title: Impact of extraction and elution media on non-size effects in size exclusion chromatography of proteins.
abstract: Size exclusion chromatography is extensively used to separate proteins and to determine their apparent molecular weights. It separates proteins based on hydrodynamic volume, but interactions between the chromatography resin and proteins lead to non-size effects. This report discusses the impact of co-solvents [salt, urea, sodium dodecyl sulfate (SDS), dithiothreitol] in extraction media when separating wheat gluten proteins, soy glycinin, bovine serum albumin and ovalbumin on a Biosep-SEC-S4000 column. With acetonitrile/water (1:1, v/v) containing 0.05% (v/v) trifluoroacetic acid as eluent, salts and SDS in the extraction media increase while urea decreases non-size effects. Most gluten and globular proteins are extractable in sodium phosphate buffer (0.050M; pH 6.8) containing 2.0% (w/v) SDS. This chromatographic medium allows analyzing mixtures of various proteins without any non-size effects.
authors.affiliation: Laboratory of Food Chemistry and Biochemistry and Leuven Food Science and Nutrition Research Centre (LFoRCe), KU Leuven, Kasteelpark Arenberg 20, B-3001 Leuven, Belgium. Electronic address: marlies.lambrecht@biw.kuleuven.be.
authors.forename: Marlies A
authors.initials: MA
authors.lastname: Lambrecht
authors.affiliation: Laboratory of Food Chemistry and Biochemistry and Leuven Food Science and Nutrition Research Centre (LFoRCe), KU Leuven, Kasteelpark Arenberg 20, B-3001 Leuven, Belgium.
authors.forename: Ine
authors.initials: I
authors.lastname: Rombouts
authors.affiliation: Laboratory of Food Chemistry and Biochemistry and Leuven Food Science and Nutrition Research Centre (LFoRCe), KU Leuven, Kasteelpark Arenberg 20, B-3001 Leuven, Belgium.
authors.forename: Lotte
authors.initials: L
authors.lastname: Van Kelst
authors.affiliation: Laboratory of Food Chemistry and Biochemistry and Leuven Food Science and Nutrition Research Centre (LFoRCe), KU Leuven, Kasteelpark Arenberg 20, B-3001 Leuven, Belgium.
authors.forename: Jan A
authors.initials: JA
authors.lastname: Delcour
mesh_terms: D000097:Acetonitriles / Q000737:chemistry; D000818:Animals; D002417:Cattle; D002850:Chromatography, Gel; D004229:Dithiothreitol / Q000737:chemistry; D005916:Globulins / Q000302:isolation & purification*; D005983:Glutens / Q000302:isolation & purification; D008970:Molecular Weight; D010047:Ovalbumin / Q000302:isolation & purification*; D010940:Plant Proteins / Q000302:isolation & purification*; D012710:Serum Albumin, Bovine / Q000302:isolation & purification*; D012965:Sodium Chloride / Q000737:chemistry; D012967:Sodium Dodecyl Sulfate / Q000737:chemistry; D012997:Solvents; D030262:Soybean Proteins / Q000302:isolation & purification*; D014269:Trifluoroacetic Acid / Q000737:chemistry; D014908:Triticum; D014508:Urea / Q000737:chemistry
chemical_list: D000097:Acetonitriles; D005916:Globulins; D010940:Plant Proteins; D012997:Solvents; D030262:Soybean Proteins; D012710:Serum Albumin, Bovine; D012967:Sodium Dodecyl Sulfate; D012965:Sodium Chloride; D005983:Glutens; D014508:Urea; D010047:Ovalbumin; C011474:glycinin; D014269:Trifluoroacetic Acid; D004229:Dithiothreitol; C032159:acetonitrile
keywords: Co-solvents; Extraction; Non-size effects; Size exclusion