Purification of recombinant human basic fibroblast growth factor

stability of selective sorbents under cleaning in place conditions

authored by

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%.

External Organisation(s)

Helmholtz Centre for Infection Research (HZI)

Type

Article

Journal

Journal of Chromatography A

Volume

711

Pages

129-139

No. of pages

11

ISSN

0021-9673

Publication date

08.09.1995

Publication status

Published

Peer reviewed

Yes

ASJC Scopus subject areas

Analytical Chemistry, Biochemistry, Organic Chemistry

Electronic version(s)

https://doi.org/10.1016/0021-9673(95)00102-S (Access: Closed)