Organic compounds -- part of the class 532-570 series – Organic compounds – Heterocyclic carbon compounds containing a hetero ring...
Reexamination Certificate
1999-08-20
2001-03-06
Trinh, Ba K. (Department: 1625)
Organic compounds -- part of the class 532-570 series
Organic compounds
Heterocyclic carbon compounds containing a hetero ring...
C549S511000
Reexamination Certificate
active
06197980
ABSTRACT:
The present invention relates to a process for the purification of taxoids by centrifugal partition chromatography.
More especially, the present invention relates to a process for the purification of Taxotere and of 10-deacetylbaccatin III.
10-Deacetylbaccatin III extracted from yew leaves is useful in the preparation of Taxotere under the conditions described, for example, in European Patents EP 0,253,738 or EP 0,336,841 or in International PCT WO 92/09,589.
10-Deacetylbaccatin III obtained by extraction of yew leaves contains, depending on the species from which it originates, impurities which are chiefly products belonging to the taxoid family, such as, for example, 2&agr;-benzoyloxy-4-acetoxy-5&bgr;,20-epoxy-1&bgr;,7&bgr;,10&bgr;,13&agr;,19-pentahydroxy-9-oxo-11-taxene, 2&agr;-benzoyloxy-4-acetoxy-5&bgr;,20-epoxy-1&bgr;,7&agr;,10&bgr;,13&agr;-tetrahydroxy-11-taxene or taxins.
Taxotere which is obtained by partial synthesis from 10-deacetylbaccatin III contains as chief impurities the esterification products of the impurities contained in 10-deacetylbaccatin III, as well as other impurities originating, for example, from the epimerization of the carbon at position 2′ of the side chain.
10-Deacetylbaccatin III and Taxotere itself may generally be purified by liquid chromatographic methods on a column, and more especially by high performance liquid chromatography (HPLC) on a silica column. However, while these methods are especially well suited to the purification of a few tens of grams, their industrial extrapolation comes up against constraints, such as the quantity of solvents consumed and the handling and destruction of the (silica) support contaminated by toxic residues, which become paramount.
In the case of taxoids which constitute products whose toxicity is often very high and whose lability is great, it is especially important to have the possibility of using purification methods which have good productivity, which do not require a solid support which is expensive to buy, use and destroy, which require the use of small quantities of solvents and which can be readily automated to permit continuous production.
It has now been found, and this forms the subject of the present invention, that the purification of taxoids, and more especially the purification of Taxotere and of 10-deacetylbaccatin III, may be achieved by carrying out centrifugal partition chromatography (CPC) or high speed countercurrent chromatography, the principle of which is described, for example, by A. Foucault and P. Rosset, Analusis, 16 (3), 157-167 (1988).
Centrifugal partition chromatography makes it possible to effect partition of the constituents of the mixture to be separated between two immiscible liquid phases, by establishing successive equilibria produced in a logical manner and automatically.
This method is characterized by a very efficient partition mechanism, a strong retention of the stationary phase and a high speed of the mobile phase, leading to excellent separations in a few hours.
Centrifugal partition chromatography requires the use of two or more solvents supplying 2 partially miscible phases. Although there is an unlimited number of solvents displaying this feature, it is especially important industrially to use non-chlorinated and nontoxic industrial solvents. Among solvents which may be used industrially incurring lower risks, alcohols such as methanol, ethers such as methyl t-butyl ether, esters such as ethyl acetate, ketones such as methyl isobutyl ketone and aliphatic hydrocarbons such as heptane or isooctane may be mentioned.
To carry out an efficient purification, it is necessary to choose a mixture of solvents whose rapid and complete separation yields two phases for which the partition coefficient is between 0.1 and 10, and preferably between 0.5 and 5, and still more especially in the vicinity of 1.
A product for which the partition coefficient (K) is, for example, equal to 10 is 10 times more soluble in the upper phase than in the lower phase. Consequently, it will be eluted very rapidly if the upper phase is mobile, and after a very long time, with a poor resolution, if the lower phase is the one which is mobile. In contrast, when K is in the region of 1, the solubility will be practically identical in both phases, and both modes of elution may be used, and where appropriate combined, to obtain maximum resolution.
In the particular case of 10-deacetylbaccatin III, mixtures of an aliphatic hydrocarbon, an ester, an alcohol and water such as a heptane/ethyl acetate/methanol/water (1:2:1:2 by volume) mixture, and more especially mixtures of aliphatic ketones and water such as a methyl isobutyl ketone/acetone/water (2:3:2 by volume) mixture, are most especially suitable.
Furthermore, it is necessary for the retention in the stationary phase, which is the fraction of the total volume of the apparatus occupied by the stationary phase at equilibrium, the flow rate of the mobile phase being fixed, to be as high as possible and equal to at least 50%.
Furthermore, in order to improve the productivity, it is especially advantageous to be able to work at a high flow rate, which is limited by the maximum pressure imposed by the rotor.
In the particular case of Taxotere, it is advantageous to use a mixture of an aliphatic hydrocarbon, an ester, an alcohol and water, such as a heptane/ethyl acetate/methanol/water (2:4:3:2 by volume) system, which has a retention of 80% and a partition coefficient in the region of 0.5, thus giving a narrower peak which is eluted rapidly, thereby leading to a very marked improvement in productivity.
The centrifugal partition chromatography may be carried out in any commercially available apparatus suitable for this purpose, such as those which have been developed by Yoichiro Ito [CRC Crit. Rev. Anal. Chem., 17, 65 (1986)] or marketed by SANKI Engineering Limited in Kyoto (Japan).
The present invention also relates to purified Taxotere and purified 10-deacetylbaccatin III when they are obtained by carrying out the process according to the invention.
The examples which follow show how the invention may be put into practice.
REFERENCES:
patent: 4814470 (1989-03-01), Colin et al.
patent: 4924012 (1990-05-01), Colin et al.
patent: 5229526 (1993-07-01), Holton
patent: 0 253 738 (1988-01-01), None
patent: 0 336 841 (1989-10-01), None
patent: WO 92/09589 (1992-06-01), None
Voegelein et al.J. Med. Chem., 34(3), pp. 992-998, 1991.
A. Foucault et al., “La chromatographie a contre-courant,”Analusis, vol. 16, No. 3, pp. 157-167, 1988.
Tian-You Zhang et al., “Rapid Separation of Flavonoids by Analytical High-Speed Counter-Current Chromatography,”Journal of Chromatography, 445, pp. 199-206, 1988 (Abstact).
Yoichiro Ito et al., “Improved Cross-Axis Synchronous Flow-Through Coil Planet Centrifuge for Performing Counter-Current Chromatography,”Journal of Chromatography, 464, pp. 305-416, 1988 (Abstract).
Molina Bhatnagar et al., “Improved Cross-Axis Synthronous Flow-Through Coil Planet Centrifuge for Performing Counter-Current Chromatography,”Journal of Chromatography, 463, pp. 317-328, 1989 (Abstract).
Yoichiro Ito et al., “Horizontal Flow-Through Coil Planet Centifuge Equipped with a Set of Multilayer Coils Around the Column Holder,”Journal of Chromatography, 457, pp. 393-397, 1988 (Abstract).
Takashi Yoshida et al., “Chromatography of Tannins,”Journal of Chromatography, 467, pp. 139-147, 1989 (Abstract).
Hisao Oka et al., “Improved Method for Continuous UV Monitoring in High-Speed Counter-Current Chromatography,”Journal of Chromatography, 475, pp. 229-235, 1989 (Abstract).
Nabil Tayar et al., “Use of Centrifugal Partition Chromatography for Assessing Partition Coefficients in Various Solvent Systems,”Journal of Chromatography, 469, pp. 91-99, 1989 (Abstract).
Hisao Oka et al., “Multilayer Coil Planet Centrifuge for Analytical High-Speed Counter-Current Chromatography,”Journal of Chromatography, 479, pp. 53-60, 1989 (Abstract).
Yoichiro Ito et al., “Improved High-Speed Counter-Current Chromatograph with Three Multilayer Coils Connected in Series,”
Durand Andre
Gerbaud Alain
Margraff Rodolphe
Finnegan Henderson Farabow Garrett & Dunner L.L.P.
Rhone-Poulenc Rorer S.A.
Trinh Ba K.
LandOfFree
4-Acetoxy-2&agr;-benzoyloxy-5&bgr;,20-epoxy-1&bgr;,7&bgr;,10... does not yet have a rating. At this time, there are no reviews or comments for this patent.
If you have personal experience with 4-Acetoxy-2&agr;-benzoyloxy-5&bgr;,20-epoxy-1&bgr;,7&bgr;,10..., we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and 4-Acetoxy-2&agr;-benzoyloxy-5&bgr;,20-epoxy-1&bgr;,7&bgr;,10... will most certainly appreciate the feedback.
Profile ID: LFUS-PAI-O-2461561