Phenotyping the quality of complex medium components by simple online-monitored shake flask experiments

Microbial Cell Factories

Research

Antje Staaden¹  Sylvia Diederichs¹  Anna Korona¹  Jochen Büchs¹  Kohsuke Honda²  Hisao Ohtake²  Wolfgang Kroutil³ 

[1] AVT – Biochemical Engineering, RWTH Aachen University, Worringerweg 1, D-52074, Aachen, Germany;Department of Biotechnology, Graduate School of Engineering, Osaka University, 2-1 Yamadaoka, 565-0871, Suita, Osaka, Japan;Department of Chemistry, Organic and Bioorganic Chemistry, NAWI Graz, University of Graz, Heinrichstrasse 28, A-8010, Graz, Austria;

关键词: RAMOS;    OTR;    Yeast extract;    Complex medium;    E. coli;    Auto-induction;    Screening;    Lactose;    PAT;   

DOI  :  10.1186/s12934-014-0149-5

 received in 2014-06-26, accepted in 2014-10-15,  发布年份 2014

来源: Springer

【 摘 要 】

BackgroundMedia containing yeast extracts and other complex raw materials are widely used for the cultivation of microorganisms. However, variations in the specific nutrient composition can occur, due to differences in the complex raw material ingredients and in the production of these components. These lot-to-lot variations can affect growth rate, product yield and product quality in laboratory investigations and biopharmaceutical production processes. In the FDA's Process Analytical Technology (PAT) initiative, the control and assessment of the quality of critical raw materials is one key aspect to maintain product quality and consistency. In this study, the Respiration Activity Monitoring System (RAMOS) was used to evaluate the impact of different yeast extracts and commercial complex auto-induction medium lots on metabolic activity and product yield of four recombinant Escherichia coli variants encoding different enzymes.ResultsUnder non-induced conditions, the oxygen transfer rate (OTR) of E. coli was not affected by a variation of the supplemented yeast extract lot. The comparison of E. coli cultivations under induced conditions exhibited tremendous differences in OTR profiles and volumetric activity for all investigated yeast extract lots of different suppliers as well as lots of the same supplier independent of the E. coli variant. Cultivation in the commercial auto-induction medium lots revealed the same reproducible variations. In cultivations with parallel offline analysis, the highest volumetric activity was found at different cultivation times. Only by online monitoring of the cultures, a distinct cultivation phase (e.g. glycerol depletion) could be detected and chosen for comparable and reproducible offline analysis of the yield of functional product.ConclusionsThis work proves that cultivations conducted in complex media may be prone to significant variation in final product quality and quantity if the quality of the raw material for medium preparation is not thoroughly checked. In this study, the RAMOS technique enabled a reliable and reproducible screening and phenotyping of complex raw material lots by online measurement of the respiration activity. Consequently, complex raw material lots can efficiently be assessed if the distinct effects on culture behavior and final product quality and quantity are visualized.

【 授权许可】

Unknown   
© Diederichs et al.; licensee BioMed Central Ltd. 2014. This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.