16-[(2-methylpropan-2-yl)oxy]-16-oxohexadecanoic acid

Conditions: temp:0.00; pressure:0.00; stir:

Conditions: temp:50.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: To a solution of hexadecandioic acid mono t-butyl ester (100 mg, 0.29 mmol) in dry DCM (2 ml), HOAt (44 mg, 0.29 mmol) and DCC (72 mg, 32 mmol) was added. The mixture was stirred at 50° C. for 1 h. The oil bath was removed. 5-Amino-isophthalic acid mono t-butyl ester (69 mg, 0.29) and DIPEA (0.07 ml, 0.32 mmol) was added. The mixture was stirred at RT over night under nitrogen. The yellow suspension was filtered and the filtrate was concentrated. The residue was redissolved in EtOAc and extracted with 0.1 N HCl (2×), brine (1×), dried (Na2SO4) and concentrated to give a white solid, which was purified twice by flash chromatography using EtOAc/Hept 1:2+5% AcOH. The title compound was obtained in 86% yield (0.140 g) as an oil containing an impurity. The oil bath was removed The mixture was stirred at RT over night under nitrogen The yellow suspension was filtered the filtrate was concentrated The residue was redissolved in EtOAc extracted with 0.1 N HCl (2×), brine (1×) dried (Na2SO4) concentrated to give a white solid, which was purified twice by flash chromatography

Conditions: temp:50.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Hexadecanedioic acid mono-tert-butyl ester (0.4 g, 1.17 mmol) was dissolved in NMP (6 mL). N-Ethyl-N-(3-dimethylaminopropyl)-carbodiimide hydrochloride (0.223 g, 0.1.17 mmol) and 1-hydroxy-7-azabenzotriazole (0.159 g, 1.17 mmol) was added and the mixture was stirred at 50° C. for 1 hour and then allowed to cool to room temperature. Diisopropylethylamin (0.6 mL, 3.5 mmol) was added followed by methyl 4-aminobenzoate (0.353 g, 2.34 mmol). The mixture was stirred overnight under nitrogen at room temperature. The mixture was poured into saturated aqueous NaCl (50 mL), washed with diethylether (3×100 mL between ethyl acetate. The organic phases were collected, dried (Na2SO4) and solvent removed in vacuo. The crude material was purified on silica using ethyl acetate/heptane (50:50), to give pure 4-(15-tert-Butoxycarbonylpentadecanoylamino)benzoic acid methyl ester (235 mg, 42%) to cool to room temperature The mixture was stirred overnight under nitrogen at room temperature washed with diethylether (3×100 mL between ethyl acetate The organic phases were collected dried (Na2SO4) and solvent removed in vacuo The crude material was purified on silica

Conditions: temp:50.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Hexadecanedioic acid mono-tert-butyl ester (0.4 g, 1.17 mmol) was dissolved in NMP (6 mL). N-Ethyl-N-(3-dimethylaminopropyl)-carbodiimide hydrochloride (0.223 g, 0.1.17 mmol) and 1-hydroxy-7-azabenzotriazole (0.159 g, 1.17 mmol) was added and the mixture was stirred at 50° C. for 1 hour and then allowed to cool to room temperature. Diisopropylethylamin (0.6 mL, 3.5 mmol) was added followed by methyl 4-aminobenzoate (0.353 g, 2.34 mmol). The mixture was stirred overnight under nitrogen at room temperature. The mixture was poured into saturated aqueous NaCl (50 mL), washed with diethylether (3×100 mL between ethyl acetate. The organic phases were collected, dried (Na2SO4) and solvent removed in vacuo. The crude material was purified on silica using ethyl acetate/heptane (50:50), to give pure 4-(15-tert-Butoxycarbonylpentadecanoylamino)benzoic acid methyl ester (235 mg, 42%) to cool to room temperature The mixture was stirred overnight under nitrogen at room temperature washed with diethylether (3×100 mL between ethyl acetate The organic phases were collected dried (Na2SO4) and solvent removed in vacuo The crude material was purified on silica

Conditions: temp:50.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Hexadecanedioic acid mono-tert-butyl ester (0.3 g, 0.88 mmol) was dissolved in ethyl acetate. N-Ethyl-N-(3-dimethylaminopropyl)-carbodiimide hydrochloride (0.167 g, 0.88 mmol) and 1-hydroxy-7-azabenzotriazole (0.119 g, 0.88 mmol) was added and the mixture was stirred at 50° C. for 1 hour. After cooling to room temperature, diisopropylethylamin (0.45 mL, 2.63 mmol) was added followed by 4-[(2-tert-Butoxycarbonylethylamino)methyl]benzoic acid (0.489 g, 1.75 mmol). The mixture was stirred overnight under nitrogen at room temperature. The mixture was separated between ethyl acetate (100 mL) and water (2×50 mL). The organic phase was dried (Na2SO4), solvent removed in vacuo. The crude product was purified by RP-HPLC on C18-column, buffer A: 0.1% TFA, buffer B: MeCN+0.1% TFA; gradient 80-100% B to yield the title compound (145 mg, 27%). After cooling to room temperature The mixture was stirred overnight under nitrogen at room temperature The mixture was separated between ethyl acetate (100 mL) and water (2×50 mL) The organic phase was dried (Na2SO4), solvent removed in vacuo The crude product was purified by RP-HPLC on C18-column, buffer A: 0.1% TFA, buffer B

Conditions: temp:50.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: To a solution of hexadecandioic acid mono t-butyl ester (100 mg, 0.29 mmol) in dry DCM (2 ml), HOAt (44 mg, 0.29 mmol) and DCC (72 mg, 32 mmol) was added. The mixture was stirred at 50° C. for 1 h. The oil bath was removed. 5-Amino-isophthalic acid mono t-butyl ester (69 mg, 0.29) and DIPEA (0.07 ml, 0.32 mmol) was added. The mixture was stirred at RT over night under nitrogen. The yellow suspension was filtered and the filtrate was concentrated. The residue was redissolved in EtOAc and extracted with 0.1N HCl (2×), brine (1×), dried (Na2SO4) and concentrated to give a white solid, which was purified twice by flash chromatography using EtOAc/Hept 1:2+5% AcOH. The title compound was obtained in 86% yield (0.140 g) as an oil containing an impurity. The oil bath was removed The mixture was stirred at RT over night under nitrogen The yellow suspension was filtered the filtrate was concentrated The residue was redissolved in EtOAc extracted with 0.1N HCl (2×), brine (1×) dried (Na2SO4) concentrated to give a white solid, which was purified twice by flash chromatography

Conditions: temp:50.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: To a solution of hexadecandioic acid mono t-butyl ester (100 mg, 0.29 mmol) in dry DCM (2 ml), HOAt (44 mg, 0.29 mmol) and DCC (72 mg, 32 mmol) was added. The mixture was stirred at 50° C. for 1 h. The oil bath was removed. 5-Amino-isophthalic acid mono t-butyl ester (69 mg, 0.29) and DIPEA (0.07 ml, 0.32 mmol) was added. The mixture was stirred at RT over night under nitrogen. The yellow suspension was filtered and the filtrate was concentrated. The residue was redissolved in EtOAc and extracted with 0.1N HCl (2×), brine (1×), dried (Na2SO4) and concentrated to give a white solid, which was purified twice by flash chromatography using EtOAc/Hept 1:2+5% AcOH. The title compound was obtained in 86% yield (0.140 g) as an oil containing an impurity. The oil bath was removed The mixture was stirred at RT over night under nitrogen The yellow suspension was filtered the filtrate was concentrated The residue was redissolved in EtOAc extracted with 0.1N HCl (2×), brine (1×) dried (Na2SO4) concentrated to give a white solid, which was purified twice by flash chromatography

Conditions: temp:50.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Hexadecanedioic acid mono-tert-butyl ester (0.4 g, 1.17 mmol) was dissolved in NMP (6 mL). N-Ethyl-N-(3-dimethylaminopropyl)-carbodiimide hydrochloride (0.223 g, 0.1.17 mmol) and 1-hydroxy-7-azabenzotriazole (0.159 g, 1.17 mmol) was added and the mixture was stirred at 50° C. for 1 hour and then allowed to cool to room temperature. Diisopropylethylamin (0.6 mL, 3.5 mmol) was added followed by methyl 4-aminobenzoate (0.353 g, 2.34 mmol). The mixture was stirred overnight under nitrogen at room temperature. The mixture was poured into saturated aqueous NaCl (50 mL), washed with diethylether (3×100 mL between ethyl acetate. The organic phases were collected, dried (Na2SO4) and solvent removed in vacuo. The crude material was purified on silica using ethyl acetate/heptane (50:50), to give pure 4-(15-tert-Butoxycarbonylpentadecanoylamino)benzoic acid methyl ester (235 mg, 42%) to cool to room temperature The mixture was stirred overnight under nitrogen at room temperature washed with diethylether (3×100 mL between ethyl acetate The organic phases were collected dried (Na2SO4) and solvent removed in vacuo The crude material was purified on silica

Conditions: temp:50.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Hexadecanedioic acid mono-tert-butyl ester (0.3 g, 0.88 mmol) was dissolved in ethyl acetate. N-Ethyl-N-(3-dimethylaminopropyl)-carbodiimide hydrochloride (0.167 g, 0.88 mmol) and 1-hydroxy-7-azabenzotriazole (0.119 g, 0.88 mmol) was added and the mixture was stirred at 50° C. for 1 hour. After cooling to room temperature, diisopropylethylamin (0.45 mL, 2.63 mmol) was added followed by 4-[(2-tert-Butoxycarbonylethylamino)methyl]benzoic acid (0.489 g, 1.75 mmol). The mixture was stirred overnight under nitrogen at room temperature. The mixture was separated between ethyl acetate (100 mL) and water (2×50 mL). The organic phase was dried (Na2SO4), solvent removed in vacuo. The crude product was purified by RP-HPLC on C18-column, buffer A: 0.1% TFA, buffer B: MeCN+0.1% TFA; gradient 80-100% B to yield the title compound (145 mg, 27%). After cooling to room temperature The mixture was stirred overnight under nitrogen at room temperature The mixture was separated between ethyl acetate (100 mL) and water (2×50 mL) The organic phase was dried (Na2SO4), solvent removed in vacuo The crude product was purified by RP-HPLC on C18-column, buffer A: 0.1% TFA, buffer B

Conditions: temp:50.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Hexadecanedioic acid mono-tert-butyl ester (0.3 g, 0.88 mmol) was dissolved in ethyl acetate. N-Ethyl-N-(3-dimethylaminopropyl)-carbodiimide hydrochloride (0.167 g, 0.88 mmol) and 1-hydroxy-7-azabenzotriazole (0.119 g, 0.88 mmol) was added and the mixture was stirred at 50° C. for 1 hour. After cooling to room temperature, diisopropylethylamin (0.45 mL, 2.63 mmol) was added followed by 4-[(2-tert-Butoxycarbonylethylamino)methyl]benzoic acid (0.489 g, 1.75 mmol). The mixture was stirred overnight under nitrogen at room temperature. The mixture was separated between ethyl acetate (100 mL) and water (2×50 mL). The organic phase was dried (Na2SO4), solvent removed in vacuo. The crude product was purified by RP-HPLC on C18-column, buffer A: 0.1% TFA, buffer B: MeCN+0.1% TFA; gradient 80-100% B to yield the title compound (145 mg, 27%). After cooling to room temperature The mixture was stirred overnight under nitrogen at room temperature The mixture was separated between ethyl acetate (100 mL) and water (2×50 mL) The organic phase was dried (Na2SO4), solvent removed in vacuo The crude product was purified by RP-HPLC on C18-column, buffer A: 0.1% TFA, buffer B

Conditions: temp:0.00; pressure:0.00; stir:

Conditions: temp:0.00; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Hexadecanedioic acid mono-tert-butyl ester can be activated with TSTU in similar fashion to the method described in Example 1, Step 6. The product can be reacted with Boc-Lys-OH in DMF at RT for 16 h. After concentrating under vacuum, AcOEt can be added the residue and the mixture can be washed with 0.2 N HCl. The organic phase is then dried over MgSO4 and concentrated under vacuum to yield the crude product, 15-((S)-5-tert-Butoxycarbonylamino-5-carboxypentylcarbamoyl)pentadecanoic acid tert-butyl ester. This can either be used without further purification or purified by flash chromatography. This compound can be activated with TSTU in similar fashion to the method described in Example 1, Step 6 to yield 15-[(S)-5-tert-butoxycarbonylamino-5-(2,5-dioxopyrrolidin-1-yloxycarbonyl) pentylcarbamoyl]pentadecanoic acid tert-butyl ester, and this compound can be coupled to insulin and deprotected to yield the title compound as described in Example 1, Step 7 using prepatative HPLC to isolate the product. After concentrating under vacuum, AcOEt can be added the residue the mixture can be washed with 0.2 N HCl The organic phase is then dried over MgSO4 concentrated under vacuum