Validation of carbon isotopologue distribution measurements by GC-MS and application to C-metabolic flux analysis of the tricarboxylic acid cycle in leaves.

The estimation of metabolic fluxes in photosynthetic organisms represents an important challenge that has gained interest over the last decade with the development of C-Metabolic Flux Analysis at isotopically non-stationary steady-state. This approach requires a high level of accuracy for the measurement of Carbon Isotopologue Distribution in plant metabolites. But this accuracy has still not been evaluated at the isotopologue level for GC-MS, leading to uncertainties for the metabolic fluxes calculated based on these fragments.

Metabolomics and modelling approaches for systems metabolic engineering.

Metabolic engineering involves the manipulation of microbes to produce desirable compounds through genetic engineering or synthetic biology approaches. Metabolomics involves the quantitation of intracellular and extracellular metabolites, where mass spectrometry and nuclear magnetic resonance based analytical instrumentation are often used. Here, the experimental designs, sample preparations, metabolite quenching and extraction are essential to the quantitative metabolomics workflow.

An optimization method for untargeted MS-based isotopic tracing investigations of metabolism.

Introduction: Stable isotope tracer studies are increasingly applied to explore metabolism from the detailed analysis of tracer incorporation into metabolites. Untargeted LC/MS approaches have recently emerged and provide potent methods for expanding the dimension and complexity of the metabolic networks that can be investigated. A number of software tools have been developed to process the highly complex MS data collected in such studies; however, a method to optimize the extraction of valuable isotopic data is lacking.

Exploring the Glucose Fluxotype of the y-ome Using High-Resolution Fluxomics.

We have developed a robust workflow to measure high-resolution fluxotypes (metabolic flux phenotypes) for large strain libraries under fully controlled growth conditions. This was achieved by optimizing and automating the whole high-throughput fluxomics process and integrating all relevant software tools. This workflow allowed us to obtain highly detailed maps of carbon fluxes in the central carbon metabolism in a fully automated manner.

Recent advances in high-throughput C-fluxomics.

The rise of high throughput (HT) strain engineering tools accompanying the area of synthetic biology is supporting the generation of a large number of microbial cell factories. A current bottleneck in process development is our limited capacity to rapidly analyze the metabolic state of the engineered strains, and in particular their intracellular fluxes. HT C-fluxomics workflows have not yet become commonplace, despite the existence of several HT tools at each of the required stages.