Cells were seeded in six-well plates at 100,000 cells per well (BT549 and CAL-120) or 200,000 cells per well (MDA-MB-468) in 2 ml of DMEM-F12 or Plasmax (both supplemented with 2.5% FBS). After 24 hours (day 1), culture medium was changed to 8 ml of DMEM containing 700 μM 13C6 arginine (Cambridge Isotope Laboratories, MA, USA) (cells seeded in DMEM-F12) or Plasmax (as such or supplemented to 700 μM Arg with 13C6 arginine; cells seeded in Plasmax). After 48 hours (day 3), cells were extracted in 400 μl of ice-cold extraction solution, and samples were analyzed by LC/MS as described above.

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