The method of receiving cyclosporine a high purity from a crude product containing complex cyclosporiasis

 

(57) Abstract:

The invention relates to a method for cyclosporine And high purity by purification of the crude product containing cyclosporiasis complex by multi-step chromatography on silica gel at high load columns from 10 to 52%, using as eluent a mixture of toluene with acetone in an amount of from 10 to 30 vol.% or toluene with ethyl acetate in an amount of from 10 to 35 vol.%, cyclosporine And high purity with content cyclosporine L, U and D less than 0.05% and the content of cyclosporine and < 0,02% vol., industrial method of purification of cyclosporin a from the crude product, containing cyclosporiasis complex. 3 S. and 7 C.p. f-crystals, 1 table.

The object of this invention is a method for the chromatographic purification of cyclosporin a from the crude product, containing cyclosporiasis complex, using a column filled with silica gel, and by applying multi-step chromatography with a column filled with normal phase silica gel and solvent mixtures containing as a main component toluene.

Cyclosporine are N-methylated in several places undecapeptide and most of the E. letters from a to Z. First are allocated cyclosporine a, which is a natural material selected from the culture liquid medium (broth) strain lypocladium inflatum Gams (Helv. Chim. Acta 59, 1075/1976/). At first, the connection became known as mild antifungal antibiotic and later attention was drawn to its immunosuppressive action. (J. F. Borel et al. Immunology 32, 1017/1977/). For this reason, it is mainly used for the transplantation of organs from other entities (lung, heart, kidney, bone marrow, skin). Pharmacological studies have confirmed that it inhibits both tissue and cellular immune response by impeding the growth of T cells and interruption of the synthesis of interleukin-2. Later it became clear that it is effective in various types of autoimmune and inflammatory diseases, for example autoimmune hematologic disease, ulcerative colitis, graves disease, multiple sclerosis, psoriasis, and rheumatoid arthritis. For the treatment of infectious diseases caused by protozoa, and tumors were conducted additional experiments. The importance of this group is shown by the fact that through modification of various amino acids and Vice ">

Most city cyclosporin produced by fermentation. As examples of cultures of microorganisms used Cylindrocarpon Lucidum Booth (patent N CH 589716); Trichoderma polysporum Rifai (patent N CH 603790); Tolypocladium varium (patent N HU 201557). At the end of fermentation, depending on the nature of the process is formed cyclosporiasis complex, which may also contain other impurities (the ingredients of the culture medium, protivovspenivayushchie substance, metabolites, and so on).

Typically, the product is separated from the broth by extraction process. It can be done by selection from the broth mycelium by centrifugation or filtration, and then the dissolution of the active ingredient from the mycelium with methanol or acetone and extraction of the filtrate vodosmeshivayuschego solvents. Another known method is the implementation of the method without filtering, which is applied to the extraction of the whole broth odonatologica organic solvent. The solvent contained in the organic phase is evaporated by vacuum distillation. However, during the extraction with an organic solvent compounds having lipid properties, is also transferred to the organic phase, which causes truncata Switzerland N 589716 or published patent application Germany N 2455859), after removal of the extracting solvent, the residue dissolved in a mixture of methanol-water and then extracted several times the same volume of petroleum ether. Portions of petroleum ether are combined and from them through a mixture of methanol-water extract cyclosporine. The active substance through a multistage extraction transferred from phase a mixture of methanol-water telengard, which is then washed with water and evaporated to a dry residue. Obtained by the above method the crude product containing cyclosporine, can be cleaned more effectively one of chromatographic methods.

In accordance with the method described in U.S. patent N 4117118, cyclosporines the first mixture is transferred to a column of Sephadex LH-20 (Sephadex) and elute with methanol, then sequentially elute in the column of aluminum oxide with a mixture of toluene and ethyl acetate (15%) and in the column of silica gel with a mixture of chloroform and ethanol (2%). Despite repeated chromatography, the obtained product is not pure, but a mixture of cyclosporine a and B.

Such chromatographic method disclosed among others in U.S. patent N 4215199, in which rough purification is carried out in a column of silica gel with a mixture of chloroform is dissolved in methanol and subjected to chromatography in a column of Sephadex LH-20 using methanol as eluent. Fractions of the eluate is evaporated to

the dry residue, and then the residue is dissolved in a mixture of chloroform and methanol at a volume ratio of components in a mixture of 98:2. Then again subjected silicagel chromatography. In the first eluate appears cyclosporine A. This and subsequent fraction is separated by evaporation of eluent get pure components.

In accordance with German patent N DD 298276 oily crude product is dissolved in a small amount of chloroform, and then subjected to chromatography in a column of alumina using chloroform. Containing cyclosporine a fraction is evaporated in vacuo, dissolved in chloroform, subjected to a similar column chromatography and then elute with chloroform. The fractions containing the active substance, again evaporated in vacuo. To the residue add hexane and crystallized cyclosporine A. the Product is washed with hexane, then dried and finally re-crystallized from a mixture of simple ether and hexane or acetone.

In accordance with the patent Hungary N 201577 crude product obtained after evaporation, can be purified in a column of silica gel by elution with a mixture of chloroform and methanol at a gradually increasing concentrationa in the eluate in the subsequent steps of about 0.5. %. Cyclosporine And elute from the column with chloroform containing methanol in the amount of 2 vol.%, cyclosporine In chloroform containing methanol at a rate of 2.5% vol., cyclosporine With chloroform containing methanol in an amount of 3 vol.%. Components obtained by evaporation of the fractions.

The above methods describe methods mainly fermentation cyclosporine where the main purpose of the purification steps is the identification of the obtained product. Therefore, the product emit only a small amount, and are only physical and chemical properties without publishing the data related to the purity of the product and the quantities of impurities.

Rueger and With. /Helv. Chim. Acta 59(4), p-1075-92 (1976)/ for the identification and structural analysis identified a small number of pure cyclosporine a and C, by repeated chromatography and other stages of purification. In accordance with this article, the crude product obtained by the fermentation of Trichoderma polysporum Rifai, containing mainly cyclosporine a and C, degreased with methanol and petroleum ether. After evaporation the residue is dissolved in chloroform and subjected to chromatography by gradient elution using as eluent a mixture of chloroform pure crystalline cyclosporin A. The fraction containing cyclosporin a, dissolved in methanol and subjected to chromatography in a column of Sephadex LP-20 using methanol as eluent. Fraction is evaporated, dissolved in toluene and subjected to chromatography in a column filled with alumina, using as eluent toluene in the presence of increasing concentrations of acetic acid. After evaporation of the fractions and processing of activated carbon in an alcohol solution get crystalline product.

The treatment method is carried out in industrial scale, is described in U.S. patent 5382655. In accordance with the method the crude product, containing various cyclosporine components are heat-treated to column chromatography with silica gel using as eluent mixtures of chloroform-ethanol-dichloromethane and chloroform-ehtilacetatnogo. Then the obtained product is subjected to further chromatography and recrystallization, the result is high-quality, clean product, suitable for injection.

Purification of the crude product containing a mixture of cyclosporin is very difficult, as impurities, having a similar chemical structure, have the chromatographic hartom. The methods mentioned earlier, prove that regardless of the applied solvent mixture and by overlapping chromatographic peaks for certain components in pure form should implement the additional phase chromatography or purification. The purification methods known so far, are generally characterized not only by the fact that in their application require a lot from the solvent, and also that it is necessary to use 3-4 different types of solvents or solvent mixtures and 2-3 types of gaskets column. As a consequence of these facts in the way required several types of chromatographic methods and ways of regeneration, which create difficulties in developing structurally simple and equally managed cost-effective technologies for implementation on an industrial scale.

When the use of chlorinated hydrocarbons from the point of view of environmental protection additional problems arise, therefore, an increasing number of countries are making attempts to limit their use.

As for the material of the packing of the column, the use aluminiumoxide gaskets for industrial purposes is doubtful, because the ü small. In addition, it is disadvantageous for industrial purposes, because aluminiumoxide gasket is hard and brittle, causing the need for special equipment and technology. Such gaskets may not be used often bleeded columns of stainless steel commonly used in the chemical industry.

Gaskets type Sephadex are very expensive and if cyclosporine complex their effectiveness is very limited, as the dimensions of the various molecules are very close.

The aim of the present invention is to provide easily applicable on an industrial scale chromatographic purification method, which is suitable for the production of the ingredient of cyclosporine And containing less impurities, consequently, it can be used safely in medical practice.

Our goal is to create such a chromatographic purification technology, which requires only one type of solvent mixture and one type of padding columns.

Due to its advantageous properties, high specific surface area, large pore size, high absorptive capacity, light gaskets, that is appropriate for the Department cyclosporine components with high selectivity, it is necessary to choose the ideal solvent mixture and method.

In our experiments, we found that our goal can be realized through a multi-stage column chromatography with silica gel using as eluent solvent mixture, the main component of which is toluene. Unexpectedly, it was found that through a three-step chromatography in a column of silica gel using as eluent toluene containing acetone, can be separated even components cyclosporine U and L, which are closest to cyclosporine A. These components differ from cyclosporine And only methyl group.

In accordance with the first example application No. WO 94/16091 solvent mixture of toluene and acetone is also used on one stage. The resulting product is recrystallized from a solvent mixture of simple ether and hexane (Yield: 66.6 per cent). Optical rotation of the product is sufficient, but its melting point is significantly lower than that described in the literature, which indicates that the product is impure. Thus, even as The method in accordance with the present invention is suitable for the separation of the most common impurities, for example, cyclosporin b and C, which are present in large quantities and, in addition, cyclosporine components of D, U and L, which are present in trace quantities. The content of cyclosporine In and in the final product the cyclosporin a obtained in this way is less than 0.02%, while the content of cyclosporin L, U and D is below 0.05%.

The object of our invention is an improved method of purification of cyclosporin a from the crude product, containing cyclosporiasis complex, which also ensures the production of high-purity cyclosporine And on a large scale chromatographic method in silicagel column using multi-step chromatography, which is carried out by using a solvent mixture containing as a main component toluene. Another new feature of this method is the use of high column load. In the conventional chromatographic practice degree column load is not more than 5-10% of the loading column, and the value for cyclosporine is lower.

Multi-step chromatography, a solvent mixture containing as a main component toluene, and high colon the application. Therefore, to obtain a very pure product should simultaneously apply all three of the above properties.

In accordance with the present invention preferably sequentially applied 2-4 chromatographic steps and more appropriate three steps.

Overloading of the column is highest in the first step of chromatography. Exact separation of the active substance and the impurities also provides high column load on two subsequent stages chromatography.

For purification it is advantageous to use a solvent mixture of toluene-acetone, which contains not more than 30 vol.% of acetone.

In accordance with other suitable method is used a solvent mixture of toluene-ethyl acetate, in which the concentration of ethyl acetate below 35% vol.

In the cleaning method favorably at least once to apply a gradient elution.

According to our experiments it was found that for cleaning cyclosporine complex in our case, it is advantageous to use 10-30 vol.% and more suitable 13-18% vol. acetone and 10-35% vol. or 15-20 vol.% ethyl acetate.

In accordance with a possible method of the present invention in the case of trestain the pout stage separates the main part of cyclosporine, at the second stage remove most cyclosporine components, L and U, and the last third stage of some number of components of L and U and other unidentifiable impurities can be reduced to a content of less than 0.05 vol.%. At the first stage of loss of cyclosporine a are minimal, however, in a pre-obtained fractions of the second and third stages together with cyclosporiasis component D, which is very close to cyclosporine a, removes a significant amount of cyclosporine A. This cyclosporine And can be retrieved at the fourth stage in a very pure form.

For comparison, the table below presents the profile of the impurity standard cyclosporine And pharmaceutical purity according to United States Pharmacopeia (USP), the active ingredient ciclosporin And injection SANDIMMUNand product data cyclosporine a, obtained by the present invention.

Standard injection SANDIMMUNaccording to United States Pharmacopeia product in accordance with example 1 (see table at the end of the description).

From the data one can see that the quality of cyclosporin a obtained by this invention is much superior to the properties of injection SANDIMMUNmoreover, ciclos is in accordance with the present invention is applicable to purification of the crude product in small, and on a large scale.

Except for the fact that the method in accordance with the invention provides for obtaining pure cyclosporin a, he also has some very significant technological advantages. Because using only one kind of technological methods (chromatography), the cleaning method is easily manageable, additionally, it can be reproduced and can be transformed into a continuous process. Moreover, there is one definite advantage consisting in the fact that at three stages of chromatography use only one type of solvent mixture, and therefore regeneration as columns and solvents simplified.

An additional benefit of the method is that during the first stage of chromatography significantly bind impurities remain in the column, which greatly facilitates the regeneration of the downloads column on two subsequent stages.

The invention is explained hereinafter, the examples are not to limit protection, and for the purpose of illustration.

In the comparative example (example 4) shows that when used in a three-step chromatography solvent mixture of dichloromethane-acetone, used so far in known ways is renovage product of the three-step chromatography using a solvent mixture of toluene-acetone.

The quality of the source cyclosporine product,%:

The content of the cyclosporine A - 60,9

The content of cyclosporine IN - 11,2

The content of cyclosporine WITH - 8,3

The content of cyclosporine L - 1,79

The content of cyclosporine U - 1,58

The content of cyclosporine D - 1,25

1-IDegree

Chromatography was performed using connected in series of two chromatographic columns, each column together with the casing had a volume of 8 l, diameter 10 cm, length 100 cm, Each of the two columns contained 3,95 kg of silica gel Kieselgel type Measurement with a grain size of 0.04-0,063 mm At the first stage chromatography both columns contained fresh silica gel. At the next stage, the first chromatography column was separated, and the second column was connected to the column containing fresh silica gel. Further, at every stage of chromatography used only one new column.

Obtaining the crude product

4.1 kg crude product purity 60,9% vol. downloaded in a couple of series-connected columns, pre-dissolving it in 15 l of toluene. Received 19 l of solution, which was applied to the top of the first column through a filter with a feed rate of 2.4 l/h After loading the material was suirable solvent mixture of Aceto the NCI amounted to 39 HP The content of cyclosporine in stemming the flow were analyzed by TLC (thin layer chromatography). In the form of waste collected fractions not containing cyclosporine. As the main faction took 28 l flowing stream after introduction of cyclosporine. The dry matter content of the resulting intermediate compound I to 3.23 kg

Composition,vol.%:

Cyclosporine A - 75

Cyclosporine IN - 10,1

Cyclosporine - 1,6

Cyclosporine L - 1,7

Cyclosporine U - 1.5

Cyclosporine D - 1,3

The output in terms of cyclosporine a was 97%.

2-IDegree

The separation was carried out in a fitted casing 8 l column, having a length of 1 m Column contained silica gel Kieselgel 60 type of Measurement (0,015-0,040 mm). The mass of cushioning material was 3,95 kg Solution of intermediate compound 1 with a volume of about 3 liters, containing 370 g of dry matter obtained in the first stage, was loaded into the column with a feed rate of 2.4 l/h, then washed in 1 l of toluene.

After loading, the column material was suirable 10 l of a mixture of acetone and toluene at a volume ratio of components in a mixture of 15:85 and then 20 l of a solution of acetone and toluene at a volume ratio of components in a mixture of 25:75. The flow rate and TLC (thin layer chromatography). 1-11 l fraction was a waste, 12-19 l fractions were considered as critical of them and took samples. By means of HPLC (high performance liquid chromatography) analyzed the profile contained impurities and their manipulated as a preliminary fractions or mixed with the main factions. This way you can get the pre-fraction containing 80 g of dry matter, which is then boiled away before the formation of the dry residue. 20-25 l fractions were combined as the main fraction. 26-31 l fractions were considered as critical fractions and, after analysis, they were combined with the main fraction or manipulated by them as the subsequent fraction.

Obtained in this way the main fraction boiled away to the dry residue in film evaporator equipped with a vibrating mixer. Received 234 g of intermediate compound P with yields of 80%, with the following quality,vol.%:

Cyclosporine AND 95

Cyclosporine U 1,2

Cyclosporine L 0,7

Cyclosporine < 0,1

Cyclosporine D 0,5

Cyclosporine 0.1

3-IDegree

Chromatography was carried out in columns with the same design and the same geometrical dimensions, which were described for the first and Eastmore, containing 220 g of dry matter, and filed on the top of the column at a rate of 2.4 l/h, then washed in 1 l of toluene.

The column was suirable a mixture of 20 l of acetone and toluene at a volume ratio of components in a mixture of 15:85, and then the cyclosporine was suirable a mixture of 20 l of acetone and toluene in a ratio of components in a mixture of 25:75. The flow rate of eluent to obtain 31 l fraction was 2.4 l/h, then starting with 32 l of fraction - 5 l/h 1-18 l fraction was a waste, 19-23 l fraction was a preliminary faction and they were viewed as critical fraction I. For analysis of dry matter content and impurity profile high-performance liquid chromatography of them took samples.

After analysis they were manipulated as a preliminary fractions or mixed with the main factions. 29-38 l fractions were combined as the main fraction, 39-41 l fractions were collected in 1 l portions and considered as a critical fraction II. After analysis, they were combined with the main fraction or manipulated as subsequent fractions II. After analysis of their mingled with the main fraction or manipulated as the subsequent fraction. In collecting fractions after evaporation to a dry residue, you can obtain 70 g of the lip 157 g of pure cyclosporine with a yield of 75%. The quality of the product was as follows,%:

Cyclosporine AND 99,6

Cyclosporine L < 0,05

Cyclosporine U < 0,05

Cyclosporine D < 0,05

Cyclosporine < 0.02

Cyclosporine < 0.02

Example 2

Purification of the crude cyclosporine product four chromatography column with a porous layer of silica gel using solvent mixtures of toluene-acetone or toluene-ethyl acetate.

Raw cyclosporiasis product was purified three-step chromatography described in example 1. The obtained three-step chromatography preliminary faction cleaned fourth step in a fixed bed solvent mixture of toluene-ethyl acetate.

4-Idegree

The design and dimensions of the chromatographic column were the same as those described in example 1. The column described in example 1, contained silica gel type Kieselgel 60 Merck (0,015-0.40 mm).

The column was treated concentrate with a feed rate of 2.4 l/h, obtained from preliminary faction in the amount of 260 g of the material dissolved in 2.5 l of toluene.

The content of cyclosporine And 80,6% vol.

The content of cyclosporine D 4,2% vol.

Processed p is omponents in a mixture of 17: 83 a flow rate of 2.4 l/h, then, elution was continued with a mixture of 40 liters of ethyl acetate and toluene at a volume ratio of components in a mixture of 28:72. 1-19 l faction while collecting fractions represented a major shift. After analysis by high-performance liquid chromatography 20-25 l fraction represented a departure or mixed with the main fraction. 26-35 l fraction was collected as the main fraction. After sampling and analysis HPLC 36-42 l fraction or combined with the main fraction or manipulated as waste. Collected 23-42 l main fraction boiled away to the dry residue. In this way received 195 g of pure cyclosporin a, containing 99,6% vol. active ingredient with a yield of 75% with the following quality,vol.%:

Cyclosporine AND 99,6

Cyclosporine D < 0,05

Cyclosporine U

Cyclosporine L

Example 3

Purification of the crude cyclosporine product two-step chromatography column with a porous layer of silica gel using a solvent mixture of toluene-acetone

Raw cyclosporiasis product was purified according to the same manner as that described in the first stage of example 1, receiving the intermediate cyclosporine connection I with the same quality. Further, the method implemented as OO 370 g of dried material, with a feed rate of 2.4 l/h, then treated sample was washed in 1 l of toluene.

After loading the column was suirable 10 l of a mixture of acetone and toluene at a volume ratio of components in a mixture of 15:85, and then elution was continued with 20 l of a mixture of acetone and toluene at a volume ratio of components in a mixture of 25: 75. The flow rate of solvent to obtain 17 l was 2.4 l/h and from 18 l - 5 l/h

In accordance with thin-layer chromatography and high performance liquid chromatography 1-11 l fraction was a waste, 12-20 l fraction was a preliminary faction, 21-24 l fractions were considered as the main fraction and fractions from 25 l to the end of manipulated as subsequent fractions. Washed with acetone fraction was a waste.

After fractionation collected basic fractions were combined and boiled away until the solids receiving 114 g of cyclosporine And with the release of 41% and with the same high quality product that is described in example 1.

Example 4

Comparative example purification of the crude product of the three-step chromatography using a solvent mixture of dichloromethane-acetone

The quality of the original raw cyclosporine cont leporina 11.2

The content of cyclosporine FROM 8.3

The content of cyclosporine L 1,79

The content of cyclosporine U 1,58

The content of cyclosporine D 1,25

1-IDegree

Chromatographic equipment and packing of the column were the same as those described in example 1. One pair connected in series columns subjected to processing 4.1 kg crude product purity 60,9% in 15 l of a solution of dichloromethane.

After loading the sample the column was suirable dichloromethane with a flow rate of 2.4 l/h to collect 35 l flowing stream.

1-10 l fraction was a waste, whereas 11-35 l fractions were considered as the main fraction.

The dry matter content of the intermediate compound 1 obtained in this way was 2.9 kg, the content of the active ingredient - 75 vol.%.

2-IDegree

Chromatographic equipment and packaging the column were the same as those described in example 1. The column was subjected to processing on top of 3 l of intermediate compound I containing 350 g of dry matter, with a feed rate of 2.4 l/h Elution was carried out 10 l of a mixture of acetone and dichloromethane at a volume ratio of components in a mixture of 1:9, then the elution was continued with 25 l of a mixture of acetone and is accordance with thin-layer chromatography volume prior fraction was 13 l, the main fraction - 22 l and the volume of subsequent fraction - 11L. 22 l the main fraction boiled away to the dry residue and consequently received 220 g of the intermediate product II, having a purity of 91%.

3-IDegree

Chromatographic equipment and packaging the column were the same as in example 1. When filing in column dichloromethane concentrate with a rate of 2.4 l/h gasket loaded 220 g of intermediate compounds P. Elution was carried out 20 l of a mixture of acetone and dichloromethane at a volume ratio of components in a mixture of 1:9, then the elution was continued with 30 l of a mixture of acetone and dichloromethane at a volume ratio of components in a mixture of 2:8 and graduated from acetone with the speed of its submission 2.4 l/h

The first 26 l fraction was considered as the prior fraction, then collected 26 l of the main fraction and finally 11 l subsequent fractions. 26 l of the main fraction boiled away to the dry residue and consequently received 140 g of the intermediate product III, with the following quality,vol.%:

Cyclosporine AND 98,6

Cyclosporine U 0,6

Cyclosporine D 0,3

Cyclosporine L 0,2

Cyclosporine 0.1

Cyclosporine From 0.1 to

1. The method of receiving cyclosporine And high purity by cleaning surgicutt solvent mixture, including toluene, characterized in that use multi-step chromatography at high load columns from 10 to 52%, elution carried out with a mixture of toluene with acetone in an amount of from 10 to about 30. % or toluene with ethyl acetate in an amount of from 10 to about 35. %.

2. The method according to p. 1, characterized in that sequentially perform 2-4 chromatographic steps.

3. The method according to p. 1, characterized in that sequentially perform 3 chromatographic steps.

4. The method according to p. 1, characterized in that the solvent mixture using toluene-acetone.

5. The method according to p. 4, characterized in that the use of toluene containing not more than 30% acetone.

6. The method according to p. 1, characterized in that use solvent mixture of toluene and ethyl acetate.

7. The method according to p. 6, characterized in that the use of toluene containing not more than about 35. % ethyl acetate.

8. The method according to p. 1, characterized in that the gradient elution is applied in the case of at least one chromatographic step.

9. Cyclosporine And high purity, where the content of cyclosporine L, U cyclosporine and cyclosporine D is less than 0.05. % and the content of Cyclos is from the crude product, containing cyclosporiasis complex by chromatography on silica gel using as eluent a mixture of solvents, including toluene, characterized in that use multi-step chromatography at high load columns from 10 to 52%, elution carried out with a mixture of toluene with acetone in an amount of from 10 to about 30. % or toluene with ethyl acetate in an amount of from 10 to about 35. %.

 

Same patents:

Cyclosporine // 2085589

The invention relates to compounds of the formula I

R1-Q1-X-Q2-R2(I)

in which Q1, Q2in each case, independently of one another are either absent or represent-NH-(CH2)n-CO-,

R1, R2in each case, independently of each other either absent or represent a cyclo-(Arg-Gly-Asp-Z), where Z is linked to the side chain of Q1or Q2or, if Q1and/or Q2missing (et), with X and where at least one of the radicals R1or R2must always be present,

X represents a-CO-R18-CO-, and if R1-Q1or R2-Q2- no, there is an R10, R13, R16, Het-CO -, or the residue of a fluorescent dye that is chemically linked through CONH-, -COO-, -NH-C(=S)-NH- -NH-C(=O)-NH-, - SO2NH -, or-NHCO-bonds

Z in each case independently represents an amino acid residue or di-, tri - or tetrapeptides balance, where amino acids independently selected from the group comprising Ala, Asn, Asp, Arg, Cys, Gln, Glu, Gly, His, Ile, Leu, Lys, Met, Phe, Pro, Ser, Thr, Trp, Tyr, Val or M

where these amino acids can be derived and amino acid residues connected to one another like xilinix groups, and where M is always present,

M represents NH(R8)-CH(R3)COOH,

R3- R5-R4, -R6-R4or-R7-R4< / BR>
R4is a HE, NH2, SH or COOH,

R5- alkylene containing 1-6 carbon atoms,

R6- alkaliphiles containing 7 to 14 carbon atoms,

R7- alkylenediamine containing 8-15 carbon atoms,

R8-H, a or alkylether containing 7-12 carbon atoms,

A is alkyl containing 1-6 carbon atoms,

R10- alkanoyl containing 1-18 carbon atoms, which is unsubstituted or contains one Deputy from among COOH, COOA, SR11or NR12R12,

R11- H or trityl, pyridyl-2-thio - or allylthiourea containing 1-6 carbon atoms,

R12, R12'each, independently of one another, represent H, alkyl containing 1-8 carbon atoms or a protective group of amino group,

R13- aroyl, which contains 7 to 11 carbon atoms and is unsubstituted or substituted and contains one or two substituent selected from the group comprising alkyl containing 1-6 carbon atoms, alkoxygroup containing 1-4 carbon atoms, alkanoyl containing 1-8 tomsche independently of one another H or A,

R16is arkanoid, which contains 7-19 carbon atoms and which is not substituted or substituted in the aryl fragment of one, two or three deputies, including Hal, alkoxygroup containing 1-6 carbon atoms, or HE, and in which the aryl fragment may also represent a group:

< / BR>
E - CH2or,

D is carbonyl or [C(R17R17')]m,

R17R17'each independently represents H or A,

R18is absent or is an R19, R20, R19-R20-R19or phenylene, which is not substituted or substituted and contains one or two substituent R5the length of the chain which is in each case independently of each other,

R19is alkylene containing 1-8 carbon atoms, where 1 or 2 methylene groups can be replaced with S, -CH=CH - or,

R20- cycloalkyl containing 3-7 carbon atoms,

Hal is F, Cl, Br or I,

Het is a monocyclic or bicyclic saturated, unsaturated or aromatic heterocycle which contains from 1 to 4 atoms of N, O and/or S, attached cherepy, including Hal, A, R3, NR4R4', CN, NO2and/or carbonyl oxygen,

n- 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 and

m is 1 or 2,

where, assuming that residues are residues of optically active amino acids and derivatives of amino acids, contain both D and L forms, and their salts

Cyclopeptide // 2171260
The invention relates to cyclopeptides and to their therapeutic use as inhibitors of the expression of adhesion molecules

The invention relates to new methods of chromatography, designed for the purification of crude extracts containing cyclosporine, for use in the pharmaceutical industry

The invention relates to cyclopentapeptide formula (I):

cyclo (A-B-C-E-F-(D)-Ala)

in which A, B, C, E and F, independently of one another, may be the same or different and represent a residue of a natural amino acid except cysteine (Cys) and tryptophan (Trp), and accordingly can be an alanine (Ala), arginine (Arg), asparagine (Asn), aspartic acid (Asp), glutamine (Gln), glutamic acid (Glu), glycine (Gly), histidine (His), isoleucine (Ile), leucine (Leu), lysine (Lys), methionine (Met), phenylalanine (Phe), Proline (Pro), serine (Ser), tryptophan (Thr), tyrosine (Tyr) or valine (Val), as well as their physiologically compatible salts

The invention relates to somatostatinomas peptides, method for their production and pharmaceutical preparations containing them

The invention relates to new cyclopeptides formula cyclo-(pad-nGly-nAsp-nD-nE), where n, D, and E have the meanings indicated in the claims, and to pharmaceutical compositions based on them having inhibitory activity againstv3and/orv5-integrins and to a method for producing a pharmaceutical composition having inhibitory activity

The invention relates to method modification, facilitating access to the treated (poly)peptides and to a method of cleaning using this method modifications

The invention relates to method modification, facilitating access to the treated (poly)peptides and to a method of cleaning using this method modifications

The invention relates to method modification, facilitating access to the treated (poly)peptides and to a method of cleaning using this method modifications

The invention relates to medicine, in particular to drugs, protein-containing, and methods for their preparation, namely natural cationic proteins from the tissues of the thymus gland of calves - histone

The invention relates to medicine, in particular to drugs, protein-containing, and methods for their preparation, namely natural cationic proteins from the tissues of the thymus gland of calves - histone

The invention relates to novel conjugates of affinity ligand - matrix containing the ligand associated with the matrix carrier, optionally via a spacer elements located between the matrix and ligand, and new conjugates affinity ligand - matrix, to their preparation and use for the purification of protein materials, such as, for example, immunoglobulins, insulin, factor VII, or human growth factor, or its analogs, derivatives and fragments, and their predecessors
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