Title of Invention

"SINGLE POT PROCESS FOR THE PREPATATION OF ARTEETHER FROM ARTEMISININ"

Abstract The present invention provides a method for the preparation of arteether from artemisinin in one pot in just about 4 hours comprising reduction of artemisinin into dihydroartemisinin by less quantity of sodium borohydride in ethanol at room temperature in the presence of a novel polyhydroxy catalyst, acylation of dihydroartemisinin in the presence of an acid catalyst, extraction of arteether from an aqueous reaction mixture using 1% ethyl acetate in n-hexane followed by workup and purification of the impure arteether to yield 80-86% (w/w) pure alpha, beta arteether.
Full Text SINGLE POT CONVERSION OF ARTEMISININ INTO ARTEETHER
Technical Field
The present invention relates to an improved single pot method for preparation of
Arteether from Artemisinin. Arteether prepared from the process is useful for the
treatment of uncomplicated, severe complicated and multi drug resistant malaria.
Background Art
Approximately, out of the 4 billion people suffering from malaria, 1-3 million people,
mostly children die every year worldwide. The rapidly spreading multidrug resistant
parasite to standard quinoline based antimalarial drugs such as chloroquine and
mefloquine based antimalarial complicate chemotherapy treatment of malaria
patients.
Artemisinin and its derivative artemether, arteether, artelinate and artesunate are a
class of antimalarials compounds derived from Artemisia annua which are now
proving their promising activity and are being used for the treatment of uncomplicated
severe complicated /cerebral and multi drug resistant malaria. Dihydroartemisinin is
derived from artemisinin, a sesquiterpene endoperoxide isolated from the plant
Artemisia annua.
Arteether, a ethyl ether derivative of dihydroartemisinin, a drug introduced in India
for the first time by Central Institute of Medicinal and Aromatic Plants (CIMAP),
Lucknow, has undergone extensive preclinical, animal, toxicological studies as well
as clinical studies on Indian subjects for drug regulatory purposes. World Health
Organization (WHO) has also recommended arteether as life saving antimalarial drug.
Arteether is more potential as compared to artemisinin and is an ideal antimalarial
drug especially for treating multi drug resistant and complicated strains of
Plasmodium falciparum. Arteether shows rapid schizoticial action with quicker
clearance rate, short fever clearance time with no side effects and low recrudescence
rate.
Brossi, et al (Brossi, A; Venugopalan, B; Domingueg, GL; Yeh,H.J.C; Flippend-
Anderson, J.L.; Buchs, P; Luo, X.D.; Milhous,W and peters, W; J. Med. Chem. 31,
646-649,1988) reported the preparation of arteether, the ethyl ether derivative of
dihydroartemisinin in two steps: First artemisinin was reduced with an excess of
sodium borohydride in methanol at 0 to -5°C in 3 hours to dihydroartemisinin in 79%
yield. In the second step, arteether is prepared by dissolving the dihydroartemisinin in
the solvent mixture of benzene and ethanol at 45°C followed by addition of BF3
etherate and refluxing the reaction mixture at 70°C for one hour. After the
completion of the reaction it was worked up, dried over anhydrous sodium
sulphate with removal of the solvent dichloromethane. The reaction yielded
arteether along with some impurities. Column chromatography of the reaction mixture
over silica gel, 1:20 ratio yielded pure alpha and beta arteether in nearly equal
qualitative yield.
EL-Feraly et al. (EL Feraly, F.S; Al-Yahya MA; Orabi, K.Y; Mc-Phail DR and Me
Phail A.T. J. Nat. Prod. 55, 878-883, 1992) reported the preparation of arteether by a
process in which anhydrodihydroartemisinin, prepared from artemisinin was
dissolved in absolute alcohol. The reaction mixture was stirred in the presence of ptoluene
sulphonic acid used as a catalyst. On workup it yielded a mixture of beta
arteether and C-ll epimer in the ratio of 3:1. In this process only beta arteether is
obtained and separation of C-ll epimer is difficult and preparation of
anhydrodihydroartemisinin is a tedious process. The reaction took 22 hours to
complete. The lewis acid catalyst used in this reaction is required in large amount
(60mg. acid catalyst by lOOmg. anhydrodihydroartemisinin.
In another method Bhakuni etal (Bhakuni, R.S.; Jain D.C and Sharma R.P. Indian. J.
Chemistry, 34B, 529-30,1995) arteether, artemether and other ether derivatives were
prepared from dihydroartemisinin in different alcohol and benzene in the presence of
chlorotrimethylsilane catalyst in 2-4 hours at room temperature. After workup of the
reaction mixture and removal of the solvent, the impure reaction products were
purified over silica gel column to obtained the pure mixture of alpha, beta ethers.
Another method is reported by Lin et al. (Lin, AJ. and Miller, R.E, J.Med Chem.
38,764-770,1995). In this method the new ether derivatives were prepared by
dissolving dihydroartemisinin in anhydrous ether and appropriate alcohol followed
by BFs - etherate. The reaction mixture was stirred at room temperature for 24 hours.
The yield of the purified products ranged from 40-90%. Purification was achieved by
the use of silica gel chromatography.
Yet another method described by Jain et al (Jain D.C, Bhakuni R.S, Saxena S, kumar,
S and Vishwakarma, R.A. ref: US patent 6,346,631, G.B. Application no 0007261.1
and German application no 10014669.4] teaches preparation of arteether from
artemisinin which comprises: Reduction of artemisinin into dihydroartemisinin,
isolation of dihydroartemisinin, conversion of dihydroartemisinin by dissolving it in
alcohol and adding trialkylorthoformate in the reaction mixture, which produce ethers
in quantitative yield in 10 hours at 40 degree C.
The above mentioned methods are not cost effective and are time consuming.
Moreover, benzene, a carcinogenic solvent, used in the previous methods is not
acceptable according to the health standard. Further, all the above methods require
atleast two separate steps to convert artemisinin into ethers i.e reduction of
artemisinin into dihydroartemisinin in the first pot followed by isolation of
dihydroartemisinin and then comes the second step of conversion of
dihydroartemisinin into different ethers in the second pot.
The Assignees co-pending US Application No. 10/105,964 filed on March 25, 2002
which is incorporated herein as prior art reference teaches a process for preparing
artemether from artemisinin. It should be noted that the aforesaid application does not
use a polyhydroxy compound as catalyst during the process of reduction of
artemisinin into dihydroartemisinin. In the present application, the applicants have
utilized a polyhydroxy compound as a catalyst during the step of reduction of
artemisinin to dihydroartemisinin. The Applicants have been successful in reducing
artemisinin to dihydroartemisinin at room temperature in the presence of the
polyhydroxy compound. It should be noted that to carry out a reaction not only the
reactants play an inportant role but also other reaction conditions like solvent used,
cooling or heating, inert atmosphere etc. are also important. Such reaction conditions
provide a particular structure /steriochemistry to the reactant molecules leading to the
desired product. Introduction of the polyhydroxy compound for the reduction of
artemisinin into dihydroartemisinin provides the ideal environment to the reactant
molecules (artemisinin or the reducing agent or both) to reeact at room temperature
(20-30°C) which other react only at temperature in the range of 0-5°C. Also, the
Applicants have found that the extraction of arteether from an aqueous reaction
mixture using 1% ethyl acetate in »-hexane avoids extraction of unwanted polar
impurities as compared to use of dihydromethane in the co-pending application.
Further, the Applicants have found that resin could not perform esterification and
hence, only unrecoverable catalysts such as chlorotrimethylsilane and p-toluene
sulphonic acid are used in the present process. Thus the process of the present
invention can not be considered as being obvious to a person of ordinary skill in the
art and present invention provides an efficient method for conversion of artemisinin to
arteether.
Object of the invention
The object of the present invention is the development of cost effective and improved
single step method for the preparation of arteether which possesses reduction of
artemisinin into dihydroartemisinin in the presence of a catalyst, conversion of
dihydroartemisinin into arteether followed by extraction of the same in a single pot.
Summary of the invention
The present invention provides a method for the preparation of arteether from
artemisinin in one pot in just about 4 hours. The process of the present invention
comprises: reduction of artemisinin into dihydroartemisinin by less quantity of
sodium borohydride in ethanol at room temperature in the presence of a novel
polyhydroxy catalyst, acylation of dihydroartemisinin in the presence of an acid
catalyst followed by extraction of arteether from an aqueous reaction mixture using
1% ethyl acetate in «-hexane. Workup of the impure arteether followed by silica gel
column chromatography in 1:5-10 ratio, yields 80-86% (w/w) pure alpha, beta
arteether.
Detailed description of the invention
Accordingly, the present invention provides a single step process for the preparation
of arteether from artemisinin in one pot comprising the steps of:
a. dissolving artemisinin and a polyhydroxy catalyst in ethanol at room
temperature to obtain a solution,
b. adding a reducing agent to step (a) solution, stirring the reaction mixture at a
room temperature (20 to 30°C) for about 0.5 to 2 hours to reduce artemisinin
into dihydroartemisinin,
c. adding an acid catalyst to the reaction mixture of step (b) with cooling,
d. stirring the reaction mixture of step (c) for about 1 to 2 hours at room
temperature,
e. adding cold water to the reaction mixture of step (d), extracting with a mixture
of ethyl acetate and w-hexane, separating the organic layer,
f. washing the organic layer of step (e) with 0.5% aqueous sodium bicarbonate
solution followed by water,
g. drying the washed organic layer of step (f) over anhydrous sodium sulphate,
filtering, evaporating the organic layer to obtain a residue, and
h. purifying the residue thus obtained by silica gel column chromatography to
obtain arteether.
In an embodiment of the present invention, the two reactions, namely reduction of
artemisinin to dihydroartemisinin and alkylation of dihydroartemisinin into arteether
are carried out in a single pot thereby avoiding the process of isolation of the
intermediate dihydroartemisinin.
In another embodiment of the present invention, the time required for conversion of
artemisinin into arteether is about 4 hours.
In yet another embodiment of the present invention, ethanol used acts as a solvent and
an alkylating agent.
In still another embodiment of the present invention, the polyhydroxy catalyst is
selected from the group consisting of pholoroglucinol, galactose or dextrose.
In a further embodiment of the present invention, the ratio of artemisinin and the
polyhydroxy catalyst is in the range of 1:2 to 1:5 w/w.
In one more embodiment of the present invention, the reducing agent is selected from
the group consisting of sodium borohydride, lithium aluminium hydride (LiAlH4),
lithium tritert-butoxy aluminium gydride (Li[OC(CH3)3]3 A1H), lithium trimethoxy
aluminium hydride (Li(OCH3)3 A1H), sodium trimethoxy borohydride (Na(OCH3)3
BH), sodium bis-2-methoxy, ethoxy aluminium hydride or a mixture of lithium or
sodium in alcohol or liquid ammonia.
In one another embodiment of the present invention, the reducing agent is preferably
sodium borohydride.
In an embodiment of the present invention, the ratio of artemisinin and sodium
borohydride is in the rage of 1:0.5 to 1:0.7 w/w.
In another embodiment of the present invention, the acid catalyst is a solid or a liquid.
In yet another embodiment of the present invention, the liquid acid catalyst is a
silylated compound.
In still another embodiment of the present invention, the silylated compound is
chlorotrimethysilane.
In one more embodiment of the present invention, the w/v ratio of artemisinin and
chlorotrimethysilane is in the range of 1:3 tol: 4.
In one another embodiment of the present invention, the solid acid catalyst is an
aromatic sulphonic acid.
In a further embodiment of the present invention, the aromatic sulphonic acid is ptoluene
sulphonic acid.
In an embodiment of the present invention, the w/w ratio of artemisinin and p-toluene
sulphonic acid is in the range of 1:3 to 1:4.
In another embodiment of the present invention, the acid catalyst is added to the
reaction mixture at a temperature in the range of from 10° to 23°C.
In yet another embodiment of the present invention, the extraction of crude arteether
from aqueous reaction mixture is carried out with a mixture of 1% ethyl acetate and nhexane
to avoid extraction of unwanted polar impurities.
In still another embodiment of the present invention, the extraction of arteether using
the mixture of 1% ethyl acetate and n-hexane may be performed more than once for
complete extraction.
In one more embodiment of the present invention, the column us eluted using a
gradient mixture of hexane-ethyl acetate having the ratio in the range of 92:8 to
99.5:0.5.
In one another embodiment of the present invention, 80-86% w/w arteether is
obtained after purification by silica gel chromatography.
In a further embodiment of the present invention, the arteether obtained is a mixture
of alpha and beta arteether in the w/w ratio range of 20:80 to 30:70 w/w.
In an embodiment of the present invention, the conversion of artemisinin into pure
arteether takes about 6-8 hours which is significantly less time consuming method.
In another embodiment of the present invention, the yield of the final product i.e. pure
alpha, beta arteether by 3-10% w/w as compared to previously reported methods.
To describe in detail, in the process of invention, artemisinin and polyhydroxy
catalyst were taken in the ratio of 1:2 to 1:5 w/w and dissolved in ethanol at room
temperature and stirred for 5 minutes. Now sodium borohydride is added slowly at the
room temperature (20 to 23°C) and the reaction mixture is stirred for about 0.5 to 1.5
hours.
After completion of the reduction of artemisinin, without workup or the isolation of
the dihydroartemisinin, a solid acid catalyst, resin or a liquid acid catalyst,
chlorotrimethysilane or trifluroacetic acid is added at 10-20°C and the reaction
mixture is further stirred for about 1 to 2 hours at room temperature.
After completion of the acylation reaction, cooled water is added to the reaction
mixture. The solid catalyst is filtered and the filtrate or the aqueous reaction mixture
extracted with 1% ethyl acetate in n-hexane. During extraction of the crude arteethers
with 1% ethyl-acetate in hexane, the polyhydroxy compound remains in the aqueous
phase and is discarded since it is insoluble in hexane-ethyl acetate mixture but soluble
in aqueous phase. The combined ethyl acetate-hexane extract is washed with 0.5%
sodium bicarbonate solution followed by water.
The extract is dried over anhydrous sodium sulphate and removal of the solvent
furnishes impure artemether. Silica gel column chromatography (1:5 to 10 ratio) with
0.5 to 8% ethyl acetate in n-hexane furnishes a mixture of alpha and beta artemether
in 80-86% w/w yield.
Detailed description of the accompanying drawings:
Fig 1 is a schematic representation of the conversion of artemisinin into arteether.
The following examples are given by way of illustration of the present invention and
should not be construed to limit the scope of the present invention.
Example 1
Artemisinin (1 g.) and polyhydroxy catalyst, dextrose (5 g.) were stirred in ethanol
(20ml) at room temperature for 5 minutes. Now sodium borohydride (600 mg) was
added slowly for 10 minutes and the reaction mixture was stirred for about 1 hour at
room temperature (20-23 °C). The reaction was monitored by TLC to check
completion of the reduction step. Acid catalyst chlorotrimethysilane (3.5 ml) was
added slowly at 10-23°C and the reaction mixture was further stirred at room
temperature for about 1 hour. Cooled water (about 150 ml) was added to the reaction
mixture the aqueous reaction mixture was extracted with 1% ethyl acetate in n-hexane
(3 x 50 ml).
The combined ethyl acetate-hexane extract was washed with 0.5% sodium
bicarbonate (100 ml) followed by water (50 ml). The n-hexane extract was dried over
anhydrous sodium sulphate and evaporation of the solvent yielded 1.038 g. of crude
arteether along with some impurities. The impure artemether purified over silica gel
(10 g.) with o.5 to 8% ethyl acetate in hexane furnished a mixture of alpha and beta
arteether 0.86 g. (86% w/w). Small portion of arteether was separated by preparative
TLC into alpha and beta isomers and characterized by Co-TLC and spectral analysis.
Example 2
Artemisinin (1 g.) and polyhydroxy catalyst, dextrose (4 g.) were stirred in ethanol
(15 ml). Sodium borohydride (500 mg.) was added slowly for 10 minutes and the
reaction mixture was stirred for 30 minutes at room temperature (20-23°C). After
completion of the reduction step, chlorotrimethysilane (3.5 ml) was added and the
reaction mixture was further stirred for 1.5 hours at room temperature. After usual
work up and purification through column chromatography (1:5 ratio) a mixture of
alpha and beta arteether (0.805 g., 80.5% w/w) were obtained.
Example 3
Artemisinin (1 g.) and polyhydroxy catalyst, dextrose (2 g.) were stirred in ethanol
(25 ml). Sodium borohydride (700 mg.) was added slowly for 10 minutes and the
reaction mixture was stirred for 1.5 hours at room temperature (20-2 3 °C). After
completion of the reduction step, chlorotrimethysilane (4 ml) was added and the
reaction mixture was further stirred for 2 hours at room temperature (20-23°C) to give
0.95 g. of crude arteether. After usual work up and purification through column
chromatography 0.95 g. of crude arteether yielded 0.825 g. of a mixture of alpha and
beta arteether (82.5% w/w).
Example 4
Artemisinin (100 g.) and polyhydroxy catalyst, dextrose (500 mg.) were stirred in
ethanol (10 ml) for 5 minutes. Sodium borohydride (65 mg.) was added slowly to the
reaction mixture and the same was stirred for 1.25 hours at room temperature (20-
23°C). After completion of the reduction step, /p-toluene sulphonic acid (300 mg.) was
added and the reaction mixture was completed in 4 hours at room temperature. After
usual work up and purification by preparative TLC, the impure reaction product
yielded 53% w/w a mixture of alpha and beta arteether.
Example 5
Artemisinin (100 g.) and polyhydroxy catalyst, galactose (300 mg.) were stirred in
ethanol (5 ml) for 5 minutes. Sodium borohydride (60 mg.) was added slowly to the
reaction mixture and the same was stirred for 1.5 hours at room temperature (20-
23°C). After completion of the reduction step, liquid acid catalyst
chlorotrimethysilane (0.35 ml) was added and the reaction mixture was fUrther stirred
for 2 hours to complete the reaction. After usual work up and purification by
preparative TLC, the impure reaction product afforded 62% w/w of a mixture of alpha
and beta arteether.
Example 6
Artemisinin (100 g.) and polyhydroxy catalyst, phloroglucinol (400 mg.) were stirred
in ethanol. Sodium borohydride (65 mg.) was added slowly to the reaction mixture
and the same was stirred for 2 hours at room temperature (20-23°C). After completion
of the reduction step, chlorotrimethysilane (0.8 ml) was added and the reaction
mixture was further stirred for 2 hours at room temperature to complete the reaction.
Work up and purification of the crude product by preparative TLC yielded 74% w/w a
mixture of alpha and beta arteether.

We Claims:
1. A single pot process for the preparation of arteether from artemisinin comprising
the steps of:
(a) dissolving artemisinin and a polyhydroxy catalyst which is selected from the group consisting of pholoroglucinal, galactose or dextrose in the range of 1:2 to 1:5 w/w in ethanol at room temperature to obtain a solution,
(b) adding a reducing agent which is selected from the group consisting of sodium borohydride, lithium aluminium hydride(LiAlH4),lithium tritert-butoxy aluminium gydride (Li[OC(CH3)3]3 A1H), lithium trimethoxy aluminium hydride (Li(OCH3)3 A1H), sodium trimethoxy borohydride (Na(OCH3)3 BH), sodium bis-2-methoxy, ethoxy aluminium hydride or a
mixture of lithium or sodium in alcohol or liquid ammonia in the range of 1:0.5 to 1:0.7 w/w to solution obtained in step (a), stirring the reaction mixture at a temperature ranging between 20 to 30°C for a period of 0.5 to 2 hours to reduce artemisinin into dihydroartemisinin,
(c) adding a liquid or a solid acid catalyst wherein liquid acid catalyst is a silylated compound is chlorotrimethysilane to the reaction mixture in the range of 1:3 to 1:4 w/w with cooling of step(b),
(d) stirring the reaction mixture of step (c) for a period of 1 to 2 hours at room temperature,
(e) adding cold water 150 ml to the reaction mixture of step (d), extracting with a mixture of 1% ethyl acetate in n-hexane(3x50ml), separating the organic layer and unwanted polar impurities in the range of 92:8 to 99.5:0.5 w/w.
(f) washing the organic layer of step (e) with 0.5% aqueous sodium bicarbonate solution followed by water,
(g) drying the washed organic layer of step (f) over anhydrous sodium sulphate,
filtering, evaporating the organic layer to obtain a residue, and
(h) purifying the residue of step (g) by silica gel column chromatography to obtain arteether.

2. A'process as claimed in claim 1, wherein the two reactions, namely reduction of artemisinin to dihydroartemisinin and alkylation of dihydroartemisinm into arteether are carried out in a single pot thereby avoiding the process of isolation of the intermediate dihydroartemisinin.
3. A process as claimed in claim 1, wherein the time required for conversion of artemisinin into arteether is for a period of 4 hours.
4. A process as claimed in claim 1, wherein ethanol used acts as a solvent and an alkylating agent.
5. A process as claimed in claim 1, herein the reducing agent is preferably socluim borohydride.
6. A process as claimed in claim 1, wherein the solid acid catalyst is an aromatic sulphonic acid.
7. A process as claimed in claim 6, wherein the aromatic sulphonic acid is p-toluenc
sulphonic acid.
8. A process as claimed in claim l,wherein the acid catalyst is added to the reaction
mixture at a temperature preferably in the range of 10to23°C.
{). A-process as claimed in claim 1. wherein 80-86% w/w arteether is obtained after
purification by silica gel chromatography.
10. A process as claimed in claim 1, wherein the arteether obtained is a mixture ol alpha and beta arteether in the w/w ratio range of 20:80 to 30:70.
11. A single pot process for the preparation of arteether from artemisinin substantially as herein describe with refrences to examples accompanying this specification



Documents:

00703-delnp-2004-abstract.pdf

00703-delnp-2004-claims.pdf

00703-delnp-2004-complete specification (granted).pdf

00703-delnp-2004-correspondence-others.pdf

00703-delnp-2004-description (complete).pdf

00703-delnp-2004-form-1.pdf

00703-delnp-2004-form-18.pdf

00703-delnp-2004-form-2.pdf

00703-delnp-2004-form-3.pdf

00703-delnp-2004-form-5.pdf

703-delnp-2004-abstract-(10-12-2008).pdf

703-delnp-2004-claims-(10-12-2008).pdf

703-delnp-2004-correspondence-others-(10-12-2008).pdf

703-delnp-2004-form-2-(10-12-2008)'.pdf

703-delnp-2004-petition-137-(10-12-2008).pdf


Patent Number 234267
Indian Patent Application Number 00703/DELNP/2004
PG Journal Number 25/2009
Publication Date 19-Jun-2009
Grant Date 14-May-2009
Date of Filing 19-Mar-2004
Name of Patentee COUNCIL OF SCIENTIFIC & INDUSTRIAL RESEARCH
Applicant Address RAFI MARG, NEW DELHI-110001, INDIA.
Inventors:
# Inventor's Name Inventor's Address
1 RAJENDRA SINGH BHAKUNI CENTRAL INSTITUTE OF MEDICINAL AND AROMATIC PLANTS, P.O.CIMAP, LUCKNOW, INDIA, 226015.
2 AMIT TEWARI CENTRAL INSTITUTE OF MEDICINAL AND AROMATIC PLANTS, P.O.CIMAP, LUCKNOW, INDIA, 226015.
3 TARUN SINGH CENTRAL INSTITUTE OF MEDICINAL AND AROMATIC PLANTS, P.O.CIMAP, LUCKNOW, INDIA, 226015.
4 SUMAN PREET SINGH KHANUJA CENTRAL INSTITUTE OF MEDICINAL AND AROMATIC PLANTS, P.O.CIMAP, LUCKNOW, INDIA, 226015.
PCT International Classification Number C07D 321/12
PCT International Application Number PCT/IB02/05074
PCT International Filing date 2002-11-29
PCT Conventions:
# PCT Application Number Date of Convention Priority Country
1 10/308,651 2002-12-02 U.S.A.