Peptide mols. bearing disulfide moieties constitute an important category of therapeutics. The establishment of the disulfide structure plays a pivotal role in the production of those peptides. In-solution cyclization is generally selected in industrial peptide production as a synthetic strategy to build disulfide from the linear deprotected precursor. This methodol. is inherently limited by the relatively low productivity, considering the minimizing intermol. oligomerization by dilution of the overall concentration of the reaction. Alternatively, the disulfide bond could be established from the thiol-protected building blocks directly on the solid support. On-resin disulfide construction was pursued for the atosiban process development. Protected linear atosiban resin was treated by I2/DMF, and the target disulfide was built rapidly and quant. within 30 min. The effects of the resin linker and thiol-protecting groups were tested by utilizing Sieber resin, Rink amide resin, and Ramage resin as solid support and Cys(Trt)/Cys(Acm) as building blocks. The I2 equivalence, as a potential critical process parameter, was subjected to a PAR (proven acceptable range) study to facilitate the envisioned large-scale manufacturing Although there was rapid and complete disulfide formation on the solid support in all tested combinations of resin and thiol-protecting groups, a reduction of the disulfide was observed in the processes of peptidyl resin cleavage/global deprotection, with the exception of the Sieber resin strategy. A dedicated investigation revealed that hydrosilane, employed as a cation scavenger in the peptide synthesis, is responsible for the disulfide degradation in the presence of TFA (trifluoroacetic acid). Eliminating silane from the TFA cleavage solution or subtly controlling its equivalence could effectively suppress the nocuous disulfide reduction in this process. In summary, on-resin disulfide cyclization has been verified to be a viable strategy for atosiban industrial production by this study.