FIELD OF THE INVENTION
The present invention relates to a novel process for the preparation Eluxadoline (1) and its intermediate. More particularly, the present invention relates to a process for the preparation of Eluxadoline with more than 99.0% purity. It further relates to the process for preparing of intermediates used herein.
BACKGROUND OF THE INVENTION
Eluxadoline is a mixed mu-opioid receptor agonist, which is being used for the treatment of irritable bowel syndrome with diarrhea (IBS-D) in human adults. Chemically it is known as 5-[[[(2S)-2-amino-3-(4-carbamoyl-2,6-dimethylphenyl) propanoyl]-[(lS)-l-(5-phenyl-lH-imidazol-2-yl) ethyl] amino] methyl]-2-methoxybenzoic acid. It is marketed as Viberzi in US and Truberzi in Europe.
The following patents and applications describe the synthesis of Eluxadoline (1)
US7741356 discloses the synthesis of Eluxadoline dihydrochloride salt. 5-({[2-tert-butoxycarbonyl methyl-3-(4-carbamoyl-2,6-dimethyl-phenyl)-propionyl]-[l-(4-phenyl-lH-imidazol-2-yl)-ethyl]-amino}-methyl)-2-methoxy-benzoic acid methyl ester was hydrolyzed and neutralized to obtain 5-({[2-tert-Butoxycarbonyl amino-3-(4-carbamoyl-2,6-dimethyl-phenyl)-propionyl]-[l-(4-phenyl-lH-imidazol-2-yl)-ethyl]-amino}-methyl)-2-methoxy-benzoic acid, which was deprotected and isolated as Eluxadoline dihydrochloride salt. The process is lengthy and time consuming, making it unsuitable for industrial use, more over does not give information about the free base Eluxadoline (1).

WO2017208156 discloses the preparation of intermediate (S)-benzyl l-oxo-l-(2-oxo-2-phenylethylamino) propan-2-ylcarbamate, by coupling (S)-2-(benzyloxycarbonylamino) propanoic acid with 2-amino-l-phenylethanone.
0 o
The above processes do not clearly mention the yield and purity of the synthesized Eluxadoline free base. Hence, the present inventors hereby report a novel process for the preparation of Eluxadoline free base of formula (1) and its intermediates with desired purity and yield.
OBJECTIVE OF THE INVENTION
In one object, the invention provides novel process for the preparation Eluxadoline (1) and its intermediates thereof.
In another embodiment, the present invention provides process for the preparation of Eluxadoline with purity greater than 99.0%.
In another object of the present invention provides a process for the preparation of intermediates used for the preparation of Eluxadoline (1).
In another object the present invention provides novel intermediates for the preparation of Eluxadoline (1), as represented by following structural formulas

SUMMARY OF THE INVENTION
Accordingly, the present invention provides a novel process for the preparation Eluxadoline (1) and its intermediates as shown in scheme-1 and scheme-2.

In one embodiment, the present invention provides a novel process for the preparation of Eluxadoline (1) comprises the steps:
a) reductive amination of benzyl-5-formyl-2-methoxybenzoate (12) with (S)-methyl-2-amino propanoate (11) to obtain compound of (R)-benzyl 2-methoxy-5-((l-methoxy-l-oxopropan-2-ylamino) methyl) benzoate (10);
b) coupling of (R)-benzyl 2-methoxy-5-((l-methoxy-l-oxopropan-2-ylamino) methyl) benzoate (10) with (S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethyl phenyl) propanoic acid (9) to obtain benzyl 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl)-N-((S)-l-methoxy-1 -oxopropan-2-yl) propanamido) methyl)-2-methoxybenzoate
(8);
c) hydrolysis of the methyl ester of 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl)-N-((S)-1 -methoxy-1 -oxopropan-2-yl) propanamido) methyl)-2-methoxybenzoate (8) to obtain ((S)-2-((S)-N-(3-(benzyloxycarbonyl)-4-methoxybenzyl)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6 dimethyl phenyl) propanamido) propanoic acid (7);
d) esterification of intermediate (7) to benzyl 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl)-N-((S)-l-oxo-1-(2-oxo-2-phenylethoxy) propan-2-yl) propanamido) methyl)-2-methoxybenzoate (5);
e) cyclization of benzyl 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl)-N-((S)-1 -oxo-1 -(2-oxo-2-phenylethoxy) propan-2-yl) propanamido) methyl)-2-methoxybenzoate (5) to obtain benzyl 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl)-N-((S)-l-(4-phenyl-lH-imidazol-2-yl) ethyl) propanamido) methyl)-2-methoxybenzoate (4);
f) deprotection of benzyl group of intermediate (4) to obtain 2 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl)-N-((S)-l-(4-phenyl-lH-imidazol-2-yl) ethyl) propanamido) methyl)-2-methoxybenzoic acid (3);

g) hydrolysis of intermediate (3) to form 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-carbamoyl-2,6-dimethylphenyl)-N-((S)-l-(4-phenyl-lH-imidazol-2-yl) ethyl) propan amido) methyl)-2-methoxybenzoic acid (2);
h) deprotecting Boc group of intermediate (2) to obtain Eluxadoline dihydrochloride salt (2a) ; and
i) conversion of Eluxadoline dihydrochloride salt (2a) to Eluxadoline (1).
In another embodiment, the present invention provides Eluxadoline with purity greater than 99%.
In another embodiment the present invention provides process for the preparation of novel intermediates 4,4'-((2S,5S)-3,6-dioxo-l-((R)-l-phenylethyl)-4-((S)-l-phenylethyl) piperazine-2,5-diyl) bis(methylene)bis(3,5- dimethylbenzonitrile) (14) and (S)-2-amino-3-(4-cyano-2,6-dimethyl phenyl) propanoic acid (13) used in the preparation of intermediate (S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl) propanoic acid (9) which can be used in the preparation of Eluxadoline (1).
NCXA AH. (^
(S^-amino-S-^-cyano-I.fi-dimcthyl V-KZS.SSJ-S^-dioxo-HM-l-phcnylcthyl)-
phcnyI)propanoic acid 4-((.S)-l-phcnylcthyl)pipcrazine-2,5-diyl)bis
(mcthylcne)bis(3,5-dimcthyIbenzonitrilc)
} (14)
Yet, in another embodiment, the present invention provides process for the preparation novel intermediate as described in scheme 2, which comprises of the following steps:
i. alkylation of (S)-phenylethylamine (19) with chloroacetyl chloride (18) to obtain 1-((R)-1 -phenylethyl)-4-((S)-l-phenylethyl) piperazine-2,5-dione
(IV);

ii. cyclization of l-((R)-l-phenylethyl)-4-((S)-l-phenylethyl) piperazine-2,5-
dione (17) to l-((R)-l-phenylethyl)-4-((S)-l-phenylethyl) piperazine-2,5-
dione (16) in presence of base and phase transfer catalyst;
iii. alkylation of l-((R)-l-phenylethyl)-4-((S)-l-phenylethyl) piperazine-2,5-
dione (16) in presence of base to form novel intermediate 4,4'-((2S,5S)-3,6-
dioxo-l-((R)-l -phenyl ethyl)-4-((S)-l-phenylethyl) piperazine-2,5-diyl)
bis(methylene) bis (3,5- dimethylbenzonitrile) (14);
iv. conversion of 4,4'-((2S,5S)-3,6-dioxo-l-((R)-l-phenylethyl)-4-((S)-l-
phenylethyl) piperazine-2,5-diyl) bis(methylene)bis (3,5-
dimethylbenzonitrile) (14) to obtain (S)-2-amino-3-(4-cyano-2,6-dimethyl
phenyl) propanoic acid (13) using suitable acid ; and
v. protecting the free amine group of intermediate (S)-2-amino-3-(4-cyano-
2,6-dimethyl phenyl) propanoic acid (13) using a suitable protecting agent
to obtain (S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-
dimethylphenyl) propanoic acid (9). In another embodiment, the present invention provides intermediates (13), intermediate (14) and intermediate (9) having purity greater than 99.0%.
DETAILED DESCRIPTION OF THE INVENTION
Accordingly, the first aspect of the present invention provides a novel process for the preparation of Eluxadoline (1) with purity greater than 99.0%
The preparation of Eluxadoline (1) comprises of following steps as outlined in Scheme-1.
In one embodiment, step a) proceeds by reductive amination of benzyl-5-formyl-2-methoxybenzoate (12) with (S)-methyl 2-aminopropanoate (11) in presence of a suitable hydride source and base in protic solvent. The said reaction may be carried out at a temperature range of about 10 °C to about 50 °C, more preferably at 25 °C to 30 °C, to obtain (R)-benzyl 2-methoxy-5-((l-methoxy-l-oxopropan-2-ylamino) methyl) benzoate (10).

In another embodiment, step b) involves coupling of (R)-benzyl 2-methoxy-5-((l-methoxy-l-oxopropan-2-yl amino) methyl) benzoate (10) with (S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl) propanoic acid (9) in presence of a suitable coupling agent in aprotic solvent to obtain benzyl 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethyl phenyl)-N-((S)-l-methoxy-l-oxopropan-2-yl)propanamido)methyl)-2-methoxy benzoate (8). Step b) was be carried out at a temperature of about 15 °C to about 45 °C, preferably at 25 °C to 30 °C.
In another embodiment, step c) involves hydrolysis of the methyl ester of intermediate (8) under alkaline condition in a suitable aprotic solvent to obtain (S)-2-((S)-N-(3-(benzyloxycarbonyl)-4-methoxybenzyl)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethyl phenyl) propanamido) propanoic acid (7).
In another embodiment, step d) proceeds with esterification of intermediate (7) to benzyl 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl)-N-((S)-l-oxo-l-(2-oxo-2-phenyl ethoxy) propan-2-yl) propanamido) methyl)-2-methoxybenzoate (5) by reacting with 2-bromo-l-phenylethanone (6) in presence of suitable base and aprotic solvent.
In another embodiment step e) involves cyclization of intermediate (5) to obtain benzyl 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl)-N-((S)-l-(4-phenyl-lH-imidazol 2-yl) ethyl) propanamido) methyl)-2-methoxybenzoate (4) by reacting with a suitable ammonium salt in a suitable aprotic solvent, preferably ammonium acetate was used in the present invention. The said reaction may undergo cyclization at a temperature of about 120 °C-170 °C, preferably at 140 °C-150 °C more preferably 110-120°C.
In another embodiment step f) proceeds with hydrogenation of benzyl group of intermediate (4) using suitable metal catalyst to obtain 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl)-N-((S)-l-(4-phenyl-lH-imidazol-2-yl) ethyl) propanamido) methyl)-2-methoxybenzoic acid (3)

In another embodiment, step g) involves hydrolysis of intermediate (3) with suitable base to form 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-carbamoyl-2,6-dimethylphenyl)-N-((S)-1 -(4-phenyl-1 H-imidazol-2-yl) ethyl) propanamido) methyl)-2-methoxybenzoic acid (2). The said reaction may be carried out at a temperature of about -10 °C to about 50 °C more preferably at about 0 °C to about 30 °C.
In another embodiment, step h) proceeds with the formation of dihydrochloride salt of intermediate 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-carbamoyl-2,6-dimethylphenyl)-N-((S)-1 -(4-phenyl-1 H-imidazol-2-yl) ethyl) propanamido) methyl)-2-methoxybenzoic acid (2a) by treating intermediate (2) with a suitable acid source and aprotic solvent. The said reaction can be carried out at a temperature of about -10 °C to about 45 °C, more preferably at about 5 °C to about 30 °C.
In another embodiment, step i) involves conversion of dihydrochloride salt of intermediate (2a) to Eluxadoline (1) by reacting with a suitable base in a suitable protic solvent.

(I)
Scheme-1 The hydride source used in step a) was selected from a group comprising of sodium cyanoborohydride (NaBFbCN), sodium triacetoxyborohydride (NaBH (CH3COO) 3), sodium borohydride (NaBHU), lithium cyanoborohydride, tetrabutylammonium cyanoborohydride ((CH3CH2CH2CH2)4N(BH3CN)), tetramethylammonium triacetoxyborohydride (NMe4BH(OAc)3 or the like, preferably sodium borohydride (NaBH4) was used in the present invention.

The suitable protic solvent used in step a), step b), step c), step d), step f), step g) and step h) were selected from a group comprising of methanol, ethanol, n-propanol, isopropyl alcohol (IPA), n-propanol, n-butanol, isobutanol, t-butanol, 2-nitro ethanol, 2-fluroethanol, ethylene glycol, propylene glycol, water or the like, preferably methanol and water can be used in the present invention.
The suitable aprotic solvents used in step a) ,step b), step c) , step d), step e), step g) and step h) were selected from a group comprising of acetone, acetonitrile, 1,4-dioxane, diethyl ether, dichloromethane, ethyl acetate, N, N-dimethylformamide, methyl tertiary butyl ether , heptane, hexane, cyclohexane, toluene, methylcyclohexane, o- xylene m- xylene, p-xylene, , tetrahydrofuran or the like, preferably ethyl acetate, N, N-dimethylformamide, xylene , toulene and tetrahydrofuran were used in the present invention.
The base used herein in step a), step b), step c), step d) and step g) were selected
from a group of alkali metal hydroxides comprising of lithium hydroxide, sodium
hydroxide, potassium hydroxide or the like. Alkali metal carbonates comprising of
sodium carbonate, potassium carbonate, lithium carbonate, cesium carbonate or the
like. Alkali metal bicarbonates comprising of sodium bicarbonate, potassium
bicarbonate, lithium bicarbonate or the like and "alkali metal alkoxides" comprising
of sodium methoxide, sodium ethoxide, sodium tert-butoxide, potassium methoxide,
potassium ethoxide, potassium tert-butoxide or the like. Other bases which may be
selected from a group comprising of ammonia or other organic bases such as
piperidine, pyridine, triethylamine, methyl amine, ethyl amine, dimethyl
aminopyridine (DMAP), isopropyl amine, diisopropyl amine,
diisopropylethylamine, l,8-diazabicyclo[5.4.0]undec-7-ene (DBU), 1,5-
diazabicycto[4.3.0] non-5-ene (DBN), tribenzylamine, N-
methylmorpholine(NMM) , N-ethylmorpholine, morpholine, , 2,6-lutidine, 2,4,6-collidine, imidazole, 1 -methyl imidazole, 1 ,2,4-triazole, 1 ,4-diazabicyclo[2.2.2]octane (DABCO) or mixtures thereof. Preferably sodium bicarbonate, sodium hydroxide, lithium hydroxide, cesium carbonate and potassium carbonate were used in the present invention.

The suitable acid source used in step b) and step h) were selected from a group comprising of hydrochloric acid gas, aqueous hydrochloric acid, dry hydrochloric acid, ethyl acetate hydrochloric acid, methanolic hydrochloric acid, ethanolic hydrochloric acid or the like. Preferably, aqueous hydrochloric acid and ethyl acetate hydrochloric acid were used in the present invention.
The suitable coupling agent used in step b) were selected from a group comprising
of N,N'-dicyclohexylcarbodiimide(DCC), N,N'-diisopropylcarbodiimide (DIC), 1-
Ethyl-3-(3-dimethyl aminopropyl)carbodiimide (EDC), N-(3-
Dimethylaminopropyl)-N'-ethylcarbodiimide hydrochloride (EDC1), alkyl or aryl
chloroformates such as ethylchloroformate, benzyl chloroformate,
diphenylphosphoroazidate (DPP A), (Benzotriazol-1 -yloxy)tris(dimethyl
amino)phosphonium hexafiuorophosphate methanesulfonyl chloride which were
optionally used in combination with Dimethylaminopyridine (DMAP), 1- hydroxy-
7-azatriazole (HOAt), 1-hydroxybenzotriazole (HOBt), l-hydroxy-lH-1,2,3-
triazole-4-carboxylate(HOCt), N-hydroxysuccinamide, N-hydroxysulfo
succinimide or mixtures thereof. Preferably, N-(3-dimethylaminopropyl)-N'-ethyl carbodiimide hydrochloride (EDC1) and 1 -hydroxy benzotriazole (HOBt) or N,N'-dicyclohexylcarbodiimide(DCC) and 4-Dimethylaminopyridine (DMAP), or N,N'-diisopropylcarbodiimide (DIC) and 1- hydroxy-7-azatriazole (HOBt), or N,N'-diisopropylcarbodiimide (DIC) and 1- hydroxy-7-azatriazole (HOAt); more preferably N,N'-diisopropylcarbodiimide (DIC) and 1- hydroxy-7-azatriazole (HOBt) and N,N'-diisopropylcarbodiimide (DIC) and 1- hydroxy-7-azatriazole (HOAt) were used in the present invention.
The suitable metal catalysts used in step f) was selected from a group comprising of Raney-Nickel, Platinum on carbon (Pt/C), Palladium on carbon (Pd/C), Rhodium on carbon with hydrogen gas, preferably Palladium on carbon (Pd/C) with hydrogen gas was used in the present invention
In some embodiment, the present invention provides process for the preparation intermediate (9), which comprises of the steps as illustrated in scheme 2.

Scheme-2 In an embodiment of the present invention provides a novel process of preparation of intermediate (13) and intermediate (14).
In another embodiment intermediate (13) and intermediate (14) were used to prepare the intermediate (9) as illustrated in scheme 2, which comprises of following steps: i. acylation of (S)-phenylethylamine (19) with chloroacetyl chloride (18) by reacting with a suitable base to form l-((R)-l-phenylethyl)-4-((S)-l-phenylethyl) piperazine-2,5-dione (17); ii. cyclization of l-((R)-l-phenylethyl)-4-((S)-l-phenylethyl) piperazine-2,5-dione (17) by treating with a suitable base and phase transfer catalyst to form 1 -((R)-1 -phenylethyl)-4-((S)-1 -phenyl ethyl) piperazine-2,5-dione (16); iii. alkylation of intermediate (16) with 4-(bromomethyl)-3,5-dimethylbenzonitrile (15) to obtain a novel intermediate 4,4'-((2S,5S)-3,6-dioxo-1 -((R)-1 -phenylethyl)-4-((S)-1 -phenylethyl) piperazine-2,5-diyl) bis (methylene)bis(3,5-dimethyl benzonitrile ) (14) using suitable base ; iv. conversion of intermediate (14) to obtain (S)-2-amino-3-(4-cyano-2,6-
dimethyl phenyl) propanoic acid (13) with a suitable acid; and v. protection of amino group of intermediate (13) using a suitable protecting agent to form (S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethyl phenyl) propanoic acid (9).

In one embodiment step i) proceeds acylation of (S)-phenylethylamine (19) by reacting with chloroacetyl chloride (18) using suitable base in a suitable protic or aprotic or mixture of solvents to form l-((R)-l-phenylethyl)-4-((S)-l-phenylethyl) piperazine-2,5-dione (17)
In another embodiment step ii) proceeds with cyclization of l-((R)-l-phenylethyl)-4-((S)-l-phenylethyl) piperazine-2,5-dione (17) by treating with a base and suitable phase transfer catalyst in a suitable aprotic solvent to afford 1-((R)-1-phenyl ethyl)-4-((S)-l-phenylethyl) piperazine-2,5-dione (16)
In another embodiment step iii) proceeds with alkylation of intermediate (16) upon treatment with 4-(bromomethyi)-3,5-dimethylbenzonitrile (15) using a suitable base in an aprotic solvent to afford intermediate 4,4'-((2S,5S)-3,6-dioxo-l-((R)-l-phenylethyl)-4-((S)-l-phenylethyl) piperazine -2,5-diyl) bis (methylene)bis (3,5-dimethyl benzonitrile) (14). The said reaction was earned out at a temperature of about -78 °C to about 60 °C, preferably at about -50 °C to about 50 °C, more preferably at about 0 °C to about 40 °C, still more preferably at about 20 °C to about 30 °C.
Further, in another embodiment step iv) proceeds with conversion of intermediate (14) to obtain (S)-2-amino-3-(4-cyano-2,6-ditnethyl phenyl) propanoic acid (13) by reacting with a suitable acid at a temperature of about 100 °C to about 140 °C, more preferably at about 120 °C to about 130 °C.
Yet in another embodiment, step v) proceeds with protecting the amino group of (S)-2-amino-3-(4-cyano-2,6-dimethyl phenyl) propanoic acid (13) with a suitable protecting agent in a suitable aprotic solvent to afford intermediate (9). The suitable aprotic solvent used in step i), step ii), step iii) and step v) were selected from a group comprising of acetone, acetonitrile, 1,4-dioxane, diethyl ether, dichloromethane, methyl acetate, ethyl acetate, N, N-dimethylformamide, methyl tertiary butyl ether , heptane, hexane, cyclohexane, toulene, pentane, cyclopentane, methylcyclohexane, o-xylene, m- xylene, p-xylene, furan, tetrahydrofuran or

mixture thereof, preferably acetone, ethyl acetate, dichloroethane, tetrahydrofuran, hexane, 1,4 -dioxane, diethyl ether were used in the present invention.
The suitable protic solvent used in step i), step ii), step iii) and step iv) were selected from a group comprising of methanol, ethanol, n-propanol, isopropyl alcohol (IP A), n-propanol, n-butanol, isobutanol, t-butanol, ethylene glycol, propylene glycol, phenol, benzyl alcohol, water or mixture thereof, preferably water was used in the present invention. The base used in step i), step ii) and step iii) until unless specified were selected from a group comprising of alkali metal hydroxides comprising of lithium hydroxide, sodium hydroxide,potassium hydroxide, alkali metal carbonates comprising of sodium carbonate, potassium carbonate, lithium carbonate, cesium carbonate or the like, alkali metal bicarbonates comprising of sodium bicarbonate, potassium bicarbonate, lithium bicarbonate or the like and alkali metal alkoxides such as sodium methoxide, sodium ethoxide, sodium tert-butoxide, potassium methoxide, potassium ethoxide, potassium tert-butoxide or the like. Lithium amides comprising of lithium diisopropylamide (LDA), lithium tetramethylpiperidide (LiTMP), lithium hexamethyldisilazide (LHMDS) or the like and other bases which may be selected from a group comprising of ammonia or other organic bases such as piperidine, pyridine , triethylamine, methyl amine, ethyl amine, dimethyl aminopyridine (DMAP), isopropyl amine, diisopropyl amine, diisopropylethylamine or the like were used. Preferably sodium carbonate, sodium hydroxide and lithium hexamethyldisilazide (LHMDS) were used in the present invention.
The protecting group used in step v) was selected from a group comprising of Fmoc (9-fluorenyl methoxy carbonyl), Boc (tert-butyloxycarbonyl), Cbz (Benzyloxycarbonyl), Bpoc (2-(4-biphenyl)-2-propyloxycarbonyl), 2,2,4,6,7-pentamethyldihydrobenzofuran-5-sulfonyl (Pbf) or the like. Preferably Boc (tert-butyloxycarbonyl) was used in the present invention.

Intermediate (13) and intermediate (12) so obtained by the above process were
having purity greater 99%. Another embodiment, of the present invention is to
provide alternative process for the preparation of Eluxadoline as depicted in scheme-
3 as below which comprises hydrolysis of intermediate (4) using a suitable base to
yield benzyl 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-carbamoyl-2,6-
dimethylpheny 1)-N-((S)-1 -(4-phenyl-1 H-imidazol-2-yl) ethyl) propanamido) methyl)-2-methoxybenzoate (20). Intermediate (20) was hydrogenated using a suitable metal catalyst in an aprotic solvent and acidified to form the trifluroacetic acid salt of 5-(((S)-2-amino-3-(4-carbamoyl-2,6-dimethylphenyl)-N-((S)-l-(4-phenyl-lH-imidazol-2-yl) ethyl) propanamido) methyl)-2-methoxybenzoic acid (21). Intermediate (21) was converted to dihydrochloride salt of Eluxadoline (2a) using suitable acid in an aprotic solvent. Eluxadoline dihydrochloride (2a) was neutralized to obtain Eluxadoline free base (1)
N III I B°cIIN 9 T I ° I O 'V hydrolysis 1 -M jl 1 hydrogcnntion U )—.
InocMiN Jf [ —'—: .. |ps*i/^-^-N — — ■" fiY^r N
. TA, ° „ Y" ° " XT
J ° O^O-Bn <21» (A°"
Bn
salt formation
Mc: -CII3
Bn: -CH2-C6H5 ^v
Boc: tcrt-butyloxycarboiiyl [ j WJi
ff^'<^f ^^ N neutralization ■"*"" J~~OH (2.,) cA°"
(1)
Scheme-3 Another embodiment, of the present invention is to provide alternative process for the preparation of Eluxadoline as depicted in scheme-4 as below, which comprises hydrolysis of intermediate (4) by using a suitable base to yield benzyl 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-carbamoyl-2,6-dimethylphenyl)-N-((S)-l-(4-phenyl-1H imidazol-2-yl) ethyl) propanamido) methyl)-2-methoxybenzoate (20). Intermediate (20) may be deprotected using a suitable acid dissolved in aprotic

and stirred for 24hrs at 25-30 °C. After completion of reaction, saturated sodium bicarbonate solution was added to the resulting solution and the solvent was removed by distillation. The residue so obtained was extracted with ethyl acetate and distilled off under reduced pressure to afford titled compound (R)-benzyl 2-methoxy-5-((l-methoxy-l-oxopropan-2-ylamino) methyl) benzoate (10). Yield: 77.0%, Purity: 99.0%
EXAMPLE-2: Preparation of benzyl 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl)-N-((S)-l-methoxy-l-oxopropan-2-yl) propanamido) methyI)-2-methoxy benzoate (8)
0.36g of (R)-benzyl 2-methoxy-5-((l-methoxy-l-oxopropan-2-ylamino) methyl) benzoate (10) and 0.32g of (S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl) propanoic acid (9)were dissolved in dimethyl formamide. 0.03g of 1-hydroxybenzotriazole (HOBt) and 0.29g of N-(3 dimethyl aminopropyl) -N'-ethylcarbodiimide hydrochloride (EDC1) were added to the reaction mixture and stirred for 12 hrs. at 25-30 °C. On completion of reaction, water was added to the resulting mixture and extracted with ethyl acetate. The organic phase was separated and washed with IN hydrochloric acid, saturated sodium bicarbonate solution and sodium chloride solution. The organic layers were combined, dried over sodium sulphate, filtered and concentrated under reduced pressure to yield 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl)-N-((S)-l-methoxy-l-oxo propan-2-yl) propanamido) methyl)-2-methoxy benzoate (8) .Yield: 40%, Purity: 99.62%.
EXAMPLE-3: Alternative process for the preparation of benzyl 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethyIphenyl)-N-((S)-l-methoxy-l-oxopropan-2-yl) propanamido) methyi)-2-methoxy benzoate (8) 0.36g of (R)-benzyl 2-methoxy-5-((l-methoxy-l-oxopropan-2-ylamino) methyl) benzoate (10) and 0.32g of (S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl) propanoic acid (9) were dissolved in dimethyl formamide. 0.03g of 1-hydroxybenzotriazole (HOAt) and 0.29g of N-(3 dimethyl aminopropyl) -N'-ethylcarbodiimide hydrochloride ( DIC), were added and stirred for 12 hrs. at 25-30 °C. On completion of reaction, water was added to the resulting mixture and

extracted with ethyl acetate. The organic phase was separated and washed sequentially with IN hydrochloric acid, saturated sodium bicarbonate solution and brine. The combined organic layers were dried over sodium sulphate, filtered and concentrated under reduced pressure to yield 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl)-N-((S)-l-methoxy-l-oxo propan-2-yl) propanamido) methyl)-2-methoxy benzoate (8).Yield: 65%, Purity: 99.20%
EXAMPLE-4: Preparation of ((S)-2-((S)-N-(3-(benzyloxycarbonyl)-4-methoxybenzyl)-2-(tert-butoxy carbonylamino)-3-(4-cyano-2,6 dimethyl phenyl) propanamido) propanoic acid (7)
0.66g of 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl)-
N-((S)-l-methoxy- 1-oxo propan-2-yl) propanamido) methyl)-2-methoxy benzoate
(8) was dissolved in tetrahydrofuran and cooled to 0-5 °C. To the reaction solution
added IN lithium hydroxide (20mL) for 12hrs at 25-30 °C. 3volumes of ethyl
acetate was added to the reaction mixture and the phases were separated. The pH
of aqueous layer was adjusted to 6.0-7.0 with IN hydrochloric acid and extracted
with 6 volumes of ethyl acetate. The combined organic phases were dried over
sodium sulphate and filtered off. The solvent of the filtrate was removed under
reduced pressure to furnish ((S)-2-((S)-N-(3-(benzyloxycarbonyl)-4-methoxy
benzyl)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6 dimethyl phenyl)
propanamido) propanoic acid (7) .Yield: 63.91%, Purity: 99.75% EXAMPLE-5: Preparation of benzyl 5-(((S)-2-(tert-butoxycarbonyIamino)-3-(4-cyano-2,6-dimethylphenyl)-N-((S)-l-oxo-l-(2-oxo-2-phenylethoxy) propan-2-yl) propanamido) methyl)-2-methoxybenzoate (5)
0.48g of Cesium carbonate was added to the reaction mixture of 0.64g ((S)-2-((S)-N-(3-(benzyloxy carbonyl)-4-methoxy benzyl)-2-(tert-butoxy carbonyl amino)-3-(4-cyano-2,6-dimethyl phenyl) propanamido) propanoic acid (7) dissolved in dimethylformamide. 0.24g of phenacyl bromide (6) was then added to the reaction mass and stirred for 12hrs at 25-30 °C. On completion of reaction, ice cold water was added the reaction mass and extracted with 90mL of ethyl acetate. The organic layer was separated, washed with water and distilled under vacuum to obtain 5-

(((S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl)-N-((S)-l-oxo-l-(2-oxo-2-phenylethoxy) propan-2-yl) propanamido) methyl)-2-methoxybenzoate (5).Yield: 51.08%, Purity: 99.45%
EXAMPLE-6: Alternative process for the preparation of benzyl 5-(((S)-2-
(tert-butoxycarbonyIamino)-3-(4-cyano-2,6-dimethylphenyl)-N-((S)-l-oxo-l-
(2-oxo-2-phenylethoxy) propan-2-yl) propanamido) methyl)-2-
methoxybenzoate (5)
0.48g of potassium carbonate was added to the reaction mixture of 0.64g of ((S)-2-
((S)-N-(3-(benzyloxy carbonyl)-4-methoxy benzyl)-2-(tert-
butoxycarbonylamino)-3-(4-cyano-2,6-dimethyl phenyl) propanamido) propanoic acid (7) dissolved in dimethylformamide and stirred for half an hour. 0.24g of phenacyl bromide (6) was then added to the reaction mass and stirred for 12hrs at 25-30 °C. On completion of reaction, ice cold water was added the reaction mass and extracted with 90mL of ethyl acetate. The organic layer was separated, washed with water and distilled under vacuum to obtain 5-(((S)-2-(tert-butoxycarbonylamino)-3 -(4-cyano-2,6-dimethylphenyl)-N-((S)-1 -oxo-1 -(2-oxo-2-phenylethoxy) propan-2-yl) propanamido) methyl)-2-methoxybenzoate (5).Yield: 70.5%, Purity: 99.0%.
EXAMPLE-7: Preparation of benzyl 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl)-N-((S)-l-(4-phenyl-lH-imidazoI-2-yl) ethyl) propanamido) methyl)-2-methoxybenzoate (4)
0.76g of 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl)-N-((S)-l-oxo-l-(2-oxo-2-phenylethoxy) propan-2-yl) propanamido) methyl)-2-methoxybenzoate (5) was dissolved in xylene and 1.54g of ammonium acetate was added to it at 20-30 °C. The reaction mass was heated at 150-160 °C and stirred for 12hrs. On completion of reaction, the reaction mass was cooled to 20-30 °C and diluted with water. The aqueous layer was separated and extracted with 3 volumes of ethyl acetate. The combined organic layers were dried over sodium sulphate and filtered. The filtrate was distilled under vacuum to afford benzyl 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl)-N-((S)-l-(4-phenyl-lH-

imidazol-2-yl) ethyl) propanamido) methyl)-2-methoxybenzoate (4).Yield:
40.48%, Purity:99.0%
EXAMPLE-8: Alternative process for the preparation of benzyl 5-(((S)-2-
(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl)-N-((S)-l-(4-
phenyl-lH-imidazol-2-yI) ethyl) propanamido) methyl)-2-methoxybenzoate
(4)
0.76g of 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl)-N-((S)-l-oxo-l-(2-oxo-2-phenylethoxy) propan-2-yl) propanamido) methyl)-2-methoxybenzoate (5) was dissolved intoulene and 1.54g of ammonium acetate was added to it at 20-30 °C. The reaction mass was heated at 115-120 °C and stirred for 12hrs. On completion of reaction, the reaction mass was cooled to 20-30 °C and diluted with water. The aqueous layer was separated and extracted with 3 volumes of ethyl acetate. The combined organic layers were dried over sodium sulphate and filtered. The filtrate was distilled under vacuum to afford benzyl 5-(((S)-2-(tert-butoxycarbonylamino)-3 -(4-cyano-2,6-dimethylphenyl)-N-((S)-1 -(4-phenyl-1H-imidazol-2-yl) ethyl) propanamido) methyl)-2-methoxybenzoate (4).Yield: 75.5%, Purity: 99.65%
EXAMPLE-9: Preparation of 5-(((S)-2-(tert-butoxycarbonyIamino)-3-(4-cyano-2,6-dimethyI phenyI)-N-((S)-l-(4-phenyI-lH-imidazoI-2-yl) ethyl) propanamido) methyl)-2-methoxybenzoic acid (3)
0.74g of 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl)-
N-((S)-1-(4-phenyl -lH-imidazol-2-yl) ethyl) propanamido) methyl)-2-
methoxybenzoate (4) was dissolved in methanol and 10% Pd/C was added to the
reaction mixture at 25-30 °C under hydrogen atmosphere. On completion of
reaction, the reaction solution was filtered through Hyflo and the filtrate removed
under vacuum to yield titled compound 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-
cyano-2,6-dimethylphenyl)-N-((S)-l-(4-phenyl-lH-imidazol-2-yl) ethyl)
propanamido) methyl)-2-methoxybenzoic acid (3).Yield: 46.89%, Purity:99.0%

EXAMPLE-10: Preparation of 5-(((S)-2-(tert-butoxycarbonyIamino)-3-(4-carbamoyl-2,6-dimethylphenyl)-N-((S)-l-(4-phenyI-lH-imidazol-2-yl) ethyl) propanamido) methyI)-2-methoxybenzoic acid (2)
0.65g of 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl)-N-((S)-l-(4-phenyl -lH-imidazol-2-yl) ethyl) propanamido) methyl)-2-methoxy benzoic acid (3) was dissolved in dimethyl sulfoxide and treated with sodium hydroxide (leq) at 0-5 °C. 30% hydrogen peroxide was added to the reaction mixture at 0-5 °C and stirred at 25-30 °C till the completion of reaction. Water was added to the reaction mass and extracted with ethyl acetate. The organic layer was separated and distilled under vacuum to obtain 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-carbamoyl-2,6-dimethylphenyl)-N-((S)-l-(4-phenyl-lH-imidazol-2-yl) ethyl) propanamido) methyl)-2-methoxybenzoic acid (2). Yield: 42.62%, Purity:99.44%.
EXAMPLE-11: Preparation of Eluxadoline free base (1) 0.57g of 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-carbamoyl-2,6-dimethyl phenyl)-N-((S)-l-(4-phenyl-lH-imidazol-2-yl) ethyl) propanamido) methyl)-2-methoxybenzoic acid (2) was dissolved in 2 volumes of ethyl acetate at 5-10 °C with addition of ethyl acetate hydrochloride and stirred at 25-30 °C. The reaction mass was filtered and washed with ethyl acetate to obtain dihydrochloride salt of 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-carbamoyl-2,6-dimethylphenyl)-N-((S)-l-(4-phenyl-lH-imidazol-2-yl) ethyl) propanamido) methyl)-2-methoxybenzoic acid (2a). The dihydrochloride salt (2a) so obtained was dissolved in 10 volumes of water and 40% sodium carbonate solution was added to adjust the pH between 7.0-7.5 at 25-30 °C. The solid so formed was washed with 2 volumes of water and filtered to obtain Eluxadoline free base (1). Yield: 68.86%, Purity: 99.75% EXAMPLE-12: Preparation of benzyl 5-(((S)-2-(tert-butoxycarbonyIamino)-3-(4-carbamoyI-2,6-dimethyIphenyl)-N-((S)-l-(4-phenyl-lH-imidazol-2-yl) ethyl) propanamido) methyl)-2-methoxybenzoate (20)
0.5g Benzyl 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethyl phenyl)-N-((S)-l-(4-phenyl-lH-imidazol-2-yl) ethyl) propanamido) methyl)-2-methoxybenzoate (4) was dissolved in 5mL methanol. 1M NaOH (5 mL) and 30%

H2O2 (1.35 mL) were added to the reaction mass at 25-30 °C. The reaction mass was heated to 40-45 °C . On completion of reaction, the reaction mass was quenched with ice cold water at 0-5 °C. The obtained precipitate was filtered and washed with water and dried under vacuum to yield the desired 5-(((S)-2-(tert-butoxycarbonylamino)-3 -(4-carbamoyl-2,6-dimethylphenyl) -N-((S)-1 -(4-phenyl-lH-imidazol-2-yl) ethyl) propanamido) methyl)-2-methoxybenzoate (20).Yield: 45%, Purity: 99.15%
EXAMPLE-13: Preparation of benzyl 5-(((S)-2-amino-3-(4-carbamoyl-2,6-dimethylphenyl)-N-((S)-l-(4-phenyl-lH-imidazol-2-yl) ethyl) propanamido) methyI)-2-methoxybenzoate (22)
0.5g of 5-(((S)-2-(tert-butoxycarbonylamino)-3-(4-carbamoyl-2,6-dimethyl phenyl)-N-((S)-l-(4-phenyl-lH-imidazol-2-yl) ethyl) propanamido) methyl)-2-methoxybenzoate (20) was added to 5 mL of isopropyl alcohol. To this hydrochloric acid dissolved in isopropyl alcohol was added at 0 °C and the reaction mass stirred at 25-30 °C. After completion of reaction, the volatiles were removed under vacuum and the solid mass triturated with methyl tert-butyl ether. The solid obtained was filtered and dried under vacuum to get benzyl 5-(((S)-2-amino-3-(4-carbamoyl-2,6-dimethylphenyl)-N-((S)-1 -(4-phenyl-1 H-imidazol-2-yl) ethyl) propanamido) methyl)-2-methoxybenzoate (22).Yield: 55%, Purity: 99.58% EXAMPLE-14: Alternative process for the preparation of Eluxadoline free base (1)
10% Pd/C was added to a solution of 5-(((S)-2-amino-3-(4-carbamoyl-2,6-dimethylphenyl)-N-((S)-1 -(4-phenyl-1 H-imidazol-2-yl) ethyl) propanamido) methyl)-2-methoxybenzoate (22) (0.5 g) in ethyl acetate. The resulting reaction mixture was stirred at 25-30 °C under a hydrogen atmosphere. On completion of reaction, the reaction solution was filtered through Hyflo and the filtrate removed under vacuum to yield Eluxadoline free base (1). Yield: 65%; Purity: 99.8% EXAMPLE-15: Preparation of l-((R)-l-phenylethyl)-4-((S)-l-phenylethyI) piperazine-2,5-dione (17)
15.90 mL of chloroacetyl chloride (18) in acetone was added to the reaction mixture of 25.7mL of S-l-phenylethylamine (19) and 30.Og of sodium carbonate dissolved

in water-acetone (1:1) mixture at 0 °C. On completion of reaction, the solvent was removed under pressure and the residue was acidified with 6M hydrochloric acid. The reaction mass so obtained was extracted with ethyl acetate and the solvent was removed under pressure. The obtained solid was crystallized from ethyl acetate/ether mixture to obtain l-((R)-l-phenylethyl)-4-((S)-l-phenylethyl) piperazine-2,5-dione (17). Yield: 91%, Purity: 99.01%
EXAMPLE-16: Preparation of l-((R)-l-phenylethyl)-4-((S)-l-phenylethyl) piperazine-2,5-dione (16)
3.55g of l-((R)-l-phenylethyl)-4-((S)-l-phenylethyl) piperazine-2,5-dione (17) was dissolved in dichloromethane and 50% of 11.5mL of sodium hydroxide solution was added at 25-30 °C. Benzyltriethylammonium chloride (TEBA) was then added to the reaction mixture and stirred for 48 hrs. On completion of reaction, water and 6M hydrochloric acid were added to the reaction mixture. The dichloroethane was removed under vacuum and aqueous layer was extracted with ethyl acetate. The combined organic layers were washed with sodium chloride solution, dried over magnesium sulphate and filtered. The filtrate was distilled under vacuum to afford l-((R)-l-phenylethyl)-4-((S)-l-phenylethyl) piperazine-2,5-dione (16). Yield: 85%, Purity: 99.46%
EXAMPLE-17: Preparation of 4,4'-((2S,5S)-3,6-dioxo-l-((R)-l-phenylethyi)-4-((S)-l-phenylethyI) piperazine-2,5-diyl) bis (methylene) bis (3,5-dimethylbenzonitrile) (14)
12mL of Lithium bis(trimethylsilyl)amide solution (LHMDS) was added to the reaction solution of 1.64g of l-((R)-l-phenylethyl)-4-((S)-l-phenylethyl) piperazine-2,5-dione (16) in dry tetrahydrofuran at -78 °C under nitrogen atmosphere and stirred for 90 mins. 2.24g of 4-(bromomethyl)-3,5-dimethylbenzonitrile (15) was dissolved in dry tetrahydrofuran and added to the above reaction mass. The temperature of the reaction mixture was raised to 25-30 °C , water was added and extracted with ethyl acetate. The organic layers were combined, washed with sodium chloride solution and dried over anhydrous sodium sulphate and filtered. The filtrate was dried under vacuum to obtain 4,4'-((2S,5S)-

3,6-dioxo-1 -((R)-1 -phenyl ethyl)-4-((S)-1 -phenylethyl) piperazine-2,5-diyl) bis (methylene) bis (3,5- dimethylbenzonitrile) (14).Yield: 75%, Purity:99.25%. EXAMPLE-18: Preparation of (S)-2-amino-3-(4-cyano-2,6-dimethyl phenyl) propanoic acid (13)
0.68g of 4,4'-((2S,5 S)-3,6-dioxo-1 -((R)-1 -phenylethyl)-4-((S)-1 -phenylethyl) piperazine-2,5-diyl) bis (methylene) bis (3,5- dimethylbenzonitrile) (14) was refluxed with lOmL of 57% hydroiodic acid for 3hrs.The solvent was removed under vacuum and 23mL aqueous sodium bicarbonate was added to the crude. The reaction mass was cooled to 0°C to yield (S)-2-amino-3-(4-cyano-2,6-dimethyl phenyl) propanoic acid (13) which was used directly in the next step without isolation.
EXAMPLE-19: Preparation of (S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl) propanoic acid (9)
0.55g of di-tert-butyl decarbonate ((Boc)20) was added to a solution of (S)-2-amino-3-(4-cyano-2,6-dimethyl phenyl) propanoic acid (13) in 1,4 -dioxane at 0 °C and stirred for 12hrs. The temperature of the reaction mixture was raised to 25-30 °C and water was added. The aqueous layer was extracted with diethyl ether. The total aqueous layer was combined, and the pH adjusted to 2 with aqueous citric acid and extracted with ethyl acetate. The combined organic layers were washed with brine solution and dried over sodium sulphate and filtered. The filtrate was removed under vacuum to obtain (S)-2-(tert-butoxycarbonylamino)-3-(4-cyano-2,6-dimethylphenyl) propanoic acid (9). Yield: 80%, Purity:99.1%

We claim:
1. A process for preparing Eluxadoline (1), comprising:
O )—CH3
7-N ,=^ CH3
H3C /—( H /Vo
)=( NH2 v—