Liquid State and HR-MAS Quantification of Absolute Concentration of Choline Derivatives by 14N-NMR

Abstract

Nuclear Magnetic Resonance (NMR) spectroscopy is a powerful tool used by biologists and chemists1. The study of the use of NMR for 14N, the most abundant nitrogen isotope, is lacking. Research here is limited because 14N is a spin 1 nucleus and therefore has a large quadrupole moment, resulting in extremely wide peaks in NMR for most compounds that are hard to handle typical conditions. However, some nitrogen containing molecules, such as choline, which are small and nearly symmetrically substituted, show strong, sharp peaks and therefore using 14N NMR to study these compounds could be a selective spectroscopic method. Choline and its derivatives, such as phosphocholine, are important biomarkers because choline plays an important role in the cell cycle. Choline metabolism is involved in activity of growth factors in cells, and abnormalities in choline metabolism have been linked to metastasis certain cancers, as well as Alzheimer’s and male infertility. Cancer treatment has potential to be tracked by phosphocholine levels dropping to normal levels, and using the unique selectivity of 14N NMR to quantify the concentrations of choline and its derivatives can prove to be a very useful tool in cancer studies and diagnosis. Several methods of quantification of samples have been developed and tested using 2D 1H,14N HSQC NMR experiments, both in liquid state and using HR-MAS technology. The clean spectra allow for concentration analysis via several methods, even in complex and heterogeneous mixtures. This work will provide the foundation for using these methods in a variety of biological samples.