(2,5-dioxopyrrolidin-1-yl) 2-[(2-methylpropan-2-yl)oxycarbonylamino]acetate

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

Procedure: To the aforesaid Boc-Gly-OSu solution the thyroxine methyl ester solution was added while ice cooling. After permitting the same to warm to room temperature, the mixture was stirred for 1.5 hr. After letting the reaction mixture stand overnight, 25 ml. of distilled water and 50 ml. of ethyl acetate were added thereto and the mixture was extracted with ethyl acetate. The extraction was repeated three times using 25 ml. of ethyl acetate. The ethyl acetate layers were combined and dried over anhydrous magnesium sulfate. The liquid was concentrated using a rotary evaporator and subjected to gel chromatography using a column filled with Sephadex LH-20 (made by Pharmacia Co., Ltd.). As the solvent, a mixture obtained by mixing acetone and methanol in a proportion of 4:1 (volume) was used. While confirming by TLC, the objective fraction was taken and concentrated under reduced pressure followed by crystallization. The product was obtained in an amount of 950 mg. (yield 82.5%). stand overnight the mixture was extracted with ethyl acetate The extraction dried over anhydrous magnesium sulfate The liquid was concentrated a rotary evaporator filled with Sephadex LH-20 (made by Pharmacia Co., Ltd.) As the solvent, a mixture obtained concentrated under reduced pressure followed by crystallization The product was obtained in an amount of 950 mg

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

Procedure: To the aforesaid Boc-Gly-OSu solution the thyroxine methyl ester solution was added while ice cooling. After permitting the same to warm to room temperature, the mixture was stirred for 1.5 hr. After letting the reaction mixture stand overnight, 25 ml. of distilled water and 50 ml. of ethyl acetate were added thereto and the mixture was extracted with ethyl acetate. The extraction was repeated three times using 25 ml. of ethyl acetate. The ethyl acetate layers were combined and dried over anhydrous magnesium sulfate. The liquid was concentrated using a rotary evaporator and subjected to gel chromatography using a column filled with Sephadex LH-20 (made by Pharmacia Co., Ltd.). As the solvent, a mixture obtained by mixing acetone and methanol in a proportion of 4:1 (volume) was used. While confirming by TLC, the objective fraction was taken and concentrated under reduced pressure followed by crystallization. The product was obtained in an amount of 950 mg. (yield 82.5%). stand overnight the mixture was extracted with ethyl acetate The extraction dried over anhydrous magnesium sulfate The liquid was concentrated a rotary evaporator filled with Sephadex LH-20 (made by Pharmacia Co., Ltd.) As the solvent, a mixture obtained concentrated under reduced pressure followed by crystallization The product was obtained in an amount of 950 mg

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

Procedure: Boc-Gly-Ala-OH was prepared by an N-hydroxysuccinimide coupling procedure. Boc-Gly-OSu (7.67 g, 28.2 mmol) was dissolved in 10 mL of dimethylsulfoxide and the resulting solution was added to a solution consisting of H-Ala-OH (3.76 g, 42.3 mmol) and triethylamine (5.89 mL, 42.3 mmol) in 20 mL of water. After stirring overnight, a precipitate was removed by filtration. The remaining solution was diluted with water and then acidified with hydrochloric acid. An aqueous phase separated from which product was extracted by addition of ethyl acetate. The ethyl acetate extract was washed with 0.2N hydrochloric acid and then saturated sodium chloride prepared in 0.2N hydrochloric acid. The washed ethyl acetate solution was then dried over sodium sulfate and filtered. Solvent was then evaporated, leaving a foamy product, which was crystallized from ethyl acetate:hexane to yield 2.88 g of Boc-Gly-Ala-OH (m.p. 128°-130°). the resulting solution was added to a solution a precipitate was removed by filtration The remaining solution was diluted with water An aqueous phase separated from which product was extracted by addition of ethyl acetate The ethyl acetate extract was washed with 0.2N hydrochloric acid saturated sodium chloride prepared in 0.2N hydrochloric acid The washed ethyl acetate solution was then dried over sodium sulfate filtered Solvent was then evaporated leaving a foamy product, which was crystallized from ethyl acetate

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

Procedure: 10.00 g (36.73 mmol) Boc-Gly-Osu (Bachem) was dissolved together with 3.35 g (36.77 mmol) 1-amino-2,3-propanediol in 50 ml DMF and stirred for 18 h. The product was purified by flash chromatography (CH2Cl2 /MeOH; 95:5 v/v). 8.94 g (98%) 19 was obtained as a colourless oil. The product was purified by flash chromatography (CH2Cl2 /MeOH; 95:5 v/v)

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

Procedure: 10.00 g (36.73 mmol) Boc-Gly-Osu (Bachem) was dissolved together with 3.35 g (36.77 mmol) 1-amino-2,3-propanediol in 50 ml DMF and stirred for 18 h. The product was purified by flash chromatography (CH2Cl2 /MeOH; 95:5 v/v). 8.94 g (98%) 19 was obtained as a colourless oil. The product was purified by flash chromatography (CH2Cl2 /MeOH; 95:5 v/v)

Conditions: temp:2.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: In a 5 L reactor, to a solution of (2S)-2-amino-4-penten-1-ol D20 (110 g of the crude title compound prepared in the description D20) in THF (660 ml) and MeOH (440 ml) stirred at 0° C. (+5° C. internal) was added triethylamine (182 ml, 1305 mmol) and 2,5-dioxo-1-pyrrolidinyl N-{[(1,1-dimethylethyl)oxy]carbonyl}glycinate (available from Sigma-Aldrich #15423) (237 g, 870 mmol) portionwise over 15 min. The reaction mixture was stirred at 2° C. (internal temperature) for 3 hours. TLC check (TLC-NH2, DCM/MeOH 95/5, potassium permanganate) showed residual starting material. Further 2,5-dioxo-1-pyrrolidinyl N-{[(1,1-dimethylethyl)oxy]carbonyl}glycinate (60 g, 220 mmol) was added and the mixture stirred at 2° C. for 1 hour. TLC check (TLC-NH2, DCM/MeOH 95/5, potassium permanganate) showed residual starting material. Further 2,5-dioxo-1-pyrrolidinyl N-{[(1,1-dimethylethyl)oxy]carbonyl}glycinate (40 g, 146 mmol) was added and the mixture stirred at 2° C. for 1 hours. TLC check (TLC-NH2, DCM/MeOH 95/5, potassium permanganate) showed residual starting material but the work-up was carried out. The reaction mixture was poured into aqueous saturated solution of NH4Cl (3400 ml) and AcOEt (1375 ml), then the phases were separated and the aqueous layer was back-extracted with AcOEt (1375 ml). The combined organic layers were washed with NaHCO3 aqueous saturated solution (1031 ml) dried (Na2SO4) and evaporated to give crude material (268 g, deep brown). This residue was triturated with Et2O (687 ml) for 1 hour at 25° C. The solid was filtered (gooch n3), washed with Et2O (200 ml) and dried under vacuum to give the title compound D21 (87 g) as pale brown solid. Mother liquors (deep brown) were evaporated and the residue chromatographed (Biotage 75 L, silica column, eluting with DCM/MeOH 98/2, 95/5) to give (34 g) of residual brown product that was triturated with Et2O (200 ml). The solid was filtered, washed with Et2O and dried under vacuum to give a further batch of the title compound D21 (26 g) as pale brown solid. MS: (ES/+) m/z: 259 (M+1). C12H22N2O4 requires 258. 1H NMR (400 MHz, CDCl3) δ ppm 1.47 (s, 9 H) 2.22-2.43 (m, 2 H) 2.68-2.83 (m, 1 H) 3.50-3.86 (m, 4 H) 3.94-4.09 (m, 1 H) 5.00-5.25 (m, 3 H) 5.64-5.89 (m, 1 H) 6.17-6.44 (m, 1 H) the mixture stirred at 2° C. for 1 hour the mixture stirred at 2° C. for 1 hours the phases were separated the aqueous layer was back-extracted with AcOEt (1375 ml) The combined organic layers were washed with NaHCO3 aqueous saturated solution (1031 ml) dried (Na2SO4) evaporated to give crude material (268 g, deep brown) This residue was triturated with Et2O (687 ml) for 1 hour at 25° C The solid was filtered (gooch n3) washed with Et2O (200 ml) dried under vacuum

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

Procedure: Boc-Gly-Ala-OH was prepared by an N-hydroxysuccinimide coupling procedure. Boc-Gly-OSu (7.67 g, 28.2 mmol) was dissolved in 10 mL of dimethylsulfoxide, and the resulting solution was added to a solution consisting of H-Ala-OH (3.76 g, 42.3 mmol) and triethylamine (5.89 mL, 42.3 mmol) in 20 mL of water. After stirring overnight, a precipitate was removed by filtration. The remaining solution was diluted with water and then acidified with hydrochloric acid. An aqueous phase separated from which product was extracted by addition of ethyl acetate. The ethyl acetate extract was washed with 0.2N hydrochloric acid and then saturated sodium chloride prepared in 0.2N hydrochloric acid. The washed ethyl acetate solution was then dried over sodium sulfate and filtered. Solvent was then evaporated, leaving a foamy product, which was crystallized from ethyl acetate:hexane to yield 2.88 g of Boc-Gly-Ala-OH (m.p. 128°-130°). the resulting solution was added to a solution a precipitate was removed by filtration The remaining solution was diluted with water An aqueous phase separated from which product was extracted by addition of ethyl acetate The ethyl acetate extract was washed with 0.2N hydrochloric acid saturated sodium chloride prepared in 0.2N hydrochloric acid The washed ethyl acetate solution was then dried over sodium sulfate filtered Solvent was then evaporated leaving a foamy product, which was crystallized from ethyl acetate

Conditions: temp:22.51; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Under ice-cooling, to a suspension of 6-(1-amino-2-oxo-2-phenylethyl)-2-isopropyl-3(2H)-pyridazinone hydrochloride (1.00 g) and tert-butyl {2-[(2,5-dioxo-1-pyrrolidinyl)oxy]-2-oxoethyl}carbamate (973 mg) in CH2Cl2 (10 ml) was added ethyl(diisopropyl)amine (0.651 ml) and stirred at the same temperature for 2 hours and at 20-25° C. for 18 hours. The mixture was washed with water, dried over MgSO4 and concentrated under reduced pressure to give syrup. The syrup was purified by column chromatography on silica gel (EtOAc only) to give tert-butyl (2-{[1-(1-isopropyl-6-oxo-1,6-dihydro-3-pyridazinyl)-2-oxo-2-phenylethyl]amino}-2-oxoethyl)carbamate as an amorphous solid (1.13 g). The mixture was washed with water dried over MgSO4 concentrated under reduced pressure to give syrup The syrup was purified by column chromatography on silica gel (EtOAc only)

Conditions: temp:22.51; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Under ice-cooling, to a suspension of 6-(1-amino-2-oxo-2-phenylethyl)-2-isopropyl-3(2H)-pyridazinone hydrochloride (1.00 g) and tert-butyl {2-[(2,5-dioxo-1-pyrrolidinyl)oxy]-2-oxoethyl}carbamate (973 mg) in CH2Cl2 (10 ml) was added ethyl(diisopropyl)amine (0.651 ml) and stirred at the same temperature for 2 hours and at 20-25° C. for 18 hours. The mixture was washed with water, dried over MgSO4 and concentrated under reduced pressure to give syrup. The syrup was purified by column chromatography on silica gel (EtOAc only) to give tert-butyl (2-{[1-(1-isopropyl-6-oxo-1,6-dihydro-3-pyridazinyl)-2-oxo-2-phenylethyl]amino}-2-oxoethyl)carbamate as an amorphous solid (1.13 g). The mixture was washed with water dried over MgSO4 concentrated under reduced pressure to give syrup The syrup was purified by column chromatography on silica gel (EtOAc only)

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

Procedure: To the Boc-Gly-OSu solution was added under ice-cooling the thyroxine methyl ester solution. The resulting mixture was allowed to stand to reach room temperature, and then stirred for 1.5 hrs. The reaction liquid was allowed to stand overnight, and to this were added 25 ml. of distilled water and 50 ml. of ethyl acetate for extraction with the ethyl acetate. The extraction with 25 ml. of ethyl acetate was repeated three times. The ethyl acetate extracts were combined and dried over anhydrous magnesium sulfate. The dry extract was concentrated in a rotary evaporator and subjected to gel chromatography using a column filled with Sephadex LH-20 (tradename of Pharmacia Corp.). In this procedure, a mixture of acetone and methanol in a volume ratio of 4:1 was employed as the developing solvent. Under confirmation by TLC, the fractions containing the desired product were collected and concentrated under reduced pressure to obtain a crystalline product. There was obtained 950 mg. (yield 82.5%) of the desired product. to reach room temperature The reaction liquid to stand overnight to this were added 25 ml of ethyl acetate for extraction with the ethyl acetate The extraction with 25 ml dried over anhydrous magnesium sulfate The dry extract was concentrated in a rotary evaporator filled with Sephadex In this procedure, a mixture of acetone and methanol in a volume ratio of 4:1 Under confirmation by TLC, the fractions containing the desired product were collected concentrated under reduced pressure to obtain a crystalline product There was obtained 950 mg

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

Procedure: To the Boc-Gly-OSu solution was added under ice-cooling the thyroxine methyl ester solution. The resulting mixture was allowed to stand to reach room temperature, and then stirred for 1.5 hrs. The reaction liquid was allowed to stand overnight, and to this were added 25 ml. of distilled water and 50 ml. of ethyl acetate for extraction with the ethyl acetate. The extraction with 25 ml. of ethyl acetate was repeated three times. The ethyl acetate extracts were combined and dried over anhydrous magnesium sulfate. The dry extract was concentrated in a rotary evaporator and subjected to gel chromatography using a column filled with Sephadex LH-20 (tradename of Pharmacia Corp.). In this procedure, a mixture of acetone and methanol in a volume ratio of 4:1 was employed as the developing solvent. Under confirmation by TLC, the fractions containing the desired product were collected and concentrated under reduced pressure to obtain a crystalline product. There was obtained 950 mg. (yield 82.5%) of the desired product. to reach room temperature The reaction liquid to stand overnight to this were added 25 ml of ethyl acetate for extraction with the ethyl acetate The extraction with 25 ml dried over anhydrous magnesium sulfate The dry extract was concentrated in a rotary evaporator filled with Sephadex In this procedure, a mixture of acetone and methanol in a volume ratio of 4:1 Under confirmation by TLC, the fractions containing the desired product were collected concentrated under reduced pressure to obtain a crystalline product There was obtained 950 mg

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

Procedure: N-Boc-glycine (7.31 g, 41.7 mmol) was dissolved in 100 mL of DCM and to the cooled (15° C.) solution N-hydroxysuccinimide (5.28 g, 45.9 mmol) was added. N,N′-dicyclohexylcarbodiimide (9.47 g, 45.9 mmol) was added to the formed suspension under vigorous stirring. After a few seconds, a cloudy white suspension formed, the mixture was allowed to reach room temperature and stirred for 1 h. It was subsequently filtrated over celite, washed with 50 mL saturated sodium bicarbonate, dried over sodium sulfate and concentrated in vacuo to yield a crystalline powder. Yield: 7.02 g, 61.8% was added After a few seconds a cloudy white suspension formed to reach room temperature It was subsequently filtrated over celite washed with 50 mL saturated sodium bicarbonate dried over sodium sulfate concentrated in vacuo to yield a crystalline powder