## 3 Results and Discussion  
### Evaluation of SAFE, SDE and SPME Extracts by GC-O  
Three different extraction techniques were compared in order to evaluate their suitability for the determination of aroma components in broccoli. Two of them were exhaustive techniques: SAFE, SDE, and one non-exhaustive: SPME. Those were selected based on their popularity in research related to aroma active components in food products. SDE, which is the most traditional extraction method, widely applied for food products was compared with the modern SAFE technique, which does not require a high-temperature application. SPME, on the other hand, is very useful in volatile analysis. It is a solventless technique, especially convenient for analysis of small to medium-mass substances. In comparison with other non-exhaustive methods, it is characterized by its simplicity and also it does not require any specific equipment for its performance. Extracts were subsequently analyzed by different chromatographic methods. First, GC-O was employed to determine aroma active compounds, and GC-MS to identify them. Then GC\(\times\)GC-ToFMS was employed to identify components not visible on GC-MS chromatogram. Additionally, results from GC\(\times\)GC-ToFMS analysis were analyzed in order to profile volatiles from the three applied extraction techniques. Potent odorants (31) were identified in the SAFE extract, 27 in the SDE extract and 21 after applying SPME (Table 1). The SAFE technique gave the best results in the detection of aroma compounds, compared to the other two methods. SPME was important for identifying compounds eluting in the first few minutes, which coelute with a solvent peak on a GC-MS chromatogram and in the other extractions that are not visible due to solvent delay in the mass spectrometer. The first 3 min brought some characteristic odors in all extracts: \"sulfur\" resulting from the presence of methanethiol--a compound with extremely low odor threshold (0.2 \(\upmu\)g/L) [15]. Putrid, cabbage-like odor notes were perceived in SDE and SPME extracts caused by dimethyl sulfide (odour threshold 12 ppb [16]) and \"buttery\" odor present in all samples, assigned as 2,3-butanedione. Those first eluting compounds were particularly relevant for the flavor, because of their low odor threshold and characteristic aromas. Identification of early eluting compounds was possible only by applying the SPME solventless technique. All sulfides: dimethyl sulfide, dimethyl disulfide, dimethyl trisulfide, and dimethyl tetrasulfide gave more intense aroma in the case of the SDE extract, than in the two remaining solutions. This was probably due to the high-temperature applied during extraction, leading to the degradation of their precursors and/or thermal formation of di-, tri-, and tetrasulfides. Moreover, the concentration of sulfides was lower in SAFE compared to SDE and SPME, illustrating the influence of sample temperature on the formation of these compounds. In the SAFE extract, S-methylmethanethiosulphonate was presented in higher amounts (relatively high peak area) than in the two other extractions. High amounts of this component might be caused by low temperature extraction and fast enzymes inactivation, caused by adding the solvent right after tissue disruption. In SDE, the broccoli pulp was mixed with water in the first step, which created the possibility of enzymatic reactions and, moreover, the high temperature may lead to the thermal-induced sulfides formation. In SPME, the enzymes were probably active during the extraction procedure however, the odor intensity was lower than in SDE.