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FTIR spectroscopy as a unified method for simultaneous analysis of intra- and extracellular metabolites in high-throughput screening of microbial bioprocesses
BACKGROUND: Analyses of substrate and metabolites are often bottleneck activities in high-throughput screening of microbial bioprocesses. We have assessed Fourier transform infrared spectroscopy (FTIR), in combination with high throughput micro-bioreactors and multivariate statistical analyses, for...
Autores principales: | , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
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BioMed Central
2017
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5683213/ https://www.ncbi.nlm.nih.gov/pubmed/29132358 http://dx.doi.org/10.1186/s12934-017-0817-3 |
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author | Kosa, Gergely Shapaval, Volha Kohler, Achim Zimmermann, Boris |
author_facet | Kosa, Gergely Shapaval, Volha Kohler, Achim Zimmermann, Boris |
author_sort | Kosa, Gergely |
collection | PubMed |
description | BACKGROUND: Analyses of substrate and metabolites are often bottleneck activities in high-throughput screening of microbial bioprocesses. We have assessed Fourier transform infrared spectroscopy (FTIR), in combination with high throughput micro-bioreactors and multivariate statistical analyses, for analysis of metabolites in high-throughput screening of microbial bioprocesses. In our previous study, we have demonstrated that high-throughput (HTS) FTIR can be used for estimating content and composition of intracellular metabolites, namely triglyceride accumulation in oleaginous filamentous fungi. As a continuation of that research, in the present study HTS FTIR was evaluated as a unified method for simultaneous quantification of intra- and extracellular metabolites and substrate consumption. As a proof of concept, a high-throughput microcultivation of oleaginous filamentous fungi was conducted in order to monitor production of citric acid (extracellular metabolite) and triglyceride lipids (intracellular metabolites), as well as consumption of glucose in the cultivation medium. RESULTS: HTS FTIR analyses of supernatant samples was compared with an attenuated total reflection (ATR) FTIR, which is an established method for bioprocess monitoring. Glucose and citric acid content of growth media was quantified by high performance liquid chromatography (HPLC). Partial least square regression (PLSR) between HPLC glucose and citric acid data and the corresponding FTIR spectral data was used to set up calibration models. PLSR results for HTS measurements were very similar to the results obtained with ATR methodology, with high coefficients of determination (0.91–0.98) and low error values (4.9–8.6%) for both glucose and citric acid estimates. CONCLUSIONS: The study has demonstrated that intra- and extracellular metabolites, as well as nutrients in the cultivation medium, can be monitored by a unified approach by HTS FTIR. The proof-of-concept study has validated that HTS FTIR, in combination with Duetz microtiter plate system and chemometrics, can be used for high throughput screening of microbial bioprocesses. It can be anticipated that the approach, demonstrated here on single-cell oil production by filamentous fungi, can find general application in screening studies of microbial bioprocesses, such as production of single-cell proteins, biopolymers, polysaccharides, carboxylic acids, and other type of metabolites. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1186/s12934-017-0817-3) contains supplementary material, which is available to authorized users. |
format | Online Article Text |
id | pubmed-5683213 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2017 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-56832132017-11-20 FTIR spectroscopy as a unified method for simultaneous analysis of intra- and extracellular metabolites in high-throughput screening of microbial bioprocesses Kosa, Gergely Shapaval, Volha Kohler, Achim Zimmermann, Boris Microb Cell Fact Research BACKGROUND: Analyses of substrate and metabolites are often bottleneck activities in high-throughput screening of microbial bioprocesses. We have assessed Fourier transform infrared spectroscopy (FTIR), in combination with high throughput micro-bioreactors and multivariate statistical analyses, for analysis of metabolites in high-throughput screening of microbial bioprocesses. In our previous study, we have demonstrated that high-throughput (HTS) FTIR can be used for estimating content and composition of intracellular metabolites, namely triglyceride accumulation in oleaginous filamentous fungi. As a continuation of that research, in the present study HTS FTIR was evaluated as a unified method for simultaneous quantification of intra- and extracellular metabolites and substrate consumption. As a proof of concept, a high-throughput microcultivation of oleaginous filamentous fungi was conducted in order to monitor production of citric acid (extracellular metabolite) and triglyceride lipids (intracellular metabolites), as well as consumption of glucose in the cultivation medium. RESULTS: HTS FTIR analyses of supernatant samples was compared with an attenuated total reflection (ATR) FTIR, which is an established method for bioprocess monitoring. Glucose and citric acid content of growth media was quantified by high performance liquid chromatography (HPLC). Partial least square regression (PLSR) between HPLC glucose and citric acid data and the corresponding FTIR spectral data was used to set up calibration models. PLSR results for HTS measurements were very similar to the results obtained with ATR methodology, with high coefficients of determination (0.91–0.98) and low error values (4.9–8.6%) for both glucose and citric acid estimates. CONCLUSIONS: The study has demonstrated that intra- and extracellular metabolites, as well as nutrients in the cultivation medium, can be monitored by a unified approach by HTS FTIR. The proof-of-concept study has validated that HTS FTIR, in combination with Duetz microtiter plate system and chemometrics, can be used for high throughput screening of microbial bioprocesses. It can be anticipated that the approach, demonstrated here on single-cell oil production by filamentous fungi, can find general application in screening studies of microbial bioprocesses, such as production of single-cell proteins, biopolymers, polysaccharides, carboxylic acids, and other type of metabolites. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1186/s12934-017-0817-3) contains supplementary material, which is available to authorized users. BioMed Central 2017-11-13 /pmc/articles/PMC5683213/ /pubmed/29132358 http://dx.doi.org/10.1186/s12934-017-0817-3 Text en © The Author(s) 2017 Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. 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. |
spellingShingle | Research Kosa, Gergely Shapaval, Volha Kohler, Achim Zimmermann, Boris FTIR spectroscopy as a unified method for simultaneous analysis of intra- and extracellular metabolites in high-throughput screening of microbial bioprocesses |
title | FTIR spectroscopy as a unified method for simultaneous analysis of intra- and extracellular metabolites in high-throughput screening of microbial bioprocesses |
title_full | FTIR spectroscopy as a unified method for simultaneous analysis of intra- and extracellular metabolites in high-throughput screening of microbial bioprocesses |
title_fullStr | FTIR spectroscopy as a unified method for simultaneous analysis of intra- and extracellular metabolites in high-throughput screening of microbial bioprocesses |
title_full_unstemmed | FTIR spectroscopy as a unified method for simultaneous analysis of intra- and extracellular metabolites in high-throughput screening of microbial bioprocesses |
title_short | FTIR spectroscopy as a unified method for simultaneous analysis of intra- and extracellular metabolites in high-throughput screening of microbial bioprocesses |
title_sort | ftir spectroscopy as a unified method for simultaneous analysis of intra- and extracellular metabolites in high-throughput screening of microbial bioprocesses |
topic | Research |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5683213/ https://www.ncbi.nlm.nih.gov/pubmed/29132358 http://dx.doi.org/10.1186/s12934-017-0817-3 |
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