Figure 1
From: Quantification of folate metabolism using transient metabolic flux analysis
tMFA protocol and validation. (a) Simplified diagram of one-carbon metabolism. (b) Schematic representation of changes in isotope fractions after addition of a tracer in the culture media at plating. The white bars represent the unlabeled fraction from the intracellular metabolite present at plating, the cyan bar the unlabeled metabolite synthesized from unlabeled precursors and the red bar the labeled metabolite synthesized from labeled precursors. f denotes the rate of synthesis and k the rate of turnover per unit of metabolite. (c) Changes in the isotope pools due to inhibition of metabolite synthesis. (d) Sketch of the tMFA protocol. (e-h) Isotope fractions of serine and glycine in MCF7 cells, untreated or treated with MTX (black, M+0; red, M+1; green, M+2, and blue, M+3). (i-l) Estimated turnover rate of purines and glutathione (GSH) in MCF7 cells, untreated or treated with MTX (black lines, experiment 1, LC-MS quantification; circle, experiment 2, CE-MS quantification). The lines/points represent the average and the error bars the standard deviation over three replicates. (m-p) Comparison between the predicted isotope fractions based on turnover estimates for Exp 2 (dashed line) and the isotope fractions measured in Exp 1 (solid line), for both purines and glutathione in untreated and MTX-treated cells.