Abstract
Triterpenic saponins represented in Calendula officinalis L. by oleanolic acid (OA) glycosides are pentacyclic triterpene compounds with a wide range of biological and medicinal properties. This report demonstrates nitrogen source impact on growth, saponin accumulation, and secretion in hairy root and suspension cultures of marigold. Hairy roots preferred nitrate as a mineral source of nitrogen, but its impact on growth, OA glycosides accumulation, and secretion were line-dependent. The best productivity of OA glycosides was found in CC16 line (74.86 mg flask−1) in ½ MS medium modified by 2.5× KNO3 and ammonium elimination with 2.5 g l−1 peptone. Organic nitrogen source at 27.5-g l−1 impairs the growth rate of hairy roots. Its effect on saponin accumulation and secretion to the surrounding medium depended on line and media composition. Nitrate:ammonium ratio of 4:2 for CC16 resulted in 5.7-fold increment of saponin secretion comparing to the standard medium. Embryo roots, apical bud, and hypocotyls explants were crucial for induction of suspension culture synthesizing saponins; however, effect of mineral form of nitrogen in cultivating medium had to be considered. The highest OA glycosides level (171.97 μg g−1 of dry weight) was recorded in the root derived culture with nitrate as a sole mineral form of nitrogen. Peptone from lactalbumin decidedly inhibited the saponin formation; however, it was essential for culture initiation, proliferation, and organ differentiation.
Highlights
Oleanolic acid (OA) naturally occurs in plants as the form of glycosidic derivatives is a pentacyclic triterpenoid which has been identified in more than 1620 of edible and medicinal species (Fai and Tao 2009; Fukushima et al 2011)
When organic form predominated or proportion of mineral nitrogen as for 1⁄2 MS was used, aggregates were smaller (< 1 mm) than in the media with standard level of edamine and with unbalanced mineral nitrogen ratio. This effect was not observed for Suspension cultures (SC) derived from cotyledon: small aggregate size was obtained in the medium with standard edamine concentration (250 mg L−1) and nitrate as a sole mineral source of nitrogen (Fig. 2c)
Results presented in this paper showed significantly that nitrate and organic source of nitrogen were essential for suspension culture initiation; nitrate strongly accelerates cell proliferation and generally saponins accumulation
Summary
Oleanolic acid (OA) naturally occurs in plants as the form of glycosidic derivatives is a pentacyclic triterpenoid which has been identified in more than 1620 of edible and medicinal species (Fai and Tao 2009; Fukushima et al 2011). Other specific biological activity of OA molecules is associated with sugar chain linked to the aglycone with glycosidic bond These complex structures, called saponins, have a better solubility in water and exhibit hemolytic, allelopathic, fungistatic (Szakiel et al 2005), antibacterial, and antiparasitic properties (Szakiel et al 2008; Doligalska et al 2011).
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