4.4.4. Extraction and Identification of Volatile Compounds in Fermented and Dried Cocoa Beans by GC-MS

The fermented and dried cocoa beans were milled (A11B, Ika) and subjected to simultaneous distillation/extraction for 2 hours using 4 mL of pentane as a solvent. The volatile concentrate obtained was stored at a temperature of 5 °C [52,53,54].

Aliquots (2 μL) of volatile concentrate was injected into a gas chromatograph coupled to mass spectrometry (GC-MS), in a Shimadzu QP-2010 Plus system, equipped with a DB-5MS column (30 m × 0.25 mm × 0.25 μm). The helium gas was used as the carrier gas with a flow rate of 1.2 mL/min. The temperature of the injector and the interface was 250 °C, and the oven temperature was adjusted to 60–250 °C, using a ramp of 3 °C/min. Electronic impact mass spectrometer at 70 eV and the ion source temperature at 220 °C were used.

Quantitative analysis of the chemical constituents was performed by peak-area normalization using a flame ionization detector (FID—Shimadzu, QP 2010 system) under the same conditions as GC-MS, except that hydrogen was used as a mobile phase. Chemical identification was carried out by comparing the mass spectra and retention indices (RI) with those of standard substances in the system libraries and with data from the literature [55,56,57]. The RIs were obtained using a homologous series of n-alkanes (C8–C24, Sigma-Aldrich Co, St. Louis, MO, USA).