Characterization of chemically synthesized human relaxin by high-performance liquid chromatography

Highly purified human relaxin, produced by combining chemically synthesized A- and B-chains, was analyzed by reversed-phase high-performance liquid chromatography, ion-exchange chromatography and tryptic mapping in order to ascertain purity of the material, presence of uncleaved protecting groups, correctness of disulfide linkages and presence of deamidated or oxidized variants. It was shown by a variety of analytical methods that the product was of high purity; a minimum purity level as judged by the most discriminating assay was greater than 98%. Components of the relaxin preparation removed during the purification were identified to be variants containing deletions arising from incomplete coupling reactions in the solid phase peptide synthesis and/or oxidized methionine residues.