Stable aqueous formulations of the compounds include cyclodextrins, connections enable-,or- cyclodextrin or its alkyl - or hydroxyalkylated and (6r)- or (6s)-5,10-methylenetetrahydrofolic acid or a salt thereof, a method of stabilizing aqueous solutions and the method of obtaining stable solutions

 

(57) Abstract:

Stable aqueous formulations of the compounds include cyclodextrins contain a) (6R)-, (6S)- or (6R,S)-5,10 - methylentetrahydrofolate acid or salt of 6R)-, (6S)- or (6R,S)-5,10 - methylenetetrahydrofolic acid, b) -, or-cyclodextrin or alkyl or hydroxyalkyl-derivatives-or-cyclodextrin or mixture, or cyclodextrin, or alkyl - or hydroxyalkylated -,- or-cyclodextrin, or a mixture,- or-cyclodextrin with alkyl - or hydroxyalprazolam -,- or-cyclodextrin. It turned out that these connections enable more stable than the 5,10-tetrahydrofolate acid. These compounds are stable in aqueous solution and suitable for receiving parenteral medicines. 5 C. and 8 C.p. f-crystals, 5 PL.

The invention relates to new compounds include, -, - or-cyclodextrin or its alkyl - or hydroxyalkylated and (6R)-, (6S) - or (6R,S)-5,10 - methylenetetrahydrofolic acid or its salts, stable solutions of compounds include cyclodextrin, a method of stabilizing aqueous solutions and the way to obtain stable solutions, which can be used in the pharmaceutical industry.

And arctonyx drugs.

Preferred hydroxypropylcellulose selected from the group: hydroxypropylcellulose (0,6), hydroxypropylcellulose (0,9), 3-hydroxypropylcellulose, 2,3-dihydroxyphenylalanine, especially 2-hydroxypropylcellulose. Salt of 5,10-methylenetetrahydrofolic acid, first of all, are salts of alkali and alkaline earth metals.

Tetrahydrofolate are biologically active forms of folic acid (Co-factors folic acid). As medicines are usually used 5-formyl-5,6,7,8 - tetrahydrofolic acid (leucovorin) in the form of calcium salts, for example, to enhance therapeutic effects of 5-fluorouracil or, for example, as Resco substances when the use of methotrexate in the treatment of cancer.

In the body (6S)-5-formyltetrahydrofolate acid into (6R)-5,10-methylentetrahydrofolate acid under the action of which as a Co-factor 5-fluorouracil (5-FU) is transformed into 5-ftordezoksianalogov (5-F-dUMP) and under the action of thymidylate synthetase (TS) cytostatic active in covalent complex: 5-F-dUMP(TS)-5,10-methylentetrahydrofolate acid. (See , W. A. Bleyer, Cancer March 15 Supplement 1989: S. 995 - 1007 compounds E. L. R. Stokstad, Folic Acid Metabolism in Health and Disease) directly Co-factor 5,10 - methylenetetrahydrofolic acid. (See, WO 91/17660, page 5, lines 24-35). To date these attempts have failed because of insufficient purity and stability 5,10 - methylenetetrahydrofolic acid and its salts. According to EP 0409125, in these compounds, the content of (6R,S)-5,10 - methylenetetrahydrofolic acid typically is in the range of 85-90%. Only from the salts of calcium and magnesium (6R, S)-5,10 - methylenetetrahydrofolic acid were obtained in connection with the content of 96.5 is 98.8%. However, the stability of solutions of salts is very critical. At pH 9 after 6 hours was observed only 85% of the initial number of 5,10-methylenetetrahydrofolic acid. (See EP 0409125, the table on page 14).

5,10-Methylentetrahydrofolate acid in solution is in equilibrium with formaldehyde and tetrahydrofolate acid. (See, L. J. Machlin, Handbook of Vitamins, 2nd Ed. (Marcel Decker Inc., N. Y./Basel), T. Brody S. 457; M. J. Osborn, et al, J. Am. Chem. Soc. 82, 4921 (1960), R. G. Kallen, et al., J. Biol. Chem. , 241, 5851 (1966), Moran et al., Proc. Natl. Acad. Sci. USA 76, 1456-1460 (1979). This equilibrium is contraindicated for parenteral use 5,10-methylenetetrahydrofolic acid. Even in WO 91/17660 that clearly shows the advantages of using 5,10-methylenetetrahydrofolic acid compared with the derivative tetrahydrofolate acid, does not refer to the possibility of stabilization of the CSOs application 5,10-methylenetetrahydrofolic acid. Known to date technology, which is used to stabilize 5-formyltetrahydrofolate acid adding ascorbic acid, parabens, mercaptopurin or trometamol, (see European patent EP 0416232), or, in the case of stabilization of the calcium salt of 5-formyltetrahydrofolate acid adding complexing agents for alkaline earth ion (see European patent EP 0401895) may not be applied to 5,10-methylenetetrahydrofolic acid or have only limited effect, for example, increase the time oxidative cleavage of tetrahydrofolate acid and the derivatives tetrahydrofolate acid. None of these methods, however, do not affect the equilibrium 5,10-methylenetetrahydrofolic acid and tetrahydrofolate acid and free formaldehyde.

It is known that the inhibition of oxidative decomposition sent using - cyclodextrin at stabilization digidrofolieva acid, which can be used as the substrate in the biochemical analysis of methotrexate (Japan patent 58-48933 Sho) and the use of cyclodextrin for stabilization of leucovorin (European patent 0427078).

Unexpectedly it was discovered that then it is carbonated the Les reactions in aqueous solution obtained stable compounds include 5,10-methylenetetrahydrofolic acid and the corresponding cyclodextrin. As a result of this first became possible therapeutic application of 5,10-methylenetetrahydrofolic acid in aqueous solutions.

The increase in stability is all the more unexpected that previously held the study of reactions include virtually showed no interaction tetrahydrofolate with cyclodextrin (see, D. W. Armstrong et al., Science, 232, 1132-5 (1986), especially pp. 134, column 2, line 2). By special measurements were able to establish that the cyclodextrin 5-formyltetrahydrofolate acid and 5-methyltetrahydrofolate acid not interacting at all or very poorly. Thus, it was impossible to expect the stabilization of 5,10-methylenetetrahydrofolic acid against chemical decomposition and, above all, in relation to shifting the equilibrium of free formaldehyde and tetrahydrofolate acid side 5,10-methylenetetrahydrofolic acid.

The present invention relates to the production and study of compounds include cyclodextrins with 5,10-methylenetetrahydrofolic acid. When it was unexpectedly found that these compounds enable more stable than the 5,10-tetrahydrofolate acid. The stability inherent in the compounds included in any form, that is by means of on -, - or - cyclodextrin or its alkyl - or hydroxyalkylated and (6R) or (6S)-5,10 - methylenetetrahydrofolic acid or its salts, when a molar ratio of from 1:1 to 1:2.

These compounds are stable in aqueous solution and suitable for receiving parenteral medicines, for example in the treatment of cancer.

Preferred compounds include, representing a compound include hydroxyalkyl-- or hydroxyalkyl -- cyclodextrin and (6R)- or (6S)-5,10 - methylenetetrahydrofolic acid or its salts, in particular representing connections include hydroxypropyl-- or hydroxypropyl--cyclodextrin and (6R)- or (6S)-5,10-methylenetetrahydrofolic acid or its salts or compounds include, representing a compound include alkyl-or alkyl-cyclodextrin and (6R)- or (6S)-5,10 - methylenetetrahydrofolic acid or its salts, in particular, representing a compound include dimethyl-- or dimethyl--cyclodextrin and (6R)- or (6S)-5,10 - methylenetetrahydrofolic acid or a salt thereof.

Due to the high solubility and good physiological compatibility for obtaining these compounds are especially suitable salts of alkaline or alkaline earth metallon 5,10-methylenetetrahydrofolic acid compared with the calcium salt with the same purity has approximately an order of magnitude greater solubility, and sodium and potassium salts more soluble. These substances are therefore particularly preferable for obtaining pharmaceutical products.

The preferred compounds are:

salts of alkali and alkaline earth metals (6R)- and (6S)-5,10-methylenetetrahydrofolic acid-cyclodextrin;

salts of alkali and alkaline earth metals (6R)- and (6S)-5,10 - methylenetetrahydrofolic acid with hydroxypropyl -- cyclodextrin;

salts of alkali and alkaline earth metals (6R)- and (6S)-5,10 - methylenetetrahydrofolic acid with hydroxypropyl -- cyclodextrin;

salts of alkali and alkaline earth metals (6R)- and (6S)-5,10-methylenetetrahydrofolic acid-cyclodextrin;

salts of alkali and alkaline earth metals (6R)- and (6S)-5,10 - methylenetetrahydrofolic acid with dimethyl-cyclodextrin.

Consequently natural (6R)-form 5,10 - methylenetetrahydrofolic acid is preferred for obtaining compounds of inclusion.

Another object of the present invention is a method of obtaining compounds include (6R)-, (6S)- or (6R,S)-5,10 - methylenetetrahydrofolic acid or their salts, which consists in the fact that you get a stable aqueous Rasta formaldehyde in the presence of -, - or - cyclodextrin or alkyl - or hydroxyalkylated -, - or - cyclodextrin, followed by separation of the compounds of cyclodextrin inclusion 5,10 - methylenetetrahydrofolic acid.

Connections include cyclodextrin is preferably produced from easily available (see EP 0495204 A) sulfate (6S)-, (6R)- and (6RS)-tetrahydrofolate acid or salt of sulfonic acid in situ or after pre-selection or isolation and purification as a result of interaction with formaldehyde in the presence of an appropriate cyclodextrin. For the quantitative conversion of tetrahydrofolate acid is needed in this case only a slight molar excess of formaldehyde, a maximum of 10-20%. The reaction is carried out at this preferably in the range of pH 8 to 9. Connection of power, however, also be obtained by introducing (6R)-, (6S)- or (6R, S)-5,10-methylenetetrahydrofolic acid in cyclodextrines solution or suspension of cyclodextrin, or by grinding (6R)-, (6S)- or (6R,S)-5,10 - methylenetetrahydrofolic acid and cyclodextrin in solid form. The resulting products are stable in solution at room temperature. They are suitable as a component of parenteral dosage forms or the military form suitable for example, for cancer therapy, for the treatment of certain forms of anemia, autoimmune diseases and nervous disorders.

The object of the present invention are also stable aqueous formulations of the compounds include cyclodextrins containing:

a) (6R)-, (6S)- or (6R,S)-5,10-methylentetrahydrofolate acid or salt of (6R)-, (6S)- or (6R,S)-5,10 - methylenetetrahydrofolic acid, and

b) -, or-cyclodextrin or alkyl - or hydroxyethylphosphonate -, - or-cyclodextrin, or

C) a mixture-or - cyclodextrin or a mixture of alkyl - or hydroxyalkylated -, - or-cyclodextrin, or a mixture-or-cyclodextrin with alkyl - or hydroxyalprazolam -, - or-cyclodextrin.

Preferred compositions containing (6R)-5,10 - methylenechloride-folic acid or a pharmaceutically compatible salt of (6R)-5,10-methylenetetrahydrofolic acid and - or - cyclodextrin or a pharmaceutically compatible alkyl - or hydroxyethylphosphonate - or - cyclodextrin.

As a pharmaceutically compatible salt of 5,10 - methylenetetrahydrofolic acid composition usually contains a salt of alkali or alkaline earth metal, preferably sodium, magnesium or calcium and the quality is oxypropyl --, hydroxypropyl-or dimethyl-cyclodextrin.

All of the above compounds have therapeutic activity.

The invention also relates to a method of stabilizing aqueous solutions of (6R)-, (6S)- or (6R,S)-5,10-methylenetetrahydrofolic acid or its salts by adding-or - cyclodextrin or alkyl - or hydroxyalkylated -, - or - cyclodextrin,

and also to the method of preparation of stable solutions of (6R)-, (6S)- or (6R, S)-5,10-methylenetetrahydrofolic acid or a salt thereof, which consists in the interaction of (6R)-, (6S)- or (6R,S)- tetrahydrofolate acid or its salts with formaldehyde in the presence-or - cyclodextrin or alkyl - or hydroxyalkylated -, - or - cyclodextrin.

Normally interact (6R)-, (6S)- or (6R,S)- tetrahydrofolate acid or its salts with formaldehyde in the presence-or - cyclodextrin or alkyl - or hydroxyalkylated -, - or - cyclodextrin and the resulting connection enable turn into salt.

The free connection of the inclusion can be obtained by transformation of the corresponding salt.

The preferred salt components 5,10 - methylenetetrahydrofolic acid or tetrahydrofurane, as sulfates, sulfonates or halides.

Preferred cyclodextrine connections enable receive with cyclodextrins such as, hydroxypropyl or -- dimethyl --, - hydroxypropyl-or dimethyl-cyclodextrin. The cyclodextrin used is preferably at least in a molar ratio to 5,10-methylenetetrahydrofolic acid, for example, 1: 1 or 2:1. The use of highly concentrated solutions of cyclodextrin is preferred. You can also use mixtures of different cyclodextrins. Depending on the purpose of use may be preferred in some salt 5,10-methylenetetrahydrofolic acid, a specific cyclodextrin or a certain attitude 5,10-methylenetetrahydrofolic acid to the cyclodextrin. Optimal conditions can be determined by simple experiments.

The interaction of tetrahydrofolate acid with formaldehyde in the presence of cyclodextrin carried out preferably in a solvent consisting of water or miscible with water, an organic solvent, as the lower aliphatic carboxylic acid or a lower alcohol.

As tetrahydrofolate acid is susceptible to oxidation, requestflags using known methods, for example, evaporation of the solvent at elevated temperature in vacuo, crystallization, liofilizirovannam or by precipitation by addition of an organic solvent.

Get cyclodextrine connection enable you rubbing 5,10-methylenetetrahydrofolic acid and cyclodextrin in solid form.

The following examples illustrate the invention.

Example 1

The stability of cyclodextrin inclusion and 5,10-methylenetetrahydrofolic acid and its salts in solution

a) Stability of solutions of salts of sodium and calcium (6R)- and (6S)- 5,10-methylenetetrahydrofolic acid at the 23oC in phosphate buffer solution (see tab.1 in the end of the description).

b) Stability of solutions of sodium salt of (6R)-5,10-methylenetetrahydrofolic acid at 60oC (stress test) in phosphate buffer solution (see tab.2 at the end of the description).

in) Stability of the solution of the calcium salt of (6R)-5,10-methylenetetrahydrofolic acid at 60oC (stress test) in phosphate buffer solution (see tab.3 at the end of the description).

d) Stability of the solution of the magnesium salt of (6R)-5,10-methylenetetrahydrofolic acid at 60oC (stress test is indicated content 5,10-methylenetetrahydrofolic acid in percent of initial values (t=0).

To determine the stability of the solution was used with the 5,10-methylenetetrahydrofolic acid with a concentration of about 4% at pH=7 obtained in examples 3-20. The aim of the experiments was to obtain, respectively, comparable to the values specified in tables a-d). Under optimal conditions selected, which can easily be determined by simple experiments by changing the concentration of 5,10-methylenetetrahydrofolic acid or cyclodextrin, or by changing the concentrations of both components, or by selecting a salt of 5,10-methylenetetrahydrofolic acid, or by choice of solvent, the stability of the inclusion is clearly higher.

To obtain the substances in the following examples were used cyclodextrins with the following content: water-cyclodextrin 11,5%, - cyclodextrin 15.6 percent dimethyl--cyclodextrin was 12.7%, hydroxypropyl-- cyclodextrin 6.4% of the cyclodextrin 9.8%, dimethyl--cyclodextrin 9.2%, hydroxypropyl--cyclodextrin 6,0% (all values are determined using thermogravimetry).

Example 2

The study of the chemical interaction of the compounds include cyclodextrin and 5,10-methylenetetrahydrofolic acid in comparison with a mixture of cyclodextrin and 5-formyltetrahydrofolate acids is or13C-signal 5-formyltetrahydrofolate acid and1H-signal 5,10-methylenetetrahydrofolic acid when the connection switch with cyclodextrin at pH 9 (see tab.5 at the end of the description).

Value ppm13C-spectra 5-formyltetrahydrofolate acid clearly shows that the cyclodextrin with this substance does not interact. On the contrary, ppm13C-spectra and1N-spectra 5,10-methylenetetrahydrofolic acid indicate a strong interaction especially in pteridines part of the molecule.

Example 3

Connection enable - cyclodextrin and (6R)-5,10 - methylenetetrahydrofolic acid

22,7 g - cyclodextrin and 4,88 g (6S)-tetrahydrofolate acid are suspended in the atmosphere of nitrogen at room temperature in a 2000 ml of water. Then added dropwise 1.3 ml of a solution of formaldehyde (36,2%). With light heating the suspension to approximately 40oC get almost clear solution, which, after vigorous filtration on a rotary evaporator at 20oC and 1 mbar evaporated to dryness.

Get to 25.8 g of compound include cyclodextrin and (6R)- 5,10-methylenetetrahydrofolic acid content of (6R)-5,10 - methylenetetrahydrofolic acid and 13.3% (ODA is tetrahydrofolate acid

a) 22,7 g - cyclodextrin suspended in 650 ml of water. At room temperature in a nitrogen atmosphere add 4.8 g (6S)- tetrahydrofolate acid. At the same time supporting pH 7-9 (16,3 ml of 1N. NaOH). Then added dropwise with 0.93 ml of formalin solution (36,2%). The resulting solution was vigorously filtered and after 20 minutes the subsequent reaction at room temperature at 20oC/1 mbar evaporated to dryness.

Get 26,3 g connections enable - cyclodextrin and sodium salt of (6R)-5,10-methylenetetrahydrofolic acid content of (6R)-5,10-methylenetetrahydrofolic acid and 17.9% (determined using HLPC).

b) the Product is similar to that described in example 4A, receive as a result of intensive grinding 10 g of wet - cyclodextrin and 2.6 g of sodium salt of (6R)-5,10-methylenetetrahydrofolic acid.

Stability of solids: the content of (6R)-5,10 - methylenetetrahydrofolic acid after 113 days, -25oC - 100,0% through 63 days at +23oC - level 98.2%.

Example 5

Connection enable - cyclodextrin and the calcium salt of (6R)-5,10-methylenetetrahydrofolic acid

5,78 g - cyclodextrin and 320 mg of calcium hydroxide suspended in 30 ml of water. In nitrogen atmosphere is added dropwise of 1.97 g of (6S)-Tetra quickly filtered and from the filtrate precipitated compound include the addition of 300 ml of ethanol at 0oC. the Product is washed with a mixture of ethanol/water, at 20oC dried.

Get to 7.7 g of compound include - cyclodextrin and the calcium salt of (6R)-5,10-methylenetetrahydrofolic acid content of (6R)-5,10-methylenetetrahydrofolic acid 25,6% (determined using HLPC).

Example 6

Connection enable - cyclodextrin and potassium salts of (6R)-5,10-methylenetetrahydrofolic acid

Connection enable - cyclodextrin and potassium salts of (6R)-5,10-methylenetetrahydrofolic acid get similar to the one described in example 4.

Example 7

Connection enable - cyclodextrin and magnesium salts of (6R)-5,10-methylenetetrahydrofolic acid

Connection enable - cyclodextrin and magnesium salts of (6R)-5,10-methylenetetrahydrofolic acid get similar to the one described in example 5.

Example 8

Connection of on - and - cyclodextrin and the calcium salt of (6R)-5,10-methylenetetrahydrofolic acid

10,85 g - cyclodextrin, 12.9 liters - cyclodextrin and 744 mg of calcium hydroxide suspended in 100 ml of water. In nitrogen atmosphere add is 4.93 g (6S)-tetrahydrofolate acid, then 31 mg of calcium hydroxide and 0.92 ml of formalin solution (36,2%). Upon completion of the reaction charikleia; it is washed with a mixture of ethanol/water and 20oC dried.

Get 29,2 g connections enable -/- cyclodextrin and (6R)-5,10-methylenetetrahydrofolic acid content of (6R)-5,10-methylenetetrahydrofolic acid of 16.5% (determined using HPLC).

Example 9

Connection enable - cyclodextrin and sodium salt of (6S)-5,10-methylenetetrahydrofolic acid

of 2.27 g - cyclodextrin suspended in 100 ml of water. At room temperature in a nitrogen atmosphere add 608 mg (6S)-5,10 - methylenetetrahydrofolic acid. This supported the pH 7-9 (2,7 ml of 1N. NaOH). The resulting solution was quickly filtered and liofilizirovanny.

Get 2,75 g connections enable - cyclodextrin and sodium salt of (6S)-5,10-methylenetetrahydrofolic acid c content (6S)-5,10-methylenetetrahydrofolic acid 16,0% (determined using HPLC).

Stability of solids: the content of (6S)-5,10 - methylenetetrahydrofolic acid through 156 days at -25oC 100,0%.

Example 10

Connection enable-cyclodextrin and sodium salt of (6R)-5,10-methylenetetrahydrofolic acid sulphate

22,7 g - cyclodextrin suspended in 1130 ml of water. At room temperature in the atmosphere is spencie 7-9 (33 ml of 1N. NaOH). The resulting solution was quickly filtered and lyophilizers.

Obtain 28.4 g of the compound include - cyclodextrin and sodium salt of (6R)-5,10-methylenetetrahydrofolic acid content of (6R)-5,10-methylenetetrahydrofolic acid and 15.3% (determined using HPLC).

Stability of solids: the content of sodium salt of (6R)-5,10-methylenetetrahydrofolic acid after 154 days at -25oC of 99.5%.

Example 11

Connection enable - cyclodextrin and (6R)-5,10 - methylenetetrahydrofolic acid from a solution

of 2.27 g - cyclodextrin is dissolved in 150 ml of water at 25oC. In nitrogen atmosphere add 608 mg (6R)-5,10-methylenetetrahydrofolic acid (pH of 3.7). The resulting solution was quickly filtered and lyophilizers.

Obtain 2.7 g of the compound include - cyclodextrin and (6R)-5,10-methylenetetrahydrofolic acid content of (6R)-5,10 - methylenetetrahydrofolic acid 15,5% (determined using HPLC).

Stability of solids: the content of (6R)-5,10 - methylenetetrahydrofolic acid through 158 days at -25oC 100,0%.

Example 12

Connection enable - cyclodextrin and sulfate (6R)-5,10 - methylenetetrahydrofolic acid

As a result of application 5 is of the form cyclodextrin and sulfate (6R)-5,10 - methylenetetrahydrofolic acid content of (6R)-5,10-methylenetetrahydrofolic acid and 15.2% (determined using HPLC).

Stability of solids: the content of (6R)-5,10 - methylenetetrahydrofolic acid through 157 days at -25oC 99,0%.

Example 13

Connection enable-cyclodextrin and benzosulfimide (6R)-5,10-methylenetetrahydrofolic acid

The application of 603 mg of salt benzosulfimide acid (6R)-5,10-methylenetetrahydrofolic acid in analogy to example 11 receive

2,94 g connections enable-cyclodextrin and benzosulfimide (6R)-5,10-methylenetetrahydrofolic acid content of (6R)-5,10-methylenetetrahydrofolic acid 13,1% (determined using HPLC).

Stability of solids: the content of benzosulfimide (6R)-5,10-methylenetetrahydrofolic acid after 72 days at -25oC 100,0%.

Example 14

Compound include calcium salts - cyclodextrin and (6R)-5,10-methylenetetrahydrofolic acid

The application of 610 mg of the calcium salt of (6R)-5,10 - methylenetetrahydrofolic acid in analogy to example 11 obtain 2.65 g of the compound include - cyclodextrin and calcium is Adelino using HPLC).

Example 15

Connection enable - cyclodextrin and magnesium salts of (6R)-5,10-methylenetetrahydrofolic acid

The application of 600 mg of the magnesium salt of (6R)-5,10 - methylenetetrahydrofolic acid in analogy to example 11 get 2,69 g connections enable - cyclodextrin and magnesium salts of (6R)-5,10-methylenetetrahydrofolic acid content of (6R)-5,10-methylenetetrahydrofolic acid 13,1% (determined using HPLC).

Stability of solids: the stability of the magnesium salt of (6R)-5,10-methylenetetrahydrofolic acid through 155 days at -25oC of 99.5%.

Example 16

Compounds include dimethyl--cyclodextrin and sodium salt of (6R)-5,10-methylenetetrahydrofolic acid

26,2 g of dimethyl-cyclodextrin are dissolved in 100 ml of water. At room temperature in a nitrogen atmosphere added 5.6 g of sulfate (6R)-5,10-methylenetetrahydrofolic acid. At the same time supporting the pH 7-9 (37 ml 1N. NaOH). The resulting solution was quickly filtered and lyophilizers.

Get a 32.6 g of compound include dimethyl-cyclodextrin sodium salt (6R)-5,10-methylenetetrahydrofolic acid content of (6R)-5,10-methylenetetrahydrofolic acid and 14.3% (determined using HPLC).

One hundred is at -25oC 100%, after 56 days at -25oC 97,5%.

Example 17

Compound include dimethyl -- cyclodextrin and salts of sulfuric acid (6R)-5,10-methylenetetrahydrofolic acid

2,62 g of dimethyl-cyclodextrin dissolved in 10 ml of water. At room temperature in a nitrogen atmosphere add 560 mg sulfate (6R)-5,10-methylenetetrahydrofolic acid and directly after that add 140 ml of water. The resulting solution was quickly filtered and lyophilizers.

Get 3,29 g of compound include dimethyl--cyclodextrin and sulfate salts of (6R)-5,10 - methylenetetrahydrofolic acid content of (6R)-5,10 - methylenetetrahydrofolic acid 13.6% as determined using HPLC).

Example 18.

Connections include hydroxypropyl -- cyclodextrin and (6R)- or (6S)-5,10-methylenetetrahydrofolic acid

The application of hydroxypropyl -- cyclodextrin by analogy with examples 3 -17 receive connections include hydroxypropyl -- cyclodextrin (6R)- or (6S)-5,10-methylentetrahydrofolate acids and their salts.

Example 19

Connections include hydroxypropyl -- cyclodextrin and (6R)- or (6S)-5,10-methylenetetrahydrofolic acid

As a result of applying hydroxypropyl (6R)- or (6S)-5,10 - methylenetetrahydrofolic acid or their salts.

Example 20

Compounds include dimethyl -- cyclodextrin and (6R)- or (6S)- 5,10-methylenetetrahydrofolic acid

The application dimethyl-cyclodextrin by analogy with examples 3-17 receive compounds include dimethyl -- cyclodextrin (6R)- or (6S)-5,10-methylenetetrahydrofolic acid or their salts.

1. Stable aqueous formulations of the compounds include cyclodextrins containing

a) (6R)-, (6S)- or (6R,S)-5,10-methylentetrahydrofolate acid or salt of (6R)-, (6S)- or (6R,S)-5,10-methylenetetrahydrofolic acid,

b) -, or-cyclodextrin or alkyl - or hydroxyethylphosphonate -,- or-cyclodextrin, or

C) a mixture-or-cyclodextrin or a mixture of alkyl - or hydroxyalkylated -,- or-cyclodextrin, or a mixture-or-cyclodextrin with alkyl - or hydroxyalprazolam -,- or-cyclodextrin.

2. The compositions according to p. 1, characterized in that the contain (6R)-5,10-methylentetrahydrofolate acid or a pharmaceutically compatible salt of (6R)-5,10-methylenetetrahydrofolic acid and - or-cyclodextrin or a pharmaceutically compatible alkyl - or hydroxyethylphosphonate - or-cyclodextrin.

3. The compounds under item 1 or 2, characterized in that as farmacia metal, preferably the salt of sodium, magnesium or calcium and as a pharmaceutically compatible alkyl - or hydroxyalkylated cyclodextrin preferably hydroxypropyl --, hydroxypropyl-or dimethyl-cyclodextrin.

4. The compositions according to the PP.1 to 3, characterized in that have therapeutic activity.

5. Connection enable-or-cyclodextrin or its alkyl - or hydroxyalkylated and (6R) or (6S)-5,10-methylenetetrahydrofolic acid or its salts, when a molar ratio of 1 : 1 to 1 : 2.

6. Connections include under item 5, a connection switch hydroxyalkyl-- or hydroxyalkyl-- cyclodextrin and (6R)- or (6S)-5,10-methylenetetrahydrofolic acid or a salt thereof.

7. Connections include under item 6, which are compounds include hydroxypropyl-- or hydroxypropyl--cyclodextrin and (6R)- or (6S)-5,10-methylenetetrahydrofolic acid or a salt thereof.

8. Connections include under item 5, which are compounds include alkyl-or alkyl-cyclodextrin and (6R)- or (6S)-5,10-methylenetetrahydrofolic acid or a salt thereof.

9. Connection turn on p. 8, representing a compound include dimethyl-- or dimerization aqueous solutions of (6R)-, (6S)- or (6R,S)-5,10-methylenetetrahydrofolic acid or its salts by adding-or - cyclodextrin or alkyl - or hydroxyalkylated -,- or-cyclodextrin.

11. Method of preparation of stable solutions of (6R)-, (6S)- or (6R,S)-5,10-methylenetetrahydrofolic acid or a salt thereof, which consists in the interaction of (6R)-, (6S)- or (6R,S)-tetrahydrofolate acid or its salts with formaldehyde in the presence-or-cyclodextrin or alkyl - or hydroxyalkylated -,- or-cyclodextrin.

12. The method of obtaining on p. 11, wherein interact (6R)-, (6S)- or (6R,S)-tetrahydrofolate acid or its salts with formaldehyde in the presence-or-cyclodextrin or alkyl - or hydroxyalkylated -,- or-cyclodextrin and the resulting connection enable turn into salt.

13. The method of obtaining compounds include (6R)-, (6S)- or (6R,S)-5,10-methylenetetrahydrofolic acid or their salts, which consists in the fact that you get a stable aqueous solution of the compounds included in the result of the interaction of (6R)-, (6S)- or (6R,S)-tetrahydrofolate acid or its salts with formaldehyde in the presence-or-cyclodextrin or alkyl - or hydroxyalkylated -,- or-zikawei acid.

 

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The invention relates to the field of biotechnology and can be used to obtain from filamentous fungi polysaccharides, including polyaminoamide - chitin and chitosan

The invention relates to the field of biotechnology and chemical technology, particularly to a method of depolymerization of dextrans, namely the method of production of dextran from srednevekovoi molecular weight (m m) 1000 D, which can be used in the medical industry, the drug is able to prevent severe anaphylactic reactions during transfusion dextranomer products with higher m m
The invention relates to a method for producing a hydrophilic gel used in medical industry for the purification of insulin from proinsulin and pronsolino proteins

The invention relates to the field of polymer chemistry, particularly the chemistry of polysaccharides

FIELD: organic chemistry, chemical technology, medicine, pharmacy.

SUBSTANCE: invention describes derivatives of 8-phenyl-6,9-dihydro[1,2,4]-triazolo[3,4-I]purine-5-one of the general formula:

wherein R1 means hydrogen atom, group -CH2-R6 wherein R6 means phenyl; R2 means (C1-C5)-alkyl or group -(CH2)n-R6 wherein n= 1 or 2; R6 means (C1-C4)-alkoxy-group or pyridyl group; R3 means (C1-C6)-alkyl; R4 means hydrogen atom or (C1-C4)-alkyl; R5 means -(CH2)n-R7 wherein n = 0-4; R7 means 3-7-membered ring comprising 1-3 heteroatoms taken among nitrogen atom (N) and oxygen atom (O), (C3-C7)-cycloalkyl or phenyl wherein indicated groups can be substituted with different substitutes; or R4 and R5 mean independently hydrogen atom (H), (C2-C6)-alkynyl or (C1-C6)-alkyl that can be substituted possibly; or R4 and R5 in common with nitrogen atom (N) form 4-7-membered ring comprising 1-2 heteroatoms taken among N and O and substituted possibly. Also, invention relates to their pharmaceutically acceptable salts, methods for preparing these compounds, intermediate substances, pharmaceutical composition and a to a method for treatment of different diseases mediated by activity of phosphodiesterase-5 (PDE-5). Described compounds of the formula (I) are inhibitor of PDE-5.

EFFECT: improved preparing method and treatment, valuable properties of compounds.

20 cl, 5 tbl, 149 ex

FIELD: organic chemistry, labeled compounds.

SUBSTANCE: invention relates to a new highly labeled compound that represent an analog of the known physiologically active compound that is the strongest toxin and inhibitor of some viable important processes, for example, sodium ions transporting. [3H]-Saxitoxin dihydrochloride highly labeled with tritium corresponds to the formula: .

EFFECT: valuable properties of compound.

1 ex

FIELD: organic chemistry, medicine, pharmacy.

SUBSTANCE: invention describes derivatives of benzodiazepine of the general formula (I): wherein R1 represents hydrogen, halogen atom, (C1-C7)-alkyl, hydroxy- or (C1-C7)-alkylthio-group; R2 represents -C(O)O-(C1-C7)-alkyl, 1,2,4-oxadiazole-3-yl or 1,2,4-oxadiazole-5-yl wherein their cyclic fragments are substituted with (C3-C7)-cycloalkyl; R3 represents hydrogen atom (C1-C7)-alkyl, -(CH2)n-(C3-C7)-cycloalkyl, -(CH2)n-halogen, -(CH2)n-pyridine-4-yl or -(CH2)n-phenyl wherein phenyl ring can be substituted with one or some substitutes chosen from the group comprising (C1-C7)-alkoxy-group, halogen atom, -SO2CH3, phenyl, -OCF3, nitro-group, -CF3, -NR2, or it means -(CH)n-indolyl optionally substituted with (C1-C7)-alkyl or (C1-C7)-alkoxy-group, or means pyrrolidinyl-5-oxo-group, -C(O)-NR2, -(CH2)n-OH, -(CH2)n-NR2 or -(CH2)n-benzo[1,3]dioxol; R represents hydrogen atom or (C1-C7)-alkyl; n = 0, 1, 2 or 3, and its pharmaceutically acceptable acid-additive salts with exception for the following compounds: 9H-imidazo[1,5-a][1,2,4]triazolo[3,4-d][1,4]benzodiazepine-10-carboxylic acid ethyl ester. 10-(3-cyclopropyl-1,2,4-oxadiazole-5-yl)-9H-imidazo[1,5-a][1,2,4]triazolo[4,3-d][1,4]benzodiazepine, 3-fluoro-9H-imidazo[1,5-a][1,2,4]triazolo[4,3-d]benzodiazepine-10-carboxylic acid ethyl ester, 10-(3-cyclopropyl-1,2,4-oxadiazole-5-yl)-3-fluoro-9H-imidazo[1,5-a][1,2,4]triazolo[4,3-d][1,4]benzodiazepine, 3-chloro-9H-imidazo[1,5-a][1,2,4]triazolo[4,3-d][1,4]benzodiazepine-10-carboxylic acid ethyl ester, 3-chloro-1-(3-cyclopropyl-1,2,4-oxadiazole-5-yl)-9H-imidazo[1,5-a][1,2,4]triazolo[4,3-d][1,4]benzodiazepine and 3-methyl-9H-imidazo[1,5-a][1,2,4]triazolo[3,4-d][1,4]benzodiazepine-10-carboxylic acid ethyl ester. Also, invention relates to pharmaceutical composition comprising both new and above enumerated and excluded compounds. New compounds possess ability for selective binding with α5-subunit of gamma-aminobutyric acid receptor A and can be used in treatment, for example, Alzheimer's diseases.

EFFECT: valuable medicinal properties of compounds and pharmaceutical composition.

13 cl, 1 tbl, 67 ex

FIELD: organic chemistry, medicine.

SUBSTANCE: invention relates to derivatives of imidazo[1,5-a]pyrimido[5,4-d][1]benzazepine of the formula (I): , wherein R1 means halogen atom or (lower)alkyl; R2 means hydrogen atom, (lower)alkyl, (C3-C7)-cycloalkyl, -(CH2)m-phenyl wherein phenyl ring can be substituted with (lower)alkoxy-group or it means -(CH2)m-indolyl; R3 means -C(O)O-(lower)alkyl, -C(O)OH or five-membered heteroaromatic group comprising nitrogen and oxygen atoms as heteroatoms and wherein rings can be substituted with (lower)alkyl or (C3-C7)-cycloalkyl; n means 0, 1 or 2; m means 0, 1 or 2, and their pharmaceutically acceptable acid-additive salts. Compounds of this class elicit the high degree and selectivity to binding sites of GABA A α5-receptor and can be used in treatment of the conception enhancer or disorders of cognitive ability similar with Alzheimer's disease.

EFFECT: valuable medicinal properties of derivatives.

10 cl, 6 sch, 41 ex

FIELD: organic chemistry, medicine, pharmacy.

SUBSTANCE: invention relates to triheterocyclic compound of the formula (I): wherein X represents carbon atom; Y represents carbon or nitrogen atom; W represents carbon or nitrogen atom; U represents -CR2, and Z represents -CR2 or nitrogen atom; ring A represents (C5-C6)-cycloalkyl ring or 5-membered heterocyclic ring comprising one nitrogen, oxygen or sulfur atom; R1 represents alkyl, alkenyl, alkynyl, -NR4R5, -OR6 and others; R3 represents phenyl ring substituted with 1-3 substitutes or pyridyl or 1,3-dioxoindanyl ring substituted with 1-2 substitutes, and its pharmaceutically acceptable salts and pharmaceutical composition containing thereof as an active component. Also, invention relates to derivatives of pyrazolopyrimidine and derivatives of pyrrolopyrimidine. Compounds of the formula (I) show antagonistic activity with respect to corticotropin-releasing factor receptors. The compound can be used in treatment and/or prophylaxis of depression, anxiety state, disorders in food intake, post-traumatic stress, ulcerous disease, irritable bowel syndrome, Alzheimer's disease, abuse in drugs using or alcoholic syndrome dependence.

EFFECT: valuable medicinal properties of compounds and pharmaceutical agent.

7 cl, 1 dwg, 24 ex

FIELD: organic chemistry, medicine, biochemistry, pharmacy.

SUBSTANCE: invention describes novel imidazo-condensed compounds of the general formula (I): wherein Z represents nitrogen atom (N); Z1 represents N whein a bond between C5 and Z1 represents a simple bond, and Z1 represents carbon atom (C) when a bond between C5 and Z1 represents a double bond; R1 represents hydrogen atom; R2 represents (C1-C6)-alkyl, (C1-C6)-hydroxyalkyl, phenyl-(C1-C4)-alkyl substituted with halogen atom, ((C1-C4)-alkyl)-SO2, (C1-C6)-alkyl, (C5-C6)-cycloalkyl possibly substituted with hydroxy-group, phenyl substituted with halogen atom, heterocyclyl possibly substituted and chosen from group consisting of tetrahydropyranyl, (N-methylsulfonyl)piperidinyl or tetrahydro-1,1-dioxide-2H-thiopyranyl; A is absent or represents -O-; a bond between C5 and Z1 is a simple or double bond; a bond between C8 and C9 is a simple or double bond; Y represents phenyl substituted with halogen atom, or their pharmaceutically acceptable salts possessing inhibitory activity with respect to p38 MAP kinase, and pharmaceutical composition containing thereof. Proposed compounds can be used, for example, in treatment/or prophylaxis of such diseases as rheumatic arthritis, fever and reduced bone resorption.

EFFECT: valuable medicinal properties of compounds and pharmaceutical composition.

16 cl, 2 tbl

FIELD: organic chemistry, medicine, pharmacy.

SUBSTANCE: invention describes novel derivatives of 5-aminopyrazolo-[4,3-e]-1,2,4-triazolo-[1,5-c]-pyrimidine of the general formula: wherein R means furanyl, possibly substituted pyrrolyl, possibly substituted pyridyl, possibly substituted phenyl or (C4-C6)-cycloalkenyl; X means (C2-C6)-alkylene or -C(O)CH2-; Y means the following groups: -N(R2)CH2CH2N(R3)-, -OCH2CH2N(R2)- wherein R2 and R3 mean hydrogen atom or (C1-C6)-alkyl, -O-, -S-, -CH2S-, -(CH2)2-NH- or compound of the formula: wherein Q means or R4 means hydrogen atom or (C1-C6)-alkyl, or two R at one carbon atom form group =O; Z means phenyl comprising from 1 to 5 of different substitutes, phenylalkyl or heteroaryl, diphenylmethyl and other values; or Z and Y in common can form substituted piperidinyl or substituted phenyl also possessing activity of antagonist of A2a adenosine receptors. Also, invention relates to a pharmaceutical composition based on these compounds, using novel compounds for preparing medicinal agents in treatment, for example, Parkinson's disease, and two methods for synthesis of intermediate compounds of formulae (II) and (IIIa) .

EFFECT: improved methods of synthesis, valuable medicinal properties of compounds and pharmaceutical composition.

17 cl, 17 tbl, 29 ex

FIELD: medicine; pharmacology.

SUBSTANCE: in formula (I) V represents -N (R1) (R2) or OR4; R4 represents H, C1-6alkyl, C1-6halogenalkyl or (C1-6alkylen)0-1R4' R4' represents C3-7cycloalkyl, phenyl, pyridyl, piperidinyl; and R4' is optionally substituted with 1 or 2 identical or different substitutes chosen from group consisting of C1-4alkyl, amino, C1-3alkylamino, C1-3dialkylamino, phenyl and benzyl; and each R1 and R2 independently represents L1, where L1 is chosen from group consisting from H, C1-6alkyl, C2-6alkenyl, C2-6alkinyl, - adamantyl, pyrrolidinyl, pyridyl, or R1 and R2 together with nitrogen atom to which attached, form X, where X represents pyrrolidinyl, piperazinyl, piperidinyl, morpholino; where X is optionally substituted with Y, where Y represents dioxolanyl, C1-9alkyl, phenyl, furanyl, pyrrolyl, pyridyl, pyrrolidinyl; and where X and Y are optionally split with Z, where Z represents -C1-3alkylen-, C1-3alkylen-. Other radical values are specified in formula of invention.

EFFECT: effective application for treatment of migraine and other headache mediated by action of CGRP-receptors.

34 cl, 11 dwg, 6 tbl, 201 ex

FIELD: chemistry.

SUBSTANCE: invention relates to new compounds of formula (I) and their pharmaceutically acceptable salts with antagonistic properties towards adenosine A2A receptor, which can be used for treating central nervous system diseases such as Parkinson's disease. In general formula (I) , R is ,, R1, R2, R3, R4 and R5 is independently selected from a group which consists of hydrogen; R6 is hydrogen, (C1-C6)alkyl or -CH2F; R7, R8 and R9 are independently selected from a group which consists of hydrogen, (C1-C6)alkyl, (C1-C6)alkoxy, haloid and -CF3; Z is R10-phenyl, R10-5-6-member heteroaryl, which contains 1 or 2 hetwroatoms, selected from nitrogen or from nitrogen and oxygen, possibly condensed with a benzene ring, or ; R10 represents 1 to 3 substitutes, independently selected from a group which consists of hydrogen, (C1-C6)-alkyl, hydroxy, (C1-C6)-alkoxy, hydroxy-(C1-C6)-alkyl, hydroxy-(C1-C6)-alkoxy, (C1-C6)-alkoxy-(C1-C6)-alkyl, (C1-C6)-alkoxy-(C1-C6)-alkoxy, (C1-C6)-alkoxy-(C1-C6)-alkoxy-(C1-C6)-alkyl, (di-(C1-C6)-alkoxy)-(C1-C6)-alkyl, (hydroxy)-(C1-C6)-alkoxy-(C1-C6)-alkyl, (C3-C6)-cycloalkyloxy, (C3-C6)-cycloalkyl-O-(C3-C6)-alkoxy, (C1-C6)-alkyl-SO2-, (C1-C6)-alkyl-SO-, haloid, -CN, cyano-(C1-C6)-alkyl, -CHF2, -CF3, -C(O)R13, -C(O)O-(C1-C6)-alkyl, -N(R11)(R12), N(R11)(R12)- (C1-C6)-alkyl, - C(O)N(R13)(R16), R11-5-6-member nitrogen-containing heteroaryl, possibly condensed with a benzene ring, R15-5-6-member heterocycoalkyl, with 1 or 2 heteroatoms selected from nitrogen and oxygen, R15-5-6-member heterocycloalkyl-(C1-C6)-alkyl, with 1 or 2 heteroatoms selected from nitrogen and oxygen, R15-5-6-member heterocycloalkyl-(C1-C6)-alkoxy, with 1 or 2 heteroatoms selected from nitrogen and oxygen, R15-5-6-member heterocycloalkyl-oxy, with 1 or 2 heteroatoms in a heterocyclic ring selected from nitrogen and oxygen, CF3-(C1-C4)alkylene-O-(C1-C6)alkyl, CF3-hydroxy(C1-C6)alkyl, cyano-(C1-C6)-alkoxy, (C1-C4)alkylene-C(O)-O-(C1-C6)alkyl, -SO2-N((C1-C4)alkyl)2, ((C3-C4)cycloalkyl)hydroxy(C1-C6)alkyl, (hydroxy(C1-C6)alkyl)-(C1-C4)alkoxy, (dihydroxy)-(C1-C6)-alkyl, (dihydroxy)(C1-C6)alkoxy, -C(=NOR17)- (C1-C6)alkyl and -C(=NOR17)-CF3; or two R10 groups, on neighbouring carbon atoms of the ring, together form -O-CH2-O-, -O-(CH2)2-O-, -CH2-O(CH2)2-O-, -O-(CH2)2-, -(CH2)3-O-, -O-(CH2)3-O, -(CH2)3-, where the ring, formed by two R10 substitutes and ring carbon atoms with which they are bonded, is substituted with R16; or two R10 groups on neighbouring ring carbon atoms, together form -O(CH2)3CH((OR18)-, each R11 is independently selected from a group which consists of hydrogen and (C1-C6)alkyl; each R12 is independently selected from a group which consists of (C1-C6)alkyl, hydroxy(C1-C6)alkyl, -C(O)-(C1-C6)alkyl, -C(O)O-(C1-C6)alkyl, ((C1-C6)alkoxy)hydroxy(C1-C6)alkyl, (C1-C6)alkoxy (C1-C6)alkyl-C(O)-, -SO2(C1-C6)alkyl; R13 is hydrogen, (C1-C6)alkyl or -CF3; R14 is (C1-C6)alkoxy-C(O)-; R15 represents 1 to 3 substitutes, independently selected from a group which consists of (C1-C6)alkoxy, hydroxy-(C1-C6)alkyl; or two R15 substitutes, taken together with the carbon atom with which they are bonded, form a -C(=O)- group; R16 is (C1-C6)alkoxy(C1-C6)alkyl, hydroxy or hydroxy(C1-C6)alkyl; R17 is hydrogen or (C1-C6)alkyl. The invention also relates to a pharmaceutical composition based on said compounds.

EFFECT: increased effectiveness of composition and method of treatment.

17 cl, 23 ex

FIELD: chemistry.

SUBSTANCE: invention relates to compounds of formula (I) and their pharmaceutically acceptable salts of formula (I) where n equals 0, 1 or 2, A is a five- or six-member aromatic ring which optionally contains one or two heteroatoms independently selected from nitrogen, oxygen or sulphur, B is a 5-9-member ring containing 0 or 1 double bonds and optionally contains an additional heteroatom selected from nitrogen and oxygen; where the ring optionally contains one or two substitutes independently selected from a group comprising C1-C6-alkoxy, C1-C6-alkoxycarbonyl, C1-C6-alkyl, carboxy, cyano, hydroxy, hydroxy-C1-C6-alkyl, di-C1-C6-alkylamino-C1-C6-alkyl, (NR4R5)-carbonyl or oxo; R1 is selected from -C(O)NR4R5 - CO2R4, 5-tetrazolyl, cyano; each R2 is independently selected from a group comprising C1-C6-alkyl, amino, benzyloxy, halogen, hydroxyl; R3 is a 5-7-member cycloalkyl ring; values of the rest of the radicals are given in the formula of invention. The invention also relates to a method for synthesis of the said compounds, a method of inhibiting HCV replicon function and a method of inhibiting functioning of the HCV NS5B protein.

EFFECT: wider field of use of the compounds.

16 cl, 4 tbl, 29 ex

FIELD: organic chemistry, drugs.

SUBSTANCE: invention relates to complex of general formula (X)n-Y, wherein X are identical or different and represent folic acid, folinic acid, dihydrofolic acid, tetrahydrofolic acid, tetrahyldopterine, pteroyl-polyglutamic acid, 2-deaminohydroxyfolic acid, 1-denitrofolic acid, 3-denitrofolic acid, 8-denitrofolic acid; Y represents polysaccharide different from arabinogalactane, selected from group containing glucanes, dextranes, polysaccharoses, fructosanes, heteropolysaccharides, homo- or heteropolysaccharide sulfates, ginseng pectin polysaccharide and other pectin polysaccharides, gumghatti, alginate, etc; or Y is selected from polyethylene oxide, methoxypolyethylene glycol and hydrophilic alcohol polymers; n >=1. Said complex permeates into in cell via pathway involving folic acid receptor on cell membrane. Complex of present invention is useful in production of pharmaceutical agents having antitumor activity.

EFFECT: new intermediates for antitumor agent production.

10 cl, 7 ex, 11 dwg, 2 tbl

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