Culturas de raízes e análise fitoquímica de Kalanchoe pinnata (Lam.) Pers.

Abstract

Kalanchoe pinnata, popularly known as saião roxo , is widely spread in tropical regions of the world due to its medicinal and ornamental importance. It possesses an expressive capacity of vegetative propagation, being considered an Invasive Species in several places around the world, what highlights the need of eradicating it from natural environments. The bioactive compound already identified for the species, mainly flavonoids, presents major importance on traditional medicine and several studies corroborate its medicinal value, demanding more studies about its utilization, as well as its controlled use. As an alternative, plant tissue culture refers to techniques which allow in vitro production of botanic material, including the ones with uncontrolled propagation, minimizing the impacts of the species multiplication on the environment and enabling its rational exploitation. Of all techniques used to produce in vitro metabolites, root cultures are very promising. Therefore, the present work aimed to: (1) Determine the best in vitro conditions in order to maintain root explant donor plants; (2) Evaluate the effect of the medium supplementation with IBA, as well as of the presence or absence of light on the growth of adventitious root cultures; (3) Analyze the effect of the addition of Methyl Jasmonate (MeJa) on the production of secondary metabolites by root cultures; (4) Characterize a phytochemical profile by thin layer chromatography (TLC) of adventitious root cultures and their mediums, comparing it with the extracts of in vivo growth plants and (5) Determine the most efficient method of root culture production according to productivity and inputs. In order to do that, were evaluated: (1) three leaves decontamination protocols (Hypochlorite; Hypochlorite + Tween 80; Ethanol + Hypochlorite) for the establishment of in vitro cultures; (2) three culture systems (solid, liquid and over paper bridge mediums) and two culture mediums (MS0 e MS + 1 mg.L-1 NAA) for the plant propagation and root production; (3) root culture conditions (Flask size, light condition, IBA concentration, MeJa elicitation), and (4) the roots drying method (laboratory oven or lyophilizer), for DW measurement and extract preparation. Phytochemicals analysis were performed with TLC technique, comparing in vivo and in vitro material, with or without elicitation. Results indicate that hypochlorite 2% for 15 minutes is enough for the in vivo decontamination, promoting better rates of regeneration. The paper bridge system over MS + 1 mg.L-1 NAA medium led to the production of plants with leaves with higher propagation capacity, and roots that, once put on liquid medium, presented higher rates of biomass accumulation. Erlenmeyer size was not considered a limiting factor over the roots growth for a period of 30 days; on the other hand, the absence of light during the same period has shown to be significant to increase biomass accumulation in all treatments evaluated. The best IBA concentration varied according to the system roots originated from. For the paper bridge over MS + 1 mg.L-1 NAA medium system, roots grew better when put on a 5 mg.L-1 AIB concentration. The drying method did not affect the dry weight, so lyophilization was chosen, once it allowed the posterior use of the material for extract preparation. Elicitation did not interfere on the biomass accumulation, but it altered the secondary metabolites production. TLC evidenced the presence of terpenoids, flavonoids and saponines on in vivo and in vitro extracts. The study of different phases of K. pinnata adventitious root culture allowed determining conditions to enhance production and make the method more efficient.