Table 4. Extraction Buffer (EB) composition.

Component		Volume added	Final concentration
HPLC grade acetonitrile	5771.25 μl		71.25%
HPLC grade methanol	1923.75 μl		23.75%
HPLC grade formic acid	15.39 μl		1.9%
~15 mg of isotopically labeled yeast extract (ISO1) dissolved in 1.5 ml of HPLC grade water. Note: After adding water to the yeast extract, dissolve the yeast extract by vortexing and/or rocking the yeast extract and water at 4 °C for approximately 30 min. Solution can be stored at -80 °C although some metabolites will degrade over time (see manufacturer’s instructions).	405 μl		5%
2 mM solution of 2H9 choline prepared in HPLC grade water (stored at -20 °C)	4.03 μl		1 μM
50 mM solution of 13C4 3-hydroxybutyrate prepared in HPLC grade water (stored at -20 °C)	0.81 μl		5 μM
200 μM solution of 13C6 15N2 cystine prepared in HPLC grade water (stored at -20 °C)	81 μl		2 μM
100 mM solution of 13C3 lactate prepared in HPLC grade water (stored at -20 °C)	16.2 μl		200 μM
57.3 mM solution of 13C6 glucose prepared in HPLC grade water (stored at -20 °C)	7.05 μl		50 μM
100 mM solution of 13C3 serine prepared in HPLC grade water (stored at -20 °C)	1.62 μl		20 μM
750 mM solution of 13C2 glycine prepared in HPLC grade water (stored at -20 °C)	1.62 μl		150 μM
2 mM solution of 13C5 hypoxanthine prepared in HPLC grade water (stored at -20 °C)	2.02 μl		0.5 μM
200 mM solution of 13C2 15N taurine prepared in HPLC grade water (stored at -20 °C)	2.02 μl		50 μM
60 mM solution of 13C3 glycerol prepared in HPLC grade water (stored at -20 °C)	2.02 μl		15 μM
4 mM solution of 2H3 creatinine prepared in HPLC grade water (stored at -20 °C)	2.02 μl		1 μM