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(13)CO₂ pulse-chase experiments monitored by high-resolution NMR spectroscopy and mass spectrometry can provide (13)C-isotopologue compositions in biosynthetic products. Experiments with a variety of plant species have documented that the isotopologue profiles generated with (13)CO₂ pulse-chase labeling are directly comparable to those that can be generated by the application of [U-(13)C₆]glucose to aseptically growing plants. However, the application of the (13)CO₂ labeling technology is not subject to the experimental limitations that one has to take into account for experiments with [U-(13)C₆]glucose and can be applied to plants growing under physiological conditions, even in the field. In practical terms, the results of biosynthetic studies with (13)CO₂ consist of the detection of pairs, triples and occasionally quadruples of (13)C atoms that have been jointly contributed to the target metabolite, at an abundance that is well above the stochastic occurrence of such multiples. Notably, the connectivities of jointly transferred (13)C multiples can have undergone modification by skeletal rearrangements that can be diagnosed from the isotopologue data. As shown by the examples presented in this review article, the approach turns out to be powerful in decoding the carbon topology of even complex biosynthetic pathways.

Decoding Biosynthetic Pathways in Plants by Pulse-Chase Strategies Using 13CO2 as a Universal Tracer