Hieracium waldsteinii (Asteraceae) and Onosma stellulata (Boraginaceae) as a Source of Antioxidant and Antimicrobial Agents.

The current study aimed to phytochemically characterize (including a detailed phenolic profile) two endemic Balkan's species (Hieracium waldsteinii and Onosma stellulata) and determine their possible application as a source of natural antioxidant and antimicrobial agents. The main phenolic compound in both species (in all examined parts) was chlorogenic acid. Eriodictyol, genistein and naringenin were quantified only in H. waldsteinii while isorhamnetin-3-O-rutinoside and sinapic acid were characteristic for O. stellulata. The highest antioxidant activity (98 mg AAE/g dry weight for TAC assay) was ascribed to the flower extract of H. waldsteinii while the lowest results (∼4.3 mg AAE/g dry weight for FRP assay) were exhibited by the extracts obtained from the plant's stem. Antimicrobial assays showed moderate antibacterial, i. e., moderate/strong activity against several tested fungi (in particular Trichoderma viride). Correlation analysis revealed strong positive connection between phenolic compounds and reducing power of extracts as well as between total phenolic and flavonoid content and the obtained minimal inhibitory concentration recorded in antibacterial assays.

Skimmed Goat's Milk Powder Enriched with Grape Pomace Seed Extract: Phenolics and Protein Characterization and Antioxidant Properties.

The aim of this research was phenolics and protein characterization and antioxidant properties evaluation of skimmed thermally treated goat's milk powder enriched with different concentration of grape pomace seed extract (SE). The dominant phenolics in SE were phenolic acids, flavan-3-ols and procyanidins. Different electrophoretic techniques together with UHPLC-MS/MS analysis revealed the presence of phenolics-protein interactions in the samples, mainly procyanidins with whey protein/caseins complexes. Addition of SE into thermally treated goat's milk significantly improved antioxidant properties of goat's milk such as TAC, FRP, DPPH• and ABTS•+ scavenging activity. Gallic acid, catechin, and procyanidins mostly contributed to these activities. The schematic representation of phenolics-casein micelles interactions in thermally treated goat's milk enriched with SE was given. The addition of SE into thermally treated goat's milk can be a promising strategy in food waste recovery and to enhance the beneficial health effects of goat's milk-based functional foods.

Polyphenol bioaccessibility and antioxidant properties of in vitro digested spray-dried thermally-treated skimmed goat milk enriched with pollen.

The aim of research was to determine polyphenols bioaccessibility and antioxidant properties of thermally-treated skimmed goat milk enriched with sunflower bee-collected pollen through in vitro digestion. HPLC analysis confirmed that pollen-enriched milk contained flavonols as the main phenolic fraction (80.7-76.2%) followed by phenolic acids (14.2-17.4%). Among individual compounds quercetin-3-O-glucoside (155.1-197.2 µg/L) and p-coumaric acid (29.5-30.7 µg/L) were the main quantified flavonols and phenolic acids, respectively. After digestion of milk/pollen sample, total polyphenols recovery was 30.71% with higher phenolic acids recovery (40.1%) compared to flavonols (28.3%) indicating strong interactions between caprine milk casein micelles and pollen polyphenols. Applied antioxidant assays (phosphomolybdenum, ABTS•+scavenging activity and ferrous-ion-chelating capacity) have confirmed complexity of prepared product- it had high ability to quench ABTS•+ radicals and to form chelating complexes with Fe2+ ions. Digestion provoked 20% reduction in total antioxidant capacity compared to the initial sample. TTSG milk/pollen powder could be good functional ingredient.

Comparative Study on the Phenolic Fingerprint and Antioxidant Activity of Strawberry Tree (Arbutus unedo L.) Leaves and Fruits.

The strawberry tree (Arbutus unedo L., Ericaceae family) is an evergreen Mediterranean shrub whose leaves and fruits are used in traditional medicine due to their antioxidant, antimicrobial, antidiabetic, diuretic, and antiproliferative properties. The health benefits are mainly attributed to the presence of phenolic compounds. The aim of this study was to compare the phenolic profiles, total phenolic content (TPC), and radical scavenging activity (RSA) of A. unedo leaves and fruits collected at two locations in Croatia. Phenolic profiles were identified using an ultra-high-performance liquid chromatograph (UHPLC) coupled with a hybrid mass spectrometer (LTQ Orbitrap MS). TPC was determined by Folin-Ciocalteu's assay, while RSA was investigated using DPPH reagent. A total of 64 phenolics (60 and 42 compounds in leaves and fruits, respectively) were identified. Hyperoside and flavan-3-ols were predominant compounds in leaves, while gallocatechin and catechin were the major compounds found in fruits. To the authors' knowledge, 16 and 5 phenolics in leaves and fruits, respectively, were reported for the first time. Principal component analysis (PCA) showed that UHPLC-LTQ Orbitrap MS could be used to identify which phenolics were able to discriminate samples regarding plant tissue and geographical origin. TPC in leaves and fruits were in the ranges of 67.07-104.74 and 16.78-25.86 mg gallic acid equivalents (GAE)/g dried weight (dw), respectively. RSA for leaves and fruits were in the ranges of 408.92-430.98 and 74.30-104.04 µmol Trolox equivalents (TE)/g dw, respectively. The number of identified phenolics was lower in fruits compared to leaves. Such a large number of bioactive phenolics identified and the strong antioxidant activity pointed to A. unedo as a promising health-promoting plant and natural food preservative.

Chemical composition and antimicrobial activity of Osage orange (Maclura pomifera) leaf extracts.

The main goal of this study was to establish the chemical profile of Osage orange (Maclura pomifera) leaf extracts, obtained by conventional maceration technique, and to examine its antimicrobial activity. The identification and quantification of the extract compounds were done using ultra-high-performance liquid chromatography, with a diode array detector coupled with triple-quadrupole mass spectrometer and gas chromatography-mass spectrometry techniques. Thirty-one polyphenolic compounds were detected and identified in the ethanolic extracts, whereby 5-O-caffeoylquinic acid was found to be the dominant compound. Among other compounds, pentacosane and palmitic acid were the most abundant compounds in the dichloromethane extract. The preliminary antimicrobial activity screening shows that Gram-positive bacteria tend to be more sensitive to the investigated extracts. The highest antimicrobial activity was determined against Enterococcus faecalis ATCC 19433 and Listeria monocytogenes ATCC 35152. From these results, Osage orange leaves can be considered as plant material with significant antimicrobial properties.

Polyphenols as Possible Agents for Pancreatic Diseases.

Pancreatic cancer (PC) is very aggressive and it is estimated that it kills nearly 50% of patients within the first six months. The lack of symptoms specific to this disease prevents early diagnosis and treatment. Today, gemcitabine alone or in combination with other cytostatic agents such as cisplatin (Cis), 5-fluorouracil (5-FU), irinotecan, capecitabine, or oxaliplatin (Oxa) is used in conventional therapy. Outgoing literature provides data on the use of polyphenols, biologically active compounds, in the treatment of pancreatic cancer and the prevention of acute pancreatitis. Therefore, the first part of this review gives a brief overview of the state of pancreatic disease as well as the procedures for its treatment. The second part provides a detailed overview of the research regarding the anticancer effects of both pure polyphenols and their plant extracts. The results regarding the antiproliferative, antimetastatic, as well as inhibitory effects of polyphenols against PC cell lines as well as the prevention of acute pancreatitis are presented in detail. Finally, particular emphasis is given to the polyphenolic profiles of apples, berries, cherries, sour cherries, and grapes, given the fact that these fruits are rich in polyphenols and anthocyanins. Polyphenolic profiles, the content of individual polyphenols, and their relationships are discussed. Based on this, significant data can be obtained regarding the amount of fruit that should be consumed daily to achieve a therapeutic effect.

The Application of Pollen as a Functional Food and Feed Ingredient-The Present and Perspectives.

Pollen is recognized as an excellent dietary supplement for human nutrition, which is why it can be found in different forms on the market (granules, capsules, tablets, pellets, and powders). But, the digestibility of pollen's nutrients is strongly affected by the presence of a pollen shell, which can decrease the bioavailability of nutrients by 50% and more. Since consumers have become more aware of the benefits of a healthy diet and the necessity to improve pollen digestibility, different pollen-based functional food products have been developed and extensive studies were done to estimate the beneficial effects of pollen-based feed on animal growth, health, and rigor mortise stage. Considering the positive effects of pollen nutrients and phytometabolites on human and animal health, the aim of this paper was to give an overview of recent achievements in the application of pollen in the formulation of functional food and animal diets. Special attention was paid to the effects of pollen's addition on the nutritional, functional, techno-functional, and sensory properties of the new formulated food products. Anti-nutritional properties of pollen were also discussed. This review points out the benefits of pollen addition to food and feed and the possible directions in the further development of functional food and feed for the wellbeing of everyone.

The functional food production: Application of stinging nettle leaves and its extracts in the baking of a bread.

Stinging nettle is an annual plant. This plant is known for applications in folk medicine, and as the human diet. The stinging nettle leaves has been used to obtain extracts, which are expected to be with a high content of biologically active compounds. Prepared microwave extracts were applied in the formulation of the functional products. The article completes preparation of the bread with addition of the stinging nettle leaves and its extract, and bread's composition in phenolic acids, flavonoids, micro elements, and macro elements. The biological activity of the prepared bread samples showed significant antioxidant activity. This was especially true against DPPH radicals. It has been shown that it holds a high cytotoxic activity. Leaves themselves decreased the quality of the bread, while the extract improved the quality. It was sensorially confirmed. The article concluded that the extract substitutes leaves in bread as a product of a high benefit.

Phenolic Composition Influences the Health-Promoting Potential of Bee-Pollen.

Information on compositional, nutritional and functional properties of bee-pollen, as a health-promoting food, is essential for defining its quality. Concerning the nutritional importance of phenolic compounds, the aim of this study was to determine the phenolic profile and antioxidant activity of twenty-four bee-pollen samples collected from different regions of Serbia. High-performance thin-layer chromatographic (HPTLC) fingerprinting was used for profiling of bee-pollen samples according to the botanical type. HPTLC hyphenated with image analysis and a pattern recognition technique confirmed the grouping of samples caused by the specific phenolic composition of pollens of different botanical origin. Flavonoid glycosides in bee-pollen samples were identified by applying ultra-high-performance liquid chromatography (UHPLC) coupled with linear ion trap-Orbitrap mass spectrometry (LTQ Orbitrap MS). Eight out of twenty-seven flavonol glycosides were identified in bee-pollen samples for the first time. All analyzed bee-pollen samples showed a high number of phenolic compounds which may have therapeutic potential.

In vitro digestion of meat- and cereal-based food matrix enriched with grape extracts: How are polyphenol composition, bioaccessibility and antioxidant activity affected?

The aim of this study was to evaluate the effect of enriching a complex food matrix (FM) with grape extracts on polyphenol content, composition, bioaccessibility and antioxidant activity during digestion. The grape extracts and FM were separately tested under the same conditions as controls. The FM by itself contains a significant amount of phenolic acids and flavonols, influencing the final recovery of polyphenols from grape extracts. The FM significantly increased the total recovery of polyphenols after digestion of grape seed extracts compared to those digested without the FM; however, a low recovery of proantocyanidins and total flavonoids was observed. Digestive fluids and FM compounds significantly increased the total polyphenol content of grape digests and significantly contributed to their ABTS+ scavenging activity and ferrous-ion-chelating capacity. The present study suggested that enrichment of meat- and cereal-based products with grape polyphenol extracts could be a good strategy to formulate a healthier diet.

Mycotoxins and Mycotoxin Producing Fungi in Pollen: Review.

Due to its divergent chemical composition and good nutritional properties, pollen is not only important as a potential food supplement but also as a good substrate for the development of different microorganisms. Among such microorganisms, toxigenic fungi are extremely dangerous as they can synthesize mycotoxins as a part of their metabolic pathways. Furthermore, favorable conditions that enable the synthesis of mycotoxins (adequate temperature, relative humidity, pH, and aw values) are found frequently during pollen collection and/or production process. Internationally, several different mycotoxins have been identified in pollen samples, with a noted predominance of aflatoxins, ochratoxins, fumonisins, zearalenone, deoxynivalenol, and T-2 toxin. Mycotoxins are, generally speaking, extremely harmful for humans and other mammals. Current EU legislation contains guidelines on the permissible content of this group of compounds, but without information pertaining to the content of mycotoxins in pollen. Currently only aflatoxins have been researched and discussed in the literature in regard to proposed limits. Therefore, the aim of this review is to give information about the presence of different mycotoxins in pollen samples collected all around the world, to propose possible aflatoxin contamination pathways, and to emphasize the importance of a regular mycotoxicological analysis of pollen. Furthermore, a suggestion is made regarding the legal regulation of pollen as a food supplement and the proposed tolerable limits for other mycotoxins.

The influence of the extraction temperature on polyphenolic profiles and bioactivity of chamomile (Matricaria chamomilla L.) subcritical water extracts.

The main goal of this research was to determine the relationship among chemical structure, bioactivity and temperature of chamomile during subcritical water extraction in isobaric conditions (45 bar) at seven different temperatures (65-210 °C). The influence of temperature on phenolic profiles was defined by UHPLC-HESI-MS/MS. The overall results indicate that the presence of conjugated double bonds, side chains, glucose moiety or ether moiety in molecules influence the efficiency of polyphenols' extraction in subcritical water. In terms of antioxidant activity, the extracts were the most active towards ABTS radicals (IC50 = 7.3-16.8 µg/mL), whereby temperature of 150 °C was optimal. On the other hand, the extracts obtained at 115 °C showed highest cytotoxicity. Inhibition of α-amylase and α-glucosidase was the highest at 65 and 85 °C, i.e. 0.51 and 4.13 mmolAE/g, respectively. Activity against tyrosinase was the highest at 210 °C (17.92 mgKAE/g). The data showed that different non-phenolic compounds may also participate in bio-activities of the extracts.

Subcritical water extraction as a cutting edge technology for the extraction of bioactive compounds from chamomile: Influence of pressure on chemical composition and bioactivity of extracts.

The study was designed to determine the relationship between chemical structure, bioactivity and pressure during the subcritical water extraction (SCW) of chamomile. Extraction was carried out at isothermal conditions (100 °C) at five different pressures (10, 30, 45, 60 and 90 bar). Twenty three polyphenolic compounds were identified in the extracts, whereby apigenin was found to be the dominant compound (61.53-1344.99 mg/kg). Results suggest that the lowest applied pressure has negligible effect on phenolic recovery from chamomile, but also the use of pressures above than 45 bar was proven as needlessly. By using in vitro assays, influence of pressure on antioxidant, cytotoxic and enzyme-inhibitory activities of the extracts was evaluated. Extracts obtained at 30, 45 and 60 bar exhibited stronger bioactivities than at 10 and 90 bar. It was concluded that pressure exert a significant influence on chemical composition of extracts, and thus on biological activity of chamomile extracts.

The polyphenolics and carbohydrates as indicators of botanical and geographical origin of Serbian autochthonous clones of red spice paprika.

Spice peppers (Capsicum annuum L.) var. Lemeska and Lakosnicka paprika were investigated to evaluate their polyphenolic and carbohydrate profiles and antioxidant activity. A total of forty-nine polyphenolics were identified using ultrahigh-performance liquid chromatography (UHPLC) coupled to LTQ OrbiTrap mass analyzer. Twenty-five of them were quantified using available standards, while the other compounds were confirmed by exact mass search of their deprotonated molecule [M-H](-) and its MS(4) fragmentation. Thirteen carbohydrates were quantified using high-performance anion exchange chromatography (HPAEC) with pulsed amperometric detection (PAD). Radical scavenging activity (RSA) ranged from 17.32 to 48.34mmol TE (Trolox equivalent)/kg DW (dry weight) and total phenolics content (TPC) was ranged between 7.03 and 14.92g GAE (gallic acid equivalents)/kg DW. To our best knowledge, five polyphenolic compounds were for the first time tentatively identified in paprika: 5-O-p-coumaroylquinic acid, luteolin 7-O-(2″-O-pentosyl-4″-O-hexosyl)hexoside, quercetin 3-O-(2″-O-hexosyl)rhamnoside, isorhamnetin 3-O-[6″-O-(5-hydroxyferuloyl)hexoside]-7-O-rhamnoside, and luteolin 7-O-[2″-O-(5'″-O-sinapoyl)pentosyl-6″-O-malonyl]hexoside.

Study of silver, selenium and arsenic concentration in wild edible mushroom Macrolepiota procera, health benefit and risk.

The content and bioaccumulation of trace (Ag, Se, As) and major elements (Ca, Mg, Na and K) in wild edible mushroom Macrolepiota procera and its corresponding soil substrates, collected from five sites in the Rasina region in central Serbia, were investigated. The content of Ag, As and Se was determined by inductively coupled plasma mass spectrometer (ICP-MS) while the amount of Ca, Mg, Na and K was determined by inductively coupled plasma optical emission spectrometer (ICP-OES). The concentrations of major elements in the mushrooms were at typical levels. As far as trace elements are concerned, M. procera bioaccumulates silver although all samples were collected from unpolluted sites. It was found that the content of Ag depended on the geographical origin and the density of fruiting body on the certain site. Principal component analysis distinguished the mushroom samples from different geographical areas and revealed the influence of soil composition on metal content in fruiting bodies. Also, a linear regression correlation test was performed to investigate correlations between Ag, Cd, Se, Pb and As in caps and stipes at different geographic sites separately. In addition, our results indicated that M. procera could serve as a good dietary source of Mg, K and Se. The content of Ag and As was low, so it could not pose a health risk for consumers.

Identification of Phenolic Compounds from Seed Coats of Differently Colored European Varieties of Pea (Pisum sativum L.) and Characterization of Their Antioxidant and In Vitro Anticancer Activities.

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 46.56 mg 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.

Identification and quantification of phenolic compounds in berry skin, pulp, and seeds in 13 grapevine varieties grown in Serbia.

Phenolic profile of 13 grapevine varieties was determined, with respect to three different parts of berries: skin, seed and pulp. Identification and quantification of the phenols was done using ultra-high performance liquid chromatography (UHPLC) coupled with a diode array detector and a triple-quadrupole mass spectrometer. The most abundant phenols in grape seeds were found to be flavan-3-ols, most of which are gallocatechin gallate and catechin. The skins were characterized mostly by flavonols, i.e. quercetin and myricetin. Characterization of anthocyanins in the berry skin by UHPLC coupled with Linear Trap Quadrupole and OrbiTrap mass analyzer revealed a total of twenty derivatives of malvidin, delphinidin, petunidin, cyanidin, and peonidin. To the extent of our knowledge this is the first work that shows the presence of malvidin 3,5-O-dihexoside in the berry skin of 'Merlot', 'Cabernet Franc', 'Shiraz' 'Sangiovese', 'Pinot Noir' and 'Prokupac', untypical for Vitis vinifera Linneo species.

Chemical profile of major taste- and health-related compounds of Oblacinska sour cherry.

BACKGROUND: Oblacinska sour cherry, an autochthonous cultivar, is the most planted cultivar in Serbian commercial orchards. Owing to its long cultivation under different agro-ecological conditions, it is a mixture of different clones. To obtain comprehensive information on Oblacinska sour cherry fruit, the total carbohydrate composition, total phenolic content, total anthocyanin content, antioxidant activity and polyphenolic profiles of 39 clones were investigated. RESULTS: Fructose was found to be the dominant sugar, followed by glucose, sorbitol and sucrose. Rutin and chlorogenic acid were the most abundant polyphenols. Some clones contained pinobanksin, hesperetin and galangin, which, to best knowledge, is the first time these compounds have been reported in sour cherry fruit. CONCLUSION: The data obtained showed huge variation within the germplasm studied, confirming the fact that Oblacinska sour cherry is not a cultivar but a population.

Analysis and characterisation of phytochemicals in mulberry (Morus alba L.) fruits grown in Vojvodina, North Serbia.

In this study, the polyphenolic profile of 11 Morus alba fruits grown in the Vojvodina region was investigated. Ultra high performance liquid chromatography (UHPLC) coupled with Linear Trap Quadrupole and OrbiTrap mass analyzer, and UHPLC coupled with a diode array detector and a triple-quadrupole mass spectrometer were used for the identification and quantification of the polyphenols, respectively. A total of 14 hydroxycinnamic acid esters, 13 flavonol glycosides, and 14 anthocyanins were identified in the extracts with different distributions and contents according to the sampling. The total phenolic content ranged from 43.84 to 326.29 mg GAE/100g frozen fruit. The radical scavenging capacity (50.18-86.79%), metal chelating ability (0.21-8.15%), ferric ion reducing power (0.03-38.45 µM ascorbic acid) and superoxide anion radical scavenging activity (16.53-62.83%) were assessed. The findings indicated that mulberry polyphenolics may act as potent superoxide anion radical scavengers and reducing agents.

Simultaneous UHPLC/DAD/(+/-)HESI-MS/MS analysis of phenolic acids and nepetalactones in methanol extracts of Nepeta species: a possible application in chemotaxonomic studies.

INTRODUCTION: Nepeta species contain a variety of secondary metabolites, including iridoid monoterpenes - nepetalactones and phenolic acids - that are considered the main bioactive constituents. This work represents the first attempt to comparatively explore variations in these two major groups of secondary metabolites within the genus. OBJECTIVE: To develop an efficient analytical methodology for simultaneous analysis of nepetalactones and phenolic acids in methanol extracts of selected Nepeta species, and to evaluate its potential application in chemotaxonomic studies. MATERIAL AND METHODS: A UHPLC combined with linear-trap quadrupole (LTQ) orbitrap MS method was used to characterise chemical diversity and complexity of phenolics among 12 selected Nepeta species. A targeted metabolomic approach using UHPLC coupled to a diode array detector (DAD) and combined with (+/-) heated electrospray ionisation (HESI) MS/MS was developed and validated for quantitative analysis of six hydroxycinnamic acid derivatives and four nepetalactones. RESULTS: Phenolic profiling provided a valuable database of bioactive compounds in the plant group studied, including phenolic acids (hydroxybenzoic and hydroxycinnamic acids) and flavonoids (flavones, flavonols and flavanones). Principal component analysis and cluster analysis suggested the applicability of 10 targeted compounds as chemomarkers for chemotaxonomic studies. Pearson's correlation analysis revealed significant positive correlations between metabolites involved in different biosynthetic pathways (phenylpropanoid or monoterpenoid). CONCLUSION: The described targeted metabolomic approach proved to be highly beneficial in designing a phytochemical overview of the genus Nepeta, and might have applications in further clarification of phylogenetic relations. Furthermore, it has the potential to be implemented in a routine quality control of plant material and herbal preparations.