Search

Introduction Ramonda serbica and R. nathaliae are resurrection plants that have the remarkable ability to survive the complete desiccation of their vegetative organs (i.e. leaves, stem, roots) during periods of drought and rapidly revive when rewatered and rehydrated. Objective To investigate metabolic changes in R. serbica and R. nathaliae during their desiccation and recovery process Methods Proton nuclear magnetic resonance (1H-NMR) and gas chromatography–mass spectrometry (GC–MS)-based metabolomics approach coupled with multivariate data analysis was utilised to identify the metabolomes of the plants from 90 biological replicates. Results Sucrose and the polyphenolic glycoside myconoside were predominant in almost equal amounts in all samples studied, regardless of their water content at sampling. During the dehydration process, a decrease in the relative content of fructose, galactose, and galactinol was observed while the contents of those metabolites were preserved in the partially rehydrated plants. Raffinose and myo-inositol were accumulated in dry samples. Conclusion Using 1H-NMR and GC–MS as two complementary analytical platforms provided a more complete picture of the metabolite composition for investigation of the desiccation and recovery process in resurrection plants.

We examined the effect of drought-tolerant Bacillus safensis SS-2.7 and B. thuringiensis SS-29.2 strains on the response of four (133, 274, California Wonder—CalW, and Matica) sweet pepper genotypes to water deficiency conditions. Pepper seeds were sown in pots with (treated) and without (control) bacterial strain inoculation. After four weeks of growth under controlled conditions and regular watering, drought was imposed by completely withholding watering for seven days. Under conditions of normal watering, genotype 274 showed better seedling establishment than genotype 133 and CalW, while the slowest was genotype Matica. Antioxidant enzyme activity under drought conditions was genotype and bacterial treatment-dependent. The best response to bacterial treatment in order to cope with severe drought was found in the CalW genotype, while in genotype 133, we determined even faster plant decay during water deficiency in treated seeds. Inoculated seeds of the Matica genotype did not show different antioxidant enzyme activity under normal and drought conditions. According to the obtained results, we concluded that under drought conditions, the most susceptible was genotype 274, moderate susceptibility was detected in genotype 133, and CalW and Matica were the most tolerant genotypes. Our study demonstrates (1) that drought-tolerant Bacillus strains showed a plant growth-promoting effect on some selected pepper genotypes; (2) that there were genotype-dependent antioxidant enzyme activities under drought conditions in response to treatment with a particular bacterial strain; and (3) that we could expect a genotype-dependent response during biostimulant application, especially under stress conditions.

In this study, for the first time, the comprehensive analysis of antiproliferative and antioxidant activities of ramson, followed by the analysis of its associated microbiota and health-promoting effects of lactic acid bacteria (LAB), was performed. Ramson (Allium ursinum) is recognized as a medicinal plant with a long history of use in traditional medicine due to its antimicrobial and antioxidant activity. In this study the influence of in vitro gastrointestinal digestion on the cytotoxic activity of A. ursinum extracts against human malignant cell lines was demonstrated. Seven sulfur compounds, the degradation products of thiosulfinates, including diallyl disulfide were shown to inhibit proliferation of malignant cells by inducing accumulation within G2/M phase as well as to induce apoptosis through activation of caspase-3 and mitochondrial signaling pathway. Further, the A. ursinum microbiota, particularly LAB with potential probiotic effects, was analyzed by culture-dependent method and culture-independent method [denaturing gradient gel electrophoresis (DGGE)]. The obtained results revealed that the most abundant genera were Streptococcus, Lactobacillus, and Bacillus. The Lactobacillus genus was mainly represented by L. fermentum. The pulsed-field gel electrophoresis (PFGE) analysis revealed the presence of two PFGE pulsotypes. The probiotic potential of the strain L. fermentum BGSR163 belonging to PFGE pulsotype 1 and the strain L. fermentum BGSR227 belonging to the PFGE pulsotype 2 was characterized. The results revealed that both strains are safe for human use, successfully survive the simulated gastrointestinal conditions, have potential to transiently colonize the gastrointestinal tract (GIT) and have a protective immunomodulatory effect, inducing the production of proinflammatory cytokine IL17 and regulatory cytokine IL10, while decreasing the production of proinflammatory cytokine IFN-γ. In conclusion, the results of this study suggest that consumption o

In this study, for the first time, the comprehensive analysis of antiproliferative and antioxidant activities of ramson, followed by the analysis of its associated microbiota and health-promoting effects of lactic acid bacteria (LAB), was performed. Ramson (Allium ursinum) is recognized as a medicinal plant with a long history of use in traditional medicine due to its antimicrobial and antioxidant activity. In this study the influence of in vitro gastrointestinal digestion on the cytotoxic activity of A. ursinum extracts against human malignant cell lines was demonstrated. Seven sulfur compounds, the degradation products of thiosulfinates, including diallyl disulfide were shown to inhibit proliferation of malignant cells by inducing accumulation within G2/M phase as well as to induce apoptosis through activation of caspase-3 and mitochondrial signaling pathway. Further, the A. ursinum microbiota, particularly LAB with potential probiotic effects, was analyzed by culture-dependent method and culture-independent method [denaturing gradient gel electrophoresis (DGGE)]. The obtained results revealed that the most abundant genera were Streptococcus, Lactobacillus, and Bacillus. The Lactobacillus genus was mainly represented by L. fermentum. The pulsed-field gel electrophoresis (PFGE) analysis revealed the presence of two PFGE pulsotypes. The probiotic potential of the strain L. fermentum BGSR163 belonging to PFGE pulsotype 1 and the strain L. fermentum BGSR227 belonging to the PFGE pulsotype 2 was characterized. The results revealed that both strains are safe for human use, successfully survive the simulated gastrointestinal conditions, have potential to transiently colonize the gastrointestinal tract (GIT) and have a protective immunomodulatory effect, inducing the production of proinflammatory cytokine IL17 and regulatory cytokine IL10, while decreasing the production of proinflammatory cytokine IFN-gamma. In conclusion, the results of this study suggest that consumpti

Interactions between plants and herbivorous insects can be shaped by light environment, resulting in differential herbivory in sun and shade. In dioecious species, the combination of plant sex and light-induced changes in defense traits and nutritive value can alter the patterns of foliar damage. We explored the combined effects of light environment and plant sex on natural herbivore damage and plant traits in the dioecious understory forb Mercurialis perennis on Mt. Kopaonik (Serbia). The role of plant traits in predicting the extent of damage was examined as well. Male and female plants from contrasting light environments (shade vs. sun) were analyzed with respect to leaf damage, as well as plant morphological and biochemical traits (size, specific leaf area, carbon-based defensive compounds and nutritional quality). We found prominent differences in herbivore damage between sun and shade conditions, but not between the sexes. Plants from the sun-exposed site had a significantly larger leaf area removed. The specific leaf area co-varied with herbivore damage in an inverse fashion, while leaf nutritional value had a moderate effect. Contrasting light conditions influenced the patterns of intersexual differences in the contents of condensed tannins and soluble proteins, with females exhibiting higher trait values. We found that factors defining risk of consumption were related to plant morphological traits and nutritive value rather than to chemical defenses.

Eight 1,3,4-oxadiazole derivatives containing phenolic acid moieties (7a-h) and eight of their diacylhydrazine precursors (6a-h) were synthesized, characterized using spectroscopic methods and examined by scavenging of stable DPPH (2,2-diphenyl-1-picrylhydrazyl) radicals. The most potent phenolic 1,3,4-oxadiazoles showed better DPPH scavenging activity in comparison with their corresponding diacylhydrazine precursors as a result of participation of both aromatic rings and a 1,3,4-oxadiazole moiety in resonance stabilization of the formed phenoxyl radical. Four diacylhydrazines (6d, 6e, 6g, and 6h) and four 1,3,4-oxadiazoles (7d, 7e, 7g and 7h) with the best DPPH scavenging activity, were chosen for further evaluation of their antioxidant potential through various assays. The investigated compounds exerted pronounced ABTS radical scavenging capacity, moderate to good H2O2 scavenging properties and strong ferric ion reducing capacity. Further in vitro evaluation of the antioxidant properties of the most active compounds demonstrated their protective effects in normal lung fibroblasts MRC-5 against hydrogen peroxide induced oxidative stress. Diacylhydrazine 6h increased two times the activity of glutathione peroxidase in treated cells in comparison with a control sample and did not affect the superoxide dismutase activity.

MicroRNAs (miRNAs), recently recognized as important regulator of gene expression at posttranscriptional level, have been found to be involved in plant stress responses. The observation that some miRNAs are up- or down regulated by stress implies that that could play vital roles in plant resistance to abiotic and biotic stress. We investigated the effect of water stress treatment during 10 days on expression of conserved miRNAs-miR398a/b and miR408 in pea plants. This time frame could reflect the changes as closely as possible those during water stress cause visible effects under field condition. It was observed that dehydration strongly down regulates the expression of both miR398a/b and miR408 in pea roots and shoots. The down-regulation of miR398a/b and the up-regulation of potential target genes copper superoxide dismutase, CSD1, highlight the involvement of this miRNA in pea stress response. On the contrary, the mRNA level of cytochrome c oxidase subunit 5 (COX5b) was not changed in roots and shoots of water-stressed plants, compared to control-well hydrated plants. This suggests that COX5b is not target of miR398, or that its expression is regulated by some other mechanism. P1B-ATPase expression increased during water deficit only in shoots of pea; in the roots there was no changes in expression. Our results might help to understand the possible role of investigated miRNAs and their contribution to pea capacity to cope with water deficit.

The extracts were prepared from the samples of 3,200-year-old charred pea and bitter vetch seeds from the site of Hissar near Leskovac, South Serbia, using two different DNA extraction procedures. We used CTAB method with some modification and obtained low quantity of ancient DNA in comparison with the second method used - commercial available kit. After the extraction, a whole genome amplification using Phi29 DNA polymerase was performed. The amplified DNAs were used for PCR reaction using primers for 26S rDNA gene, which is located on the nuclear genome. The single band corresponding to 26S rDNA fragment from modern relatives was obtained. We conclude that DNA from charred pea and vetch seed can be extracted and used for further archaeobotanical analysis at the molecular level., Primenom dve različite procedure ekstrahovana je drevna DNK iz ugljenisanih semena graška i urova starih 3200 godina sa lokaliteta Hisar kod Leskovca. Korišćena je modifikovana CTAB metoda i dobijena je relativno mala količina drevne DNK u poređenju sa drugom primenjenom metodom - komercijalno dostupnim kitom za izolaciju DNK. Nakon ekstrakcije primenjeno je umnožavanje celog genoma pomoću Phi29 DNK polimeraze. Umnožena DNK je korišćena za PCR reakciju primenom prajmera za 26S rDNK gen, koji je lociran u jedarnom genomu. Dobijen je fragment iste veličine kao i PCR fragment 26S rDNK savremenih srodnika graška i urova. Može se zaključiti da je moguće ekstrahovati drevnu DNK iz ugljenisanih semena graška i urova i koristiti je za dalje arheobotaničke analize na molekularnom nivou.

To date little has been done on identification of major phenolic compounds responsible for anticancer and antioxidant properties of pea (Pisum sativum L.) seed coat extracts. In the present study, phenolic profile of the seed coat extracts from 10 differently colored European varieties has been determined using ultrahigh-performance liquid chromatography-linear trap quadrupole orbitrap mass spectrometer technique. Extracts of dark colored varieties with high total phenolic content (up to 46.56mg GAE/g) exhibited strong antioxidant activities (measured by 2,2-diphenyl-1-picrylhydrazyl or DPPH assay, and ferric ion reducing and ferrous ion chelating capacity assays) which could be attributed to presence of gallic acid, epigallocatechin, naringenin, and apigenin. The aqueous extracts of dark colored varieties exert concentration-dependent cytotoxic effects on all tested malignant cell lines (human colon adenocarcinoma LS174, human breast carcinoma MDA-MB-453, human lung carcinoma A594, and myelogenous leukemia K562). Correlation analysis revealed that intensities of cytotoxic activity of the extracts strongly correlated with contents of epigallocatechin and luteolin. Cell cycle analysis on LS174 cells in the presence of caspase-3 inhibitor points out that extracts may activate other cell death modalities besides caspase-3-dependent apoptosis. The study provides evidence that seed coat extracts of dark colored pea varieties might be used as potential cancer-chemopreventive and complementary agents in cancer therapy.

To date little has been done on identification of major phenolic compounds responsible for anticancer and antioxidant properties of pea (Pisum sativum L.) seed coat extracts. In the present study, phenolic profile of the seed coat extracts from 10 differently colored European varieties has been determined using ultrahigh-performance liquid chromatography-linear trap quadrupole orbitrap mass spectrometer technique. Extracts of dark colored varieties with high total phenolic content (up to 46.56mg GAE/g) exhibited strong antioxidant activities (measured by 2,2-diphenyl-1-picrylhydrazyl or DPPH assay, and ferric ion reducing and ferrous ion chelating capacity assays) which could be attributed to presence of gallic acid, epigallocatechin, naringenin, and apigenin. The aqueous extracts of dark colored varieties exert concentration-dependent cytotoxic effects on all tested malignant cell lines (human colon adenocarcinoma LS174, human breast carcinoma MDA-MB-453, human lung carcinoma A594, and myelogenous leukemia K562). Correlation analysis revealed that intensities of cytotoxic activity of the extracts strongly correlated with contents of epigallocatechin and luteolin. Cell cycle analysis on LS174 cells in the presence of caspase-3 inhibitor points out that extracts may activate other cell death modalities besides caspase-3-dependent apoptosis. The study provides evidence that seed coat extracts of dark colored pea varieties might be used as potential cancer-chemopreventive and complementary agents in cancer therapy.

Porodica mahunarki (Fabaceae) obiluje ekonomski i poljoprivredno značajnim vrstama među kojima se po zastupljenosti gajenja u Srbiji izdvaja grašak (Pisum sativum L.) sa udelom od 4,5% u ukupnoj povrtarskoj proizvodnji. Semenjače graška, posebno genotipova sa izrazito pigmentisanim tamnim semenima predstavljaju značajan izvor fenolnih jedinjenja čiji su sastav i biološke aktivnosti nedovoljno ispitani, te stoga ni potencijal ovog materijala nije iskorišćen na odgovarajući način. Semenjače graška čine i do 10% suve mase semena i uglavnom predstavljaju otpadni materijal koji nastaje u procesu industrijske izolacije proteina semena koji se koriste u humanoj i animalnoj ishrani. U ovom radu je ispitivan sadržaj fenolnih jedinjenja prisutnih u semenjačama deset evropskih genotipova graška kao i njihove biološke aktivnosti. Pored toga je ispitivan i uticaj različitih rastvarača, termičke obrade i in vitro digestije semena graška na prinos fenolnih jedinjenja i antioksidativne aktivnosti dobijenih ekstrakata. Dobijeni rezultati su pokazali da termički tretman i in vitro digestija dovode do statistički značajnog povećanja sadržaja fenolnih jedinjenja, kako u vodenim ekstraktima semena graška tako i u ekstraktima pripremljenim pomoću organskog rastvarača. Utvrđeno je da peptidi koji nastaju u toku in vitro digestije proteina semena graška mogu značajno interferirati sa testovima za određivanje antioksidativne aktivnosti, pogotovo u vodenim rastvorima, pošto i sami poseduju detektabilne antioksidativne aktivnosti. Pokazano je da kombinacija rastvarača koja sadrži 70% acetona, 29% destilovane vode i 1% sirćetne kiseline najpodesnije sredstvo za ekstrakciju ukupnih fenola iz semenjače graška, koje daje najveći prinos kako kod genotipova sa bledom, tako i kod genotipova sa izrazito tamno pigmentisanom semenjačom. Sadržaj ukupnih fenola u ekstraktima dobijenim primenom pomenutog rastvarača se kod pojedinih genotipova (Assas, MBK 168 и Aslaug) kretao u opsegu 41,8-46,6 mg GAE/ g s, The family of legumes (Fabaceae) is rich in economicaly and agriculturally important species among which the most abundant in Serbia stands peas (Pisum sativum L.) with a share of 4.5% of the total vegetable production. Pea seed coats, especially in genotypes with highly pigmented dark seeds are an important source of phenolic compounds, whose composition and biological activity have not been tested in detail, thus this valuable material is still underutilized. Pea seed coat makes up to 10% of the seed dry weight and largely represents waste material generated in the process of industrial isolation of seed proteins, which are used in human and animal nutrition. In this study the content of phenolic compounds in seeds of ten European pea genotypes, as well as their biological activities has been examined. In addition, the effect of different solvents, heat treatment and in vitro digestion of pea seeds on yield of phenolic compounds and antioxidant activity in extracts was also studied. The obtained results showed that the thermal treatment and in vitro digestion led to a statistically significant increase of the total phenolic content, both in aqueous extracts of pea seeds and in the extracts prepared by using an organic solvent. It was shown that peptides which were formed during the in vitro digestion of the pea seed proteins can significantly interfere with the assays for determination of antioxidant activity, especially in aqueous solutions, since they possess a detectable antioxidant activity. It has been shown that the combination of solvents containing 70% acetone, 29% distilled water and 1% acetic acid was the most suitable means for the extraction of total phenols from pea seed coats, which gave the highest yield in both bright colored and dark colored seed genotypes. Total phenolic content of extracts obtained by mentioned solvent in certain genotypes (Assas, MBK 168 and Aslaug) ranged from 41.8 to 46.6 mg GAE / g s.m., which are the highest recorded values

The phenolic composition of seed coats in four differently colored pea varieties (Pisum sativum L.) was investigated using UHPLC-LTQ OrbiTrap MS. The obtained findings revealed that the seed coats of the examined pea genotypes possess a unique phenolic composition compared to previously studied European cultivars. In total, 41 phenolic compounds have been identified. The seed coats of the studied cultivars contained certain amounts of rosmarinic acid, rutin, galangin, morin, naringin, hesperetin and pinocembrin as well as ten flavonol glycosides that had not been reported previously. Additionally, the total phenolic content, antioxidant activity and metal chelating capacity of extracts was determined using Folin-Ciocalteu's method, 2,2-diphenyl-1-picrylhydrazyl radical scavenging assay, ferric ion-reducing capacity and ferrous ion-chelating capacity assay, respectively. Dark colored genotypes MBK 168 and MBK 173 possessed the highest total phenolic contents as well the strongest antioxidant activities. On the other hand, bright colored genotypes MBK 88 and MBK 90 exhibited the strongest metal-chelating capacities. The examined pea seed coats may be considered as important potential contributors to human health due to the presence of bioactive phenolic constituents. In addition, our results could be used as a guideline for breeding new pea cultivars with high antioxidant activities applicable in the formulation of functional food products.

Nine Lactobacillus strains known for surface proteinase activity were chosen from our collection and tested for their ability to grow in pea seed protein-based medium, and to hydrolyze purified pea proteins in order to produce peptides with antioxidant (AO) activity. Two strains, Lactobacillus rhamnosus BGT10 and Lactobacillus zeae LMG17315, exhibited strong proteolytic activity against pea proteins. The AO activity of the pea hydrolysate fraction, MW lt 10 kDa, obtained by the fermentation of purified pea proteins with Lactobacillus rhamnosus BGT10, was tested by standard spectrophotometric assays (DPPH, ABTS, Fe3+-reducing capacity) and the recently developed direct current (DC) polarographic assay. The low molecular weight fraction of the obtained hydrolysate was separated using ion exchange chromatography, while the AO activity of eluted fractions was determined by means of a sensitive DC polarographic assay without previous concentration of samples. Results revealed that the fraction present in low abundance that contained basic peptides possessed the highest antioxidant activity. Based on the obtained results, it can be concluded that Lactobacillus rhamnosus BGT10 should be further investigated as a candidate strain for large-scale production of bioactive peptides from legume proteins.

Although more than 400 microsatellite loci are currently available for Vicia faba L. (faba bean), an important food and feed grain crop legume, they have not yet been used for comprehensive molecular characterization of this crop. We report a three-step procedure for rapid and cost-effective delineation and utilization of informative genomic nuclear SSRs for paralleled genotyping in faba bean suitable also for other species: (i) pre-selection of loci generating PCR products of expected lengths which are potentially polymorphic (achieved by PCR amplification in bulked samples); (ii) exclusion of loci burdened with persistent null alleles and multilocus amplification products (based on PCR amplification of pre-selected loci in individual genotypes), and (iii) multiplexing. We demonstrate also that genomic SSRs are promising molecular tools for molecular characterization of faba bean required also for crop improvement.

Nine Lactobacillus strains known for surface proteinase activity were chosen from our collection and tested for their ability to grow in pea seed protein-based medium, and to hydrolyze purified pea proteins in order to produce peptides with antioxidant (AO) activity. Two strains, Lactobacillus rhamnosus BGT10 and Lactobacillus zeae LMG17315, exhibited strong proteolytic activity against pea proteins. The AO activity of the pea hydrolysate fraction, MW lt 10 kDa, obtained by the fermentation of purified pea proteins with Lactobacillus rhamnosus BGT10, was tested by standard spectrophotometric assays (DPPH, ABTS, Fe3+-reducing capacity) and the recently developed direct current (DC) polarographic assay. The low molecular weight fraction of the obtained hydrolysate was separated using ion exchange chromatography, while the AO activity of eluted fractions was determined by means of a sensitive DC polarographic assay without previous concentration of samples. Results revealed that the fraction present in low abundance that contained basic peptides possessed the highest antioxidant activity. Based on the obtained results, it can be concluded that Lactobacillus rhamnosus BGT10 should be further investigated as a candidate strain for large-scale production of bioactive peptides from legume proteins.

The phenolic composition of seed coats in four differently colored pea varieties (Pisum sativum L.) was investigated using UHPLC-LTQ OrbiTrap MS. The obtained findings revealed that the seed coats of the examined pea genotypes possess a unique phenolic composition compared to previously studied European cultivars. In total, 41 phenolic compounds have been identified. The seed coats of the studied cultivars contained certain amounts of rosmarinic acid, rutin, galangin, morin, naringin, hesperetin and pinocembrin as well as ten flavonol glycosides that had not been reported previously. Additionally, the total phenolic content, antioxidant activity and metal chelating capacity of extracts was determined using Folin-Ciocalteu's method, 2,2-diphenyl-1-picrylhydrazyl radical scavenging assay, ferric ion-reducing capacity and ferrous ion-chelating capacity assay, respectively. Dark colored genotypes MBK 168 and MBK 173 possessed the highest total phenolic contents as well the strongest antioxidant activities. On the other hand, bright colored genotypes MBK 88 and MBK 90 exhibited the strongest metal-chelating capacities. The examined pea seed coats may be considered as important potential contributors to human health due to the presence of bioactive phenolic constituents. In addition, our results could be used as a guideline for breeding new pea cultivars with high antioxidant activities applicable in the formulation of functional food products.

Bitter vetch (Vicia ervilia (L.) Willd.) was a part of the everyday diet of the Eurasian Neanderthal population and the modern human Palaeolithic hunter-gatherers at the end of the last Ice Age. The major criteria to determine the domestication in bitter vetch and other ancient grain legumes are non-dehiscent pods, larger seed size and smooth seed testa. Bitter vetch seeds were found among the earliest findings of cultivated crops at the site of Tell El-Kerkh, Syria, from 10th millennium BP. Along with cereals, pea and lentil, bitter vetch has become definitely associated with the start of the 'agricultural revolution' in the Old World. Bitter vetch entered Europe in its south-east regions and progressed into its interior via Danube. Its distribution was rapid, since the available evidence reveals its presence in remote places at similar periods. Recently the first success has been obtained in the extraction of ancient DNA from charred bitter vetch seeds. The linguistic evidence supports the fact that most of Eurasian peoples have their own words denoting bitter vetch, meaning that its cultivation preceded the diversification of their own proto-languages.