Rapid propagation in vitro and accumulation of active substances of endangered Dendrobium cariniferum Rchb. f

Abstract

Dendrobium cariniferum is a valuable ornamental and medicinal plant rich with polysaccharides, alkaloid, and other bioactive compounds, which are potential raw materials for pharmacological utilization. In this study, an efficient protocol for the rapid propagation of D. cariniferum was developed. By using the tissue culture protocol, the effects of pH, hormone combinations, temperatures, light intensity, culture time protocorm proliferation, seedlings rooting, and accumulation of biomass with bioactive compounds were investigated. The experiments showed that the medium [1/2 MS + activated carbon1.0 g/L+ agar strip 7.5 g/L + sucrose 25 g/L] effectively promoted the germination of D. cariniferum seeds. The optimal culture conditions were found at pH 5.7, temperature 23 ± 2°C, and light intensity of 1000 Lx in the protocorm proliferation stage. Adding 1.5 g/L peptone in the medium effectively promoted the seedling rooting. The optimal culture conditions for accumulation of bioactive compounds (polysaccharides and alkaloids) of seedlings were found at temperature of 25 ± 2°C, light intensity of 1500-2000 Lx after the 60-day (d). Our study constructed a rapid propagation system in vitro for D. cariniferum, as well as the methods for efficient accumulation of active substances in seedling culture, which will serve as guidance for industrial production of D. cariniferum seedlings for both medicinal raw materials and ornamental plants. In addition, our study provided a new idea that we can directly use the high bioactive compound seedlings to extract medicinal components in industry conditions without transferring to the field.

Keywords: Dendrobium cariniferum; bioactive compounds; medium; propagation in vitro; tissue culture.

Figure 1.

(a) Flowers of Dendrobium cariniferum after pollination. Fruit capsule (b) and seeds (c) of D. cariniferum. (d) Seed germination in one-half MS medium. (e) Seed germination in MS medium.

Figure 2.

Effects of different factors on the proliferation of protocorms. (a) different medium pH; (b) different hormone combinations (A: 0.1 mg/L NAA+2.0 mg/L 6-BA; B: 0.5 mg/L NAA+2.0 mg/L 6-BA; C: 0.5 mg/L NAA; D: 0.1 mg/L NAA; E: 0.5 mg/L NAA+1.0 mg/L 6-BA; F: 0.1 mg/L NAA+1.0 mg/L 6-BA); (c) different temperatures; (d) different light intensities. The proliferation rate = the fresh weight after protocorm proliferation/fresh weight before protocorm inoculation. *Significance was determined by ANOVA (the same letter marks mean the difference was not significant: P ≥ 0.05, while different letters mean significant difference: P < 0.05).

Figure 3.

Effects of different concentrations of peptone on seedling rooting. *Significance was determined by ANOVA (the same letter marks mean the difference was not significant: P ≥ 0.05, while the different letters mean significant difference: P < 0.05).

Figure 4.

The influence of different culture time on the accumulation of biomass (a), polysaccharides (b), as well as alkaloids (c) in seedling culture. *Significance was determined by ANOVA (the same letter marks mean the difference was not significant: P ≥ 0.05, while the different letters mean significant difference: P < 0.05).

Figure 5.

The influence of different degrees of culture temperature on the accumulation of biomass (a), polysaccharides (b), and alkaloids (c) in seedling culture. *Significance was determined by ANOVA (the same letter marks mean the difference was not significant: P ≥ 0.05, while the different letters mean significant difference: P < 0.05).

Figure 6.

The influence of different levels of light intensity on the accumulation of biomass (a), polysaccharides (b), and alkaloids (c) in seedling culture. *Significance was determined by ANOVA (the same letter marks mean the difference was not significant: P ≥ 0.05, while the different letters mean significant difference: P < 0.05).