%0 Journal Article
%T Large-scale purification of a deprotected macrocyclic peptide by supercritical fluid chromatography (SFC) integrated with liquid chromatography in discovery chemistry.
%A Li P
%A Wu DR
%A Yip SH
%A Sun D
%A Pawluczyk J
%A Smith A
%A Kempson J
%A Mathur A
%J J Chromatogr A
%V 1730
%N 0
%D 2024 Aug 16
%M 38972253
%F 4.601
%R 10.1016/j.chroma.2024.465112
%X A macrocyclic peptide A was successfully purified in large quantities (∼30 g) in >95 % purity by an integrated two-step orthogonal purification process combining supercritical fluid chromatography (SFC) with medium-pressure reverse-phase liquid chromatography (MP-RPLC). MP-RPLC was used to fractionate the crude peptide A, remove unwanted trifluoroacetic acid (TFA) originating from the peptide A cleavage off the resin, and convert the peptide A into ammonium acetate salt form, prior to the final purification by SFC. A co-solvent of methanol/acetonitrile containing ammonium acetate and water in CO2 was developed on a Waters BEH 2-Ethylpyridine column. The developed SFC method was readily scaled up onto a 5 cm diameter column to process multi-gram quantities of the MP-RPLC fraction to reach > 95 % purity with a throughput/productivity of 0.96 g/h. The incorporation of SFC with MP-RPLC has been demonstrated to have a broader application in other large-scale polypeptide purifications.