## 3 Discussion  
### Comparison of Method Sensitivity, Selectivity, Reproducibility, and Efficiency  
In matrices such as milk where the flavour profile is mild and volatile concentrations are low, recovery of adequate concentrations of volatiles for detection and quantification can be a challenge [11]. The present work optimized and compared four volatile extraction techniques based on their respective benefits and biases. The primary limitations of these techniques were a polarity bias for the PDMS stir bar techniques (HSSE and SBSE) and a volatility bias for the headspace techniques (HSSE and SPME). One of the major drawbacks of headspace techniques is that they tend to lack sensitivity for larger, less volatile compounds compared with liquid extraction techniques [30]. The volatility bias was  
Figure 3: Apparent concentration of dimethyl sulphone extracted by Solvent Assisted Flavour Evaporation (SAFE), Solid Phase Microextraction (SPME), Headspace Sportive Extraction (HSSE) and Stir Bar Sportive Extraction (SBSE). Error bars represent the standard deviations. evident in the results of this experiment with HSSE and SPME extracting lower overall concentrations of volatiles from the matrix, largely due to poor of extraction of lactones and carboxylic acids. For example, excluding lactones, SBSE extracted 297 \(\upmu\)g/kg and HSSE extracted 271 \(\upmu\)g/kg of volatiles in sheep milk. However, with lactones, SBSE extracted more than twice the apparent total concentration of volatiles as the headspace technique, with 657 \(\upmu\)g/kg and 271 \(\upmu\)g/kg extracted respectively. HSSE and SPME were only able to extract trace levels of \(\updelta\)-octalactone and \(\updelta\)-decalactone. On the other hand, the SBSE technique detected eight lactones and SAFE extracted four lactones. The non-polar bias of the PDMS phase appeared to be beneficial for the extraction of lactones and may explain, at least in part, why SBSE alone was able to extract \(\updelta\)-tetradcalactone and \(\updelta\)-hexadeclactone, due to immersion in the liquid phase for extraction. As expected, due to its lower extraction efficiency, SPME detected lower total apparent concentrations of volatiles from the sheep milk than the other three methods. It has been noted in the literature that both highly polar and higher molecular weight volatiles can be well extracted by the SAFE approach depending on the solvent employed, and this study confirmed that finding [31]. SAFE was the only extraction technique that was able to extract high apparent concentrations of both the small polar dimethyl sulphone and the larger less volatile \(\updelta\)-dodeclactone. The non-polar PDMS stir bar methods, HSSE and SBSE, extracted only low concentrations of sulphur compounds while SAFE and SPME excelled. However, the PDMS phase is not exclusively biased against polar compounds. SBSE detected substantial concentrations of dimethyl sulphone while HSSE was only able to detect trace concentrations. However, the concentration detected in both instances was low compared to SAFE and SPME. Possibly due to the broad selectivity of the triple phase DVB/CAR/PDMS fibre, SPME was also the only method to detect pentanal and dimethyl sulphide. There was likely a polarity bias against pentanal and dimethyl sulphide by the PDMS phase of the adsorptive stir bars and the peaks were likely lost under the solvent peak for the SAFE method. It can be difficult to discuss reproducibility without also addressing automation and efficiency. [PERSON] et al. found that the SAFE method excelled in terms of efficiency compared to other distillation techniques [31]. Since then, advances in software and hardware have enabled techniques like SBSE, HSSE, and SPME to become simple, highly automated, and efficient methods for extracting volatiles from food matrices. SAFE required significantly more time (approximately 12 h per sample) and effort than the other methods, mostly due to the extensive glassware preparation, manual sample manipulations, and sample concentration steps.