Metabolism of Choline and Deuterated Choline Detected by ¹H–¹⁴N 2D Heteronuclear Single-Quantum Coherence (HSQC) NMR

Choline is an essential nutrient that plays a critical role in tumor growth. Choline levels can be detected by proton (¹H) MR spectroscopy (MRS) <i>in vivo</i>, whereas active (dynamic) choline metabolism can be studied with deuterated choline and ²H MRS. The detected ¹H and ²H choline signals represent the sum of choline, phosphocholine (PC) and glycerophosphocholine (GPC), preventing a detailed characterization of choline metabolism. Here we have developed a two-dimensional (2D) NMR method that allows the simultaneous detection of all protonated and deuterated choline-related compounds in excised tissue. The methodology relies on the high ¹H detection sensitivity and chemical shift dispersion to distinguish between choline types, the sensitivity of the ¹⁴N chemical shift toward deuteration of nearby methyl groups and the presence of a ¹H-¹⁴N scalar coupling between ¹⁴N and CH₂ groups. With optimized sequence parameters, the utility of the method is demonstrated on extracts from cultured cancer cells, blood plasma and rat brain and brain tumor tissues.