Fabrication of a guanidine ligand-based anion exchange monolithic stationary phase in a 3D printed polypropylene housing for protein chromatography

Abstract

A monolithic anion exchange support is synthesized based on the incorporation of γ -guanidinobutyric acid an alkaloid with three resonant amino groups- as ligand onto a poly(EDMA-co-GMA) monolith. The created monolithic anion exchange support is referred as M-Gnd. Monolith was synthesized in a one-step polymerization reaction in a designed and printed polypropylene housing within a 5x20mm i.d. channel. Functionalization of the poly-methacrylate monolith with γ -guanidinobutyric acid was done by Shift-based method using diethylenediamine as a spacer. Homogeneous surface morphology was appreciated by SEM image, and FTIR analysis confirmed the presence of ligand. Highest value of Dynamic binding capacity (12.7mg BSA/mL monolith) was obtained at 0.5 mL/min when a 2 mg/mL protein concentration was used. The estimated ligand for M-Gnd was 1.23 mmol/g dry support. The M-Gnd support allowed to purity a red fluorescent protein up to 75%, presenting a high efficiency (N and HETP values). The characterized M-Gnd showed positive results as anion exchange chromatography support for protein purification. However, the functionalization of γ-guanidinobutyric acid can be improved to have a comparable chromatographic performance to commercial anion exchange monolith.