NZ242431A - Mixture of sulphated oligosaccharides derived from depolymerised heparin, and pharmaceutical compositions - Google Patents
Mixture of sulphated oligosaccharides derived from depolymerised heparin, and pharmaceutical compositionsInfo
- Publication number
- NZ242431A NZ242431A NZ242431A NZ24243192A NZ242431A NZ 242431 A NZ242431 A NZ 242431A NZ 242431 A NZ242431 A NZ 242431A NZ 24243192 A NZ24243192 A NZ 24243192A NZ 242431 A NZ242431 A NZ 242431A
- Authority
- NZ
- New Zealand
- Prior art keywords
- heparin
- mixture
- depolymerised
- oligosaccharides
- gel
- Prior art date
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Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/70—Carbohydrates; Sugars; Derivatives thereof
- A61K31/715—Polysaccharides, i.e. having more than five saccharide radicals attached to each other by glycosidic linkages; Derivatives thereof, e.g. ethers, esters
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K31/00—Medicinal preparations containing organic active ingredients
- A61K31/70—Carbohydrates; Sugars; Derivatives thereof
- A61K31/715—Polysaccharides, i.e. having more than five saccharide radicals attached to each other by glycosidic linkages; Derivatives thereof, e.g. ethers, esters
- A61K31/726—Glycosaminoglycans, i.e. mucopolysaccharides
- A61K31/727—Heparin; Heparan
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
- A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P7/00—Drugs for disorders of the blood or the extracellular fluid
- A61P7/02—Antithrombotic agents; Anticoagulants; Platelet aggregation inhibitors
-
- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08B—POLYSACCHARIDES; DERIVATIVES THEREOF
- C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
- C08B37/006—Heteroglycans, i.e. polysaccharides having more than one sugar residue in the main chain in either alternating or less regular sequence; Gellans; Succinoglycans; Arabinogalactans; Tragacanth or gum tragacanth or traganth from Astragalus; Gum Karaya from Sterculia urens; Gum Ghatti from Anogeissus latifolia; Derivatives thereof
- C08B37/0063—Glycosaminoglycans or mucopolysaccharides, e.g. keratan sulfate; Derivatives thereof, e.g. fucoidan
- C08B37/0075—Heparin; Heparan sulfate; Derivatives thereof, e.g. heparosan; Purification or extraction methods thereof
- C08B37/0078—Degradation products
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- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- General Health & Medical Sciences (AREA)
- Medicinal Chemistry (AREA)
- Pharmacology & Pharmacy (AREA)
- Veterinary Medicine (AREA)
- Public Health (AREA)
- Animal Behavior & Ethology (AREA)
- Molecular Biology (AREA)
- Epidemiology (AREA)
- Organic Chemistry (AREA)
- Engineering & Computer Science (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Diabetes (AREA)
- Hematology (AREA)
- General Chemical & Material Sciences (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Biochemistry (AREA)
- Materials Engineering (AREA)
- Polymers & Plastics (AREA)
- Dermatology (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
- Polysaccharides And Polysaccharide Derivatives (AREA)
Description
New Zealand Paient Spedficaiion for Paient Number £42431
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242431
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NEW ZEALAND PATENTS ACT, 1953
No.: Date:
. ?ATErtT Gf;
22 APR 199.
'IFCEIV'O
COMPLETE SPECIFICATION SULPHATED POLYSACCHARIDES. THEIR PREPARATION. PHARMACEUTICAL COMPOSITIONS CONTAINING THEM AND THEIR
USE
We, RHONE-POULENC RORER SA, a French body corporate, of 20 Avenue Raymond ARon, F 92165, Antony, France hereby declare the invention for which we pray that a patent may be granted to us, and the method by which it is to be performed, to be particularly described in and by the following statement:-
24 2 4
The present invention relates to low-molecular weight polysaccharides, and, more specifically, to oligosaccharide compositions possessing useful pharmacological properties.
Generally, antithrombotic treatments require the use of two main categories of agents, namely anticoagulatory agents and antiplatelet agents.
Antivitamin K compounds constitute a very important family of the anticoagulatory agents. Given that 10 these compounds are active via the oral route, they are used in numerous indications. However, their use is still limited by certain disadvantages and in particular the risks of haemorrhages caused by them and the difficulty of adapting the dosage to long-term treatment. 15 Heparins constitute a second category of anticoagulatory agents. They are biological substances of the glycosaminoglycan family obtained by extraction, which are oligosaccharide compounds with varying chain lengths and degrees of sulphation. Heparins are used in various 20 types of thromboses, in particular in the treatment or prevention of venous thromboses, optionally combined with other therapies.
The disadvantage of heparins lies in their high anticoagulatory activity which may cause haemorrhages, and 25 in their sensitivity to certain serum factors such as pf4, which requires the use of relatively high doses.
Moreover, heparins are very heterogeneous products. It is therefore difficult to evaluate their mode
of action, to assess the contribution of each of the components in the overall activity of heparin, and, consequently, to increase the antithrombotic activity without increasing the side effects.
A first solution to the abovementioned disadvantages has been provided by low-molecular weight heparins. These heparins are obtained by fragmentation (depolymerisation) of oligosaccharide chains using chemical or enzymatic agents. In particular, depolymerisation has 10 been described by a treatment of a heparin ester in the presence of a strong base (EP 40144). It may also be carried out by treating heparin in the presence of nitrous acid, or by the action of a heparinase (EP 64452). These various methods lead to mixtures of oligosaccharides having 15 the general structure of the polysaccharides which make up heparin, but having a mean molecular weight which is smaller by weight. More particularly, the investigations were directed mainly towards heparin-derived mixtures having very short oligosaccharide chains. Thus, Patent EP 20 27089 indicates that heparin-derived oligosaccharide mixtures containing not more than 8 saccharide units possess an antithrombotic specific activity which is greater than heparin. Similarly, hexasaccharides have been prepared and their antithrombotic properties studied (EP 25 64452). More recent Patents, EP 84 999 and EP 301 618, on heparin-derived polysaccharides such as hexa-, penta- and tetrasaccharides, may also be mentioned.
However, the products described so' far have not
2 4 51
enabled the problems encountered with heparins to be resolved in a completely satisfactory manner. In particular, it has not been possible to confirm in vivo the correlation between the mean molecular mass of the products 5 and their side effects.
The applicant has now shown that it is possible to obtain, from native or depolymerised heparins, oligosaccharide mixtures having greatly improved antithrombin properties, and therefore better therapeutic 10 potential.
In effect, the applicant has shown, unexpectedly, that a substantial part of the antithrombotic activity of heparin is present in a small and homogeneous fraction.
The present invention results more particularly 15 from the identification of monodisperse fractions of heparin having a mean molecular mass of about 6 kD and possessing a high antithrombin activity.
As illustrated in the examples, it is therefore possible to obtain mixtures possessing a particularly high 20 antithrombotic activity by calibrating the molecular mass and by reducing the polydispersity.
One subject of the invention is a mixture of sulphated oligosaccharides having the general structure of the constituent oligosaccharides of heparin, the said 25 mixture having a weight average molecular mass of 6 + 0.6 kD and a polydispersity of about l, and possessing the capacity to inhibit the generation of thrombin.
The polydispersity corresponds to the ratio of ^
/■v i;
24 2 4 31
the weight average molecular weight of the mixture to its number average molecular weight. It is a measure of the molecular homogeneity of the mixture. The closer this value is to 1, the more homogeneous is the mixture.
In addition to their antithrombin properties, the mixtures of the invention possess particularly advantageous pharmacokinetic properties. More particularly, compared with native heparin and its depolymerised forms, the mixtures of the invention exhibit a lower sensitivity to serum factors such as pf4, which increases their therapeutic potential.
Other advantages of the mixtures of the invention lie in particular in the reduction of certain undesirable side effects such as:
- thrombocytopenic effect. One of the disadvantages of the known mixtures derived from heparin stems from the drop in the number of platelets which they can bring about. This undesirable effect is substantially reduced when the mixtures of the invention are used.
- immunogenic reactions. When such reactions are too intense, it is evident that the therapeutic efficacy of the products is reduced. The weak immunogenicity of the mixtures of the invention constitutes another of their very advantageous pharmacological characteristics.
Furthermore, the mixtures of the invention have excellent plasmatic bioavailability and half-life.
The properties described above permit a particularly effective pharmacological use especially in
242431
the prophylaxis and treatment of venous or arterial thromboses. Moreover, they should permit the use of higher doses in vivo without increasing the risks of haemorrhage.
In a preferred mode, the mixtures of the 5 invention are more particularly depolymerised heparin fractions.
As indicated above, the depolymerised heparin may be obtained by any chemical, enzymatic or other technique known to a person skilled in the art, which enable the 10 oligosaccharide chains of heparin to be fragmented. In particular, the methods described in Patents EP 40144, EP 64452, EP 37319 or EP 337327 are suitable for use in the invention.
Preferably, the mixtures of the invention consist 15 of oligosaccharides having a 2-0-sulpho-4-enopyranosuronic acid at one of their ends.
A particularly advantageous mixture consists of a heparin fraction which has been depolymerised by the action of a base on a heparin ester.
The antithrombin activity of the mixtures of the invention may be demonstrated in a test in which the generation of thrombin is initiated in the presence of human thromboplastin (extrinsic route) or by contact (intrinsic route). Such a test has been described 25 previously (Hemker et al., Thromb. Haemostas. 56, 9-17, 1986).
This activity may be expressed quantitatively as the amount of product required for 25 % inhibition of the
2 & ? 4 '3 4
generation of thrombin. Thus, the increase in activity of the mixtures of the invention is clearly evident since their specific antithrombin activity in vitro is surprisingly increased by a factor above 100 % compared 5 with the heparin starting material. Taking into account the particularly advantageous pharmacokinetic properties of the mixtures of the invention, this increase in specific activity is even greater in vivo.
More particularly, the mixtures of the invention 10 permit, in a test carried out on plasma substantially free of platelets, a
% inhibition of the generation of thrombin at concentrations below 300 ng/ml.
Another subject of the invention is a method of preparing a mixture as defined above, which comprises 15 ^fractionating a heparin or a depolymerised heparin by gel filtration.
The process of the invention brings into play several parameters whose control makes it possible to calibrate the molecular mass of the final mixture and to 20 determine its polydispersity. These parameters are in particular the ionic strength of the eluant and the nature of the support used.
More preferably, the fractionation comprises, successively, the stages of (i) dissolving the starting 25 heparin or depolymerised heparin in the eluant, (ii)
passing the solution thus obtained through at least one column containing the solid support for the gel filtration, equilibrated beforehand with the same eluant, and (iii)
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0
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24 2 4 31
recovering the fractions of the desired molecular weight. The starting material is preferably depolymerised heparin.
Even more preferably, a heparin depolymerised by the action of a base on a heparin ester is used. In 5 particular, the depolymerisation may be carried out in an aqueous medium or in an inert organic solvent under the action of an organic or inorganic base such as for example sodium or potassium hydroxide, an alkali metal carbonate or a tertiary amine (triethylamine, triethylenediamine and the 10 like). The action of the base on the ester makes it possible to carry out a partial and controlled depolymerisation of the heparin without modifying its general structure.
The depolymerisation conditions described in 15 Patent EP 40144 may be used to produce the starting material of the present invention.
Various types of saline solutions such as solutions of sodium chloride may be mentioned as eluant which may be used in the method of the invention. However, 20 the applicant has shown that in order to obtain fractions with the best qualities, it is particularly advantageous to carry out the fractionation using an eluant chosen from phosphate buffers such as in particular potassium phosphate, sodium phosphate or NH4H2P04. It is also possible 25 to use NaC104 or NH4N03 solutions which make it possible to obtain mixtures with excellent characteristics.
The concentration of the eluant, and therefore its ionic strength, are adjusted to the final mixture
24 2 4 31
desired. In particular, the concentration of the eluant is advantageously less than 1M and, even more preferably,
between 0.1 and 0.5 M.
When a phosphate buffer is used, it is 5 particularly advantageous to carry out the procedure at concentrations of about 0.2 M.
In the second stage of the method of the invention, the support used is generally chosen as a function of the mean molecular mass of the starting mixture 10 (native or depolymerised heparin and the like), of the final product desired and of the behaviour of the starting mixture in the eluant used. Advantageously, a polyacrylamide-agarose type gel is used as support. The gels AcA 54, AcA 202, Sephadex G-25 or G-50 or 15 alternatively Biogel P30, which give excellent results, may be mentioned by way of example.
In a first particularly advantageous embodiment of the method of the invention, the solid support is divided among several columns arranged in series, during 20 the second stage of the fractionation. This variant of the invention makes it possible to use substantial final amounts of gel filtration support without the disadvantages of the prior art, namely, essentially, the phenomena of settling. Thus, the separation is substantially more 25 distinct, including in the high molecular weight range, in a single fractionation operation, and the supports are more easily regenerated.
The number of columns used is adjusted by a
person skilled in the art as a function of the volume and the nature of the gel used so as to obtain the best balance between efficiency of the separation and the adverse effect due to the settling of the gel.
For practical considerations relating to the implementation, the preferred number of columns generally used in the second stage of the process is less than 20. By way of illustration, 40 litres of AcA 202 gel may be divided into 10 4-litre columns.
In another particularly advantageous embodiment of the method of the invention, at least 2 types of supports having differing separation characteristics are used successively in the second stage of the fractionation. This variant of the invention makes it possible to obtain a 15 final fractionation of better quality. By way of example, the fractionation may be carried out on the following sequence of gels: AcA 202 - AcA 54 - AcA 202.
For a better implementation of the invention, it is important to use high amounts of gel so as to achieve a 20 more distinct separation and to obtain greater homogeneity. However, given the fairly slow flow rates used for this type of gel filtration, the gel volume should be adapted to the amount of product to be separated so as to obtain the best equilibrium between the separation and the effect of 25 longitudinal diffusion.
Advantageously, in the method of the invention, the starting heparin (g)/ gel volume (1) ratio is less than 2, and more preferably between 0.5 and 1.5.
£c 4 £ 3
11
The invention also relates to a method of preparation of weakly dispersed mixtures of oligosaccharides with a predetermined molecular weight which comprises fractionating heparin or depolymerised heparin by 5 gel filtration on a solid support, divided among several columns arranged in series. •
Another subject of the invention is a pharmaceutical composition having a mixture as defined 10 above as active ingredient. Such a composition may be used in a particularly advantageous manner in the prophylaxis or treatment or prevention of thrombotic accidents. More specifically, it may be used:
- in the prevention of venous thromboses in 15 situations where a risk exists,
- in the prevention of arterial thrombotic accidents, especially in the case of myocardial infarction,
- in post-operative regime, in the prevention of venous thromboses in surgical patients, or alternatively,
- in the prevention of thromboses in surgical material.
The present invention is illustrated by the following examples.
Example 1: Preparation of mixtures according to 25 the invention.
- Depolymerisation of heparin
A solution of benzethonium chloride (25 g) in water (125 ml) is added to a solution of sodium heparinate.
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t\ 17 AUG |p?j'
242431
(10 g) in water (100 ml). The product obtained at room temperature is filtered off, washed with water and then dried. The benzethonium heparinate (15 g) thus obtained is dissolved in methylene chloride (75 ml) to which benzyl 5 chloride (15 ml) is added. The solution is heated at a temperature of between 25 and 35°C for 25 hours. A 10 % solution of sodium acetate in methanol (90 ml) is then added, and the precipitated solid is filtered off, washed with methanol and dried. The heparin benzyl ester (10 g) 10 obtained in the form of a sodium salt under the conditions described above, is dissolved in water (250 ml). Sodium hydroxide (0.9 g) is added to this solution heated to about 60°C. The temperature is maintained for 1 hour 30 minutes at about 60°C and the reaction mixture is then cooled to 15 around 20°C and neutralised by adding dilute hydrochloric acid. The mixture is then adjusted to a sodium chloride concentration of 10 % and the product is precipitated in methanol (750 ml), filtered off and dried.
- Several glass columns are used for the 20 fractionation:
(a) 1 column with a diameter of 95 mm and a height of 2 m containing the AcA 202 gel (14 litres) (gel in the form of polyacrylamide-agarose beads, with a diameter of between 60 and 140 /xm) ,
(b) 1 column with a diameter of 50 mm and a height of 2 m containing the AcA 54 gel (4 litres) (gel in the form of polyacrylamide-agarose beads, with a diameter of between 60 and 140 jum)/
242431
(c) 2 columns with a diameter of 50 mm and a height of 1 m containing the AcA 202 gel (2 litres).
A solution containing heparin (20 g)
depolymerised under the conditions described above is 5 placed at the top of the column (a) and eluted using a mobile phase consisting of a 0.33M solution of NaCl at a flow rate of 210 ml/hour.
The fractions are collected at the outlet of the column (a) and loaded onto the top of the column (b). The 10 elution is carried out with the same solution and the fractions collected are passed successively through the 2 columns (c).
This treatment enables a fraction having the following characteristics to be separated efficiently and 15 recovered at the outlet of the column (c):
Molecular weight: 6100 +/- 200
Polydispersity: 1.01
Example 2:
- 10 columns with an internal diameter of 2.5 cm 20 and a height of 50 cm, each containing the AcA 2 02 gel
(about 0.25 litre), are connected in series,
- a solution containing heparin (2 g) which is depolymerised under the conditions of Example 1, is loaded onto the top of the device and eluted using a 0.2M aqueous
solution of KH2P04 at a flow rate of 0.42 ml/min,
- 113 fractions of 12.6 ml are collected starting from 21 hours.
The characteristics of these fractions are given
24 2 4 5 1
in Table 1, in which the mean molecular mass was determined by refractometry.
Example 3:
The procedure is as in Example 2: 5 - 10 columns with an internal diameter of 10 cm and a height of 50 cm, each containing the AcA 202 gel (3 to 4 litres), are connected in series,
- a solution containing heparin (30 g) which is depolymerised under the conditions of Example 1, is loaded 10 onto the top of the device and eluted using a 0.2M aqueous solution of KH2P04 at a flow rate of 6.8 ml/min.
Fractions having the desired polydispersity characteristics are obtained.
Example 4:
The antithrombin activity of the mixtures of the invention is measured on plasma stimulated by human thromboplastin (extrinsic route) or by contact (phospholipids + kaolin : intrinsic route) under the conditions described above (cf Hemker et al., mentioned 20 above). The activity is estimated by the decrease in the peak of the thrombin generation curve relative to a control carried out in the presence of buffer alone. The results are expressed as the IC25 : concentration required to obtain 25 % inhibition of the generation of thrombin. 25 Procedure:
50 mM tris-HCl buffer, 0.1 M NaCl, pH 7.35 (1/4 volume) with bovine albumin (0.5 mg/ml), containing various concentrations of test samples, is added to plasma (1
242 4 5
volume). After incubating for 5 min at 37°C, the generation of thrombin is initiated by the addition of thromboplastin (1/4 volume) 1:40 diluted in 0.1 M CaCl2 (extrinsic system) or by phospholipids (6 iXH) (20 % phosphatidylserine, 80 % 5 phosphatidylcholine) and kaolin (0.15 mg/ml) in 0.1 M CaCl2 (intrinsic system). The generation of thrombin is obtained by measuring, at regular intervals (15-30 sec), the amidolytic activity on the substrate S2238, a 405 nM chromogenic substrate specific for thrombin. Various 10 concentrations of the samples are tested in order to obtain 25 % inhibition of the control.
Results:
On plasma substantially free from platelets
1) extrinsic route Depolymerised starting heparin : IC25 = 450 ng/ml Mixture prepared in Example l : IC25 = 200 ng/ml
Activity gain: 125 %
2) intrinsic route
Depolymerised starting heparin : IC25 = 550 ng/ml Mixture prepared in Example l : IC25 = 250 ng/ml
Activity gain: 120 %
On plasma high in platelets
Depolymerised starting heparin : IC25 = 1100 ng/ml Mixture prepared in Example 1 : IC25 = 500 ng/ml
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24 2 4 3
Under the same conditions, native heparin (nondepolymerised, nonfractionated) possesses no inhibitory activity at 2500 ng/ml.
TABLE 1
FRACTION NO.
MEAN MOLECULAR MASS
POLYDISPERSITY
14-17
712
1.027
18-20
8 400
1.013
21-22
7 519
1.020
23-24
6 986
1.011
26-27
6 365
1.008
28-31
874
1.009
32-35
295
1.011
36-40
4 761
1.012
42-46
4 192
1.013
48-53
3 608
1.016
56-61
2 988
1.019
64-70
2 359
1.023
75-80
1 758
1.029
83-85
1 476
1.028
88-94
1 176
1.027
Claims (22)
1. A mixture of sulphated oligosaccharides having the general structure of the constituent oligosaccharides of heparin, the said mixture having a 5 weight average molecular mass of 6 ± 0.6 kD and a polydispersity of about 1, and possessing the capacity to inhibit the generation of thrombin.
2. The mixture according to claim 1, which is a depolymerised heparin fraction. 10
3. The mixture according to claim 2, which consists of oligosaccharides having a 2-0-sulpho-4-enopyranosuronic acid at one of their ends.
4. The mixture according to claim 3, which is a heparin fraction depolymerised by the action of a base on a 15 heparin ester.
5. The mixture according to any one of claims l to 4, which has the capacity, in a test on plasma substantially free from platelets stimulated by human thromboplastin or by contact, to inhibit by 25 % the 20 generation of thrombin at concentrations less than 300 ng/ml.
6. A method of preparing a mixture according to any one of claims 1 to 5, which comprises fractionating a heparin or a depolymerised heparin by gel filtration. 25
7. The method according to claim 6, which comprises, successively, the following stages (i) dissolving the starting heparin or depolymerised heparin in the eluant, (ii) passing the solution thus obtained through ; i > o ■ >\ . . 17Aug:;93"; 24 2 43 1 - 18 - at least one column containing the solid support for the gel filtration, equilibrated beforehand with the same eluant, and (iii) recovering the fractions of the desired molecular weight. 5
8. The method according to claim 7,. wherein a--— depolymerised heparin is used.
9. The method according to claim 8, wherein a heparin depolymerised by the action of a base on a. heparin ester is used.
10 10. The method according to claim 7, wherein the eluant consists of a phosphate buffer, or of a solution of NaCl04 or NHaN03.
11. Method according to claim 10 wherein the phosphate buffer is a sodium phosphate or potassium 15 ^phosphate buffer.
12. Method according to claim 7, wherein, during stage (ii) the support is divided among several columns arranged in series.
13. Method according to claim 12, wherein the 20 number of columns used is less than 20.
14. Method according to claim 7, wherein stage (ii) is carried out using, successively, at least 2 types of supports having different separation characteristics.
15. Method according to any one of claims 7, 12, 25 13 and 14, wherein the support used is a polyacrylamide- agarose type gel.
16. Method according to claim 15, wherein the starting heparin (g)/gel volume (1) ratio is less than 2. // < Q \ r -9 MAR 1994 7 f; $ / '•J> fl i 0 - 19 -
17. Method according to claim 16 wherein the said ratio is between 0.5 and 1.5.
18. Method of preparing a mixture of oligosaccharides having a polydispersity of about 1 and a predetermined 5 molecular weight which comprises fractionating. Heparin ' ~— or depolymerised heparin by gel filtration on a solid support divided among several columns arranged in series.
19. Method of preparing a mixture of sulphated oligosaccharides as claimed in claim 1 substantially as 10 described in any one of Examples 1 to 3.
20. A mixture of sulphated oligosaccharides as claimed in claim 1 when prepared by a method as claimed in any one of claims 6 to 19.
21. A pharmaceutical composition having as 15 active ingredient a mixture of oligosaccharides according to any one of claims 1 to 5 and 20.
22. Use of a mixture of oligosaccharides according to any one of claims 1 to 5 and 20, in the prevention of thromboses in surgical material separated 20 from the human body. By the authorised agents A J PARK & SON "■ * r A c iv'
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
FR919104991A FR2675806B1 (en) | 1991-04-23 | 1991-04-23 | SULPHATE POLYSACCHARIDES, METHOD OF PREPARATION, PHARMACEUTICAL COMPOSITION AND USE. |
Publications (1)
Publication Number | Publication Date |
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NZ242431A true NZ242431A (en) | 1994-04-27 |
Family
ID=9412159
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
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NZ242431A NZ242431A (en) | 1991-04-23 | 1992-04-22 | Mixture of sulphated oligosaccharides derived from depolymerised heparin, and pharmaceutical compositions |
Country Status (10)
Country | Link |
---|---|
EP (2) | EP0581846A1 (en) |
JP (1) | JPH06506968A (en) |
AU (1) | AU1748592A (en) |
CA (1) | CA2108363A1 (en) |
FR (1) | FR2675806B1 (en) |
IE (1) | IE921287A1 (en) |
MX (1) | MX9201846A (en) |
NZ (1) | NZ242431A (en) |
WO (1) | WO1992018544A1 (en) |
ZA (1) | ZA922874B (en) |
Families Citing this family (8)
Publication number | Priority date | Publication date | Assignee | Title |
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FR2687158B1 (en) * | 1992-02-07 | 1995-06-30 | Rhone Poulenc Rorer Sa | SULPHATE POLYSACCHARIDES, METHOD OF PREPARATION, PHARMACEUTICAL COMPOSITION AND USE. |
US5763427A (en) * | 1995-03-31 | 1998-06-09 | Hamilton Civic Hospitals Research Development Inc. | Compositions and methods for inhibiting thrombogenesis |
US5744457A (en) * | 1995-03-31 | 1998-04-28 | Hamilton Civic Hospitals Research Development Inc. | Compositions and methods for inhibiting thrombogenesis |
US6001820A (en) * | 1995-03-31 | 1999-12-14 | Hamilton Civic Hospitals Research Development Inc. | Compositions and methods for inhibiting thrombogenesis |
US5767269A (en) * | 1996-10-01 | 1998-06-16 | Hamilton Civic Hospitals Research Development Inc. | Processes for the preparation of low-affinity, low molecular weight heparins useful as antithrombotics |
AU7753898A (en) * | 1997-06-06 | 1998-12-21 | Hamilton Civic Hospitals Research Development, Inc. | Modified low molecular weight heparin that inhibits clot associated coagulation factors |
WO1999010746A2 (en) * | 1997-08-26 | 1999-03-04 | The University Of North Carolina At Chapel Hill | Method of monitoring blood low molecular weight heparin and heparin |
AU2011303081B2 (en) * | 2010-09-14 | 2015-08-13 | Fuso Pharmaceutical Industries, Ltd. | High purity heparin and production method therefor |
Family Cites Families (4)
Publication number | Priority date | Publication date | Assignee | Title |
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FR2548672A1 (en) * | 1983-07-04 | 1985-01-11 | Pharmuka Lab | SULPHATE OLIGOSACCHARIDES AND THEIR USE AS MEDICAMENTS |
DE3608685A1 (en) * | 1986-03-15 | 1987-09-17 | Sandoz Ag | Stable low molecular weight heparin |
DK196886D0 (en) * | 1986-04-30 | 1986-04-30 | Novo Industri As | PREPARATION OF POLYSACCHARIDES |
EP0337327A1 (en) * | 1988-04-09 | 1989-10-18 | Bioiberica, S.A. | Process for the preparation of new oligosaccharide fractions by controlled chemical depolimerization of heparin |
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1991
- 1991-04-23 FR FR919104991A patent/FR2675806B1/en not_active Expired - Fee Related
-
1992
- 1992-04-21 WO PCT/FR1992/000352 patent/WO1992018544A1/en not_active Application Discontinuation
- 1992-04-21 CA CA002108363A patent/CA2108363A1/en not_active Abandoned
- 1992-04-21 ZA ZA922874A patent/ZA922874B/en unknown
- 1992-04-21 EP EP92910133A patent/EP0581846A1/en not_active Ceased
- 1992-04-21 AU AU17485/92A patent/AU1748592A/en not_active Abandoned
- 1992-04-21 EP EP92401111A patent/EP0511075A1/en active Pending
- 1992-04-21 JP JP4509237A patent/JPH06506968A/en active Pending
- 1992-04-22 IE IE128792A patent/IE921287A1/en not_active Application Discontinuation
- 1992-04-22 MX MX9201846A patent/MX9201846A/en unknown
- 1992-04-22 NZ NZ242431A patent/NZ242431A/en unknown
Also Published As
Publication number | Publication date |
---|---|
JPH06506968A (en) | 1994-08-04 |
IE921287A1 (en) | 1992-11-04 |
WO1992018544A1 (en) | 1992-10-29 |
ZA922874B (en) | 1992-12-30 |
AU1748592A (en) | 1992-11-17 |
EP0511075A1 (en) | 1992-10-28 |
FR2675806B1 (en) | 1994-06-10 |
EP0581846A1 (en) | 1994-02-09 |
FR2675806A1 (en) | 1992-10-30 |
MX9201846A (en) | 1993-02-01 |
CA2108363A1 (en) | 1992-10-24 |
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