Untargeted metabolomics-based approach using UHPLC-HRMS to authenticate carrots (Daucus carota L.) based on geographical origin and production mode.

Carrots produced in different agricultural regions with organic or conventional mode were analyzed by untargeted UHPLC-HRMS using reversed-phase and HILIC modes. Data were first treated separately, and further combined to possibly improve results. An in-house data processing workflow was applied to identify relevant features after peak detection. Based on these features, discrimination models were built using chemometrics. A tentative annotation of chemical markers was performed using online databases and UHPLC-HRMS/MS analyses. An independent set of samples was analyzed to assess the discrimination potential of these markers. Carrots produced in the New Aquitaine region could be successfully discriminated from carrots originating from the Normandy region by an OLPS-DA model. Arginine and 6-methoxymellein could be identified as potential markers with the C18-silica column. Additional markers (N-acetylputrescine, l-carnitine) could be identified thanks to the polar column. Discrimination based on production mode was more challenging: some trend was observed but model metrics remained unsatisfactory.

Optimized slice-selective 1H NMR experiments combined with highly accurate quantitative 13C NMR using an internal reference method.

Isotope ratio monitoring by 13C NMR spectrometry (irm-13C NMR) provides the complete 13C intramolecular position-specific composition at natural abundance. It represents a powerful tool to track the (bio)chemical pathway which has led to the synthesis of targeted molecules, since it allows Position-specific Isotope Analysis (PSIA). Due to the very small composition range (which represents the range of variation of the isotopic composition of a given nuclei) of 13C natural abundance values (50‰), irm-13C NMR requires a 1‰ accuracy and thus highly quantitative analysis by 13C NMR. Until now, the conventional strategy to determine the position-specific abundance xi relies on the combination of irm-MS (isotopic ratio monitoring Mass Spectrometry) and 13C quantitative NMR. However this approach presents a serious drawback since it relies on two different techniques and requires to measure separately the signal of all the carbons of the analyzed compound, which is not always possible. To circumvent this constraint, we recently proposed a new methodology to perform 13C isotopic analysis using an internal reference method and relying on NMR only. The method combines a highly quantitative 1H NMR pulse sequence (named DWET) with a 13C isotopic NMR measurement. However, the recently published DWET sequence is unsuited for samples with short T1, which forms a serious limitation for irm-13C NMR experiments where a relaxing agent is added. In this context, we suggest two variants of the DWET called Multi-WET and Profiled-WET, developed and optimized to reach the same accuracy of 1‰ with a better immunity towards T1 variations. Their performance is evaluated on the determination of the 13C isotopic profile of vanillin. Both pulse sequences show a 1‰ accuracy with an increased robustness to pulse miscalibrations compared to the initial DWET method. This constitutes a major advance in the context of irm-13C NMR since it is now possible to perform isotopic analysis with high relaxing agent concentrations, leading to a strong reduction of the overall experiment time.