Impact of external calcium and calcium sensors on ginsenoside Rb₁ biosynthesis by Panax notoginseng cells

Abstract

The effects of external calcium concentrations on biosynthesis of ginsenoside Rb1 and several calcium signal sensors were quantitatively investigated in suspension cultures of Panax notoginseng cells. It was observed that the synthesis of intracellular ginsenoside Rb1 in 3-day incubation was dependent on the medium Ca2+ concentration (0-13 mM). At an optimal Ca2+ concentration of 8 mM, a maximal ginsenoside Rb1 content of 1.88 +/- 0.03 mg g(-1) dry weight was reached, which was about 60% and 25% higher than that at Ca2+ concentrations of 0 and 3 mM, respectively. Ca2+ feeding experiments confirmed the Ca2+ concentration-dependent Rb1 biosynthesis. In order to understand the mechanism of the signal transduction from external Ca2+ to ginsenoside biosynthesis, the intracellular content of calcium and calmodulin (CaM), activities of calcium/calmodulin-dependent NAD kinase (CCDNK) and calcium-dependent protein kinase (CDPK), and activity of a new biosynthetic enzyme of ginsenoside Rb1, i.e., UDPG:ginsenoside Rd glucosyltransferase (UGRdGT), in the cultured cells were all analyzed. The intracellular calcium content and CCDNK activity were increased with an increase of external Ca2+ concentration within 0-13 mM. In contrast, the CaM content and activities of CDPK and UGRdGT reached their highest levels at 8 mM of initial Ca2+ concentration, which was also optimal to the ginsenoside Rb1 synthesis. A similar Ca2+ concentration-dependency of the intracellular contents of calcium and CaM and activities of CCDNK, CDPK, and UGRdGT was confirmed in Ca2+ feeding experiments. Finally, a possible model on the effect of external calcium on ginsenoside Rb1 biosynthesis via the signal transduction pathway of CaM, CDPK, and UGRdGT is proposed. Regulation of external Ca2+ concentration is considered a useful strategy for manipulating ginsenoside Rb1 biosynthesis by P. notoginseng cells.

Production of ginseng saponins: elicitation strategy and signal transductions

“…The external Ca 2+ similarly changed specific UGRdGT activity and synthesis of ginsenoside Rb1 together with CaM content and CDPK activity in P. notoginseng cells which suggests that CaM and CDPK may regulate biosynthesis of ginsenoside Rb1 in response to external calcium signal by triggering UGRdGT activity (Yue and Zhong 2004).…”

“…In case of copper treatment, higher amounts of Rg1 group are accumulated after treatment with CuSO 4 (Ali et al 2006a). Regulation of external calcium was found as an efficient strategy for biosynthesis of ginsenoside Rb1 (Yue and Zhong 2004). Taken together, alteration of ginsenoside heterogeneity can be manipulated in ginseng tissue cultures by utilization of specific elicitors for each purpose (Table 1).…”

Pharmacological activity of sanchi ginseng (Panax notoginseng)

“…Addition of methyl jasmonate to P. notoginseng cell culture increased the production of ginsenosides (Wang et al 2005). Ginsenoside biosynthesis was also affected by the calcium concentration of the medium (Yue & Zhong 2005). Zhang & Zhong (2004) described scaling up of a centrifugal impellar bioreactor for hyperproduction of saponin and polysaccharide by cultured P. notoginseng cells.…”

Protopanaxadiol 6‐hydroxylase and its role in regulating the ginsenoside heterogeneity in Panax notoginseng cells

“…Suspension cultures of P. notoginseng cells were maintained as described previously (Yue and Zhong, 2005a). Reduced form of nicotinamide-adenine dinucleotide phosphate (NADPH), phenobarbital sodium salt, (AE) miconazole nitrate salt, ancymidol, cytochrome C, glucose oxidase, b-Dglucose and catalase were obtained from Sigma (St. Louis, MO).…”