Elite 5ms column

Volatile compounds (VOC) were extracted from Pecorino cheese samples through solid-phase microextraction (SPME), and the analysis was performed with a gas chromatograph (Clarus 580; Perkin Elmer, Waltham, MA, USA) coupled with a mass spectrometer (SQ8S; Perkin Elmer, Waltham, MA, USA). The gas chromatograph was equipped with an Elite-5MS column (length × internal diameter: 30 × 0.25 mm; film thickness: 0.25 μm; Perkin Elmer, Waltham, MA, USA). The samples preparation and the settings relating to the thermal program used for the analysis were performed as previously reported by Ianni et al. [21 (link)]. Five grams of cheese previously grated were mixed with 10 mL of saturated NaCl solution (360 g/L). After the addition of 10 μL of internal standard solution (4-methyl-2-heptanone; 10 mg/kg in ethanol), the vials were sealed and stirred at 50 °C; VOCs were extracted from the headspace with a divinylbenzene-carboxen-polydimethylsiloxane SPME fiber (length: 1 cm; film thickness: 50/30 μm; Supelco, Bellefonte, PA, USA) with an exposition time of 60 min. VOCs were identified by comparison with mass spectra of a library database (NIST Mass Spectral library, Search Program version 2.0, National Institute of Standards and Technology, U.S. Department of Commerce, Gaithersburg, MD, USA) and by comparing the eluting order with Kovats indices.

Before the commencement of mass spectrometry, the crude sample was subjected to preliminary phytochemical analysis. A Perkin Elmer model Clarus 600 T (Perkin Elmer Waltham, Massachusetts, USA) combined with a single quadrupole mass spectrometer was used for gas chromatography–mass spectrometry (GC–MS) analysis. The chromatographic column was an Elite 5MS column by Perkin Elmer Waltham, Massachusetts, U.S.A (30 m × 0.25 mm × 0.25 µm film thickness), with high-purity helium as the gas carrier, at a flow rate of 1 mL/min. The injector temperature was 280 °C and it was equipped with a splitless injector at 20:1. The temperature was set initially to 40 °C (held for 2 min), then was increased to 150 °C at 10 °C per min (held for 2 min), then increased further to 300 °C at 10 °C per min for 2 min. The MS ion source temperature was 220 °C and inlet line temperature was set to 240 °C. The scan range was set at 40 to 600 mass ranges at 70 eV electron energy and the solvent delay of 4/min. Finally, unknown compounds were identified by comparing the spectra with that of the NIST 2005 (National Institute of Standard and Technology library) and the Wiley 2006 library. The total time required for analyzing a single sample was 32 min.

Volatile compounds (VOCs) present in cheese at 60 days of ripening were extracted by solid-phase microextraction (SPME) and the separation was carried out by gas chromatography (GC Clarus 580; Perkin Elmer, Waltham, MA, USA) coupled with a mass spectrometer (SQ8S, Perkin Elmer). The GC was equipped with an Elite-5MS column (length × internal diameter: 30 × 0.25 mm; film thickness: 0.25 µm; Perkin Elmer). For all samples, 4 g of earlier shredded cheese was mixed with 10 mL of saturated NaCl solution (360 g/L) containing the internal standard (4-methyl-2-heptanone). VOCs were extracted using a divinyl-benzene-carboxen-polydimethylsiloxane SPME fibre (length: 1 cm; film thickness: 50/30 215 μm; Supelco) at 60 °C with a 40 min exposition time. Helium was used as a carrier gas at a flow rate of 1 mL/min, and VOCs identification was performed by comparison with the mass spectra of a library database (NIST Mass Spectral library, Search Program version 2.0, National Institute of Standards and Technology, US Department of Commerce, Gaithersburg, MD, USA) and by comparing the eluting order with Kovats indices. Data were expressed as the mean relative percentage of the total VOCs identified.