title: Allantoin as a solid phase adsorbent for removing endotoxins.
abstract: In this study we present a simple and robust method for removing endotoxins from protein solutions by using crystals of the small-molecule compound 2,5-dioxo-4-imidazolidinyl urea (allantoin) as a solid phase adsorbent. Allantoin crystalline powder is added to a protein solution at supersaturated concentrations, endotoxins bind and undissolved allantoin crystals with bound endotoxins are removed by filtration or centrifugation. This method removes an average of 99.98% endotoxin for 20 test proteins. The average protein recovery is ∼80%. Endotoxin binding is largely independent of pH, conductivity, reducing agent and various organic solvents. This is consistent with a hydrogen-bond based binding mechanism. Allantoin does not affect protein activity and stability, and the use of allantoin as a solid phase adsorbent provides better endotoxin removal than anion exchange, polymixin affinity and biological affinity methods for endotoxin clearance.
authors.affiliation: Bioprocessing Technology Institute, A*STAR (Agency for Science, Technology and Research), 20 Biopolis Way #06-01 Centros, Singapore 138668, Singapore. Electronic address: Vincent_Vagenende@bti.a-star.edu.sg.
authors.forename: Vincent
authors.initials: V
authors.lastname: Vagenende
authors.forename: Tim-Jang
authors.initials: TJ
authors.lastname: Ching
authors.forename: Rui-Jing
authors.initials: RJ
authors.lastname: Chua
authors.forename: Pete
authors.initials: P
authors.lastname: Gagnon
mesh_terms: D000327:Adsorption; D000481:Allantoin / Q000737:chemistry* / Q000378:metabolism; D002850:Chromatography, Gel; D004731:Endotoxins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D006860:Hydrogen Bonding; D006863:Hydrogen-Ion Concentration; D055550:Protein Stability; D011506:Proteins / Q000737:chemistry / Q000302:isolation & purification; D012965:Sodium Chloride; D052616:Solid Phase Extraction / Q000379:methods*
chemical_list: D004731:Endotoxins; D011506:Proteins; D000481:Allantoin; D012965:Sodium Chloride
keywords: Endotoxins; Hydrogen-bond based adsorption; Lipopolysaccharides; Protein purification; Supersaturated allantoin