HRP20030654A2 - Method for purifying 20(s)-camptothecin - Google Patents

Abstract

The invention relates to a method for purifying 20(S) camptothecin by transforming the lactone ring of the 20(S) camptothecin into a carboxylate salt with the help of an aqueous base, hydrogenating the mixture that is obtained in the presence of a transition metal catalyst, acidifying the aqueous phase to form camptothecin crystals, adding a polar aprotic solvent, optionally heating and cooling the mixture and separating the camptothecin crystals.

Description

The invention relates to a process for cleaning (S)-camptothecin, which is contaminated with a vinyl-camptothecin derivative.

Background of the invention

20(S)-camptothecin, (20(S)-CPT), is a naturally occurring alkaloid of formula (I)

[image]

in which R represents ethyl.

20(S)-CPT and its derivatives as inhibitors of topoisomerase 1 have tumor inhibitory activity (eg Giovanelle, B.C. et al., Cancer Research, 51: 302-3055, 1991, European patent applications EP 0 074 256, EP 0 088 642, US Patents US 4,473,692, US 4,545,880, US 4,604,463 and International Patent Application WO 92/05785).

20 (S)-CPT can be obtained as a crude product, inter alia, from the Chinese tree Camptotheca acuminata (Nyssaceae) (Wall M. et al., J. Am. Chem. Soc. 88: 3888-3890, 1966) or from of the Indian tree Nothapodytes foetida (nimmoniana) (earlier name: Mappie foetida Miers) (Govindachari, T.R. et al., Phytocheraistry 11: 3529-3531, 1972).

These crude products, especially those obtained from Nothapodytes foetida, have 20 (S) CPT contaminated with a CPT-derivative of formula (I) in which R 1 represents vinyl, (20-vinyl-CPT).

A common way is to purify the crude product by expensive chromatographic procedures or by converting camptothecin into the aqueous phase and removing impurities by extraction with water-insoluble solvents (eg WO 94/19353). In any case, contamination with 20-vinyl-CPT cannot be effectively removed by this process.

Therefore, the task of the proposed invention is to provide a process that enables the purification of the 20(S)-CPT starting product without applying expensive chromatographic procedures.

Description of the invention

Surprisingly, it was found that 20(S)-CPT can be almost completely freed from contamination with 20-vinyl-CPT, where the starting material is first treated with an aqueous base, then hydrated, and finally acidified and the product isolated.

Therefore, the subject of the invention is a process for cleaning 20(S)-camptothecin, which includes the following steps:

(a) the starting material, containing 20(S)-camptothecin, is mixed with an aqueous base, whereby the lactone ring of 20(S)-camptothecin is converted into a carboxylate salt;

(b) the resulting mixture is hydrogenated in the presence of a transition metal catalyst;

(c) the aqueous phase is acidified, whereby camptothecin crystals are formed;

(d) adding at least one polar aprotic solvent; and

(e) Camptothecin crystals are separated.

A further object of the invention is a process for the production of 20 (S)-camptothecin of formula (I), in which R1 represents ethyl, from 20-vinyl-camptothecin of formula (I), in which R represents vinyl, which includes the following steps:

(a) the starting material containing 20-vinyl-camptothecin is mixed with an aqueous base, whereby the compound of formula (II) is obtained,

[image]

where

R1 represents vinyl; and

Met is a metal;

(b) the resulting mixture is hydrogenated in the presence of a transition metal catalyst;

(c) the aqueous phase is acidified, whereby camptothecin crystals are formed;

(d) adding at least one polar aprotic solvent; and

(e) Camptothecin crystals are separated.

Description of the invention in detail

The term "camptothecin-containing starting material", as used herein above and below, refers to contaminated material containing 20(S)-CPT, crude camptothecin, plant extract containing camptothecin, synthetic camptothecin, camptothecin derivatives such as for example those described in international patent application WO 92/05785, or camptothecin containing reaction products.

The starting material is primarily a natural raw product, which is obtained specifically from Nothapodites foetida. As a rule, it is a mixture of a compound of formula (I), in which R is ethyl, and a compound of formula (I), in which R is vinyl. As a rule, it contains 0.9 to 1.5 wt. %, preferably 1.0 to 1.4 wt. % vinyl compound. In addition, the starting material may also contain other camptothecin derivatives such as, for example, 9-methoxy-CPT, 10-methoxy-CPT, 11-methoxy-CPT, 10-hydroxy-CPT and 11-hydroxy-CPT. As a rule, the starting material contains up to l mass. % of one or more of these additional CPT derivatives, especially 0.2 to 0.8 wt. % 9-methoxy-CPT.

The term "aqueous base", as used hereinabove and below in connection with step (a) of the cleaning process according to the invention, refers to an aqueous agent which, preferably in pure water, has sufficient hydroxide ions to completely convert the lactone group contained in the camptothecin derivative into starting material, into the corresponding hydroxycarboxylate. Metal hydroxides, especially hydroxides of alkali metals or alkaline earth metals such as lithium hydroxide, sodium hydroxide, potassium hydroxide or calcium hydroxide, are advantageous. The greatest preference is given to sodium hydroxide.

Metal hydroxide is preferably used in the form of a diluted aqueous solution, preferably in the form of a 1 to 25%, especially a 3 to 10% aqueous solution. As a rule, enough metal hydroxide is used that the camptothecin derivative completely passes into the solution. Preferably 1 to 20 moles are used, especially preferably 5 to 15 moles, especially 7.5 to 12.5 moles of metal hydroxide per 1 mole of starting material.

A catalyst based on a transition metal, preferably a catalyst based on heterogeneous transition metals, especially platinum, platinum oxide, nickel, palladium or rhodium on a support such as activated carbon or aluminum oxide, is added to the mixture obtained in step (b). Preference is given to palladium on activated carbon with a content of 1 to 15 wt. %, preferably 2 to 10 wt. %, especially approximately 5 wt. % palladium.

The amount of transition metal-based catalyst is chosen so that complete hydrogenation of the vinyl CPT derivative is enabled. Preferably, 0.01 to 0.50 parts by mass, especially 0.02 to 0.10 parts by mass of transition metal-based catalyst are used in relation to 1 part by mass of the starting material.

Gaseous hydrogen is passed through the thus obtained mixture preferably at a temperature of -20°C to 100°C, especially from 10°C to 40°C, preferably at room temperature.

Hydrogen pressure is not crucial, so hydration is preferably carried out under normal pressure or under slightly elevated pressure, especially under 0.9 to 5.0 bar, preferably under approx. 1 bar.

Under the mentioned conditions, hydration is usually completed in 1 to 20 hours, preferably in 4 to 15 hours, especially in 6 to 10 hours.

After complete hydrogenation, the catalyst based on the transition metal is removed, preferably by filtration, and the resulting reaction mixture is acidified in step (c). It can be acidified with inorganic or organic acid. Preference is given to the use of mineral acids, such as HCl, HBr, HJ, HNO3, H3PO4, H2SO4 or aliphatic carboxylic acids such as acetic acid and trifluoroacetic acid or a mixture of these acids, especially concentrated hydrochloric acid. Adjust the pH value to 3.0 to 6.0, preferably 3.5 to 5.0, especially from approx. 4.0 to 4.5. As a rule, the conversion with acid takes place at a temperature of 0°C to 100°C, preferably from 30°C to 80°C, especially from 50°C to 60°C.

In a particularly advantageous embodiment, it is acidified with 2 to 20 parts by mass, preferably with 4 to 9 parts by mass, especially 6 to 8 parts by mass of concentrated hydrochloric acid in relation to 1 part by mass of the starting material.

Under the mentioned conditions, lactonization in CPT is usually completed in 10 to 180 minutes, preferably 15 to 60 hours, especially in approx. 30 minutes.

As a rule, the reaction mixture obtained by acidification is a pure suspension. To improve crystallization in step (d), the mixture is mixed with one or more polar aprotic solvents. As such solvents, sulfoxides, such as dimethyl sulfoxide (DMSO) or amides and urea derivatives of the formula

[image]

where

R2 represents hydrogen or a C1-C4 alkyl group;

R3 and R4 in each case independently of each other represent a C1-C4 alkyl group; or

R2 and R3 together form the group -(CH2)m- or -NR5-(CH2)n-, in which

R5 represents a C1-C4 alkyl group;

m is 3 or 4; and

n is 2 or 3,

especially solvents selected from the group consisting of N,N-dimethylformamide (DMF), N,N-dimethylacetamide (DMA), N-methylpyrrolidone (NMP), N,N-dimethylethyleneurea (DMEU) and N,N-dimethylpropylene-urea (DMPU) or mixtures of such solvents, preferably DMF.

As a rule, 10 to 100 parts by mass, preferably 20 to 80 parts by mass, especially 30 to 50 parts by mass of polar aprotic solvent are used in relation to 1 part by mass of the starting material used.

Treatment with a polar aprotic solvent can be carried out at any temperature. The reaction mixture is preferably stirred at a temperature of 30°C to 120°C, especially from 80 to 100°C and then slowly cooled to room temperature.

The CPT crystals obtained in this way can be easily separated from the liquid phase in step (e), whereby the separation is primarily performed by decantation, centrifugation, whisking, pressing or filtration, especially filtration.

As a rule, the CPT crystals thus obtained are washed with alcohol, preferably methanol, ethanol or isopropanol, especially methanol, and dried.

The advantage of the process according to the invention is the high utilization of space and time, as well as the high utilization and purity of the obtained 20(S)-camptothecin, which without chromatographic purification is obtained mainly without impurities containing the vinyl group.

The following examples serve to illustrate the implementation of the process for purifying camptothecin according to the invention. They are only possible examples of the implementation of the process and should not be considered that the invention is limited to their content.

Example 1

10.45 g of the crude product obtained from Nothapodites foetida., which contains camptothecin and further contains 1.33% 20-vinyl-CPT and 0.47% 9-methoxy-CPT, is placed in 260 ml of 2 normal sodium hydroxide solution and mixed with 0.6 g of palladium on activated carbon (5%). Hydrogen is passed through the mixture for 8 hours at room temperature and under a pressure of 1 bar.

The reaction mixture is then filtered and mixed with 80 ml of concentrated hydrochloric acid at 50-60°C and adjusted to a pH value of 4.0 to 4.5.

The resulting suspension was mixed with 400 ml of DMF and stirred for 2.5 hours at 90-100°C. The resulting suspension is slowly cooled to room temperature and filtered. The obtained CPT crystals are washed with 100 ml of methanol and dried at 55°C in a vacuum.

9.85 g (94.2% of the charge) of 20(S)-camptothecin containing less than 0.05% 20-vinyl-CPT and 0.11% 9-methoxy CPT are obtained.

Example 2

10.45 g of the crude product obtained from Nothapodites foetida, containing camptothecin and further containing 1.33% 20-vinyl-CPT and 0.47% 9-methoxy-CPT was placed in 260 ml of 2 normal sodium hydroxide solution and mixed with 0.6 g of palladium on activated carbon (5% vol.). Hydrogen is passed through the mixture for 8 hours at room temperature and under a pressure of 1 bar.

The reaction mixture is then filtered and mixed with 300 ml of 10% sulfuric acid at 50-60°C and adjusted to a pH value of 4.0 to 4.5.

The resulting suspension was mixed with 500 ml of DMF and stirred for 2.5 hours at 90-100°C. The resulting suspension is slowly cooled to room temperature and filtered. The obtained CPT crystals are washed with 100 ml of methanol and dried at 55°C in a vacuum.

9.67 g (92.6% of the added amount) of 20 (S)-camptothecin containing 0.09% of 9-methoxy CPT are obtained, whereby the content of 20-vinyl-CPT is below the detection limit.

Claims (10)

1. A process for cleaning 20(S)-camptothecin, characterized in that it includes the following steps: (a) the starting material, containing 20(S)-camptothecin, is mixed with an aqueous base, whereby the lactone ring of 20(S)-camptothecin is converted into a carboxylate salt; (b) the resulting mixture is hydrogenated in the presence of a transition metal catalyst; (c) the aqueous phase is acidified, whereby camptothecin crystals are formed; (d) adding at least one polar aprotic solvent; and (e) Camptothecin crystals are separated. 2. The method for cleaning 20(S)-camptothecin according to claim 1, characterized in that the starting material, which contains 20(S)-camptothecin, is a natural plant product. 3. Process for cleaning 20(S)-camptothecin according to claim 1 or 2, characterized in that the starting material containing 20(S)-camptothecin consists mainly of a mixture of compounds of formula (I), [image] wherein R 1 represents ethyl or vinyl. 4. A process for the purification of 20(S)-camptothecin according to any preceding claim, characterized in that the base in step (a) is sodium hydroxide. 5. A process for purifying 20(S)-camptothecin according to any preceding claim, characterized in that the mixture obtained in step (a) is hydrogenated in the presence of a palladium catalyst at a temperature from 0°C to 100°C and under a pressure of 0, 5 bar to 5.0 bar. 6. A process for purifying 20(S)-camptothecin according to any preceding claim, characterized in that the aqueous phase obtained in step (b) is treated with an acid selected from the group consisting of HCl, HBr, HJ, HNO3, H3PO4, H2SO4, acetic acid and trifluoroacetic acid or with a mixture of these acids at a temperature of 30°C to 80°C. 7. A process for purifying 20(S)-camptothecin according to any preceding claim, characterized in that the aqueous phase obtained in step (c) is treated with one or more polar aprotic solvents of the formula [image] where R2 represents hydrogen or a C1-C4 alkyl group; R3 and R4 in each case independently of each other represent a C1-C4 alkyl group; or R2 and R3 together form the group -(CH2)m- or -NR5-(CH2)n-; where R5 represents a C1-C4 alkyl group; m is 3 or 4; and n is 2 or 3, at a temperature of 30°C to 120°C. 8. The method for cleaning 20(S)-camptothecin according to claim 7, characterized in that the polar aprotic solvent is selected from the group consisting of N,N-dimethylformamide (DMF), N,N-dimethylacetamide (DMA), N-methylpyrrolidone ( NMP), N,N-dimethylethyleneurea (DMEU) and N,N-dimethylpropyleneurea (DMPU) or a mixture of these solvents. 9. A process for purifying 20(S)-camptothecin according to any preceding claim, characterized in that the crystals of 20(S)-camptothecin in step (d) are separated by filtration. 10. A process for the production of 20(S)-camptothecin of formula (I), in which R represents ethyl, from 20-vinyl-camptothecin of formula (I), in which R1 represents vinyl, characterized by the fact that it includes the following stages: (a) the starting material containing 20-vinyl-camptothecin is mixed with an aqueous base, whereby the compound of formula (II) is obtained, [image] where R1 represents vinyl; and Met is a metal; (b) the resulting mixture is hydrogenated in the presence of a transition metal catalyst; (c) the aqueous phase is acidified, whereby camptothecin crystals are formed; (d) adding at least one polar aprotic solvent; and (e) Camptothecin crystals are separated.