Essential oils from plants and in vitro shoot cultures of Hypericum androsaemum L., H. perforatum L. and H. undulatum Schousboe ex. Wild
Year: 2009
Type: dissertation
Abstract: Apical buds and nodal segments, from plants grown in Nature were used as primary explants to establish in vitro shoot cultures of Hypericum androsaemum and Hypericum undulatum, respectively. Hypericum perforatum shoot cultures were established from nodal segments of axenic seedlings grown from seeds germinated asseptically on MS medium devoid of growth regulators. Shoot multiplication was performed by subculturing nodal segments on MS medium devoid of growth regulators, in the cases of H. perforatum and H. undulatum, and on MS medium supplemented with IAA and KIN, in the case of H. androsaemum. A modified Mg medium was used in parallel with the MS medium both devoid of growth regulators, in the case of H. undulatum. After 60 days on MS basal medium, H. undulatum cultures were characterized by a higher number of shoots and roots, comparing with the cultures grown on Mg basal medium. Essential oils (EO) from in vivo plants and in vitro cultures of H. androsaemum, H. perforatum and H. undulatum were isolated by hydrodistillation in a Clevenger type apparatus and analyzed by Gas Chromatography (GC) and Gas Chromatography-Mass Spectrometry (GC-MS). More than 70 compounds were identified in EO from plants of H. androsaemum L. cultivated in two places (Arouca and Arcos de Valdevez) and harvested with intervals of 2 months, over one year. Seasonal variations on the content of sesquiterpene hydrocarbons, the most represented group of compounds, were registered (42.8-72.7% in plants grown in Arouca and 43.4-78.5% in plants grown in Arcos de Valdevez). A high number of intermediate to long chain n-alkanes and 1-alkenes was recorded in EO of H. androsaemum plants grown in Arouca, in the month of February, as well as plants grown in Arcos de Valdevez, during the spring and in the end of winter. In most of the EO of this species, (Ε)-caryophyllene, β-gurjunene and γ- elemene were the major compounds independently of the experimental field. Ripened seed capsules and stems were the H. androsaemum organs with the highest and the lowest EO contents, respectively. Sesquiterpene hydrocarbons dominated the EO of leaves and stems of H. androsaemum, while monoterpene hydrocarbons dominated the EO of the ripened seed capsules. From the five most represented compounds in the EO of H. androsaemum no one was common to all the three organs (leaves, stems and ripened seed capsules). Almost 80% of the total EO of in vitro shoots of H. androsaemum was represented by sesquiterpene hydrocarbons, with γ-elemene as the only major constituent common to EO from in vitro shoots and from in Nature cultivated plants. Essential oils from aerial parts of H. perforatum plants of two cultivars (common cultivar and cv. ‘Topaz’), grown in two experimental fields and sampled over one year, revealed high levels of sesquiterpene hydrocarbons, and low levels of oxygenated compounds. Germacrene D, (E)-caryophyllene and β-selinene were the major compounds. The highest EO content was found in flowers (~12-17 mg/g of dry biomass), in which sesquiterpene hydrocarbons was the major compound group and 2-methyloctane the most represented compound (22-29%). Alkanes which represented no more than 9% of the total EO from in Nature cultivated plants, was the second major group in the EO of in vitro shoots, in which nnonane accounted for more than 24% of the total EO. Essential oils of plants and in vitro shoots of H. undulatum Schousboe ex Willd had nnonane as the major constituent, accounting for more than 40% in most of them. This compound was that most contributed for the high level of n-alkanes group in the EO of the different plant organs, notwithstanding sesquiterpene hydrocarbons constituted the dominant group. The highest yield of H. undulatum EO was obtained from leaves, followed by ripened seed capsules, flowers and stems. The EO contents observed in in vitro H. undulatum shoots (4.9-10.5 mg/g of dry weight, for MS basal medium and 4.1-9.5 mg/g of dry weight, for Mg basal medium) were higher than those observed in aerial parts of in Nature growing plants. Although variations in the composition of the EO from shoots grown on two different basal media had been registered, over the 60 days of culture, the group of alkanes was the major one independently of the culture conditions. The highest contents of n-nonane were recorded in the EO from shoots grown on Mg basal medium. In order to get hairy root cultures of H. androsaemum, H. perforatum and H. undulatum, the influence of several factors (effect of explant pre-culture, bacterial density, explant wounding, addition of acetosyringone to the bacterial suspension and co-culture medium, as well as co-culture period) were evaluated, using the A. rhizogenes-mediated transformation as the main approach. Notwithstanding the several assays performed, hairy roots production was not achieved in any of the tested explants (leaves, internodal segments and roots).
Author: Ana P. Guedes
Language: English
Subfield: Plant Science
Domain: Life Sciences
Open Access status: green
OpenAlex ID: W2262290868