Microcapsulation by interchange of multiple emulsions

SOOIETE CORTIAL, SA

PROCESS FOR PREPARATION OF NEW 3 ', 4' DERIVATIVES,

5,7-TETRAHYDROXIFLAVONE AND PHARMACEUTICAL COMPOSITIONS CONTAINING THEM wherein R 4 is H or OH and is alkoxy ^çi^çl^{oalco: x}i ^ 5 ™

-Cg, methanesulfonyloxy or paratoluenesulfonyloxy carac. characterized by a) when methanesulfonyloxy or to toluenesulfonyloxy, blocking free phenolic groups in an analog of (I) wherein O-glycosyl replaces Rg is released and the esterifying group or b) when Rg is Cg alkoxy -C 6 or C 1 -C 6 cycloalkoxy, RgCH 3 CN is reacted with 1,3-di- or 1,3,5-tri-thiiroxybenzylenium, followed by reaction with 3,4-dibenzyloxybenzoic acid anhydride and then debenzylation.

Process for the preparation of compositions and medicaments containing these compounds.

These compounds are useful, for example, in treating or preventing eye and nerve complications of diabetes.

The present invention relates to novel differently substituted hydroxy flavones in oxygen at position 3, their method of preparation and their therapeutic application.

The novel products forming the object of the present invention have the general formula

<img file="PT78337B_D0001.tif" />

<img file="PT78337B_D0002.tif" />

OH ·

ΟΙΙ (I) wherein R4H or OH and R₂= lower alkoxy of C2 to C6, cycloalkoxy of 05 to C8, methanesulfonyloxy or paratoluenesul '

phonyloxy under the condition that R3 is different from OH if R3₂= lower alkoxy from 02 to 04.

Products such that R1 = OH are derivatives of quercetin and products such that R1 = OH are derivatives of fisetin.

Some methyl ethers, and in particular 3-O-methyl, of flavools are already known. A number are natural derivatives. Societe Biosedra had registered on May 21, 1970, under No. 70184-58 the pentabenzylquercetin used in therapy in the classic flavonoid indications (inhibition of hypermeability and reduction of capillary fragility).

Societe Hoffman-la-Roche filed 9,1980 according to the English priority of 10.4.1979 a European patent No. 19081, which discloses the activity 3-alkoxy-5,7-dihydroxy flavones viruses, and in particular the preducts corresponding to general formula I, where R₁= 0H and R₂= lower alkoxy of C1 to C4 and preferably equal to methoxy or ethoxy. These products, although included in the general formula as set out in the European patent, are not, however,

-4·

<img file="PT78337B_D0003.tif" />

<img file="PT78337B_D0004.tif" />

described in the examples illustrating the invention.

It has now been found that the flavones of formula I, wherein Rj = H or OH and R₂= lower alkoxy. from 02 to 06, cycloalkoxy from 05 to 08, methanesulfonyloxy or paratoluenesulfonyloxy under the proviso that R 2 is different from OH if R 2₂= lower alkoxy from 02 to 04, showing inhibitory properties of aldose reductase, allowing its use in the prevention of ocular and nerve complications, in particular diabetes. These new compounds are equally useful as bipoglycemic and hypolipidemic.

The products which constitute the object of the present invention are prepared as follows:

- Products of formula I such as R₂= methanesulphonyloxy or paratoluenesulfonyloxy are prepared from the flavone glycosides such that H or OH and R₂= 0-glycosyl. Glycoside-free phenolic clusters are blocked, for example as benzyl esters; aglycone is obtained by the action of concentrated hydrochloric acid. The 3-OH grouping of flavorol is then esterified by mesityl chloride or paratoluenesulfonic acid chloride. Benzoate clusters are eliminated with a soda treatment.

Products of formula I such as Rj = H or OH and

R₂= lower alkoxy from 02 to 06 or cycloalkoxy from 05 to 08, provided that Rj is different from OH if R₂= lower alkoxy I from 02 to 04 are prepared by the action of formula &lt;

<img file="PT78337B_D0005.tif" />

no^J; L: '. Li.

ηιΚΤΙΙΙψί '·'. 'y_{í (}i'm ¹ ' \ ·

R₂CH₂CN and

<img file="PT78337B_D0006.tif" />

in the presence of hydrochloric acid, leading to

<img file="PT78337B_D0007.tif" />

This derivative is reacted with 3,4-dibenzyloxy-benzoic acid anhydride to yield a product of formula

<img file="PT78337B_D0008.tif" />

This ester is transformed into a product of formula I by hydrogenolysis in the presence, for example of palladium / BaSO4.

The present invention will be illustrated by the following examples:

EXAMPLE 1

Synthesis of 5-cyclohexyloxy-5,7,5,4-tetrahydroxy flavone; product of formula I, wherein R = O, O, code name COR 1985.

Synthesis of cyclohexyloxyacetonitrile

In an x ​​G-rignard reactor

500 cm 2, 100 g of cyclohexanol, 40 g of trio x _z xane, 200 cm @ 3 of toluene. This mixture is brought to 0 ° C under stirring. Gaseous hydrochloric acid is bubbled for four hours. The water formed is decanted.

Chloromethoxycyclohexane is dried and then distilled off. PF = 75^fiC under 27 millibars. A mixture of 62 g of this derivative and 56 g of cupro cyanide is heated at reflux at 120 ° C for 4 hours. Cyclohexylacetonitrile is distilled off. MP 60-7020 under 0.014 millibars. Yield 75%.

Synthesis of 2-cyclohexylglycoloyl floroglucinol

Dissolve in 500 cm @ 3 of anhydrous ether, 20 g of cyclohexyloxyacetonitrile and 18 g of anhydrous glucinol. Gaseous hydrochloric acid is bubbled for 4 hours at 0 ° C and stirred overnight at 0 ° C. It filters out. Ketimine hydrochloride is straight to 250 cm² of boiling ethanol. The mixture is heated at reflux for three hours. The reaction is followed by thin layer chromatography. Once cooled, the 2-cyclohexylglycolyl floroglucinol is quenched as an oil, then crystallized. It is recrystallized from an alcohol / water mixture, taken up in chloroform, then precipitated in petroleum ether. Yield 60%.

<img file="PT78337B_D0009.tif" />

Synthesis of 3,4-dibenzyloxy benzoic acid anhydride

To a 2 liter Grignard reactor 138 g of 3,4-dihydroxybenzaldehyde, 138 g of potassium carbonate and one liter of dimethylformam are introduced. (DMF). The mixture is refluxed under stirring. 253 g of chloromethylbenzene are added dropwise.

The mixture is stirred and refluxed for 6 hours. Once cooled, the precipitated salt is filtered off and washed with a little DMF. The reaction medium is poured in small quantities into 15 liters of ice water while shaking well. 3,4-Dibenzyloxy-benzaldehyde precipitates as a paste, then crystallizes. The solid is filtered then taken up in boiling methanol. After hot filtration, the methanolic solution is cooled under stirring and the precipitate formed is filtered off, washed with ice cold methanol, ethyl ether, then dried.

Yield: 77%.

In a 4 liter flask, the mixture consisting of 1.5 liters of water, 700 cm² of pyridine and 245 g of 3,4-dibenzyloxy-benzaldehyde is heated to r. flow. Little by little, 162 g of potassium permanganate are added. The mixture is heated at reflux for one hour, then filtered, hot and washed with one liter of boiling water. The first filtrate is concentrated to remove pyridine. After acidification by E ^ SO ^. In 1/2 of the filtrate set, the precipitate is filtered, then washed with water and dried.

Yield 85%.

The mixture consisting of one liter of benzene, 130 g of 3,4-dibenzyloxy benzoic acid and 45 cm @ 3 of thionyl chloride is heated at reflux for two hours and stirred. After evaporation is taken up in ether, the acid chloride is filtered off and washed with cold ether. Yield 90%.

In a flask fitted with one of these singings, the mixture consisting of 1,5 l of anhydrous ether,

290 in pyridine 100.2 g of 3,4-dibenzyloxy-benzoic acid and 105.7 g of 3,4-dihenzyloxy-benzoic acid chloride. The mixture is stirred for 24 hours at room temperature. The mixture is then poured into 8 liters of ice water. After 45 min of stirring, the precipitate is filtered off, washed with one liter of ice-cold 0.1 N hydrochloric acid and then with one liter of ice water. 0 solid is solubilized in CHCl3; decant, dry and evaporate and the anhydride is recrystallized from ethyl acetate.

Yield 88%.

Synthesis of 3 ', 4'-Dibenzyloxy-5,7-dihydroxy-3-cyclohexyloxy flavone

To a 2 liter flask 13.3 g of 2-cyclohexylglycolyl floroglucinol prepared according to 1b, 130 g of anhydride prepared according to 10, and 28 cm of triethylamine are introduced. The reaction mixture is dipped in a 170 ° C bath._O It is heated for 3 hours under stirring, then cooled to 100 ° C. After the addition of 1 liter of ethanol then 60 cm of 60% pots is heated for 45 min at reflux. After cooling to 30 ° C 1.5 liters of cold water are added. The precipitate formed is filtered off. The aqueous phase is extracted 3 times in ether. The solution is brought to pH 10 by a stream of COg θ extracted three times in ethyl acetate. The organic phase is dried and evaporated.

residue is taken up in ether.

Yield: 64%.

<img file="PT78337B_D0010.tif" />

-tetrahydroxy-flav

Summary of 5-οίο1ο-Ηβχί1οχί-5.7.5 ', 4'

<img file="PT78337B_D0011.tif" />

27 g of 3 ', 4'-dibenzyloxy-5 °, 7 ° -dihydroxy-3-cyclohexyloxy flavone, 500 cm @ 2 acetone 500 cm @ -1 methanol 8 are added to a two liter flask. palladium spatulas over 5% BaSO 4 · The mixture is stirred under a hydrogen atmosphere for 3 hours. The reaction This is followed by thin layer chromatography. After filtration over millipores the solution is evaporated._OYield: 98%.

NMR Spectrum: DMS0D6: 0.8-2.3 ppm, 10 protons, complex mass, CH₂4.0-4.6 ppm, 1 proton, massive complex, HCO-; 6.2 ppm, 1 proton doublet, H-6; 6.4 ppm, 1 doublet proton, H-8; 6.9 ppm, 1 proton, doublet, H-5 '&gt; 7.5-7.8 ppm, 2 protons, massive complex, H-2' + H-6 '; 9,9 ppm, 3 protons ”, dome 0H-7 + 0H-3 '+ 0H-4', exchangeable with D₂0;

12.8 ppm, 1 singlet proton, D exchangeable OH-5₂0 j

PP = 260-270 ° C (Kofler bank). {

Elemental microanalysis: Calculated C 65.62%; H5.24%; 0 1

29,13%.

Found: C 65.63%; H 5.5%;

29,40%.

i

Prepare the same way 0

3-ethoxy-7,5 ', 4'-trihydroxy flavone, product of formula I wherein R 2 = H and R₂= OC ^; code number COR 1987.

NMR spectrum in DMSOD6: 1.3 ppm, 3 triplet protons CH2; 4.1 ppm, 2 quadruple protons, CH₂, 6.7-8.1 ppm, 6 protons, massive complex aromatic protons; 9.9 ppm, 3 protons, broad peak, OH, exchangeable with D₂0.

PP = 25420 (Mettler apparatus).

<img file="PT78337B_D0012.tif" />

EXAMPLE 2

<img file="PT78337B_D0013.tif" />

<img file="PT78337B_D0014.tif" />

Synthesis of 3-methanesulfonyloxy-3 ', 4', 5,7-tetrahydroxy flavone; product of formula I wherein R1 = OH, 11 = CH3 -SO4 -0; code name COR 1988.

Synthesis of 3 '«4¹5,7-tetrabenzoyloxy flavone

800 g of potassium carbonate are dissolved in 7 liters of water. 100 is added; g of rutin under agitation. When the mixture becomes clear, 400 cm @ 2 of benzoyl chloride is added twice. The mixture is kept under strong stirring for two hours. The solid is filtered off and washed to neutral. It is vacuum dried over Ρ £ θ5 »ⁱⁿ It is then suspended in 3 to 4 liters of ethyl ether; The mixture is stirred vigorously, the solid is filtered off and this preparation is repeated a second time. 205 g of crude rutin tetrabenzoate thus obtained. These are dissolved in 1.3 liters of boiling ethanol. 400 cm @ 3 of concentrated hydrochloric acid are added at one time. The mixture is stirred while refluxing with ethanol for one hour. The alcohol phase is separated from the solid. than precipitated as a viscous mass. 0 The solid is ground in a mortar and washed with water if necessary. ! It is then dissolved in chloroform. After washing in | water, drying and evaporation, the residue is taken up to 700 cm² of acetone. The mixture is refluxed under stirring and filtered hot. The solid is washed with some acetone.

Yield: 28.2% (relative to rutin).

Synthesis of 3-methanesulfonyloxy-3 ', 4', 5,7-tetrahydroxy flavone g of either ketone tetrabenzoate are dissolved in 180 cm @ 2 of pyridine. Add 10 cm swimwear⁵ of mesityl chloride. The mixture is kept under stirring for one hour at room temperature.

A further 10 cm @ 3 amount of mesityl chloride is added again and stirring is continued at room temperature for three hours. The mixture is poured into a cold slurry under stirring. After filtration, the solid is washed with water and dissolved in chloroform. The formic chlorine phase is dried and evaporated. 600 cm @ 2 of acetone is again added to the evaporation residue. The mixture is refluxed under stirring, then filtered hot. Yield: 54.3% · g of 3-methanesulfonyloxy flavone tetrabenzoate are dissolved in a mixture of

1.5 liters of acetone and boiling methanol under argon. To the reaction mixture is added 2 g of soda ash in methanol. After the thin layer chromatography has verified that the reaction is complete, the mixture is cooled and the pH is brought to about 9 with the aid of a stream of CO 2.₂. The solvents are evaporated. Ethyl acetate is added to the residue. The mixture is stirred, then the solvent is decanted.

residue is solubilized in water. The solution is neutralized using C0₂ and the precipitate is filtered off. It is dried under vacuum over P20.

Yield 38%.

NMR spectrum in DMSOD6: 3.5 ppm, 3 protons, singlet CH 3; 6.3 ppm, 1 proton, doublet, H-6; 6.5 ppm, 1 proton 1 doublet, H-8; 7.0 ppm, 1 proton, 1 doublet, H-5; 7.3-7.7 ppm, 2 protons, massive H-2 '+ H-6 complex; 10.0 ppm, proton dome 0H-7 + 0H-3 '+ 0H-4' exchangers with D₂0; 12.1 ppm, 1 proton, OH-5 dome, exchangeable with D₂0.

MP = 233^yesC (Mettler apparatus).

It is also prepared to

3-paratoluenes sulfonyloxy-5,7,3 ', 4'-tetrahydroxy flavone

<img file="PT78337B_D0015.tif" />

product of formula I, wherein R 1 = OH and R₂= paratoluenesulphonyloxy; code name COR 1990.

The pharmacological properties of the products which are the object of the present invention | are set forth below.

Interest of the products of the present invention in the treatment and prophylaxis of diabetes complications has been highlighted in vitro by; determination of aldose reductase inhibitory activity and in vivo in the model of zotocin strep-induced neuropathy.

<img file="PT78337B_D0016.tif" />

Inhibition of aldose reductase in vitro

i

The enzyme is extracted from the oxen lens by the method

I is described by Hayman S. et Kinoshita JH (J.Biol.Chem. J

1965, 240,2,877) · Percent inhibition of the enzyme's ability to reduce glyceraldehyde to glycerol under the effect of the test product is determined by spectrophotometric dosing of the amount of NALPH reacting according to the method described by Hayman et Kinoshita . Present | below in each parenthesis after each product is the log value 1/0150 where IC 50 represents the concentration expressed in mole / 1, leading to 50% inhibition of enzymatic activity: COR 1983 (5,8), COR 1987 (6.3),

COR 1988 (6.8), COR 1990 (5.5) · These values ​​are to be compared with the corresponding value for keratin (5.0).

Streptozotocin Neuropathy Lots of 10 rats weighing about 200 g are treated from J-2 to J + 3 by a single portion of the test product. OJ the animals receive an injection. 100 mg / kg streptozotocin dissolved in a citrate buffer. Blood glucose is determined 24 hours and 3 days after the injection of the diabetic agent. Rats are sacrificed at 3 days after streptozine kin injection and sciatic nerves are raised early to determine sorbitol and inositol rates. Tested under these conditions COR 1988 leads to a decrease in sorbitol rate from 0.819 + 0.215 mg / kg in control animals to 0.608 + 0.049 mg / kg in animals treated with 50 mg / kg and 0.482 + 0.091 mg / kg. kg animal levels treated with 100 mg / kg. Similarly, the rate of inositol increased from 2,174 +0.481 mg / kg in control animals to 1.593 + .109 mg / kg in animals treated with 25 mg / kg to 1.430 +, 0.104 mg / kg in animals treated with 50 mg. / kg and to 1.402 +, 0.130 mg / kg in animals treated with 100 mg / kg. Sorbitol and insitol rates after treatment are compared with control rates by the Mann and Whitney U test (Schwartz D. Méthodes Statistiques à l'usag | and des medecins et biologists, Ed. Plammarion Paris 1963).

The results obtained are significant with pL 1%. The blood glucose of the animators. munha is 4.72 +, 0.60 g / 1 at J3. It goes from 3.93 +0.82 g / 1 in animals treated with 50 mg / kg COR 1988 and to 3.61 + 0.75 g / 1 in animals treated with 100 mg / kg. These blood glucose drops are significant.

The toxicity of the products of the present invention is determined in mice.

COR 1983 administered orally in solution in water in the presence of 6% gum does not determine any mortality at the dose of 1 g / kg. Adminis, administered intraperitoneally in solution at 5% Tween does not cause any mortality up to 100 mg / kg and leads to 10% mortality at the dose of 200 mg / kg.

COR 1988 orally administered in solution in water in the presence of a / o gum causes no mortality up to a dose of 3 g / kg.

The products which are the subject of the present invention are useful as hypolipidemics. Thus, COR 1983 administered twice a dose 400 mg / kg orally, 20 hours apart, to hypercholesterolemic mice leads to 24% colonic decrease, terolemia.

Given their properties together with low toxicity, the products of the present invention are useful in human and veterinary therapy, for example, in the treatment and prevention of ocular and nerve complications of diabetes or in the treatment of diabetes. diabetes and hyperlipidemias. The products which are the object of the present invention may be used alone or in combination with antidiabetics. In combination with appropriate vehicles and excipients, they will be administered orally in the form of dragees, tablets, syrups, drinkable rectal suppositories, parenterally in subcutaneous, intramuscular intravenous injections, topically. in the form of ointments or jellies. They may also enter ophthalmic compositions in the form of eye drops or ointments. Doses administered will vary according to the indication and the individual from 5 to 500 mg / day in 2 to 6 oral doses, from 5 to 500 mg / day in one or two rectal doses of 0.5 to 50 mg by parenteral injection.