## 4 Discussion
Recently, some biotechnological approaches have been successfully implemented, such as the overexpression of a potato \(\alpha\)-copaene synthase gene, resulting in enhanced levels (i.e., up to 15-fold higher than controls) or the \(\alpha\)-copaene synthesis from glucose by _Escherichia coli_ transformation with a gene that heterologously expressed _Piper nigrum_\(\alpha\)-copaene synthase [72; 73]. These novel bioengineered approaches are promising, but their feasibility should be verified in pest control. Moreover, the extraction of \(\alpha\)-copaene directly from plants, although present in many species, has the difficulty that its content is usually low [44; 52]. Trying to solve these issues, in this work, a chemical approach has been performed using fractional distillation, a method that is extensively used in industry (e.g., biorefineries, desalination, or separation of contaminants, among many others) and whose base is the separation of components by their boiling points (i.e., using heat) [74; 75; 76; 77; 78]. In fact, some variants like hydrodistillation (HD) and steam distillation (SD) have been one of the most widely used techniques in the extraction of EOs; and successfully applied to clove oil extraction [46; 79; 80; 81; 82]. However, some other innovative techniques have also been applied to extract EOs from clove oil, such as microwave-assisted hydrodistillation, microwave-assisted steam distillation (i.e., MDAs), hydrodistillation assisted by ohmic heating (HDA), or supercritical fluid extraction assisted by cold pressing (SFEA). These modern techniques saved extraction time compared to conventional methods (i.e., cold pressing, HD, or SD), although not differing in major components' composition (i.e., eugenol, \(\beta\)-cariophyllene, \(\alpha\)-humulene, and eugenyl acetate in clove oil), and yields varied with conditions (i.e., temperature, time, and power); however, no results on \(\alpha\)-copaene were provided in this research [46; 83; 84]. Related to this, it was possible to extract low percentages of copaene (although it is not indicated which, \(\alpha\) or \(\beta\) stereoisomer) with SD (3.41%) and superheated-SD (0.55%) techniques [85]. The yield and quality of extracted EOs from plants depends not only on the extraction method itself but many other factors, like plant organ used (e.g., aerial parts, inflorescence, or roots), agronomic practices (e.g., fertilization, irrigation, date of harvest), or cultivation location (e.g., microclimate, soil composition) [46]. As a handicap, the distillation method, although easy to operate and highly reproducible, requires a long extraction time and large volumes of solvent and energy [46]. At the industrial level, distillation necessitates a thoroughly rigorous process control and systems optimization to be effective and prevent contamination [86]. In this research, based on this background, the fractional distillation process has made it possible to concentrate clove oil into fractions with different contents in \(\alpha\)-copaene, despite the difficulties of using plants as a source of this compound. In fact, the major component of clove oil is eugenol (ca. 50-70%), followed by eugenyl acetate, \(\beta\)-caryophyllene, and \(\alpha\)-humulene (these three components together ca. 10-40%), with the \(\alpha\)-copaene content not exceeding 10% (i.e., within minority compo nents) [46,87]. Clove oil has been tested in the field as a source of attractant pheromones to _C. capitata_. [PERSON] et al. [88] used white Jackson traps with a piece of cotton saturated with diluted clove oil concentrations (12.5, 25, 50, 75, and 100%) in paraffin oil. Clove oil at a concentration of 50% produced the highest attraction for males compared with the other different concentrations of clove oil or trimedlure, but the components responsible for the attractiveness of the clove oil were not specified. Also, clove oil was demonstrated to be effective in the attraction of other related tephritids [89].