A novel headspace gas chromatography-mass spectrometry (HS-GC/MS) method for identifying and quantifying 3,3-dimethyl-1-butanol (DMB), a bioactive compound, in extra virgin olive oil was developed. In this study, solvents with similar properties to DΜB but potentially different molecular masses, namely isopropanol, n-butanol, n-pentanol, and 2-pentanol, were tested and 2-pentanol was selected as the internal standard due to both chromatographic separation and distinctive mass spectrum pattern. The method for DMB determination and quantification was validated according to Eurochem CITAC Guide. Sensitivity, specificity, linearity, accuracy, and precision parameters were evaluated. The validation process included an assessment of the method’s robustness and repeatability, ensuring that it produces reliable results for future analyses of olive oil. A linear regression equation was developed for DMB concentrations ranging from 100 to 5,000 µg/L, expressed as y = x.11.462.10⁻⁶, where y represents the peak area ratio and x is the DMB concentration in µg/L. This equation has a high correlation coefficient of 0.9989 (R²), and the uncertainty budget was estimated to be 8.22% at a confidence level of k = 2. In addition, the lower concentration range of 0 to 100 µg/L was characterized by the equation y = x.17.860.10⁻⁶ which has a correlation coefficient of 0.9983 (R²). The uncertainty of DMB measurements in extra virgin olive oil was estimated to be 7.0% at a confidence level of k = 2. This new method was applied to determine the presence of DMB in various olive oil samples from Greece and Türkiye. The DMB values for olive oil samples were: N1(Plus health blue multi varietal) 9.7 μg/L, Ν2 (Plus health DMB multi varietal) 11.4 μg/L, N3 (Fyllikon first harvest organic) 8.3 μg/L, N4 (Plus health green multi varietal) 7.8 μg/L, N5 (Agourelaio early harvest organic) 6.8 μg/L, N6 (unknown) 4.8 μg/L, N7 (Armonia monovarietal organic) 4.8 μg/L and N8 (Edremit type olive oil from Kırkağaç, Manisa) 8.8 μg/L and N9 (Market product in Türkiye) 1.5 μg/L.

Olives and olive oil have established, through centuries, strong bonds with the prosperity and well being of the Mediterranean people. Numerous studies have featured them as main antioxidant sources, based on their phenolic profile, which includes phenolic acids and alcohols, polyphenols, lignans, secoiridoids, oleacein and oleocanthal. Several factors, such as cultivar, fruit maturation, processing methods, storage conditions etc. affect the presence of these significant constituents in the products. Re-use of olive mill waste water has been studied to enrich the phenolic profile of the final product. Olive oil is considered a functional food with extensive use in the food industry. The biological effects of olives and virgin olive oil components include protection against cardiovascular diseases, anti-inflammation action, neuro and endothelial protection etc. Research studies, in vitro and in vivo, in humans and in animals have been performed to better understand the metabolism and bioactivity of olives and olive oil phenolics. Olives and olive oil antioxidants are not only absorbed by the body, but can also attach to the lining of the digestive tract, contributing to the health impact afforded by these products. Laboratory methods and techniques used so far for the qualitative and quantitative identification of these compounds are cited.