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How metabolomics can contribute to bio-processes: a proof of concept study for biomarkers discovery in the context of nitrogen-starved microalgae grown in photobioreactors

Abstract

Microalgae appear to be one of the most promising sustainable resources as alternative crops for the production of renewable transport fuel. The exploitation of this bioresource requires, however, a fine monitoring of the culture conditions, for example by using more relevant control variables than usual macroscopic indicators (biomass or pigment estimation). In this proof of concept study, we propose to search potential biomarkers of progressive nitrogen regime culture conditions using an untargeted metabolomic approach based on LC-HRMS combined to a non-invasive analysis based on FTIR spectroscopy. One microalgae model was investigated i.e. Chlamydomonas reinhardtii to characterize the effect of progressive nitrogen regime in batch culture conditions on its metabolome. FTIR allowed assessing the intracellular macrometabolic perturbations, highlighting the over-accumulation of carbohydrates. LC-HRMS complemented the macromolecular information by revealing the dependence of microalgae metabotypes on nitrogen regime conditions tested for cells culture. Patterns of significantly modulated metabolites were also detected during those slight contrasted nitrogen regimes and interesting features were structurally elucidated. This included metabolites belonging to the pantothenate, branched chain and aromatic amino acids pathways. In the last step of this proof of concept study, amino acid targets proposed by metabolomic investigations were assessed on nitrogen-limited continuous culture on photobioreactors. This was performed to test the validity of proposed targets in real small-scale industrial production conditions. Results were very encouraging and suggested the possibility of using potentially relevant metabolites as intracellular biomarkers only (tryptophan) or as both intra and extracellular biomarkers (e.g. 2-methylbutyric acid and ketoleucine).

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Abbreviations

ATR:

Attenuated total reflectance

BCAAs:

Branched-chain amino acids

CE:

Capillary electrophoresis

FTIR:

Fourier transform infrared spectroscopy

GC:

Gas chromatography

HC:

Hierarchical clustering

HPLC:

High performance liquid chromatography

HTS-XT:

High throughput screening eXTension

LC-HRMS:

Liquid chromatography—high resolution mass spectrometry

MS:

Mass spectrometry

NMR:

Nuclear magnetic resonance

PLS-DA:

Partial least square-discriminant analysis

RSD:

Relative standard deviation

TAP:

Tris acetate phosphate

TIC:

Total ionic current

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Acknowledgments

The authors wish to acknowledge CORSAIRE metabolomics analytical facilities from BIOGENOUEST network (http://www.biogenouest.org/). Part of this work has been funded by the French National Research Agency project ALGOMICS (ANR-08-BIOE-002).

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Correspondence to Olivier Gonçalves.

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Courant, F., Martzolff, A., Rabin, G. et al. How metabolomics can contribute to bio-processes: a proof of concept study for biomarkers discovery in the context of nitrogen-starved microalgae grown in photobioreactors. Metabolomics 9, 1286–1300 (2013). https://doi.org/10.1007/s11306-013-0532-y

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Keywords

  • LC-HRMS
  • FTIR spectroscopy
  • Nitrogen progressive regime
  • Chlamydomonas reinhardtii
  • Biofuel
  • Photobioreactors