## 3 Discussion
Cyclization reaction occurs with the use of dammarenediol-II synthase (DS). Our result demonstrated the up-regulation of the gene coding dammarenediol-II synthase. What is interesting is the fact that there are reports that confirm a relationship between the overexpression of the \(DS\) gene and the enhanced production of ginseng in _P. ginseng_[40; 46; 47]. The formation of dammarenediol is an important element of ginseng biosynthesis, considering the fact that dammarenediol, via oxidation, hydroxylation, and glycosylation, can be converted to dammaran-type ginseng. The results of this paper showed enhanced _FPS_, \(SS\), and \(DS\) expression and improved ginseng accumulation. Our findings also demonstrated that carvacrol affected the production of protopanaxadiol and protopanaxatriol derivatives in a different way. It can be concluded that the accumulation of saponins belonging to the Rb group decreased, whereas the accumulation of saponins belonging to the Rg group increased. This effect could be associated with the different activation of protopanaxadiol synthase (PPDS, CYP716A47) and protopanaxatriol synthase (PPTS, CYP716A53v2). Both the enzymes are key for the formation of PPD and PPT aglycones. In addition, CYP716A53v2 is known to enable the conversion of PPD-type ginsenosides into PPT-type ginsenosides [49; 50]. [PERSON] et al. [51] indicated that the overexpression of _PPTS_ in hairy roots cultures of _P. ginseng_ contributed to the enhanced accumulation of _PPTS_ mRNA in transgenic roots compared to the control--nontransgenic roots. In contrast, the silencing of _PPTS_ mRNA in RNA reduced _PPTS_ transcription [51]. The biosynthesis of individual ginsenosides is possible via glycosylation, performed by UDP-dependent glycosyltransferases (UGTs) (Figure 8). UGTs form a glycosidic bond using sugar moieties [52]. Up to now, UGTs, involved in ginseng biosynthesis, have not been fully recognized [53; 54; 55; 56; 57; 58]. In _P. ginseng_, UDP-dependent glycosyltransferases add monosaccharides to triterpene aglycones, mainly at positions C-3 and/or C-20 in PPD-type ginsenosides or C-6 and/or C-20 in PPT-type ginsenosides [59]. [PERSON] et al. [58] showed that MJ treatment up-regulated 11 UGTs and down-regulated 3 UGTs in American ginseng. The results of our study revealed a significant increase in the Re metabolite content and the induction of Rg2 biosynthesis as well as the reduced accumulation of Rb-group saponin. Hence, we can conclude that carvacrol possibly influences the regulation of UGT genes and, in consequence, UDP-dependent glycosyltransferases activity in studied cultures of _P. quinquefolium_ hairy root. The ginseng biosynthesis pathway proposed by [PERSON] et al. [42] and [PERSON] et al. [52] shows that the Rg1 metabolite is a substrate for Re biosynthesis. In addition, Rg2 saponin can be generated via the conversion of Rh1 ginseng with the use of a suitable glycosyltransferase. In light of the knowledge concerning ginseng biosynthesis and the obtained result of individual metabolite contents, it seems possible to make a hypothesis about the potential stimulating influence of carvacrol on glycosyltransferases' activity, converting metabolite Rg1 to Re, regulating Rg2 generation and inhibiting its influence on glycosyltransferases activity, and leading to the obtainment of Rd, Rb1, Rb2, or Rb3 saponins. However, the potential influence of carvacrol on glycosyltransferases, which are involved in ginsenoside biosynthesis, will have to be verified in further studies after identifying these enzymes and preparing their detailed characteristics. The obtained results also indicated that the tested overexpression genes were achieved at different concentrations of carvacrol than the maximum level of the studied metabolites. This investigation proved that carvacrol up-regulated _FPS_, _SS_, and _DS_ genes. Probably, other factors play an important role in the regulation of ginsenoside biosynthesis.