Enhancement of ginsenoside biosynthesis in cell cultures of Panax ginseng by N,N′-dicyclohexylcarbodiimide elicitation

Abstract

In this work, the effect of N,N′-dicyclohexylcarbodiimide (DCCD) on ginsenoside biosynthesis in suspension cultures of Panax ginseng cells was investigated. The optimal concentration and timing of DCCD addition were found to be 10μM and on day 4 of cultivation. Under this condition, the maximal content of total ginsenosides increased to 3.0-fold that of untreated control, and the contents of Rg-group (Rg1 and Re) ginsenosides and Rb1 were 2.5- and 8.9-fold higher, respectively, which coincided with elevated activities of protopanaxatriol biosynthetic enzyme protopanaxadiol 6-hydroxylase and UDPG-ginsenoside Rd glucosyltransferase that converts Rd to Rb1. In addition, DCCD treatment induced the activity of defense response enzyme, phenylalanine ammonia lyase. To gain a better understanding of the molecular processes underlying the elicitation, we examined nitric oxide (NO) content and expression levels of the triterpene biosynthetic genes encoding squalene synthase (sqs), squalene epoxidase (se), and dammarenediol-II synthase (ds). It was found that DCCD up-regulated NO generation and transcription levels of sqs, se and ds. Interestingly, these effects of DCCD were compromised by an NO biosynthetic inhibitor, while an NO donor alone recapitulated the elicitation effect of DCCD on ginsenoside biosynthesis. These results suggest that DCCD may induce the ginsenoside biosynthesis via NO signaling in the P. ginseng cells. The information obtained might also be helpful to hyperproduction of valuable secondary metabolites in other plant cell cultures.