FORM 2
THE PATENTS ACT 1970
(Act 39 of 1970)
&
THE PATENTS RULE, 2003
COMPLETE SPECIFICATION
(SECTION 10 and Rule 13)
TITLE OF THE INVENTION
"AN IMPROVED PROCESS FOR PREPARATION OF
GADOBENATE DIMEGLUMINE"
Emcure Pharmaceuticals Limited,
an Indian company, registered under the Indian Company's Act 1957
and having its registered office at
Emcure House, T-184, M.I.D.C., Bhosari, Pune-411026, India.
THE FOLLOWING SPECIFICATION DESCRIBES THE NATURE OF THE INVENTION AND THE MANNER IN WHICH IT IS TO BE PERFORMED

FIELD OF THE INVENTION
The present invention relates to an improved process for the preparation of Gadobenate dimeglumine conforming to regulatory specifications. The invention specifically relates to preparation of 4-carboxyl-5,8,l l-tri(carboxymethyl)-l-phenyl-2-oxa-5,8,ll-triazatridecan-13-oic acid (BOPTA) of formula (I), a key intermediate in synthesis of Gadobenate dimeglumine. The synthetic method for compound (I) comprises reaction of tertiary-butylcarboxymethyl substituted diethylenetriamine with tertiary butyl(3-benzyloxy-2-bromo)propionate, followed by alkylation with tertiary-butylbromoacetate and subsequent deprotection to give the desired intermediate (I).
BACKGROUND OF THE INVENTION
Gadobenate dimeglumine, also known as GdBOPTA Dimeg (Multihance®) belongs to the class of magnetic resonance imaging (MRI) contrast agents, which are complex paramagnetic salts and which, when administered in the body tissues help in improving the diagnostic information obtained from the imaging technique. Gadobenic acid, which is administered as its dimeglumine salt, is a complex of the transition metal gadolinium with the ligand, 4-carboxyl-5,8,l l-tri(carboxymethyl)-l-phenyl-2-oxa-5,8,l l-triazatridecan-13-oic acid (BOPTA). The active ingredient functions as a gadolinium based MRI contrast agent, and is employed for the detection of focal liver lesions in liver cancer patients as well as in MRI of central nervous system.

EP 230893 discloses synthesis of BOPTA which comprises reaction of diethylenetriamine (DETA) with 3-phenylmethoxy-2-chloropropionic acid to give a serine derivative, which on carboxymethylation with bromoacetic acid and subsequent purification by resin treatment gives BOPTA (I).

US 6,162,947 discloses a process wherein diethylenetriamine is treated with potassium salt of 2-chloro-3-(phenylmethoxy)propionic acid and the resulting intermediate, upon further carboxymethylation with bromoacetic acid at basic pH, gives BOPTA.
Both the methods involve monoalkylation of the unsubstituted polyamine DETA wherein there is an inherent danger of polyalkylation and associated reactions, which leads to undesired by-products. Although measures such as use of large excess of DETA, careful control on pH of the reaction mixture etc. have been incorporated in the procedures, they seem to be inadequate since the method still involves resin purification step for the intermediates at each stage of synthesis. Control of pH is quite critical for the reaction because at lower pH, formation of quaternary ammonium salts is likely to compete with the desired reaction while at higher pH values, higher quantities of bromoacetic acid are required due to substitution of the functional group bromine with a hydroxyl group which is also accompanied by degradation of benzyloxypropionic moiety. Further, during the reaction, the undesired condensation of the free carboxylic acid moiety in the molecule with adjacent amine group gives rise to a six-membered lactam impurity, which is very difficult to separate and hampers purity of the desired product.
US 7,592,482 also discloses the synthesis of BOPTA, wherein monoalkylation of DETA is carried out using potassium salt of 2-chloro-3-methylphenoxypropionic acid. However, further reaction of the benzyloxy substituted polyamine is effected using a large excess of the alkylating agent, bromoacetic acid (molar ratio of acid : amine is between 7:1 and 9:1). Also, the procedure involves a stage-wise, sequential, and alternate dosing of aliquots of aqueous solutions of bromoacetic acid and sodium hydroxide at different temperatures. Although the process discloses a synthetic route wherein any critical pH control is not needed, the involvement of highly cumbersome mode of addition of stipulated quantities of the aliquots of reagent solutions having specific concentrations in alternate manner, at different temperatures, makes the process unsuitable for commercial scale operations. Also, the alkylation reaction being extremely sensitive to pH, any minor change in mode of addition, temperature, concentrations of reagent solutions and time of their addition could lead to a high percentage of undesired side products, thereby leading to additional purification steps, which increase the cost exorbitantly.

Hence, there is a need for an efficient and economical process for the synthesis of gadobenic acid on industrial scale, which is capable of restricting the formation of undesired side products which are difficult to separate, avoids high excesses of reagents, cumbersome operational procedures and yields the desired product with purity conforming to regulatory norms.
While carrying out experimentation for developing an industrially suitable process for gadobenic acid, the present inventors surprisingly observed that the aforementioned shortcomings in the prior art could be circumvented by carrying out the alkylation reaction with N-substituted diethylenetriamine derivative. When diethylenetriamine derivative tetrasubstituted or protected with tertiarybutyl carboxymethyl group was subjected to reaction with tertiary butyl -2-bromo-3-benzyloxy propionate, alkylation was selective and did not require excessive diethylenetriamine to give the corresponding alkylated derivative in quantitative yield and with desired purity. In the case of diethylenetriamine trisubstituted with tertiarybutyl carboxymethyl group, alkylation was again selective and subsequent reaction with tertiarybutyl bromopropionate gave the desired tetrasubstituted diethylenetriamine derivative. The tetrasubstituted DETA obtained in both the cases was subjected to deprotection of the tertiary butyl group to give BOPTA having desired purity.
CN 102603550 discloses reaction of diethylenetriamine with tertiarybutyl-2-bromo-3-benzyloxy propionate and polyalkylation of resulting compound using tertiarybutyl bromoacetate followed by deprotection reaction to yield BOPTA. However, the method does not mention any remedy for the undesired polyalkylated impurities, which are reported in most of the prior art references that deal with monoalkylation of polyamines such as diemylenetriamine. Further, the method for preparation of 2-bromo-3-benzyloxy tertiary butyl propionate from benzyloxy serine is quite uneconomical and does not have practical utility. This route necessitates protection and deprotection of nitrogen and carboxyl functionalities in the starting material serine, and also involves low-yielding, hazardous reactions like diazotization, which adds to the cost of the desired product.

It is pertinent to mention that replication of the said method for preparing BOPTA yielded numerous associated impurities based on TLC. Further, none of the compounds isolated from the reaction mass matched the mass spectrum of the desired compound, BOPTA.
Therefore, the present inventors embarked on developing a process which avoided the drawbacks of prior art methods. The developed process for preparation of 4-carboxy-5,8,1 l-tri(carboxymethyl)-l-phenyl-2-oxa-5,8,ll-triazatridecan-13-oic acid (BOPTA) comprises alkylation of trisubstituted diethylenetriamine with tertiarybutyl-2-bromo-3-benzyloxy propionate, followed by reaction of the resulting compound with tertiary-butyl bromoacetate and subsequent treatment with trifluoroacetic acid to provide BOPTA (I) in good yield and with purity conforming to regulatory specifications. BOPTA having desired purity was also obtained from tetrasubstituted DETA by following similar reaction sequence.
OBJECT OF THE INVENTION
An objective of the present invention is to provide a convenient, industrially viable process for the synthesis of 4-carboxyl-5,8,ll-tri(carboxymethyl)-l-phenyl-2-oxa-5,8,ll-triazatridecan-13-oic acid (BOPTA) wherein excessive use of reactants is avoided and formation of impurities is restricted below the desired limits.
Another object of the invention is to provide an economical method for synmesis of BOPTA which comprises monoalkylation of methyl tertiary-butyl carboxylated diethylenetriamines, optional reaction of the alkylated amine with bromoacetate, which is carried out only in case of trisubstituted diethylenetriamine, followed by facile deprotection of the resulting compound to yield BOPTA of formula (I) having purity conforming to regulatory specifications.
SUMMARY OF THE INVENTION
The present invention relates to a novel process for the preparation of 4-carboxyl-5,8,l 1-tri (carboxymethyl)-l-phenyl-2-oxa-5,8,ll-triazatridecan-13-oic acid (BOPTA) (I), wherein associated impurities are minimized below regulatory limits.

An aspect of the present invention relates to a process for preparation of 4-carboxyl-5,8,l 1-tri (carboxymemyl)-l-phenyl-2-oxa-5,8,ll-triazatridecan-13-oic acid (BOPTA) of formula (I) comprising reaction of tertiarybutylcarboxymethyl substituted diethylenetriamine (II) with tertiary butyl-2-bromo-3-benzyloxy propionate in presence of diisopropylethylamine and an organic solvent such as acetonitrile or dimethylformamide in the temperature range of 60-80°C, to give compound (III or IIIa), which on treatment with tertiary butyl bromoacetate in presence of diisopropylethylamine and solvent dimethylformamide at 25-30°C, followed by subsequent reaction of the resulting compound with trifluoroacetic acid and isolation of die product gives BOPTA of formula (I). The tertiarybutylcarboxymethyl substituted diethylenetriamines are 4,7,7'-N-tris-(tertiarybutylcarboxymethyl) diethylenetriamine (IIa) or 1,4,7,7' N-tetra-(tertiary butyl carboxymethyl) diethylenetriamine (lib), which, when subjected to reaction with tertiary butyl-2-bromo-3-benzyloxy propionate give (IIIa) and (III) respectively and only compound (IIIa) is subjected to reaction with tertiary butyl bromoacetate, prior to reaction with trifluoroacetic acid.
These objectives of the present invention will become more apparent from the following detailed description.
DETAILED DESCRD?TION OF THE INVENTION
The present inventors have carried out extensive experimentation for development of an economical, industrially viable synthetic route for BOPTA. During the process of development, it was observed that although diethylenetriamine (DETA) was an appropriate starting material for synthesis of BOPTA, however the presence of multiple nitrogen functions in DETA led to polyalkylation resulting in various undesired products in significant proportions. Hence, efforts were directed to the selection of appropriate protecting group for the nitrogen sites which would prevent undesired polyalkylation. The highly stable methyl tertiarybutyl carboxyl group was selected for protecting active nitrogen sites in DETA as it could withstand the highly basic reaction medium and was stable to autoclave conditions as well. The resulting polysubstituted DETA was used as a starting material for synthesis of BOPTA.
The inventors observed that the methyl tertiarybutyl carboxyl group protected DETA helped in control of undesired poly alkylated side products and avoided excessive quantities

of DETA to restrict polyalkylation. Further, the lactam impurity arising from condensation of free amine group with carboxylic acid functionality was also minimized because of the protected amine functionalities. It was also evident that both trisubstituted and tetrasubstituted diethylenetriamines could be used as starting materials in the synthesis of BOPTA provided that in case of trisubstituted diemylenetriamine, product obtained after monoalkylation step was subjected to reaction with tertiary butyl bromoacetate, prior to the deprotection step.

Scheme 1: Method embodied in the present invention for the preparation of BOPTA (I)
In an embodiment, 4 7,7-N-tris-(tertiarybutylcarboxymethyl) diethylenetriamine of formula (IIa) was treated with tertiary butyl-(2-bromo-3-benzyloxy) propionate in presence of a base and an organic solvent in the temperature range of 65 to 85°C, preferably 75-80°C. The base for above reaction was diisopropylethylamine while the organic solvent was acetonitrile.

Upon completion of the reaction, as monitored by TLC, the reaction mass was cooled and concentrated to obtain a residue, which was purified by column chromatography on silica gel using ethyl acetate / n-hexane mixture as eluent to give l-phenyl-2-oxa-4-tertiarybutyl-carboxylate-8,1 l-di(tertiarybutylcarboxymethyl)-5,8,l l-trazatridecan-13-tertiary-butylcarboxylate of formula (IIIa)
Further reaction of compound (Ilia) with tertiarybutyl bromoacetate was carried out at 25 to 30°C, in an organic solvent, in presence of diisopropylethylamine as base to give 1-phenyl-2-oxa-4-tertiary-butyl-carboxylate-5,8,1 l-tri-(teitiarybutylcarboxymethyl)-5,8,l 1-triazatridecan-13-tertiary-butylcarboxylate of formula (III).
The organic solvent was selected from the group comprising of dimethyl formamide, dimethyl acetamide, dimethyl sulfoxide etc.
After completion of reaction, as monitored by TLC, the reaction mass was quenched with water and extracted with toluene. Distillation of the organic layer gave an oily residue, which was purified by column chromatography on silica gel column using ethyl acetate and n-hexane as eluent.
Compound (III). was also obtained by reaction of 1,4,7,7'-N-tetra-(tertiarybutylcarboxymethyl) diethylenetriamine of formula (lib) with tertiary butyl-(2-bromo-3-benzyloxy) propionate in presence of diisopropylethylamine at around 60°C using dimethylformamide as solvent. After completion of the reaction, as monitored by TLC, reaction mass was cooled, quenched with water and extracted with toluene, followed by. concentration of the organic layer to give a residue. The desired compound (III) was obtained by subjecting the residue to column chromatographic separation on silica gel column using ethyl acetate and n-hexane mixture for elution.
In a further embodiment, trifluoroacetic acid was added to compound (III) at room temperature and the reaction mixture was stirred at the same temperature till completion of the reaction, as monitored by TLC. The reaction mass was concentrated under reduced pressure to obtain a residue. Water was added to this residue, followed by addition of

aqueous sodium hydroxide solution till pH 12 was attained. The solution was treated with Amberlite IRA 120 (H+ form), followed by filtration and further treatment of the resin with aqueous ammonia solution. The resin was filtered off and the aqueous filtrate was acidified to pH 2 using hydrochloric acid. Separation of the water layer and crystallization of the oily residue yielded BOPTA having desired purity.
The compounds 4,7,7-N-uisKtertiarybutylcarboxymemyl)diemylenetriamine (Ha), 1,4,7,7'-N-terra-(tertiaiybutylcarboxymemyl)diemylenetriamine (lib), tertiarybutyl-(2-bromo-3-benzyloxy) propionate and tertiarybutyl bromoacetate were prepared based on procedures reported in the literature.
The following examples are meant to be illustrative of the present invention. These examples exemplify the invention and are not to be construed as limiting the scope of the invention.
EXAMPLES
Example 1:
1 -Phenyl-2-oxa-4-tertiarybutyl-carboxylate-8,l l-di(tertiarybutylcarboxymethyl)-
5,8,11-triazatridecan-13-tertiary-butylcarboxylate(IIIa)
4,7,7-N-tris-(tertiarybutylcarboxymethyl) diethylenetriamine (Ha; 200.8gms) was added to a mixture of diisopropylethylamine (116.0gms), tertiarybutyl-(2-bromo-3-benzyloxy) propionate (211.7gms) and acetonitrile (1000ml) at room temperature. The reaction mixture was heated to 70-80°C and continued till completion, as monitored by TLC. The reaction mass was cooled to 25-35C and concentrated under reduced pressure to obtain a residue which was purified by column chromatography using ethyl acetate and hexane as the eluents. Yield: 180.7gms

Example 2:
l-Phenyl-2-oxa-4-tertiary-butyl-carboxylate-5,8,ll-tri-(tertiarybutylcarboxymethyl)-
5,8,ll-triazatridecan-13-tertiary-butylcarboxylate(III)
Tertiarybutyl bromoacetate (56.6 gms) was added to a mixture of compound IIIa (180.2 g) dissolved in a mixture of diisopropylethylamine (68.4 g) and dimethyl formamide (500 ml) at 25-30°C and stirred at the same temperature till completion of reaction as monitored by TLC. The reaction mass was quenched with water and extracted with toluene. Concentration of the organic layer under reduced pressure gave an oily residue, which was purified by column chromatography on silica gel using ethyl acetate and hexane to give compound III. Yield: 100.5 gms. Example 3:
l-Phenyl-2-oxa-4-tertiary-butyl-carboxylate-5,8,ll-tri-(teriiarybutyIcarboxymethyl) -5,8,ll-triazatridecan-13-tertiary-butylcarboxyIate (in) 1,4,7,7 N-tetra-(tertiary butyl carboxymethyl) diethylenetriamine (IIb, 251.6 gms) was added to a mixture of diisopropylethylamine (116.1 gms), tertiarybutyI-(2-bromo-3-benzyloxy) propionate (213.5 gms) and dimethylformamide (900 ml) at room temperature. The reaction mixture was heated to 55-60 C and reaction was continued till completion, as monitored by TLC. After completion, the reaction mass was cooled, quenched with water and extracted with toluene. After concentrating the organic layer, a residue was obtained which was purified by column chromatography using ethyl acetate and hexane as the eluents.
Yield: 198.3 gms Example 4:
4-carboxyl-5,8,11-tri(carboxymethyl)-l-phenyl-2-oxa-5,8,ll-triazatridecan-13-oic acid (BOPTA) (I)
A mixture of compound III (100.3 g) in trifluoroacetic acid (500 ml) was stirred at 25-30°C till completion of the reaction as monitored by TLC. The reaction mass was concentrated under reduced pressure to obtain a residue, which was then dissolved in water (100 ml) and treated with aqueous solution of sodium hydroxide till pH 12. Amberlite IRA 120 (H+ form) was added to the resulting solution, stirred and filtered. The filtrate was treated with aqueous ammonia solution and filtered. The aqueous filtrate was concentrated under

reduced pressure to obtain a residue, which was dissolved in water and the solution was
acidified to pH 2 using hydrochloric acid. Separation of the water layer and crystallization
of the oily residue from ethanol gave BOPTA (I).
Yield: 40.5 g
Purity: 99.6%
Example 5:
Gadobenate dimeglumine
The compound (I), BOPTA obtained in Example 4 (50.5 gms) was suspended in water (300
ml) and stirred with aqueous solution of N-methyl glucamine (36.4 gms in 180 ml water) at
40 to 50°C until complete dissolution. The solution was treated with Gd2O3 (18.0 gms) at
70 to 85°C monitoring completion of the complex formation by HPLC, to obtain
Gadobenate dimeglumine.
Purity: 95 %
Example 6:
Tertiary butyl-(2-bromo-3-benzyIoxy)propionate
Solution of bromine (246.5gms) in dichloromethane (250 ml) was slowly added to a stirred
solution of tertiary butyl acrylate (200 g) in dichloromethane (1500 ml) at -5 to 0°C, and
the reaction was continued at the same temperature till completion, as monitored by TLC.
The reaction mass was concentrated under vacuum to obtain an oily residue of
tertiarybutyl-2,3-dibromopropionate. The residue was dissolved in dimethylformamide
(180 ml). A mixture of benzyl alcohol (120 ml), sodium hydroxide (12.4 g) and water (15
ml) was heated in an inert atmosphere at 90°C and after cooling to ambient temperature
was gradually added to the DMF solution of the dibromopropionate residue at 0 to 5°C and
stirred at the same temperature. Completion of the reaction was monitored by TLC. The
reaction mixture was quenched and extracted with toluene. Separation and concentration of
the organic layer gave an oily liquid, which upon column chromatographic purification on
silica gel using a mixture of ethyl acetate and hexane as eluant yielded tertiary butyl-2-
bromo-3 benzyloxy propionate.
Yield : 140 g
Purity: 97%

Example 7:
4,7,7-N-tris-(tertiarybutylcarboxymethyl)diethylene triamjne (Ha)
A mixture of trityl chloride (200.2 g) in dichloromethane (350 ml) was gradually added to a solution of diethylenetriamine (370.8gms) in 200 ml dichloroinethane at 25 to 30°C and the reaction mixture was stirred at the same temperature till completion of the reaction as monitored by TLC. The reaction mixture was quenched with water. Separation and concentration of organic layer gave tritylated diethylenetriamine (trityl-DETA). Yield: 240 gms.
Diisopropylethylamine (DEPEA), (261.4g) was gradually added to a solution of trityl-DETA, (200.2 g) in DMF (600 ml) at 25 to 30°C, followed by drop wise addition of tertiarybutyl bromoacetate (361.2 g) at the same temperature. The reaction was monitored by TLC and after completion, the reaction mass was mixed with water and extracted with toluene. The organic layer was separated, washed with a dilute acid solution, was separated, and concentrated to give l-N-trityl-4,7,7'-N-tris-.(tertiarybutylcarboxymethyl) diethylenetriamine (trityl-tri-Boc-DETA), which separated as an oil Yield: 385.6 gms
A mixture of methanol (5000ml) and trityl-tri-Boc-DETA (350.5 g) was charged in the autoclave, followed by addition of 10%Pd/C (12g). The reaction was carried out at 50-60°C under hydrogen pressure of 4-6 kg/cm . After completion of the reaction, as monitored by HPLC, the reaction mass was filtered and the filtrate was concentrated under vacuum, followed by addition of hexane to the residue to yield 4,7,7'-N-tris-(tertiarybutylcarboxymethyl) diethylenetriamine (IIa) Yield: 170.3 gms. Example 8:
1,4,7,7' N-tetra-(tertiary butylcarboxymethyl) diethylenetriamine (lib)
Di-tertiary butyl dicarbonate (34.9 g) in methanol (400 ml) was gradually added to a
mixture of diethylenetriamine (15.0 g) and methanol (600 ml) at 0°C and the reaction
mixture was stirred till completion of the reaction as monitored by TLC. The reaction mass
was concentrated under reduced pressure to obtain N-
tertiarybutylcarboxydiethylenetriamine, (Boc-DETA) Yield: 25.7 gms

Benzaldehyde (13.7 g) was added to a mixture of Boc-DETA (24.0 g) and ethanol (100ml) and the mass was stirred at 50-60°C till completion of the reaction as monitored by TLC. Sodium borohydride (7.1g) was added to the reaction mass and stirred at 50-60°C. After completion of the reaction, as monitored by TLC, the mass was cooled, quenched with water and extracted with dichloromethane. The organic layer was separated, acidified with hydrochloric acid and stirred further at room temperature. The aqueous layer was separated, basified using aqueous sodium hydroxide solution and extracted with dichloromethane. Concentration of the organic layer yielded N-benzyl diethylenetriamine, (benzyl- DETA). Yield: 12.5 gms.
Tertiary-butyl bromoacetate (44.6 g).was gradually added to a mixture of benzyl-DETA (lO.lg) and dimethylformamide (60 ml) at room temperature. The reaction mass was stirred at the same temperature till completion of the reaction as monitored by TLC. After completion, the reaction mass was quenched with water and extracted with toluene. The organic layer was separated, concentrated to give N-l-benzyl,l,4,7,7-N-tetra-(tertiary butylcarboxymethyl)diethylenetriamine, (benzyl-tetraBoc-DETA). Yield: 15.6 gms.
A mixture of methanol (100 ml), benzyl-tetraBoc-DETA (10.3 g) was charged in the
autoclave, followed by addition of Pd/C (10%, lg). The reaction was carried out at 50-60°C
under hydrogen pressure of 3-4 Kg/cm .
After completion of the reaction, as monitored by HPLC, the reaction mass was filtered and
the filtrate was concentrated under vacuum to yield 1,4,7,7 N-tetra-(tertiary butyl
carboxymethyl) diethylenetriamine (IIb).
Yield: 5.8 gms

We claim:
1. A process for preparation of 4n;arboxyl-5,8,ll-tri(carboxymethyl)-l-phenyl-2-oxa-5,8,11-triazatridecan-13-oic acid (BOPTA) of formula (I) comprising reaction of tertiarybutylcarboxymethyl substituted diethylenetriamine (II) with tertiary butyl-2-bromo-3-benzyloxy propionate in presence of a base and an organic solvent to give compound (III or IIIa), which on treatment with tertiarybutyl bromoacetate in presence of diisopropylethylamine and solvent dimethylformamide at 25-30°C, followed by subsequent reaction of the resulting compound with trifluoroacetic acid and isolation of the product gives BOPTA of formula (I).
2. A process according to claim 1 wherein tertiarybutylcarboxymethyl substituted diethylenetriamines are 4,7,7'-N-tris-(tertiarybutylcarboxymethyl) diethylenetriamine (Ha) or 1,4,7,7' N-tetra-(tertiary butyl carboxymethyl) diemylenetriamine (IIb), which, when subjected to reaction with tertiary butyl-2-bromo-3-benzyloxy propionate give (IIIa) and (III) respectively.

3. A process according to claims 1 and 2 wherein only compound (IIIa) is subjected to reaction with tertiary butyl bromoacetate, prior to reaction with trifluoroacetic acid.
4. A process according to claim 1 wherein the base is diisopropylethylamine, the organic solvent is selected from acetonitrile and dimethylformamide and the reaction is carried out in the temperature range of 60-80°C.