Optimizing Industrial Solid-Phase Peptide Synthesis: Integration of Raman Spectroscopy as Process Analytical Technology

Peptide therapeutics have exploded in popularity in recent years, motivating the need for advanced manufacturing methods which can be applied across the solid-phase peptide synthesis (SPPS) process. The Food and Drug Administration's Process Analytical Technology (PAT) initiative offers a platform to implement advanced methods to improve the efficiency and understanding of pharmaceutical manufacturing processes and shows great promise in application toward industrial SPPS. In this work, Raman spectroscopy was used as the main PAT tool to implement methods for on-line and real-time monitoring of the entire SPPS process, from Fmoc removal, to coupling, and through the extensive solvent-washing steps. Raman spectroscopy is a rapid, specific, and nondestructive technique that can provide rich real-time information for SPPS processes and can be used to improve efficiency in solvent use and save process time. Specifically, this work reports on the development of PAT methods for monitoring of amino acid coupling during the coupling stage and residual piperidine concentration during the post-deprotection washing stage of SPPS, to help reduce solvent use and better understand the coupling reaction and its time frame. We show a significant reduction in solvent use is possible by employing Raman spectroscopy along with a partial least squares model to predict the piperidine concentration in real time during continuous wash. In addition, Raman spectroscopy offers a greater understanding of coupling reaction kinetics during SPPS, which could lead to significant improvements in total SPPS process time.