Purification of recombinant and human apolipoprotein A-1 using surfactant micelles in aqueous two-phase systems: Recycling of thermoseparating polymer and surfactant with temperature-induced phase separation
(1999) In Biotechnology and Bioengineering 65(4). p.371-381- Abstract
- An effective system has been developed for purification of apolipoprotein A-1 from Escherichia coli fermentation solution and human plasma using aqueous two-phase extraction and thermal-phase separation. The system included non-ionic surfactants (Triton or Tween) and as top phase-forming polymer a random copolymer of ethylene oxide (50%) and propylene oxide (50%), Breox PAG 50A 1000, was used. The bottom phase-forming polymer was either hydroxypropyl starch, Reppal PES 100 and PES 200, or hydroxyethyl starch, Solfarex A 85. The top-phase-forming polymer and the surfactants are thermoseparating in water solution, i.e., when heated a water phase and a polymer/surfactant phase are formed. Recombinant apolipoprotein A-1, the Milano variant,... (More)
- An effective system has been developed for purification of apolipoprotein A-1 from Escherichia coli fermentation solution and human plasma using aqueous two-phase extraction and thermal-phase separation. The system included non-ionic surfactants (Triton or Tween) and as top phase-forming polymer a random copolymer of ethylene oxide (50%) and propylene oxide (50%), Breox PAG 50A 1000, was used. The bottom phase-forming polymer was either hydroxypropyl starch, Reppal PES 100 and PES 200, or hydroxyethyl starch, Solfarex A 85. The top-phase-forming polymer and the surfactants are thermoseparating in water solution, i.e., when heated a water phase and a polymer/surfactant phase are formed. Recombinant apolipoprotein A-1, the Milano variant, was extracted from E. coli fermentation solution in a primary Breox-starch phase system followed by thermal separation of the Breox phase where the target protein was recovered in the water phase. Both in the Breox-starch system and in the water-Breox system Triton X-100 was partitioned to the Breox phase. The addition of non-ionic surfactants to the Breox-starch system had strong effect on the purification and yield of the amphiphilic apolipoprotein A-1. In a system containing 17% Breox PAG 50A 1000, 12% Reppal PES 100 and addition of 1% Triton X-100 the purification factor was 7.2, and the yield 85% after thermal separation of the Breox phase. Recycling of copolymer and surfactant was possible after thermal separation of copolymer phase. Approximately 85% of the copolymer and surfactant could be recycled in each extraction cycle. DNA could be strongly partitioned to the starch phase in the primary-phase system. This resulted in a 1000-fold reduction of E. coli DNA in the apolipoprotein A-1 solution obtained after thermoseparation. In extraction from human plasma containing low concentrations of apolipoprotein A-1, it was possible to reach a purification factor of 420 with 98% yield. By reducing the volume ratio to 0.1 Apo A-1 could be concentrated in a small volume of top phase (concentration factor 10) with a yield of 85% and a purification factor of 110. © 1999 John Wiley & Sons, Inc. Biotechnol Bioeng 65: 371-381, 1999. (Less)
Please use this url to cite or link to this publication:
https://lup.lub.lu.se/record/125377
- author
- Persson, Josefine ; Nyström, Lena ; Ageland, Hans and Tjerneld, Folke LU
- organization
- publishing date
- 1999
- type
- Contribution to journal
- publication status
- published
- subject
- keywords
- downstream processing, apolipoprotein A-1, purification, thermoseparating polymer, surfactant, recycling, aqueous two-phase system
- in
- Biotechnology and Bioengineering
- volume
- 65
- issue
- 4
- pages
- 371 - 381
- publisher
- John Wiley & Sons Inc.
- external identifiers
-
- scopus:0033589830
- ISSN
- 1097-0290
- language
- English
- LU publication?
- yes
- id
- c4ad1347-b48e-42f9-b779-d3e8e23d85fc (old id 125377)
- alternative location
- http://www3.interscience.wiley.com/cgi-bin/abstract/71003550/ABSTRACT
- date added to LUP
- 2016-04-01 12:35:36
- date last changed
- 2022-01-27 07:11:51
@article{c4ad1347-b48e-42f9-b779-d3e8e23d85fc,
abstract = {{An effective system has been developed for purification of apolipoprotein A-1 from Escherichia coli fermentation solution and human plasma using aqueous two-phase extraction and thermal-phase separation. The system included non-ionic surfactants (Triton or Tween) and as top phase-forming polymer a random copolymer of ethylene oxide (50%) and propylene oxide (50%), Breox PAG 50A 1000, was used. The bottom phase-forming polymer was either hydroxypropyl starch, Reppal PES 100 and PES 200, or hydroxyethyl starch, Solfarex A 85. The top-phase-forming polymer and the surfactants are thermoseparating in water solution, i.e., when heated a water phase and a polymer/surfactant phase are formed. Recombinant apolipoprotein A-1, the Milano variant, was extracted from E. coli fermentation solution in a primary Breox-starch phase system followed by thermal separation of the Breox phase where the target protein was recovered in the water phase. Both in the Breox-starch system and in the water-Breox system Triton X-100 was partitioned to the Breox phase. The addition of non-ionic surfactants to the Breox-starch system had strong effect on the purification and yield of the amphiphilic apolipoprotein A-1. In a system containing 17% Breox PAG 50A 1000, 12% Reppal PES 100 and addition of 1% Triton X-100 the purification factor was 7.2, and the yield 85% after thermal separation of the Breox phase. Recycling of copolymer and surfactant was possible after thermal separation of copolymer phase. Approximately 85% of the copolymer and surfactant could be recycled in each extraction cycle. DNA could be strongly partitioned to the starch phase in the primary-phase system. This resulted in a 1000-fold reduction of E. coli DNA in the apolipoprotein A-1 solution obtained after thermoseparation. In extraction from human plasma containing low concentrations of apolipoprotein A-1, it was possible to reach a purification factor of 420 with 98% yield. By reducing the volume ratio to 0.1 Apo A-1 could be concentrated in a small volume of top phase (concentration factor 10) with a yield of 85% and a purification factor of 110. © 1999 John Wiley & Sons, Inc. Biotechnol Bioeng 65: 371-381, 1999.}},
author = {{Persson, Josefine and Nyström, Lena and Ageland, Hans and Tjerneld, Folke}},
issn = {{1097-0290}},
keywords = {{downstream processing; apolipoprotein A-1; purification; thermoseparating polymer; surfactant; recycling; aqueous two-phase system}},
language = {{eng}},
number = {{4}},
pages = {{371--381}},
publisher = {{John Wiley & Sons Inc.}},
series = {{Biotechnology and Bioengineering}},
title = {{Purification of recombinant and human apolipoprotein A-1 using surfactant micelles in aqueous two-phase systems: Recycling of thermoseparating polymer and surfactant with temperature-induced phase separation}},
url = {{http://www3.interscience.wiley.com/cgi-bin/abstract/71003550/ABSTRACT}},
volume = {{65}},
year = {{1999}},
}