Microwave-Assisted Automated Peptide Synthesis

Microwave-assisted automated solid-phase peptide synthesis (Liberty Blue CEM Corporation, Matthews, NC, USA) was used to synthesize PS1-2, PS1-5, and PS1-6 (PS1-2: KWYKKWYKKWYK-CONH₂, PS1-5: RWYRRWYRRWYR-CONH₂, and PS1-6: KWLKKWLKKWLK-CONH₂). Rink Amide resin (Novabiochem) (0.55 mmol/g) was used and Fmoc deprotection was assessed by 20% piperidine in dimethylformamide (DMF). Each coupling step of Fmoc amino acids was achieved using microwave heating in the presence of DIC and Oxyma pure in DMF. Resin-synthesized peptide was transferred to a conical tube, washed with dichloromethane, and allowed to air-dry. Peptide cleavage was performed by treatment with trifluoroacetic acid (TFA)/triisopropylsilane/DiH₂O (95:2.5:2.5, v/v/v) for 2 h at room temperature. The cleaved peptide–TFA solution was precipitated with diethyl ether and then dried under a vacuum pump (Edwards RV5, Seongnam-si, Korea) to obtain a powder. The synthesized peptides were purified using a Zorbax C₁₈ column (21.2 × 250 mm, 300 Å, 7 μm) on a Shimadzu Preparative HPLC system (Kyoto, Japan) using 5%–60% acetonitrile gradient in water with 0.05% TFA. Molecular masses were confirmed using a matrix-assisted laser desorption ionization mass spectrometer (MALDI II, Kratos Analytical Ltd., Manchester, UK) [34 (link)].