Journal of Medicinal Chemistry Ethyl 2-[(1-tert-Butoxycarbonylpiperidin-3-yl)amino]acetate ((rac)-1). NaBH(OAc)3 (5.3 g, 25.0 mmol) was added to a solution of 1-Boc-piperidin-3-one (4.98 g, 25.0 mmol), acetic acid (2.25 g, 37.5 mmol), glycine ethyl ester hydrochloride (3.5 g, 25.0 mmol), and molecular sieves (4.5 g) in dichloromethane (100 mL). The reaction mixture was stirred at room temperature for 20 h, diluted with dichloromethane (100 mL), and quenched by the addition of aqueous NaOH (3 M) (20 mL). The aqueous phase was extracted with dichloromethane (2 × 100 mL). The combined organic phases were dried (Na2SO4), filtered, and concentrated in vacuo. Purification by flash chromatography (heptane/ethyl acetate 1:2, Rf = 0.18) gave (rac)-1 as a transparent oil (4.4 g, 62%). Resolution of a portion of the racemate (2.0 g) by chiral preparative HPLC gave the enantiomers of 1. (−)-Ethyl 2-[(1-tert-Butoxycarbonyl-(R)-piperidin-3-yl)- amino]acetate ((R)-1). Resolution of (rac)-1 (2.0 g) gave enantiomerically pure (R)-1 (779 mg). Analytical chiral HPLC (hexane/2-propanol 95:5) tR 9.4 min; 95% purity, ee 100%. 1H NMR δ 4.18 (q, J = 7.1 Hz, 2H), 3.80−3.73 (m, 2H), 3.55−3.41 (AB system, 2H), 2.86−2.51 (m, 3H), 2.20−1.80 (m, 2H), 1.73−1.64 (m, 1H), 1.47−1.21 (m, 2H), 1.44 (s, 9H), 1.26 (t, J = 7.1 Hz, 3H). 13C NMR δ 172.2, 154.6, 79.2, 60.6, 53.3, 48.6, 48.1, 43.9, 31.1, 28.2, 23.2, 14.0; [α]D20 −8.5 (c 1.0, CHCl3). IR (neat) v 3515, 3333, 2976, 2933, 2860, 1738, 1691, 1423, 1157 cm−1. HRMS (FT-ICR-MS) calcd for C14H26N2O4 [M + H]+ 287.1965; found 287.1962. For NMR spectral assignments, see the Supporting Information. (+)-Ethyl 2-[(1-tert-Butoxycarbonyl-(S)-piperidin-3-yl)- amino]acetate ((S)-1). Resolution of (rac)-1 (2.0 g) gave enantiomerically pure (S)-1 (802 mg). Analytical chiral HPLC (hexane/2-propanol 95:5) tR 8.7 min, 94% purity, ee 100%. 1H NMR δ 4.18 (q, J = 7.1 Hz, 2H), 3.81−3.73 (m, 2H), 3.51−3.37 (AB system, 2H), 2.93−2.48 (m, 3H), 2.03−1.80 (m, 3H), 1.73−1.64 (m, 1H), 1.49−1.21 (m, 10H), 1.26 (t, J = 7.1 Hz, 3H). 13C NMR δ 172.2, 154.6, 79.1, 60.5, 53.3, 48.9, 48.1, 43.9, 31.0, 28.2, 23.2, 13.9; [α]D20 +8.5 (c 1.0, CHCl3). IR (neat) v 3515, 3333, 2976, 2933, 2860, 1738, 1691, 1423, 1157 cm−1. HRMS (FT-ICR-MS) calcd for C14H26N2O4 [M + H]+ 287.1965; found 287.1962. Both enantiomers of 1 were also obtained separately by reductive alkylation of 3-amino-1-tert-butyloxycarbonylpiperidine with defined stereochemistry. These products were used as reference material. (−)-Ethyl 2-[(1-tert-Butoxycarbonyl-(R)-piperidin-3-yl)- amino]acetate ((R)-1). Ethyl glyoxylate in toluene (50%) (0.5 mL, 2.5 mmol) was added dropwise to a mixture of (R)-3-amino-1-tert- butyloxycarbonylpiperidine (500 mg, 2.5 mmol) and Na2SO4 (900 mg) in dichloromethane (9 mL). The reaction mixture was stirred at room temperature for 2 h. NaBH(OAc)3 (529 mg, 2.5 mmol) and AcOH (225 mg, 3.74 mmol) were added, and the stirring continued for 90 min. The mixture was diluted with dichloromethane (20 mL) and quenched by addition of aqueous NaOH (3 M) (8 mL). The aqueous phase was extracted with dichloromethane (1 × 20 mL). The combined organic phases were dried (Na2SO4) and filtered and the solvent was removed in vacuo. Purification by flash chromatography (heptane/ethyl acetate 1:3, Rf = 0.17) gave (R)-1 as a transparent oil (410 mg, 60%). Analytical chiral HPLC (hexane/2-propanol 95:5) tR 9.3 min; ee 90%. NMR data were in agreement with those reported for the pure enantiomer (R)-1. (+)-Ethyl 2-[(1-tert-Butoxycarbonyl-(S)-piperidin-3-yl)- amino]acetate ((S)-1). Compound (S)-1 was synthesized from (S)- 3-amino-1-tert-butyloxycarbonylpiperidine (500 mg, 2.5 mmol) using the same procedure as described for (R)-1 to afford (S)-1 as a transparent oil (380 mg, 56%). Analytical chiral HPLC (hexane/2- propanol 95:5) tR 8.5 min; ee 86%. NMR data were in agreement with those reported for the pure enantiomer (S)-1. (−)-2-[(1-tert-Butoxycarbonyl-(R)-piperidin-3-yl)amino]- acetamide ((R)-2). Compound (R)-2 was synthesized from (R)-1 (210 mg, 0.73 mmol) and NH3 in methanol (7 N) (4.2 mL) as described for (S)-2 to give the product as a white solid (174 mg, 92%). Analytical HPLC (hexane/ethanol 97:3) tR 84.4 min, purity 100%. 1H NMR δ 7.05 (br s, 2H), 5.53 (br s, 1H), 3.97−3.54 (m, 2H), 3.38− 3.25 (AB system, 2H), 3.16−2.67 (m, 2H), 2.56 (app tt, J = 8.1, 8.1, Article 3.7, 3.7 Hz, 1H), 1.97−1.84 (m, 1H), 1.73−1.63 (m, 1H), 1.51−1.23 (m, 2H), 1.45 (s, 9H). 13C NMR δ 175.1, 154.6, 79.2, 53.7, 49.5, 48.7, 43.8, 30.9, 28.1, 22.9; [α]D20 −4.6 (c 1.0, CHCl3). IR (KBr) v 3398, 2977, 2933, 1683, 1638, 1423, 1262, 1180, 1155 cm−1. HRMS (FT- ICR-MS) calcd for C12H23N3O3 [M + H]+ 258.1812; found 258.1809. For NMR spectral assignment, see the Supporting Information. (+)-2-[(1-tert-Butoxycarbonyl-(S)-piperidin-3-yl)amino]- acetamide ((S)-2). NH3 in methanol (7 N) (4.3 mL) was added to (S)-1 (214 mg, 0.75 mmol), and the reaction mixture was stirred at room temperature for 72 h. The solvent and excess of reagent were removed in vacuo. The remaining was trituated with hexane/diethyl ether (1:1) (1 mL) to give (S)-2 as a white solid (173 mg, 90%). Analytical chiral HPLC (hexane/ethanol 97:3) tR 76.0 min, purity 100%. 1H NMR δ 7.06 (br s, 2H), 5.62 (br s, 1H), 3.94−3.56 (m, 2H), 3.39−3.24 (AB system, 2H), 3.32−2.72 (m, 2H), 2.56 (app tt, J = 8.1, 8.1, 3.7, 3.7 Hz, 1H), 1.95−1.86 (m, 1H), 1.73−1.57 (m, 2H), 1.45 (s, 9H), 1.39−1.23 (m, 2H). 13C NMR δ 175.0, 154.5, 79.1, 53.6, 49.4, 48.6, 43.7, 30.8, 28.0, 22.8; [α]D20 +5.1 (c 1.0, CHCl3). IR (KBr) v 3398, 2977, 2933, 1683, 1638, 1423, 1262, 1180, 1155 cm−1. HRMS (FT-ICR-MS) calcd for C12H23N3O3 [M + H]+ 258.1812; found 258.1810. (−)-2-((R)-Piperidin-3-ylamino)acetamide × 2TFA ((R)-pipG) ((R)-3). Deprotection of (R)-2 (137 mg, 0.53 mmol) using TFA (364 mg, 3.2 mmol) was performed as described for (S)-pipG ((S)-3) to give the TFA salt of (R)-pipG ((R)-3) as a white solid (210 mg, quant). 1H NMR (CD3OD) δ 4.00−3.88 (AB system, 2H), 3.73 (dd, J = 12.1, 4.0 Hz, 1H), 3.63−3.51 (app tt, J = 11.4, 11.4, 4.1, 4.1 Hz, 1H), 3.40 (app dd, J = 12.9, 3.6 Hz, 1H), 3.08 (app t, J = 11.7 Hz, 1H), 2.96 (dt, J = 12.7, 12.7, 3.2 Hz, 1H), 2.32−2.24 (m, 1H), 2.14−2.04 (m, 1H), 1.87−1.78 (m, 1H), 1.71 (app ddt, J = 12.5, 12.5, 3.6 Hz, 1H). 13C NMR (CD3OD) δ 168.5, 53.0, 46.6, 45.0, 44.6, 26.2, 21.6; [α]D20 −9.3 (c 0.41, CH3OH). IR (neat) v 3399, 3012, 2822, 1683, 1436, 1205, 1130 cm−1. HRMS (FT-ICR-MS) Calcd for C7H15N3O [M + H]+ 158.1288; found 158.1287; pKa1 = 5.13, pKa2 = 8.46. For NMR spectral assignment, see the Supporting Information. (+)-2-((S)-Piperidin-3-ylamino)acetamide × 2TFA ((S)-pipG) ((S)-3). TFA (375 mg, 3.3 mmol) was added to (S)-2 (141 mg, 0.55 mmol) in dichloromethane (3 mL), and the reaction mixture was stirred at room temperature for 12 h. The solvent and excess of reagent were removed in vacuo to give the TFA-salt of (S)-pipG ((S)- 3) as a white solid (210 mg, quant). 1H NMR (CD3OD) δ 4.00−3.85 (AB system, 2H), 3.76−3.65 (m, 1H), 3.56 (app tt, J = 11.4, 11.4, 3.7, 3.7, 1H), 3.44−3.34 (m, 1H), 3.08 (app t, J = 11.7 Hz, 1H), 2.96 (app dt, J = 12.8, 12.8, 3.3 Hz, 1H,), 2.33−2.21 (m, 1H), 2.15−2.01 (m, 1H), 1.88−1.60 (m, 2H). 13C NMR (CD3OD) δ 168.4, 53.0, 46.6, 45.0, 44.6, 26.2, 21.6; [α]D20 +6.6 (c 0.41, CH3OH). IR (neat) v 3383, 3014, 2824, 1676, 1432, 1202, 1130 cm−1. HRMS (FT-ICR-MS) calcd for C7H15N3O [M + H]+ 158.1288; found 158.1287. The purity was 90% according to UPLC analysis. pKa1 = 5.21, pKa2 = 8.53. (−)-Ethyl 2-[(Benzyloxycarbonyl)(1-tert-butoxycarbonyl-(R)- piperidin-3-yl)amino]acetate ((R)-4). Compound (R)-4 was s ynthesiz e d fro m (R )- 1 (802 mg, 2.8 mmol) a nd N - (benzyloxycarbonyloxy)succinimide (698 mg, 2.8 mmol) with Et3N (312 mg, 3.1 mmol) in dichloromethane (11 mL) as described for (S)- 4 to give the product as a transparent oil (0.98 g, 82%). Analytical chiral HPLC (hexane/2-propanol 95:5) tR 19.8 min, 98% purity, ee 100%. 1H NMR (rotamers) δ 7.59−7.20 (m, 5H), 5.19 and 5.10 (rot, s, 2H), 4.34−3.72 (m, 6H), 2.80−2.43 (m, 3H), 2.03−1.87 (m, 1H), 1.80−0.98 (m, 1H), 1.43−1.17 (m, 2H), 1.43 and 1.37 (rot, s, 9H), 1.26 and 1.17 (rot, t, J = 7.1 Hz, 3H). 13C NMR δ 169.7, 155.8, 155.1, 154.5, and 154.3 (rot), 136.1, 128.3, and 128.2 (rot), 127.8, 127.5, 79.4, 67.4, and 67.1 (rot), 60.9, 53.1, and 52.9 (rot), 47.0 and 46.5, 45.6 and 45.3, 43.2, 28.7, 28.1, 24.4, 13.9; [α]D20 −14.9 (c 1.0, CHCl3). IR (neat) v 3500, 2976, 2864, 1749, 1693, 1416, 1152 cm−1. HRMS (FT-ICR-MS) calcd for C22H32N2O6 [M + H]+ 421.2333; found 421.2333. For NMR spectral assignment, see the Supporting Information. (+)-Ethyl 2-[(Benzyloxycarbonyl)(1-tert-butoxycarbonyl-(S)- piperidin-3-yl)amino]acetate ((S)-4). Et3N (290 mg, 2.9 mmol) in dichloromethane (2 mL) was added to a solution of (S)-1 (745 mg, 2556 dx.doi.org/10.1021/jm201517a | J. Med. Chem. 2012, 55, 2549−2560