*4.7. Identification of the Chemical Constituents of the Essential Oil*

#### 4.7.1. Quantitative and Qualitative Analysis

The dentification of the chemical constituents of the essential oil was carried out using an Agilent gas chromatograph (GC) (6890N series, Agilent Technologies, Santa Clara, CA, USA). For the quantitative analysis gas chromatograph was equipped with a flame ionization detector (FID) and for qualitative analysis gas chromatograph was coupled to a mass spectrometer (quadrupole) detector (MS) (model Agilent 5973 inert series, Agilent Technologies, Santa Clara, CA, USA). The GC-FID and GC-MS analyses were performed according to the procedure described by Valarezo et al. [35]. The injection of the samples was carried out by an automatic injector (Agilent 7683 automatic liquid sampler, Agilent Technologies, Santa Clara, CA, USA) in split mode. Chromatographic runs were performed using a nonpolar and a polar column. The nonpolar was an Agilent J&W DB-5ms Ultra Inert GC column with stationary phase 5%-phenyl-methylpolyxilosane and the polar was an Agilent J&W HP-INNOWax GC column with stationary phase polyethylene glycol. Both columns with a length of 30 m, an outer diameter of 0.25 mm and a stationary phase thickness of 0.25 μm. Identification of the EO compounds was based on a comparison of relative retention indices (RIs) and mass spectra data with those of the published literature [38,39] according as described by Valarezo et al. [36].

#### 4.7.2. Enantioselective Analysis

The enantiomeric distribution was performed using an enantioselective column with stationary phase 2,3-diethyl-6-tert-butyldimethylsilyl-β-cyclodextrin. The chromatographic run was performed with a temperature ramp of 2 ◦C/min from 50 ◦C (maintained for 2 min) to 220 ◦C (maintained for 2 min) in an Agilent gas chromatograph (model 6890N series, Agilent Technologies, Santa Clara, CA, USA) coupled to a mass spectrometer (quadrupole) detector (model Agilent 5973 inert series, Agilent Technologies, Santa Clara, CA, USA). The injection of enantiomerically pure standards was used to determine the order of elution of the enantiomers.
