CHARACTERIZATION OF DRY EXTRACTS OF EVERLASTING INFLORESCENCES, Helichrysum plicatum DC., AND ASSAYING THEIR ANTIOXIDANT, CYTOTOXIC, SPASMOLYTIC AND ANTIMICROBIAL ACTIVITY

Dubravka Bigović

Abstract


In this doctoral dissertation, the characterization of dry extracts of everlasting inflorescences, Helichrysum plicatum DC., and assay of their antioxidant, cytotoxic, spasmolytic and antimicrobial activity were carried out. Special attention was paid to investigation of correlation between chemical composition and pharmacological effects of dry extracts to determine potentially active substances or active markers.

In the preformulation study, the herbal drug Helichrysi flos (Helichrysum plicatum DC.) was precisely defined, identified and tested. The dry extracts were prepared from powdered herbal drug by triple percolation method and purification of obtained liquid extracts by means of re-extraction method and vacuum evaporation. The following factors varied in the extraction procedures at three levels: degree of fineness of herbal drug, type of extragens (40%, 50%, 60% ethanol/water ratio) and re-extraction solvent (5:5, 9:1, 100:0 ethilacetate/ethanol ratio; v/v).

The HPTLC fingerprint method was developed for rapid and reliable identification of glycosides and aglicone flavonoids present in the herbal drugs and extracts H.plicatum. This identification was based on the color fluorescence, and application of the standards and the tentative LC-UV-MS analysis of purified extracts. .

 In the chemical characterization of Helichrysi flos (Helichrysum plicatum DC.) dry extracts, the total content of phenols has been determined in all extract samples.

Naringenin, apigenin and kaempeferol content was determined by HPLC methods in hydrolyzed and nonhydrolysed samples of dry extracts, and by application of standards. Extraction process optimization, based on factor analysis, showed different optimum extraction performance depending on the extraction target. In order to increase the dry extract yields, the extraction process optimization was based on the influence of various factors (degree of fineness of herbal drug, type of extragens and re-extraction solvent) on the extraction yields.

Antioxidant activity of dry extract samples was evaluated on the basis of the ability of DPPH radical reduction.

Cytotoxic activity of selected extracts of Helichrysi flos (Helichrysum plicatum DC.) and standard substances (naringenin, apigenin and kaempeferol) was tested in vitro on three types of human cancer cells: Cervix Aden carcinoma Hela cells, PC3 prostate cancer cell lines and myelogenous leukemia K562 cells.

The in vitro spasmolytic activity of selected extract of Helichrysi flos (Helichrysum plicatum DC.), assayed on isolated rat ileum, spasmolytic effect of dry extract on spontaneous ileum contractions and contractions induced by acetylcholine, histamine, barium and potassium ions.

Antimicrobial activity of selected extract of Helichrysi flos (Helichrysum plicatum DC.) was tested by in vitro micro dilution method against 20 microorganisms, including bacerias, pests and phytopathogene yeasts.

The results of Helichrysi flos (Helichrysum plicatum DC.) chemical composition and pharmacological activity analyses indicated on potential active substances or active markers used for H.flos extract standardization (biological source Helichrysum plicatum DC.).

The results, as demonstrated above, justify the traditional use of Helichrysum plicatum DC., and indicate the similarity to H. arenarium (L.) Moench, which could be considered as a good basis for further investigations.

 


Keywords


everlasting; Helichrysum plicatum; extraction process optimization; phenols compounds; flavonoids; antioxidant activity; antimicrobial activity; cytotoxic activity; spasmolytic activity.

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Copyright (c) 2017 Dubravka Bigović



ISSN 0455-6224 (Print)
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