Intracellular Metabolite 13C Isotopomer Analysis

For the ¹³C isotopomer analysis of intracellular metabolites by GC-MS, snap-frozen NHAs and HCNs were thawed and centrifuged to remove the protein precipitations. Sodium 2-oxobutyrate (50 nmol) was added to the supernatants as an internal standard. The samples were evaporated and derivatized by trimethylsilylation (Tri-Sil HTP, Thermo Scientific). A total of 3 µL of the derivatized solution was injected onto an Agilent 6970 gas chromatograph equipped with a fused silica capillary GC column (30-m length/0.25-mm diameter) coupled with an Agilent 5973 mass selective detector [51 (link)]. The measured distributions of the carbon isotopomers were corrected for the natural abundance of the ¹³C isotopomer (1.1%).