Get 20M+ Full-Text Papers For Less Than $1.50/day. Start a 14-Day Trial for You or Your Team.

Learn More →

Design of experiments reveals critical parameters for pilot‐scale freeze‐and‐thaw processing of L‐lactic dehydrogenase

Design of experiments reveals critical parameters for pilot‐scale freeze‐and‐thaw processing of... Freezing constitutes an important unit operation of biotechnological protein production. Effects of freeze‐and‐thaw (F/T) process parameters on stability and other quality attributes of the protein product are usually not well understood. Here a design of experiments (DoE) approach was used to characterize the F/T behavior of L‐lactic dehydrogenase (LDH) in a 700‐mL pilot‐scale freeze container equipped with internal temperature and pH probes. In 24‐hour experiments, target temperature between –10 and –38°C most strongly affected LDH stability whereby enzyme activity was retained best at the highest temperature of –10°C. Cooling profile and liquid fill volume also had significant effects on LDH stability and affected the protein aggregation significantly. Parameters of the thawing phase had a comparably small effect on LDH stability. Experiments in which the standard sodium phosphate buffer was exchanged by Tris‐HCl and the non‐ionic surfactant Tween 80 was added to the protein solution showed that pH shift during freezing and protein surface exposure were the main factors responsible for LDH instability at the lower freeze temperatures. Collectively, evidence is presented that supports the use of DoE‐based systematic analysis at pilot scale in the identification of F/T process parameters critical for protein stability and in the development of suitable process control strategies. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Biotechnology Journal Wiley

Design of experiments reveals critical parameters for pilot‐scale freeze‐and‐thaw processing of L‐lactic dehydrogenase

Download PDF
12 pages

Loading next page...
 
/lp/wiley/design-of-experiments-reveals-critical-parameters-for-pilot-scale-7P5k06ERo0

References (38)

Publisher
Wiley
Copyright
Copyright © 2015 Wiley Subscription Services, Inc., A Wiley Company
ISSN
1860-6768
eISSN
1860-7314
DOI
10.1002/biot.201400766
pmid
25820483
Publisher site
See Article on Publisher Site

Abstract

Freezing constitutes an important unit operation of biotechnological protein production. Effects of freeze‐and‐thaw (F/T) process parameters on stability and other quality attributes of the protein product are usually not well understood. Here a design of experiments (DoE) approach was used to characterize the F/T behavior of L‐lactic dehydrogenase (LDH) in a 700‐mL pilot‐scale freeze container equipped with internal temperature and pH probes. In 24‐hour experiments, target temperature between –10 and –38°C most strongly affected LDH stability whereby enzyme activity was retained best at the highest temperature of –10°C. Cooling profile and liquid fill volume also had significant effects on LDH stability and affected the protein aggregation significantly. Parameters of the thawing phase had a comparably small effect on LDH stability. Experiments in which the standard sodium phosphate buffer was exchanged by Tris‐HCl and the non‐ionic surfactant Tween 80 was added to the protein solution showed that pH shift during freezing and protein surface exposure were the main factors responsible for LDH instability at the lower freeze temperatures. Collectively, evidence is presented that supports the use of DoE‐based systematic analysis at pilot scale in the identification of F/T process parameters critical for protein stability and in the development of suitable process control strategies.

Journal

Biotechnology Journal – Wiley

Published: Sep 1, 2015

Keywords: ; ; ; ;

There are no references for this article.