Cp sil 8 cb low bleed

A volume of 1 μl aliquot of derivatized sample was injected in splitless mode and separated on a 30 m × 0.25 mm DB-5MS capillary column coated with a 0.25-µm CP-Sil 8 CB low bleed (Varian Inc., Palo Alto, CA, USA). Helium was used as the carrier gas. The front inlet purge flow was 3 ml min⁻¹. The gas flow rate through the column was 1 ml min⁻¹. The initial temperature was kept at 50°C for 1 min. Then, it was raised to 310°C at a rate of 10°C min⁻¹ and kept for 8 min at 310°C. The injection, transfer line, and ion source temperatures were 280°C, 280°C, and 250°C, respectively. The energy was −70 eV in the electron impact mode. The mass spectrometry data were acquired in the full-scan mode with the 50–500-m/z range at 12.5 spectra per second after a solvent delay of 6.27 min.

Each FAME sample (1 μL) was injected into the Agilent 7890A GC with an Agilent 7683B autosampler (Agilent, Atlanta, GA, USA) with a split ratio of 25 and separated in a 30 m × 0.25 mm I.D. fused-silica capillary column coated with 0.25 μm CP-SIL 8 CB Low Bleed (Varian Inc., Palo Alto, CA, USA). The injector temperature was 230 °C. The helium gas flow rate through the column was 1.0 mL/min. The temperature program was as follows: starting temperature 80 °C, maintained for 2 min, followed by an increase to 320 °C at 15 °C/min and a 10 min hold at 320 °C. The transfer line and ion-source temperatures were 250 and 200 °C, respectively. The scanned mass range was 85–600 m/z, and the detector voltage was set at 1700 V.

Dried extracts were derivatized using a method previously described [8 (link), 17 (link)]. A fused silica capillary column (CP-Sil 8 CB low bleed, 30 m × 0.25 mm i.d., Film Thickness 0.25 µm; Varian Inc., Palo Alto, CA) was used for separation of the metabolites. A Shimadzu GCMS-QP-2010 system (Kyoto, Japan) was used for the detection of ALA and other metabolites. GC–MS analysis was performed using a modified method previously described [8 (link)]. In particular, m/z 174 was added to the ion monitoring channels for detection of ALA. A calibration curve was obtained from the ratio of ion peak areas of ALA and adipic acid as an internal standard detected at m/z 111.