Teodora Janković

Optimization of polyphenols extraction from dried chokeberry using maceration as traditional technique

Traditional maceration method was used for the extraction of polyphenols from chokeberry (Aronia melanocarpa) dried fruit, and the effects of several extraction parameters on the total phenolics and anthocyanins contents were studied. Various solvents, particle size, solid-solvent ratio and extraction time have been investigated as independent variables in two level factorial design. Among examined variables, time was not statistically important factor for the extraction of polyphenols. The optimal extraction conditions were maceration of 0.75mm size berries by 50% ethanol, with solid-solvent ratio of 1:20, and predicted values were 27.7mgGAE/g for total phenolics and 0.27% for total anthocyanins. Under selected conditions, the experimental total phenolics were 27.8mgGAE/g, and total anthocyanins were 0.27%, which is in agreement with the predicted values. In addition, a complementary quantitative analysis of individual phenolic compounds was performed using HPLC method. The study indicated that maceration was effective and simple technique for the extraction of bioactive compounds from chokeberry fruit.

In vitro cytotoxic and antioxidative activity of Cornus mas and Cotinus coggygria

Cytotoxicity and antioxidant properties of the methanol extracts of leaves and flowers of Cornus mas and Cotinus coggygria were studied, together with their chemical composition. Extracts of C. coggygria flowers and leaves showed better antioxidant activity in reaction with 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical as well as in inhibition of lipid peroxidation (LP) than C. mas extracts. Preliminary results showed that all extracts possessed potential cytotoxic activity towards HeLa and LS174 human cancer cell lines in vitro, with stronger inhibition against growth of HeLa cell growth than against LS174 cell growth. Cytotoxic activity of C. coggygria extracts showed good correlation with their antioxidative activity in vitro. In the brine shrimp lethality test, there were no significant changes in toxicity.

Antioxidant and antiproliferative activity of chokeberry juice phenolics during in vitro simulated digestion in the presence of food matrix

Chokeberry juice was subjected to in vitro gastric digestion in the presence of food matrix in order to determine the changes in polyphenol content and antioxidant activity. Addition of food matrix immediately decreased the total phenolic content, anthocyanin content, DPPH scavenging activity as well as total reducing power by 36%, 90%, 45% and 44%, respectively. After in vitro digestion, total phenolic content, anthocyanin content and reducing power are slightly elevated, but they are still lower than in initial non-digested juice. The effect of digested juice on Caco-2 cells proliferation was also studied, and the reduction of proliferative rate by approximately 25% was determined. Our results suggested that although a large proportion of chokeberry phenolics undergo transformation during digestion they are still potent as antioxidant and antiproliferative agents.

Neuropharmacological evaluation of diethylether extract and xanthones of Gentiana kochiana

Diethylether extract of aerial parts of Gentiana kochiana mostly consists of two tetraoxygenated xanthones: gentiacaulein (1,7-dihidroxy-3,8-dimethoxyxanthone; 76.1%) and gentiakochianin (1,7,8-trihidroxy-3-methoxyxanthone; 14.2%). The extract and these xanthones were evaluated for the CNS pharmacological activity in rodents. In vitro assays on rat brain preparations revealed insignificant interaction of the compounds with the specific dopamine and serotonin receptors or synaptosomal uptake of serotonin. However, the extract and gentiacaulein strongly inhibited rat microsomal MAO A (IC50=0.22 microg/ml and 0.49 microM, respectively). Their effects on MAO B and a gentiakochianin blocking potential on both MAO enzymes were moderate. Behavioral examinations on mice showed that 10 day s.c. administration of the extract (20 mg/kg) significantly decreased immobility score in a forced swimming test and strongly inhibited ambulation and stereotypy in an open-field test. These effects resembled those induced by 10 mg/kg imipramine. The ex vivo MAO A activity in crude brain mitochondrial fraction of mice treated with 20 mg/kg of the extract was significantly elevated, whilst that outside brain nerve terminals was declined. This study suggests some antidepressant therapeutic potential of G. kochiana, particularly of gentiacaulein, with an ambiguity whether pharmacological mechanism could be related only to the central inhibition of MAO A.

SECOIRIDOIDS AND XANTHONES IN THE SHOOTS AND ROOTS OF CENTAURIUM PULCHELLUM CULTURED IN VITRO

SummaryThe qualitative and quantitative composition of secondary metabolites was studied in the shoots and roots of Centaurium pulchellum cultured in vitro. Secoiridoids (gentiopierin, swertiamarin, and sweroside) and xanthones (methylbellidifolin, demethyleustomin, and deccussatin) were isolated. Sweroside was found to be the major secoiridoid compound in the aerial parts of plants growing in nature. while swertiamarin dominated in plants cultured in vitro. In roots of all plants, genciopicrin was the major compound. Xanthone demethyleustomin was the major compound both in the shoots and roots of plants growing in nature and cultured in vitro. Different sugars (glucose, fructose, and sucrose) added in different concentrations in the medium affected the production of secondary compounds.

Antiglioma action of xanthones from Gentiana kochiana: Mechanistic and structure–activity requirements

The present study identifies xanthones gentiakochianin and gentiacaulein as the active principles responsible for the in vitro antiglioma action of ether and methanolic extracts of the plant Gentiana kochiana. Gentiakochianin and gentiacaulein induced cell cycle arrest in G(2)/M and G(0)/G(1) phases, respectively, in both C6 rat glioma and U251 human glioma cell lines. The more efficient antiproliferative action of gentiakochianin was associated with its ability to induce microtubule stabilization in a cell-free assay. Both the xanthones reduced mitochondrial membrane potential and increased the production of reactive oxygen species in glioma cells, but only the effects of gentiakochianin were pronounced enough to cause caspase activation and subsequent apoptotic cell death. The assessment of structure-activity relationship in a series of structurally related xanthones from G. kochiana and Gentianella austriaca revealed dihydroxylation at positions 7, 8 of the xanthonic nucleus as the key structural feature responsible for the ability of gentiakochianin to induce microtubule-associated G(2)/M cell block and apoptotic cell death in glioma cells.

Antioxidant, Cytotoxic, and Antimicrobial Activity of Alnus incana (L.) ssp. incana Moench and A. viridis (Chaix) DC ssp. viridis Extracts

Antioxidant, cytotoxic, and antimicrobial activities of leaves, bark, and cone extracts of Alnus incana (L.) Moench ssp. incana and endemic species A. viridis (Chaix) DC ssp. viridis were evaluated. All extracts were found to be strong 1,1-diphenyl-2-picrylhydrazyl free radical scavengers, exhibiting 50% inhibitory concentration (IC(50)) values of 3.3-18.9 microg/mL, and also showed activity in inhibition of lipid peroxidation with IC(50) values ranging from 38.5 to 157.4 microg/mL. A. incana and A. viridis extracts exhibited significant cytotoxic effects toward HeLa cells, with IC(50) values ranging from 26.02 to 68.5 microg/mL. The most active extract of A. incana bark also contained great amounts of total phenolics (316.2 mg of gallic acid equivalents/g). In our experiment all extracts were virtually nontoxic on brine shrimps. Extracts were screened for activity against 15 microorganisms, and all extracts investigated showed antimicrobial activity. The most active were dry extracts of cones of A. incana and A. viridis with minimum inhibitory concentration values ranging from 0.117 to 0.129 mg/mL.

Radioprotective Effects of Gentianella austriaca Fractions and Polyphenolic Constituents in Human Lymphocytes

The aim of this study was to identify active principles of Gentianella austriaca responsible for the reduction of the incidence of micronuclei in irradiated lymphocytes in vitro. The radioprotective effects of ether (EF) and methanolic (MeF) fractions, water-soluble xanthones demethylbellidifolin (1), demethylbellidifolin 8-O-glucoside (2), bellidifolin 8-O-glucoside (3), and flavonoid swertisin (4) against chromosomal damage induced by gamma-rays were determined using the micronucleus test. EF and MeF showed better protection in treatment of human lymphocytes after gamma-irradiation than did isolated compounds. Among the isolated compounds, the effectiveness in reduction of the frequency of micronuclei followed the order 4>3>2>1. The anti-lipoperoxidant activity was in the order 2>4>1, while 3 slightly increased the level of malondialdehyde. In addition, the effectiveness in induction of apoptosis followed the order, 3>2>4, while 1 had no proapoptotic effect. These results suggest that the antioxidative properties of the polyphenols tested may contribute to the radioprotective effects of G. austriaca.

Secoiridoid content of Blackstonia perfoliata in vivo and in vitro

SummaryThis study reports the analysis of secondary metabolites of gentiopicrin, swertiamarin, and sweroside in shoot and root cultures of yellow wort (Blackstonia perfoliata), which were initiated from seeds, grown on Murashige and Skoog (MS) medium. Shoot cultures of B. perfoliata were inoculated with suspension of Agrobacterium rhizogenes strain A4M70GUS and hairy roots appeared at the infected sites after 3 wk of inoculation. Tips of adventitious roots of B. perfoliata were grown on hormone-free MS medium and three clones of the transformed roots regenerated shoots spontaneously. Gentiopicrin, swertiamarin, and sweroside were detected in both roots and shoots of B. perfoliata in vitro and in vivo, but gentiopicrin was found to be the major compound. The concentration of growth regulator in the medium affected the production of secoiridoids in B. perfoliata in vitro, where the level of gentiopicrin was higher in plants grown in the presence of indole-3-butyric acid, but the presence of 6-benzylaminopurine was inhibitory to secoiridoid production.

Biochemical Properties of the Fresh and Frozen Black Currants and Juices

Thirteen black currant varieties (Ribes nigrum L.) cultured in Serbia were characterized for their pomological properties and chemical composition (total phenolics, total anthocyanins, anthocyanin aglycones, sugars, and vitamin C). The average amount of vitamin C varied from 122.4 to 193.2 mg/100 g fresh weight (FW), while concentration of invert sugars ranged from 6.3% to 11.1%. The highest amounts of total phenolics and anthocyanins were detected in variety Ometa (278.9 mg of gallic acid equivalents per 100 g of FW [mg GAE/100 g FW] and 135.4 mg/100 g, respectively). Quantitative analyses of anthocyanin aglycones in berries were performed using high-performance liquid chromatography, and delphinidin was found to be dominant compound in 11 varieties. Total phenolics and anthocyanins contents decreased during the processing of berry fruits to juices, and the reduction of anthocyanins was more pronounced, 12%-80%. The radical scavenging activity of black currant juices was investigated by 1,1-diphenyl-2-picrylhydrazyl, and the IC(50) value ranged from 1.9 to 4.0 mg/mL. Our results also showed that freezing as a way of preservation and storage could save important phytochemicals and health benefits of berries and berry juices. The amount of total phenolics in berries increased during 1 year of storage by 46.09%-171.76% and in juices by even 107.58%, while the amount of total anthocyanins in berries and juices decrease by 5.63%-52.76% and 13.04%-36.82%, respectively.