# Discrimination of Aroma Characteristics for Cubeb Berries by Sensomics Approach with Chemometrics
## 2 Results and Discussion
### Identification of Aroma Compounds  
To obtain a wider volatile profile and better discriminate the two cubeb berries, three extraction methods (HD, SDE, and SPME) were used (Table 1). In total, 90 volatile compounds were identified by HD, SDE, and SPME combined with GC-MS. Seventy-three volatile compounds belonged to different  
Figure 1: Effects of different factors—(**a**) desorption time; (**b**) incubation time; (**c**) extraction temperature; and (**d**) extraction time—on the peak areas of different volatile compounds of cubeb berries captured by DVB/CAR/PDMS. Different letters (a, b, c, and d) on the top of columns indicate significant differences (\(p<0.05\))  
chemical families: terpenes (8.82-80.65% for _C. cubbea_; 18.69-52.27% for _P. cubbea_), ketones (1.49-3.24% for _C. cubbea_; 17.97-20.99% for _P. cubbea_), alcohols (4.16-10.56 % for _C. cubbea_; 16.68-28.97% for _P. cubbea_), aldehydes (2.02-23.79% for _C. cubbea_; 1.71-6.43% for _P. cubbea_), esters (0-1.06% for _C. cubbea_; 0.98-2.57% for _P. cubbea_), and acids (2.39-48.55% for _C. cubbea_; 0-18.69% for _P. cubbea_). In order to highlight the differences between the two cubbe fruit with different extraction techniques in a simple and immediate way, Figure 2 showed the comparison of the relative percentages of the main chemical families present in cubeb berries. One of the most abundant chemical families identified in cubeb fruits was terpenes. To our knowledge, the literature dealing with the comparison of different extraction methods of volatile compounds in cubeb berries is scare. [PERSON] et al. [20] studied the chemical composition of the essential oil obtained only by HD of different parts (root, stem, leaf, flower, and fruit) of _L. cubbea_ and showed that citral and limonene were the main constituents. In the present work, for _L. cubbea_, d-limonene was one of the main terpenes for the SDE (10.57%) and SPME extracts (38.89%), but it was not detected in the HD extract. [PERSON] et al. [21] reported that some other terpenes, such as \(\alpha\)-pinene, \(\beta\)-pinene, and \(\beta\)-caryophyllene, were also detected as the main volatile compounds in the bio-oils produced from _L. cubbea_ seed by hydrothermal liquefaction. \(\alpha\)-Pinene (5.74%), \(\beta\)-pinene (6.22%), d-limonene (9.84%), and caryophyllene (5.57%) were also the main terpenes for the SPME extract of _P. cubbea_. \(\gamma\)-Terpinene (A13, 0.12-0.39%), \(\alpha\)-copaene (A19, 0.13-1.15%), caryophyllene (A23, 5.53-9.11%), and humulene (0.77-4.33%) were the common terpenes detected in all the three different extracts of two cubbe berries. Three terpen could only be detected in all the extracts of _P. cubbe_, including \(\alpha\)-cubebeene (A18, 0.4-0.52%), bicyclosesquiphellandrene (A27, 2.41-5.28%), and \(\alpha\)-farnesene (A30, 0.13-0.16%). Cubebene was also identified in the direct analysis in real time mass spectrometry (DART-MS) fingerprint of _P. cubbea_ studied by [PERSON] et al. [7]. The extracts obtained by SPME were rich in terpenes (80.65% for _L. cubbea_ and 52.27% for _P. cubbea_) in comparison with those from HD (8.82% for _L. cubbea_ and 18.69% for _P. cubbea_) and SDE (29.9% for _L. cubbea_ and 35.4% for _P. cubbea_). However, SDE with solvents tends to extract a higher amount of the volatile monoterenes than SPME in bay leaf [22], French beans [23], and wines [24]. These differences may be due to the matrix effect in releasing volatile compounds as each spice had a characteristic plant tissue structure. Meanwhile, strong oxidation and degradation of terpenes may occur for HD and SDE extracts because of higher temperature and longer time [25]. Figure 2: Chemical compositions of volatile compounds in different extracts (HD, SDE, and SPME) of _L. cubbea_ and _P. cubbea_. Different letters (a, b, c, d, and e) indicate significant differences (\(p<0.05\)).