Five Agilent 1290 Infinity II UHPLCs (Agilent Technologies, Santa Clara, CA, USA) were used for retention time measurement of the 4549 macrocyclic peptides from physiochemical screening. All UHPLCs were equipped with Supelco Ascentis Express C18 (Merck KGaA, Darmstadt, Germany) columns (2.7 µm, 3.0 × 30 mm). A 1.3 minute gradient was used, with acetonitrile (Fisher Scientific, Fair Lawn, NJ, USA) as the organic phase and a pH 7 potassium phosphate buffer (Merck KGaA) as the aqueous phase and with the column temperature at 30 ºC. Injections were done at 2 µL volumes with spectrophotometric detection monitoring wavelengths of 215 and 238 nm.