7890a gc system

The fatty acids from the collected cells were extracted using Folch mixture [57 (link)] (2:1, chloroform:methanol), and heneicosanoic acid (21:0) was added as an internal standard to the collected cells. The fatty acids were saponified and methylated with KOH and BF₃ in methanol. Extraction of the obtained fatty acid methyl esters was then carried out with hexane. They were then analyzed by gas chromatography, with the use of an Agilent Technologies 7890A GC System (SUPELCOWAX™ 10 Capillary GC Column (15 m × 0.1 mm × 0.1 μm)) (Supelco, Bellefonte, PA, USA). The following chromatographic conditions were applied: from an initial temperature of 60 °C increasing at a rate of 40 °C/min to 160 °C, then increasing at a rate of 30 °C/min to 190 °C, and then increasing at a rate of 30 °C/min to 230 °C for 2.6 min, where it was maintained for 4.9 min. The total analysis took approximately 8 min. The gas flow rate was 0.8 ml/min; the carrier gas was comprised of hydrogen. The identification of fatty acids was done by comparing their retention times with those of commercially available standards. The fatty acid concentrations were determined based on standard curves and were expressed in mg/ml.

Gas chromatography was performed using the Agilent Technologies 7890A GC System (SUPELCOWAX™ 10) using a Capillary GC Column (15 mm × 0.10 mm, 0.10 μm; Supelco, Bellefonte, PA, United States). The following chromatographic conditions were used: initial temperature of 60°C for 0 min, increased at a rate of 40°C/min to 160°C (0 min), next increased at a rate of 30°C/min to 190°C (0.5 min) and then increased at a rate of 30°C / min to 230°C for 2.6 min, and then maintained for 4.9 min. The total analysis lasted ca. 8 min, and the gas flow rate was 0.8 ml/min. Hydrogen was used as the carrier gas. Fatty acids were identified by comparing their retention times with those of commercially available standards. C:18:2n6t, C:18:4 and C:20:3n3 cis-11 were not isolated in either the PCOS or control group.

Aliquots of 400 μL of urine sampled from the collection container and from the blank sample were mixed with 40 μL of an internal standard (myo-inositol, 20 mg/mL) and lyophilized. Subsequently, we added 200 μL of anhydrous pyridine in hydroxylamine (25 mg/mL) to samples, mixed them, and heated them to 70 °C for 1 h; then, we centrifuged the mixture at 800× g for 5 min and collected 200 μL of supernatant. Then, we silylated the sugars with 100 µL of N-trimethylsilylimidazole for 30 min at 70 °C.
Finally, we assayed sugars by gas chromatography with an Agilent Technologies 7890A GC System and capillary column (15 m × 0.530 mm, 1.50 μm), (Supelco, Bellefonte, PA, USA). Chromatographic conditions were an initial temperature of 220 °C for 5 min, an increase at a rate of 10 °C/min for 2 min, an increase at a rate of 5 °C/min for 4 min and eventually an increase at a rate of 3.5 °C/min for 4 min to a final temperature of 274 °C, which was maintained for 7 min.
Hydrogen was the carrier gas. Lactulose, mannitol, and myo-inositol retention times in the samples were compared to those produced by commercially available standards.