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Download Article (PDF) Advances in Biological Sciences Research, volume 7 1st International Symposium Innovations in Life Sciences (ISILS 2019) Phytochemical Studies and Standardization of Scabiosa Succisa L. Lyudmila Efremencko Anastasiya Malyutina Dmitriy Pisarev Belgorod State National Research Belgorod State National Research Рeoples’ Friendship University of University University Russia Belgorod, Russia Belgorod, Russia Moscow, Russia [email protected] [email protected] [email protected] Valentina Scorbach Nataliya Shestopalova Valentina Kasakova Belgorod State National Research Belgorod State National Research Belgorod State National Research University University University Belgorod, Russia Belgorod, Russia Belgorod, Russia [email protected] [email protected] [email protected] Dariya Fadeeva Belgorod State National Research University Belgorod, Russia [email protected] Abstract—The aim of this work is phytochemical studies insoluble in hydrochloric acid, microbiological purity), and standardization of Scabiosa succisa L. By reverse phase impurities, quantitative content of the main active high performance liquid chromatography of ethanol extract substances. The development of standards for these substances belonging to polyphenolic compounds were found, parameters is especially important for the first time- among which hydroxycinnamic acids predominate. considered plants, which are promising sources of Chlorogenic acid and flavonoid glycosides of apigenin and biologically active substances [4,7-9]. luteolin were identified. Microscopic analysis revealed the main diagnostic features of the Scabiosa succisa L. herb. One such plant is a member of the Caprifoliaceae family As a result of research component composition of Scabiosa - Scabiosa succisa L. It is a perennial herbaceous plant succisa L. polyphenols was partially installed, which common in Western Europe and throughout the European substantiates its anti-inflammatory properties, and part of Russia, which prefers moist, slightly acidic soils of microdiagnostics signs were discovered, which can determine meadows, forests, thickets and forest edges. Scabiosa the plant authenticity. succisa L. has a long erect stem, branched at the top, Keywords—Scabiosa succisa L., Caprifoliaceae, glabrous at the base and pubescent to the top. Leaves are standardization, high-performance liquid chromatography, ovate-lanceolate, glabrous, with petioles, upper leaves are microscopy, phenolic compounds sessile, linear-lanceolate. Flowers have a four-lobed purple corolla. They are collected in apical hemispherical inflorescences, surrounded by involucre of a double leaf I. INTRODUCTION row. Fruit is an achene [10,11]. According to information provided by the world health organization, most people in the world, choosing medicines In the middle ages a property of Scabiosa succissa L. to over-the-counter, prefer drugs based on phytocomponents. heal a toothache was described by Pope John XXI in the The growing demand for herbal medicines is also typical for sourcebook of traditional medicine "Thesaurus pauperum". the Russian pharmaceutical market, while biologically A Scabiosa succisa L. decoction was used as an antidote active additives (herbal teas, fees, capsules, tablets with against the poisonous snakes bites. It was also used to treat plant extracts) and parapharmaceutical products (creams, headaches and stomach cramps. In Russia, the plant leaves lotions, shampoos, etc.) are also very popular. It’s connected were applied to ulcers, on the skin for scabies and eczema with the population’s centuries-old commitment to treatment and were used as a wound healing agent. In Europe folk by means of natural origin, which, in addition, have a medicine expectorant and anthelmintic properties of number of advantages over synthetic ones: they are non- Scabiosa succisa L. are valued. Researchers conducted by toxic, have a minimum of side effects and can be used for a modern scientists have shown that the plant also has long treatment course [1 – 6]. The herbal remedies market is antifungal activity comparable to nystatin [11-13]. updated, new combinations of plant components and All these properties give reason to believe that this plant medicines based on new plant sources appear, and the may be interesting for medicine and the pharmaceutical questions of ways to confirm the medicinal plant raw industry as a promising source of biologically active materials quality remain relevant. substances. However, the available data of Scabiosa succisa Quality standards of medicinal plant raw materials are L. chemical composition is not enough, from the literary described in almost all pharmacopoeias, including source it is known only that it contains tannins, saponins and International one. The State Pharmacopoeia of Russian flavonoids, without specifying their qualitative and Federation normalizes such parameters as authenticity quantitative composition [14]. And the presence of some testing (raw material external signs, microscopy, similar species, which are difficult to distinguish from determination of the main groups of biologically active Scabiosa succisa L. only by external description, makes it substances), numerical indicators (humidity, total ash, ash necessary to search for a number of microdiagnostic signs Copyright © 2019, the Authors. Published by Atlantis Press. This is an open access article under the CC BY-NC license (http://creativecommons.org/licenses/by-nc/4.0/). 97 Advances in Biological Sciences Research, volume 7 that can confirm the authenticity of Scabiosa succissa L. computer programs "Adobe Photoshop CS5 Extended" and raw material. "Microsoft Office Picture Manager". Therefore, the aim of this research is phytochemical III. RESULTS AND DISCUSSION study and standardization of Scabiosa succissa L. During chromatography 21 peaks of various substances II. EXPERIMENTAL were registered, which are typical for the complex of polyphenolic compounds (fig. 1). Among them hydroxy- The investigation object was a dried whole Scabiosa cinnamic acid predominate, flavonoid glycosides are found succissa L. herb, harvested in 2018 in the Ivanovo region in in smaller amounts. C-glycosides of luteolin and apigenin the flowering period. were identified by characteristic retention times, among The study of the herb component composition was acids – chlorogenic acid (table II). carried out by high-performance liquid chromatography on The UV profile of chlorogenic acid is shown in fig. 2. the "Agilent Technologies 1200 Infinity" device. The device has an automatic sampler Agilent 1200, a vacuum The carrier of important anatomical and diagnostic signs microcapacitor, gradient pump and thermostat of the same of the plant is the stem. In its microscopic analysis, attention series. Chromatographic column is «Ascentis express is paid to such characteristics as the primary and secondary C182,7μм × 100 mm × 4.6 mm». structure, the thickness of the collenchyma, the presence of a starch sheath, the type of vascular bundle and the nature of Ethyl alcohol and water of HPLC qualification were the conductive tissues location [15]. used as the mobile phase, formic acid was an acid modifier. As a result of the Scabiosa succisa L. stem cross section Chromatography was performed under the following microscopic analysis, the following microdiagnostic signs conditions: were revealed. column thermostat temperature - +35 ° C; DAD1 D, Sig=325,4 Ref=off (2018-04-27\SCABIOSA.D) mAU 6.148 mobile phase speed-0.5 ml / min; 120 the sample volume is 1 µl. 100 Gradient elution conditions are given in table I. 80 60 TABLE I. CONDITIONS FOR A GRADIENT ELUTION OF SCABIOSA SUCCISA L. POLYPHENOLS 40 15.687 18.892 Time, min А,% B,% 20 11.123 12.581 9.012 7.892 15.112 14.861 22.326 2.449 3.730 7.041 16.384 17.106 37.258 4.324 9.664 5.118 30.452 0 90 10 0 27.661 10 80 20 20 70 30 0 5 10 15 20 25 30 35 min 30 50 50 Fig. 1. Chromatogram of the separation of Scabiosa succisa L. 40 10 90 polyphenolic complex (diode matrix detection, λ=325 nm). The absorption spectra were recorded using a diode- matrix detector of the Agilent 1200 series (wavelength range TABLE II. CHARACTERISTICS OF SUBSTANCES ISOLATED FROM THE SCABIOSA SUCCISA L. HERB ALCOHOLIC EXTRACT from 190 to 950 nm), the scanning period was 2 nm. Retention Quantitative relation, Detection was carried out using the following wavelengths: Substance for flavones – 336 nm, for hydroxycinnamic acids – 310, time, min % 325 nm. Chlorogenic acid 6,146 42,17 The spectra and chromatograms were processed using C-glycoside of 11,123 3,26 Agilent Chem Station software. Identification of separated apigenin polyphenolic compounds was performed by comparing the C-glycoside of 9,012 3,10 retention times of the peaks of the test sample with the luteolin retention time of the standards. The quantitative content of substances was assessed by internal normalization of the *DAD1, 6.146 (149 mAU, - ) Ref=5.886 & 6.746 of SCABIOSA.D peak area. mAU 140 For the purpose of chromatography, the extraction from 120 the dried raw material was prepared as follows. 2.5 g of Scabiosa succisa L. herb, crushed to a particle size of 1 mm, 100 was filled with 70% ethyl alcohol in an amount of 50 ml, 80 placed in a 100 ml volumetric flask and extracted in a water 60 bath with reflux condenser for 30 minutes. 40 The microscopic structure of the Scabiosa succisa L. 20 herb was analyzed using the methods set out in the XIV 0 State Pharmacopoeia of Russian Federation, using a 200 220 240 260 280 300 320 340 360 380 nm
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