Phenolic compounds of Bassia prostrata (Chenopodiaceae)

The composition and content of phenolic compounds i plants of the polymorphic species Bassia prostrata (Chenopod iaceae) from geographically distant populations were studied by high-performance liquid chromatography. The material for the study w as the aboveground part of the plant. The plants were collected in the flowering phase in Russia (Novosibirsk region, Republic of Tyva, Khakassia, Bur yatia, Altai), Kazakhstan, Armenia. The phenolic compounds were ex tracted by double extraction with 70% ethanol. Component composition of the phenolic complex was investigated by Agilent 1200 chromatogr aph. Eleven phenolic compounds, including isoquercitrin, kaempf erol-3-glycoside, isoramnetin-3-rutinoside, and luteolin were found i the composition. The quantitative content of each compound could vary fr om 0.1 to 10.8 mg/g in different populations. Chemotypes were determined fo r the qualitative and quantitative content of phenolic compounds


Introduction
Bassia prostrata (L.) A.J. Scott s. l. (≡ Kochia prostrata (L.) Schrad.) is a species of the Chenopodiaceae family. It is a drought-and salt-tolerant species that is a valuable pasture plant. It is used in breeding to improve or create pastures in desert and arid zones. The main range of B. prostrata extends from the northern Black Sea region, the Caucasus to Central Asia. In Russia, the eastern border of the range passes through Zabaikalski Krai and eastern Mongolia.
B. prostrata s. l. is characterized by the presence of several cytotypes. Different ploidy levels of this species have been repeatedly reported in the literature. Diploids (2n = 18) have been reported from the Republics of Tyva, Altai, Krasnoyarskii Krai, Mongolia, Kazakhstan, Uzbekistan, and Iran; tetraploids (2n = 36) from Kazakhstan and China. Hexaploids (2n = 54) found in Uzbekistan, Pakistan, and Iran were noted much less frequently.
Mainly the study of the chemical composition of extracts of the aboveground parts of plants of the Bassia genus was carried out for pharmacological research, with the detection of different biological activity of the extracts and production of new medicinal preparations. There is information on the anthelmintic, cardiotonic, tonic, and other activities of extracts *Correspondingauthor:pet.a@mail.ru of above-ground parts of plants of genus Bassia in folk and traditional medicine. Scientific and clinical studies have been conducted on hypotensive, hypolipidemic, analgesic, anticancer, and other properties [7][8]. For the species B. prostrata, B. scoparia (L.) A.J. Scott, B. laniflora (S.G. Gmel.) A.J. Scott, B. iranica (Litv. ex Bornm.) Bornm., B. muricata (L.) Asch., data on flavonoids, tannins, saponins, alkaloids are given [9][10][11][12]. B. prostrata has been the subject of joint pharmacological studies in Kazakhstan and Pakistan [13]. 20 amino acids, 8 fatty acids, and 4 phenolic acids -protocatechuic, vanillinic, isovanillinic, and p-coumaric -were isolated and identified in the studied objects.
This work aimed to study the composition and content of phenolic compounds of B. prostrata plants from geographically distant populations of different cytotypes by highperformance liquid chromatography methods. The aim was to investigate the variability of phytochemical traits in natural populations of B. prostrata and to analyze the use of biochemical (composition and content of phenolic compounds) traits as additional markers in the identification of diploid and tetraploid cytotypes.

Materials and methods
The object of the study is the leaves of the plants of B. prostrata in the flowering phase collected in the period 2019-2020 in Novosibirskaja Oblast', the Republics of Khakassia, Buryatia, Altai, Tyva, etc. (Table 1). The raw materials were dried in well-ventilated rooms, crushed to a particle size of 2 mm, mixed and an average sample was taken.
Double extraction with 70% ethanol in a water bath (60-70°C for 1.0-1.5 h.) was carried out to extract the phenolic compounds. An exact weight (0.1 g) of the crushed material was extracted with 30 ml of aqueous ethanol for 30 min followed by 20 ml for 20 min. The residue on the filter was washed with 5 ml of 70% ethanol. The combined extract was concentrated to 10-15 ml (exact volume). To free from impurities, 1 ml of the extract was diluted with bidistilled water to 5 ml and passed through a Diapack C16 cartridge.
The component composition of the phenolic complex of the samples was studied by high-performance liquid chromatography on an analytical HPLC system consisting of an Agilent 1200 liquid chromatograph. The chromatographic separation was conducted at 25°C on a Zorbax SB-C18 Column (4.6 × 150 mm, 5 µm internal diameter) with the Agilent Guard Column Hardware Kit. Detection was performed at a wavelength of λ = 360 nm [11]. Quantification of individual components in plant samples was carried out using the external standard method.

Results
Using high-performance liquid chromatography to separate the complex of phenolic compounds, we were able to conduct a comparative study of the qualitative composition and content of phenolic compounds in samples of B. prostrata from geographically distant regions of Russia and neighboring countries, 14 samples in total were examined ( Table 1).
The quantitative content of each compound in different populations as a whole could vary from 0.1 to 10.8 mg/g. The highest content of isoquercitrin (3.4 mg/g) is characteristic of plants from Armenia (RU-AR2) (hexaploid). The high content of kaempferol-3glycoside was observed both in diploid cytotype, from Tyva (RU-TU2) -6.4 mg/g, and in hexaploid from Armenia (RU-AR2) -4.8 mg/g. The highest content of isoramnetin-3rutinoside was observed in hexaploid from Armenia (10,3 mg/g), tetraploid from Novosibirsk region (RU-NO1) (7,4 mg/g), and diploid from Tyva (RU-TU2) (6,5 mg/g). The highest luteolin content was observed in hexaploid from Armenia and the lowest in tetraploid from Kazakhstan (KZ-A1). In general, almost all compounds showed instability in quantitative content, indicating heterogeneity in populations. In almost all plants collected in Kazakhstan from both diploid and tetraploid cytotypes the compounds №№ 1, 2, 7 (kaempferol-3-glycoside), 8 (isoramnetin-3-rutinoside) were the predominant components. In samples from Buryatia (diploid) similarity was observed for the predominance of component № 3 and 6, in samples from Tyva -only for component № 3.