pSIRM workflow including the determination and calculation of stable isotope incorporation. (A) Data analysis workflow from GC-MS raw data up to quantification and the calculation of 13C-label incorporation. (B) Map of the central carbon metabolism highlighting detection and quantification of intermediates as well as evaluation of stable isotope enrichment after application of 13C-glucose or 13C-glutamine in cell culture experiments. (C) Visualization of mass isotopomer fractions (MIF) of a mass fragment of 12C-glucose (left) and a 1:1 mixture of 12C- and 13C1-glucose (right). Targeted (Formula 1.1–1.3) and position-independent (Formula 2) calculation strategies provide the determination of 13C-label incorporation (L in (%)). (D) Calculation strategies were validated by the analysis of known ratios of 12C-glucose: 13C1-glucose. Shown are expected versus measured stable isotope incorporation comparing both correction strategies.