Targeted Compound Selection with Increased Sensitivity in ¹³C-Enriched Biological and Environmental Samples Using...

Chemical characterization of complex mixtures by Nuclear Magnetic Resonance (NMR) spectroscopy is challenging due to a high degree of spectral overlap and inherently low sensitivity. Therefore, NMR experiments that reduce overlap and increase signal intensity hold immense potential for the analysis of mixtures such as biological and environmental media. Here, we introduce a ¹³C version of DREAMTIME (Designed Refocused Excitation And Mixing for Targets In Vivo and Mixture Elucidation) NMR, which, when analyzing ¹³C-enriched materials, allows the user to selectively detect only the compound(s) of interest and remove all other peaks in a ¹³C spectrum. Selected peaks can additionally be "focused" into sharp "spikes" to increase sensitivity. ¹³C-DREAMTIME is first demonstrated at high field strength (500 MHz) with simultaneous selection of eight amino acids in a ¹³C-enriched cell free amino acid mixture and of six metabolites in an extract of ¹³C-enriched green algae and demonstrated at low field strength (80 MHz) with a standard solution of ¹³C-d-glucose and ¹³C-l-phenylalanine. ¹³C-DREAMTIME is then applied at high-field to analyze metabolic changes in ¹³C-enrichedDaphnia magna after exposure to polystyrene "microplastics," as well as at low-field to track fermentation of ¹³C-d-glucose using wine yeast. Ultimately, ¹³C-DREAMTIME reduces spectral overlap as only selected compounds are recorded, resulting in the detection of analyte peaks that may otherwise not have been discernable. In combination with focusing, up to a 6-fold increase in signal intensity can be obtained for a given peak. ¹³C-DREAMTIME is a promising experiment type for future reaction monitoring and for tracking metabolic processes with ¹³C-enriched compounds.