5
2. Materials and Methods 2.1. Chemicals and reagents A mixture of normal alkanes (C7
-C30
USA) dissolved in hexane (HPLC grade, MilliporeSigma®
) (Supelco, , USA) was used for calculating
the linear retention index (LRI) for confirming peak identity. A divinylbenzene/carboxen/
polydimethylsiloxane (DVB/CAR/PDMS) df 50/30 µm 1 cm length fibre was used (offered by Millipore Sigma, Bellefonte, PA, USA).
Extra-virgin olive was purchased in a local supermarket (Gembloux, Belgium).
2.2. HS-SPME procedure for regular and reduced pressure conditions
The fibre was conditioned as suggested by the manufacturer before the first use. Blanks were run periodically to verify the absence of carryover.
1.5 g of oil sample was weighed into a 20 mL screw top vial (Restek, Bellefonte, USA). The sample was allowed to equilibrate with the headspace for 5 min at the temperature set for extraction (30°C and 43°C). Then, the SPME fibre was exposed to headspace and sampling was performed under agitation (250 rpm) at the selected sampling time (10, 20, 30, 40 min).
The fibre was desorbed at 250°C for 2 min (split 1:5) into the GC-MS inlet equipped with an SPME glass liner. All experiments were run in triplicate.
The difference between regular and Vac- HS-SPME is in the vial cap used. For the former, metallic caps with a central hole and a polytetrafluoroethylene (PTFE)/silicone septa (Restek, Bellefonte, USA) were used. For Vac- HS-SPME a custom-made closure design was implemented (provided by the Laboratory of Aquatic Chemistry, School of Environmental
mol-1
Engineering, Technical University of Crete [18]). The air inside the vial was evacuated for 1 min using a MD 4C diaphragm vacuum pump (7 mbar = 0.007 atm ultimate vacuum without gas ballast) (Vacuubrand GmbH & Co KZ, Wertheim, Germany). Then a 5 mL gastight syringe (SGE, Australia) was used to introduce 1.5 g of oil samples in the vial (Figure 1).
2.3. Central Composite Design
Central composite experimental design (CCD) was used to optimise extraction temperature and time, both for regular and Vac-HS-SPME. The extraction temperature was tested between 30°C and 55°C, and the exposition time from 10 to 30 minutes, based on conditions reported in previous works [19,20].
2.4. GC-MS analysis
An Agilent 7890B GC coupled to a 5977 MSD was used for all analyses. Carrier gas: helium at 1 mL/min flow rate. GC column: a 30 m x 0.25 mm i.d. x 0.5 µm df SLB-5ms capillary column [(silphenylene polymer, practically equivalent in polarity to poly(5%diphenyl/95% methylsiloxane)] kindly obtained from MilliporeSigma (Bellefonte, PA, USA). GC oven temperature program: 35°C (hold 2 min) to 250°C at 3°C/min and to 300°C at 25°C/min.
2.5. Data elaboration and statistical analysis
Twelve compounds were selected over the entire chromatogram (Table 1). All the chemical-physical properties of the 12 compounds selected (reported in and obtained from the ChemSpider website (
http://www.chemspider.com/).
All statistical analyses were performed using R v3.3.2 (R Foundation for Statistical Computing, Vienna, Austria) and Minitab 19 (
https://www.minitab.com/ en-us/) and
https://software.broadinstitute. org/morpheus/.
Table 1: List of the 12 selected compounds together with their Chemical Abstracts Service (CAS) number, boiling point (Bp), Henry’s constant (KH, atm m3 octanol-air partition constant (Koa, atm m3 ) and Molecular volume (VM, cm3
), octanol-water partition constant (Kow, atm m3 mol -1 # Compound Name
v1 v2 v3 v4 v5 v6 v7
v8 v9
v10 v11 v12
Acetic acid
Penten-3-one Hexanal
1-Hexanol
2(E)-Heptenal Benzaldehyde Octanal
Hex-(3Z)-enyl acet- ate
ß-Ocimene,(E) Nonanal
Methyl salicylate
α-Farnesene, (E,E) mol-1 ), Vapour pressure (Vp CAS 64-19-7
1629-58-9 66-25-1 111-27-3
18829-55-5 100-52-7 124-13-0
3681-71-8 13877-91-3
124.19-6 119-36-8 502-61-4
MW (g mol -1
60.1 84.1
) VM (cm3 ) 56.2 BP (°C) 117.1 103.3 104.3
100.2 124.9 127.0 102.2 125.0 158.2 112.1 135.0 166.0 106.1 101.1 178.7 128.2 157.9 163.4
142.2 157.7 175.2 136.2 175.5 175.2
142.2 174.4 190.8 152.1 125.8 222.0 204.3 251.5 279.6
Log KH (atm m3
mol-1 -5.54 -3.71 -3.45 -4.76 -3.81 -4.64 -3.20
)
log Koa (atm m3
mol-1 5.218 3.748 3.84
) ) , and linear retention index (IT) experimentally calculated and reported in the literature.
log Kow (atm m3
5.185 4.341 4.442 4.457
-3.35 -0.62
-3.10 -5.24 -0.19
4.195 3.398
4.793 4.947 5.067
mol-1 0.09 0.90 1.80 1.82 2.07 1.71 2.78
2.61 4.80
3.27 2.60 7.10
) Vp
(mmHg at 25°C)
m/z IT ex IT Lib
13.9 60 634 641 31.1 55 691 683 10.9 56 801 801 0.9 1.8
mol-1 ), , mmHg at 25°C), Molecular weight (MW , g
Figure 1: Schematic of the Vac-HSSPME
MS: in EI mode at 70 eV; source temperature: 230°C; quadrupole temperature: 150°C; full scan mode in the 35-500 m/z range.
56 872 867 83 960 956
1.0 106 965 960 2.1
1.4 1.6
0.5
84 1004 1006 67 1005 1008 93 1047 1046
57 1105 1107
0.1 120 1197 1192 0.0
93 1506 1504
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