Scaled-up Synthesis and Characterization of Oxytocin Trisulfide

The title peptide, oxytocin trisulfide (3), was required on a multi-100 mg scale, as part of an analytical method validation process for the clinically and commercially important product “Oxytocin Injection,” which is a sterile isotonic injectable formulation of oxytocin (2). We report here that previous chemistry from our academic laboratory can be successfully scaled up in the biotechnology sector to indeed provide pure 3 (overall yield 6–7%, > 95% purity). The scaled-up synthesis also gave rise, somewhat unexpectedly, to modest levels of oxytocin tetrasulfide (4), and even higher homologues. Protocols for synthesis, purification, and characterization [by HPLC, mass spectrometry and amino acid analysis] are described and discussed, along with possible mechanistic explanations for our findings.