title: Sequential separation of immunoglobulin Y and phosvitin from chicken egg yolk without using organic solvents.
abstract: A study was conducted to develop a simple sequential separation protocol to separate phosvitin and IgY from egg yolk without using organic solvents. Egg yolk was diluted with 2 volumes of distilled water (DW), homogenized, and centrifuged. The precipitant was collected and homogenized with 4 volumes of 10% NaCl (wt/vol) in 0.05 N NaOH solution to extract phosvitin. The pH of the homogenate was adjusted to 4.0 and the precipitate was removed by centrifugation. The supernatant was collected and then heat-treated at 70°C for 30 min and centrifuged to remove impurities. The supernatant containing phosvitin was collected, had salts removed, and was concentrated and then freeze-dried. The supernatant from the centrifugation of diluted egg yolk was diluted again with 3 volumes of DW, and the precipitate was removed by centrifugation. The resulting supernatant was concentrated using ultrafiltration and then IgY was precipitated using 20% saturated (NH₄)₂SO₄+ 15% NaCl (wt/vol). The precipitant was collected after centrifugation at 3,400 × g for 30 min at 4°C and dissolved with DW, had salts removed, and then was freeze-dried. The purity of separated phosvitin and IgY was checked using SDS-PAGE and the proteins were verified using Western blotting. The purity of phosvitin and IgY was 97.2 and 98.7%, and the yield was 98.7 and 80.9%, respectively. The ELISA results indicated that the activities of separated IgY and phosvitin were 96.3 and 98.3%, respectively. This study proved that both phosvitin and IgY can be separated in sequence from egg yolk without using an organic solvent. Also, the method is very simple and has a high potential for scale-up processing.
authors.affiliation: WCU Biomodulation, Department of Agricultural Biotechnology, Seoul National University, Seoul 151-921, South Korea Center for Nutraceutical and Pharmaceutical Materials, Myongji University, Cheoin-gu, Yongin, Gyeonggi-Do 449-728, Korea.
authors.forename: Hyun Yong
authors.initials: HY
authors.lastname: Lee
authors.affiliation: WCU Biomodulation, Department of Agricultural Biotechnology, Seoul National University, Seoul 151-921, South Korea Department of Animal Science, Uva Wellassa University, Badulla, Sri Lanka 90000.
authors.forename: E D N S
authors.initials: ED
authors.lastname: Abeyrathne
authors.affiliation: Functional Materials Research Group, Food Research Institute, Sungnam, Gyeonggi-Do, 463-746, Korea.
authors.forename: Inwook
authors.initials: I
authors.lastname: Choi
authors.affiliation: Center for Nutraceutical and Pharmaceutical Materials, Myongji University, Cheoin-gu, Yongin, Gyeonggi-Do 449-728, Korea duahn@iastate.edu jwsuh@mju.ac.kr.
authors.forename: Joo Won
authors.initials: JW
authors.lastname: Suh
authors.affiliation: WCU Biomodulation, Department of Agricultural Biotechnology, Seoul National University, Seoul 151-921, South Korea Department of Animal Science, Iowa State University, Ames 50011 Department of Animal Science and Technology, Sunchon National University, Sunchon 540-742, South Korea duahn@iastate.edu jwsuh@mju.ac.kr.
authors.forename: Dong Uk
authors.initials: DU
authors.lastname: Ahn
mesh_terms: D000818:Animals; D015153:Blotting, Western; D011232:Chemical Precipitation; D002645:Chickens; D004527:Egg Proteins / Q000737:chemistry / Q000302:isolation & purification*; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D005511:Food Handling / Q000379:methods*; D007136:Immunoglobulins / Q000737:chemistry / Q000302:isolation & purification*; D010774:Phosvitin / Q000737:chemistry / Q000302:isolation & purification*; D012997:Solvents / Q000737:chemistry*
chemical_list: D004527:Egg Proteins; C016512:IgY; D007136:Immunoglobulins; D012997:Solvents; D010774:Phosvitin
keywords: egg yolk; immunoglobulin Y; phosvitin; sequential separation