(M1330-01-03) FM1000 Surfactant is Compatible with Downstream Bioprocessing

Purpose: Proteins, such as monoclonal antibodies, are subjected to various stresses during downstream processing which can impact product quality. Generally, surfactants cannot be utilized in downstream unit operations due to poor filterability and an inability to remove prior to final formulation.¹ This work explored whether the novel surfactant, N-myristoyl phenylalanine polyether diamide (FM1000) could be compatible with downstream processing steps.

Methods: FM1000 was synthesized as previously reported.^2,3 All other materials were from commercially available sources. IgG was bovine IgG from MP Biomedical. Surfactant concentrations were determined using an HPLC-CAD method developed in-house, and protein quantification was done using SEC with a UV detector. UF/DF was performed on a Minimate TFF system (Pall). Water contact angles were measured on a Ossila goniometer following immersion of the substrate in a surfactant-IgG solution. Surfactant binding to filters and columns was measured by passing solutions over the material in question using a syringe pump and measuring concentration on aliquots collected.

Results: Pumping studies showed a greater than 10-fold decrease in sub-visible particle generation with FM1000 present compared to control samples. FM1000 was able to be removed from a protein solution via diafiltration and did not appreciably concentrate during subsequent ultrafiltration. In contrast, PS80 was not effectively removed under the same conditions. FM1000, at 0.05 mg/mL, dropped in concentration by 94%, while the PS80 dropped by only 36%. In measuring the ability of IgG to foul various surfaces, FM1000 was effective at blocking fouling at lower concentrations than both PS80 and PS20, with effectiveness below 0.02 mg/mL. FM1000 also bound 25% less to filters (i.e. syringe filters) and eluted consistently from the filters as a single HPLC peak, whereas the polysorbates eluted as multiple peaks each at different rates. Finally, FM1000 did not interfere with IgG’s ability to bind to ProteinA.

Conclusion: This study explored the use of FM1000 in various applications related to downstream processing operations. Unlike with polysorbates, FM1000 can be effectively and efficiently removed during the buffer exchange step of UF/DF. In pumping studies, FM1000 kept particle counts in IgG solutions at levels close to non-pumped controls. FM1000 is effective at preventing protein adsorption on a variety of pharma-relevant surface chemistries, and FM1000 concentration recovers uniformly and quickly after flowing through filters and columns. Its inclusion was also shown to have no interference with protein yield in bind-and-elute experiments. Together, these observations indicate that the scope of usability for FM1000 includes final formulations as well as downstream unit operations where improved efficiency, higher yields, or higher protein quality is required. Future studies can further build on these investigations to account for specific unit operations and the diverse chemistries of various antibodies to identify the specific applications that would bring the most benefit to a process engineer.

References: 1. Katz, J.S.; Chou, D.K.; Christian, T.R.; Das, T.K.; Patel, M.; Singh, S.N.; and Wen, Y. “Emerging Challenges and Innovations in Surfactant-Mediated Stabilization of Biologic Formulations.” Journal of Pharmaceutical Sciences. 2022, 111, 4, 919-932.
2. Katz, J.S.; Nolin, A.; Yezer, B.A.; and Jordan, S. “Dynamic Properties of Novel Excipient Suggest Mechanism for Improved Performance in Liquid Stabilization of Protein Biologics.” Mol. Pharmaceutics, 2019, 16, 1, 282-291.
3. Katz, J.S.; Tan, Y.; Kuppannan, K.; Song, Y.; Brennan, D.J.; Young, T.; Yao, L.; and Jordan, S. “Amino-Acid-Incorporating Nonionic Surfactants for Stabilization of Protein Pharmaceuticals.” ACS Biomaterials Science & Engineering. 2016. 2:7, 1093-1096.


Subvisible particle formation following 24 hours pumping stress of an IgG solution with and without FM1000 present.


Removal of FM1000 and PS80 during diafiltration (solid symbols) and concentration during ultrafiltration (hollow symbols). FM1000 shows significantly better removal during diafiltration compared to PS80.


FM1000 concentration impact on IgG recovery during ProA bind and elute study. FM1000 had no impact on protein recovery when included in bind or elute steps.