To a solution of 2-hydroxy-2-(2-pyridyl)acetic acid from the previous step (160 mg as a 1:1 mixture with NaCl, 0.71 mmol), N-[5-chloro-2-(1H-tetraazol-1-yl)benzyl]-L-prolinamide (215 mg, 0.71 mmol, HPLC RT=2.25 min, method A), and HOBt hydrate (110 mg, 0.71 mmol) in DMF (5 mL) was added EDC (175 mg, 0.92 mmol). Diisopropylethylamine was then added slowly in portions (˜0.1 mL total) to bring the pH of the solution to 6–7 as measured on wetted E. Merck pH indicator strips. The mixture was stirred at ambient temperature for 18 h, and the solvent was removed under reduced pressure. The residue was partitioned between EtOAc (75 mL) and saturated aqueous NaHCO3 (20 mL). The EtOAc layer was separated, dried over anhydrous MgSO4, and filtered. The filtrate solvent was removed under reduced pressure and the residue was purified by pressurized silica gel column chromatography using a gradient elution of 2%, 3%, 4%, 5%, 6% MeOH in dichloromethane. Product-containing fractions were combined and the solvent was removed under reduced pressure to give two diastereomers of the title compound, both as amorphous solids (first eluting diastereomer: 96 mg; TLC Rf=0.3 (96:4 CH2Cl2:MeOH); HPLC RT=2.32 min, method A; LC-MS m/z=456; second eluting diastereomer: TLC Rf=0.2 (96:4 CH2Cl2:MeOH); HPLC RT=2.33 min, method A; LCMS m/z=456).
