Purification of recombinant human basic fibroblast growth factor

stability of selective sorbents under cleaning in place conditions

verfasst von: F. Birger Anspach, Henning Spille, Ursula Rinas
Abstract: Human basic fibroblast growth factor (bFGF) was produced from recombinant Escherichia coli by high-cell-density cultivation. In order to develop a purification strategy for large-scale purification, chromatographic sorbents with different anionic functional groups were compared in terms of selectivity for bFGF and stability under cleaning in place (CIP) conditions. Heparin-Sepharose CL-6B, Fractogel EMD-SO₃^- 650 (S) and SP-Sepharose (high performance) were found suitable for this purpose with decreasing selectivity in that order. Each sorbent was treated eight times under CIP conditions employing both 0.2 and 1.0 M NaOH, in order to study modifications of these sorbents. Heparin-Sepharose displayed more than 50% loss of capacity after the first CIP treatment and decreasing selectivity with each cycle. Both cation exchangers displayed almost constant results regarding selectivity and capacity. The Fractogel EMD-SO₃^- exhibited only slightly lower selectivity for bFGF than Heparin-Sepharose and the highest capacity of all sorbents tested. Agglomeration of bFGF at low salt concentrations was a serious problem. By direct application of pooled fractions from Fractogel EMD-SO₃^- onto Heparin-Sepharose a highly pure product was obtained; however, the recovery after Heparin-Sepharose was only 30%.
Externe Organisation(en): Helmholtz-Zentrum für Infektionsforschung GmbH (HZI)
Typ: Artikel
Journal: Journal of Chromatography A
Band: 711
Seiten: 129-139
Anzahl der Seiten: 11
ISSN: 0021-9673
Publikationsdatum: 08.09.1995
Publikationsstatus: Veröffentlicht
Peer-reviewed: Ja
ASJC Scopus Sachgebiete: Analytische Chemie, Biochemie, Organische Chemie
Elektronische Version(en): https://doi.org/10.1016/0021-9673(95)00102-S (Zugang: Geschlossen)

BibTeX

@article{106eb891086d4e86a655b469e50147a5,
title = "Purification of recombinant human basic fibroblast growth factor: stability of selective sorbents under cleaning in place conditions",
abstract = "Human basic fibroblast growth factor (bFGF) was produced from recombinant Escherichia coli by high-cell-density cultivation. In order to develop a purification strategy for large-scale purification, chromatographic sorbents with different anionic functional groups were compared in terms of selectivity for bFGF and stability under cleaning in place (CIP) conditions. Heparin-Sepharose CL-6B, Fractogel EMD-SO3- 650 (S) and SP-Sepharose (high performance) were found suitable for this purpose with decreasing selectivity in that order. Each sorbent was treated eight times under CIP conditions employing both 0.2 and 1.0 M NaOH, in order to study modifications of these sorbents. Heparin-Sepharose displayed more than 50% loss of capacity after the first CIP treatment and decreasing selectivity with each cycle. Both cation exchangers displayed almost constant results regarding selectivity and capacity. The Fractogel EMD-SO3- exhibited only slightly lower selectivity for bFGF than Heparin-Sepharose and the highest capacity of all sorbents tested. Agglomeration of bFGF at low salt concentrations was a serious problem. By direct application of pooled fractions from Fractogel EMD-SO3- onto Heparin-Sepharose a highly pure product was obtained; however, the recovery after Heparin-Sepharose was only 30%.",
author = "{Birger Anspach}, F. and Henning Spille and Ursula Rinas",
year = "1995",
month = sep,
day = "8",
doi = "10.1016/0021-9673(95)00102-S",
language = "English",
volume = "711",
pages = "129--139",
journal = "Journal of Chromatography A",
issn = "0021-9673",
publisher = "Elsevier",
number = "1",
}