Agilent gc ms 5975 7890

Following induction and incubation, cell cultures were centrifuged, and the supernatant decanted. In a typical pentadecane assay, 3 ml of supernatant vigorously mixed with 1 ml of 1M NaCl and 1 ml pure high-performance liquid chromatography (HPLC) grade ethyl acetate (Sigma Aldrich). After 1 h of shaking, the ethyl acetate extract was separated by centrifugation (10 min at 4000 g) and obtaining the top layer. Extracts were analyzed by gas chromatography–mass spectrometry (GC-MS) on an Agilent GC-MS 5975/7890 (Agilent Technologies) with the HP-5MS column (30 m length, 0.50 mm diameter). Following a 1–3 μl split-less injection, the inlet temperature was held at 100°C for 3 min and ramped up to 320°C at 30°C per minute. For quantification, standard curves of pentadecane and hexadecanol were generated, by using the pure compounds (Sigma Aldrich). Succinic acid production assays were similarly performed on decanted supernatant. Roche (r-biopharm) enzymatic succinic acid assay kits (CN: 10176281035) were used to measure production levels using a Victor3V 1420 Multilabel plate reader (Perkin Elmer).