Bpx 5

This measurement reference to the previous our reported methods [46 (link)]. GC-MS analysis was performed using a GC-MS QP2010 Ultra spectrometer (Shimadzu) with a fused silica capillary column (BPX-5; 30 m × 0.25 mm inner diameter, film thickness: 0.25 μm; Shimadzu) and a front inlet temperature of 25 °C, and a helium gas flow rate through the column of 39.0 cm/s. The column temperature was held at 60 °C for 2 min, then raised by 15 °C/min to 330 °C and maintained for 3 min. The interface and ion source temperatures were 280 and 200 °C, respectively. To perform a semi-quantitative assessment, the peak height of each quantified ion was calculated and normalized using citrate-d4 and 2-isopropylmalate peak height and protein concentration. Protein concentrations were determined with Bio-Rad Quick Start Bradford 1x Dye Reagent according to the manufacturer’s instructions. The retention times and SRM conditions for the derivatized metabolites are summarized in Table S3.

These measurements were performed with reference to the previously reported methods [43 (link),44 (link),45 (link)]. GC-MS analysis was performed using a GC-MS QP2010 Ultra (Shimadzu, Kyoto, Japan) with a fused silica capillary column (BPX-5; 30 m × 0.25 mm inner diameter, film thickness: 0.25 μm; Shimadzu), a front inlet temperature of 250 °C, and a helium gas flow rate through the column of 39.0 cm/s. The column temperature was maintained at 60 °C for 2 min, then raised by increments of 15 °C/min to 330 °C, and maintained at that temperature for 3 min. The interface and ion source temperatures were 280 °C and 200 °C, respectively. All data obtained by GC-MS analysis were analyzed using MetaboAnalyst software (v. 5.0; Reifycs, Inc., Tokyo, Japan). The retention times indicated in the Smart Metabolites Database (Shimadzu) were used as references to create a library for data analysis. To perform a semi-quantitative assessment, the peak area of each quantified compound was calculated and normalized using the 2-isopropylmalate peak area.

This measurement was based on our previously reported methods [48 (link)]. GC-MS analysis was performed using a GC-MS QP2010 Ultra (Shimadzu Corp., Kyoto, Japan) with a fused silica capillary column (BPX-5; 30 m × 0.25 mm inner diameter, film thickness: 0.25 μm; Shimadzu Corporation, Kyoto, Japan) and a front inlet temperature of 250 °C and helium gas flow rate through a column of 39.0 cm/s. The column temperature was held at 60 °C for 2 min, then raised by 15 °C/min to 330 °C and maintained for 3 min. The interface and ion-source temperatures were 280 °C and 200 °C, respectively. To perform a semi-quantitative assessment, the peak height of each quantified ion was calculated and normalized using the citrate-d₄ and 2-isopropylmalate peak heights and protein concentrations. The retention times and selected reaction monitoring conditions of the derivatized metabolites are summarized in Table S1.