Compatibility of column inlet and adsorbent designs for processing of corn endosperm extract by expanded bed adsorption

• Published in: Biotechnology and bioengineering 2004-08, Vol.87 (3), p.324-336
• Main Authors: Menkhaus, T.J, Glatz, C.E
• Format: Article
• Language: English
• Subjects: Adsorption; Biotechnology; Chromatography; Chromatography, Ion Exchange - instrumentation; Chromatography, Ion Exchange - methods; column design; Contaminants; Corn; corn endosperm; corn solid elution; EBA resin; Endosperm; Equipment Design; Equipment Failure Analysis; expanded bed adsorption (EBA); Gene expression; Ion exchange; Ion Exchange Resins - chemistry; Particle Size; Plant Extracts - chemistry; Plant Extracts - isolation & purification; Plant Proteins - chemistry; Plant Proteins - genetics; Plant Proteins - isolation & purification; Protein Binding; protein purification; Proteins; Q1; Recombinant Proteins - chemistry; Recombinant Proteins - isolation & purification; Resins; Salts; Sedimentation; Seeds - embryology; Seeds - genetics; Seeds - metabolism; Styles; Substance P; Transgenic plants; Ultrafiltration - instrumentation; Ultrafiltration - methods; Velocity; Zea mays - embryology; Zea mays - genetics; Zea mays - metabolism
• ISSN: 0006-3592; 1097-0290
• DOI: 10.1002/bit.20117

Corn has emerged as a viable host for expression of recombinant proteins; targeted expression to the endosperm has received particular attention. The protein extracts from corn endosperm differ from those of traditional hosts in regard to the nature of residual solids and extracted matrix contaminants. Each of these differences presents reasons for considering expanded bed adsorption for product capture and new considerations for limitations of the method. In this work three inlet-flow distribution devices (mesh, glass ballotini, and localized mixing) and six adsorbents with different physical (size and density), chemical (ligand), and base matrix properties were evaluated to determine conditions compatible with processing of crude corn endosperm extract by expanded bed adsorption. Of the inlet devices evaluated, the design with localized mixing at the inlet (as produced commercially by UpFront Chromatography A/S, Copenhagen, DK) allowed solids up to 550 micrometer into the column without clogging for all flow rates evaluated. A mesh at the inlet with size restriction of either 50 micrometer or 80 micrometer became clogged with very small corn particles (< 44 micrometer). When glass ballotini was used, large particles (550 micrometer) passed through for high flow rates (570 cm/h), but even small (< 44 micrometer) particles became trapped at a lower flow rate (180 cm/h). The physical and chemical properties of the resin determined whether solids could be eluted. The denser UpFront adsorbents allowed for complete elution of larger and more concentrated corn solids than the currently available Amersham Streamline adsorbents (Amersham Biosciences, Piscataway, NJ) as a result of the former's higher flow rate for the desired 2x expansion (570 cm/h for UpFront vs. 180 cm/h for Streamline). All corn solids < 162 micrometer eluted through nonderivatized UpFront resin. Larger corn solids began to accumulate due to their elevated sedimentation velocities. Feeds of < 44 micrometer solids at 0.45% and 2.0% dry weight successfully eluted through ion exchange adsorbents (DEAE and SP) from UpFront. However, significant accumulation occurred when the solids size increased to a feed of < 96 micrometer solids, thus indicating a weak interaction between corn solids and both forms of ion exchange ligands. Expanded beds operated with Streamline ion exchange adsorbents (DEAE and SP) did not allow full elution of corn solids of < 44 micrometer. A hyperdiffuse style EBA resin prod