Focus gc fid

A Thermo Scientific Focus GC-FID with an auto-sampler (AS3000) was used for detection of ethanol production. A set of ethanol standards were prepared to determine the concentration of ethanol produced during the fermentation. A 5 % (v/v) n-propanol internal standard was added to each of the standards and samples to determine reproducibility of instrument response. Stored frozen samples were thawed and the cells pelleted by centrifugation. Then 500 µL of the supernatant were transferred to a GC vial and 450 µL of nanopure water and the internal standard (50 µL) were added for a final volume of 1 mL. Duplicate injection volumes of 0.7 µL were used; the inlet and detector were set to 250 °C. The carrier gases, helium (175 kPa), compressed air (220 kPa), and nitrogen (500 kPa), were at a constant flow of 1.2 mL/min. Each GC separation was run for 2.5 min at 75 °C.

An aliquot (1 μL) of organic fraction of each sample was submitted for analysis of volatile components. The analyses were carried on a GC-MS Thermo Focus DSQ II (Thermo Fisher Scientific, Austin, TX, USA), under the following conditions: DB-5ms (30 m × 0.25 mm; 0.25 mm film thickness, Agilent, Santa Clara, CA, USA) fused-silica capillary column; programmed temperature, 60–240 °C (3 °C/min); injector temperature, 250 °C; carrier gas, helium, adjusted to a linear velocity of 32 cm/s (measured at 100 °C); injection type, split (2 μL); split flow was adjusted to yield 20:1 ratio; septum sweep was a constant 10 mL/min; EIMS (Electron Impact Mass Spectrometry), electron energy, 70 eV; temperature of the ion source and connection parts, 200 °C. The quantitative data regarding the volatile constituents were obtained by peak area normalization using a FOCUS GC/FID (Thermo Fisher Scientific, Austin, TX, USA) operated under similar conditions of the GC-MS, except the carrier gas, which was nitrogen. The retention indices were calculated for all the volatile constituents using a homologous series of n-alkanes (C8–C32, Sigma-Aldrich, St. Louis, MO, USA) [56 (link)].

The quantitative analysis was performed by gas chromatography with flame ionization detector (Focus GC-FID, Thermo Scientific^™). Sample solutions in hexane (2 μL/1000 μL) were prepared and 1.0 μL were injected under the following conditions: silica capillary column DB-5 MS (30 m × 0.25 mm × 0.25 μm), carrier gas: nitrogen (flow rate: 1 2 mL/min) injection mode split (20:1) temperature of column 60 to 240°C (range of 3°C/min) and injector and detector temperatures 250°C. For qualitative analysis of the essential oils gas chromatography-mass spectrometry (DSQ II GC-MS, Thermo Scientific^™) was used. The analysis conditions for the injector and column were the same for GC-FID. The ionization source is electron impact (70 eV); transfer line temperature 200°C, helium carrier gas. The structural identification was made by comparison of their mass spectra and retention index to existing data in system libraries NIST 2011 and Adams 2007 [20 ,21 ].