Characterization of Antioxidant and Antimicrobial Activity and Phenolic Compound Profile of Extracts from Seeds of Different Vitis Species.

Seeds of Vitis vinifera L. with a high content of bioactive compounds are valuable by-products from grape processing. However, little is known about the bioactivity of seeds from other Vitis species. The aim of this study has been to compare the phenolic composition, antimicrobial activity, and antioxidant activity of extracts from seeds of four Vitis species (V. riparia Michx., V. californica Benth., V. amurensis Rupr., and V. vinifera L.). Antioxidant activities were assessed as ferric-reducing antioxidant power (FRAP), 2,2-diphenyl-1-picrylhydrazyl radical (DPPH•) scavenging activity, and oxygen radical absorbance capacity (ORAC). The antimicrobial activity was determined using the microdilution method against some Gram-negative (Escherichia coli, Salmonella enterica ser. Typhimurium, and Enterobacter aerogenes) and Gram-positive (Enterococcus faecalis and Staphylococcus aureus) bacteria. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) was used to evaluate the phenolic profile of extracts. Flavan-3-ols, procyanidins, phenolic acids, flavonols, anthocyanins, and stilbenoids were detected. (+)-Catechin and (-)-epicatechin turned out to be the most abundant in the phenolic profile of V. amurensis seed extract. Phenolic acids prevailed in the extract from V. vinifera seeds. The V. riparia and V. californica seed extracts had higher contents of most individual phenolics compared to the other Vitis species. They also showed a higher total phenolic content, DPPH• scavenging activity, ORAC, and overall antibacterial activity. Total phenolic content significantly correlated with antioxidant activity and antimicrobial activity against E. coli. The principal component analysis (PCA) showed discrimination between V. vinifera, V. amurensis, and clustered V. riparia and V. californica with respect to variables. To recapitulate, this research demonstrates that seeds of different Vitis species, especially V. riparia and V. californica, are sources of molecules with antioxidant and antimicrobial activities that can be used in different sectors, such as in the food, cosmetic, and pharmaceutical industries.

Changes in the proteome of grapevine leaves (Vitis vinifera L.) during long-term drought stress.

The essence of exploring and understanding mechanisms of plant adaptation to environmental stresses lies in the determination of patterns of the expression of proteins, identification of stress proteins and their association with the specific functions in metabolic pathways. To date, little information has been provided about the proteomic response of grapevine to the persistent influence of adverse environmental conditions. This article describes changes in the profile of protein accumulation in leaves of common grapevine (Vitis vinifera L.) seedlings in response to prolonged drought. Isolated proteins were separated by two-dimensional electrophoresis (2 DE), and the proteins whose level of accumulation changed significantly due to the applied stress factors were identified with tandem mass spectrometry MALDI TOF/TOF type. Analysis of the proteome of grapevine leaves led to the detection of many proteins whose synthesis changed in response to the applied stressor. Drought caused the most numerous changes in the accumulation of proteins associated with carbohydrate and energy metabolism, mostly connected with the pathways of glycolysis and photosystem II protein components. The biological function of the identified proteins is discussed with reference to the stress of drought. Some of the identified proteins, especially the ones whose accumulation increased during drought stress, may be responsible for the adaptation of grapevine to drought.

Comparative proteomic analysis of ß-aminobutyric acid-mediated alleviation of salt stress in barley.

The non-protein amino acid ß-aminobutyric acid (BABA) is known to induce plant resistance to a broad spectrum of biotic and abiotic stresses. This is the first study describing the effect of BABA seed priming on physiological and proteomic changes under salt stress conditions in barley (Hordeum vulgare). The aim of our study was to investigate the changes of fresh weight, dry weight and relative water content (RWC) as well as root proteome changes of two barley lines contrasting in salt tolerance (DH14, DH 187) in response to salt stress after seed priming in water or in 800 µM BABA. Seed priming with BABA significantly increased (p ≤ 0.05) RWC in both barley lines, which indicates considerably lower water loss in BABA-primed plants than in the non-primed control plants. Dry and fresh matter increased significantly in line DH 187, whereas no changes were detected in line DH14. BABA-primed plants of both lines showed different proteomic patterns than the non-primed control plants. The root proteins exhibiting significant abundance changes (1.75-fold change, p ≤ 0.05) were separated by two-dimensional polyacrylamide gel electrophoresis (2D- PAGE). Thirty-one spots, representing 24 proteins, were successfully identified by MALDI TOF/TOF mass spectrometry. The most prominent differences include the up-regulation of antioxidant enzymes (catalase, peroxidase and superoxide dismutase), PR proteins (chitinase, endo-1,3-ß-glucosidase), and chaperones (cyclophilin, HSC 70). Our results indicate that BABA induces defence and detoxification processes which may enable faster and more effective responses to salt stress, increasing the chances of survival under adverse environmental conditions.

Alterations in root proteome of salt-sensitive and tolerant barley lines under salt stress conditions.

Salinity is one of the most important abiotic stresses causing a significant reduction of crop plants yield. To gain a better understanding of salinity tolerance mechanisms in barley (Hordeum vulgare), we investigated the changes in root proteome of salt-sensitive (DH14) and tolerant (DH187) lines in response to salt-stress. The seeds of both barley lines were germinating in water or in 100mM NaCl for 6 days. The root proteins were separated by two-dimensional gel electrophoresis. To identify proteins regulated in response to salt stress, MALDI-TOF/TOF mass spectrometry was applied. It was demonstrated that the sensitive and tolerant barley lines respond differently to salt stress. Some of the identified proteins are well-documented as markers of salinity resistance, but several proteins have not been detected in response to salt stress earlier, although they are known to be associated with other abiotic stresses. The most significant differences concerned the proteins that are involved in signal transduction (annexin, translationally-controlled tumor protein homolog, lipoxygenases), detoxification (osmotin, vacuolar ATP-ase), protein folding processes (protein disulfide isomerase) and cell wall metabolism (UDP-glucuronic acid decarboxylase, ß-d-glucan exohydrolase, UDP-glucose pyrophosphorylase). The results suggest that the enhanced salinity tolerance of DH187 line results mainly from an increased activity of signal transduction mechanisms eventually leading to the accumulation of stress protective proteins and cell wall structure changes.

Analysis of phenolic compounds and antioxidant abilities of extracts from germinating Vitis californica seeds submitted to cold stress conditions and recovery after the stress.

The material for this study consisted of stratified seeds of Vitis californica submitted to germination under optimum conditions (+25 °C) or under chill stress (+10 °C), also followed by recovery. It has been determined that the germinating seeds contain considerable amounts of tannins, catechins as well as phenolic acids such as gallic, p-coumaric, caffeic and ferulic acids. Gallic acid appeared in the highest amount in the germinating seeds (from 42.40-204.00 µg/g of fresh weight (FW)), followed by caffeic acid (from 6.62-20.13 µg/g FW), p-coumaric acid (from 2.59-5.41 µg/g FW), and ferulic acid (from 0.56-0.92 µg/g FW). The phenolic acids occurred mostly in the ester form. Under chill stress, the germinating seeds were determined to contain an elevated total amount of phenolics, as well as raised levels of condensed tannins, catechins, gallic acid, and gafeic acid. The levels of p-coumoric and ferulic acids were found to have decreased. In extracts isolated from a sample exposed to low temperature, increased antioxidant activity and reduction potential were also demonstrated. Tissue of the germinating seeds which underwent post-stress recovery was found to have less total phenolics.

Proteomic changes in the roots of germinating Phaseolus vulgaris seeds in response to chilling stress and post-stress recovery.

Plants respond to different environmental cues in a complex way, entailing changes at the cellular and physiological levels. An important step to understand the molecular foundation of stress response in plants is the analysis of stress-responsive proteins. In this work we attempted to investigate and compare changes in the abundance of proteins in the roots of bean (Phaseolus vulgaris L.) germinating under long continuous chilling conditions (10°C, 16 days), exposed to short rapid chilling during germination (10°C, 24h), as well as subjected to recovery from stress (25°C, 24h). The results we obtained indicate that germination under continuous chilling causes alterations in the accumulation of the proteins involved in stress response, energy production, translation, vesicle transport, secondary metabolism and protein degradation. The subsequent recovery influences the accumulation of the proteins implicated in calcium-dependent signal transduction pathways, secondary metabolism and those promoting cell division and expansion. Subjecting the germinating bean seeds to short rapid chilling stress resulted in a transient changes in the relative content of the proteins taking part in energy production, DNA repair, RNA processing and translation. Short stress triggers also the mechanisms of protection against oxidative stress and promotes expression of anti-stress proteins. Subjecting bean seeds to the subsequent recovery influences the abundance of the proteins involved in energy metabolism, protection against stress and production of phytohormones. The exposure to long and short chilling did not result in the alterations of any proteins common to both treatments. The same situation was observed with respect to the recovery after stresses. Bean response to chilling is therefore strongly correlated with the manner and length of exposure to low temperature, which causes divergent proteomic alterations in the roots.

Changes in the protein patterns in pea (Pisum sativum L.) roots under the influence of long- and short-term chilling stress and post-stress recovery.

Amongst many factors restricting geographical distribution of plants and crop productivity, low temperature is one of the most important. To gain better understanding of the molecular response of germinating pea (Pisum sativum L.) to low temperature, we investigated the influence of long and short chilling stress as well as post-stress recovery on the alterations in the root proteomes. The impact of long stress was examined on the pea seeds germinating in the continuous chilling conditions of 10 °C for 8 days (LS). To examine the impact of short stress, pea seeds germinating for 72 h in the optimal temperature of 20 °C were subjected to 24-h chilling (SS). Additionally, both stress treatments were followed by 24 h of recovery in the optimal conditions (accordingly LSR and SR). Using the 2D gel electrophoresis and MALDI-TOF MS protein identification, it was revealed, that most of the proteins undergoing regulation under the applied conditions were implicated in metabolism, protection against stress, cell cycle regulation, cell structure maintenance and hormone synthesis, which altogether may influence root growth and development in the early stages of plant life. The obtained results have shown that most of detected alterations in the proteome patterns of pea roots are dependent on stress duration. However, there are some analogical response pathways which are triggered regardless of stress length. The functions of proteins which accumulation has been changed by chilling stress and post-stress recovery are discussed here in relation to their impact on pea roots development.

Differences in the phenolic composition and antioxidant properties between Vitis coignetiae and Vitis vinifera seeds extracts.

Phenolic compounds were extracted from European and Japanese grapevine species (Vitis vinifera and V. coignetiae) seeds using 80% methanol or 80% acetone. The total content of phenolic compounds was determined utilizing Folin-Ciocalteu's phenol reagent, while the content of tannins was assayed by the vanillin and BSA precipitation methods. Additionally, the DPPH free radical and ABTS cation radical scavenging activities and the reduction power of the extracts were measured. The HPLC method was applied to determine the phenolic compounds, such as phenolic acids and catechins. The seeds contained large amounts of tannins and gallic acid and observable quantities of catechins, p-coumaric, ferulic and caffeic acids. The dominant form of phenolic acids in the extracts was the ester-bound form. The content of total phenolics was higher in the European grape V. vinifera seeds, which also contained more tannins, catechins and phenolic acids, except for caffeic acid. Extracts from V. vinifera seeds showed better radical scavenger properties and stronger reducing power. The total contents of phenolic compounds and tannins in acetone extracts were higher than in methanolic extracts. Acetone extracts also exhibited stronger antiradical properties as well as stronger reducing power.

Proteomic analysis of response to long-term continuous stress in roots of germinating soybean seeds.

Germination is a complex process, highly dependent on various environmental factors, including temperature and water availability. Germinating soybean seeds are especially vulnerable to unfavorable environmental conditions and exposure to long-term abiotic stresses may result in diminishing much of the yield and most importantly - restrained germination. In the present study, a proteomic approach was employed to analyze influence of cold and osmotic stress on roots of germinated soybean (Glycine max, L.) seeds. Seeds were germinating under continuous conditions of cold stress (+10°C/H2O), osmotic stress (+25°C/-0.2MPa) as well as cold and osmotic stress combined (+10°C/-0.2MPa). Proteome maps established for control samples and stress-treated samples displayed 1272 CBB-stained spots. A total of 59 proteins, present in both control and stress-treated samples and showing significant differences in volume, were identified with LC/nanoESI-MS. Identified proteins divided into functional categories, revealed 9 proteins involved in plant defense, 8 proteins responsible for plant destination and storage and 10 proteins involved in various tracks of carbohydrate metabolism. Furthermore, a number of proteins were assigned to electron transport, range of metabolic pathways, secondary metabolism, protein synthesis, embryogenesis and development, signal transduction, cellular transport, translocation and storage. By analyzing differences in expression patterns, it was possible to trace the soybean response to long-term abiotic stress as well as to distinguish similarities and differences between response to cold and osmotic stress.

Extracts of phenolic compounds from seeds of three wild grapevines-comparison of their antioxidant activities and the content of phenolic compounds.

Phenolic compounds were extracted from three wild grapevine species: Vitis californica, V. riparia and V. amurensis seeds using 80% methanol or 80% acetone. The total content of phenolic compounds was determined utilizing the Folin-Ciocalteu's phenol reagent while the content of tannins was assayed with the vanillin and BSA precipitation methods. Additionally, the DPPH free radical scavenging activity and the reduction power of the extracts were measured. The RP-HPLC method was applied to identify the phenolic compounds in the extracts, such as phenolic acids and catechins. The seeds contained large amounts of tannins, catechins and gallic acid and observable quantities of p-coumaric acid. The total content of phenolic compounds and tannins was similar in the extracts from V. californica and V. riparia seeds. However, the total content of total phenolic compounds and tannins in the extracts from V. californica and V. riperia seeds were about two-fold higher than that in the extracts from V. amurensis seeds. Extracts from seeds of the American species (V. californica and V. riparia) contained similarly high concentrations of tannins, whereas extracts from seeds of V. amurensis had approximately half that amount of these compounds. The content of catechin and epicatechin was similar in all extracts. The highest DPPH(•) anti-radical scavenging activity was observed in the acetonic and methanolic extracts of V. californica and V. riparia seeds- while the acetonic extract from the V. californica seeds was the strongest reducing agent.