(2S)-1-[2-[(2-methylpropan-2-yl)oxycarbonylamino]acetyl]pyrrolidine-2-carboxylic acid

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

Procedure: Compound 28 was synthesized from compound 11 (0.471 mmol) and compound 27 (0.707 mmol), following the procedure as described for compound 23, as an off-white solid in 46% yield. MS (ESI, EI+) m/z=471.99-474.01 (MH+).

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

Procedure: Compound 27 was synthesized from compound 26, following the procedure as described for compound 3a, as a white solid. MS (ESI, EI−) m/z=271 (MH−).

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

Procedure: Compound 27 was synthesized from compound 26, following the procedure as described for compound 3a, as a white solid. MS (ESI, EI−) m/z=271 (MH−).

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

Procedure: 1.4 ml (11 mmol) of N-methylmorpholine are added to 1.15 g (10 mmol) of proline in 50 ml of DMF. The mixture is cooled with an ice bath, 3.06 g (10 mmol) of Boc-Gly-OSu are then added and the pH is adjusted to between 6 and 7 throughout the reaction (14 h) by successive additions of small amounts of N-methylmorpholine (this substance is used as a proton acceptor). The DMF is evaporated off and the evaporation residue is taken up with AcOEt; after precipitation in petroleum ether and then filtration, the expected product is collected. The mixture is cooled with an ice bath The DMF is evaporated off after precipitation in petroleum ether filtration the expected product is collected

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

Procedure: 1.4 ml (11 mmol) of N-methylmorpholine are added to 1.15 g (10 mmol) of proline in 50 ml of DMF. The mixture is cooled with an ice bath, 3.06 g (10 mmol) of Boc-Gly-OSu are then added and the pH is adjusted to between 6 and 7 throughout the reaction (14 h) by successive additions of small amounts of N-methylmorpholine (this substance is used as a proton acceptor). The DMF is evaporated off and the evaporation residue is taken up with AcOEt; after precipitation in petroleum ether and then filtration, the expected product is collected. The mixture is cooled with an ice bath The DMF is evaporated off after precipitation in petroleum ether filtration the expected product is collected

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

Procedure: A portion (10.3g= 0.05 mole) of dicyclohexylcarbodiimide (DCC) was added dropwise under stirring at 0° C. to a solution of N-Boc-glycyl-L-proline prepared in accordance with the method described in J. Chem. Soc. (C), 954 (1969) (27.2g= 0.1 mole) in dried THF (100ml) and the reaction mixture was kept under continuous stirring for 45 minutes. To the filtrate obtained by filtering-off of dicyclohexyl urea precipitated in the mixture, was added at 0° C. a solution of 7-amino-4-methylcoumarin (8.75g= 0.05 mole) in dimethylformamide (DMF) (60ml). After standing overnight at room temperature, the reaction mixture was concentrated in vacuo to give a residue which was dissolved in ethyl acetate (300ml.). The solution was washed successively with 1 N hydrochloric acid, aqueous 5% sodium bicarbonate and water, dried with anhydrous magnesium sulfate, and concentrated in vacuo. The residue was recrystallized from a binary mixture of ethyl acetate (150ml) and n-hexance (70ml) to give 20.1g (yield, 93.4%) of crystalline 7-(n-BOC-glycyl-L-prolyl)-amino-4-methylcoumarin. prepared in accordance with the method

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

Procedure: A portion (10.3g= 0.05 mole) of dicyclohexylcarbodiimide (DCC) was added dropwise under stirring at 0° C. to a solution of N-Boc-glycyl-L-proline prepared in accordance with the method described in J. Chem. Soc. (C), 954 (1969) (27.2g= 0.1 mole) in dried THF (100ml) and the reaction mixture was kept under continuous stirring for 45 minutes. To the filtrate obtained by filtering-off of dicyclohexyl urea precipitated in the mixture, was added at 0° C. a solution of 7-amino-4-methylcoumarin (8.75g= 0.05 mole) in dimethylformamide (DMF) (60ml). After standing overnight at room temperature, the reaction mixture was concentrated in vacuo to give a residue which was dissolved in ethyl acetate (300ml.). The solution was washed successively with 1 N hydrochloric acid, aqueous 5% sodium bicarbonate and water, dried with anhydrous magnesium sulfate, and concentrated in vacuo. The residue was recrystallized from a binary mixture of ethyl acetate (150ml) and n-hexance (70ml) to give 20.1g (yield, 93.4%) of crystalline 7-(n-BOC-glycyl-L-prolyl)-amino-4-methylcoumarin. prepared in accordance with the method

Conditions: temp:0.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Boc-Gly-Pro-OH (3.85 g, 0.014 mol), the HCl salt of Nleu-OEt (2.76 g, 0.014 mol) and HOBt (2.8 g, 0.02 mol.) were dissolved in 100 ml DMF and the solution was chilled to 0° C. on a water-ice bath. TEA (2.8 ml, 0.02 mol.) was added to the solution. After stirring the solution for 10 minutes, EDC (4.0 g, 0.021 mol.) was added. The stirring was continued overnight. The DMF was removed under reduced pressure and the resulting mixture was poured into 200 ml ethyl acetate. The organic layer was washed by H2O 2×30 ml, saturated NaHCO3 3×10 ml, brine 2×10 ml, saturated NaHSO4 3×15 ml and brine again until the pH value of the brine layer was approximately 7. The ethyl acetate solution was dried using Na2SO4 and the solvent was removed by distillation under reduced pressure. Column chromatography (elution solvents: E/H, 2:1) was performed to obtain the pure product (yellowish oil, 4.7 g, 81%). TLC Rf=0.54 (developing solvents: CMA, 85:15:3) and Rf=0.34 (developing solvents: E/H, 2:1). The stirring was continued overnight The DMF was removed under reduced pressure the resulting mixture was poured into 200 ml ethyl acetate The organic layer was washed by H2O 2×30 ml, saturated NaHCO3 3×10 ml, brine 2×10 ml, saturated NaHSO4 3×15 ml and brine again until the pH value of the brine layer The ethyl acetate solution was dried the solvent was removed by distillation under reduced pressure Column chromatography (elution solvents: E/H, 2:1) to obtain the pure product (yellowish oil, 4.7 g, 81%)

Conditions: temp:0.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Boc-Gly-Pro-OH (3.85 g, 0.014 mol), the HCl salt of Nleu-OEt (2.76 g, 0.014 mol) and HOBt (2.8 g, 0.02 mol.) were dissolved in 100 ml DMF and the solution was chilled to 0° C. on a water-ice bath. TEA (2.8 ml, 0.02 mol.) was added to the solution. After stirring the solution for 10 minutes, EDC (4.0 g, 0.021 mol.) was added. The stirring was continued overnight. The DMF was removed under reduced pressure and the resulting mixture was poured into 200 ml ethyl acetate. The organic layer was washed by H2O 2×30 ml, saturated NaHCO3 3×10 ml, brine 2×10 ml, saturated NaHSO4 3×15 ml and brine again until the pH value of the brine layer was approximately 7. The ethyl acetate solution was dried using Na2SO4 and the solvent was removed by distillation under reduced pressure. Column chromatography (elution solvents: E/H, 2:1) was performed to obtain the pure product (yellowish oil, 4.7 g, 81%). TLC Rf=0.54 (developing solvents: CMA, 85:15:3) and Rf=0.34 (developing solvents: E/H, 2:1). The stirring was continued overnight The DMF was removed under reduced pressure the resulting mixture was poured into 200 ml ethyl acetate The organic layer was washed by H2O 2×30 ml, saturated NaHCO3 3×10 ml, brine 2×10 ml, saturated NaHSO4 3×15 ml and brine again until the pH value of the brine layer The ethyl acetate solution was dried the solvent was removed by distillation under reduced pressure Column chromatography (elution solvents: E/H, 2:1) to obtain the pure product (yellowish oil, 4.7 g, 81%)

Conditions: temp:0.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Boc-Gly-ONp (50 g, 0.17 mol.) and proline (20.7 g, 0.175 mol.) were dissolved in 300 ml DMF. 20 ml TEA and 30 ml water were added while stirring the solution. The stirring was continued overnight and TLC analysis showed no more starting materials. The solvents were removed under reduced pressure and the resulting mixture was dissolved in 250 ml saturated Na2CO3. The aqueous solution was extracted by 3×100 ml ethyl acetate to remove p-nitrophenol. Then, after chilling to 0° C., the aqueous solution was acidified to about pH=3 by addition of concentrated HCl slowly (vigorously swirling the solution at the same time). The product was extracted from the aqueous phase by 3×150 ml ethyl acetate. The organic phase was combined and washed by saturated NaCl (brine) 3×20 ml, and dried using Na2SO4. TLC analysis showed there was still p-nitrophenol in the solution. The ethyl acetate was removed under reduced pressure and column chromatography (elution solvents: E/H, 2:1) was carried out to obtain the pure product Boc-Gly-Pro-OH (white solid, 41.2 g, 88%). TLC Rf=0.25 (CMA, 85:15:3). FAB-MS, observed (M+H)+=273. 1H-NMR (360 MHZ, DMSO-d6, 20° C.) δ12.55 (s, 1H, carboxyl), 6.79 (s, NH-major), 6.44 (s, NH-minor), 4.50 (dd, 0.25H, Pro-α-minor), 4.20 (dd, 0.71H, Pro-α-major), 3.81-3.62 (m, 1.7H, Gly-α-major), 3.47-3.33 (m, 2.5H, Pro-δ and Gly-α-minor), 2.25-2.00 (m, 1.2H, Pro-β), 1.96-1.60 (m, 2.7H, Pro-β and Pro-γ), 1.36 (s, 9H, Boc-CH3)b. The solvents were removed under reduced pressure the resulting mixture was dissolved in 250 ml saturated Na2CO3 The aqueous solution was extracted by 3×100 ml ethyl acetate to remove p-nitrophenol the aqueous solution was acidified to about pH=3 by addition of concentrated HCl slowly ( The product was extracted from the aqueous phase by 3×150 ml ethyl acetate washed by saturated NaCl (brine) 3×20 ml dried The ethyl acetate was removed under reduced pressure and column chromatography (elution solvents: E/H, 2:1)

Conditions: temp:0.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Boc-Gly-ONp (50 g, 0.17 mol.) and proline (20.7 g, 0.175 mol.) were dissolved in 300 ml DMF. 20 ml TEA and 30 ml water were added while stirring the solution. The stirring was continued overnight and TLC analysis showed no more starting materials. The solvents were removed under reduced pressure and the resulting mixture was dissolved in 250 ml saturated Na2CO3. The aqueous solution was extracted by 3×100 ml ethyl acetate to remove p-nitrophenol. Then, after chilling to 0° C., the aqueous solution was acidified to about pH=3 by addition of concentrated HCl slowly (vigorously swirling the solution at the same time). The product was extracted from the aqueous phase by 3×150 ml ethyl acetate. The organic phase was combined and washed by saturated NaCl (brine) 3×20 ml, and dried using Na2SO4. TLC analysis showed there was still p-nitrophenol in the solution. The ethyl acetate was removed under reduced pressure and column chromatography (elution solvents: E/H, 2:1) was carried out to obtain the pure product Boc-Gly-Pro-OH (white solid, 41.2 g, 88%). TLC Rf=0.25 (CMA, 85:15:3). FAB-MS, observed (M+H)+=273. 1H-NMR (360 MHZ, DMSO-d6, 20° C.) δ12.55 (s, 1H, carboxyl), 6.79 (s, NH-major), 6.44 (s, NH-minor), 4.50 (dd, 0.25H, Pro-α-minor), 4.20 (dd, 0.71H, Pro-α-major), 3.81-3.62 (m, 1.7H, Gly-α-major), 3.47-3.33 (m, 2.5H, Pro-δ and Gly-α-minor), 2.25-2.00 (m, 1.2H, Pro-β), 1.96-1.60 (m, 2.7H, Pro-β and Pro-γ), 1.36 (s, 9H, Boc-CH3)b. The solvents were removed under reduced pressure the resulting mixture was dissolved in 250 ml saturated Na2CO3 The aqueous solution was extracted by 3×100 ml ethyl acetate to remove p-nitrophenol the aqueous solution was acidified to about pH=3 by addition of concentrated HCl slowly ( The product was extracted from the aqueous phase by 3×150 ml ethyl acetate washed by saturated NaCl (brine) 3×20 ml dried The ethyl acetate was removed under reduced pressure and column chromatography (elution solvents: E/H, 2:1)

Conditions: temp:0.01; pressure:0.00; stir:; See reaction.notes.procedure_details.

Procedure: Boc-Gly-ONp (50 g, 0.17 mol.) and proline (20.7 g, 0.175 mol.) were dissolved in 300 ml DMF. 20 ml TEA and 30 ml water were added while stirring the solution. The stirring was continued overnight and TLC analysis showed no more starting materials. The solvents were removed under reduced pressure and the resulting mixture was dissolved in 250 ml saturated Na2CO3. The aqueous solution was extracted by 3×100 ml ethyl acetate to remove p-nitrophenol. Then, after chilling to 0° C., the aqueous solution was acidified to about pH=3 by addition of concentrated HCl slowly (vigorously swirling the solution at the same time). The product was extracted from the aqueous phase by 3×150 ml ethyl acetate. The organic phase was combined and washed by saturated NaCl (brine) 3×20 ml, and dried using Na2SO4. TLC analysis showed there was still p-nitrophenol in the solution. The ethyl acetate was removed under reduced pressure and column chromatography (elution solvents: E/H, 2:1) was carried out to obtain the pure product Boc-Gly-Pro-OH (white solid, 41.2 g, 88%). TLC Rf=0.25 (CMA, 85:15:3). FAB-MS, observed (M+H)+=273. 1H-NMR (360 MHZ, DMSO-d6, 20° C.) δ12.55 (s, 1H, carboxyl), 6.79 (s, NH-major), 6.44 (s, NH-minor), 4.50 (dd, 0.25H, Pro-α-minor), 4.20 (dd, 0.71H, Pro-α-major), 3.81-3.62 (m, 1.7H, Gly-α-major), 3.47-3.33 (m, 2.5H, Pro-δ and Gly-α-minor), 2.25-2.00 (m, 1.2H, Pro-β), 1.96-1.60 (m, 2.7H, Pro-β and Pro-γ), 1.36 (s, 9H, Boc-CH3)b. The solvents were removed under reduced pressure the resulting mixture was dissolved in 250 ml saturated Na2CO3 The aqueous solution was extracted by 3×100 ml ethyl acetate to remove p-nitrophenol the aqueous solution was acidified to about pH=3 by addition of concentrated HCl slowly ( The product was extracted from the aqueous phase by 3×150 ml ethyl acetate washed by saturated NaCl (brine) 3×20 ml dried The ethyl acetate was removed under reduced pressure and column chromatography (elution solvents: E/H, 2:1)