DESC:CROSS-REFERENCE TO RELATED APPLICATIONS
This application claims the benefit of the earlier filing date of Indian Provisional Patent Application No. IN202141046771 filed on Oct 13, 2021.

BACKGROUND OF THE INVENTION

FIELD OF THE DISCLOUSRE
The present disclosure relates to a process for the preparation of liraglutide by fragment condensation in solution phase.
DESCRIPTION OF THE RELATED ART
Liraglutide developed by Novo Nordisk, glucagon-like peptide -1 (G LP-1) receptor agonist, as a subcutaneous formulation, can play a good role in lowering blood glucose. Liraglutide is marketed under the brand name Victoza in the U.S, India, Canada, Europe and Japan. The molecular formula of liraglutide is C172H265N43O51 and the molecular weight is 3751.2 Daltons. The peptide sequence of the liraglutide can be represented in terms of chemical formula (I) as follows:
His-Ala-Glu-Gly-Thr-Phe-Thr-Ser-Asp-Val-Ser-Ser-Tyr-Leu-Glu-Gly-Gln-Ala-Ala-Lys(Glu-palmitoyl)-Glu-Phe-Ile-Ala-Trp-Leu-Val-Arg-Gly-Arg-Gly-OH.
Formula (I)

Liraglutide first disclosed in US6268343B1, in which Liraglutide is prepared by solid-liquid synthetic method, the intermediate GLP-l (7-37)-OH needs reverse phase HPLC purification; and then reaction taken place with Na-hexadecanoyl-Glu (ONSu)-OtBu under liquid phase condition to get liraglutide. The deprotection of side chain leads to formation of impurities and purification is difficult due to long chain peptide.

WO2013037266 describes solid phase synthetic method by using 2-CTC resin or wang resin. According to this method, the liraglutide main chain peptide sequence is prepared by coupling N terminal Fmoc protected, and side chain protected amino acids, including lysine Fmoc-Lys (Alloc)-OH; removing Alloc protecting group of lysine side chain and coupling with Palmitoyl-Glu-OtBu; followed by deprotection and cleavage of resin to obtain crude liraglutide and further purification to get pure liraglutide. It is difficult to assemble full length peptide in an acceptable yield and purity using sequential solid phase synthesis.

Solid phase peptide synthesis (SPPS) in general shows low yields compared to liquid phase peptide synthesis (LPPS). However, conventional SPPS methods for producing large quantity of long peptides like liraglutide are time-consuming and are not able to achieve satisfying yield in large scale.

In view of all these disadvantages, there is a significant need to develop a cost effective, stable, commercially viable, large-scale process for the preparation of liraglutide with good yield. In hybrid approach, both SPPS and LPPS are employed at appropriate places, targeted peptide is assembled by fragment condensation in liquid phase whereas the fragments are generated through conventional solid phase peptide synthesis.

SUMMARY OF THE DISCLOSURE
A first aspect of the present invention provides a process for the preparation of liraglutide, which comprises:
a) synthesis of suitable fragments by Solid phase peptide synthesis SPPS and liquid phase peptide synthesis LPPS;
b) coupling of the suitable fragments in presence of coupling agents and solvent; and
c) deprotecting the peptide to get liraglutide.

In one aspect the present invention provides a process for the preparation of liraglutide, which comprises:
a) preparing Y-His(X)-Ala-Glu(X)-Gly-Thr-Phe-Thr(X)-Ser(X)-Asp(X)-Val-Ser-Ser(X)-Tyr(X)-Leu-Glu(X)-Gly-OH (Fragment-I), Y-Gln(X)-Ala-Ala-Lys(Palmitoyl-Glu-OtBu)-Glu(X)-Phe-Ile-Ala-Trp(X)-Leu-Val-Arg(Z)-Gly-OH (Fragment-II) by solid phase synthesis and H-Arg(Z)-Gly-OtBu (Fragment-III) in a solvent;
b) condensing Fragment-II and Fragment-III in presence of coupling agents and solvent followed by deprotection to get H-Gln(Trt)-Ala-Ala-Lys(Palmitoyl-Glu-OtBu)-Glu(X)-Phe-Ile-Ala-Trp(X)-Leu-Val-Arg(Z)-Gly-Arg(Z)-Gly-OtBu;
c) condensing the peptide obtained in step (b) H-Gln(X)-Ala-Ala-Lys(Palmitoyl-Glu-OtBu)-Glu(X)-Phe-Ile-Ala-Trp(X)-Leu-Val-Arg(Z)-Gly-Arg(Z)-Gly-OtBu with Fragment-I in presence of coupling agents and solvent to obtain protected liraglutide; and
d) deprotecting the peptide obtained in step (c) to get Liraglutide.
wherein, Y represents amino protecting group, X represents carboxyl, phenol and alcoholic protecting group, Z represents guanidine protecting group.

Another aspect of the present invention is to provide a process for the preparation of liraglutide, which comprises:
a) preparing Boc-His(Trt)-Ala-Glu(OtBu)-Gly-Thr-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Val-Ser-Ser(tBu)-Tyr(tBu)-Leu-Glu(OtBu)-Gly-OH (Fragment-Ia), Fmoc-Gln(Trt)-Ala-Ala-Lys(Palmitoyl-Glu-OtBu)-Glu(OtBu)-Phe-Ile-Ala-Trp(Boc)-Leu-Val-Arg(Pbf)-Gly-OH (Fragment-IIa) by solid phase synthesis and H-Arg(Pbf)-Gly-OtBu (Fragment-IIIa) in a solvent;
b) condensing Fragment-IIa and Fragment-IIIa in presence of coupling agents and solvent followed by deprotection to get H-Gln(Trt)-Ala-Ala-Lys(Palmitoyl-Glu-OtBu)-Glu(OtBu)-Phe-Ile-Ala-Trp(Boc)-Leu-Val-Arg(Pbf)-Gly-Arg(Pbf)-Gly-OtBu;
c) condensing the peptide obtained in step (b) H-Gln(Trt)-Ala-Ala-Lys(Palmitoyl-Glu-OtBu)-Glu(OtBu)-Phe-Ile-Ala-Trp(Boc)-Leu-Val-Arg(Pbf)-Gly-Arg(Pbf)-Gly-OtBu with Fragment-Ia in presence of coupling agents and solvent to obtain protected liraglutide; and
d) deprotecting the peptide obtained in step (c) to get Liraglutide.

BRIEF DESCRIPTION OF ABBREVIATIONS
CTC - Chlorotrityl chloride
DCM – Dichloromethane
DIEA - N,N-Diisopropylethylamine
DIPE - Diisopropyl ether
DMAC – Dimethylacetamide
DMF - N,N-dimethylformamide
DMS - Dimethyl sulfide
DMT - dimethoxy trityl
DTT – Diothreitol
MeOH – Methanol
MMT - Methoxytrityl
MTBE - Methyl tert-butyl ether
NMP - N-Methyl pyrrolidine
TFA - Trifluoro acetic acid
THF - Tetrahydrofuran
TIPS - Triisopropyl silane
TIS - Triisopropyl silane
Trt - Trityl
Boc - tert-butoxycarbonyl
Cbz - benzyloxycarbonyl
Bpoc - 2-(4-biphenyl)-2-propyloxycarbonyl
Fmoc - 9-fluorenylmethoxycarbonyl
Palmitoyl - hexadecanoyl
Pbf - pentamethyldihydrobenzofurane-5-sulfonyl
Pmc - 2,2,5,7,8-pentamethylchroman-6-sulfonyl

DETAILED DESCRIPTION OF THE DISCLOSURE
It is to be understood that the description of the present invention has been simplified to illustrate elements that are relevant for a clear understanding of the invention, while eliminating, for purposes of clarity, other elements that may be well known.

The present invention provides an improved process for the preparation of Liraglutide by a hybrid approach.

The present invention relates to a process for the preparation of Liraglutide by fragment condensation in solution phase whereas the fragments are generated through the conventional solid phase synthesis and liquid phase synthesis. Both solid phase synthesis and liquid phase synthesis are employed at appropriate places.

One embodiment of the present invention is to provide a process for the preparation of liraglutide, which comprises:
a) synthesis of suitable fragments by Solid phase peptide synthesis SPPS and liquid phase peptide synthesis LPPS;
b) coupling of the suitable fragments in presence of coupling agents and solvent; and
c) deprotecting the peptide to get liraglutide.

According to the present invention, Fragment-I, Fragment-II are prepared by solid phase synthesis and Fragment-III is prepared in a solvent. These fragments are coupled in presence of coupling reagents, solvent to get protected liraglutide and deprotected to get liraglutide.

Within the context of the present invention Fragment-I, Fragment-II and Fragment-III are as represented below:

Fragment-I: Y-His(X)-Ala-Glu(X)-Gly-Thr-Phe-Thr(X)-Ser(X)-Asp(X)-Val-Ser-Ser(X)-Tyr(X)-Leu-Glu(X)-Gly-OH

Fragment-II: Y-Gln(X)-Ala-Ala-Lys(Palmitoyl-Glu-OtBu)-Glu(X)-Phe-Ile-Ala-Trp(X)-Leu-Val-Arg(Z)-Gly-OH

Fragment III: H-Arg(Z)-Gly-OtBu

In other embodiment the present invention is to provide a process for the preparation of liraglutide, which comprises:
a) preparing Y-His(X)-Ala-Glu(X)-Gly-Thr-Phe-Thr(X)-Ser(X)-Asp(X)-Val-Ser-Ser(X)-Tyr(X)-Leu-Glu(X)-Gly-OH (Fragment-I), Y-Gln(X)-Ala-Ala-Lys (Palmitoyl-Glu-OtBu)-Glu(X)-Phe-Ile-Ala-Trp(X)-Leu-Val-Arg(Z)-Gly-OH (Fragment-II) by solid phase synthesis and H-Arg(Z)-Gly-OtBu (Fragment-III) in a solvent;
b) condensing Fragment-II and Fragment-III in presence of coupling agents and solvent followed by deprotection to get H-Gln(Trt)-Ala-Ala-Lys(Palmitoyl-Glu-OtBu)-Glu(X)-Phe-Ile-Ala-Trp(X)-Leu-Val-Arg(Z)-Gly-Arg(Z)-Gly-OtBu;
c) condensing the peptide obtained in step (b) H-Gln(X)-Ala-Ala-Lys(Palmitoyl-Glu-OtBu)-Glu(X)-Phe-Ile-Ala-Trp(X)-Leu-Val-Arg(Z)-Gly-Arg(Z)-Gly-OtBu with Fragment-I in presence of coupling agents and solvent to obtain protected liraglutide; and
d) deprotecting the peptide obtained in step (c) to get Liraglutide.
wherein, Y represents amino protecting group, X represents carboxyl, phenol and alcoholic protecting group, Z represents guanidine protecting group.

Within the context of the present invention, the amino protecting groups are selected from but not limited to a group comprising of Fmoc, Boc, Cbz, Bpoc, and the like.
Within the context of the present invention, the carboxyl, phenolic and alcoholic protecting groups are selected from but not limited to a group comprising of DMT, MMT, Trt, tert-butyl, t-butoxy carbonyl, and the like.

Within the context of the present invention, the guanidine protecting groups are selected from but not limited to a group comprising of Pbf and Pmc.

Another embodiment of present invention provides a process for the preparation of liraglutide, which comprises:
a) preparing Boc-His(Trt)-Ala-Glu(OtBu)-Gly-Thr-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Val-Ser-Ser(tBu)-Tyr(tBu)-Leu-Glu(OtBu)-Gly-OH (Fragment-Ia), Fmoc-Gln(Trt)-Ala-Ala-Lys(Palmitoyl-Glu-OtBu)-Glu(OtBu)-Phe-Ile-Ala-Trp(Boc)-Leu-Val-Arg(Pbf)-Gly-OH (Fragment-IIa) by solid phase synthesis and H-Arg(Pbf)-Gly-OtBu (Fragment-IIIa) in a solvent;
b) condensing Fragment-IIa and Fragment-IIIa in presence of coupling agents and solvent followed by deprotection to get H-Gln(Trt)-Ala-Ala-Lys(Palmitoyl-Glu-OtBu)-Glu(OtBu)-Phe-Ile-Ala-Trp(Boc)-Leu -Val-Arg(Pbf)-Gly-Arg(Pbf)-Gly-OtBu;
c) condensing the peptide obtained in step (b) H-Gln(Trt)-Ala-Ala-Lys(Palmitoyl-Glu-OtBu)-Glu(OtBu)-Phe-Ile-Ala-Trp(Boc)-Leu-Val-Arg(Pbf)-Gly-Arg(Pbf)-Gly-OtBu with Fragment-Ia in presence of coupling agents and solvent to obtain protected liraglutide; and
d) deprotecting the peptide obtained in step (c) to get Liraglutide.

Within the context of the present invention, the coupling agents are selected from the group comprising of hydroxybenzotriazole (HOBt); 1-Hydroxy-7-azabenzotriazole (HOAt), O-(7-azabenzotriazol-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate (HATU), 1,3-dicyclohexylcarbodiimide (DCC), 1-(dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC HCl), diisopropylcarbodiimide (DIC), isopropylchloroformate (IPCF), O-(benzotriazol-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate (HBTU) or mixtures thereof.

Within the context of the present invention, the solvents employed is selected from the group comprising of dimethylformamide (DMF), dichloromethane (DCM), tetrahydrofuran (THF), N-Methyl-2-pyrrolidone (NMP), dimethylacetamide (DMAC), methanol, ethanol, isopropanol, dichloroethane, 1,4-dioxane, 2-methyl tetrahydrofuran ethyl acetate, acetonitrile, acetone, and the like or a mixture of the listed solvents.

Within the context of the present invention, in step (b) amino protecting group is deprotected using a base. The base is organic or inorganic base. The inorganic base is selected from the group comprising of potassium carbonate, lithium carbonate, sodium carbonate, sodium ethoxide, sodium bicarbonate, potassium bicarbonate, sodium hydroxide, potassium hydroxide, ammonium hydroxide, and mixtures thereof; the organic base is selected from the group comprising of diisopropylamine, N,N-diisopropylethylamine triethylamine, dimethylamine, trimethyl amine, isopropyl ethylamine, tert butylamine, pyridine, N-methyl morpholine and mixtures thereof.

Within the context of the present invention, the protected liraglutide is de-protected with a mixture of reagents selected from the group comprising of trifluoroacetic acid (TFA), tri-isopropylsilane (TIS), tri-isopropylsilyl ether (TIPS), dithiothreitol (DTT), ethane-1,2-dithiol (EDT), thioanisole, ammonium iodide, 2,2'-(ethylenedioxy)diethane and acetyl cystein, DMS, phenol, cresol and thiocresol.

Within the context of the present invention, the preparation of liraglutide is described in the following scheme-I:

Scheme-I

Within the context of the present invention, Fragment-Ia is prepared by solid phase synthesis as the process described in the scheme-II:

Scheme-II

Within the context of the present invention, Fragment-IIa is prepared by solid phase synthesis as the process described in the scheme-III:

Scheme-III
Within the context of the present invention, Fragment-IIIa is prepared by liquid phase synthesis as the process described in the scheme-IV:

Scheme-IV
In view of the above description and the examples below, one of ordinary skill in the art will be able to practice the invention as claimed without undue experimentation. The foregoing will be better understood with reference to the following examples that detail certain procedures for the preparation of molecules, compositions and Formulations according to the present invention. All references made to these examples are for the purposes of illustration. The following examples should not be considered exhaustive, but merely illustrative of only a few of the many aspects and embodiments contemplated by the present disclosure.

EXAMPLES
Example 1: Preparation of Fragment Ia:

2- Chlorotrityl chloride resin (50 g) was charged to a peptide reactor and swelled with DCM for 1 hour. After draining DCM solvent, the resin was washed with DMF. A precooled solution of Fmoc-Gly-OH (10.4 g) and DIEA (34.8 mL) in DMF was added to the resin containing fresh lot of DCM. The mixture was stirred under nitrogen for 3-4 hours at temperature. 25-30°C. Then, remaining active sites are end-capped with a with a mixture of DMF/DIEA/MeOH (85:5:10) and the resin was washed with DMF, DCM. Next, the resin was treated with precooled solution of 20% piperidine in DMF (twice) to deprotect the Fmoc- group and then the resin was washed with DMF (2 ×500 mL) followed by DCM (2 ×500 mL). Next amino acid in the sequence, Fmoc-Glu(OtBu)-OH (51 g) was coupled to the resin in presence of oxyma (17.0 g) and di-isopropylcarbodimide (21.7 mL), and DMF (500 mL) solvent at temp. 25-30°C. The progress of coupling reaction was monitored by Kaiser test. After completion of the reaction, the resin was washed with DMF (3 × 500 mL). Subsequent, Fmoc-deprotections are carried out with 20% piperidine in DMF and sequential coupling of amino acids Fmoc-Leu-OH, Fmoc-Tyr(tBu)-OH, Fmoc-Ser(tBu)-OH, Fmoc-Val-Ser (psi Me, Me Pro)-OH, Fmoc-Asp(OtBu)-OH, Fmoc-Ser(tBu)-OH, Fmoc-Thr(tBu)-OH, Fmoc-Phe-OH, Fmoc-Gly-Thr(psi Me, Me Pro)-OH, Fmoc-Ala-OH, Boc-His(Trt)-OH are carried out with the Oxyma, di-isopropylcarbodimide, and DMF as a solvent at temp. 25-30°C. After completion of synthesis, the resin was washed with DMF (3×500 mL), DCM (2×500 mL) followed by MTBE (2 ×500 mL) and it was dried under vacuum for 12 h at temparature.35-40°C to get resin containing peptide (112 g). The protected Segment 1 selectively cleaved from CTC resin using 1% TFA-DCM (4 ×500 mL). The DCM containing product was neutralized with DIEA, evaporated under vacuum and isolated with water to provide white solid. Further, the solid was made slurry with DIPE and dried under reduced pressure at 35-40°C to give 60 g of Fragment Ia.
Weight: 60 g

Example 2: Preparation of Fragment IIa:

2- Chlorotrityl chloride resin (50 g) was charged to a peptide reactor and swelled with DCM for 1 h. After draining DCM solvent, the resin was washed with DMF. A precooled solution of Fmoc-Gly-OH (10.4 g) and DIEA (34.8 mL) in DMF was added to the resin containing fresh lot of DCM. The mixture was stirred under nitrogen for 3-4 hours at temperature 25-30°C. Then, remaining active sites are end-capped with a with a mixture of DMF/DIEA/MeOH (85:5:10) and the resin was washed with DMF, DCM. Next, the resin was treated with precooled solution of 20% piperidine in DMF (twice) to deprotect the Fmoc- group and then the resin was washed with DMF (2 ×500 mL) followed by DCM (2 ×500 mL). Next amino acid in the sequence, Fmoc-Arg(Pbf)-OH (77.8 g) was attached to the resin in presence of oxyma (17.0 g) and di-isopropyl carbodimide (21.7 mL), and DMF (500 mL) solvent at temp. 25-30°C. The progress of coupling reaction was monitored by Kaiser test. After completion of the reaction, the resin was washed with DMF (3 ×500 mL). Subsequent, Fmoc-deprotections are carried out with 20% piperidine in DMF and sequential coupling of amino acids Fmoc-Val-OH, Fmoc-Leu-OH, Fmoc-Trp(Boc)-OH, Fmoc-Ala-OH, Fmoc-Ile-OH, Fmoc-Phe-OH, Fmoc-Glu(OtBu)-OH, Fmoc-Lys(Palmitoyl-Glu-OtBu)-OH, Fmoc-Ala-OH. Fmoc-Ala-OH, Fmoc-Gln(Trt)-OH are carried out with the Oxyma, di-isopropylcarbodimide, and DMF as a solvent at temp. 25-30°C. After completion of synthesis, the resin was washed with DMF (3×500 mL), DCM (2×500 mL) followed by MTBE (2×500 mL) and it was dried under vacuum for 12 h at temparature.35-40°C to get resin containing peptide (119 g). The protected Segment 2 selectively cleaved from CTC resin using 1% TFA-DCM (4 ×1000 mL). The DCM containing product was neutralized with DIEA, evaporated under vacuum and isolated with water to provide white solid. The solid was further dried under reduced pressure at 35-40°C to give 64.8 g of Fragment IIa.
Weight: 64.8 g

Example 3: Preparation of Fragment IIIa:

Step 1 (Protected Fragment IIIa); A solution of Fmoc-Arg(Pbf))-OH (100 g), H-Gly-OtBu.HCl (28.4 g) and HOBt (20.8 g) in DCM (1200 mL) was cooled to 5-10°C. Coupling reagent HBTU (64.2 g) was added to the above reaction mixture. After maintenance of 5 minutes, DIEA (56.3 mL) was added dropwise at 5-10°C and the reaction was stirred for 4 hours. at room temperature. Reaction mixture was quenched with aq. NaHCO3 solution. The DCM layer was separated and washed with water and brine. Evaporate the DCM layer and the product precipitated with DIPE. Filter the solid and washed with DIPE. Dry the product under vacuum at 35-40°C to get 101 g of Protected Fragment IIIa.
Weight: 101 g

Step 2 (Fragment IIIa); To a cooled solution of protected segment 3 (100 g) in DMF (500 mL),
t-BuNH2 (20.8 mL) was added, and the reaction was stirred for 4 hours. After completion, the reaction mixture was quenched with aq. KHSO4 solution. The aqueous layer washed with DIPE
and the organic extracts were discarded. The pH of aqueous layer was adjusted 7 to 8 using 5% aqueous NaHCO3 solution and extracted with ethyl acetate (3 x 600 mL). The ethyl acetate layer was washed with water, brine. The solvent was removed under reduces pressure, and the residue was co-distilled with under reduced pressure and the product was precipitated with ethyl acetate and n-heptane. Filter the solid and washed with n-heptane. Dry the product under vacuum at 35-40°C to get Fragment IIIa.
Weight: 44 g

Example 4: Preparation of Fmoc-AA17-31-OtBu:

To a suspension of Fragment IIa (Fmoc-AA17-29-OH) (5 g) in NMP-THF solvent, HOAt (366 mg) followed by 1.16 g of Fragment IIIa (H-AA30-31-OtBu) were added. The solution was cooled to 0 to 5°C and maintained for 5-10 minutes. Next, EDC.HC1 (515 mg) was added at 0 to 5°C, and the reaction was stirred for 12 hours. After the completion of reaction, it was quenched with 2.5% aqueous NaHCO3 solution at 0 to 5°C. The precipitated solid was filtered and washed with water and DIPE. The solid was dried under reduced pressure at 40°C to furnish 4.5 g of Fmoc-AA17-31-OtBu.
Weight: 4.5 g

Example 5: Preparation of H-AA17-31-OtBu:

A suspension of Fmoc-AA17-31-OtBu (4 g) in NMP-THF solvent, was cooled to 0 to 5°C and treated with tert. Butylamine (1.3 mL). Further, the reaction was stirred for 12 hours. After the completion of reaction, it was quenched with aqueous KHSO4 solution at 0 to 5°C. The precipitated solid was filtered and washed with water and MTBE. The solid was dried under reduced pressure at 40°C to furnish 3.5 g of H-AA17-31-OtBu.
Weight: 3.5 g

Example 6: Preparation of Protected Liraglutide (Boc-AA1-31-OtBu):
To a suspension of H-AA17-31 (3 g) and Fragment Ia (Boc-AA1-16-OH) (2.5 g) in NMP-THF solvent, HOAt (198 mg) was added. The solution was cooled to 0 to 5°C and DIEA was added dropwise at 0 to 5°C. Next, EDC.HC1 (277.9 mg) was added at 0 to 5°C, and the reaction was stirred for 12 hours. After the completion of reaction, it was quenched with 2.5% aqueous NaHCO3 solution at 0 to 5°C. The precipitated solid was filtered and washed with aqueous KHSO4, water and DIPE. The solid was dried under reduced pressure at 40°C to furnish 4.5 g of Fmoc-AA11-31-OtBu.
Weight: 4.5 g

Example 7: Preparation of Liraglutide Crude:
Protected Liraglutide (2.0 g) was treated with a mixture of trifluoroacetic acid (16.5 mL), tri-isopropylsilane (TIPS, 0.5 mL), DTT (0.5 g), Phenol (0.5 mL), Thioanisole (1 mL) and water (1 mL), for 1 hours at 0 to 5°C. The reaction mixture allowed to warm to 25 to 30°C, and stirring was continued for 4 hours at 25-30°C. Next, the reaction was cooled to 0 to 5°C, and quenched with precooled MTBE at 0-5°C, and the suspension was stirred for 1 hour at 0 to 5°C. The solid was filtered and washed with MTBE and dried under reduced pressure at 40°C to furnish 1.2 g crude Liraglutide with purity of ~64% by HPLC.
Weight: 1.2 g

,CLAIMS:We Claim:
1. A process for the preparation of liraglutide, which comprises:
a) preparing Y-His(X)-Ala-Glu(X)-Gly-Thr-Phe-Thr(X)-Ser(X)-Asp(X)-Val-Ser-Ser(X)-Tyr(X)-Leu-Glu(X)-Gly-OH (Fragment-I), Y-Gln(X)-Ala-Ala-Lys(Palmitoyl-Glu-OtBu)-Glu(X)-Phe-Ile-Ala-Trp(X)-Leu-Val-Arg(Z)-Gly-OH (Fragment-II) by solid phase synthesis and H-Arg(Z)-Gly-OtBu (Fragment-III) in a solvent;
b) condensing Fragment-II and Fragment-III in presence of coupling agents and solvent followed by deprotection to get H-Gln(Trt)-Ala-Ala-Lys(Palmitoyl-Glu-OtBu)-Glu(X)-Phe-Ile-Ala-Trp(X)-Leu-Val-Arg(Z)-Gly-Arg(Z)-Gly-OtBu;
c) condensing the peptide obtained in step (b) H-Gln(X)-Ala-Ala-Lys(Palmitoyl-Glu-OtBu)-Glu(X)-Phe-Ile-Ala-Trp(X)-Leu-Val-Arg(Z)-Gly-Arg(Z)-Gly-OtBu with Fragment-I in presence of coupling agents and solvent to obtain protected liraglutide; and
d) deprotecting the peptide obtained in step (c) to get Liraglutide.

2. The process as claimed in claim 1, wherein, Y represents amino protecting group, X represents carboxyl, phenol and alcoholic protecting group, Z represents guanidine protecting group.
3. The process as claimed in claim 2, wherein the amino protecting groups are selected from to a group comprising of Fmoc, Boc, Cbz, Bpoc, and the like; wherein the carboxyl, phenolic and alcoholic protecting groups are selected from to a group comprising of DMT, MMT, Trt, tert-butyl, t-butoxy carbonyl, and the like; and the guanidine protecting groups are selected from to a group comprising of Pbf and Pmc.

4. A process for the preparation of liraglutide, which comprises:
a) preparing Boc-His(Trt)-Ala-Glu(OtBu)-Gly-Thr-Phe-Thr(tBu)-Ser(tBu)-Asp(OtBu)-Val-Ser-Ser(tBu)-Tyr(tBu)-Leu-Glu(OtBu)-Gly-OH (Fragment-Ia), Fmoc-Gln(Trt)-Ala-Ala-Lys(Palmitoyl-Glu-OtBu)-Glu(OtBu)-Phe-Ile-Ala-Trp(Boc)-Leu-Val-Arg(Pbf)-Gly-OH (Fragment-IIa) by solid phase synthesis and H-Arg(Pbf)-Gly-OtBu (Fragment-IIIa) in a solvent;
b) condensing Fragment-IIa and Fragment-IIIa in presence of coupling agents and solvent followed by deprotection to get H-Gln(Trt)-Ala-Ala-Lys(Palmitoyl-Glu-OtBu)-Glu(OtBu)-Phe-Ile-Ala-Trp(Boc)-Leu-Val-Arg(Pbf)-Gly-Arg(Pbf)-Gly-OtBu;
c) condensing the peptide obtained in step (b) H-Gln(Trt)-Ala-Ala-Lys(Palmitoyl-Glu-OtBu)-Glu(OtBu)-Phe-Ile-Ala-Trp(Boc)-Leu-Val-Arg(Pbf)-Gly-Arg(Pbf)-Gly-OtBu with Fragment-Ia in presence of coupling agents and solvent to obtain protected liraglutide; and
d) deprotecting the peptide obtained in step (c) to get Liraglutide.

5. The process as claimed in claim 1 and 4, wherein the coupling agents are selected from the group comprising of hydroxybenzotriazole (HOBt); 1-Hydroxy-7-azabenzotriazole (HOAt), O-(7-azabenzotriazol-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate (HATU), 1,3-dicyclohexylcarbodiimide (DCC), 1-(dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC HCl), diisopropylcarbodiimide (DIC), isopropylchloroformate (IPCF), O-(benzotriazol-1-yl)-1,1,3,3-tetramethyluronium hexafluorophosphate (HBTU) or mixtures thereof.

6. The process as claimed in claim 1 and 4, wherein the solvents employed is selected from the group comprising of dimethylformamide (DMF), dichloromethane (DCM), tetrahydrofuran (THF), N-Methyl-2-pyrrolidone (NMP), dimethylacetamide (DMAC), methanol, ethanol, isopropanol, dichloroethane, 1,4-dioxane, 2-methyl tetrahydrofuran ethyl acetate, acetonitrile, acetone, and the like or a mixture of the listed solvents.

7. The process as claimed in claim 1 and 4, wherein, the protected liraglutide is de-protected with a mixture of reagents selected from the group comprising of trifluoroacetic acid (TFA), tri-isopropylsilane (TIS), tri-isopropylsilyl ether (TIPS), dithiothreitol (DTT), ethane-1,2-dithiol (EDT), thioanisole, ammonium iodide, 2,2'-(ethylenedioxy)diethane and acetyl cystein, DMS, phenol, cresol and thiocresol.