 13.25; H, 2.39; S, 19.05. Found: C,l 3.16,; H, 2.43; S, 19.27.

Example 4 3,6,9, 12.15-Pentaoxa-heptadecane- 1 , 17-diyl bis(O-α-D-glucopyranosyl- (l-- 4)-β-D-glucopyranoside) sulfate (PF#4) The glycosylation was performed as described in Example 2 using hexaethylene glycol (0.28 g), 2,3,6-tri-O-benzoyl-4-O-(2,3,4,6-tetra-O-benzoyl-α-D-glucopyranosyl)-α-D- glucopyranosyl bromide (3.45 g), silver trifluoromethanesulfonate (0.962 g), and 2,4,6- collidine (0.1 mL). Column chromatography (toluene-ethyl acetate 8:2— >7:3) afforded 3,6,9, 12,15-pentaoxa-heptadecane-l,17-diyl bis[O-(2,3,4,6-tetra-O-benzoyl-α-D- glucopyranosyl)-(l→4)-2,3,6-tri-O-benzoyl-β-D-glucopyranoside] (1.34 g, 56%). [α]_D +54.1° (c 1.06, chloroform). Η-NMR data (CDC1₃): δ 3.30-3.57 (m, 10H, OCH₂); 3.73 (m, 1Η, OCH₂); 3.94 (m, 1Η, OCH₂); 4.87 (d, 1Η, Η-l 7^=7.7 Hz); 5.74 (d, IH, H-l' 7^=3.9 Hz); 7.14-8.15 (m, 35H, Ph). ¹³C-NMR data (CDC1₃): δ 62.49 (C-6); 63.46 (C-6'y_ 69.29, 70.17, 70.36, 70.52, (O H₂); 69.09, 69.85, 70.87, 72.25, 72.83, 73.12, 74.91 (C- 2,3,4,5,2',3',4',5'); 96.38 (C-l'); 100.84 (C-l); 128.07-133.49 (Ph); 165.00, 165.03, 165.15, 165.41, 165.63, 165.82, 166.16 (COPh).

Debenzoylation of the product as described in Example 2, followed by column chromatography (chloroform-90% aqueous methanol 1:1) gave 3,6,9, 12,15-pentaoxa- heptadecane-l,17-diyl bis(O-α-D-glucopyranosyl-(l→4)-β-D-glucopyranoside) (0.45 g, 96%). [α]_D +49.6° (c 1.32, methanol). Η-NMR (CD₃OD): δ 4.34 (d, IH, H-l 7^=7.8 Hz); 5.16 (d, IH, H-l' 7^=3.7 Hz). ¹³C-NMR (D₂O): δ 62.20, 62.75 (C-6, C-6'); 69.70, 71.45, 71.51 (OCH₂); 74.14, 74.64, 74.79, 75.07, 76.60, 77.68, 81.32 (C-2,3,4,5,2',3',4',5'); 102.94 (C-l'); 104.33 (C-l).

The compound was sulfated as described in Example 1 to give 3,6,9,12,15- pentaoxa-heptadecane-l,17-diyl bis(O-α-D-glucopyranosyl-(l*→4)-β-D-glucopyranoside) sulfate (PF#4). Η-NMR data (D₂O): δ 5.01 (d, IH, H-l 7^=4.3 Hz); 5.59 (d, IH, H-l' 7_r =3.3 Hz). ¹³C-NMR data (D₂O): δ 66.28, 67.65 (C-6, C-6'); 68.56, 69.50, 69.66 (OCH₂); 71.76, 71.94, 73.45, 73.86, 75.21, 75.93, 77.12 (C-2,3,4,5,2',3',4',5'); 94.39 (C-l'); 99.73 (C-l). Anal. Calc. for C₃₆H₅₂O₆₉S₁₄Na₁₄ x 10H₂O: C, 17.02; H, 2.85; S, 17.67. Found: C, 16.89; H, 2.86; S, 17.83.

Example 5

Ethane- 1,2-diyl bis(O-α-D-glucopyranosyl-(l— >4)-O-α-D-glucopyranosyl- (1— »4)-β-D-glucopyranoside) sulfate (PF#5). The glycosylation was performed as described in Example 2 using ethylene glycol (0.128g), O-(2,3,4,6-tetra-O-benzoyl-α-D-glucopyranosyl)-(l→4)-O-(2,3,6-tri-O-benzoyl-α- D-glucopyranosyl)-(l→4)-2,3,6-tri-O-benzoyl-α-D-glucopyranosyl bromide (6.74 g), silver trifluoromethane sulfonate (1.61 g), 2,4,6-collidine (0.05 mL). The product, ethane- 1,2-diyl bis[O-(2,3,4,6-tetra-O-benzoyl-α-D-glucopyranosyl)-(l→4)-O-(2,3,6-tri-O-benzoyl-α-D- glucopyranosyl)-(l→4)-2,3,6-tri-O-benzoyl-β-D-glucopyranoside] (4.27 g, 73%) was isolated by column chromatography (toluene- ethyl acetate, 95:5->9:l). It had [α]_D +88.4° (c 0.96, chloroform). Η-NMR data (CDC1₃): δ 3.67 (dd, 2H, OCH₂); 4.44 (d, 1Η, Η-l 7^=7.7 Hz); 5.54 and 5.80 (2d, 2 X IH, H-l' and H-l" 7=3.6 Hz 7=3.8 Hz, respectively). ¹ C-NMR data (CDC1₃): δ 62.24 (C-6); 62.93, 62.97 (C-6',C-6"); 68.89 (OCH₂); 68.98, 69.11, 69.98, 70.89, 71.66, 72.19, 72.59, 73.30, 73.76, 74.59 (C- 2,3,4,5,2',3',4',5',2",3",4",5"); 96.62, 96.72 (C-1',C-1"); 100.59 (C-l). The compound was debenzoylated as described in Example 2 to give ethane- 1,2-diyl bis(O-α-D-glucopyranosyl-(l-→4)-O-α-D-glucopyranosyl-(l*→4)-β-D-glucopyranoside) (1.28g, 82%). [α]_D +96.5° (c 0.96, methanol). Η-NMR data (D₂O 300): δ 4.53 (d, IH, H-l 7^=8.0 Hz); 5.32 (2d, 2H, H-l' and H-l", 7=3.5 Hz and 7=3.7 Hz, respectively). ¹³C-NMR data (D₂O): 60.71, 60.85 (C-6,6',6"); 69.25 (OCH₂); 69.56, 71.45, 71.72, 72.00, 72.96, 73.12, 73.20, 73.54, 74.75, 76.30, 77.10, 77.28 (C-2,3,4,5,2',3',4',5',2",3",4",5"); 99.80, 100.06 (C-l',1"); 102.44 (C-l).

Sulfation as described in Example 1 gave ethane- 1,2-diyl bis(O-α-D- glucopyranosyl-(l→4)-O-α-D-glucopyranosyl-(l→4)-β-D-glucopyranoside) sulfate (PF#5). Η-NMR data (D₂O): 4.95 (d, IH, H-l, J₁₂= 5.5 Hz), 5.54, 5.65 (2d, 2 X IH, H-l',1", 7=3.5 Hz and 7= 3.3 Hz, respectively). ¹³C-NMR data: 66.46, 66.52 (C-6,6',6"); 69.03 (OCH₂); 70.13, 70.30, 70.35, 72.03, 72.20, 72.47, 73.25, 73.39, 74.92, 76.26 (C- 2,3,4,5,2',3',4',5',2",3",4",5"); 94.16, 94.52 (C-l',1"); 100.61 (C-l). Anal. Calc for C₃₈H₄₆O₉₂S₂₀Na₂₀ x 18H₂O: C, 13.42; H, 2.42; S, 18.86. Found: C, 13.84; H, 2.55; S, 18.89.

Example 6 3,6,9, 12,15-Pentaoxa-heptadecane-l,17-diyl bis(O- -D-glucopyranosyl-

( 1 -→4)-O-α-D-glucopyranosyl-( 1 — 4)-β-D-glucopyranoside) sulfate (PF#6) Glycosylation of hexaethylene glycol (0.141g) with O-(2,3,4,6-tetra-O-benzoyl-α-D- glucopyranosyl)-(l-→4)-O-(2,3,6-tri-O-benzoyl- -D-glucopyranosyl)-(l*→4)-2,3,6-tri-O- benzoyl-α-D-glucopyranosyl bromide (1.68g), as described above, afforded 3,6,9,12,15- pentaoxa-heptadecane-l,17-diyl bis[O-(2,3,4,6-tetra-O-benzoyl-α-D-glucopyranosyl)-(l-→4)- O-(2,3,6-tri-O-benzoyl-α-D-glucopyranosyl)-(l*→4)-2,3,6-tri-O-benzoyl-β-D- glucopyranoside] (0.98g, 59%). [α]_D +65.8 (c 1.18, chloroform). Η-NMR data (CDC1₃): δ 3.31-3.35 (m, 2H, OCH₂); 3.38-3.43 (m, 2Η, OCH₂); 3.45-3.56 (m, 6Η, OCH₂); 3.72 (m, 1Η, OCH₂); 3.92 (m, 1Η, OCH₂); 4.84 (d, 1Η, Η-l 7^=7.6 Hz); 5.61 and 5.76 (2d, 2 X IH, H-l' and H-l", 7=3.4 Hz and 7=3.3 Hz, respectively). ¹³C-NMR data (CDC1₃): δ 62.25, 63.00, 63.13 (C-6,6',6"); 69.24, 70.16, 70.38, 70.58 (OCH₂); 69.00, 69.11, 69.94, 70.02,70.71, 70.88, 71.70, 72.28, 72.85, 73.62, 73.66, 74.95 (C-

2,3,4,5,2',3',4',5',2",3",4",5"); 96.53, 96.76 (C-l',1"); 100.75 (C-l); 164.75, 164.80, 165.04, 165.14, 165.31, 165.52, 165.78, 165.88, 166.12 (COPh). Debenzoylation of the product as described in Example 2 gave 3,6,9,12,15- pentaoxa-heptadecane- 1 , 17-diyl bis(O-α-D-glucopyranosyl-( 1 —* 4)-O-α-D-glucopyranosyl- (l→4)-β-D-glucopyranoside) (0.35g, 96%). [α]_D +83.3° (c 1.10, methanol). Η-NMR data (CD₃OD): δ 4.36 (d, IH, H-l 7^=7.7 Hz); 5.17 (2d, 2H, H-l',1", 7=3.5 Hz and 7=3.7 Hz, respectively). ¹³C-NMR data (CD₃OD): δ 62.13, 62.24, 62.71 (C-6,6',6"); 69.69, 71.42, 71.46 (OCH₂); 73.31, 73.70, 74.17, 74.59, 74.74, 74.93, 75.04, 76.53, 77.61, 81.20, 81.28 (C-2,3,4,5,2',3',4',5',2",3",4",5"); 102.65, 102.82 (C-l',1"); 104.24 (C-l).

Sulfation as described in Example 1 gave 3,6,9,12,15-pentaoxa-heptadecane-l,17-diyl bis(O-α-D-glucopyranosyl-(l→4)-O-α-D-glucopyranosyl-(l*→4)-O-β-D-glucopyranoside) sulfate (PF#6). Η-NMR data (D₂O): δ 4.91 (d, IH, H-l 7^=5.2 Hz); 5.54, 5.64 (2d, 2 X IH, H-l',1" 7=4.4 Hz and 7=3.4 Hz, respectively). ¹³C-NMR data (D₂O): δ 66.04, 66.48, 67.72 (C-6,6',6"); 68.58, 68.83, 69.69, 69.64 (OCH₂); 69.93, 71.88, 72.18, 72.55, 73.17, 73.46, 73.60, 74.93, 76.01, 76.36, 77.74 (C-2,3,4,5,2',3',4',5',2",3",4",5"); 94.19, 94.55 (C- l',l"); 100.06 (C-l). Anal. Calc. for C₄₈H₆₆O₉₇S₂₀Na₂₀ x 20H₂O: C, 15.76; H, 2.91; S, 17.53. Found: C, 15.47; H, 3.01; S, 17.34.

Example 7

Ethane- 1 ,2-diyl bis(O-α-D- glucopyranosvH 1 —>4)-O-α-D-glucopyranosyl- ( 1 *-- 4)-β-D-glucopyranosyl-( 1 — >4)-O- -D-glucopyranosyl-( 1 *→4) -O-β-D-glucopyranoside) sulfate (PF#7) Ethane- 1,2-diyl bis(O-α-D-glucopyranosyl-(l->4)-β-D-glucopyranoside) (2.011 g) was dissolved in DMF (40 mL), α,α-dimethoxy-toluene (1.8 mL) and 10-camphorsulfonic acid (0.07 g) were added and the mixture was stirred at 60 °C for 2h in vacuo. The mixture was neutralized with NaHCO₃, and was evaporated to dryness. The residue was subjected to column chromatography (chloroform-methanol 95:5—>4:l) to give ethane- 1,2- diyl bis[O-(4,6-O-benzylidene-α-D-glucopyranosyl)-(l→4)-β-D-glucopyranoside] (2.373 g, 94%). [α]_D +19.1° (c 0.98, methanol). Η-NMR data (DMSO): δ 4.26 (d, IH, H-l 7^=7.7 Hz); 5.13 (d, IH, H-l' 7^=3.7 Hz); 5.57 (s, IH, PhCH); 7.34-7.38 (m, 3Η, Ph); 7.42-7.46 (m, 2H, Ph). ¹³C-NMR data (DMSO): δ 60.66 (C-6); 68.10, 68.21 (C-6' and OCH₂); 63.40, 70.11, 73.04, 73.14, 75.15, 76.44, 79.52, 81.00 (C-2,3,4,5,2',3',4',5'); 100.93, 101.18 (C-l', PhCH); 102.99 (C-l); 126.50, 128.13, 128.97, 137.85 (Ph). The 4,6-O-benzylidene derivative (2.37 g) was dissolved in DMF (25 mL), 80% sodium hydride (1.60 g) was added, and the mixture was cooled to 0 °C. Benzyl bromide (4.76 mL) was added, and the mixture was stirred overnight at room temperature. Methanol was added, the solution was evaporated to dryness, the residue was partitioned between chloroform and water, the organic layer was dried and evaporated. Column chromatography (toluene-ethyl acetate 9:1) gave ethane- 1,2-diyl bis[O-(2,3-di-O-benzyl-4,6- O-benzylidene-α-D-glucopyranosyl)-(l→4)-(2,3,6-tri-O-benzyl-β-D-glucopyranoside] (2.63g, 55%). [α]_D +10.8° (c 1.01, chloroform). ¹³C-NMR (CDC1₃): δ 68.83, 68.88 (C-6, C- 6', OCH₂); 63.20, 71.83, 74.12, 78.64, 82.14, 82.21, 84.75 (C-2,3,4,5,2',3',4',5'); 75.2, 74.5, 73.72, 72.66, 73.4 (5 PhCH₂); 97.18 (C-l'); 101.07 (PhCH); 103.76 (C-l); 137.52, 137.78, 138.13, 138.20, 138.61, 138.70 (Ph).

Reductive ring-cleavage of the 4,6-O-benzylidene derivative (0.98g) with NaCNBH₃-HCl in tetrahydrofuran afforded ethane- 1,2-diyl bis[O-(2,3,6-tri-O-benzyl-α-D- glucopyranosyl)-(l→4)-(2,3,6-tri-O-benzyl-β-D-glucopyranoside] (0.62g, 62%). [α]_D +18.0° (c 0.96, chloroform). ¹³C-NMR (CDC1₃): δ 68.82, 69.07, 69.59 (C-6, C-6', OCH₂); 70.59, 71.26, 72.31, 74.39, 78.84, 81.21, 82.04, 84.66 (C-2,3,4,5,2',3',4',5'); 72.96, 73.28, 73.52, 73.72, 74.42, 75.25 (6 PhCH₂); 96.51 (C-l'); 103.75 (C-l); 137.80, 137.89, 138.25, 138.70 (Ph).

Glycosylation of the above compound (0.479 g) with O-(2,3,4,6-tetra-O-benzoyl- - D-glucopyranosyl)-(l->4)-O-(2,3,6-tri-O-benzoyl-α-D-glucopyranosyl)-(l-→4)-2,3,6-tri-O- benzoyl-α-D-glucopyranosyl bromide (1.288 g) as described in Example 2 afforded the bridged decasaccharide, ethane- 1,2-diyl bis[O-(2,3,4,6-tetra-O-benzoyl-α-D-glucopyranosyl)- (l*→4)-O-(2,3,6-tri-O-benzoyl-α-D-glucopyranosyl)-(l->4)-O-(2,3,6-tri-O-benzoyl-β-D- glucopyranosyl)-(l*→4)-O-(2,3,6-tri-O-benzyl-α-D-glucopyranosyl)-(l*→4)-O-(2,3,6-tri-O- benzyl-β-D-glucopyranoside)] (1.047 g, 80.8%) after column chromatography (toluene-ethyl acetate 9:1→85:15). [α]_D +50.6° (c 1.03, chloroform). ¹³C-NMR data (CDC1₃): 103.7, 99.7 (2 β C-l), 96.78, 96.71, and 96.3 (3 α C-l). The bridged decasaccharide (1.005 g) was debenzoylated as described in Example 2 to give ethane- 1,2-diyl bis[O-α-D-glucopyranosyl-(l*→4)-O-α-D-glucopyranosyl-(l→4)-O- β-D-glucopyranosyl-(l→4)-O-(2,3,6-tri-O-benzyl-α-D-glucopyranosyl)-(l→4)-O-(2,3,6-tri- O-benzyl-β-D-glucopyranoside)] (0.45 g, 78.8%) after column chromatography (chloroform- 90% aqueous methanol 4:1). [α]_D +87.8° (c 1.07, methanol). ¹³C-NMR data (CD₃OD): 104.8, 103.3, 102.8, 102.6, 97.5 (5 C-l).

The above compound (0.45 g) was hydrogenated with 10% palladium on charcoal catalyst (0.4 g) in a mixture of ethanol (20 mL), water (10 mL), and acetic acid (10 mL) at atmospheric pressure for 2 days. After filtration and evaporation of the solvent, the residue was passed through a Biogel P-2 column using water as the eluent to give ethane- 1,2-diyl bis(O-α-D-glucopyranosyl-(l-→4)-O-α-D-glucopyranosyl-(l*→4)-β-D-glucopyranosyl-

(l→4)-O-α-D-glucopyranosyl-(l— -»4)-O-β-D-glucopyranoside)the title compound (0.220 g, 80.2%). ¹³C-NMR data (D₂O): 102.5, 102.4 (2 β C-l), 100.0, 99.7, 99.5 (3 α C-l), 69.2 (CH₂ ethane- 1,2-diyl).

Sulfation of this compound ( 0.168 g) as described in Example 1 afforded the title compound (0.371 g). FAB-MS data: [M-Na-2SO₃Na]^" 4725.6; [M-Na-3-SO₃Na]^" 4620.2. Example 8 Methyl 4-O--f4-O-r4-O-(α-D-glucopyranosyl)- ( 1— >4)-β-D-glucopyranosyloxyethyll- -D-glucopyranosyl I - (l-->4)-β-D-glucopyranoside sulfate (PF#8) A mixture of ethylene glycol (3.10 g), mercuric cyanide (3.55 g), mercuric bromide

(0.39 g) and 4 A molecular sieves powder (3 g) in a mixture of nitromethane (100 mL) and benzene (100 mL) was distilled at atmospheric pressure until about half of the solvent was distilled off. The mixture was cooled to room temperature, 2,3,6-tri-O-acetyl-4-O- (2',3',4',6'-tetra-O-acetyl-α-D-glucopyranosyl)-α-D-glucopyranosyl bromide (6.99 g) was added, and the mixture was stirred at room temperature overnight. The mixture was filtered through Celite, the filtrate was evaporated and the residue was taken up in chloroform (500 mL) and was washed with 10% aqueous KI, saturated aqueous NaHCO₃, water, was dried and evaporated. Column chromatography (toluene- acetone 4:1) gave 2- hydroxyethyl (2,3,4,6-tetra-O-acetyl-α-D-glucopyranosyl)-(l*→4)-2,3,6-tri-O-acetyl-β-D- glucopyranoside (2.51 g, 37%). [α]_D +53.5 (c 0.97, chloroform). Η-NMR data (CDC1₃): δ 2.00, 2.01, 2.03, 2.04, 2.11, 2.15 (21H, COCH₃); 4.58 (d, IH, H-l 7^=7.9 Hz); 5.41 (d, IH, H-l' 7^=4.0 Hz). ¹³C-NMR data (CDC1₃): δ 20.58, 20.67, 20.76, 20.90 (7 COCH₃); 61.54, 61.93, 62.76 (C-6,6', CH₂OH); 68.01, 68.60, 69.32, 69.98, 72.18, 72.26, 72.74, 75.24 (C-2,3,4,5,2',3',4',5'); 73.17 (OCH₂); 95.62 (C-l'); 100.94 (C-l); 169.44, 169.78, 169.95, 170.20, 170.50, 170.55 (7 COCH₃).

To a solution of 2-hydroxyethyl (2,3,4,6-tetra-O-acetyl-α-D-glucopyranosyl)-(l—>4)- 2,3,6-tri-O-acetyl-β-D-glucopyranoside (2.51 g) in a mixture of dichloromethane (30 mL) and tetrahydrofuran (20 mL) 4-methoxyphenol (1.19 g) and triphenylphosphine (2.51 g) were added and the solution was stirred at 0 °C. Diethyl-azodicarboxylate (1.33 mL) was added dropwise, and the mixture was stirred overnight at room temperature. The reaction mixture was diluted with chloroform (300 mL) and was successively washed with water, saturated aqueous NaHCO₃, water, then was dried and evaporated. Column chromatography (toluene-acetone 95:5 -→ 9:1) gave 2-(4-methoxyphenyl)-ethyl (2,3,4,6- tetra-O-acetyl-α-D-glucopyranosyl)-(l→4)-2,3,6-tri-O-acetyl-β-D-glucopyranoside (3.35 g, 100%). [α]_D +29.9 (c 0.99, chloroform). Η-NMR data (CDC1₃): δ 1.93, 2.00, 2.01, 2.02, 2.05, 2.10, 2.13 (COCH₃); 3.76 (s, 3H, OCH^; 4.64 (d, 1Η, Η-l 7^=7.9 Hz); 5.41 (d, IH, H-l' 7^= .0 Hz); 7.14-7.28 (m, 4H, Ph). ¹³C-NMR data (CDC1₃): δ 20.57, 20.68, 20.82, 20.90 (COCH₃); 55.68 (OCH₃); 61.50, 62.78 (C-6,6'); 67.88, 68.45 (OCH₂CH₂); 68.02, 68.49, 69.35, 70.00, 72.11, 72.17, 72.70, 75.37 (C-2,3,4,5,2',3',4',5'); 95.56 (C-l'); 100.55 (C-l); 114.66, 115.64, 152.73, 154.05 (Ph); 169.47, 169.77, 170.00, 170.26, 170.53, 170.57 (COCH₃). This compound (3.32 g) was deacteylated as described in Example 1 to afford 2-(4- methoxyphenyl)-ethyl O-(α-D-glucopyranosyl-(l*→4)-β-D-glucopyranoside) (2.08 g, 100%). [α]_D +48.3 (c 1.02, methanol). Η NMR data (CD₃OD): δ 3.78 (s, 3H, OCH,); 4.45 (d, 1Η, Η-l 7^=7.7 Hz); 5.15 (d, IH, H-l' 7^=3.6 Hz); 6.82-6.91 (m, 4H, Ph). ¹³C-NMR data (CD₃OD): δ 55.83 (OCH₃); 60.89, 61.69 (C-6,6'); 68.39 (2C, OCH₂); 70.23, 72.73, 73.27, 73.42, 73.84, 75.32, 76.18, 80.07 (C-2,3,4,5,2',3',4',5'); 101.91 (C-l'); 103.19 (C-l); 114.94, 116.03, 152.74, 154.43 (Ph).

The above compound (2.08 g) was dissolved in DMF (20 mL). 80% sodium hydride (1.78 g) was added and the mixture was cooled to 0°C. Benzyl bromide (5.35 mL) was added dropwise and the mixture was stirred overnight at room temperature. Methanol was added, the solution was evaporated to dryness, the residue was taken up chloroform and water. The organic layer was washed with water, was dried and evaporated. The product, 2- (4-methoxyphenyl)-ethyl O-(2,3,4,6-tetra-O-benzyl-α-D-g!ucopyranosyl)-(l→4)-2,3,6-tri-O- benzyl-β-D-glucopyranoside (4.72 g, 98%) was isolated by column chromatography (toluene-ethyl acetate 95:5 -*- 9:1). [α]_D +25.7 (c 1.09, chloroform). Η-NMR data (CDC1₃): δ 3.70 (s, 3H, OCH₃); 4.54 (d, 1Η, Η-l 7^=8.0 Ηz); 5.69 (d, 1Η, Η-l' 7^=3.5 Ηz); 6.76- 6.85 (m, 4Η, Ph). ¹³C-NMR data (CDC1₃): δ 55.62 (OCH₃); 67.76, 68.23, 68.14, 69.07 (C- 6,6' OCH₂CH₂); 70.97, 72.41, 74.46, 77.65, 79.27, 81.95, 82.07, 84.66 (C-2,3,4,5,2',3',4',5'); 73.17, 73.30, 73.43, 73.84, 74.50, 74.94, 75.46 (PhCH₂); 96.64 (C-l'); 103.73 (C-l); 114.59 (Ph); 126.59-127.27 (PhCH₂); 137.89, 138.24, 138.37, 138.70, 138.74 (PhCH₂); 152.82, 153.89 (Ph).

Removal of the 4-methoxyphenyl group from the compound above (4.72 g) with ammonium cerium(IV) nitrate (4.6 g) in 90% aqueous acetonitrile (50 mL) afforded 2- hydroxyethyl O-(2,3,4,6-tetra-O-benzyl- -D-glucppyranosyl)-(l-→4)-2,3,6-tri-O-benzyl-β-D- glucopyranoside (2.60 g, 62%). [α]_D +28.1 (c 1.09, chloroform). Η-NMR data (CDC1₃): δ 4.45 (d, IH, H-l 7^=7.7 Hz); 5.63 (d, IH, H-l' 7^=3.8 Hz). ¹³C-NMR data (CDC1₃): δ 62.46 (CH₂OH); 68.08, 69.10 (C-6, C-6'); 71.11, 72.97, 74.07, 77.60, 79.16, 81.89, 82.29, 84.60 (C-2,3,4,5,2',3',4',5'); 73.26, 73.36, 73.45, 73.64, 73.96, 74.74, 74.92, 75.48 (OCH₂ and 7 Ph H₂); 96.89 (C-l'); 104.07 (C-l); 126.51-128.31 (PhCH₂); 137.72, 137.81, 137.85, 138.09, 138.30, 138.63 (PhCH₂).

The 2-hydroxyethyl glycoside was converted into the 2-bromoethyl glycoside via the 2-tosyloxyethyl derivative.

The above described compound was treated with /?-toluenesulfonyl chloride (4.3 g) in a mixture of dichloromethane (40 mL) and pyridine (2 mL) at 0 °C to give 2- tosyloxyethyl 2,3,6,2',3',4',6'-hepta-O-benzyl-β-D-maltoside (2.59 g, 86%). [α]_D +22.7 (c 1.09, chloroform). Η-NMR data (CDC1₃): δ 2.34 (d, 3H, CH₃); 4.51 (d, IH, H-l 7^=7.1 Hz); 5.66 (d, IH, H-l' 7^=3.6 Hz). ¹³C-NMR data (CDC1₃): δ 21.55 (CH₃); 66.81

(CH,OTs); 68.17, 68.86, 68.98 (C-6,6/ OCH₂); 71.02, 72.44, 74.45, 77.66, 79.26, 81.94, 84.46 (C-2,3,4,5,2',3',4',5'); 73.22, 73.29, 73.45, 73.89, 74.56, 74.97, 75.48 (PhCH₂); 96.72 (C-l'); 103.40 (C-l); 126.56-129.79 (Ph); 132.90 (Ph-tosyl); 137.88, 138.06, 138.15, 138.35, 138.68 (PhCH₂); 144.77 (Ph-tosyl). This derivative was dissolved in acetonitrile (60 mL), tetrabutylammonium bromide

(3.82 g) was added and the mixture was stirred at 50 °C for 4 hours. The solution was evaporated to dryness, the product was isolated by column chromatography (toluene-ethyl acetate 9:1) to give 2-bromoethyl O-(2,3,4,6-tetra-O-benzyl-α-D-glucopyranosyl)-(l→4)- 2,3,6-tri-O-benzyl-β-D-glucoρyranoside (2.44 g, 94%). [α]_D +26.1 (c 1.00,chloroform). Η- NMR data (CDC1₃): δ 4.47 (d, IH, H-l 7^=7.3 Hz); 5.67 (d, IH, H-l' 7/',2'=3.5 Hz). ¹³C- NMR data (CDC1₃): δ 30.34 (CH₂Br); 68.18, 68.98 (C-6,6'); 69.55 (OCH₂); 71.03, 72.49, 74.51, 77.67, 79.28, 81.96, 82.02, 84.57 (C-2,3,4,5,2',3',4',5'); 73.22, 73.30, 73.46, 73.91, 74.69, 74.98, 75.49 (PhCH₂); 96.74 (C-l'); 103.58 (C-l); 126.59-128.28 (PΛCH₂); 136.90, 138.18, 138.37, 138.71 (PhCH₂). This bromoethyl derivative was used as the alkylating agent for the synthesis of the title compound.

The acceptor, methyl O-(2,3,6-tri-O-benzyl-α-D-glucopyranosyl)-(l→4)-2,3,6-tri-O- benzyl-β-D-glucopyranoside was synthesized as follows.

Zemplen-deacetylation of methyl 2,3,6,2',3',4',6'-hepta-O-acetyl-β-maltoside gave methyl-β-D-maltoside. This compound (5.92 g) was dissolved in DMF (60 mL), α,α- dimethoxytoluene (4.97 mL) and camphorsulfonic acid (0.1 g) were added, and the mixture was stirred at 50 °C for 2 hours in vacuo. The mixture was neutralized with NaHCO, and evaporated to dryness. The residue was subjected to column chromatography (chloroform- methanol 95;5 — 9:1) to give methyl O-(4,6-O-benzylidene-α-D-glucopyranosyl)-(l—>4)-β- D-glucopyranoside (3.37 g, 45%). [α]_D +42.6 (c 1.02, methanol). Η-NMR data (CD₃OD): δ 3.53 (s, 3H, OCH,); 4.20 d, 1Η, Η-l 7^=7.7 Ηz); 5.20 (d, 1Η, Η-l' 7^=3.8 Ηz); 5.56 (s, 1Η, PhCH). ¹³C-NMR data (CD₃OD): δ 57.38 (OCΗ₃); 62.26 (C-6); 69.81 (C-6'); 65.01, 72.10, 74.60, 74.73, 76.50, 77.73, 81.65, 82.48 (C-2,3,4,5,2',3',4',5'); 103.00, 103.45 (C-l', PhCΗ); 105.31 (C-l); 127.54, 129.05, 129.93, 139.13 (Ph).

The benzylation of the 4,6-benzylidene derivative in DMF, with benzyl bromide (6.78 mL) in the presence of 80% sodium hydride (1.82 g) afforded methyl O-(2,3-di-O- benzyl-4,6-O-benzylidene-α-D-glucopyranosyl)-(l→4)-2,3,6-tri-O-benzyl-β-D- glucopyranoside (6.55 g, 96%). [α]_D +3.99 (c 1.43 chloroform). Η-NMR data (CDC1₃): δ 3.62 (s, 3Η, OCH,); 4.39 (d, 1Η, Η-l 7^=7.7 Ηz); 5.58 (s, 1Η, PhCH); 5.76 (d, 1Η, Η-l' 7^=3.8 Ηz). ¹³C-NMR data (CDC1₃): δ 56.90 (OCΗ₃); 68.78, 68.91 (C-6,6'); 63.23, 71.90, 74.19, 78.67, 78.70, 82.24, 82.33, 84.82 (C-2,3,4,5,2',3',4',5'); 73.41, 73.70, 73.79, 74.55, 75.23 (PhCH₂); 97.20 (C-l'); 101.09 (PhCH); 104.51 (C-l); 126.02-128.81 (Ph); 137.54, 137.80, 138.22, 138.27, 138.64, 138.71 (PhCH₂, PhCH).

The 4,6-benzylidene ring was cleaved as described in Example 7 to give methyl O- (2,3,6-tri-O-benzyl-α-D-glucopyranosyl)-(l—»4)-2,3,6-tri-O-benzyl-β-D-glucopyranoside (3.44 g, 52%). [α]_D +26.4° (c 1.20, chloroform). Η-NMR data (CDC1₃): δ 3.57 (s, 3H, OCH,); 4.33 (d, 1Η, Η-l 7^=7.6 Ηz); 5.68 (d, 1Η, Η-l' 7^=3.5 Ηz). ¹³C-NMR data (CDC1₃): δ 56.98 (OCΗ₃); 69.05, 69.63 (C-6,6'); 70.58, 71.32, 72.40, 74.46, 78.85, 81.22, 82.22, 84.72 (C-2,3,4,5,2',3',4',5'); 72.98, 73.27, 73.52, 73.80, 74.99, 75.25 (PhCH2); 96.51 (C-l'); 104.51 (C-l); 126.59-128.43 (Ph); 137.80, 137.90, 138.29, 138.33, 138.71 (PA). Alkylation of the above described product (0.52 g) with 2-bromoethyl O-(2,3,4,6- tetra-O-benzyl-α-D-glucopyranosyl)-(l→4)-2,3,6-tri-O-benzyl-β-D-glucopyranoside (1.00 g) in DMF (10 mL), in the presence of 80% sodium hydride afforded methyl 4-O-{4-O-[4-O- (2,3,4,6-tetra-O-benzyl-α-D-glucopyranosyl)-(l-→4)-(2,3,6-tri-O-benzyl-β-D- glucopyranosyloxyethyl)]-(2,3,6-tri-O-benzyl-α-D-glucopyranosyl}-(l*→4)-(2,3,6-tri-O- benzyl-β-D-glucopyranoside (0.37 g, 34%). [α]_D +30.0 (c 1.05, chloroform). Η-NMR data (CDCI₃): δ 5.64, 5.71 (2d, 2H, H-l", H-l"' 7=3.5 Hz, 7=3.5 Hz). ¹³C-NMR data (CDC1₃): δ 56.90 (OCHj); 68.19, 69.07 (4C, C-6,6',6",6'"); 71.77 (2C, OCH₂); 71.01, 71.09, 72.57, 74.48, 74.66, 77.70, 78.21, 79.22, 79.32 81.88, 82.00, 82.15, 82.27, 84.64, 84.68 (C- 2,3,4,5,2',3',4',5',2",3",4",5",2^,",3^,",4"^,,5^/"); 73.22, 73.26, 73.35, 73.40, 73.48, 73.83, 73.98, 74.46, 74.54, 75.00, 75.50 (13 PhCH₂, OCH₂); 96.73, 96.90 (C-l',1'"); 103.57 (C-l"); 104.53 (C-l); 126.63-128.39 (Ph); 137.93, 137.96, 138.00, 138.06, 138.27, 138.40, 138.43, 138.66, 138.75, 138.84 (PhCH₂).

The tethered tetrasaccharide (0.210 g) was hydrogenated with 10% palladium on charcoal catalyst (0.5 g) in a mixture of methanol (10 mL), acetic acid (2 mL), water (2 mL) at atmospheric pressure for four days, to give methyl 4-O-{4-O-[4-O-(α-D- glucopyranosyl)-(l— >4)-β-D-glucopyranosyloxyethyl]-α-D-glucopyranosyl }-( l*→4)-β-D- glucopyranoside (0.04 g, 50%). [α]_D +3.7 (c 1.01, water). Η-NMR data (D₂O): δ 3.51(s, 3H, OCH,); 4.33, 4.46 (2d, 2Η, H-1,1" 7=7.8 Hz and 7=8.1 Hz, respectively); 5.33, 5.35 (2d, 2H, H-l',1"' 7=3.5 Hz and 7=3.6 Hz, respectively). ¹³C-NMR data (D₂O): δ 57.42 (OCH₃); 60.41, 60.75, 60.93, 60.98 (C-6,6',6",6"'); 69.42 (2C, OCH₂); 69.59, 71.88, 71.93, 72.97, 73.05, 73.10, 73.22, 73.28, 74.77, 74.81, 76.43, 76.48, 77.02, 77.08, 77.10, 78.44 (16C, C-2,3,4,5,2',3',4',5',2",3",4",5",2"',3^,",4^,",5'"); 99.73, 99.86 (C-l',1"'); 102.30, 103.34 (C-1,1").

This compound was sulfated as described in Example 1 to give methyl 4-O-{4-O- [4-O-(α-D-glucopyranosyl)-(l*→4)-β-D-glucopyranosyloxyethyl]-α-D-glucopyranosyl}- (l→4)-β-D-glucopyranoside sulfate (PF#8). ¹³C-NMR-data (D₂O 300): δ 56.85 (OCH₃); 66.30, 67.79, 68.72 (C-6,6',6",6"'); 71.60 (OCH₂); 69.79, 70.32, 70.46, 71.80, 72.25, 72.36, 72.56, 73.39, 73.52, 74.86, 75.01, 76.05, 76.54 (C-

2,3,4,5,2',3',4,'5',2",3",4",5",2'",3^/",4^,",5^/"); 94.50, 95.37, 100.68 (C-l,l',l",l"'). Anal. Calc. for C^ sSuNan _x 8H₂O: C, 16.28; H, 2.68; S, 17.77. Found: C, 16.26; H, 2.66; S, 18.28.

Example 9 2.7-Naphthyl bis(O-β-D-galactopyranosyl-(l— >4)-β-D-glucopyranoside) sulfate (PF#9) To a mixture of 2,3,6-tri-O-benzoyl-4-O-(2,3,4,6-tetra-O-benzoyl-β-D- galactopyranosyl)-α-D-glucopyranosyl bromide (2.30g), 2,7-dihydroxynaphthalene (0.160g) and 4 A molecular sieves powder (2g) in dichloromethane (10 mL), a solution of silver trifluoromethanesulfonate (0.642g) and 2,4,6-collidine (0.02 mL) in dichloromethane- toluene (1:1, 20 mL) was added dropwise with stirring at -30 °C. The reaction was stopped by the addition of pyridine, the mixture was diluted with dichloromethane and filtered through a pad of Celite. The filtrate was succesively washed with water, 10% aqueous Na₂S₂O₃, water, 2M HC1, water, aqueous NaHCO₃, water, then was dried and evaporated. Column chromatography (toluene-ethyl acetate 9:1) gave 2,7-naphthyl bis[O-(2,3,4,6-tetra- O-benzoyl-β-D-galactopyranosyl)-(l—»4)-2,3,6-tri-O-benzoyl-β-D-glucopyranoside] (1.61g, 71%). [α]_D +42.2° (c 1.00 chloroform). Η-NMR data (CDC1₃): δ 4.92 (d, IH, H-l' 7^=7.8 Hz); 5.32 (d, IH, H-l 7^=7.4 Hz). ¹³C-NMR data (CDC1₃): δ 61.04, 62.61 (C-6,6'); 67.54, 69.91, 71.45, 71.82, 72.81, 73.29, 76.33 (C-2,3,4,5,2',3',4',5'); 98.82, 101.23 (C-1,1'); 110.58 (C, Ar); 117.38 (C₃ Ar); 128.87 (C₁₀ Ar); 129.41 (C₄ Ar); 134.82 (C₉ Ar); 155.06 (C₂ Ar); 164.84, 165.18, 165.23, 165.42, 165.57, 165.69 (COPh).

The benzoate (1.61g) was debenzoylated as described in Example 2, the product was purified by column chromatography (chloroform-90% aqueous methanol 1:1) to give 0.45g (78%) 2,7-naphthyl bis(O-β-D-galactopyranosyl-(l→4)-β-D-glucopyranoside). [α]_D - 6.60 (c 1.0, pyridine). Η-NMR data (D₂O): δ 4.66 (d, IH, H-l' 7^=7.1 Hz); 5.33 (d, IH, H-l 7^=7.8 Hz); 7.38 (dd, IH, Ar); 7.57 (d, IH, Ar); 7.94 (d, IH, Ar). ¹³C-NMR data (D₂O): δ 61.04, 62.06 (C-6,6'); 69.67, 72.06, 73.77, 75.43, 76.03, 76.44, 79.30 (C- 2,3,4,5,2',3',4',5'); 101.21 (C-l'); 104.12 (C-l); 111.10 (C, Ar); 117.95 (C₃ Ar); 127.29 (C₁₀ Ar); 130.40 (C₄ Ar); 136.15 (C₉ Ar); 156.38 (C₂ Ar).

The compound was sulfated as described in Example 1 to give 2,7-naphthyl bis(O- β-D-galactopyranosyl-(l→4)-β-D-glucopyranoside) sulfate (PF#9). Η-NMR data (D₂O): δ 5.74 (d, IH, H-l 7^=6.0 Hz); 7.35 (d, IH, Ar); 7.59 (s, IH, Ar); 7.94 (d, IH, Ar). ¹³C- NMR data (D,O): δ 66.16, 66.76 (C-6,6'); 72.05, 73.10, 75.10, 75.34, 75.70, 75.92, 77.30, 77.92 (C-2,3,4,5,2',3',4',5'); 98.54 (C-l'); 101.108 (C-l); 111.16 ( Ar); 117.58 (C₃ Ar); 126.69 (C₉ Ar); 129.87 (C₄ Ar); 135.15 (C₁₀ Ar); 155.08 (C₂ Λr). Anal. Calc. for C^A^Na,, x 14H₂O: C, 16.40; H, 2.50; S, 18.03. Found: C, 16.14; H, 2.77; S, 17.43. Example 10 2.7-Naphthyl bis(O-β-D-galactopyranosyl-(l→3)-α-D-arabinopyranoside) sulfate (PF#10) The glycosylation was performed as described in Example 9 using 2,7- dihydroxynaphthalene (0.160g), 2,4-di-O-benzoyl-3-O-(2,3,4,6-tetra-O-benzoyl-β-D- galactopyranosyl)-β-D-arabinopyranosyl bromide (2.0g), silver trifluoromethanesulfonate (0.642g) and 2,4,6-collidine 90.02 mL). Column chromatography (toluene-ethyl acetate 9:1) gave 2,7-naphthyl bis[O-(2,3,4,6-tetra-O-benzoyl-β-D-galactopyranosyl)-(l*→3)-2,4-di-O- benzoyl-α-D-arabinopyranoside] (1.14g, 57%). [α]_D -25.5° (c 0.95 chloroform). Η-NMR data (CDC1₃): δ 5.11 (d, IH, H-l' 7 _>2<=7.7 Hz); 5.46 (d, IH, H-l 7^=5.2 Hz). ¹³C-NMR data (CDC1₃): δ 61.22 (C-5); 61.76 (C-6'); 67.86, 67.88; 69.67, 70.84, 71.37, 71.76, 74.72 (C-2,3,4,2',3',4',5'); 97.97 (C-l); 100.36 (C-l'); 110.41 (C, Ar); 117.70 (C₃ Ar); 128.88 (C₁₀ Ar); 129.45 (C₄ r); 135.16 (C_g Ar); 155.08 (C, Ar); 164.75, 165.08, 165.42, 165.52, 165.68, 165.90 (COPh). The compound was debenzoylated as described in Example 2 to give 2,7-naphthyl bis(O-β-D-galactopyranosyl)-(l->3)-α-D-arabinopyranoside (0.49g, 93%), [α]_D -61.6 (c 0.99, methanol). Η-NMR data (CD₃OD): δ 4.51 (d, IH, H-l' 7^=7.5 Hz); 5.07 (d, IH, H-l 7^=6.6 Hz); 7.20 (dd, IH, Ar); 7.40 (d, IH, Ar); 7.75 (d, IH, Ar). ¹³C-NMR data (CD₃OD): δ 62.46 (C-5); 67.01 (C-6'); 67.23, 70.16, 70.38, 72.21, 74.25, 76.82, 80.74 (C- 2,3,4,2',3',4',5'); 102.30, 102.42 (C-1,1'); 111.31 (C, Ar); 118.26 (C₃ Ar); 127.42 (C₁₀ Ar); 130.31 (C Ar); 136.51 (C₉ Ar); 156.81 (C₂ Ar).

Sulfation as described in Example 1 gave 2,7-naphthyl bis(O-β-D-galactopyranosyl)- (l→3)-α-D-arabinopyranoside) sulfate (PF#10). Η-NMR data (D₂O): δ 5.07 (d, IH, H-l' 7^=6.5 Hz); 5.82 (d, IH, H-l, 7^=2.0 Hz); 7.42 (dd, IH, Ar); 7.61 (s, IH, Ar); 7.95 (d, IH, Ar). ¹³C-NMR data (D₂O): δ 58.03 (C-5); 66.77 (C-6'); 71.11, 72.15, 73.61, 75.35, 75.45, 75.65, 76.08 (C-2,3,4,2',3',4',5'); 96.50 (C-l); 101.40 (C-l'); 111.84 (C_ Ar); 118.81 (C₃ Ar); 126.76 (C₉ Ar); 129.92 (C₄ Ar); 135.14 (C₁₀ Ar); 154.25 (C₂ Ar). Anal. Calc. for C₃₂H₃₂O₅₆S₁₂Na₁₂ x 12H₂O: C, 17.55; H, 2.57; S, 17.57. Found: C, 18.36; H, 2.65; S, 17.68. Example 11 1,5-Naphthyl bis(O-β-D-galactopyranosyl)-(l—>3)-α-D-arabinopyranoside) sulfate (PF#11) The glycosylation was performed as described in Example 9, using 1,5- dihydroxynaphthalene (0.160 g) 2,4-di-O-benzoyl-3-O-(2,3,4,6-tetra-O-benzoyl-β-D- galactopyranosyl)-β-D-arabinopyranosyl bromide (2.0 g), silver trifluoromethanesulfonate (0.642 g) and 2,4,6-collidine (0.02 mL). The product, 1,5-naphthyl bis[O-(2,3,4,6-tetra-O- benzoyl-β-D-galactopyranosyl)-(l→3)-2,4-di-O-benzoyl-α-D-arabinopyranoside] (0.661 g, 31%) was isolated by column chromatography (toluene-ethyl acetate 9:1). It had [α]_D -28.3° (c 1.00, chloroform). Η-NMR data (CDC1₃): δ 5.17 (d, IH, H-l' 7^=7.9 Hz); 5.41 (d, IH, H-l 7^=5.5 Hz). ¹³C-NMR data (CDC1₃): δ 61.83 (C-5,6'); 67.85, 67.88, 69.62, 70.78, 71.37, 72.01, 75.52 (C-2,3,4,2',3',4',5'); 98.93 (C-l); 100.43 (C-l'); 109.80 (C, Ar); 117.09 (C₄ Ar); 125.49 (C₃ Ar); 127.15, 128.62 (2C, C₉,C₁₀ r); 152.32 (C. Ar); 164.66, 165.26, 165.43, 165.49, 165.76, 165.88 (COPh). Debenzoylation of the product as described in Example 2 gave 1,5-naphthyl bis(O- β-D-galactopyranosyl)-(l→3)-α-D-arabinopyranoside) (0.27 g, 95%), [α]_D -65.8 (c 0.96, methanol). Η-NMR data (CD₃OD): δ 4.50 (d, IH, H-l 7^=7.5 Hz); 5.05 (d, IH, H-l' 7^=6.5 Hz). ¹³C-NMR data (CD₃OD): δ 62.32 (C-5); 66.98 (C-6'); 67.15, 70.10, 70.25, 72.21, 74.22, 76.84, 80.76 (C-2,3,4,2',3',4',5'); 102.36, 102.42 (C-1,1'). Sulfation as described in Example 1 gave 1,5-naphthyl bis(O-β-D-galactopyranosyl)-

(l-»3)-α-D-arabinopyranoside) sulfate (PF#11).

Example 12 Bis(4-O-β-D-galactopyranosyl-(l— »4)-β-D-glucopyranosyl)-l,3 -dithio-benzene sulfate (PF#12)

A mixture of 1,3-benzenedithiol (0.142 g) and 80% sodium hydride (0.048 g) in DMF (3 mL) was stirred at 0 °C and 2,3,6-tri-O-benzoyl-4-O-(2,3,4,6-tetra-O-benzoyl-β-D- galactopyranosyl)-oc-D-glucopyranosyl bromide (2.3 g) in DMF (8 mL) was added dropwise. The mixture was neutralized with AG 50W-X8 [H⁺] resin, the resin was filtered off, the filtrate was diluted with chloroform and washed with water and then concentrated. Column chromatography (toluene-ethyl acetate 9:1) gave bis[4-O-(2,3,4,6-tetra-O-benzoyl- β-D-galactopyranosyl)-(l→4)-(2,3,6-tri-O-benzoyl-β-D-glucopyranosyl)-l,3-dithio-benzene (1.48 g, 67%). [α]_D +36.4 (c 1.00, chloroform). Η-NMR data (CDC1₃): δ 4.84 (d, IH, H-l' 7^=7.7 Hz); 4.87 (d, IH, H-l 7^=8.8 Hz). ¹³C-NMR data (CDC1₃): δ 60.91, 62.51 (C- 6,6'); 67.43, 69.96, 70.36, 71.36, 71.77, 73.93, 75.91, 77.00 (C-2,3,4,5,2',3',4',5'); 85.78 (C- 1); 101.04 (C-l'); 125.29-136.32 (Ph); 164.78, 165.08, 165.20, 165.34, 165.38, 165.52, 165.71 (COPh).

The compound was debenzoylated as described in Example 2, to give bis(4-O-β-D- galactopyranosyl-(l— >4)-β-D-glucopyranosyl)-l,3-dithio-benzene (0.50 g, 94%). [α]_D -35.13 (c 1.02, methanol). Η-NMR data (D₂O): δ 4.35 (d, IH, H-l' 7^=7.6 Hz); 4.71 (d, IH, H- 1 7^=9.9 Hz); 7.25-7.45 (m, 3H, Ph). ¹³C-NMR data (D₂O): δ 60.62, 61.45 (C-6,6'); 69.05, 71.42, 72.03, 73.10, 75.81, 76.36, 78.46, 79.25 (C-2,3,4,5,2',3',4',5'); 87.45 (C-l); 103.41 (C-l').

Sulfation as described in Example 1 gave bis(4-O-β-D-galactopyranosyl-(l— >4)-β-D- glucoρyranosyl)-l,3-dithio-benzene sulfate (PF#12). Η-NMR data (D₂O): δ 5.35 (d, IH, H-l 7^₂=7.3 Hz); 7.38-7.66 (m, 3H, Ph). ¹³-NMR data (D₂O): δ 66.40 (C-6,6'); 71.82, 75.07, 75.38, 75.76, 75.98, 76.40. 77.60 (C-2,3,4,5,2',3',4',5'); 84.69 (C-l); 101.39 (C-l'). Anal. Calc. for C₃₀H₃₂O₆₂S₁₆Na₁₄ x 14H₂O: C, 14.57; H, 2.44; S, 20.75. Found: C, 14.46; H, 2.41; S, 20.32.

Example 13 Biological Activity Of Sulfated Bridged Mono- And Oligosaccharides Several of the sulfated bridged mono- and oligosaccharides of the present invention were tested for their antithrombotic, anticoagulant, antiangiogenic and anti-inflammatory activity.

The UC polysaccharide binding assay (UC-PBA) tests for the interaction of heparin- like molecules with the basic fibroblast growth factor. The assay operates by measuring the ability of heparin-like molecules to inhibit the interaction of transfected human lymphoblastoid cells UC 729-6 (UC) with plates coated with basic fibroblast growth factor. The UC-PBA assay is described in Ishihara et al. Anal. Biochem. (1992) 202:310 which is incorporated herein by reference.

The ACE cell growth inhibition assay (ACE-CGI) tests for an agent's ability to inhibit growth of adrenocortical endothelial (ACE) cells. The ACE-CGI assay is described in Ishihara et al. J. Biol Chem. (1992) 268:4675 which is incorporated herein by reference. The ACE growth stimulation assay (ACE-CGS) tests for an agent's ability to restore the mitogenic activity of basic fibroblast growth factor to chlorate treated ACE cells. The ACE-CGS assay is described in Ishihara et al. J. Biol Chem. (1992) 268:4675 which is incorporated herein by reference. The activated partial thromboplastin time (APTT) assay is a citrated plasma-based clotting assay based on the activation of the intrinsic system through factor XII using either a chemical or particulate activator. The APTT assay is described in depth in J. M. Walenga et al., "In Vitro Evaluation Of Heparin Fractions: Old vs. New Methods" CRC Critical Reviews in Clinical Laboratory Systems 22 at 362 which is incorporated herein by reference.

The results of these tests are provided in Table 1.

TABLE 1

Biolo gical Test Results Of Bridged Sulfatoids

COMPOUND UC-PBA ACE-CGI ACE-CGS APTT

[IC₅₀] [IC₅₀]

PF#1 >50 nd nd 0.5%

PF#2 >50 nd nd 0.2%

PF#3 2 >64 ia 10.1%

PF#4 6 >64 ia 0.9%

PF#5 <1 8 a 17.1%

PF#6 1 nd nd 13.2%

PF#7 <1 nd nd 21.3%

PF#12 <1 nd nd 6.4%

nd: not determined ia: inactive a: active

While the present invention is disclosed by reference to the details of above examples, it is to be understood that this disclosure is intended in an illustrative rather than limiting sense, as it is contemplated that modifications will readily occur to those skilled in the art, within the spirit of the invention and the scope of the appended claims.

Claims

What is claimed is:

1. A bridged saccharide compound, comprising the structure:

R'-P-^-R X^R³ wherein R¹, R² and R³ are independently one or more saccharide(s); X¹ and X² are independently difunctional or polyfunctional alkyl, aryl or aralkyl compounds capable of covalently joining together said saccharides; and n is an integer of zero to ten.

2. The bridged saccharide compound of Claim 1 wherein said saccharide(s) R\ R² and/or R³ comprise sulfateable hydroxyl or amine groups, and at least certain of said hydroxyl and amine groups are sulfated, and said covalent joining of said saccharides is by an ether, thioether, glycosidic or thioglycosidic bond.

3. The bridged saccharide compound of Claim 2 wherein certain of said amine groups are sulfated.

4. The bridged saccharide compound of Claim 2 wherein certain of said hydroxyl groups are sulfated.

5. The bridged saccharide compound of Claim 2 wherein n is greater than zero and X¹ and X² join said saccharide(s) R\ R² and R³ by a glycosidic bond.

6. The bridged saccharide compound of Claim 2 wherein n = 0 and X² joins said saccharide(s) R¹ and R³ by a glycosidic bond and/or an ether bond.

7. The bridged saccharide compound of Claim 2 wherein said saccharide(s) R¹,

R² and R³ are glucose, and X¹ and X² are ethylene glycol or hexaethylene glycol.

8. The bridged saccharide compound of Claim 2 wherein n is greater than zero,

R¹, R², and R³ are a-D-glucopyranosyl (l→4)-β-D-glucopyranoside and X¹ and X² is ethylene glycol or O[CH₂CH₂O]₆.

9. The bridged saccharide compound of Claim 8 wherein X¹ and X² joins said saccharide(s) R¹, R² and R³ by a glycosidic bond.

10. The compounds ethane- 1,2-diyl bis(β-D-glucopyranoside) or ethane- 1,2-diyl bis(β-D-glucopyranoside) sulfate.

11. The compounds 3,6,9,12,15-Pentaoxa-heptadecane-l,17-diyl bis(β-D- glucopyranoside) or 3,6,9,12,15-Pentaoxa-heptadecane-l,17-diyl bis(β-D-glucopyranoside) sulfate.

12. The compounds ethane- 1,2-diyl bis(O-α-D-glucopyranosyl-(l-+4)-β-D- glucopyranoside) or ethane- 1,2-diyl bis(O-α-D-glucopyranosyl-(l_→4)-β-D-glucopyranoside) sulfate.

13. The compounds 3,6,9,12,15-Pentaoxa-heptadecane-l,17-diyl bis(O-oc-D- glucopyranosyl-(l-→4)-β-D-glucopyranoside) or 3,6,9, 12,15-Pentaoxa-heptadecane-l,17-diyl bis(O-α-D-glucopyranosyl-( l→4)-β-D-glucopyranoside) sulfate.

14. The compounds ethane- 1,2-diyl bis(O- -D-glucopyranosyl-(l_→4)-O-α-D- glucopyranosyl-(l→4)-β- D- glucopyranoside) or ethane- 1,2-diyl bis(O-α-D-glucopyranosyl- (l_+4)-0-α-D-glucopyranosyl-(l→4)-β- D-glucopyranoside) sulfate. 15. The compounds 3,6,9,12,15-Pentaoxa-heptadecane-l,17-diyl bis(O-α-D- glucopyranosyl-(l-→4)-O-α-D-glucopyranosyl-(l→4)-β- D-glucopyranoside) or 3,6,9,12,

15- Pentaoxa-heptadecane- 1 , 17-diyl bis(O-α-D-glucopyranosyl-( 1 →4)-O-α-D-glucopyranosyl- (l→4)-β-D-glucopyranoside) sulfate.

16. The compounds ethane- 1,2-diyl bis(O-α-D-glucopyranosyl-(l_→4)-O-α-D- glucopyranosyl-(l→4)-β- D-glucopyranosyl-(l_→4)-O-α-D-glucopyranosyl-(l_→4)-O-β-D- glucopyranoside) or ethane- 1,2-diyl bis(O-α-D-glucopyranosyl-(l_→4)-O-α-D- glucopyranosyl-(l-→4)-β- D-glucopyranosyl-(l— *4)-O-α-D-glucopyranosyl-(l→4)-O-β-D- glucopyranoside) sulfate.

17. The compounds methyl 4-O-{4-O-[4-O-(α-D-glucopyranosyl)-(l-→4)-β-D- glucopyranosyloxyethyl]-α-D-glucopyranosyl}-(l→4)-β-D-glucopyranoside or methyl 4-O-

{4-O-[4-O-(α-D-glucopyranosyl)-(l→4)-β-D-glucopyranosyloxyethyl]-α-D-glucopyranosyl}- ( l→4)-β-D-glucopyranoside sulfate.

18. The compounds 2,7-Naphthyl bis(O-β-D-galactopyranosyl-(l_→4)-β-D- glucopyranoside) or 2,7-Naphthyl bis(O-β-D-galactopyranosyl-(l_→4)-β-D-glucopyfanoside) sulfate.

19. The compounds 2,7-naphthyl bis(O-β-D-galactopyranosyl-(l_→3)-α-D- arabinopyranoside) or 2,7-naphthyl bis(O-β-D-galactopyranosyl-(l_→3)-α-D- arabinopyranoside) sulfate.

20. The compounds 1,5-naphthyl bis(O-β-D-galactopyranosyl)-(l-→3)- -D- arabinopyranoside) or 1,5-naphthyl bis(O-β-D-galactopyranosyl)-(l-→3)-α-D- arabinopyranoside) sulfate.

21. The compounds bis(4-O-β-D-galactopyranosyl-(l→4)-β-D-glucopyranosyl)- 1,3-dithio-benzene or bis(4-O-β-D-galactopyranosyl-(l→4)-β-D-glucopyranosyl)-l,3-dithio- benzene sulfate.

22. A pharmaceutical composition comprising a bridged saccharide compound of Claim 1.

23. A pharmaceutical composition comprising a bridged saccharide compound of Claim 2.

24. A method for treating disease wherein said disease is beneficially treated with heparin, comprising administering an effective amount of the compound of Claim 1 to treat said disease.

25. A method for treating disease wherein said disease is beneficially treated with heparin, comprising administering an effective amount of the compound of Claim 2 to treat said disease.