(2,5-dioxopyrrolidin-1-yl) (2S)-3-methyl-2-[(2-methylpropan-2-yl)oxycarbonylamino]butanoate

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

Procedure: A mixture of t-Boc-Val-OSu (3.14 g, 10 mmol) and H-Lys(CBZ)-OCH3.HCl (3.31 g, 10 mmol) in THF (300 ml) was treated at room temperature with triethylamine (1.38 ml) and left stirring for 17 hr. The resulting solution was evaporated in vacuo and the residue was dissolved in EtOAc (500 ml). The organic solution was washed successively with water (2×200 ml), 5% citric acid (2×200 ml), water (2×200 ml) and dried over Na2SO4. The dried solution was filtered and evaporated in vacuo to give (VI) (4.9 g; 99%) as a foam. T.l.c. RfE4 =0.64. left The resulting solution was evaporated in vacuo the residue was dissolved in EtOAc (500 ml) The organic solution was washed successively with water (2×200 ml), 5% citric acid (2×200 ml), water (2×200 ml) dried over Na2SO4 The dried solution was filtered evaporated in vacuo

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

Procedure: To a solution of Citrulline (2.54 g, 14.50 mmole) and Sodium Bicarbonate (1.28 g) in water (40 mL) was added Boc-Val-OSu (4.34 g, 13.81 mmole) dissolved in dimethoxyethane (DME). To aid the solubility of the mixture tetrahydrofuran (10 mL) was added. The mixture thus obtained was let stir overnight at room temperature. Aqueous citric acid (15%, 75 mL) was added and the mixture was extracted with 10% 2-propanol/ethyl acetate (2×100 mL). The organic layer was washed with brine (2×150 mL) and the solvents were removed on the rotovap. The resulting white solid was dried in vacuo for 5 hours and then treated with ether (100 mL). After brief sonication and trituration, the white solid product was collected by filtration (1.39 g, 27%). 1H NMR (CD3OD)δ 0.91 (dd, 3H), 0.98 (dd, 3H), 1.44 (s, 9H), 1.70 (m, 2H), 1.87 (m, 2H), 2.02 (m, 2H), 3.11 (t, 2H), 3.89 (t, 1H), 4.39 (q, 1H), 8.22 (d, 1H) ppm; LC-MS (ESI) 375 (M+H+). was added The mixture thus obtained the mixture was extracted with 10% 2-propanol/ethyl acetate (2×100 mL) The organic layer was washed with brine (2×150 mL) the solvents were removed on the rotovap The resulting white solid was dried in vacuo for 5 hours treated with ether (100 mL) After brief sonication and trituration the white solid product was collected by filtration (1.39 g, 27%)

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

Procedure: To a solution of Citrulline (2.54 g, 14.50 mmole) and Sodium Bicarbonate (1.28 g) in water (40 mL) was added Boc-Val-OSu (4.34 g, 13.81 mmole) dissolved in dimethoxyethane (DME). To aid the solubility of the mixture tetrahydrofuran (10 mL) was added. The mixture thus obtained was let stir overnight at room temperature. Aqueous citric acid (15%, 75 mL) was added and the mixture was extracted with 10% 2-propanol/ethyl acetate (2×100 mL). The organic layer was washed with brine (2×150 mL) and the solvents were removed on the rotovap. The resulting white solid was dried in vacuo for 5 hours and then treated with ether (100 mL). After brief sonication and trituration, the white solid product was collected by filtration (1.39 g, 27%). 1H NMR (CD3OD)δ 0.91 (dd, 3H), 0.98 (dd, 3H), 1.44 (s, 9H), 1.70 (m, 2H), 1.87 (m, 2H), 2.02 (m, 2H), 3.11 (t, 2H), 3.89 (t, 1H), 4.39 (q, 1H), 8.22 (d, 1H) ppm; LC-MS (ESI) 375 (M+H+). was added The mixture thus obtained the mixture was extracted with 10% 2-propanol/ethyl acetate (2×100 mL) The organic layer was washed with brine (2×150 mL) the solvents were removed on the rotovap The resulting white solid was dried in vacuo for 5 hours treated with ether (100 mL) After brief sonication and trituration the white solid product was collected by filtration (1.39 g, 27%)

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

Procedure: Boc-Asp(OBzl)-Thr(Bzl)-Ser(Bzl)-Ser(Bzl)-Glu(OBzl)-OH (50.0 g, 46 mmol) was deprotected with HCl (4.15 N) in THF and worked up as usual to give 45.4 g of white solid. It was dissolved in THF (1.5 liters) and treated with ether (7 liters). On standing at 0° overnight a white solid powder was obtained (44.0 g, mp 179°-184°). Part of this material, HCl.H-Asp(OBzl)-Thr(Bzl)-Ser(Bzl)-Ser(Bzl)-Glu(OBzl)-OH (43.7 g, 42.7 mmol) was then dissolved in 500 ml of DMF, cooled to 0°, and treated with Boc-Val-OSu (15.4 g, 49 mmol) and Et3N (10 ml). The mixture was stirred for 15 hours during which time more Et3N (7.9 ml) was added in several portions maintaining the reaction slightly basic. The insoluble matter was removed by filtration and the filtrate evaporated to dryness. The oily residue was dissolved in CH2Cl2, washed with 5% HOAc, H2O, dried (Na2SO4), and evaporated to a smaller volume (0.5 liter) when treated with petroleum ether. The product crystallized slowly during overnight standing. It was recrystallized from THF and i-PrOH to yield, 26.3 g (51.4%); mp 174°-177°; α]D25 =+0.84° (c, 1, THF). a white solid powder was obtained (44.0 g, mp 179°-184°) cooled to 0° was added in several portions maintaining the reaction slightly basic The insoluble matter was removed by filtration the filtrate evaporated to dryness The oily residue was dissolved in CH2Cl2 washed with 5% HOAc, H2O dried (Na2SO4) evaporated to a smaller volume (0.5 liter) when treated with petroleum ether The product crystallized slowly during overnight It was recrystallized from THF and i-PrOH to yield

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

Procedure: Boc-Asp(OBzl)-Thr(Bzl)-Ser(Bzl)-Ser(Bzl)-Glu(OBzl)-OH (50.0 g, 46 mmol) was deprotected with HCl (4.15 N) in THF and worked up as usual to give 45.4 g of white solid. It was dissolved in THF (1.5 liters) and treated with ether (7 liters). On standing at 0° overnight a white solid powder was obtained (44.0 g, mp 179°-184°). Part of this material, HCl.H-Asp(OBzl)-Thr(Bzl)-Ser(Bzl)-Ser(Bzl)-Glu(OBzl)-OH (43.7 g, 42.7 mmol) was then dissolved in 500 ml of DMF, cooled to 0°, and treated with Boc-Val-OSu (15.4 g, 49 mmol) and Et3N (10 ml). The mixture was stirred for 15 hours during which time more Et3N (7.9 ml) was added in several portions maintaining the reaction slightly basic. The insoluble matter was removed by filtration and the filtrate evaporated to dryness. The oily residue was dissolved in CH2Cl2, washed with 5% HOAc, H2O, dried (Na2SO4), and evaporated to a smaller volume (0.5 liter) when treated with petroleum ether. The product crystallized slowly during overnight standing. It was recrystallized from THF and i-PrOH to yield, 26.3 g (51.4%); mp 174°-177°; α]D25 =+0.84° (c, 1, THF). a white solid powder was obtained (44.0 g, mp 179°-184°) cooled to 0° was added in several portions maintaining the reaction slightly basic The insoluble matter was removed by filtration the filtrate evaporated to dryness The oily residue was dissolved in CH2Cl2 washed with 5% HOAc, H2O dried (Na2SO4) evaporated to a smaller volume (0.5 liter) when treated with petroleum ether The product crystallized slowly during overnight It was recrystallized from THF and i-PrOH to yield

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

Procedure: H-Thr(Bzl)-OBzl hemioxalate (58 g, 168.4 mmol) was suspended in DMF (700 ml) and stirred with Boc-Val-OSu (48 g, 153.1 mmol) and Et3N (16 ml) at 0° C for 1 hr, 4° C for 72 hrs and 25° C for 24 hrs. It was necessary during this time to add at intervals additional Et3N in order to maintain the pH between 7 and 8. The reaction mixture was worked up as in Example 2 to yield an oil which crystallized from EtOAc with petroleum ether, yield 40.8 g (53.5%); mp 99°-100.5° C; [α]D25 -26.0° (c 1, MeOH). Anal. Calcd for C28H38N2O6 (498.62): C, 67.45; H, 7.68; N, 5.62. Found: C, 67.57; H, 7.55; N, 5.70. to maintain the pH between 7 and 8 to yield an oil which crystallized from EtOAc with petroleum ether, yield 40.8 g (53.5%)

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

Procedure: A suspension of H-Thr(Bzl)-OH (3.0 g, 14.34 mmol) in DMF (60 ml) was mixed with Et3N (2 ml) and Boc-Val-OSu (4.96 g, 15.78 mmol) at 0° C for 2 hours and then at 25° C for 24 hours. Acetic acid was added to about pH 3 and the solvents removed under reduced pressure. The product was extracted into EtOAc, washed with 5% AcOH and water, dried (MgSO4) and evaporated to clear oil which crystallized from EtOAc with petroleum ether to yield 4.9 g (83.1%); mp 132°-135° C; [α]D25 +21.5° (c, 1, CHCl3). Anal. Calcd for C21H32N2O6 (408.49): C, 61.74; H, 7.89; N, 6.85. Found: C, 61.61; H, 7.76; N, 6.80. the solvents removed under reduced pressure The product was extracted into EtOAc washed with 5% AcOH and water dried (MgSO4) evaporated to clear oil which crystallized from EtOAc with petroleum ether to yield 4.9 g (83.1%)

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

Procedure: To a solution of Citrulline (2.54 g, 14.50 mmole) and Sodium Bicarbonate (1.28 g) in water (40 mL) was added Boc-Val-OSu (4.34 g, 13.81 mmole) dissolved in dimethoxyethane (DME). To aid the solubility of the mixture tetrahydrofuran (10 mL) was added. The mixture thus obtained was let stir overnight at room temperature. Aqueous citric acid (15%, 75 mL) was added and the mixture was extracted with 10% 2-propanol/ethyl acetate (2×100 mL). The organic layer was washed with brine (2×150 mL) and the solvents were removed on the rotovap. The resulting white solid was dried in vacuo for 5 hours and then treated with ether (100 mL). After brief sonication and trituration, the white solid product was collected by filtration (1.39 g, 27%). 1H NMR (CD3OD) δ 0.91 (dd, 3H), 0.98 (dd, 3H), 1.44 (s, 9H), 1.70 (m, 2H), 1.87 (m, 2H), 2.02 (m, 2H), 3.11 (t, 2H), 3.89 (t, 1H), 4.39 (q, 1H), 8.22 (d, 1H) ppm; LC-MS (ESI) 375 (M+H+). was added The mixture thus obtained the mixture was extracted with 10% 2-propanol/ethyl acetate (2×100 mL) The organic layer was washed with brine (2×150 mL) the solvents were removed on the rotovap The resulting white solid was dried in vacuo for 5 hours treated with ether (100 mL) After brief sonication and trituration the white solid product was collected by filtration (1.39 g, 27%)

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

Procedure: Boc-Asp(OBzl)-Thr(Bzl)-Ser(Bzl)-Ser(Bzl)-Glu(OBzl)-OH (50.0 g, 46 mmol) was deprotected with HCl (4.15 N) in THF and worked up as usual to give 45.4 g of white solid. It was dissolved in THF (1.5 liters) and treated with ether (7 liters). On standing at 0° overnight a white solid powder was obtained (44.0 g, mp 179°-184°). Part of this material, HCl.H-Asp(OBzl)-Thr(Bzl)-Ser(Bzl)-Ser(Bzl)-Glu(OBzl)-OH (43.7 g, 42.7 mmol) was then dissolved in 500 ml of DMF, cooled to 0°, and treated with Boc-Val-OSu (15.4 g, 49 mmol) and Et3N (10 ml). The mixture was stirred for 15 hours during which time more Et3N (7.9 ml) was added in several portions maintaining the reaction slightly basic. The insoluble matter was removed by filtration and the filtrate evaporated to dryness. The oily residue was dissolved in CH2Cl2, washed with 5% HOAc, H2O, dried (Na2SO4), and evaporated to a smaller volume (0.5 liter) when treated with petroleum ether. The product crystallized slowly during overnight standing. It was recrystallized from THF and i-PrOH to yield, 26.3 g (51.4%); mp 174°-177°; [α]D25 = +0.84° (c l, THF). a white solid powder was obtained (44.0 g, mp 179°-184°) cooled to 0° was added in several portions maintaining the reaction slightly basic The insoluble matter was removed by filtration the filtrate evaporated to dryness The oily residue was dissolved in CH2Cl2 washed with 5% HOAc, H2O dried (Na2SO4) evaporated to a smaller volume (0.5 liter) when treated with petroleum ether The product crystallized slowly during overnight It was recrystallized from THF and i-PrOH to yield

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

Procedure: Boc-Val-OSu (12.6 g, 40 mmol) and H-Val-OH (4.68 g, 40 mmol) were condensed in DMF (250 ml) for 96 hours in the presence of 2 ml Et3N. More Et3N was added when needed to maintain the reaction slightly basic. The remaining insoluble material was filtered off and the filtrate evaporated to dryness (45°). The residue was partitioned between ether and dilute H2SO4 (~1%) and the organic layer washed with water (3×), dried over Na2SO4 and evaporated to a foamy glass. The product was crystallized from ether and petroleum ether: Yield, 12.2 g (96.4 %); mp 155°-158°; [α]D25 = +1.10° (c 1, DMF. to maintain the reaction slightly basic The remaining insoluble material was filtered off the filtrate evaporated to dryness (45°) The residue was partitioned between ether and dilute H2SO4 (~1%) the organic layer washed with water (3×) dried over Na2SO4 evaporated to a foamy glass The product was crystallized from ether and petroleum ether Yield

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

Procedure: A mixture of t-Boc-Val-OSu (3.14 g, 10 mmol) and H-Lys(CBZ)-OCH3.HCl (3.31 g, 10 mmol) in THF (300 ml) was treated at room temperature with triethylamine (1.38 ml) and left stirring for 17 hr. The resulting solution was evaporated in vacuo and the residue was dissolved in EtOAc (500 ml). The organic solution was washed successively with water (2×200 ml), 5% citric acid (2×200 ml), water (2×200 ml) and dried over Na2SO4. The dried solution was filtered and evaporated in vacuo to give (VI) (4.9 g; 99%) as a foam. T.l.c. RfE4 =0.64. left The resulting solution was evaporated in vacuo the residue was dissolved in EtOAc (500 ml) The organic solution was washed successively with water (2×200 ml), 5% citric acid (2×200 ml), water (2×200 ml) dried over Na2SO4 The dried solution was filtered evaporated in vacuo

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

Procedure: To a solution of Citrulline (2.54 g, 14.50 mmole) and Sodium Bicarbonate (1.28 g) in water (40 mL) was added Boc-Val-OSu (4.34 g, 13.81 mmole) dissolved in dimethoxyethane (DME). To aid the solubility of the mixture tetrahydrofuran (10 mL) was added. The mixture thus obtained was let stir overnight at room temperature. Aqueous citric acid (15%, 75 mL) was added and the mixture was extracted with 10% 2-propanol/ethyl acetate (2×100 mL). The organic layer was washed with brine (2×150 mL) and the solvents were removed on the rotovap. The resulting white solid was dried in vacuo for 5 hours and then treated with ether (100 mL). After brief sonication and trituration, the white solid product was collected by filtration (1.39 g, 27%). 1H NMR (CD3OD)δ 0.91 (dd, 3H), 0.98 (dd, 3H), 1.44 (s, 9H), 1.70 (m, 2H), 1.87 (m, 2H), 2.02 (m, 2H), 3.11 (t, 2H), 3.89 (t, 1H), 4.39 (q, 1H), 8.22 (d, 1H) ppm; LC-MS (ESI) 375 (M+H+). was added The mixture thus obtained the mixture was extracted with 10% 2-propanol/ethyl acetate (2×100 mL) The organic layer was washed with brine (2×150 mL) the solvents were removed on the rotovap The resulting white solid was dried in vacuo for 5 hours treated with ether (100 mL) After brief sonication and trituration the white solid product was collected by filtration (1.39 g, 27%)