Search

The purpose of this study was to compare the polyphenol, antioxidant and antidiabetic potential of powdered Matcha and bagged Sencha tea during in vitro digestion. Total phenols (TP), flavonoids (TF), flavanols (TFLA), antioxidant and antidiabetic (α-glucosidase inhibition) activity were higher in Matcha tea before and in most in vitro digestion phases. Upon gastric digestion, in Matcha tea TP, TF, TFLA were 2.6, 1.4 and 1.2 times significantly higher (p ≤ 0.05), respectively; gallic acid, gallocatechin, epigallocatechin, quercetin and kaempferol 1.5, 1.6, 1.8, 1.7, 1.2 times, respectively; whereas antioxidant activity was significantly (p ≤ 0.05) higher 3.2 and 1.1 times with ABTS and FRAP and α-glucosidase inhibition 1.8 times. After the intestinal phase, TP and TFLA were 3.4 and 1.7 times significantly (p ≤ 0.05) higher, respectively, antioxidant activity was significantly (p ≤ 0.05) higher 2.4 and 2.0 times with ABTS and FRAP, respectively, while inhibition of α-glucosidase was 1.7 time significantly (p ≤ 0.05) higher in Matcha tea, but the differences in TF, TP and identified phenolics (with the exception of gallic acid) between Matcha and Sencha tea were neutralized. Our results are the first to demonstrate that, during digestion of Matcha powder together with its water extract, Matcha polyphenols are more bioavailable and possess higher antioxidant and antidiabetic activity compared to Sencha. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s13197-021-05086-5.

Research background. Citrus limon (L.) Burm lemon juice is rich in many important natural chemical components (flavonoids, citric acid and vitamin C) and its use in traditional medicine is well known. Formulations of lemon juice with fruit polyphenols in beverages have been investigated, but there is very little information about their ability to modulate the digestive behaviour of polyphenols. The goal of this study is to determine the stability and digestive availability of spinach (Spinacia oleracea L.) polyphenols by adding different volume fractions of lemon juice (0, 2, 5, 10 and 20%) during in vitro digestion. Experimental approach. The content of polyphenols and other abundant compounds including nitrates, oxalic acid and l-ascorbic acid in spinach formulation with various volume fractions of lemon juice were measured in predigested and digested samples using in vitro human digestion model. Antioxidant and α-amylase inhibitory activities of spinach lemon juice formulation were also measured. Results and conclusions. The highest increases in total polyphenols, total flavonoids, total phenolic acids, oxalic acid and nitrate content were noted in predigested and almost all digested spinach samples formulated with the highest volume fraction of lemon juice. In the same sample, the content of individual compounds significantly increased after salivary (l-ascorbic acid), initial (p-coumaric acid) and intestinal (quercetin) phase of digestion. High bioaccessibility of polyphenols and l-ascorbic acid in all phases of digestion was observed in almost all spinach lemon juice formulations, with the exception of nitrates in gastric and intestinal phases and oxalic acid in the intestinal phase, which had moderate bioaccessibility. Novelty and scientific contribution. For the first time the stability and digestive availability of spinach polyphenols, oxalic acid, nitrates and l-ascorbic acid were tested with the addition of different volume fractions of lemon juice. The pH of lemon juice and its l-ascorbic acid content increase the stability and availability of polyphenols in spinach lemon juice formulation during in vitro digestion. Antioxidant and α-amylase inhibitory activities increase in dose-dependent manner after lemon juice addition. Accordingly, spinach formulated with 20% of lemon juice appears as the best source of dietary polyphenols with antioxidant and antidiabetic activities and nitrates that may be used as a functional drink., Pozadina istraživanja. Limunov (Citrus limon (L.) Burm) sok bogat je mnogim važnim prirodnim kemijskim spojevima (flavonoidima, limunskom kiselinom i vitaminom C) i njegova je upotreba u tradicionalnoj medicini dobro poznata. Formulacije limunovog soka s voćnim polifenolima u pićima su istražene, ali postoji vrlo malo informacija o njihovoj sposobnosti moduliranja dostupnosti polifenola u probavi. Svrha je ovog istraživanja bila utvrditi stabilnost i dostupnost polifenola špinata (Spinacia oleracea L.) u probavi dodavanjem različitih volumnih udjela soka limuna (0, 2, 5, 10 i 20 %). Eksperimentalni pristup. Sadržaj polifenola i drugih spojeva (nitrata, oksalne kiseline i L-askorbinske kiseline) izmjeren je u formulacijiama špinata s različitim volumnim udjelima limunovog soka u uzorcima prije i nakon in vitro modela ljudske probave. Također su mjerene antioksidacijska sposobnost i svojstvo inhibicije aktivnosti α-amilaze u formulacijama špinata s limunovim sokom. Rezultati i zaključci. Najizraženije povećanje udjela ukupnih polifenola, flavonoida i fenolnih kiselina, te oksalne kiseline i nitrata zabilježeno je prije probave i u gotovo svim probavljenim uzorcima špinata formuliranim s najvećim volumnim udjelom limunovog soka. U istom uzorku udio pojedinih spojeva značajno se povećao nakon faze probave u ustima (L-askorbinska kiselina), inicijalnoj (p-kumarinska kiselina) i intestinalnoj fazi (kvercetin). Visoka biodostupnost polifenola i L-askorbinske kiseline u svim fazama probave primijećena je u gotovo svim formulacijama špinata s limunovim sokom, s izuzetkom nitrata u želučanoj i crijevnoj fazi te oksalne kiseline u crijevnoj fazi, koji su imali umjerenu biodostupnost. Novina i znanstveni doprinos. Po prvi put je ispitana stabilnost i dostupnost u probavi polifenola špinata, oksalne kiseline, nitrata i L-askorbinske kiseline uz dodatak različitih volumnih udjela soka limuna. pH-vrijednost i L-askorbinska kiselina limunovog soka povećavaju stabilnost i dostupnost polifenola u formulaciji špinata s limunovim sokom tijekom in vitro probave. Antioksidacijska i sposobnost inhibicije aktivnosti α-amilaze povećavaju se ovisno o povećanju doze limunovog soka. Sukladno tome, formulacija špinata s 20 % limunovog soka predstavlja najbolji izvor dijetetskih polifenola s antioksidacijskim i antidijabetičkim djelovanjem te nitrata koji se mogu koristiti kao funkcionalni napitak.

Ethnopharmacological relevance: Arctostaphylos uva-ursi (L.) Spreng. (bearberry) is a well-known traditional herbal plant used as a urinary tract disinfectant. Its antiseptic and diuretic properties can be attributed to hydroquinone, obtained by hydrolysis of arbutin. Aim of the study: This study aimed to determine the toxic profile of free hydroquinone on urinary bladder cells (T24) as a target of therapeutic action. Materials and methods: Quantitative and qualitative analysis of the extract and the digestive stability and bioavailability of arbutin and hydroquinone were performed by HPLC assay and simulated in vitro digestion, respectively. Cytotoxic effect, reactive oxygen species induction and proteome changes in T24 cells after hydroquinone treatment were determined using Neutral red assay, 2′ , 7′ -dichlorofluorescein-diacetate (DCFH-DA) assay and mass spectrometry, respectively. Results: Through in vitro digestion, arbutin was stable, but hydroquinone increased after pepsin treatment (109.6%) and then decreased after the small intestine phase (65.38%). The recommended doses of Uva-ursi had a cytotoxic effect on T24 cells only when all hydroquinone conjugates were converted to free hydroquinone (320 and 900 μg/mL) and the toxic effect was enhanced by recovery. One cup of the therapeutic dose had a prooxidative effect after 4 h of incubation. Shorter time of cell exposure (2 h) to hydroquinone did not have any impact on reactive oxygen species induction. Proteomic analysis found 17 significantly up-regulated proteins compared to control. Hydroquinone activated proteins related to oxidative stress response, stress-adaptive signalling, heat shock response and initiation of translation. Conclusions: Despite the therapeutic properties of bearberry, up-regulated T24 cell proteins are evidence that plant compounds, although from a natural source, may exhibit negative properties.

The content of polyphenols and other abundant compounds including nitrates, oxalic acid and l-ascorbic acid in spinach formulation with various volume fractions of lemon juice were measured in predigested and digested samples usingThe highest increases in total polyphenols, total flavonoids, total phenolic acids, oxalic acid and nitrate content were noted in predigested and almost all digested spinach samples formulated with the highest volume fraction of lemon juice. In the same sample, the content of individual compounds significantly increased after salivary (l-ascorbic acid), initial (For the first time the stability and digestive availability of spinach polyphenols, oxalic acid, nitrates and l-ascorbic acid were tested with the addition of different volume fractions of lemon juice. The pH of lemon juice and its l-ascorbic acid content increase the stability and availability of polyphenols in spinach lemon juice formulation during

The high demand for healthy food in recent years has led to an increasing need for highly bioactive plant materials. One such plant, Chinese cabbage, possesses flavonoids with antioxidant and antidiabetic properties, but they appear in low quantities. The interspecific transfer of metabolites is a promising technique that could contribute to the increase of the beneficial properties of food. The objective of the study was to determine how an interspecific source-sink phytochemical transfer from donor extracts to the Chinese cabbage affects its phenolic and vitamin C profile and intestinal bioaccessibility, hypoglycemic potential and antioxidant capacity. In addition, sprouts treated with Rosa sp. and Hypericum perforatum extracts showed better antiproliferative effect towards human breast adenocarcinoma cells than untreated sprouts. The results suggest that treatment of plants with donor extracts is a promising approach to improve the nutritional and phytochemical profile and bioactive properties of acceptor plants.

The phytochemical and antioxidant properties of mature (head stage) Chinese cabbage (Brassica rapa ssp. pekinensis) are known; however, data on the phenolic profile, vitamin C (L-ascorbic acid) content and antioxidant capacity of its fresh sprouts are lacking. Since the human consumption of fresh cruciferous sprouts has significantly increased in recent years, their nutritional characterization has become a somewhat urgent matter. Therefore, in this study the contents of total phenolics, flavonols and hydroxycinnamic acids were measured spectrophotometrically, whereas individual flavonoids, phenolic acids and vitamin C were identified and quantified using a newly-developed high performance liquid chromatography method. Also, the antioxidant capacity of five Chinese cabbage sprout growth stages was determined. These stages contained either cotyledons only (seedlings), cotyledons and two leaves, four leaves, six leaves, or ten leaves. Principal component analysis (PCA) and hierarchical clustering (HC) were implemented in order to visualize the classification trend between the stages. Seedlings contained more sinapic acid and vitamin C than older plants. Plants containing six or ten leaves had more ferulic acid and isorhamnetin than younger ones. Total phenolics, flavonols, hydroxycinnamic acids, quercetin and antioxidant capacity did not statistically differ between seedlings and stages with six or ten leaves and their concentrations were significantly higher than in stages with two or four leaves. PCA and HC confirmed the higher phytochemical similarity between seedlings and plants with six or ten leaves than plants with two or four leaves. Therefore, Chinese cabbage seedlings and plants with six or ten leaves should be preferred over plants with two or four leaves, which were ultimately shown to be of lesser nutritional quality. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s13197-019-04103-y) contains supplementary material, which is available to authorized users.

Airborne fly ash and related hazardous particles derived from coal combustion contaminate soil and groundwater, negatively affecting ecosystems. The aim of this study was chemical and toxicological evaluation of aqueous extracts of soil collected from the vicinity of a coal-fired Plomin power plant (PPP), using Lemna (Lemna minor L.) bioassay and additional biochemical indicators - photosynthetic pigments, lipid peroxidation, antioxidative enzymes and glutathione. Topsoil samples were collected from distances of 200, 300, 400 and 800 m from the PPP in accordance with the prevailing SW wind direction. Elevated levels of polycyclic aromatic hydrocarbons (up to 15,765 ng L−1) and potentially toxic trace elements were detected in the Plomin soil extracts (PEs) in comparison to control soil extract (CE). Trace elements accumulated in L. minor were mostly in accordance with their concentrations in PEs. The results demonstrate that PEs induced significant growth inhibition and other phytotoxic effects. Those effects can be related to damage caused by increased production of reactive oxygen species and impaired antioxidant levels. The connection among the phytotoxicity, a distribution of analyzed contaminants, and distances from the PPP is clearly established.

Introduction The phytochemical composition and biological activity of non-volatile components of Centaurea ragusina L. has not been studied previously. Objectives Our aim was to evaluate the phytochemical and bioactive potential (including interactions with polynucleotides) of C. ragusina L. depending on the origin of plant material (in vivo – leaves from natural habitats, ex vitro – leaves from plants acclimated from culture media, in vitro – leaves and calli from plants grown in culture media) and polarity of solvents used in extract preparation (80 and 96% ethanol and water combinations or single solvents). Methodology The polyphenol composition was determined by spectrophotometric and HPLC analysis. Biological activity of extracts was evaluated by following methods: 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS) and 1,1-diphenyl-2-picrylhydrazyl (DPPH) methods for antioxidative activity, 2,3,5-triphenyl tetrazolium chloride (TTC) microdilution method for antibacterial activity, crystal-violet test for cytotoxic activity and thermal denaturation (TD) and circular dichroism (CD) for DNA/RNA interactions. Results Conditions for the most efficient polyphenol extraction were determined: the 80% ethanol/water solvent system was the most suitable for callus and leaf ex vitro samples and 80 or 96% ethanol for leaf in vivo samples. Significantly higher levels of chlorogenic acid and naringenin were detected in callus tissue than in vivo plant. Ethanolic extracts exhibited the significant antibacterial activity against Staphylococcus aureus ATCC 25923. DNA/RNA active compounds in plant extracts were detected by TD and CD methods. Conclusions Callus tissue and ex vitro leaves represent a valuable source of polyphenols as in vivo leaves. TD and CD can be applied for detection of DNA/RNA active compounds in extracts from natural resources. Copyright © 2017 John Wiley & Sons, Ltd.

Salinity is a major abiotic stress negatively affecting plant growth and consequently crop production. The effects of short-term salt stress were evaluated on seedlings of three globally important Brassica crops&mdash, Chinese cabbage (Brassica rapa ssp. pekinensis), white cabbage (Brassica oleracea var. capitata), and kale (Brassica oleracea var. acephala)&mdash, with particular focus on phenolic acids. The physiological and biochemical stress parameters in the seedlings and the levels of three main groups of metabolites (total glucosinolates, carotenoids, and phenolics) and individual phenolic acids were determined. The salt treatments caused a dose-dependent reduction in root growth and biomass and an increase in stress parameters (Na+/K+ ratio, reactive oxygen species (ROS) and glutathione (GSH)) in all seedlings but most prominently in Chinese cabbage. Based on PCA, specific metabolites grouped close to the more tolerant species, white cabbage and kale. The highest levels of phenolic acids, particularly hydroxycinnamic acids, were determined in the more tolerant kale and white cabbage. A reduction in caffeic, salicylic, and 4-coumaric acid was found in Chinese cabbage and kale, and an increase in ferulic acid levels was found in kale upon salinity treatments. Phenolic acids are species-specific among Brassicaceae, and some may participate in stress tolerance. Salt-tolerant varieties have higher levels of some phenolic acids and suffer less from metabolic stress disorders under salinity stress.

Abiotic stressors such as extreme temperatures, drought, flood, light, salt, and heavy metals alter biological diversity and crop production worldwide. Therefore, it is important to know the mechanisms by which plants cope with stress conditions. Polyphenols, which are the largest group of plant-specialized metabolites, are generally recognized as molecules involved in stress protection in plants. This diverse group of metabolites contains various structures, from simple forms consisting of one aromatic ring to more complex ones consisting of large number of polymerized molecules. Consequently, all these molecules, depending on their structure, may show different roles in plant growth, development, and stress protection. In the present review, we aimed to summarize data on how different polyphenol structures influence their biological activity and their roles in abiotic stress responses. We focused our review on phenolic acids, flavonoids, stilbenoids, and lignans.

Owing to its proven toxicity and mutagenicity, arsenic is regarded a principal pollutant in water used for drinking. The objective of this study was the toxicological and chemical evaluation of groundwater samples obtained from arsenic enriched drinking water wells before and after electrochemical and ozone-UV-H2O2- based advanced oxidation processes (EAOP). For this purpose, acute toxicity test with Daphnia magna and chronic toxicity test with Lemna minor L. were employed as well as in vitro bioassays using human peripheral blood lymphocytes (HPBLs). Several oxidative stress parameters were estimated in L.minor. Physicochemical analysis showed that EAOP treatment was highly efficient in arsenic but also in ammonia and organic compound removal from contaminated groundwater. Untreated groundwater caused only slight toxicity to HPBLs and D. magna in acute experiments. However, 7-day exposure of L. minor to raw groundwater elicited genotoxicity, a significant growth inhibition and oxidative stress injury. The observed genotoxicity and toxicity of raw groundwater samples was almost completely eliminated by EAOP treatment. Generally, the results obtained with L. minor were in agreement with those obtained in the chemical analysis suggesting the sensitivity of the model organism in monitoring of arsenic- contaminated groundwater. In parallel to chemical analysis, the implementation of chronic toxicity bioassays in a battery is recommended in the assessment of the toxic and genotoxic potential of such complex mixtures.

Hydra (Cnidaria) is a cosmopolite freshwater invertebrate that establishes symbiosis with unicellular algae. Its simple body is comprised of three layers: epidermis, mesoglea and gastrodermis. The goal of this research was to determine the effect of flavonoids quercetin and naringenin upon hydra – alga symbiosis, to determine how motivated teachers are to perform such a mode of teaching and whether this motivation depends on their age, years of working experience, the school they work in, available time, space and material. An anonymous survey on a sample of 100 biology teachers was used in the research. Hydras and algae were treated with quercetin and naringenin in concentrations of 0.2, 0.25 and 0.3 gL−1 and compared to the control, and Chlorella bioassay was used. Morphological changes included migration, reaction to mechanical stimuli, asexual reproduction, secretion of mucus and mortality. Cyto-histological analysis revealed changes of all three layers and growth inhibition. Naringenin exerted altogether higher impact than quercetin. Survey results indicated that teachers are willing to use this experiment in school, and that most schools have the minimum required equipment. Because of the possibility of adapting this experiment in different temporal and spatial conditions, its performance in school is suggested. Key words: learning; motivation; naringenin; quercetin; teaching. --- Sažetak Hydra (Cnidaria) je kozmopolitski slatkovodni beskralježnjak koji uspostavlja simbiozu s jednostanicnim algama. Njezino jednostavno tijelo sastoji se od tri sloja: epiderma, mezogleje i gastroderma. Cilj ovoga istraživanja bio je utvrditi utjecaj flavonoida kvercetina i naringenina na simbiozu hidre i alge, odrediti koliko su nastavnici motivirani za primjenu takvog nacina poucavanja te ovisi li navedena motivacija o njihovoj dobi, godinama radnog iskustva, skoli u kojoj rade, raspoloživom vremenu, prostoru i materijalima. U istraživanju je upotrijebljena anonimna anketa na uzorku od 100 nastavnika biologije. Hidre i alge tretirane su kvercetinom i naringeninom pri koncentracijama od 0,2, 0,25 i 0,3 gL-1 i uspoređene su s kontrolnim uzorkom, a koristio se bioloski test Chlorella test. Morfoloske promjene ukljucivale su migraciju, reakciju na mehanicke podražaje, nespolno razmnožavanje, izlucivanje sluzi i smrtnost. Citolosko-histoloskom analizom utvrđene su promjene svih triju slojeva, kao i inhibicije rasta. Naringenin je imao ukupno veci ucinak od kvercetina. Rezultati dobiveni anketom pokazali su da su nastavnici spremni izvoditi navedeni eksperiment u skoli te da vecina skola ima minimalnu potrebnu opremu. Zbog mogucnosti prilagodbe opisanog eksperimenta razlicitim vremenskim i prostornim uvjetima, predlaže se njegova primjena u skoli. Kljucne rijeci: ucenje; motivacija; naringenin; kvercetin; nastava. Normal 0 21 false false false HR X-NONE AR-SA

Drought is one of the major abiotic stresses affecting the productivity of Brassica crops. To understand the role of phytohormones in drought tolerance, we subjected Chinese cabbage (B. rapa ssp. pekinensis), white cabbage (B. oleracea var. capitata), and kale (B. oleracea var. acephala) to drought and examined the stress response on the physiological, biochemical and hormonal levels. The phytohormones abscisic acid (ABA), auxin indole-3-acetic acid (IAA), brassinosteroids (BRs), cytokinins (CKs), jasmonates (JAs), and salicylic acid (SA) were analyzed by ultra-high-performance liquid chromatography–tandem mass spectrometry (UHPLC-MS/MS). Based on the physiological and biochemical markers the Chinese cabbage exhibited the lowest tolerance, followed by the white cabbage, while the kale appeared to be the most tolerant to drought. The drought tolerance of the kale correlated with increased levels of SA, ABA, IAA, CKs iP(R) and cZ(R), and typhasterol (TY), a precursor of active BRs. In contrast, the drought sensitivity of the Chinese cabbage correlated with a significant increase in ABA, JAs and the active BRs castasterol (CS) and brassinolide (BL). The moderately tolerant white cabbage, positioned between the kale and Chinese cabbage, showed more similarity in terms of the phytohormone patterns with the kale. We concluded that the drought tolerance in Brassicaceae is mostly determined by the increased endogenous levels of IAA, CKs, ABA and SA and the decreased levels of active BRs.

Hydra (Cnidaria) is a cosmopolite freshwater invertebrate that establishes symbiosis with unicellular algae. Its simple body is comprised of three layers: epidermis, mesoglea and gastrodermis. The goal of this research was to determine the effect of flavonoids quercetin and naringenin upon hydra – alga symbiosis, to determine how motivated teachers are to perform such a mode of teaching and whether this motivation depends on their age, years of working experience, the school they work in, available time, space and material. An anonymous survey on a sample of 100 biology teachers was used in the research. Hydras and algae were treated with quercetin and naringenin in concentrations of 0.2, 0.25 and 0.3 gL−1 and compared to the control, and Chlorella bioassay was used. Morphological changes included migration, reaction to mechanical stimuli, asexual reproduction, secretion of mucus and mortality. Cyto-histological analysis revealed changes of all three layers and growth inhibition. Naringenin exerted altogether higher impact than quercetin. Survey results indicated that teachers are willing to use this experiment in school, and that most schools have the minimum required equipment. Because of the possibility of adapting this experiment in different temporal and spatial conditions, its performance in school is suggested., Hydra (Cnidaria) je kozmopolitski slatkovodni beskralježnjak koji uspostavlja simbiozu s jednostaničnim algama. Njezino jednostavno tijelo sastoji se od tri sloja: epiderma, mezogleje i gastroderma. Cilj ovoga istraživanja bio je utvrditi utjecaj flavonoida kvercetina i naringenina na simbiozu hidre i alge, odrediti koliko su nastavnici motivirani za primjenu takvog načina poučavanja te ovisi li navedena motivacija o njihovoj dobi, godinama radnog iskustva, školi u kojoj rade, raspoloživom vremenu, prostoru i materijalima. U istraživanju je upotrijebljena anonimna anketa na uzorku od 100 nastavnika biologije. Hidre i alge tretirane su kvercetinom i naringeninom pri koncentracijama od 0,2, 0,25 i 0,3 gL-1 i uspoređene su s kontrolnim uzorkom, a koristio se biološki test Chlorella test. Morfološke promjene uključivale su migraciju, reakciju na mehaničke podražaje, nespolno razmnožavanje, izlučivanje sluzi i smrtnost. Citološko-histološkom analizom utvrđene su promjene svih triju slojeva, kao i inhibicije rasta. Naringenin je imao ukupno veći učinak od kvercetina. Rezultati dobiveni anketom pokazali su da su nastavnici spremni izvoditi navedeni eksperiment u školi te da većina škola ima minimalnu potrebnu opremu. Zbog mogućnosti prilagodbe opisanog eksperimenta različitim vremenskim i prostornim uvjetima, predlaže se njegova primjena u školi.

The endemic Croatian species Centaurea ragusina L., like other species from the genus Centaurea, has been traditionally used in Croatia as an antibacterial agent and for the treatment of gastrointestinal and urogenital disorders. In several chromatographic steps, three flavonoids and three sesquiterpene lactones (STLs) were isolated and identified from the most active fractions of the ethanol extract. Two STLs, one for which we created the trivial name ragusinin, and hemistepsin A are here reported for the first time as constituents of the genus Centaurea. All six compounds were screened for their effect on several tumor and one normal cell lines. Among them, ragusinin showed the best bioactivity and high specificity to affect tumor murine SCCVII, human HeLa and Caco-2 cell lines, but not the viability of normal V79 fibroblasts. Due to these characteristics the action of ragusinin was investigated in more detail. Since DNA is the primary target for many drugs with antibacterial and anticancer activity, we studied its interaction with ragusinin. Rather moderate binding affinity to DNA excluded it as the primary target of ragusinin. Due to the possibility of STL interaction with glutathione (GSH), the ubiquitous peptide that traps reactive compounds and other xenobiotics to prevent damage to vital proteins and nucleic acids, its role in deactivation of ragusinin was evaluated. Addition of the GSH precursor N- acetyl-cysteine potentiated the viability of HeLa cells, while the addition of GSH inhibitor L-buthionine sulfoximine decreased it. Moreover, pre-treatment of HeLa cells with the inhibitor of glutathione-S-transferase decreased their viability indicating the detoxifying role of GSH in ragusinin treated cells. Cell death, derived by an accumulation of cells in a G2 phase of the cell cylce, was shown to be independent of poly (ADP-ribose) polymerase and caspase-3 cleavage pointing toward an alternative cell death pathway.

Native Crocus taxa are abundant in regions with moderate and dry climate ; however, their potential in the food industry has thus far been neglected. The objective of this study was to compare the quantitative and qualitative flavonoid content and antioxidant and cytotoxic activity in organs of several populations of Croatian native taxa and assess their potential for application in the food industry. The dominant flavonoids in the native Crocus taxa were kaempferol and quercetin. Tepals contained similar or higher concentrations of flavonoid- glycosides than other organs. Tepals from Cvv1 and Cvv2 populations contained more flavonoid- glycosides than the commonly used spice saffron (C. sativus stigmas). The FRAP antioxidant activity of Cvv1 and Cvv5 tepals was similar to that of standard Trolox. DPPH inhibition of Cvv1 and Cvv3 tepals was within the range of that recorded for saffron. Cvv1 tepals significantly reduced reactive oxygen species in the broadest concentration range (50–1000 μg/ml), and showed considerable antioxidant activity in the ABTS assay, equal to 82% of standard Trolox antioxidant activity. A significantly higher concentration of kaempferol-rutinoside was recorded in this than in other taxa. The flavonoid showed a very strong or strong correlation with antioxidant assays results, and a negative correlation with cellular reactive oxygen species concentration. We therefore presumed that kaempferol- rutinoside is one of the main antioxidant phenolics in Crocus tepals. None of the tested extracts showed cytotoxicity toward Caco-2 cells. The results revealed that Cvv tepals have potential as a food supplement and are a promising material for further food safety tests.

Salinity is one of major abiotic stresses affecting Brassica crop production. Here we present investigations into the physiological, biochemical, and hormonal components of the short-term salinity stress response in Chinese cabbage seedlings, with particular emphasis on the biosynthesis and metabolism of auxin indole-3-acetic acid (IAA). Upon salinity treatments (50- 200 mM NaCl) IAA level was elevated in a dose dependent manner reaching 1.3-fold increase at the most severe salt treatment in comparison to the control. IAA precursor profiling suggested that salinity activated the indole-3-acetamide and indole-3-acetaldoxime biosynthetic pathways while suppressing the indole-3-pyruvic acid pathway. Levels of the IAA catabolites 2-oxoindole-3-acetic acid and indole-3-acetic acid-aspartate increased 1.7- and 2.0-fold, respectively, under the most severe treatment, in parallel with those of IAA. Conversely, levels of the ester conjugate indole-3-acetyl-1-O-s-d-glucose and its catabolite 2-oxoindole-3-acetyl-1-O-s-d-glucose decreased 2.5- and 7.0-fold, respectively. The concentrations of stress hormones including jasmonic acid and jasmonoyl-isoleucine (JA and JA-Ile), salicylic acid (SA) and abscisic acid (ABA) confirmed the stress induced by salt treatment: levels of JA and JA-Ile increased strongly under the mildest treatment, ABA only increased under the most severe treatment, and SA levels decreased dose-dependently. These hormonal changes were related to the observed changes in biochemical stress markers upon salt treatments: reductions in seedling fresh weight and root growth, decreased photosynthesis rate, increased levels of reactive oxygen species, and elevated proline content and the Na+/K+ ratio. Correlations among auxin profile and biochemical stress markers were discussed based on Pearson’s coefficients and principal component analysis (PCA).

Summary Leachates from active and closed municipal solid waste landfills can be a major source of contamination to groundwater and surface waters. In the present study the toxic and genotoxic potential of leachate from an old sanitary landfill prior to and following chemical and electrochemical treatments were assessed using Lemna, Allium, and comet tests. Photosynthetic pigments, malondialdehyde (indicator of lipid peroxidation) and antioxidant enzyme activities were evaluated as additional indicators of toxicity in duckweed. Following duckweed exposure to 25 % dilution of landfill leachate, growth rate and photosynthetic pigments content significantly decreased while lipid peroxidation increased despite stimulation of antioxidative defence mechanisms. Diluted leachate induced DNA strand breaks in duckweed cells as evidenced by the comet assay. Regarding the Allium test, untreated leachate caused inhibition of Allium cepa cell division and induction of mitotic and chromosomal aberrations. Although both water treatments completely reduced genotoxicity of leachate, the electrochemical method was found to be more efficient in removing toxic substances present in landfill leachate and thus more suitable for treating such leachates prior to their discharge into the environment. As landfill leachates pose a risk to human health and environment in general due to their (geno)toxicity, the present study demonstrates that the ecotoxicity/genotoxicity assays should be used in leachate risk assessment together with physicochemical analysis.

Drought is one of the major abiotic stresses affecting the productivity of Brassica crops. To understand the role of phytohormones in drought tolerance, we subjected Chinese cabbage (B. rapa ssp. pekinensis), white cabbage (B. oleracea var. capitata), and kale (B. oleracea var. acephala) to drought and examined the stress response on the physiological, biochemical and hormonal levels. The phytohormones abscisic acid (ABA), auxin indole-3-acetic acid (IAA), brassinosteroids (BRs), cytokinins (CKs), jasmonates (JAs), and salicylic acid (SA) were analyzed by ultra-high-performance liquid chromatography&ndash, tandem mass spectrometry (UHPLC-MS/MS). Based on the physiological and biochemical markers the Chinese cabbage exhibited the lowest tolerance, followed by the white cabbage, while the kale appeared to be the most tolerant to drought. The drought tolerance of the kale correlated with increased levels of SA, ABA, IAA, CKs iP(R) and cZ(R), and typhasterol (TY), a precursor of active BRs. In contrast, the drought sensitivity of the Chinese cabbage correlated with a significant increase in ABA, JAs and the active BRs castasterol (CS) and brassinolide (BL). The moderately tolerant white cabbage, positioned between the kale and Chinese cabbage, showed more similarity in terms of the phytohormone patterns with the kale. We concluded that the drought tolerance in Brassicaceae is mostly determined by the increased endogenous levels of IAA, CKs, ABA and SA and the decreased levels of active BRs.

Screening for mutagens in environmental complex mixtures, such as surface water or industrial wastewater, is gradually being accepted as a routine methodology in monitoring program. In the present work, the simplified A. cepa root assay was employed in order to evaluate the possible cyto- and genotoxic effects of surface and wastewaters collected along or near the Sava River (Croatia) over a 3-month monitoring period. Physicochemical characterization of the water pollution was determined by measuring conductivity, chemical and biological oxygen demand, levels of suspended matter and salts, nitrate, nitrite, ammonium, total nitrogen and total phosphorus. Morphological modifications of the A. cepa roots, inhibition of root growth, cell division and induction of mitotic and chromosomal aberrations were observed. The most polluted water samples (industrial effluents) caused an inhibition of root growth over 50%, a decrease of mitotic index over 40% and considerable increase of chromosomal aberrations in comparison to control. The measured biological effects of some water samples appeared related to the physicochemical characteristics. Therefore, together with the conventional chemical analysis, mutagenicity/genotoxicity assays should be included as complementary parameters in water quality monitoring programs. Their use could permit the quantification of mutagenic hazards in surface and wastewaters.

The genus of Centaurea (Asteraceae) represents an attractive source of bioactive substances [1]. The goal of this work is to determine polyphenol content and antioxidant activity in endemic Croatian plant species Centaurea ragusina L. cultivated in vitro (½MS 2.9 μM GA3+0.5 μM BA and ½MS 2.5 μM IBA) [2] and collected at natural habitats (Katalinic brig - K and Sustipan - S). To clarify biological activity of C. ragusina, interactions of extracts with double stranded polynucleotides (poly A – poly U and ctDNA) were studied. The highest level of total phenols, flavonoids and flavonols was determined in calli ethanol/aqueous extracts cultivated in vitro (2.9 μM GA3 + 0.5 μM BA) while the highest value of hydroxycinnamic acids and proanthocyanidins was detected in ethanol/aqueous extracts of leaves after acclimatization from culture media (½MS 2.5 μM IBA). Significant antioxidant activity measured by DPPH and ABTS methods was observed in almost all extracts with respect to gallic acid as a standard. Extracts of leaves after acclimatization showed the highest stabilization on poly A - poly U (18.36) and ctDNA (5.75) evaluated by changing of melting temperature of the polynucleotide (ΔTm). Results of circular dichroism (CD spectroscopy) indicate significant impact of all tested extracts on the conformation of both polynucleotides. The results obtained suggest that the plant species C. ragusina grown in vitro can be efficiently used as a potential source of polyphenols and antioxidants in food, pharmacological and cosmetics industry as it is equally rich with phytochemicals as its wild grown counterparts.

In the present study toxicity of Frangula alnus Mill. bark, widely used as laxative, was investigated. Human peripheral blood lymphocytes (HPBLs) were treated with F. alnus bark extract or emodin (emodin is bark component with laxative property), and cytotoxicity, genotoxicity and parameters of oxidative stress were assessed. Also, polyphenol content of bark extract and antioxidant activity of the extract and emodin measured by DPPH, ABTS and FRAP methods were examined. The bark extract (500 μg/ml) produced cell death and DNA damage, while level of ROS changed at 250 μg/ml. Emodin induced cell death and DNA damage at 150 μg/ml and 200 μg/ml, respectively, and the increase of ROS was observed at 25 μg/ml. These results suggest that both, bark extract and emodin, are cyto/genotoxic to HPBLs and that oxidative stress is involved in the mechanism of their toxicity. The results on antioxidant activity showed that, unlike emodin, bark extract possess moderate antioxidant capacity (44.6%, 46.8% and 2.25 mmol Fe(2+)/g measured by DPPH, ABTS and FRAP assay, respectively) that can be related to relatively high phenolic content (116.07 mg/g). However, due to toxicological properties use of F. alnus bark as well as emodin-containing preparations should be taken with caution.

Leachates from active and closed municipal solid waste landfills can be a major source of contamination to groundwater and surface waters. In the present study the toxic and genotoxic potential of leachate from an old sanitary landfill prior to and following chemical and electrochemical treatments were assessed using Lemna, Allium, and comet tests. Photosynthetic pigments, malondialdehyde (indicator of lipid peroxidation) and antioxidant enzyme activities were evaluated as additional indicators of toxicity in duckweed. Following duckweed exposure to 25 % dilution of landfill leachate, growth rate and photosynthetic pigments content significantly decreased while lipid peroxidation increased despite stimulation of antioxidative defence mechanisms. Diluted leachate induced DNA strand breaks in duckweed cells as evidenced by the comet assay. Regarding the Allium test, untreated leachate caused inhibition of Allium cepa cell division and induction of mitotic and chromosomal aberrations. Although both water treatments completely reduced genotoxicity of leachate, the electrochemical method was found to be more efficient in removing toxic substances present in landfill leachate and thus more suitable for treating such leachates prior to their discharge into the environment. As landfill leachates pose a risk to human health and environment in general due to their (geno)toxicity, the present study demonstrates that the ecotoxicity/genotoxicity assays should be used in leachate risk assessment together with physicochemical analysis., Procjedne vode s aktivnih i zatvorenih odlagališta krupnog otpada mogu biti glavni izvor onečišćenja podzemnih i površinskih voda. U ovom su radu putem Lemna-, Allium- i komet-testa procijenjeni toksični i genotoksični učinci procjedne vode prikupljene sa starog odlagališta otpada Piškornica (Koprivnica) prije i nakon kemijskog ili elektrokemijskog pročišćavanja. Dodatni pokazatelji toksičnosti praćeni u vodenoj leći bili su sadržaj fotosintetskih pigmenata i malondialdehida (indikator lipidne peroksidacije) te aktivnosti antioksidacijskih enzima. U toj je biljci 25 %-tno razrjeđenje procjedne vode uzrokovalo smanjenje rasta i sadržaja pigmenata dok je lipidna peroksidacija porasla unatoč stimulaciji antioksidacijskih obrambenih mehanizama. Komet testom je utvrđeno oštećenje molekule DNA u stanicama vodene leće. Glede Alium testa, nepročišćena procjedna voda uzrokovala je inhibiciju diobe u luku (Allium cepa) te pojavu mitotskih i kromosomskih aberacija. Iako su obje metode pročišćavanja bile vrlo učinkovite u uklanjanju genotoksičnih tvari prisutnih u odlagališnom eluatu, elektrokemijska metoda pokazala se učinkovitija u uklanjanju toksičnih tvari iz eluata, te time i pogodnija za obradu takvih vrsta eluata prije njihova ispuštanja u okoliš. Kako su odlagališni eluati zbog svoje genotoksičnosti opasni za ljudsko zdravlje i okoliš općenito, rezultati ovog istraživanja pokazuju da bi se testovi toksičnosti i genotoksičnosti trebali upotrebljavati u procjeni rizika odlagališnih eluata, zajedno s fizikalno-kemijskom analizom.

Water deficit in the soil leads to osmotic stress in plants. The type of stress affects plant water relations, osmolyte accumulation and oxidative stress balance. The present study aimed to investigate the effects of osmotic stress on the Croatian perennial species Fibigia triquetra (DC.) Boiss, adapted to a hot and dry habitat. Plants grown in culture conditions were subjected to isoosmotic concentrations of mannitol and polyethylene glycol (PEG) and certain physiological and oxidative stress parameters were analyzed during a period of 14 days. Dry weight and proline content in Fibigia triquetra shoots increased in response to osmotic stress while the relative water content decreased. After an initial rise, chlorophyll and carotenoid levels in treated plants dropped to untreated plant levels. Oxidative damage to proteins and especially to lipids was evident upon PEG-induced osmotic stress. Superoxide dismutase and ascorbate peroxidase appear to play an essential protective role in stressed plants. Regardless of the osmotic agent, accumulation of heatshock proteins of 70 kDa was noticed under osmotic stress. The tolerance of the plant species to osmotic stress seems to be associated with increased capacity of the antioxidative system and efficient photoprotective system. Normal 0 21 false false false HR X-NONE X-NONE /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Table Normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-qformat:yes; mso-style-parent:""; mso-padding-alt:0cm 5.4pt 0cm 5.4pt; mso-para-margin-top:0cm; mso-para-margin-right:0cm; mso-para-margin-bottom:10.0pt; mso-para-margin-left:0cm; line-height:115%; mso-pagination:widow-orphan; font-size:11.0pt; font-family:"Calibri","sans-serif"; mso-ascii-font-family:Calibri; mso-ascii-theme-font:minor-latin; mso-fareast-font-family:"Times New Roman"; mso-fareast-theme-font:minor-fareast; mso-hansi-font-family:Calibri; mso-hansi-theme-font:minor-latin;}

Acid mine drainage (AMD) is a by-product of the mining industry that has a detrimental effect on aquatic plant and animal life due to high load of heavy metals and sulfates. In the present study, the toxic and genotoxic potential of AMD prior to and following combination of neutralization/electrocoagulation processes was evaluated using several bioassays and selected parameters. Regardless of pH correction of AMD prior to Daphnia bioassay, high acute toxicity was observed in Daphnia magna. The mine leachate also induced strong phyto-, cyto- and genotoxicity to Allium cepa roots. Short term exposure to AMD inhibited duckweed growth and chlorophyll a content and simultaneously promoted lipid peroxidation and DNA damage despite duckweed capability to upregulate antioxidative defense mechanisms. The results show that observed (geno)toxicity could be related to oxidative stress most probably induced by toxic metal action. However, influence of low pH as a contributing factor in the phytotoxicity of AMD cannot be excluded. The application of combined treatment eliminated genotoxicity and was highly efficient in reducing toxicity of AMD. Thus, the method seems to be suitable for treatment of AMD waters enabling their safe discharge to an aquatic environment.

The genotoxic and toxic potential of polluted surface water exposed to a fertilizer factory effluent was evaluated using assays with fish (Cyprinus carpio) and plant (Lemna minor) model organisms. Beside classical physicochemical parameters, the contents of fluorides, some heavy metals and polycyclic aromatic hydrocarbons were analyzed as well. Surface water caused inhibition of plant growth and decrease of photosynthetic pigment content. Regarding DNA damage and oxidative stress parameters, both fish and plants showed similar response to the surface water. In confirmation to biochemical markers, histopathological analysis of gill and liver tissues revealed a higher incidence of lesions in fish exposed to polluted surface water. Generally, results obtained by biological monitoring were mostly in agreement with chemical analysis of the surface water, although several discrepancies were observed which might be due to difference in sensitivity of model organisms or in experimental conditions (laboratory and field exposure). The results imply that conventional chemical analysis should be extended to genotoxicity /toxicity assays as measured biological effects and the potential health hazard cannot be predicted based on the physicochemical characteristics of water samples alone.

Background and purpose: Beside being rich with sweet glycosides, Stevia rebaudiana Bertoni is an interesting source of flavonoids and phenols. The current study aimed to assess the potential for increasing total phenols and flavonoids in S. rebaudiana tissue by varying concentrations of plant growth regulators (PGR) and pH levels of media. Materials and methods: The culture was established from seeds and propagated shoots were cultured on media of different pH levels (4.6, 5.8 and 7.4) and PGR. Total polyphenolics and free radical scavenging capability of leaf, callus and root extracts were determined. Shoot height, root length, shoot and root number were also recorded. Results: Shoot elongation and root development was stimulated by singly applied growth regulators though the values of both parameters were also satisfactory in PGR-free media . In the latter media, pH value of 4.6 was a main factor for increasing leaf metabolite levels. A most significant rise in phenols and flavonoids was evident in response to combination of BA either with GA3 or IAA compared to singly applied regulators indicating synergistic effects of PGR (especially of auxins and cytokinins). However, polyphenolics levels and their distribution between different tissues were also influenced by medium pH value. A positive correlation was found between antioxidant activity of S. rebaudiana extracts and phenols and flavonoids. Conclusions: The results show that PGR and medium pH value strongly affect accumulation of secondary metabolites and can lead to significant enhancement in productivity of bioactive polyphenolics in S. rebaudiana plant cultures. Keywords: Stevia, phenols, flavonoids, plant hormones, antioxidant