FORM 2
THE PATENTS ACT, 1970
(39 of 1970)
&
THE PATENTS RULES, 2003
COMPLETE SPECIFICATIONS
(See section 10, rule 13)
"PROCESS FOR PREPARATION OF MONOISOAMYL MESO-2,3-DIMERCAPTOSUCCINATE (MiADMSA)"
CADILA PHARMACEUTICALS LIMITED
Cadila Corporate Campus", Sarkhej - Dholka Road, Bhat, Ahmedabad - 382210,
Gujarat, India
The following specification particularly describes the invention and the manner in which it is to be performed.

FIELD OF THE INVENTION:
The present invention relates to a process for the preparation of monoisoamyl meso-2, 3-dimercaptosuccinate (MiADMSA) with high purity. More specifically, it relates to an industrially viable process for purification of monoisoamyl meso-2, 3-dimercaptosuccinate (MiADMSA) by using a solvent mixture of hydrocarbon and ether.
BACKGROUND OF THE INVENTION:
Pharmocol. and toxicol. 1992, 70, 336-343 discloses monoalkyl ester of meso-2, 3-dimercaptosuccinic acid (DMSA). DMSA monoesters are useful in cadmium, nickel, lead, mercury and arsenic intoxication. In vivo studies suggested that DMSA monoesters could be of great medicinal importance as being more efficient metal mobilizer than DMSA and could be potential drugs for chelation therapy in metal intoxication. The maximum mobilization of metal is observed with Monoisoamyl ester of meso-2, 3-dimercaptosuccinic acid (MiADMSA) (formula I).

Chem. Res. Toxicol. 1991, 4,107-114 describes the preparation of monomethyl ester of DMSA by refluxing DMSA in 1M methanolic HCI for 1hr followed by quenching with water. Product so obtained contains monomethylester, dimethylester and DMSA. Presence of impurities like diacid and dimethylester in final product is undesirable and reduces clinical value of the product.
Pharmocol. and toxicol. 1992, 70, 336-343 describes the preparation of MiADMSA by refluxing DMSA and isoamyl alcohol in aqueous HCI. It overcomes the shortcomings of Chem. Res. Toxicol: 1991, 4, 107-114 by using aqueous HCI instead of methanolic HCI to minimize the formation of diester (Formula III).


To remove these impurities, the crude solid is stirred with hexane to wash off the diester impurity. Hexane purified residue is further subjected to Soxhlet extraction with chloroform for 24 hrs to obtain purified monoester. The process requires multiple extractions for prolonged period making it unsuitable for large scale industrial application. Prolonged heating and multiple extractions could lead to oxidation of thiol functional group generating additional oxidized impurities.
In view of above, there is need for an industrially viable process which overcomes the shortcomings of prior art processes and which purifies MiADMSA in effective way with lesser process time.
OBJECT OF THE INVENTION:
The main object of the invention is to provide an industrially viable process for the preparation of MiADMSA with high purity.
Another object of the invention is to provide a process for the purification of monoisoamyl meso- 2, 3-dimercaptosuccinate (MiADMSA).
Yet another object of the invention is to provide an industrial process for separation of MiADMSA and unreacted DMSA.
SUMMARY OF THE INVENTION:
In accordance with the objectives, the present invention provides a cost effective synthesis of highly pure monoisoamyl meso-2, 3-dimercaptosuccinate (MiADMSA).
The process involves the esterification of meso-2,3-dimercaptosuccinic acid with isoamyl alcohol by using concentrated hydrochloric acid. Reaction mass containing esterified product was quenched with water, basified and washed with toluene. The pH of aqueous layer was adjusted and then extracted using solvent mixture of hydrocarbon and ether. MiADMSA was isolated from the extract by conventional means. If required it can be purified further from a suitable solvent or solvent mixture.
DETAILED DESCRIPTION OF THE INVENTION:
The invention provides an industrially viable process for the preparation of MiADMSA in high purity. This process is particularly advantageous in comparison with known methods because the purification is carried out by using less toxic solvents for generation of ICH

grade MiADMSA without performing prolonged Soxhlet extraction with toxic solvent like chloroform. The present invention eliminates the risk of handling toxic solvents, reduces purification time, thus making the process more economical and industrially viable.
In context of the present invention, the term "high purity" is intended to mean that MiADMSA having purity of 96% or more.
A process for the preparation of monoisoamyl meso-2,3-dimercaptosuccinate (MiADMSA) in high purity comprises the steps of:
(a) esterifying meso-2,3-dimercaptosuccinic acid with isoamyl alcohol using
concentrated hydrochloric acid;
(b) quenching the reaction mass containing esterified product with water;
(c) basifying the quenched reaction mass till pH of 8 - 8.5;
(d) washing the basified reaction mass with toluene;
(e) adjusting the pH of aqueous layer between 1 to 4 using acid;
(f) extracting the acidified aqueous layer using solvent mixture of hydrocarbon and ether;
(g) isolating MiADMSA from the extract; and
(h) optionally purifying isolated MiADMSA
Esterification in step (a) is carried out at a temperature ranging from room temperature to elevated temperature such as the reflux temperature of isoamyl alcohol, preferably at a temperature ranging from 110-115°C. Isoamly alcohol used for the esterification is having purity of more than 99% as determined by gas chromatography.
Basification in step (c) is carried out using base selected from sodium bicarbonate, potassium bicarbonate, calcium bicarbonate, sodium carbonate, potassium carbonate, cesium carbonate, calcium carbonate, sodium hydroxide, potassium hydroxide, cesium hydroxide, barium hydroxide or their like. Preferably, the base is sodium bicarbonate.
The acid used in step (e) to adjust the pH of the reaction mass is selected from inorganic acid such as hydrochloric acid, nitric acid, sulfuric acid, p-toluene sulfonic acid or like; organic acid such as tartaric acid, oxalic acid, mandelic acid, fumaric acid, acetic acid, formic acid or their like. Preferably, the acid is hydrochloric acid.
Hydrocarbon used in step (f) is selected from the group consisting of C1 to C8 hydrocarbons, preferably aliphatic hydrocarbons such as hexane, heptane or cyclohexane; aromatic hydrocarbons such as toluene or xylene. Preferable, the hydrocarbon is toluene.

Ether used in step (f) is selected from the group consisting of methyl tert-butyl ether (MTBE), tetrahydrofuran (THF), 2-methyl tetrahydrofuran, diethyl ether or diisopropyl ether (DIPE). Preferably, the ether is methyl tert-butyl ether (MTBE).
Isolation in step (g) is carried out by cooling, drying, adding an anti-solvent or seeding. Preferably, the isolation is done by drying under reduced pressure.
The purified MiADMSA obtained in step (g) can be used as it is or after further purification by a conventional manner.
The pH adjustment during step (e), before extraction with mixture of hydrocarbon and ether is very crucial. The pH of aqueous layer is adjusted by using acid within the range of 1 to 4. Preferably, the pH of aqueous layer is between 2.5 to 3.
The ratio of hydrocarbon to ether used in step (f) for extraction is selected from the range of 80:20 % to 90:10 % by volume. The ratio of hydrocarbon to ether used in the process is properly determined. Optimum ratio of hydrocarbon to ether is very vital for selective extraction of MiADMSA from the mixture of MiADMSA and DMSA. Higher amount of ether leads to ineffective extraction of MiADMSA, wherein MiADMSA is contaminated by DMSA and use of lower amount of ether leads to incomplete extraction of MiADMSA and reducing the yield.
In preferred embodiment, the extraction in step (f) is carried out by using 83:17 % by volume mixture of toluene:MTBE, wherein the pH of aqueous layer is between 2.5 to 3.
The process for the separation and purification used in the present invention is not restricted to meso-2,3-dimercaptosuccinic acid derivatives but is also applicable to optically active derivatives i.e. R,R and S,S isomers.
The invention is further illustrated by following non-limiting example:
Example-1: Process for the separation and purification of MiADMSA:
meso-2,3-dimercaptosuccnic acid (100 g) was taken in isoamyl alcohol (1500 ml). To this, concentrated hydrochloric acid (50 ml) was charged at 25-30°C. The contents were stirred for 35-40 minutes at 110-115°C. The reaction mass was then cooled at 50-60°C. The reaction mass was concentrated under reduced pressure. The residue was quenched with water (200 ml), basified with NaHCO3 till pH of 8 - 8.5 and washed with toluene (300+200 ml). The pH of aqueous layer was adjusted to 2.5 - 3 by adding concentrated hydrochloric acid. The aqueous layer was then extracted with a 5:1 mixture of toluene:MTBE (600 ml). The clear toluene:MTBE extract was then concentrated under reduced pressure to give titled compound (Yield - 26.03%). Purity by HPLC - 99.33%. Total impurities - 0.67%. DMSA -not detected.

In analogy to reaction conditions o: follow: example 1, example 2 to 10 can be summarized as
Example No pH adjusted byHCI Solvent for extraction Yield
(%) Purity by HPLC
Toluene MTBE MiADMSA DMSA Total Impurities
Ex.2 2.5-3 5.0 1.0 26.03 99.33 Not detected 0.67
Ex.3 2.5-3 5.0 1.0 27.26 99.51 0.03 0.49
Ex.4 2.5-3 5.0 1.5 23.86 97.75 1.86 2.25
Ex.5 2.5-3 5.0 0.75 25.45 98.51 0.03 1.49
Ex.6 2.5-3 6.0 1.0 29.21 98.90 0.01 1.10
Ex.7 2.5-3 4.0 1.0 28.34 98.91 0.02 1.09
Ex.8 1-1.5 5.0 1.0 27.48 94.57 4.85 5.43
Ex.9 1.5-2 5.0 1.0 31.81 96.58 2.46 3.42
Ex.10 3.5-4 5.0 1.0 15.91 98.98 0.04 1.02

We claim,
1. A process for the preparation of monoisoamyl meso-2,3-dimercaptosuccinate (MiADMSA) with high purity comprises the steps of:
(a) esterifying meso-2,3-dimercaptosuccinic acid with isoamyl alcohol using
concentrated hydrochloric acid;
(b) quenching the reaction mass containing sterified product with water;
(c) basifying the quenched reaction mass till pH of 8 - 8.5;
(d) washing the basified reaction mass till pH of 8- 8.5;
(e) adjusting the pH of aqueous layer petween 1 to 4 using acid;
(f) extracting the acidified aqueous layer using solvent mixture of hydrocarbon and
(g) isolating MiADMSA from the extract; and (h) optionally, purifying isolated MiADMSA
2. The process as claimed in claim 1, wherein esterification in step (a) is carried out at a temperature ranging from room temperature to reflux temperature of isoamyl alcohol, preferably the temperature is ranging from 110-115°C.
3. The process as claimed in claim 1, wherein basification in step (c) is carried out by using base selected from sodium bicarbonate, potassium bicarbonate, calcium bicarbonate, sodium carbonate, potassium carbonate, cesium carbonate, calcium carbonate, sodium hydroxide, potassium hydroxide, cesium hydroxide or barium hydroxide; preferably, sodium bicarbonate.
4. The process as claimed in claim 1, wherein the acid used in step (e) to adjust thepH of the reaction mass is selected from inorganic acid such as hydrochloric acid, nitric acid, sulfuric acid, p-toluene sulfonic acid and like organic acid such tartaric acid, oxalic acid, mandelic acid, fumaric acid, acetic acid or formic acid; preferably hydrochloric acid.
5. The process as claimed in claim 1, wherein hydrocarbon used in step (f) is selected from the group consisting of aliphatic hydrocarbons such as hexane, heptane or cyclohexane; aromatic hydrocarbons such as toluene or xylene; preferably toluene.
6. The process as claimed in claim 1, wherain ether used in step (f) is selected from methyl tert-butyl ether (MTBE), tetrahydrofuran (THF)- 2-methyl tetrahydrofuran, diethyl ether or diisopropyl ether (DIPE); preferably, methyl tert-butyl ether (MTBE).

7. The process as claimed in claim 1, wherein isolation in step (g) is carried out by cooling, drying, adding an anti-solvent or seeding; preferably, by drying under reduced pressure.
8. The process as claimed in claim 1, wherein the pH of aqueous layer in step (e) is adjusted within the range of 1 to 4; preferably, the pH of aqueous layer is between 2.5 to 3.
9. The process as claimed in claim 1, wherein the ratio of hydrocarbon to ether in step (f) is selected from the range of 80:20 % to 90:10 % by volume.
10. The process as claimed in claim 1, wherein the extraction is step (f) is carried out by using 83:17 % by volume mixture of toluene:MTBE and pH of aqueous layer is between 2.5 to 3.