The revised version of cancer hallmarks, depicting the biological properties acquired during tumor development and progression, includes the capability to modify or reprogram cellular metabolism. High-resolution multinuclear magnetic resonance spectroscopy (MRS) provides noninvasive means of monitoring metabolites that play a central role in several pathways, measuring the rates at which reactions within the pathways take place, and investigating how these rates are controlled in such a way that a metabolic precursor and cofactors are provided according to the demand. Here we describe comprehensive methods for assaying the activity of key enzymes involved in the phosphatidylcholine metabolic pathways responsible for phosphocholine accumulation in ovarian cancer cells using high-resolution 1 H and 31 P MRS analyses of cell lysates or cytosolic preparations.