Zb 624 capillary column

Frozen samples of semen were thawed at room temperature, and subsequently, the vials were immersed in a water bath onto a magnetic stirrer hotplate at 60 °C overnight.
After this incubation, a Carboxen^®/Polydimethylsiloxane (CAR/PDMS) fiber (57318, Supelco) was exposed to headspace for 15 min, as reported in [32 (link)]. GC-MS analysis of extracted volatiles was performed using GC (6890N series Agilent Technologies) coupled to MS (5973 series Agilent Technologies) equipped with a ZB-624 capillary column (Phenomenex); the injector temperature was set at 250 °C to allow thermal desorption of VOCs. The carrier gas was high-purity helium with a flow rate of 1 mL min⁻¹. The MS analyses were carried out in full-scan mode with a scan range of 30–500 amu at 3.2 scans/s. Chromatograms were analyzed by Enhanced Data Analysis (MSD Chemstation E.02.02, Agilent Technologies, Santa Clara, CA, USA), and the identification of the volatile compounds was achieved by comparing mass spectra with those of the data system library (NIST14, p > 60%) and confirmed by the injection of external standards corresponding to most recurrent compounds. To quantify the identified VOCs, a semiquantitative method based on the internal standard (I.S.) 1,4-Dichlorobenzene-D4 (EPA-8260C) was followed. A frequency analysis was performed to identify the most frequently present VOCs.

Air samples were collected at the appropriate time intervals with the quartz cold trap [120 mm long, 2.9 mm o.d., 1.0 mm i.d. (inlet), 2.0 mm i.d. (outlet); Markes International Ltd., Llantrisant, UK] connected to each chamber and the air was collected for 5 min at 5 mL·min^–1. An automated thermal desorption system with Air Server autosampler (UNITY; Markes International Ltd.) was connected to the injection port of the gas chromatograph–mass spectrometer (TRACE DSQ; Thermo Electron Co., Waltham, MA) (Kim et al. 2008 , 2011 ). The desorbed sample was cryofocused at 5 °C for 5 min on the first few centimeters of the column, desorbed at 280 °C, and separated using a ZB-624 capillary column (30-m length × 0.25-mm i.d., 1.40-μm film thickness of 6 % cyanopropylphenyl, 94 % dimethylpolysiloxane; Phenomenex, Torrance, CA). The injection port temperature was 180 °C with a split ratio of 29:1. Helium was used as the carrier gas at a flow rate of 1.0 mL·min^–1. The column temperature was held at 45 °C for 1 min and increased at a rate of 15 °C·min^–1 to 100 °C and held for 1 min and then increased at a rate of 5 °C·min^–1 to 135 °C. However, the transfer line temperature was 180 °C; the ion source temperature was 280 °C. The mass range was m/z 45–120 and the mode of detection was EI mode (70 eV).