NMR-Based Isotopologue Profiling of Microbial Carotenoids

C-Isotopologue profiling is a powerful tool to determine on a quantitative basis the biosynthetic origin of carotenoids in microorganisms. To this aim, the carotenoid-producing microorganism is grown in medium containing ¹³ C-labeled glucose. After growth, the ¹³ C-isotopologue distribution in a given biosynthetic carotenoid is determined by quantitative NMR spectroscopy. The labeling pattern provides a fingerprint of processes involved in the metabolism of glucose and the formation of the carotenoid. For example, the ¹³ C-profile shows whether the isoprenoid precursors, isopentenyl diphosphate and dimethylallyl diphosphate, are formed by the mevalonate or the non-mevalonate pathway. The labeling data also specify the pathways of glucose utilization, e.g., via the Entner-Doudoroff pathway or glycolysis. The method is exemplified with the analysis of zeaxanthin biosynthesis in the Alphaproteobacterium, Paracoccus zeaxanthinifaciens .