Hydrogen flame ionization detector

The preparation of fatty acid methyl esters were performed by the methylation of samara oil following the AOCS method [10 ]. The separation of fatty acid methyl esters was conducted on an Agilent 6890A gas chromatography combined with a hydrogen flame ionization detector (Agilent, Santa Clara, CA, USA) and a VF-23MS capillary column (30 m × 0.25 mm i.d., 0.25 μm film thickness; Agilent, Santa Clara, CA, USA). High-purity nitrogen (99.999%) was used as carrier gas at a constant flow of 1 mL/min. The split ratio was 100:1, and the injection volume was 1 μL. The injection and detector temperatures were set at 260 °C. The temperature program was as follows. The initial temperature was 110 °C for 3 min, then increased to 220 °C at a rate of 4 °C/min, and held for 15 min at 220 °C. The peaks of fatty acid methyl esters were identified using their corresponding standards such as palmitic, stearic, oleic, linoleic, and linolenic acid. The results were expressed as a percentage of the oil.

To determine the fatty acid composition of the oil, we opted for the cold transesterification method. A portion of 0.1 g of the oil sample was mixed with 2 mL of heptane and 0.2 mL of methanolic potassium hydroxide solution (2 N) according to AOAC (2000) method 965.33 [22 ]. The tube was closed and shaken vigorously for 30 s. After standing, the upper part of the solution, which contained the methyl esters, cleared following its decantation. The heptane solution was thus ready for injection into the GC. Analyses of fatty acid methyl ester (FAME) were carried out using an AGILENT 8860 Series (G2790A) gas chromatograph (AGILENT technologies, Waldbronn, Germany) equipped with a hydrogen flame ionization detector (AGILENT) and a capillary column (50 m × 0.25 mm × 0.20 µm Agilent, Waldbronn, Germany). The column temperature was programmed from 180 to 220 °C at 5 °C/min and the injector and detector temperatures were set at 240 °C. Identification and quantification of FAME were accomplished by comparing the retention times of peaks with those of pure standards purchased from Sigma-Aldrich (Sigma Chemical Co., Sofia, Bulgaria) and analyzed using the same conditions. The results were expressed as a percentage of individual fatty acids in the lipid fraction.