Tissues (15 mg) were homogenized in a solvent mixture of MeOH:ACN:H2O (50:30:20) in a TissueLyser II (Qiagen) at 4°C, followed by flash freezing in liquid nitrogen, and then centrifuged and passed through a Nanosep 3K molecular weight cutoff filter (Life Sciences Technologies). Tissue lysates were mixed 1:1 with 13C labeled internal standard mix balanced for the metabolites of interest. The balanced internal standard was generated by combining IROA yeast extract (IROA Technologies) with 13C-labeled lysates from several human cell lines grown in the presence of 5.5 mM U-13C glucose (Cambridge Isotopes) for 3 passages.

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