## 4 Discussion
Variability in melon fruit VOCs could be associated with differences in physiological maturity, but also fruit sampling, particularly the start of the ripening process in the placental tissue containing the seeds [28; 29] because fruit ripening enhances the presence of alcohols. Some of the alcohols, such as hexan-1-ol, are good substrates for alcohol acetyltransferases or AAT [30], and potential accumulation in some fruits can be a sign of a lack of activity of the enzyme compared with aromatic cultivars [31], or a fruit harvested in a stage when the activity of the enzyme is still low [30]. On the other hand, the presence of terpen or acids is sometimes associated with skin tissue [32], though terpen have been found at higher levels in PS than in other cultivars [33]. According to our experience and other authors, acids are rarely detected in pulp samples by the HS-SPME test [5; 33], but, by using stir-bar sorptive extraction (SBSE), it is possible to detect them as a precursor of aromas in whole PS melons, and also in the pulp of climacteric cultivars [20; 34] and in other juices [35]. Previous publications have applied PCA and PLS-DA to the classification of and comparison between VOCs of melon near-isogenic lines and the corresponding parental [5; 6], the PLS-DA providing a better classification with fewer variables involved. For a metabolomics approach, PCA combined with multiblock hierarchical PCA has been used [36]. Another possibility in the formation of outliers is the onset of analytical artifacts during the process, for example, due to septum bleeding, or the saline solution used (always stored at 2 \({}^{\circ}\)C and prepared before each season). No artifacts were observed after a blank SPME of the saturated saline solution or water used to prepare the saline solution alone, in agreement with general recommendations [8; 33]. From a physiological point of view, the higher the relative acetate ester concentration in flesh tissue, the more senescent the PS melon flesh [37]. This was the case with some samples when area counts were observed, but, in the percentage of total area counts, the outliers apparently disappeared. The esters, particularly isobutyl acetate or phenylmethyl acetate, and, to a lesser extent isopropyl acetate, are typical of climacteric cultivars [6]. Other compounds, such as methyl 2-methylpropanoate, or the sulfur-derived esters, which are very specific to some cultivars and more abundant in climacteric ones [38; 39], can also help detect outliers. A few esters can develop with certain abundance in some non-climacteric cultivars synthetized by ethylene-independent pathways, [40], particularly in certain seasons [6]. The high level of these esters dramatically reduced the proportion of the aldehydes typical of non-climacteric cultivars and accessions based on total area counts [8; 9; 37]. Another factor of importance is whether esters were found in the sampling procedure, particularly in large experiments involving climacteric aromatic and non-climacteric non-aromatic cultivars or lines for breeding purposes [39]. In the case of non-climacteric cultivars, placental tissue or local over-mature tissue should be avoided [3; 28; 36]. Overall, variables in the percentages of total area counts, such as total level of acids, terpenes, or alcohols, increased during PS melon fruit ripening, while aldehydes decreased, although non-acetate esters, alcohols and aldehydes also slightly increased close to senescence [34; 37]. The detection of the excess of some of these compounds within a sample of different fruit of one parental may help to provide more reliable control data for QTL mapping of this type of compound at harvest using near-isogenic lines [4; 5]. Several strategies are feasible to reduce fruit sampling outliers, although improving the signal to noise ratio in GC-MS analysis is probably the best by using techniques such as dynamic headspace purge and trap methodology for extraction [41].