Sulfated Diosmin Derivative Sulfatierte Diosminderivate Derives De Diosmin Sulfurizes

Europaisches Patentamt (19) European Patent Office Office europeenpeen des brevets EP 0 558 435 B1

(12) EUROPEAN PATENT SPECIFICATION

(45) Date of publication and mention (51) intci.6: C07H 17/07, A61K 31/70 of the grant of the patent: 01.09.1999 Bulletin 1999/35

(21) Application number: 93500023.2

(22) Date of filing: 24.02.1993

(54) Sulfated diosmin derivative Sulfatierte Diosminderivate Derives de diosmin sulfurizes

(84) Designated Contracting States: (72) Inventors: AT BE CH DE DK FR GB GR IT LI LU NL PT SE • Orjales-Venero, Aurelio Neguri (Vizcaya) (ES) (30) Priority: 25.02.1992 ES 9200414 • Mosquera-Pestana, Ramon Las Arenal (Vizcaya) (ES) (43) Date of publication of application: 01.09.1993 Bulletin 1993/35 (74) Representative: Blumbach, Kramer & Partner GbR (73) Proprietor: FABRICA ESPANOLA DE Radeckestrasse 43 PRODUCTOS 81245 Miinchen (DE) QUIMICOS Y FARMACEUTICOS, S.A. (FAES) E-48940 Leioa-Lamiaco (Vizcaya) (ES) (56) References cited: EP-A- 0 302 155 EP-A- 0 505 937 WO-A-82/00586

DO lO CO ^- 00 LO LO Note: Within nine months from the publication of the mention of the grant of the European patent, any person may give notice the Patent Office of the Notice of shall be filed in o to European opposition to European patent granted. opposition a written reasoned statement. It shall not be deemed to have been filed until the opposition fee has been paid. (Art. a. 99(1) European Patent Convention). LU

Printed by Jouve, 75001 PARIS (FR) EP 0 558 435 B1

Description

[0001] The present invention relates to the diosmin heptakis (hydrogensulfate) aluminium complex (formula I), useful in the treatment of ulcerous processes, and to a process for its preparation 5

OR OR

Formula I, R= S03[AI2(OH)5]

Formula II, R= S03Na 25 [0002] An earlier patent by the same authors (Spanish patent no. 8702270) corresponding to the document EP-A- 0 302 1 55 describes the preparation of a related substance, the diosmin octakis (hydrogensulfate) aluminium complex, by reacting diosmin with an excess of a sulfating agent and treating the obtained product with aluminium hydroxychloride . 30 [0003] The present invention concerns a diosmin-heptakis(hydrogensulfate) sodium or diosmin-heptakis(hydrogensulfate) aluminum complex of the general formulae I and II

Formula I: R = S03[AI2(OH)5]

Formula II: R = S03Na.

[0004] The present invention also relates to said compounds for use as a cytoprotective agent or for use in the treatment of gastric diseases and ulcerous diseases, preferably of duodenal ulcer. 55 [0005] The invention also concerns pharmaceutical compositions comprising an effective amount of at least one of the above compounds of the general formulae (I) and (II), optionally together with appropriate excipients. [0006] The invention also concerns a process for the preparation of the above compounds of the above formulae (I) and (II), said process comprising the steps of

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reacting diosmin with a sulfur trioxide complex with picolines or alkyl-substituted picolines in a molar ratio of diosmin : sulfur trioxide of about 1 : 7 in said picoline or alkyl-substituted picoline as the solvent;

neutralizing the resulting solution with an aqueous alkaline sodium ion-containing solution;

precipitating, from the aqueous phase of the resulting two phase mixture, the sodium salt of the formula II;

Formula II: R=S03Na

and optionally

transforming said precipitate of the formula II into the aluminum complex of the formula I.

Formula I: R = S03[AI2(OH)5]

[0007] This invention relates to a new sulfated diosmin derivative obtained by a controlled reaction in anhydrous pyridine or alkylpyridine of one mole of diosmin with seven moles of a sulfating agent and isolated as the sodium salt (formula II) and as the aluminium complex (formula I). The molar ratio of the sulfating agent to diosmin, is critical to obtain the diosmin heptakis (hydrogensulfate). The sulfating agent is selected from sulfur trioxide complexes with picolines and alkyl-substituted picolines, especially 2-picoline and 5-ethyl-2-picoline, which favour to control the process and afford exclusively the diosmin heptakis (hydogensulfate), which is isolated as the sodium salt from the reaction medium and transformed into diosmin heptakis (hydrogensulfate) aluminium complex. [0008] Both the sodium salt and the aluminium complex of the diosmin heptakis (hydrogensulfate) are of particular interest because of their pharmacological activities, especially the aluminium complex which has proved to be very effective as antiulcerous agent. The diosmin heptakis (hydrogensulfate) aluminium complex is a yellow solid, insoluble in water and organic solvents. Heated up to 350°C it does not melt nor undergoes any changes. The diosmin heptakis (hydrogensulfate) sodium salt is a white solid which melts at 143-6°C. The hydroxyl group which remains [0009] unsulfated is that of the 5 position (formulas I and II), as confirmed by proton nuclear magnetic resonance EP 0 558 435 B1

(NMR) determinations. Hydroxyls at 5 position in flavonoids are the least reactive in the molecule and the study pub- lished in Z. Naturforsch., 43c, 625-630 (1988), which refers to the preparation of sulfated derivatives of a group of flavonoids (kaempferol, quercetin, tamarixetin, rhamnetin, eupalitin, eupatolitin and veronicafolin), none of which has the said hydroxyl sulfated, is a good reference. However, under appropriated reaction conditions, persulfation is 5 achieved, as referred in the Spanish patent 8702270 for diosmin octakis (hydrogensulfate) and in international patent WO 82/00586 for rutin deca (hydrogensulfate) sodium salt. In any case, if one of the hydroxyls remains unsubstituted it is always that of the 5 position, as related also in the international patent WO 82/00586, which describes how, with appropriated reaction conditions, rutin nona (hydrogensulfate) sodium salt with the free hydroxyl at 5 position is obtained. A free hydroxyl at 5 position in a flavone is easily detected by means of NMR, because the corresponding 10 proton appears at a very low field position, approximately at 12-13 ppm (K. R. Markham and T. J. Mabry, "The Flavo- noids", J. Harborne, T. Marby and H. Marby Eds., page 69, Chapman and Hall Ltd., London (1975)). In the diosmin heptakis (hydrogensulfate) sodium salt NMR spectrum the 5 position hydroxyl appears at 13.1 ppm. [0010] The following examples of the preparation of the sodium salt and the aluminium complex of diosmin heptakis (hydrogensulfate) illustrate the invention but are not exhaustive. 15 EXAMPLE 1

Diosmin heptakis (hydrogensulfate) sodium salt

20 [0011] 60 ml of 5-ethyl-2-picoline are introduced into a reaction flask provided with a condenser and a pressure compensated separatory funnel. A nitrogen stream is continously flushed through the system, while is externally ice- water cooled. Under good stirring 4.58 ml of chlorosufonic acid are added dropwise. Then 6 g of anhidrous diosmin are introduced in the flask and the mixture heated during 3 hours at 60°C. The solution is then neutralized with aqueous sodium hydroxyde solution to a pH of 7.0-8.0 and the two layers obtained are separated by using a separating funnel. 25 The aqueous layer is washed with ethyl acetate and again separated, being the diosmin heptakis (hydrogensulfate) sodium salt precipitated by adding slowly and under stirring 20 ml of ethanol. The resulting solid is filtered and washed several times with ethanol and dried. It could be purified by dissolution in water and precipitation by adding ethanol. 7.1 g of the product are obtained, m. p. 143-6°C.

Elemental analysis for C28H25Na7036S7: C Na S Calculated (%) 25.43 12.17 16.93 Found (%) 25.02 12.56 16.68 1H NMR (DMSO-d6): 8 13.1 (1H,s,5).

EXAMPLE 2

[0012] To a diluted aqueous solution of 13.4 g of diosmin heptakis (hydrogensulfate) sodium salt 95 ml of a 15% aluminium hydroxychloride aqueous solution are added. The solid which forms is filtered and washed several times with water, then suspended in water and pH adjusted between 5.5 and 6.5 with diluted aqueous sodium hydroxyde solution. The solid is filtered, washed with water and dried at room temperature under vacuum, yielding 1 6.8 g of the diosmin heptakis (hydrogensulfate) aluminium complex as a yellow solid. M. p.: It does not melt at 350°C.

Elemental analysis for C28H60AI14O71S7: C Al S Calculated (%) 15.75 17.70 10.49 Found 16.14 17.40 10.82 50 (%)

[0013] The diosmin heptakis (hydrogensulfate) aluminium complex has shwon to be pharmacologically active in experimental gastric and duodenal ulcer models, induced by different mechanisms (acid gastric hypersecretion, non steroidal antiinflammatories, cysteamine, stress and cold, etc.). It also shows an excellent citoprotector activity against gastric lesions induced by absolute ethanol, with a mechanism of action partially related to an increase in the synthesis and/or intraluminal release of protective prostaglandins. [0014] The results of two pharmacological studies carried out with diosmin heptakis (hydrogensulfate) aluminium complex are described for illustrative purpouses:

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To study its protective action against indomethacin-induced gastric lesions in the rat the method of Brodie and col. (Toxicol. Appl. Pharmacol., V7, 615-624 (1970)) was used. Male Wistar rats weighing 220 g were used and gastric lesions were induced by oral administration of indomethacin (40 mg/kg) suspended in water. A control group received only water. Treated animals received the diosmin heptakis (hydrogensulfate) aluminium complex 30 minutes 5 before inducing the lesions. In table I the obtained results are shown. Code F-316 corresponds to the diosmin heptakis (hydrogensulfate) aluminium complex.

Table I.- F-316 action on indomethacin-induced gastric lesions, administered 30 minutes before the ulcerogenic agent. 10 1 1 1 PRODUCT DOSE mg/kg LESION INDEX (mm) Mean + S.D. INHIBITION PERCENTAGE VEHICLE - 0.00 VEHICLE 15 + 18.571 ±1.98 INDOMETHACIN 40 F-316 100 + 10.568 ±3.37 -43.09* INDOMETHACIN 40 on F-316 200 + 7.965 ±2,26 -57.11** INDOMETHACIN 40 F-316 400 2S + 6.536 ± 2.75 - 64.85** INDOMETHACIN 40 * p < 0.05 ** p < 0.01 according to Student's t test. 30 Diosmin heptakis (hydrogensulfate) aluminium complex action on oesophagic lesions caused by simultaneous ligation of the pylorus and the glandular-aglandular transition area of the stomach of the rat has been studied, according to the procedure of Nakamura and col. (Japan J. Pharmacol., 32, 445-456 (1 982)). It has been verified that the diosmin heptakis (hydrogensulfate) aluminium complex significatively protects the oesophagic mucose 35 against gastric juice induced lesions. In the same way, the disomin heptakis (hydrogensulfate) sodium salt also attains similar protection values. In table II are shown the obtained results. Code F-316 corresponds to diosmin heptakis (hydrogensulfate) aluminium complex and F-706 to diosmin heptakis (hydrogensulfate) sodium salt.

Table II.- Effect of compounds F-31 6 and F-706 on acute oesophagic lesions induced by simultaneous ligation of the pylorus and the glandular-aglandular transition area in the stomach of the rat. Reference product: Aluminium hydroxide. Treatment Dose Lesion grade Lesion Protection (mg/kg) Index (LI) % 45 0 12 3 4 Vehicle - 1 1 1 1 16 87.5 25 9 0 0 0 1 10 88.51 50 F-316 50 10 0 0 0 1 0 100 100 10 0 0 0 0 0 100 Aluminium 200 2 1 3 3 1 50 42.86 Hydroxide 25 0 82.8 55 7 1 1 1 15 F-706 50 10 0 0 0 0 0 100 100 10 0 0 0 0 0 100

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[0015] Based on their pharmacological properties, the diosmin heptakis (hydrogensulfate) sodium salt and the diosmin heptakis (hydrogensulfate) aluminium complex both are useful in the therapy of ulcerous processes. They can be administered alone or formulated with appropriated excipients, using conventional pharmaceutical forms . Suitable pharmaceutical forms for oral administration include solid and liquid unit oral dosage forms such as tablets, capsules, suspensions, powders and the like, whereas for dermic application formulations as cream and ointment are preferred.

Claims

1. Diosmin-heptakis(hydrogensulfate) sodium or diosmin-heptakis(hydrogensulfate) aluminum complex of the general formulae I and II

Formula I: R = S03[AI2(OH)5]

Formula II: R = SOgNa.

2. The compounds of the general formulae I or II for use as a cytoprotective agent.

3. A pharmaceutical composition comprising an effective amount of at least one of the compounds of the general formulae I or II, optionally together with appropriate excipients.

4. The parmaceutical compositions of claim 3 for use in the treatment of gastric diseases and ulcerous diseases, preferably of duodenal ulcer.

5. A process for the preparation of any one of the compounds of the formulae I or II, said process comprising the steps of

neutralizing the resulting solution with an aqueous alkaline sodium ion-containing solution;

precipitating, from the aqueous phase of the resulting two phase mixture, the sodium salt of the formula II; EP 0 558 435 B1

OCH. r— o — cel

OR • OR

Formula II: R = S03Na

and optionally

transforming said precipitate of the formula II into the aluminum complex of the formula I.

OCH. I — 0 —CEL

OR • OR OR

Formula I: R = S03[AI2(OH)5]

6. The process according to claim 5, wherein the picolines are selected from 2-picoline and 5-ethyl-2-picoline.

7. The process according to claim 5 or claim 6, wherein the sulfating reaction is carried out at 60°C.

Patentanspriiche

1. Diosmin-heptakis(hydrogensulfat)natrium- oder Diosmin-heptakis(hydrogensulfat)aluminium-Komplex der meinen Formeln I und II EP 0 558 435 B1

Formell: R = S03[AI2(OH)5]

Formelll: R = S03Na.

Verbindungen der allgemeinen Formeln I und II zur Verwendung als zellschutzendes Mittel.

Pharmazeutische Zusammensetzung, die eine wirksame Menge wenigstens einer der Verbindungen der allgemeinen Formeln I oder II umfaBt, gegebenenfalls zusammen mit geeigneten Tragern.

Pharmazeutische Zusammensetzungen nach Anspruch 3 zur Verwendung bei der Behandlung von Magenerkran- kungen und Ulkuserkrankungen, vorzugsweise von Zwolffingerdarmgeschwur.

Verfahren zur Herstellung jeder der Verbindungen der Formeln I oder II, worin das Verfahren die Schritte umfaBt, dal3 man

Diosmin mit einem Schwefeltrioxid-Komplex mit Pikolinen oder alkylsubstituierten Pikolinen in einem Molver- haltnis Diosmin : Schwefeltrioxid von etwa 1 : 7 in dem Pikolin oder alkylsubstituierten Pikolin als Losungsmittel umsetzt; die resultierende Losung mit einer waBrigen alkalischen Natriumionen enthaltenden Losung neutralisiert; aus der waBrigen Phase der resultierenden Zweiphasen-Mischung das Natriumsalz der Formel II ausfallt;

Formelll: R=S03Na

und gegebenenfalls den Niederschlag der Formel II in den Aluminium-Komplex der Formel I umwandelt.

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0 — CH, OCH.

OR • OR OR OH o

Formel I: R = S03[AI2(OH)5]

6. Verfahren nach Anspruch 5, worin die Pikoline gewahlt sind aus 2-Pikolin und 5-Ethyl-2-pikolin.

7. Verfahren nach Anspruch 5 oder Anspruch 6, worin die Sulfatier-Reaktion bei 60 °C durchgefuhrt wird.

Revendications

1 . Complexe diosmine-heptakis (hydrogenosulfate) de sodium ou diosmine-heptakis (hydrogenosulfate) d'aluminium repondant auxformules generales I et II

oca.

lRO

OR - OR OR 0

Formule I : R = S03[AI2(OH)5]

Formule II : R= S03Na.

2. Composes repondant aux formules generales I et II destines a etre utilises en tant qu'agents cytoprotecteurs.

3. Composition pharmaceutique comprenant une quantite efficace d'au moins un des composes repondant aux formules generales I et II, facultativement melange avec des excipients appropries.

4. Compositions pharmaceutiques selon la revendication 3, destinees a etre utilisees dans le traitement des maladies gastriques et des maladies ulcereuses, de preference dans le traitement de I'ulcere duodenal.

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Procede de preparation de I'un quelconquedes composes repondant aux formules I et II, ledit procede comprenant les etapes de :

mise en reaction de diosmine avec un complexe de trioxyde de soufre et de picolines ou de picolines substi- tutes par un groupe alkyle dans un rapport stoechiometrique entre la diosmine et le trioxyde de soufre d'environ 1/7 dans ladite picoline ou picoline substitute par un groupe alkyle utilisee en tant que solvant ;

neutralisation de la solution resultante avec une solution aqueuse basique contenant des ions sodium ;

precipitation, a partir de la phase aqueuse du melange biphasique ainsi obtenu, du sel de sodium repondant a la formule II ;

JLr ' OR OR ORw ' OK 0

Formule II: R = SOgNa

et facultativement,

transformation dudit precipite repondant a la formule II en le complexe d'aluminium repondant a la formule I.

Formule I : R = S03[AI2(OH)5]

Procede selon la revendication 5, dans lequel les picolines sont choisies parmi la 2-picoline et la 5-ethyl-2-picoline.

Procede selon la revendication 5 ou la revendication 6, dans lequel la reaction de sulfatation est conduite a 60°C.