Development of method of qualitative analysis of bird cherry fruit for inclusion in the monograph of state pharmacopoeia of Ukraine

Abstract

Introduction.Bird cherryPadusaviumMill, Rosaceae,is widespread in Ukraine, especially in forests and forest-steppe areas. Bird cherry fruits have long been used in medicine and is a valuable medicinal raw materials. They stated to posess astringent, anti-inflammatory, phytoncidal properties.Bird cherry fruits are included in the USSR Pharmacopoeia IX ed., The State Pharmacopoeia of the Russian Federation, The State Pharmacopoeia of Republic of Belarus. In Ukraine there are no contemporary normative documents for this medicinal plant material, therefore it is the actual to develop projects inthe national monographs "dry bird cherry fruit" and "fresh bird cherry fruit" to be included in the State Pharmacopoeia of Ukraine. According to European Pharmacopoeiarecommendation method of thin-layerchromatography (TLC) is prescribed only for the identification of the herbal drug. The principles of thin-layerchromatography and application of the technique in pharmaceutical analysis aredescribed in State Pharmacopoeia of Ukraine. As it is effective and easy to perform, and the equipment required is inexpensive, the technique is frequently used for evaluating medicinal plant materials and their preparations. The TLC is aimed at elucidating the chromatogram of the drug with respect to selected reference compounds that are described for inclusion as reagents. Aim of this study wasto develop methods of qualitative analysis of bird cherry fruits for a monograph in the State Pharmacopoeia of Ukraine (SPU).MaterialsandMethods. The object of our study was dried bird cherry fruits (7 samples) and fresh bird cherry fruits (7 samples) harvested in 2013-2015 in Kharkiv, Poltava, Luhansk, Sumy, Lviv, Mykolaiv regions and the city Mariupol. Samples were registered in the department of SPU State Enterprise "Pharmacopeia center". In accordance with the Ph. Eur. and SPU requirements in "identification C" determination was performed by TLC. TLC was performed on glass-backed silica gel F₂₅₄Merck plates, size 20x10 cm. Test solutions from samples of dried raw material were prepared by extraction of 1% solution of hydrochloric acid in 95% ethanol with ultrasound for 60 minutes at 50º C. As mobile phase solvent mixture ethyl acetate - anhydrous acetic acid - formic acid - water ratio (100: 10: 10: 25) was selected. For preparation the reference solution 1 mg chrysantheminwas dissolved in 10 mL of 1% solution of hydrochloric acid in 95% ethanol. Fresh bird cherry fruits test solutions of the samples were crushed to mash (free of seeds), 10 mL of methanol was added and treated with ultrasound for 15 minutes. As mobile phase solvent mixture formic acid anhydrous-water-butanol ratio (16:19:65) was selected. Two reference solutions were used. First was prepared from 2 mg chrysanthemum dissolved in 5 mL of methanol and second was Pharmacopoeial Reference Standard SPU of bird cherry extract dissolved in 0.2 mL of methanol with ultrasound bath. Test solutions and the reference solutions were placed onto the chromatographic plates and placed into the chamber. When the chromatograms developed, the plates were taken out, then observed the spots in daylight.Results and discussion. In the chromatogram it was observed for all dry bird cherry fruits samples two pink-red color zones, one of which coincided for Rf and color of chrysanthemin, and the other one was slightly lower. The chromatogram for all samples of fresh raw material showed two pinkish-violet zones, one of which had Rf and color corresponded chrysanthemin, and the second one was slightly lower. Compared with bird cherry extract, it was observed two pinkish-violet zones, coincided with Rf and color of bird cherry (SPU) extract zones.In accordance with the requirements of Ph. Eur. identification by TLC is obligatory. Division of anthocyanins of plant raw materials was observed sufficiently in our chosen solvent systems and methods of analysis that allows to recommend these systems and methods of analysis for inclusion in the relevant sections of monographs for bird cherry fruit, dried and fresh for SPU.Conclusion. It was confirmed the opportunity to include in the national monographs "Bird cherry fruits fresh " and " Bird cherry fruits dry" in section "Identification C" analysis of anthocyanin by TLC in chosen systems and methods.