Periodic Reporting for period 4 - SUMMIT (Site-specific Ultrasensitive Magnetic resonance of Mixtures for Isotopic Tracking)
Período documentado: 2024-04-01 hasta 2025-09-30
Observing and understanding the matter around us is at the very heart of analytical chemistry. Nuclear Magnetic Resonance (NMR) spectroscopy is a unique approach that provides a snapshot of the molecular content of complex samples. This includes in particular biological samples (biofluids, cell or plant extracts, bacterial or cell culture media, agri-food matrices, etc.). NMR spectroscopy is particularly well suited to metabolomics approaches, which aim to collect a maximum of measurable data on the nature and concentration of small molecules involved in the chemistry of living organisms. It also provides access to valuable information on the nature and abundance of certain stable isotopes, thus providing data to better interpret metabolic pathways.
In this context, the aim of the SUMMIT project was to develop new analytical chemistry approaches in order to improve the sensitivity and resolution of NMR spectroscopy for the analysis of complex biological mixtures, and in particular for applications in metabolomics. The developments were based on a set of innovative methodological developments. These included, in particular, hyperpolarization methods relying on a prototype nuclear dynamic polarization equipment.
The SUMMIT project also included the development of novel signal detection methods based on ultrafast two-dimensional NMR spectroscopy, a method that makes it possible to better separate the signals of compounds in a mixture. The development of signal processing methods was also an objective.
Finally, we aimed at evaluating the potential of these sensitive and rapid detection methods for a broad variety of biological issues. More specifically, the project goal was to demonstrate the potential of these methods for omic sciences such as metabolomics.
In this context, the aim of the SUMMIT project was to develop new analytical chemistry approaches in order to improve the sensitivity and resolution of NMR spectroscopy for the analysis of complex biological mixtures, and in particular for applications in metabolomics. The developments were based on a set of innovative methodological developments. These included, in particular, hyperpolarization methods relying on a prototype nuclear dynamic polarization equipment.
The SUMMIT project also included the development of novel signal detection methods based on ultrafast two-dimensional NMR spectroscopy, a method that makes it possible to better separate the signals of compounds in a mixture. The development of signal processing methods was also an objective.
Finally, we aimed at evaluating the potential of these sensitive and rapid detection methods for a broad variety of biological issues. More specifically, the project goal was to demonstrate the potential of these methods for omic sciences such as metabolomics.
During the project, we have set up the new analytical methodologies which form the basis of the SUMMIT project, and we explored some groundbreaking applications
On the methodological side, the main achievements include:
• The fine optimization of our prototype dissolution-DNP experimental setting for the robust and reproducible analysis of complex metabolic mixtures;
• The development of novel ultrafast 2D NMR experiments for the rapid screening of complex chemical samples (which turned out to be useful for other applications such as reaction monitoring);
• The development of a new NMR detection approach for hyperpolarized experiment, consisting in recording 1H and 13C spectra of a single hyperpolarized sample within the same experiment;
• The comprehensive description of concepts underlying ultrafast 2D NMR for analytical chemistry applications;
• The development of dedicated processing tools to extract relevant data from 2D spectra of complex mixtures.
On the application side, we have demonstrated for the first time that dissolution-DNP can provide relevant information in a full metabolomics study based on 13C NMR at natural isotopic abundance. First, we demonstrated that our approach could extract biologically relevant information on plant metabolism by analyzing tomato fruit extracts at different development stages. Then, thanks to a careful optimization of experimental parameters, we demonstrated that 13C spectra of urine at natural abundance could be recorded in a single scan with a sensitivity equivalent to that of a 1H NMR experiment, with an analytical performance that made it possible to quantify metabolites that could not be measured with conventional NMR. In the last part of the project, we collaborated with clinicians in Nantes to perform the first clinical metabolomics study involving hyperpolarized NMR metabolomics. We demonstrated that hyperpolarized 13C at natural abundance provided complementary information compared to conventional NMR, leading to the detection of two biomarkers that were not detected by conventional 1H NMR metabolomics. This final result, which is beyond the expectations of the SUMMIT project, opens many application perspectives. It paves the way to many different applications which will be explored in the near future, through the MetaboHub national research infrastructure cand through various funded projects.
In addition to these scientific advances, we organized two international symposiums, one on hyperpolarized NMR for the analysis of mixtures (in 2022), and another one on NMR metabolomics (in 2025). These events were a unique opportunity to present the project results to a broad audience of top international experts, to update the team on the latest advances in the field, to discuss potential collaborations, and for students to network and discuss career opportunities.
In conclusion, the results of the SUMMIT project should have a broad impact for the many applications that require for the analysis of complex samples by NMR spectroscopy, in particular metabolomics. It provides new analytical tools for a broad community of researchers in chemistry and biology.
On the methodological side, the main achievements include:
• The fine optimization of our prototype dissolution-DNP experimental setting for the robust and reproducible analysis of complex metabolic mixtures;
• The development of novel ultrafast 2D NMR experiments for the rapid screening of complex chemical samples (which turned out to be useful for other applications such as reaction monitoring);
• The development of a new NMR detection approach for hyperpolarized experiment, consisting in recording 1H and 13C spectra of a single hyperpolarized sample within the same experiment;
• The comprehensive description of concepts underlying ultrafast 2D NMR for analytical chemistry applications;
• The development of dedicated processing tools to extract relevant data from 2D spectra of complex mixtures.
On the application side, we have demonstrated for the first time that dissolution-DNP can provide relevant information in a full metabolomics study based on 13C NMR at natural isotopic abundance. First, we demonstrated that our approach could extract biologically relevant information on plant metabolism by analyzing tomato fruit extracts at different development stages. Then, thanks to a careful optimization of experimental parameters, we demonstrated that 13C spectra of urine at natural abundance could be recorded in a single scan with a sensitivity equivalent to that of a 1H NMR experiment, with an analytical performance that made it possible to quantify metabolites that could not be measured with conventional NMR. In the last part of the project, we collaborated with clinicians in Nantes to perform the first clinical metabolomics study involving hyperpolarized NMR metabolomics. We demonstrated that hyperpolarized 13C at natural abundance provided complementary information compared to conventional NMR, leading to the detection of two biomarkers that were not detected by conventional 1H NMR metabolomics. This final result, which is beyond the expectations of the SUMMIT project, opens many application perspectives. It paves the way to many different applications which will be explored in the near future, through the MetaboHub national research infrastructure cand through various funded projects.
In addition to these scientific advances, we organized two international symposiums, one on hyperpolarized NMR for the analysis of mixtures (in 2022), and another one on NMR metabolomics (in 2025). These events were a unique opportunity to present the project results to a broad audience of top international experts, to update the team on the latest advances in the field, to discuss potential collaborations, and for students to network and discuss career opportunities.
In conclusion, the results of the SUMMIT project should have a broad impact for the many applications that require for the analysis of complex samples by NMR spectroscopy, in particular metabolomics. It provides new analytical tools for a broad community of researchers in chemistry and biology.
Several significant advances beyond the state of the art have been made.
On the methodological side, the main advances are:
-The fine optimization of a dissolution-DNP experimental setting for 13C of metabolic samples with high repeatability and sensitivity (Dey et al., Magn. Reson.2022 3, 183-202)
-The development of a new NMR method to detect highly sensitive 1H and 13C spectra from a single hyperpolarized sample at natural abundance (Dey et al., Anal. Chem. 2023, 95, 16861-16867)
-The development of tailored ultrafast 2D NMR experiments for the analysis of hyperpolarized samples (Praud et al., Anal. Methods 2023, 15, 6209-6219)
-The development of a 2D data processing workflow for 2D NMR metabolomics (publication in preparation)
On the application side, the main progress beyond the state of the art includes:
-The first proof-of-concept that dissolution-DNP can provide relevant information in a full metabolomics study, opening the way to many applications in the SUMMIT project and beyond (Dey et al, Anal. Chem. 2020, 92, 14867);
-The first demonstration that 13C hyperpolarized spectra of urine can be recorded at natural abundance with high resolution and sensitivity (Ribay et al., Angew. Chem. Int. Ed. 2023, e202302110)
-The first clinical metabolomics study (of chronic kidney disease urine samples) relying on the hyperpolarized 13C NMR methodology developed in the SUMMIt project (Ribay et al., J. Am. Chem. Soc. 2025, 147, 644-650)
On the methodological side, the main advances are:
-The fine optimization of a dissolution-DNP experimental setting for 13C of metabolic samples with high repeatability and sensitivity (Dey et al., Magn. Reson.2022 3, 183-202)
-The development of a new NMR method to detect highly sensitive 1H and 13C spectra from a single hyperpolarized sample at natural abundance (Dey et al., Anal. Chem. 2023, 95, 16861-16867)
-The development of tailored ultrafast 2D NMR experiments for the analysis of hyperpolarized samples (Praud et al., Anal. Methods 2023, 15, 6209-6219)
-The development of a 2D data processing workflow for 2D NMR metabolomics (publication in preparation)
On the application side, the main progress beyond the state of the art includes:
-The first proof-of-concept that dissolution-DNP can provide relevant information in a full metabolomics study, opening the way to many applications in the SUMMIT project and beyond (Dey et al, Anal. Chem. 2020, 92, 14867);
-The first demonstration that 13C hyperpolarized spectra of urine can be recorded at natural abundance with high resolution and sensitivity (Ribay et al., Angew. Chem. Int. Ed. 2023, e202302110)
-The first clinical metabolomics study (of chronic kidney disease urine samples) relying on the hyperpolarized 13C NMR methodology developed in the SUMMIt project (Ribay et al., J. Am. Chem. Soc. 2025, 147, 644-650)