The Head Space Solid Phase Microextraction Method (HS-SPME), coupled with Gas Chromatography and Mass Spectrometry (GC-MS) was optimized to quantify Volatile Organic Compounds (VOC’s) in a fresh Oaxaca cheese during its shelf life of 22 days. Optimal extraction conditions were ascertained by means of a multiple linear regression analysis and the generation of a response surface. Conditions to reach the equilibrium of VOC’s at the headspace were established to be 30 minutes of ultrasonic agitation at 50°C. The conditions for extraction were exposure of the fiber at room temperature for 30 minutes. Optimal extraction of volatile compounds released by the Oaxaca cheese allowed identification of 21 volatile compounds: 8 acids, 1 alcohol, 4 aldehydes, 4 ketones, 1 ester, and 3 lactones. Quantification of VOC’s was carried out using the technique of standard addition. The limits of quantification ranged from 0.0033 to 0.0311 ng/μg. Ten compounds were identified with the heaviest concentration and are listed as follows, from heaviest to lightest: propionic acid, acetoin, acetic acid, diacetyl, capric acid, caprylic acid, δ-decalactone, δ-dodecalactone, γ-dodecalactone, butyraldehyde. During storage of the cheese, the most significant changes in the concentration of VOC’s took place between day 1 and day 15. Based on the results obtained, it can be concluded that the optimization of the SPME makes it possible to isolate and quantify VOC’s of fresh cheeses in an efficient and reproducible manner.