## 4 Conclusions  
In this study, a HS-SPME/GC-MS method by Design of Experiments (DoE) and Response Surface Methodology (RSM) approaches was successfully optimized. The optimized parameters were sample amount, sample stirring, extraction temperature, conditioning time, extraction time, desorption temperature, and desorption time. The optimized HS-SPME/GC-MS method allowed the identification and semi-quantification of a high number of compounds. Of the 92 VOCs identified, 4 are found for the first time in EVOO while others, mainly terpenoid compounds, are rarely identified in EVOO, with the use of SPME/GC-MS. Statistical analysis allowed for identifying markers with significance in the geographical classification of Cretan EVOO samples. The method is deemed useful for the analysis of EVOO, providing easy, automated, efficient, and economic sample preparation and analysis along with information-rich GC-MS data.  
**Supplementary Materials:** The following supporting information can be downloaded at [[https://www.mdpi.com/article/10.3390/metabo12020114/s1](https://www.mdpi.com/article/10.3390/metabo12020114/s1)]([https://www.mdpi.com/article/10.3390/metabo12020114/s1](https://www.mdpi.com/article/10.3390/metabo12020114/s1)), Table S1: Quadratic model parameters including term estimates, standard error (Std.Error), t values and \(p\)-values. Term significance is provided in the column Signif, in a scale given at the footnote of the table. Extraction temperature (x1), conditioning time (x2), extraction time (x3), desorption time (x4) and desorption temperature (x5). Example: x1 is a first order term, x1:x2 is a two-way interaction term and x1^2 is a quadratic term, Table S2: Quality characteristics of EVOO samples, Table S3: Complete Circumscribed Central Composite design indicating the run order along with the levels of each factor. Units for extraction temperature (Ex.temp) are \({}^{\circ}\)C, conditioning time (Co.time) mins, extraction time (Ex.time) mins, desorption temperature (Des.temp) \({}^{\circ}\)C and desorption time (Des.time) mins, Figure S1: Contour plot for factors desorption temperature (Des.temp) and desorption time (Des.time). Green colour indicates lower response values and pink colour higher response values. Contours include corresponding response levels. The contour plot is a slice at the selected points for extraction temperature (Ex.temp) and extraction time (Ex.time), and the centre point for conditioning time (Co.time), Figure S2: PCA score plot showing the examined groups together with QC samples. QC samples are clustering together, showing system's stability, Figure S3: Geographical origin of Cretan EVOOs analysed in our study.  
**Author Contributions:** A.L., I.S. and C.V., methodology, formal analysis, data curation, and writing--original draft preparation; V.T.P., A.S. and K.G.Z., Formal analysis, Writing--original draft; G.T., data curation, writing--original draft preparation, writing--review and editing, visualization, supervision and project administration. All authors have read and agreed to the published version of the manuscript.  
**Funding:** This research was supported by the project FoodOmicsGR \"Comprehensive Characterization of Foods\" (MIS 5029057), which is implemented under the action \"Reinforcement of the Research and Innovation Infrastructure\", funded by the Operational Programme \"Competitiveness, Entrepreneurship and Innovation\" (NSRF 2014-2020) and co-financed by Greece and the European Union (European Regional Development Fund). All authors have read and agreed to the published version of the manuscript.  
**Institutional Review Board Statement:** Not applicable.  
**Informed Consent Statement:** Not applicable.  
**Data Availability Statement:** Not applicable.  
**Acknowledgments:** We are grateful to the Sensory Evaluation Laboratory of Crete ACR in Rethymnon Crete, Greece for providing the organoleptic data of the VOO samples.  
**Conflicts of Interest:** The authors declare no conflict of interest.