One-step method for isolation and purification of native ß-lactoglobulin from bovine whey

M. Stojadinovic, L. Burazer, Dilek Ercili-Cura, A. Sancho, Johanna Buchert, T. Cirkovic Velickovic, D. Stanic-Vucinic (Corresponding Author)

Research output: Contribution to journalArticleScientificpeer-review

15 Citations (Scopus)

Abstract

BACKGROUND: The major whey protein β‐lactoglobulin (BLG) has been widely studied for its functional properties. The aim of this study was to develop an efficient, inexpensive and rapid one‐step method for the isolation and purification of BLG while preserving its native structure.

RESULTS: BLG was purified from defatted whey obtained from raw cow's milk by anion exchange chromatography. Protein purity and identity were determined using reverse phase high‐performance liquid chromatography and mass spectrometry. Total BLG yield was 80% with protein purity from 97 to 99%. BLG isoforms A and B were separated into fractions of 91 and 99% purity respectively. The structure and native conformation of the isolated BLG were compared with those of standard commercial BLG by circular dichroism spectrometry, susceptibility to various crosslinking enzymes and enzyme‐linked immunosorbent assay inhibition.

CONCLUSION: The proposed method is very useful for the rapid preparation of BLG suitable for studying antigenic and molecular characteristics of this protein, as well as the effect of food processing on these properties. The procedure requires only 1 day for the purification of about 300 mg of BLG from a single run using a small column (2.5 cm × 20 cm) of diethylaminoethyl Sephadex and has potential for scaling up.
Original languageEnglish
Pages (from-to)1432-1440
Number of pages9
JournalJournal of the Science of Food and Agriculture
Volume92
Issue number7
DOIs
Publication statusPublished - 2012
MoE publication typeA1 Journal article-refereed

Fingerprint

lactoglobulins
Lactoglobulins
purification methods
isolation techniques
whey
purity
cattle
circular dichroism spectroscopy
Immunosorbents
Proteins
Food Handling
proteins
reversed-phase liquid chromatography
Reverse-Phase Chromatography
Circular Dichroism
raw milk
whey protein
crosslinking
food processing
rapid methods

Keywords

  • native ß-lactoglobulin
  • isolation
  • anion exchange chromatography
  • purification

Cite this

Stojadinovic, M., Burazer, L., Ercili-Cura, D., Sancho, A., Buchert, J., Cirkovic Velickovic, T., & Stanic-Vucinic, D. (2012). One-step method for isolation and purification of native ß-lactoglobulin from bovine whey. Journal of the Science of Food and Agriculture, 92(7), 1432-1440. https://doi.org/10.1002/jsfa.4722
Stojadinovic, M. ; Burazer, L. ; Ercili-Cura, Dilek ; Sancho, A. ; Buchert, Johanna ; Cirkovic Velickovic, T. ; Stanic-Vucinic, D. / One-step method for isolation and purification of native ß-lactoglobulin from bovine whey. In: Journal of the Science of Food and Agriculture. 2012 ; Vol. 92, No. 7. pp. 1432-1440.
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title = "One-step method for isolation and purification of native {\ss}-lactoglobulin from bovine whey",
abstract = "BACKGROUND: The major whey protein β‐lactoglobulin (BLG) has been widely studied for its functional properties. The aim of this study was to develop an efficient, inexpensive and rapid one‐step method for the isolation and purification of BLG while preserving its native structure. RESULTS: BLG was purified from defatted whey obtained from raw cow's milk by anion exchange chromatography. Protein purity and identity were determined using reverse phase high‐performance liquid chromatography and mass spectrometry. Total BLG yield was 80{\%} with protein purity from 97 to 99{\%}. BLG isoforms A and B were separated into fractions of 91 and 99{\%} purity respectively. The structure and native conformation of the isolated BLG were compared with those of standard commercial BLG by circular dichroism spectrometry, susceptibility to various crosslinking enzymes and enzyme‐linked immunosorbent assay inhibition. CONCLUSION: The proposed method is very useful for the rapid preparation of BLG suitable for studying antigenic and molecular characteristics of this protein, as well as the effect of food processing on these properties. The procedure requires only 1 day for the purification of about 300 mg of BLG from a single run using a small column (2.5 cm × 20 cm) of diethylaminoethyl Sephadex and has potential for scaling up.",
keywords = "native {\ss}-lactoglobulin, isolation, anion exchange chromatography, purification",
author = "M. Stojadinovic and L. Burazer and Dilek Ercili-Cura and A. Sancho and Johanna Buchert and {Cirkovic Velickovic}, T. and D. Stanic-Vucinic",
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Stojadinovic, M, Burazer, L, Ercili-Cura, D, Sancho, A, Buchert, J, Cirkovic Velickovic, T & Stanic-Vucinic, D 2012, 'One-step method for isolation and purification of native ß-lactoglobulin from bovine whey', Journal of the Science of Food and Agriculture, vol. 92, no. 7, pp. 1432-1440. https://doi.org/10.1002/jsfa.4722

One-step method for isolation and purification of native ß-lactoglobulin from bovine whey. / Stojadinovic, M.; Burazer, L.; Ercili-Cura, Dilek; Sancho, A.; Buchert, Johanna; Cirkovic Velickovic, T.; Stanic-Vucinic, D. (Corresponding Author).

In: Journal of the Science of Food and Agriculture, Vol. 92, No. 7, 2012, p. 1432-1440.

Research output: Contribution to journalArticleScientificpeer-review

TY - JOUR

T1 - One-step method for isolation and purification of native ß-lactoglobulin from bovine whey

AU - Stojadinovic, M.

AU - Burazer, L.

AU - Ercili-Cura, Dilek

AU - Sancho, A.

AU - Buchert, Johanna

AU - Cirkovic Velickovic, T.

AU - Stanic-Vucinic, D.

PY - 2012

Y1 - 2012

N2 - BACKGROUND: The major whey protein β‐lactoglobulin (BLG) has been widely studied for its functional properties. The aim of this study was to develop an efficient, inexpensive and rapid one‐step method for the isolation and purification of BLG while preserving its native structure. RESULTS: BLG was purified from defatted whey obtained from raw cow's milk by anion exchange chromatography. Protein purity and identity were determined using reverse phase high‐performance liquid chromatography and mass spectrometry. Total BLG yield was 80% with protein purity from 97 to 99%. BLG isoforms A and B were separated into fractions of 91 and 99% purity respectively. The structure and native conformation of the isolated BLG were compared with those of standard commercial BLG by circular dichroism spectrometry, susceptibility to various crosslinking enzymes and enzyme‐linked immunosorbent assay inhibition. CONCLUSION: The proposed method is very useful for the rapid preparation of BLG suitable for studying antigenic and molecular characteristics of this protein, as well as the effect of food processing on these properties. The procedure requires only 1 day for the purification of about 300 mg of BLG from a single run using a small column (2.5 cm × 20 cm) of diethylaminoethyl Sephadex and has potential for scaling up.

AB - BACKGROUND: The major whey protein β‐lactoglobulin (BLG) has been widely studied for its functional properties. The aim of this study was to develop an efficient, inexpensive and rapid one‐step method for the isolation and purification of BLG while preserving its native structure. RESULTS: BLG was purified from defatted whey obtained from raw cow's milk by anion exchange chromatography. Protein purity and identity were determined using reverse phase high‐performance liquid chromatography and mass spectrometry. Total BLG yield was 80% with protein purity from 97 to 99%. BLG isoforms A and B were separated into fractions of 91 and 99% purity respectively. The structure and native conformation of the isolated BLG were compared with those of standard commercial BLG by circular dichroism spectrometry, susceptibility to various crosslinking enzymes and enzyme‐linked immunosorbent assay inhibition. CONCLUSION: The proposed method is very useful for the rapid preparation of BLG suitable for studying antigenic and molecular characteristics of this protein, as well as the effect of food processing on these properties. The procedure requires only 1 day for the purification of about 300 mg of BLG from a single run using a small column (2.5 cm × 20 cm) of diethylaminoethyl Sephadex and has potential for scaling up.

KW - native ß-lactoglobulin

KW - isolation

KW - anion exchange chromatography

KW - purification

U2 - 10.1002/jsfa.4722

DO - 10.1002/jsfa.4722

M3 - Article

VL - 92

SP - 1432

EP - 1440

JO - Journal of the Science of Food and Agriculture

JF - Journal of the Science of Food and Agriculture

SN - 0022-5142

IS - 7

ER -