Preparation of dipeptides
Boc-Aib-Ala-OMe (1); typical procedure
Boc-Aib-OH (0.406 g, 2 mmol) and NMM (0.110 mL, 1 mmol) were added to a vigorously stirred solution of DMT/NMM/TosO− (0.826 g, 2 mmol) in CH2Cl2 (10 mL) cooled to 0 °C. Stirring was continued until the condensing reagent disappeared (TLC analysis, staining with 0.5% solution of NBP) (1 h), after which HCl*Ala-OMe (0.278 g, 2 mmol) and NMM (0.220 mL, 2 mmol) were added. The mixture was stirred for an additional 2 h at 0 °C and left overnight at room temperature. The mixture was diluted with CH2Cl2 (10 mL), then the solution was washed successively with water, 0.5 M aqueous NaHSO4, water, 0.5 M aqueous NaHCO3 and water again. The organic layer was dried with Na2SO4, filtered and concentrated to dryness. The residue was dried under a vacuum with P2O5 and KOH to constant weight, to obtain the neutral peptide. Finally, 0.559 g was obtained, corresponding to a 97% yield.
IR (film): ν = 3678, 3608, 3430, 3017, 2965, 2398, 1734, 1672, 1512, 1451, 1420 [cm−1].
1H-NMR (250 MHz; CDCl3) δ = 1.39 (s,9H, (CH3)3-C-); 1.41 (d, 3H, J = 7.2 Hz, CH3 CH-); 1.57 (s, 6H, (CH3)2-C-); 3.72 (s, 3H, CH3O-); 4.36 (q, 1H, J = 7.2 Hz, CH3-CH-) [ppm].
Anal. RP-HPLC (10–97%B in 30 min): tR = 9.55 min (purity 98.8%).
Boc-Ala-Ala-OMe (1b)
Starting materials: Boc-Ala-OH (0.378 g, 2 mmol), HCl*Ala-OMe (0.278 g, 2 mmol), DMT/NMM/TosO− (0.826 g, 2 mmol), NMM (0.330 mL, 3 mmol). Product: Boc-Ala-Ala-OMe. Activation time: 1 h, yield 0.527 g (96%).
IR (film): ν = 3119, 3008, 1740, 1718, 1675, 1504, 1450, 1376, 1314 [cm−1].
1H-NMR (250 MHz; CDCl3) δ = 1.36 (d, 3H, J = 6.8 Hz, CH3-CH-); 1.41 (d, 3H, J = 6.8 Hz, CH3 CH-); 1.45 (s, 9H, (CH3)3-C-); 3.75 (s, 3H, CH3O-); 4.57 (q, 1H, J = 6.8 Hz, CH3 CH-); 4.61 (q, 1H, J = 6.8 Hz, CH3 CH-); 4.97 (1H, bs, -NH-), 6.49 (1H, bs, -NH-) [ppm].
Anal. RP-HPLC (10–97%B in 30 min): tR = 9.03 min (purity 98.1%).
Boc-Ser-Ala-OMe (1c)
Starting materials: Boc-Ser-OH (0.410 g, 2 mmol), HCl*Ala-OMe (0.278 g, 2 mmol), DMT/NMM/TosO− (0.826 g, 2 mmol), NMM (0.330 mL, 3 mmol). Product: Boc-Ser-Ala-OMe. Activation time: 1.5 h, yield 0.540 g (93%).
IR (film): ν = 3110, 3002, 1748, 1722, 1648, 1501, 1444, 1372, 1314 [cm−1].
1H-NMR (250 MHz; CDCl3) δ =1.38 (d, 3H, J = 7.12 Hz, CH3-CH-); 1.43 (s, 9H, (CH3)3-C-); 3.65 (dd, 2H, J1 = 6.88 Hz, J2 = 3.22 Hz, HO-CH2-); 3.72 (s, 3H, CH3O-); 4.35 (q, 1H, J = 7.1 Hz, CH3-CH-); 4.37 (t, 1H, J = 7.2 Hz, -CH-CH2-); 4.55 (t, 1H, J = 6.88 Hz, -CH2-CH-) [ppm].
Anal. RP-HPLC (10–97%B in 30 min): tR = 7.25 min (purity 98.7%).
Fmoc-Lys(Boc)-Ala-OMe (1d)
Starting materials: Fmoc-Lys(Boc)-OH (0.937 g, 2 mmol), HCl*Ala-OMe (0.278 g, 2 mmol), DMT/NMM/TosO− (0.826 g, 2 mmol), NMM (0.330 mL, 3 mmol). Product: Fmoc-Lys(Boc)-Ala-OMe. Activation time: 1 h, yield 1.041 g (94%).
IR (film): ν = 3401, 3087, 2994, 1753, 1748, 1710, 1674, 1546, 1452, 1373 [cm−1].
1H-NMR (250 MHz; CDCl3) δ = 1.41 (d, 3H, J = 7.2 Hz, CH3-CH-); 1.43 (s, 9H, (CH3)3C-); 1.48–1.55 (m, 4H, -CH2-CH2-); 1.88 (qu, 2H, J = 7.4 Hz, -CH-CH2-); 3.21 (t, 2H, J = 7.0 Hz, NH-CH2-); 3.71 (s, 3H, CH3O-); 4.36 (q, 1H, J = 7.2 Hz, CH3-CH-); 4.46 (t, 1H, J = 3.1 Hz, -CH-CH2-); 4.62 (d, 2H, J = 3.1 Hz, -CH2-O-); 7.36–8.11 (m, 8H, arom.) [ppm].
Anal. RP-HPLC (10–97%B in 30 min): tR = 10.25 min (purity 98.1%).
Fmoc-Asp(OtBu)-Ala-OMe (1e)
Starting materials: Fmoc-Asp(OtBu)-OH (0.823 g, 2 mmol), HCl*Ala-OMe (0.278 g, 2 mmol), DMT/NMM/TosO− (0.826 g, 2 mmol), NMM (0.330 mL, 3 mmol). Product: Fmoc-Asp(OtBu)-Ala-OMe. Activation time: 1 h, yield 0.934 g (94%).
IR (film): ν = 3410, 3090, 2990, 1750, 1740, 1710, 1675, 1540, 1450, 1375 [cm−1].
1H-NMR (250 MHz; CDCl3) δ = 1.40 (d, 3H, J = 7.1 Hz, CH3- CH-); 1.46 (s, 9H, -OC(CH3)3); 2.57 (dd, 1H, J1 = 17.2 Hz, J2 = 7.3 Hz, -OCO-CH2-CH-); 2.92 (dd, 1H, J1 = 17.2 Hz, J2 = 4.1 Hz, -OCO-CH2-CH-); 3.73 (s, 3H, CH3O-); 4.26 (t, 1H, J = 7.1 Hz, -CH-CH2O-); 4.42 (d, 2H, J = 7.1 Hz, -CH-CH2O-); 4.48–4.60 (m, 2H, CH3CH- + OCO-CH2-CH-); 5.97 (d, 1H, J = 7 Hz, NH); 7.07 (d, 1H, J = 5 Hz, NH); 7.31 (t, 2H, J = 7 Hz, arom.); 7.41 (t, 2H, J = 7 Hz, arom.); 7.58 (d, 2H, J = 7 Hz, arom.); 7.77 (d, 2H, J = 7 Hz, arom.) [ppm].
Anal. RP-HPLC (10–97%B in 30 min): tR = 10.05 min (purity 97.9%).
Fmoc-Trp(Boc)-Ala-OMe
Starting materials: Fmoc-Trp(Boc)-OH (1.053 g, 2 mmol), HCl*Ala-OMe (0.278 g, 2 mmol), DMT/NMM/TosO− (0.826 g, 2 mmol), NMM (0.330 mL, 3 mmol). Product: Fmoc-Trp(Boc)-Ala-OMe. Activation time: 1 h, yield 1.187 g (97%).
IR (film): ν = 3412, 3088, 2987, 1754, 1743, 1711, 1679, 1538, 1452, 1375 [cm−1].
1HNMR (250 MHz; CDCl3) δ = 1.41 (d, 3H, J = 7.2 Hz, CH3-CH-); 1.43 (s, 9H, (CH3)3C-); 3.15 (d, 2H, J = 6.7 Hz, -CH-CH2-); 3.71 (s, 3H, CH3O-); 4.91 (t, 1H, J = 6.7 Hz, CH-CH2-); 4.36 (q, 1H, J = 7.2 Hz, CH3-CH-); 4.46 (t, 1H, J = 3.1 Hz, -CH-CH2-); 4.62 (d, 2H, J = 3.1 Hz, -CH2-O-); 7.03–8.11 (m, 13H, arom.) [ppm].
Anal. RP-HPLC (10–97%B in 30 min): tR = 11.14 min (purity 98.1%).
Synthesis of 4a-f from DCMT and dipeptides
Synthesis of 2-chloro-4-methoxy-6-(NH-Aib-Ala-OMe)-1,3,5-triazine (4a). General procedure
To a vigorously stirred slurry of solid NaHCO3 (0.42 g, 5 mmol) in dichloromethane (DCM) (3 mL) cooled to 0 °C was added 2,4-dichloro-6-methoxy-1,3,5-triazine (DCMT) (0.270 g, 1.5 mmol) in DCM (3 mL), followed by the slow addition of a solution of HCl*NH-Aib-Ala-OMe (0.337 g, 1.5 mmol). Stirring was continued until complete consumption of the DCMT, as shown by the disappearance of a DCMT spot stained with 0.5% solution of NBP in ethanol at room temperature, monitored by TLC analysis. After filtration of the solid deposit, the solvent was removed using vacuum evaporation and the residue dried under a vacuum with P2O5 and KOH to a constant weight, giving 0.491 g of 2-chloro-4-methoxy-6-(NH-Aib-Ala-OMe)-1,3,5-triazine (4a), yield = 98.7% as colorless oil.
Analysis: for C12H18ClN5O4 (331.76): calculated: C 43.4%, H 5.5%, Cl 10.7%, N 21.1%, found: C 43.2%, H 5.6%, Cl 10.7%, N 21.2%.
1H-NMR (250 MHz; CDCl3) δ = 1.36 (s, 6H, ((CH3)2-C-); 1.41 (s, 3H, d, J = 7.2 Hz, CH3-CH-); 3.72 (s, 3H, CH3O-); 4.02 (s, 3H, CH3O-); 4.36 (q, 1H, J = 7.2 Hz, CH3-CH-) [ppm].
13C-NMR (75 MHz, CDCl3): δ = 17.7, 27.1, 48.1, 52.3, 54.6, 58.1, 159.7, 166.0, 172.9, 175.6 [ppm].
Synthesis of 2-chloro-4-methoxy-6-(NH-Ala-Ala-OMe)-1,3,5-triazine (4b)
Starting material: solid NaHCO3 (0.42 g, 5 mmol), DCMT (0.270 g, 1.5 mmol), HCl*H-Ala-Ala-OMe (0.316 g, 1.5 mmol). Product: 0.465 g 2-chloro-4-methoxy-6-(NH-Ala-Ala-OMe)-1,3,5-triazine (4b), yield = 97.6%, colorless oil.
Analysis: for C11H16ClN5O4 (317.73): calculated: C 41.6%, H 5.1%, Cl 11.2%, N 22%, found: C 41.5%, H 5.2%, Cl 11.2%, N 22.1%.
1H-NMR (250 MHz; CDCl3) δ = 1.39 (d, 3H, J = 7.0 Hz, CH3-CH-); 1.41 (d, 3H, J = 7.2 Hz, CH3-CH-); 3.71 (s, 3H, CH3O-); 4.02 (s, 3H, CH3O-); 4.35 (q, 1H, J = 7.2 Hz, CH3-CH-); 4.38 (q, 1H, J = 7.0 Hz, CH3-CH-) [ppm].
13C-NMR (75 MHz, CDCl3): δ = 17.4, 17.6, 47.8, 49.2, 52.3, 54.6, 159.8, 166.0, 172.2, 172.8 [ppm].
Synthesis of 2-chloro-4-methoxy-6-(NH-Ser-Ala-OMe)-1,3,5-triazine (4c)
Starting material: solid NaHCO3 (0.42 g, 5 mmol), DCMT (0.270 g, 1.5 mmol), HCl*H-Ser-Ala-OMe (0.340 g, 1.5 mmol). Product: 0.480 g 2-chloro-4-methoxy-6-(NH-Ser-Ala-OMe)-1,3,5-triazine (4c), yield = 95.8%, colorless oil.
Analysis: for C11H16ClN5O5 (333.73): calculated: C 39.6%, H 4.8%, Cl 10.6%, N 21%, found: C 39.7%, H 4.7%, Cl 10.6%, N 21%.
1H-NMR (250 MHz; CDCl3) δ = 1.41 (d, 3H, J = 7.2 Hz, CH3-CH-); 3.67 (dd, 2H, J1 = 6.8 Hz, J2 = 3.2 Hz, -CH-CH2-OH); 3.78 (s, 3H, CH3O-); 4.05 (s, 3H, CH3O-); 4.35 (1, 1H, J = 7.2 Hz, CH3-CH-); 4.48 (t, 1H, J = 6.8 Hz, -CH-CH2-) [ppm].
13C-NMR (75 MHz, CDCl3): δ = 17.6, 38.3, 47.8, 52.3, 54.6, 61.4, 159.5, 166.0171.3, 172.9 [ppm].
Synthesis of 2-chloro-4-methoxy-6-(NH-Lys(Boc)-Ala-OMe)-1,3,5-triazine (4d)
Starting material: solid NaHCO3 (0.42 g, 5 mmol), DCMT (0.270 g, 1.5 mmol), H-Lys(Boc)-Ala-OMe (0.497 g, 1.5 mmol). Product: 0.685 g 2-chloro-4-methoxy-6-(NH-Lys(Boc)-Ala-OMe)-1,3,5-triazine (4d), yield = 96.2%, colorless oil.
Analysis: for C19H31ClN6O6 (474.94): calculated: C 48%, H 6.6%, Cl 7.5%, N 17.7%, found: C 48.1%, H 6.7%, Cl 7.4%, N 17.6%.
1H-NMR (250 MHz; CDCl3) δ = 1.26–1.57 (m, 4H, -CH2-CH2-); 1.41 (d, 3H, d, J = 7.2 Hz, CH3-CH-); 1.43 (s, 9H, (CH3)3C-); 1.93 (dt, 2H, J1 = 7.5 Hz, J2 = 5.1 Hz, -CH-CH2-); 3.22 (t, 2H, J = 7.1 Hz, -CH2-NH-); 3.71 (s, 3H, CH3O-); 4.05 (s, 3H, CH3O-); 4.28 (t, 1H, J = 7.5 Hz, -CH-CH2-); 4.36 (q, 1H, J = 7.2 Hz, CH3-CH-) [ppm].
13C-NMR (75 MHz, CDCl3): δ = 17.7, 21.4, 28.2, 29.3, 29.6, 40.5, 47.8, 52.3, 53.6, 54.6, 80.5, 156.3, 159.5, 166.0, 172.5, 172.9 [ppm].
Synthesis of 2-chloro-4-methoxy-6-(NH-Asp(OtBu)-Ala-OMe)-1,3,5-triazine (4e)
Starting material: solid NaHCO3 (0.42 g, 5 mmol), DCMT (0.270 g, 1.5 mmol), H-Asp(OtBu-Ala-OMe (0.411 g, 1.5 mmol). Product: 0.607 g 2-chloro-4-methoxy-6-(NH-Asp(OtBu)-Ala-OMe)-1,3,5-triazine (4e), yield = 96.9%, colorless oil.
Analysis: for C16H24ClN5O6 (417.84): calculated: C 46%, H 5.8%, Cl 8.5%, N 16.8%, found: C 46%, H 5.9%, Cl 8.5%, N 16.8%.
1H-NMR (250 MHz; CDCl3) δ = 1.38 (s, 9H, (CH3)3C-); 1.41 (d, 3H, J = 7.2 Hz, CH3-CH-); 3.01 (d, 2H, J = 6.3 Hz, -CH-CH2-); 3.72 (s, 3H, CH3O-); 4.02 (s, 3H, CH3O-); 4.36 (1, 1H, J = 7.2 Hz, CH3-CH-); 5.02 (t, 1H, J = 6.3 Hz, -CH-CH2-) [ppm].
13C-NMR (75 MHz, CDCl3): δ = 17.7, 28.1, 37.9, 47.8, 52.3, 53.6, 54.6, 81.4, 159.6, 166.0, 172.3, 172.5, 172.9 [ppm].
Synthesis of 2-chloro-4-methoxy-6-(NH-Trp(Boc)-Ala-OMe)-1,3,5-triazine (4f)
Starting material: solid NaHCO3 (0.42 g, 5 mmol), DCMT (0.270 g, 1.5 mmol), H-Trp(Boc)-Ala-OMe (0.584 g, 1.5 mmol). Product: 0.776 g 2-chloro-4-methoxy-6-(NH-Trp(Boc)-Ala-OMe)-1,3,5-triazine (4f), yield = 97.1%, colorless oil.
Analysis: for C24H29ClN6O6 (532.98): calculated: C 54.1%, H 5.5%, Cl 6.7%, N 15.8%, found: C 54%, H 5.4%, Cl 6.7%, N 15.7%.
1H-NMR (250 MHz; CDCl3) δ = 1.41 (d, 3H, J = 7.2 Hz, CH3-CH); 1.44 (s, 9H, (CH3)3C-); 3.15 (d, 2H, J = 6.7 Hz, -CH-CH2-); 3.71 (s, 3H, CH3O-); 4.02 (s, 3H, CH3O-); 4.37 (q, 1H, J = 7.2 Hz, CH3-CH-); 4.91 (t, 1H, J = 6.7 Hz, CH-CH2-); 7.03–7.98 (m, 5H, arom) [ppm].
13C-NMR (75 MHz, CDCl3): δ = 17.7, 27.7, 28.2, 47.9, 52.3, 53.6, 54.6, 80.6, 115.4, 117.1, 118.8, 120.1, 122.8, 123.7, 129.6, 135.4, 149.2, 166.0, 162.1, 172.3, 172.9 [ppm].
Synthesis of 2-[4-(2-chloroethyl)piperazin-1-yl]-4-methoxy-6-(NH-dipeptydyl)-1,3,5-triazines 7a-f
Synthesis of 2-[4-(2-chloroethyl)piperazin-1-yl]-4-methoxy-6-(NH-Aib-Ala-OMe)-1,3,5-triazine (7a). General procedure
To a vigorously stirred solution of 2-chloro-4-methoxy-6-(NH-Aib-Ala-OMe)-1,3,5-triazine (4a) (0.332 g, 1 mmol) in DCM (10 mL) cooled to 0 °C in an ice-water bath was added 1,4-diazabicyclo[2.2.2]octane (DABCO, 5) (0.112 g, 1 mmol). Stirring at 0 °C was continued until complete consumption of 4a, as shown by TLC analysis (Rf = 0.45, DCM; staining with 0.5% solution of NBP in ethanol). The cooling bath was removed and stirring was continued at room temperature until all 6a salt was consumed, as shown by TLC analysis (Rf = 0.00, pure DCM, colored with 0.5% solution of NBP in ethanol). The solvent was removed by evaporation and the residue was dried under a vacuum with P2O5 and KOH to constant weight, giving 0.442 g 2-[4-(2-chloroethyl) piperazin-1-yl]-4-methoxy-6-(NH-Aib-Ala-OMe)-1,3,5-triazine (7a), yield = 99.1%, colorless oil.
Analysis: for C18H30ClN7O4 (443.93): calculated: C 48.7%, H 6.8%, Cl 8%, N 22.1%, found: C 48.6%, H 6.7%, Cl 8%, N 22%.
1H-NMR (250 MHz; CDCl3) δ = 1.36 (s, 6H, (CH3)2-C-); 1.41 (d, 3H, J = 7.2 Hz, CH3-CH-); 2.66 (t, 2x2H, J = 7.8 Hz, -N-CH2-); 2.85 (t, 2H, J = 6.2 Hz, -N-CH2-); 3.60 (t, 2x2H, J = 7.8 Hz, -N-CH2-); 3.86 (2, 2H, J = 6.2 Hz, Cl-CH2-); 3.71 (s, 3H, CH3O-); 4.02 (s, 3H, CH3O-); 4.36 (q, 1H, J = 7.2 Hz, CH3-CH-) [ppm].
13C-NMR (75 MHz, CDCl3): δ = 17.7, 27.1, 41.2, 44.1, 48.1, 51.8, 52.3, 53.1, 54.6, 58.1, 167.8, 171.7, 172.9, 175.6 [ppm].
LC/MS: 444.9373 ([M + H]+, C18H31ClN7O4+; calc. 443.93).
Synthesis of 2-[4-(2-chloroethyl)piperazin-1-yl]-4-methoxy-6-(NH-Ala-Ala-OMe)-1,3,5-triazine (7b)
Starting material: 2-chloro-4-methoxy-6-(NH-Ala-Ala-OMe)-1,3,5-triazine (4b) (0.318 g, 1 mmol), DABCO (5) (0.112 g, 1 mmol). Product: 0.419 g 2-[4-(2-chloroethyl)piperazin-1-yl]-4-methoxy-6-(NH-Ala-Ala-OMe)-1,3,5-triazine (7b), yield = 97.4%, colorless oil.
Analysis: for C17H28ClN7O4 (429.91): calculated: C 47.5%, H 6.6%,Cl 8.2%, N 22.8%, found: C 47.6%, H 6.7%,Cl 8.2%, N 22.7%.
1H-NMR (250 MHz; CDCl3) δ = 1.37 (d, 3H, J = 7.0 Hz, CH3-CH-); 1.41 (d, 3H, J = 7.2 Hz, CH3-CH-); 2.66 (t, 2x2H, J = 7.8 Hz, -N-CH2-); 2.85 (t, 2H, J = 6.2 Hz, -N-CH2-); 3.60 (t, 2x2H, J = 7.8 Hz, -N-CH2-); 3.86 (2, 2H, J = 6.2 Hz, Cl-CH2-); 3.71 (s, 3H, CH3O-); 4.02 (s, 3H, CH3O-); 4.35 (q, 1H, J = 7.2 Hz, CH3-CH-); 4.38 (q, 1H, J = 7.0 Hz, CH3-CH-) [ppm].
13C-NMR (75 MHz, CDCl3): δ = 17.4, 17.7, 41.2, 44.0, 47.8, 49.2, 51.8, 52.3, 53.0, 163.4, 171.1, 172.1, 172.9 [ppm].
MS: 430.9118 ([M + H]+, C17H29ClN7O4+; calc. 429.91).
Synthesis of 2-[4-(2-chloroethyl)piperazin-1-yl]-4-methoxy-6-(NH-Ser-Ala-OMe)-1,3,5-triazine (7c)
Starting material: 2-chloro-4-methoxy-6-(NH-Ser-Ala-OMe)-1,3,5-triazine (4c) (0.334 g, 1 mmol), DABCO (5) (0.112 g, 1 mmol). Product: 0.438 g 2-[4-(2-chloroethyl)piperazin-1-yl]-4-methoxy-6-(NH-Ser-Ala-OMe)-1,3,5-triazine (7c), yield = 98.2%, colorless oil.
Analysis: for C17H28ClN7O5 (445.90): calculated: C 45.8%, H 6.3%, Cl 8%, N 22%, found: C 45.7%, H 6.4%, Cl 8%, N 22%.
1H-NMR (250 MHz; CDCl3) δ = 1.40 (d, 3H, J = 7.2 Hz, CH3-CH); 2.66 (t, 2x2H, J = 7.8 Hz, -N-CH2-); 2.85 (t, 2H, J = 6.2 Hz, -N-CH2-); 3.60 (t, 2x2H, J = 7.8 Hz, -N-CH2-); 3.68 (d, 2H, J = 6.8 Hz, -CH2-OH); 3.86 (2, 2H, J = 6.2 Hz, Cl-CH2-); 3.71 (s, 3H, CH3O-); 4.02 (s, 3H, CH3O-); 4.35 (q, 1H, J = 7.2 Hz, CH3-CH-); 4.47 (t, 1H, J = 6.8 Hz, -CH-CH2-OH) [ppm].
13C-NMR (75 MHz, CDCl3): δ = 17.6, 38.3, 41.2, 44.0, 47.8, 51.8, 52.3, 53.0, 54.6, 61.4, 163.5, 171.3, 171.7, 172.9 [ppm].
MS: 446.9342 ([M + H]+, C17H29ClN7O5+; calc. 445.90).
Synthesis of 2-[4-(2-chloroethyl)piperazin-1-yl]-4-methoxy-6-(NH-Lys(Boc)-Ala-OMe)-1,3,5-triazine (7d)
Starting material: 2-chloro-4-methoxy-6-(NH-Lys(Boc)-Ala-OMe)-1,3,5-triazine (4d) (0.475 g, 1 mmol), DABCO (5) (0.112 g, 1 mmol). Product: 0.580 g 2-[4-(2-chloroethyl)piperazin-1-yl]-4-methoxy-6-(NH-Lys(Boc)-Ala-OMe)-1,3,5-triazine (7d), yield = 98.8%, colorless oil.
Analysis: for C25H43ClN8O6 (587.11): calculated: C 51.1%, H 7.4%, Cl 6%, N 19.1%, found C 51.1%, H 7.4%, Cl 6%, N 19.1%.
1H-NMR (250 MHz; CDCl3) δ = 1.26–1.57 (m, 4H, -CH2-CH2-); 1.41 (d, 3H, d, J = 7.2 Hz, CH3-CH-); 1.43 (s, 9H, (CH3)3C-); 1.93 (dt, 2H, J1 = 7.5 Hz, J2 = 5.1 Hz, -CH-CH2-); 2.66 (t, 2x2H, J = 7.8 Hz, -N-CH2-); 2.85 (t, 2H, J = 6.2 Hz, -N-CH2-); 3.22 (t, 2H, J = 7.1 Hz, -CH2-NH-); 3.60 (t, 2x2H, J = 7.8 Hz, -N-CH2-); 3.86 (2, 2H, J = 6.2 Hz, Cl-CH2-); 3.71 (s, 3H, CH3O-); 4.02 (s, 3H, CH3O-); 4.28 (t, 1H, J = 7.5 Hz, -CH-CH2-); 4.36 (q, 1H, J = 7.2 Hz, CH3-CH-) [ppm].
13C-NMR (75 MHz, CDCl3): δ = 17.7, 21.4, 28.2, 29.3, 29.9, 40.5, 41.2, 44.0, 47.8, 51.8, 52.3, 53.1, 53.6, 54.6, 80.1, 156.2, 162.2, 171.7, 172.5, 172.9 [ppm].
MS: 588.2016 ([M + H]+, C25H44ClN8O6+; calc. 587.11).
Synthesis of 2-[4-(2-chloroethyl)piperazin-1-yl]-4-methoxy-6-(NH-Asp(OtBu)-Ala-OMe)-1,3,5-triazine (7e)
Starting material: 2-chloro-4-methoxy-6-(NH-Asp(OtBu)-Ala-OMe)-1,3,5-triazine (4e) (0.418 g, 1 mmol), DABCO (5) (0.112 g, 1 mmol). Product: 0.520 g 2-[4-(2-chloroethyl)piperazin-1-yl]-4-methoxy-6-(NH-Asp(OtBu)-Ala-OMe)-1,3,5-triazine (7e), yield = 98.2%, colorless oil.
Analysis: for C22H36ClN7O6 (530.02): calculated: C 49.9%, H 6.8%, Cl 6.7%, N 18.5%, found: C 49.8%, H 6.7%, Cl 6.7%, N 18.5%.
1H-NMR (250 MHz; CDCl3) δ = 1.38 (s, 9H, (CH3)3C-); 1.41 (d, 3H, J = 7.2 Hz, CH3-CH-); 3.01 (d, 2H, J = 6.3 Hz, -CH-CH2-); 2.66 (t, 2x2H, J = 7.8 Hz, -N-CH2-); 2.85 (t, 2H, J = 6.2 Hz, -N-CH2-); 3.60 (t, 2x2H, J = 7.8 Hz, -N-CH2-); 3.86 (2, 2H, J = 6.2 Hz, Cl-CH2-); 3.71 (s, 3H, CH3O-); 4.02 (s, 3H, CH3O-); 4.36 (1, 1H, J = 7.2 Hz, CH3-CH-); 5.02 (t, 1H, J = 6.3 Hz, -CH-CH2-) [ppm].
13C-NMR (75 MHz, CDCl3): δ = 17.7, 37.9, 41.1, 44.0, 47.8, 51.8, 52.3, 53.1, 53.5, 54.6, 81.1, 162.2, 171.7, 172.1, 172.5, 172.9 [ppm].
MS: 531.1256 ([M + H]+, C22H37ClN7O6+; calc. 530.02).
Synthesis of 2-[4-(2-chloroethyl)piperazin-1-yl]-4-methoxy-6-(NH-Trp(Boc)-Ala-OMe)-1,3,5-triazine (7f)
Starting material: 2-chloro-4-methoxy-6-(NH-Trp(Boc)-Ala-OMe)-1,3,5-triazine (4f) (0.533 g, 1 mmol), DABCO (5) (0.112 g, 1 mmol). Product: 0.638 g 2-[4-(2-chloroethyl)piperazin-1-yl]-4-methoxy-6-(NH-Trp(Boc)-Ala-OMe)-1,3,5-triazine (7f), yield = 98.9%, colorless oil.
Analysis: for C30H41ClN8O6 (645.15): calculated: C 55.9%, H 6.4%, Cl 5.5%, N 17.4%, found: C 55.9%, H 6.4%, Cl 5.5%, N 17.4%.
1H-NMR (250 MHz; CDCl3) δ = 1.41 (d, 3H, J = 7.2 Hz, CH3-CH-), 1.44 (9H, s, (CH3)3-C-), 2.66 (t, 2x2H, J = 7.8 Hz, -N-CH2-); 2.85 (t, 2H, J = 6.2 Hz, -N-CH2-); 3.15 (d, 2H, J = 6.7 Hz, -CH2-CH-); 3.60 (t, 2x2H, J = 7.8 Hz, -N-CH2-); 3.86 (2, 2H, J = 6.2 Hz, Cl-CH2-); 3.72 (s, 3H, CH3O-); 4.02 (s, 3H, CH3O-); 4.36 (q, 1H, J = 7.2 Hz, CH3-CH-); 4.91 (t, 1H, J = 6.7 Hz, -CH2-CH-); 7.03–7.98 (m, 5H, arom) [ppm].
13C-NMR (75 MHz, CDCl3): δ = 17.7, 27.7, 28.1, 41.2, 44.1, 47.9, 51.8, 52.3, 53.0, 53.6, 54.5, 81.1, 115,1, 117.1, 118.7, 120.6, 122.7, 123.7, 129.6, 135.4, 149.5, 163.3, 172.4, 172.9 [ppm].
MS: 646.15864 ([M + H]+, C30H42ClN8O6+; calc. 645.15).