In the control experiment where no extraction was performed during the final step, peptide alkyl thioester 6a was obtained in a low yield (Fig. , green dots), highlighting the significance of eliminating the excess p-hydroxythiophenol to attain yields suitable for practical applications. Figure further illustrates that the thiol-thioester exchange process between pre-synthesized peptide aryl thioester 4 and 5a progressed at a slower rate compared to the adapted hydrazide method, reaching similar equilibrium position after more than 500 minutes of reaction (Fig. , black dots). Intriguingly, performing the thiol-thioester exchange reaction between 4 and 5a while extracting the released phydroxythiophenol had a minor yet positive impact on the rate (see Supplementary Information). All these experiments demonstrate that while it is essential to extract the excess p-hydroxythiophenol during the final step of the adapted hydrazide method to achieve satisfactory yields in peptide alkyl thioester, the extraction procedure alone does not explain the rapid rate of its formation.