CONSTITUENTS OF THE ESSENTIAL OIL IN SOLIDAGO CANADENSIS L . FROM EURASIA

Hydro distilled essential oils in air-dry samples of aerial parts of Solidago canadensis L., (Asteraceae) from eight local invasive populations were investigated by GC-MS analysis. A comparative study on quantity and composition of the essential oils obtained from plants, growing in different ecological and climatic conditions, ontogenesis phase and different plant organs was carried out The major compounds detected in oil samples of S. canadensis were α-pinene (1.3 – 61.27%), limonene (0.5 – 22.5%), bornyl acetat (3.4 – 29.8%) and germacrene D (1.8 – 39.2%). Samples from inflorescences contained the maximal percentage of monoterpene hydrocarbons, while the leaves' samples showed the maximal cumulative percentage of sesquiterpene and monoterpene hydrocarbons. Data obtained from our studies confirm the availability of alien invasive species Solidago canadensis for medicine and many other purposes. The variability of the qualitative and quantitative composition of essential oils in different geographical locations will allow futher selection of form containing the maximum amount of active substances.


INTRODUCTION
Nowdays alien invasive plants spread in the secondary distribution range in mass and can displace native species especially in the antropogenically transformated sites.The resource potential of these «new species» didn't studyied in detail yet.The direct and indirect effects of occurrence of invasive plants on food production and other positive externalities of its appearence have been analysed only for a few species (Fehér et al., 2016).In this regard not without interest is genus Solidago L. that comprises about 100 species, five of them occurring in the Old World.Solidago virgaurea L. is the only one native for Eurasia.The other four -S.canadensis L., S. gigantea Ait., S. rupestris Raf. and S. graminifolia (L.) Salisb.(=Euthamia graminifolia (L.) Nutt.) are of North American origin (Vinogradova et al., 2010).S. canadensis and S. gigantea are the invasive plants, widely distributed in Russia and almost all European countries.The plant material of medicinal usage, known as Herba Solidaginis, includes S. virgaurea, S. gigantea and S. canadensis (Kalemba and Thiem, 2004).This material, its extracts and derivatives serve as components in many medications, applicable while treatment of the urinary tract and prostate diseases.
S. canadensis is used in medicine for treatment of several diseases.The flowers contain analgesic, astringent, febrifuge compounds, infusion of the dried powdered herb is used as an antiseptic agent and root is applied as poultice to burns (Moerman, 1998).The species has been used in European phytotherapy for over 200 years for treatment of chronic nephritis, cystitis, urolithiasis, rheumatism and as an antiphlogistic drug (Apati et al., 2003).The chemical composition of the essential oil in S. canadensis is poorly known.The essential oils obtained from fresh green parts, dried aerial parts, inflorescences, leaves and roots of S. canadensis collected from Poland, India, Egypt and China were previously investigated (Kalemba et al., 1990;Weyerstahl et al., 1993;Xia et al., 1999;Mishra et al., 2010Mishra et al., , 2011;;El-Sherei et al., 2014).Results of those investigations showed that inflorescences of S. canadensis yielded 1.7%, 0.9% (leaves), and 0.2% (steams) of the essential oil.This oil contains 85% hydrocarbons and 15% terpenoids.Earlier, α-pinene, limonene, bornyl acetate, germacrene D, myrecene and γ-cadinene had been identified.

Scientific hypothesis
The major goal of this work was the determination chemical composition of the essential oils and the characterization essential oil chemical polymorphism of S.canadensis invasive populations from Eurasia.

Locating plants and data collection
The specimens of S. canadensis (Figure 1) were collected in eight local invasive populations from Austria, Ukraine, Kazakhstan and Russia (Moscow, Penza, Tver and Tula regions) geographically distant from each other (Table 1).The specimens were collected both at vegetative and flowering buds' formation phases (June -August 2015).For the purpose of phytochemical analysis, one to three specimens (all the aerial shoots) were sampled within 0.2 m 2patches in all the examined local populations, simultaneously with collection of the herbarium specimens.The herbarium specimens are kept in the herbarium of Main Botanical Garden RAS (MHA).

Chemicals
Various reference chemicals used in this study were obtained from Sigma Chemical Co.(St Louis, MO, USA).All other chemicals used in this study were purchased from Merck (Darmstadt, Germany).

Methods of the Essential Oil Composition Study
Essential oil was obtained from an average sample of aerial parts (leaves, inflorescences and a mixture of inflorescences and leaves) of plants.The oil was extracted from chopped air-dried material by hydrodistillation according to Ginzberg (1932).Qualitative composition of the oil was determined by gas chromatography at the Center for Collective Use of the Federal Research Center "Fundamentals of Biotechnology", Russian Academy of Sciences, Moscow (RFMEFI62114X0002), using a Shimadzu GS 2010 gas chromatograph with a GCMS-QP 2010 mass detector and SPB-1 nonpolar column (solidphase-bound methyl silicone) (Supelco, Sigma-Aldrich Corporation) with the length of 30 m and diameter of 0.25 mm.The samples were dissolved in benzene at a ratio of 1 : 150 and chromatographed with the temperatureprogramming mode under the following conditions: injector temperature, 180 °C; interface, 205 °C; detector, 200 °C.The carrier gas was helium.The flow through the column was 1 cm 3 /min and flow division was 1 : 10.Mass detector settings: logging mode, TIC; mass range, 30 -400 m/z.The temperature regime for the substances with the retention index of 1300 consisted of the thermostat at 60 °C for 3 min; then, 2 °C.min - until 230 °C; and isotherm for 2 min.The temperature regime for the substances with a retention index higher than 1300  consisted of the thermostat at 60 °C for 1 min; then, 4 °C.min - until 230 °C; and isotherm for 2 min.The percentage composition of the oils was computed by the normalization method from the GC peak areas, which were calculated as mean values of two injections of each oil sample, without using response factors.The identity of the components was assigned by comparison of their retention indices, relative to a C9 -C17 hydrocarbon standard mixture.

Statistical analyses
Results are expressed as mean of triplicate trials.Data were analyzed by one-way analysis of variance (ANOVA) on the means of values (p <0.05).The essential oils composition was used to determine the relationship between the different samples by cluster analysis using PAST 2.17.

RESULTS AND DISCUSSION
Essential oil contents of S. canadensis varied between 0.07 and 0.70%.Accumulation of essential oil of this species depends on the ecological and climatic conditions of growth, ontogenesis phase and testing plant's organs.
The maximum content of oil was found in the leaves collected at vegetative phase (0.7%).Inflorescences collected at flowering buds' formation phase yielded more oil than the leaves (Table 1).
The trace amount of essential oil (0.1%) was recorded in the plants at the northernmost collection point-Tver region, Kimry district; the maximal quantity (0.4%)in the plants from three local invasive populations (As, Pz and Tl).
Qualitative and quantitative variations in the oils' composition in leaves, inflorescences and aerial parts for all the specimens collected were observed (Table 2, 3).
The composition of essential oil significantly varies in aerial parts of S. canadensis, depending on the geographical locality of the invasive population, too.The monoterpene hydrocarbons were significantly higher in all the samples compared to sesquiterpenes.
Cluster analysis (Figure 2), confirmed a high chemical correlation among all populations (Scorr >0.80%).Two clusters were defined in the bases of the amount of monoterpenes and germacrene D. In cluster 1, which included 6 out of the 9 samples, germacrene D ranged from 3 to 24%, whereas the three sample of cluster 2 showed a higher percentage (32 -36%).Correlation was detected between the clusters and the geographical collection site.The most part of samples the essential oils studied belonged to the "European" chemotype.And only the three samples (Kz, Bs, Sh) belonged to the "Asian" chemotype.Sub-cluster a has a relative amount between 15.9 and 24.0%, sub-cluster b between 8.7 and 10.3% and sub-cluster c (Tv) has the lower relative amount (3%).No significant correlation was detected between the clusters and the geographical collection site (p = -0.35,r= 0.49).
The presence of germacrene D as the major component in overwhelming majority oil samples is in agreement with the previous reports on its occurrence as a major constituent in the oils obtained from the fresh green parts (23.8%), leaves (28.4 and 64.06%) and fresh flowers (29.5%) of S. canadensis collected from Poland (Kalemba et al., 1990;Weyerstahl et al., 1993;Kalemba and Thiem, 2004)

CONCLUSION
There is a great potential in finding new antimicrobial drugs from the wild.Natural crude drug extracts and biologically active compounds obtained from plant species used in traditional medicine can be prolific resources for new drugs.Moreover, according to literature, some monoterpenes and sesquiterpenes as α-pinene, myrcene, limonene and germacrene D have cytotoxic effects against different cell lines.We can support usage of such natural products as potent preservative agents, as well as possible candidates for new medical preparations.Data from our studies confirm the availability of Solidago canadensis: its aerial part containes from 0.1 to 0.7% of essential oil in the leaves and from 0.1 to 0.4% of essential oil in the inflorescens.The major compounds detected in oil samples of S. canadensis were α-pinene (1.3 -61.27%), limonene (0.5 -22.5%), bornyl acetat (3.4 -29.8%) and germacrene D (1.8 -39.2%).Samples from inflorescences contained the maximal percentage of monoterpene hydrocarbons, while the leaves' samples showed the maximal cumulative percentage of sesquiterpene and monoterpene hydrocarbons.The variability of the qualitative and quantitative composition of essential oils in different geographical locations will allow futher selection of form containing the maximum amount of active substances.

Figure 1
Figure 1 Solidago canadensis L. in the Moscow district, photo by Yu.Vinogradova, September 2016.

Figure 2
Figure 2 Dendrogram obtained by cluster analysis of the percentage composition of essential oils from the Solidgo canadensis L. samples examined.For abbreviations, seeTable 1 and the text.

Petrová et al., 2015; Foltinová et al., 2017). For example
, essential oil as an potential sources of antimicrobial ingredients are applied for chicken thighs meat treatment (

Kačániová et al., 2016). Essential
oils can be used as well for the control

Table 1
Examined invasive populations of Solidago canadensis L.

Table 2
Essential oil composition (%) of the leaves and inflorescens Solidgo canadensis L.within different phases of plant's ontogenesis.

:
RIretention index relative to C9-C17 n-alkanes on SPB-1 column (Supelco); «-»component is not present.identified in considerable amounts (3.4 -26.3%) in the oil of aerial parts of the plants, collected in Russia, Austria, Ukraine and Kazakhstan.