Ona Ragažinskienė

Evaluation of phytochemical composition of fresh and dried raw material of introduced Chamerion angustifolium L. using chromatographic, spectrophotometric and chemometric techniques

Due to the wide spectrum of biological activities, Chamerion angustifolium L. as medicinal plant is used for the production of food supplements. However, it should be kept in mind that quality (biological activity) of the herb depends on its geographic origin, the way of raw material preparation or extraction and chemotype. The purpose of this study was to evaluate and compare the compositions of volatile, non-volatile compounds and antioxidant activities of C. angustifolium grown in Kaunas Botanical Garden after the introduction from different locations in Lithuania. The compositions of fresh and air-dried samples were compared. The profile of volatile compounds was analyzed using headspace solid phase microextraction coupled with GC/MS. trans-2-Hexenal (16.0-55.9% of all volatiles) and trans-anethole (2.6-46.2%) were determined only in the dried samples, while cis-3-hexenol (17.5-68.6%) only in fresh samples. Caryophyllenes (α- and β-) were found in all analyzed samples, contributing together from 2.4% to 52.3% of all volatiles according to the origin and preparation (fresh or dried) of a sample. Total amount of phenolic compounds, total content of flavonoids and radical scavenging activity (using 2,2-diphenyl-1-picrylhydrazyl (DPPH)) were determined using spectrophotometric assays. The variation of total phenolic compounds content was dependent on the sample origin, moreover, drying reduced amount of phenolics 1.5-3.5 times. The DPPH free radical scavenging activity was in the range of 238.6-557.1mg/g (expressed in rutin equivalents) in the fresh samples and drastically reduced to 119.9-124.8 mg/g after drying. The qualitative analysis of phenolic compounds in the aqueous methanolic extracts of C. angustifolium was performed by means of HPLC with UV detection. Oenothein B and rutin were predominant in the samples; also caffeic and chlorogenic acids, and quercetin were determined. Chemometric methods, namely principal component analysis, hierarchical cluster analysis and K-means clustering analysis, were applied for evaluation of the results. Chemometric analysis showed existence of different chemotypes of C. angustifolium L. and their relation to the geographic origin.

Flavonoids of willow herb (Chamerion angustifolium (L.) Holub) and their radical scavenging activity during vegetation

PURPOSE Willow herb has been traditionally used in folk medicine and currently it is a potential raw material for production of phytopharmaceuticals. The aim of this work was to determine the highest amount of flavonoids and the highest radical scavenging activity of willow herb, which was collected in different vegetation phases (intensive growing, bud, massive blooming, ripening of fruits (seeds) and the end of vegetation) and in different parts of the plant (blooms, leaves and stems). MATERIAL/METHODS Raw material was collected at Kaunas Botanical garden of Vytautas Magnus University. Willow herb was extracted using methanol/water mixture (75/25 v/v, %). Methanolic extracts were purified using solid-phase extraction. For determination of the radical scavenging activity of compounds the HPLC system with the on-line post-column DPPH (1,1-diphenyl-2-picrylhydrazyl) radical reaction detection was used. RESULTS Five flavonoids were identified and their quantitative distribution and radical scavenging activity were evaluated. The highest total amount of flavonoids and radical scavenging activity were determined in willow herb collected during the massive blooming phase (11.12 ± 0.34 mg/g and 8.71 ± 0.29 mg/g, respectively). CONCLUSIONS The highest amount of flavonoids and radical scavenging activity was determined for raw material collected during the massive blooming phase. Evaluation of different parts of the plant during the massive blooming phase revealed that the highest amount of flavonoids and radical scavenging activity are characteristic for blooms of the plant.

Screening of antioxidant activity and volatile compounds composition of Chamerion angustifolium (L.) Holub ecotypes grown in Lithuania

Since biological activity of medicinal plants is dependent on cultivation area, climatic conditions, developmental stage, genetic modifications and other factors, it is important to study flora present in different growing sites and geographical zones. This study was focused on screening of antioxidant activity of C. angustifolium harvested in six different locations in Lithuania. The total contents of phenolic compounds, flavonoids and 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity were evaluated by spectrophotometric methods. A correlation between radical scavenging activity and total phenolic compounds content was observed (correlation coefficient 0.98). HPLC with online post-column DPPH radical scavenging reaction detection was used for the separation of extracts. Oenothein B, rutin and one unidentified compound were predominant. Volatile compounds were analysed using solid-phase microextraction coupled with gas chromatography–mass spectrometry. Based on the analysis of volatiles, all samples were classified into two chemotypes: (I) with predominant α- and β-caryophyllenes and (II) with predominant anethole.

Analysis of antiproliferative effect of Chamerion angustifolium water extract and its fractions on several breast cancer cell lines

PURPOSE To evaluate the antiproliferative effect of the aerial part of Chamerion angustifolium (L.) Holub. (Onagraceae) extract and its fractions in vitro. This is the first study on the anti-proliferative effect of C. angustifolium on 3 distinct breast cancer cell lines. MATERIAL/METHODS Breast cancer cell lines MCF7, MDA-MB-468 and MDA-MB-231 were exposed to different concentrations of the water extract of C. angustifolium, where DPPH radical scavenging activity was 0.018-0.443mg/ml, expressed in rutin equivalents. Cell growth was analyzed after 24, 48 and 72h of incubation. Solid-phase extraction was applied for the fractionation of C. angustifolium water extract and MDA-MB-468 cell line growth was tested using different fractions. RESULTS The concentrations corresponding to radical scavenging activity of 0.117 and 0.266mg/ml caused MCF7 cells growth inhibition, while in the samples exposed to the highest concentration (0.355 and 0.443mg/ml) no proliferation was register, suggesting cell death. MDA-MB-468 cell analysis showed similar responses. MDA-MB-231 demonstrated cell growth inhibition following the exposure to all analyzed high extract doses (0.117-0.443mg/ml). MDA-MB-468 cells were selected to evaluate the effect of fractions. In the samples exposed to the fraction containing the highest amount (91%) of oenothein B, at the concentration of 0.117mg/ml a pronounced cell growth inhibition while at higher concentrations (0.266 and 0.443mg/ml) no cell proliferation was observed. CONCLUSIONS The consumption of C. angustifolium herb can be advantageous, alongside with conventional breast cancer treatment.

Confirmation of the antiviral properties of medicinal plants via chemical analysis, machine learning methods and antiviral tests: a methodological approach

Medicinal plants are reported to possess antiviral activity, but finding the substances that are responsible for antiviral activity in the complex mixture of the plant extract is an extremely difficult task. In this paper a methodology related to the determination of the antiviral properties of medicinal plant extracts and based on phytochemical analysis, antiviral tests and machine learning methods is described. 16 potentially antiviral medicinal plants were selected, and their chemometric characteristics and antiviral properties were investigated. Three different analytical methods were used for chemical analysis: (i) spectrophotometry, (ii) capillary electrophoresis with contactless conductivity detection, and (iii) gas chromatography-mass spectrometry. 14 attributes were obtained describing the composition of the plant extracts. Viral growth inhibition properties were investigated and 8 candidate plant extracts were selected as being active against viruses. Infectious bronchitis virus was used as a model virus. Machine learning techniques including deep neural network classification, classification and regression tree induction and hierarchical clusterization were used for mining the factors that are responsible for antiviral effects. It was determined that (i) phenolic compounds providing high radical scavenging activity and fractions containing high content of phenolic compounds are positively related to antiviral activity in plant extracts, (ii) hydrophilic compounds that are positively charged (pKa > 4.7) in acidic media and possess medium and low electrophoretic mobility properties are negatively related to antiviral activity in medicinal plants, (iii) phenolic acids with pKa lower than 4.7 are not related to antiviral activity in the extracts, and (iv) volatile compounds in the extracts, including diversity, quantity and different volatility properties, do not affect the antiviral activity of plant extracts. Following the proposed methodological approach, it is possible to confirm which chemometric attributes are responsible for antiviral activity in medicinal plant extracts.

Stable isotope ratio method for the characterisation of the poultry house environment.

ABSTRACT Stable isotope analysis was applied to describe the poultry house environment. The poultry house indoor environment was selected for this study due to the relevant health problems in animals and their caretakers. Air quality parameters including temperature, relative humidity, airflow rate, NH3, CO2 and total suspended particles, as well as mean levels of total airborne bacteria and fungi count, were measured. Carbon isotope ratios (13C/12C) were obtained in size-segregated aerosol particles. The carbon (13C/12C) and nitrogen (15N/14N) isotope ratios were measured in feed, litter, scrapings from the ventilation system, feathers and eggs. Additionally, the distribution of δ13C and δ15N values in different tissues of the chicken was examined. The airborne bacteria and fungi extracted from the air filters collected from poultry farms were grown in the laboratory in media with known isotope values and measured for stable isotope ratios. Analysis of isotope fractionation between microorganisms and their media indicated the applicability of stable isotope analysis in bulk samples for the identification of source material. The analysed examples imply that stable isotope analysis can be used to examine the indoor environment along with its biology and ecology, and serve as an informative bioanalytical tool.