Chromatographic Characterization on Flavonoids and Triterpenes of Leaves and Flowers of 15 Crataegus L. Species The Hawthorn (Crataegus L.) is the most commonly used herb for treating cardiovascular diseases. The purpose of this paper was the analysis and comparison of HPTLC chromatographic profiles of flavonoids and triterpenes of leaves and flowers of 15 Crataegus L. species occurring throughout Eurasia and North America, and determination of specific distinguishing markers of its fingerprints. The optimal chromatographical conditions for evaluation of flavonoids and triterpenes of 15 Hawthorn species were proposed, the specific HPTLC fingerprints were obtained. The flavonoids fingerprints were qualitatively much the same for the pharmacopoeial species C.laevigata/oxyacantha and C.monogyna and other related non-pharmacopoeial European and Asian Hawthorn species including C.curvisepala, C.pseudokyrtostyla, C.fallacina, C.leiomonogyna, C.ambigua, C.sanguinea, C.almaatensis, C.turkestanica. The specific distinctive features for Hawthorn species native to North America, including C.punctata var aurea, C.pringlei, C.festiva, C.douglasii and C.holmesiana, were determined. The evaluation of the triterpenes profiles showed that all 15 analyzed Hawthorn species had quite similar fingerprints.
Authentication of Rhodiola rosea, Rhodiola quadrifida and Rhodiola rosea liquid extract from the Ukrainian market using HPTLC chromatographic profiles Rhodiola rosea and Rhodiola quadrifida are widely distributed and sold in Eastern Europe. The purpose of this paper was to identify R.rosea, R.quadrifida and Rhodiola rosea liquid extract (RRLE) in the Ukrainian market and bring out adulteration cases using chromatographic characterisation by HPTLC. The multiple samples of R.rosea, R.quadrifida and RRLE were compared; the optimal chromatographic conditions for identification of R.rosea and RRLE based on the presence of rosavins and salidroside as well as for identification of R.quadrifida based on the presence of salidroside were proposed; the specific HPTLC fingerprints were obtained; the acceptance criteria for each product were set. The adulteration cases for R.rosea and RRLE samples were established. The dependence on handling R.rosea and presence of rosavins was determined. It was assumed that low-quality raw materials or inefficient technology process were used for RRLE. The consistency of HPTLC fingerprints for R.quadrifida samples was established.
The plants from the Lamiaceae family are traditional remedies of folk and scientific medicine. They are present at the domestic pharmaceutical market as herbal raw material (HRM); dietary supplements; aromatherapy; as well as being included in the composition of drugs for oral use etc. According to the modern direction of international harmonization of conditions for standardizations of herbal drugs, the topical issue is the development of unified methods of HPTLC identification for HRM, including the Lamiaceae family. The aim of the work was the selection of the unified HPTLC chromatographic conditions for identification of chromatographic profiles of flavonoids and hydroxycinnamic acids of HRM of some Lamiaceae species of Ukraine’s flora, presented at domestic pharmaceutical market, such as Sage, Wild Thyme, Thyme, Oregano, Rosemary, Java tea, Peppermint, Lavender, Anise hyssop, Melissa, Hyssop, White horehound, Motherwort; and comparison of their chromatographic fingerprints. As objects of investigation, the HRM of Lamiaceae family presented at the Ukrainian pharmaceutical market were used. The investigation was conducted by HPTLC procedure using an automatic herbal system of CAMAG, Switzerland at the base of CAMAG laboratory, Muttenz, Switzerland and laboratory of ESTCCTI ESIPP, NUPh, Kharkiv, Ukraine. The harmonized chromatographic conditions for determination of flavonoids and hydroxycinnamic acids for HRM of Lamiaceae species were chosen, these were: stationary phase – HPTLC Si 60 F254; mobile phase – ethyl acetate, formic acid anhydrous, water (8:1:1); sample preparation: solvent – methanol; method of extraction – shaking for 10 min; relative humidity: 33%, temperature 22 ± 5 °C; derivatization with Natural product and PEG reagents; detection – 254 nm, 366 nm, white light before derivatization: 366 nm and white light after derivatization. The reproducibility of results in these chromatographic conditions was ∆RF ≤ 0,05. It was established, that for all investigated species of the family the zone corresponding with the position and colour to rosmarinic acid was detected, thus it can be used as standard substance for «Identification», «Limit-test» and for chromatographic system suitability test. For investigated samples of domestic HRM, the specific zones that can be used for setting acceptance criteria of the «Identification» and «Foreign matters» tests were conducted. After validation investigation involving greater numbers of samples of each species from different growing regions of Ukraine, the obtained HPTLC chromatographic fingerprints could be recommended as reference images for quality control of home-grown HRM of Lamiaceae family.
The aim of this work was to improve the approaches for standardization of Thymus L. species by development of HPTLC identification of phenolic compounds (flavonoids and phenylpropanoids) and terpenoids (essential oils and triterpenoids) and quantitative determination of rosmarinic acid and flavonoids for six Thymus L. species of Ukraine flora, as well as determination of chromatographic profiles of Thymus L. species’ extracts obtained using different extraction methods. Materials and methods. Objects were: samples of dry herb of Th.serpyllum, Th.vulgaris, Th.Marschallianus, Th.Pallasianus, Th.calcareus, and Th.moldavicus of Ukraine origin collected in 2019-2021 years or acquired commercially on Ukraine market. Instruments: CAMAG HPTLC Herbal System, visionCats 2.5. The analytical grade reagents were used. Reference substances were purchased from Extrasynthese, Sigma Aldrich. Chromatography was performed on HPTLC plates Si 60 F254, Merck according to the developed methods. Results. A new approach for quality control of different Thymus L. species of Ukraine flora included the development of HPTLC methods for identification of main groups of bioactive substances of these species, such as flavonoids and phenylpropanoids, essential oils, triterpenoids; development of quantification method of rosmarinic acid and assay of total flavonoids, expressed as luteolin-7-O-glucoside. The characteristic HPTLC fingerprints of six Thymus L. species in three mobile phases of different polarities that cover a wide range of bioactive substances were established. The content of rosmarinic acid in different Thymus L. species samples was in the range of 0,11-0,72 %: Th.moldavicus – 0,11 %; Th.Marschallianus – 0,19-0,27 %; Th.serpyllum – 0,38 %; Th.vulgaris – 0,51 %; Th.calcareus – 0,56 %; T.Pallasianus – 0,72 %. The total flavonoids content, expressed as luteolin-7-O-glucoside, was in the range of 0,8-2,72 %: Th.moldavicus – 0,8 %; Th.serpyllum – 0,87 %; Th.vulgaris – 1,06 %; Th.Pallasianus – 1,28 %; Th.Marschallianus – 1,89 %; Th.calcareus – 2,72 %. Conclusions. The proposed scientific approach for quality evaluation of Thymus L. species using HPTLC allows to determine comprehensive information of chemical composition and content of active substances of multiple samples in parallel, in a cost and time-efficient manner
It was known that hawthorn - Crataegus L. is a polymorphic genus. Two hawthorn species and their hybrids are included in the European Pharmacopoeia, twelve – in Ukrainian pharmacopoeia. Determination of chromatographic profiles of hawthorn fruits species native to Ukraine and other countries that are non-pharmacopoeial, but have sufficient plant raw material base, is essential for quality control of drugs. Aim. To analyze and compare the chromatographic profiles of fruits of 23 Crataegus L. species on phenolic compounds, evaluated by means of high-performance thin-layer chromatography procedure (HPTLC), and determine the specific features of chromatographic fingerprints. Materials and Methods. A total of 39 samples of fruits of 23 hawthorn species that are native to Europe, Asia and North America, such as Crataegus monogyna, C. laevigata/C. oxyacantha, C. leiomonogyna, C. curvisepala, C. pseudokyrtostyla, C. fallacina, C. subrotunda, C. ambigua, C. pentagyna, C. sanguinea, C. chlorosarca, C. almaatensis, C.pseudoheterophylla subsp. turkestanica, C. pinnatifida, C. pentagyna subsp. pseudomelanocarpa, C. punctata, C. pringlei, C. festiva, C. douglasii, C. holmesiana, C. submollis, C. flabellata, C. canadensis were investigated. The analysis has been done following the TLC method from European Pharmacopeia modified into HPTLC, using automated HPTLC herbal system (CAMAG, Switzerland). The results have shown that chromatographic profiles of phenolic constituents of nine Crataegus L. species of Europe, both pharmacopoeial and non-pharmacopoeial, were quite similar, despite the significant morphological distinctions. The chromatographical profiles of three species of Asia were similar to the pharmacopoeial species; three other species looked different and had specific marker zones. In addition, eight Crataegus L. species of North America had specific markers helping for discriminative analysis from pharmacopoeial species. Conclusion. The findings could help to identify the possible adulterations and prevent the falsification of finished products. The results will be taken into consideration during revision of the Ukrainian national pharmacopoeial monograph for hawthorn fruits.
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