Db ffap column

Methional was enriched by SPE and quantified by GC–PFPD. DB-FFAP column (30 m × 0.32 mm i.d., 1 μm film thickness, J&W Scientific Inc., Folsom, CA, USA) was used. The flow of the helium was 2 mL/min. Temperature of oven was programmed at 35 °C for 3 min and was increased to 150 °C at 10 °C/min, and held for 5 min. Then, it was increased at 20 °C/min to a final temperature of 220 °C and held for 3 min. Temperature of the GC injection and the detector was 250 °C. Sulfur gate time was 6–24.9 ms, and pulse frequency was approximately three pulses/s. Isopropyl disulfide (48.62 mg/L, 8 μL) was used as an internal standard (IS6) [12 (link)].

Quantitation of carbonyl compounds after derivatization was performed by GC−MS on a DB-FFAP column (60 m × 0.25 mm i.d., 0.25 μm film thickness, J&W Scientific) modified from literature. A sample of 8 mL saturated with NaCl was placed in a 20-mL standard headspace vial. A total of 10 μL of p-Fluorobenzaldehyde (internal standard, 100 mg/L) and 300 μL of PFBHA (20 g/L in water) were added. A 50/30-μm DVB/CAR/PDMS fiber (Supelco, Inc.,Bellefonte, PA) was used. The sample was equilibrated at 65 °C for 10 min, then extracted for 45 min under stirring at 250 rpm and desorption at 250 °C for 300 s. Sample was injected in a splitless mode. Carrier gas for the column was helium at a constant flow rate of 1 mL/min. Temperature for the injector was 280 °C. The oven temperature was 50 °C for 2 min and was increased to 100 °C at 6 °C/min for 0.1 min, then was increased to 160 °C at a rate of 2 °C/min for 0.1 min, and finally at 5 °C/min to 230 °C for 10 min. The electron impact energy was 70 eV with SIM. The ion monitored for p-Fluorobenzaldehyde after derivatization was m/z 319. The standard curve, LOD and recovery of carbonyl compounds, were measured by the method mentioned above.

Methyl esters of fatty acids (FAME) of D. neapolitana (~50 mg) were prepared following the method described by Aued-Pimentel et al. [38 (link)] (through transmethylation of FA using a mixture of methanolic solution KOH (2 M) and saturated NaCl). The resulting FAME were analyzed in a QP2010 Ultra Shimadzu gas chromatography–mass spectrometry, equipped with an auto-sampler a DB-FFAP column with 30 m length, 0.32 mm internal diameter, and 0.25 µm film thickness (J&W Scientific, Folsom, CA, USA). The column was initially programmed to 80 °C, increasing 25 °C min⁻¹ until 160 °C, 2 °C min⁻¹ from 160 to 220 °C, and 30 °C min⁻¹ from 220 to 250 °C, using helium as the carrier gas, at a flow of 1.8 mL min⁻¹. FAME identification was accomplished through comparison of retention times with those mixed FAME standards (C4–C24, Supelco 37 Component Fame Mix) and by comparison of the mass spectrum of each relative to standard spectra from the library “AOCS Lipid Library” (http://lipidlibrary.aocs.org/ (accessed on 5 January 2024)).

Procedures for SCFA analysis were described by a previous study (Williams et al., 2011 (link)). The gas chromatography GC-17A (Shimadzu, Kyoto, Japan) with a flame ionization detector (FID) equipped with a DB-FFAP column (30 m × 0.53 mm) (J&W Scientific) was applied to detect SCFA.

β-damascenone was extracted using CH₂Cl₂ as the organic solvent, and L-menthol (100 mg/L) was used as the internal standard. Solid samples (10 g) were added to 25 mL sterile saline (0.85% NaCl and 1% CaCl₂), ultrasonically treated for 30 min in an ice-water mixture, and then centrifuged at 8,000 × g for 5 min (4°C) to obtain supernatants. For liquid samples, yeast fermentation broths were centrifuged at 8,000 × g for 5 min (4°C). For the quantitation of β-damascenone, CH₂Cl₂ (2 mL), internal standard (20 μL), and NaCl (7 g) were added to the supernatants (20 mL). The mixture was stirred for 10 min at ambient temperature and sonicated in an ice bath for 10 min. The above obtained sample was centrifuged and refrigerated overnight to separate the organic phase. Next, the lower organic phase was collected and dried over anhydrous sodium sulfate. The extracted organic phase was analyzed using a gas chromatography-mass spectrometry (GC-MS) system (Agilent Technologies Inc., Santa Clara, CA, USA) equipped with a DB-FFAP column (30.0 m × 0.25 mm × 0.25 μm, J&W Scientific, Santa Clara, CA, USA). The standard solution was made from aqueous alcohol solution and applied to construct a standard curve. All standard solutions were analyzed in the same manner as described for samples. The conditions for GC-MS were based on a previously reported method (Sun X. Z. et al., 2022 (link)).

Serum samples were thawed and well mixed prior to analysis. Each sample (50 uL) was vortex mixed with 100 μL of phosphoric acid solution (36% v/v) for three minutes, followed by adding 150 μL of methyl tert-butyl ether analytical standard (MTBE standard solution). The mixture was vortexed for 3 min again and ultrasonicated for 5 min. After that, the mixture was centrifuged, and the supernatant was collected for GC-MS analysis. An Agilent 7890B GC system coupled to a 7000D GC/MS with a DB-FFAP column (30 m length × 0.25 mm inner diameter × 0.25 μm film thickness; J&W Scientific, USA) was used to detect the SCFAs and neurotransmitters in the serum samples.

For identification of the VOCs present in the distillates, GC-MS/O was performed using a Trace GC Ultra and a Trace dual stage quadrupole (DSQ) mass spectrometer (both Thermo Fisher Scientific GmbH) equipped with a DB-FFAP column (30 m × 0.32 mm, 0.25 μm film thickness, J&W Scientific, Waldbronn, Germany). The distillates (2 µL) were injected automatically by a multipurpose autosampler MPS 2 (Gerstel GmbH & Co. KG, Mülheim an der Ruhr, Germany) using the cold on-column technique. The initial temperature of 40 °C was held for 2 min, raised at 6.0 °C/min to 235 °C, and held for 5 min. The flow rate of the helium carrier gas was 2.2 mL/min. At the end of the capillary column, the effluent was split between an ODP and the MS using deactivated fused silica capillaries (50 cm × 0.2 mm). Mass spectra were recorded in positive electron ionization (EI) mode at 70 eV (m/z range 35–250).