Profiling of Taxoid Compounds in Plant Cell Cultures of Different Species of Yew (Taxus spp.)

Abstract

Plant cell cultures of various yew species are a profitable source of taxoids (taxane diterpenoids) with antitumor activity. So far, despite intensive studies, the principles of the formation of different groups of taxoids in cultured in vitro plant cells have not been fully revealed. In this study, the qualitative composition of taxoids of different structural groups was assessed in callus and suspension cell cultures of three yew species (Taxus baccata, T. canadensis, and T. wallichiana) and two T. × media hybrids. For the first time, 14-hydroxylated taxoids were isolated from the biomass of the suspension culture of T. baccata cells, and their structures were identified by high-resolution mass spectrometry and NMR spectroscopy as 7β-hydroxy-taxuyunnanin C, sinenxane C, taxuyunnanine C, 2α,5α,9α,10β,14β-pentaacetoxy-4(20), 11-taxadiene, and yunnanxane. UPLC-ESI-MS screening of taxoids was performed in more than 20 callus and suspension cell lines originating from different explants and grown in over 20 formulations of nutrient media. Regardless of the species, cell line origin, and conditions, most of the investigated cell cultures retained the ability to form taxane diterpenoids. Nonpolar 14-hydroxylated taxoids (in the form of polyesters) were predominant under in vitro culture conditions in all cell lines. These results, together with the literature data, suggest that dedifferentiated cell cultures of various yew species retain the ability to synthesize taxoids, but predominantly of the 14-OH taxoid group compared to the 13-OH taxoids found in plants.

Keywords: 14-hydroxylated taxoids; Taxus spp.; plant cell culture; taxoids.

Figure 1

Left—Growth curves of the suspension cell culture of Taxus baccata lines Tb-msu/B5-PB-pvp (B5PBpvp-f in the legend) and Tb-msu/B5-NB (B5NB-f) during cultivation in flasks and line Tb-msu/B5-NB during cultivation in 20-L bioreactor (B5NB-b) under semi-continuous regime (the third cycle of cultivation). Right—photograph of cell suspension cultures in 250-mL flasks and microphotograph of cells of Tb-msu/B5-NB line.

Figure 2

UPLC–ESI-MS chromatogram (total ion current, positive ion mode) of an alcohol extract from the biomass of T. baccata callus cell culture (Tb-nbg /R-NB-ac line grown in a B5-PB-ac medium, 74 days of 42nd subcultivation). The description of peaks 1–6 is given in Table 3.

Figure 3

UPLC–ESI-MS chromatogram (total ion current, positive ion mode) of an alcohol extract from the biomass of T. baccata cell suspension culture (Tb-msu/B5-NB-Car line grown in a 20-L bioreactor). The description of peaks 1–8 is given in Table 4.

Figure 4

Structure and key ¹H–¹H COSY (bold lines) and ¹H–¹³C HMBC (arrows) correlations of taxoid I isolated from T. baccata cell suspension culture.