Construction of Highly Functionalized 2-Styrylfurans by N-Heterocyclic Carbene/Brønsted Acid Catalysis.

This research presents an original method for synthesizing styrylfurans using N-heterocyclic carbenes (NHCs) and Brønsted acid catalysis. By exploiting 2,4-dioxoesters as conjugated 1,3-dicarbonyls, we have developed a technique allowing the efficient formation of highly functionalized styrylfurans with interesting photochemical properties, through a NHC-catalyzed cross-benzoin reaction followed by a Brønsted acid-driven Paal-Knorr-like condensation. This approach permits the integration of various substituents on the furan ring, with preliminary biological studies indicating potential as fluorescent dyes.

Distribution of exchangeable Ca2+ during the process of Larix decidua Mill. pollination and germination.

The involvement of Ca2+ ions in angiosperms sexual processes is well established, while in gymnosperms, such knowledge remains limited and is still a topic of discussion. In this study, we focused on Larix decidua, using Alizarin-red S staining and the pyroantimonate method to examine the tissue and subcellular distribution of free and loosely bound Ca2+ ions at different stages of the male gametophyte's development and its interaction with the ovule. Our findings show that in larch, both the germination of pollen grains and the growth of pollen tubes occur in an environment rich in Ca2+. These ions play a crucial role in the adhesion of the pollen grain to the stigmatic tip and its subsequent movement to the micropylar canal. There is a significant presence of free and loosely bound Ca2+ ions in both the fluid of the micropylar canal and the extracellular matrix of the nucellus. As the pollen tube extends through the nucellus, we observed a notable accumulation of Ca2+ ions just above the entry to the mature archegonium, a region likely crucial for the male gametophyte's directional growth. Meanwhile, the localized presence of free and loosely bound Ca2+ ions within the egg cell cytoplasm may inhibit the pollen tubes growth and rupture, playing an important role in fertilization.

Comparative Studies of Extracts Obtained from Brassica oleracea L. Plants at Different Stages of Growth by Isolation and Determination of Isothiocyanates: An Assessment of Chemopreventive Properties of Broccoli.

The main goal of this work was to develop analytical procedures for the isolation and determination of selected isothiocyanates. As an example, particularly sulforaphane from plants of the Brassicaceae Burnett or Cruciferae Juss family. The applied methodology was mainly based on classical extraction methods and high-performance liquid chromatography coupled with tandem mass spectrometry. Moreover, the effect of temperature on the release of isothiocyanates from plant cells was considered. The cytotoxic activity of the obtained plant extracts against a selected cancer cell line has also been included. The results allow evaluating the usefulness of obtained plant extracts and raw sprouts regarding their content of isothiocyanates-bioactive compounds with chemopreventive properties.

Enantioselective Synthesis of Highly Substituted Fluoroalkylated Benzopyranones and 3-Coumaranones via N-Heterocyclic Carbene-Catalyzed Intramolecular Annulations.

A highly enantioselective intramolecular NHC-catalyzed approach for the synthesis of fluoroalkylated benzopyranones and 3-coumaranones with all-carbon quaternary stereocenters is presented. This reaction is catalyzed by N-heterocyclic carbenes (NHCs) and involves annulation reactions between in situ generated acyl anion intermediates and highly substituted trifluoromethyl-ß,ß-disubstituted Michael acceptors. The method can also be extended to perfluoroalkyl homologues.

Functionalization of Alpha-Lactalbumin by Zinc Ions.

Alpha-lactalbumin (α-LA) and binding of zinc cations to protein were studied. Molecular characteristics of protein was determined by MALDI-TOF/MS and electrophoresis SDS-PAGE, and also, for complexes, it was determined by spectroscopic techniques (ATR-FT-IR and Raman) and microscopic techniques (SEM along with an EDX detector and also TEM). The pH dependence of zeta potential of α-LA was determined in saline solution. The zinc binding to the protein mechanism was investigated; zinc binding to protein kinetics, the molecular modeling by the DFT method, and electron microscopy (SEM and TEM) for microstructure observation were performed. The experiments performed indicate a quick binding process (equilibrium takes place after 2 min of incubation) which occurs onto the surface of α-LA. Zinc cations change the conformation of the protein and create spherical particles from the morphological point of view. DFT studies indicate the participation of acidic functional groups of the protein (aspartic acid and glutamic acid residues), and these have a decisive influence on the interaction with zinc cations. Application studies of general toxicity and cytotoxicity and bioavailability were conducted.

Isolation and Identification of Lactococcus lactis and Weissella cibaria Strains from Fermented Beetroot and an Investigation of Their Properties as Potential Starter Cultures and Probiotics.

The presence of certain microorganisms in dairy products or silage is highly desirable. Among them are probiotic strains of lactic acid bacteria (LAB), which show many beneficial features, including antimicrobial properties that support the development of beneficial microflora; in addition, owing to their biochemical activity, they influence the nutritional, dietary, and organoleptic properties of food products. Before being placed on the market, each strain requires separate testing to determine its probiotic properties and effectiveness. The aim of this study was to isolate LAB strains from a pickled beetroot sample that could be used in the dairy industry and with the potential to be considered as a probiotic in the future. Two strains identified using the MALDI technique were selected-Lactococcus lactis and Weissella cibaria. The optimal growth conditions of the strains were determined, and their proteolytic properties were assessed with the use of the o-PA reagent and spectrophotometry. The lipid profile was analyzed using the SALDI (surface-assisted laser desorption/ionization) technique and silver nanoparticles. High-performance liquid chromatography was used to assess the ability of the strains to synthesize beneficial metabolites, such as B vitamins (B2, B3, and B9) or lactic acid, and gas chromatography was used to analyze the substances responsible for organoleptic properties. Moreover, the ability to inhibit the growth of pathogenic strains was also tested in the selected strains. Both tested strains demonstrated the desired properties of starter cultures for future use in functional food production, showing that fermented plant products can serve as valuable potential probiotic sources.

Synthesis and physicochemical characterization of bovine lactoferrin supersaturated complex with iron (III) ions.

The aim of the study was to investigate the process of Fe3+ binding to bLTF. Moreover, the physicochemical characterization of the respective supersaturated complex was studied. The knowledge should be important for the description of processes that may take place in dairy products fortified with iron. Additionally, the synthesized complex can be utilized as a dietary supplement for the treatment of iron deficiency anemia (IDA). Finally, it was shown that formation of supersaturated iron-protein structures which include LTF often accompanies development of neurodegenerative diseases such as Alzheimer or Parkinson. Thus, the study can reveal some aspects of its pathogenesis process. The methodology of the investigation comprised the utilization of batch sorption study and applying Freundlich and Langmuir models. The complex also was characterized by numerous techniques: spectrometric (ICP-MS), spectroscopic (UV-Vis, ATR-FTIR), electron microscopy (TEM-EDX), SDS-PAGE. Based on obtained results the potential mechanisms of iron interaction with protein were described. Moreover, the molecular docking was applied to visualize possible metal binding sites. The respective complex contains ≈ 33.0 mg/g of iron which is nearly 50 Fe3+ per one protein molecule. The cytotoxicity of the obtained complex was evaluated by MTT reduction and LDH release assays on Caco-2 and nL929 cell lines.

Synthesis, Physicochemical Characterization, and Antibacterial Performance of Silver-Lactoferrin Complexes.

Antibiotic-resistant bacteria pose one of the major threats to human health worldwide. The issue is fundamental in the case of chronic wound treatment. One of the latest trends to overcome the problem is the search for new antibacterial agents based on silver. Thus, the aim of this research was to synthesize the silver-lactoferrin complex as a new generation of substances for the treatment of infected wounds. Moreover, one of the tasks was to investigate the formation mechanisms of the respective complexes and the influence of different synthesis conditions on the features of final product. The batch-sorption study was performed by applying the Langmuir and Freundlich isotherm models for the process description. Characterization of the complexes was carried out by spectroscopy, spectrometry, and separation techniques, as well as with electron microscopy. Additionally, the biological properties of the complex were evaluated, i.e., the antibacterial activity against selected bacteria and the impact on L929 cell-line viability. The results indicate the formation of a heterogeneous silver-lactoferrin complex that comprises silver nanoparticles. The complex has higher antibacterial strength than both native bovine lactoferrin and Ag+, while being comparable to silver toxicity.

Nanostructured Layer of Silver for Detection of Small Biomolecules in Surface-Assisted Laser Desorption Ionization Mass Spectrometry.

A facile approach for the synthesis of a silver nanostructured layer for application in surface-assisted laser desorption/ionization mass spectrometry of low-molecular-weight biomolecules was developed using electrochemical deposition. The deposition was carried out using the following silver salts: trifluoroacetate, acetate and nitrate, varying the voltage and time. The plate based on trifluoroacetate at 10 V for 15 min showed intense SALDI-MS responses for standards of various classes of compounds: fatty acids, cyclitols, saccharides and lipids at a concentration of 1 nmol/spot, with values of the signal-to-noise ratio ≥50. The values of the limit of detection were 0.71 µM for adonitol, 2.08 µM for glucose and 0.39 µM for palmitic acid per spot. SEM analysis of the plate showed anisotropic flower-like microstructures with nanostructures on their surface. The reduced chemical background in the low-mass region can probably be explained by the absence of stabilizers and reducing agents during the synthesis. The plate synthesized with the developed approach showed potential for future use in the analysis of low-molecular-weight compounds of biological relevance. The absence of the need for the utilization of sophisticated equipment and the synthesis time (10 min) may benefit large-scale applications of the layer for the detection of various types of small biomolecules.

Lacticaseibacillus paracasei as a Modulator of Fatty Acid Compositions and Vitamin D3 in Cream.

Butter is an important source of essential fatty acids, lipid-soluble vitamins, and antioxidants in the diet. However, this study showed that the presence of the Lacticaseibacillus paracasei strain has a great influence on the fatty acid profile as well as provitamin D3 and vitamin D3 content in the cream-the raw material from which the butter is obtained. The addition of this lactic acid bacteria enriches the cream in 9-hexadecenoic acid, oleic acid, octadeca-9,12-dienoic acid, and conjugated linoleic acid, which exhibit antimutagenic and anticarcinogenic properties. Moreover, a higher level of monounsaturated fatty acids can extend the shelf life of butter in the future. In the present work, we observed that the presence of lactic acid bacteria contributed to an increase in the level of provitamin D after 6 h of incubation and an increase in the levels of vitamin D3 after 24 and 48 h. Fatty acid profiles and the content of vitamins were largely dependent on the presence of light and mixing, which are probably associated with the status of lipid peroxidation.

New sources of lactic acid bacteria with potential antibacterial properties.

In the face of the growing demand for functional food, the search for new sources of lactic acid bacteria (LAB) becomes a priority. In our research, we used multiplied culture conditions followed by identification via the matrix-assisted laser desorption ionization-time of flight mass spectrometry for seeking LAB strains in plant- and animal-derived sources. Furthermore, the selected LAB isolates were examined for their proteolytic activity as well as antimicrobial action against different bacterial pathogens. The applied method appeared to be useful tool for searching LAB strains within different types of the biological matrices. The best source of the LABs was from calf. Comparing properties of the two selected LABs, those isolated from calf demonstrated the greatest proteolytic and antibacterial properties suggesting that gastrointestinal microbiota are the most valuable LAB source. Nevertheless, second selected strain derived from pickled cucumber juice may be also treated as a promising source of potential probiotic strains.

Tissue sample preparations for preclinical research determined by molecular imaging mass spectrometry using matrix-assisted laser desorption/ionization.

Matrix-assisted laser desorption/ionization - imaging mass spectrometry is an alternative tool, which can be implemented in order to obtain and visualize the "omic" signature of tissue samples. Its application to clinical study enables simultaneous imaging-based morphological observations and mass spectrometry analysis. Application of fully informative material like tissue allows obtaining the complex and unique profile of analyzed samples. This knowledge leads to diagnosing disease, studying the mechanism of cancer development, selecting the potential biomarkers as well as correlating obtained images with prognosis. Nevertheless, it is worth noticing that this method is found to be objective but the result of the analysis is mainly influenced by the sample preparation protocol, including the collection of biological material, its preservation, and processing. However, the application of this approach requires a special sample preparation procedure. The main goal of the study is to present the current knowledge on the clinical application of matrix-assisted laser desorption/ionization with imaging mass spectrometry in cancer research, with particular emphasis on the sample preparation step. For this purpose, several protocols based on cryosections and formalin-fixed paraffin-embedded tissue were compiled and compared, taking into account the measured metabolites of potential diagnostic importance for a given type of cancer.

Modern Methods of Pre-Treatment of Plant Material for the Extraction of Bioactive Compounds.

In this review, recent advances in the methods of pre-treatment of plant material for the extraction of secondary metabolites with high biological activity are presented. The correct preparation of the material for extraction is as important as the selection of the extraction method. This step should prevent the degradation of bioactive compounds as well as the development of fungi and bacteria. Currently, the methods of preparation are expected to modify the particles of the plant material in such a way that will contribute to the release of bioactive compounds loosely bonded to cell wall polymers. This review presents a wide range of methods of preparing plant material, including drying, freeze-drying, convection drying, microwave vacuum drying, enzymatic processes, and fermentation. The influence of the particular methods on the structure of plant material particles, the level of preserved bioactive compounds, and the possibility of their release during the extraction were highlighted. The plant material pre-treatment techniques used were discussed with respect to the amount of compounds released during extraction as well their application in various industries interested in products with a high content of biologically active compounds, such as the pharmaceutical, cosmetics, and food industries.

Investigation of the mechanism of zearalenone metabolization in different systems: Electrochemical and theoretical approaches.

Mycotoxins are toxic metabolites produced by mold fungi, which commonly contaminate cereal crops. These compounds include zearalenone (ZEA), which may disturb the proper functioning of the endocrine system in mammals. The metabolism of ZEA plays a key role in its toxic properties. The type and amount of produced metabolites may contribute to both the reduction and increase in its pathogenic effect. Therefore, it is extremely important to investigate the possible pathways of zearalenone metabolism. The electrochemical approach may prove alternatives for the in vitro and in vivo methods. For this reason, in this study the electrochemical, theoretical and in vitro assays were applied to determine the possible ZEA metabolism mechanism. Electrochemical processes were conducted in the EC/ESI-MS system. The application of HPLC-MS/MS allowed to characterize the obtained electrochemical products specific for phase I and II. Appropriate conditions for redox identification processes were optimized with regard to such parameters as the potential value, mobile phase and working electrode. In addition, the data obtained by instrumental techniques for ZEA metabolism were compared with those obtained by in vitro methods for HepG2 cell lines. Furthermore, results from quantum mechanical calculations allowed to explain the mechanism and the conformational stability of the reduction products.

The potential application of supercritical CO2 in microbial inactivation of food raw materials and products.

The purpose of this study was to review the possibility of using supercritical CO2 as a green and sustainable technology for microbial inactivation of raw material for further application in the food industry. The history of the development of supercritical CO2 microbial inactivation has been widely described in this article. The fundamental scientific part of the process like mechanism of bactericidal action of CO2 or inactivation of key enzymes were characterized in detail. In summary, this study provides an overview of the latest literature on the use of supercritical carbon dioxide in microbial inactivation of food raw materials and products.

NHC-catalyzed enantioselective C2-functionalization of 3-hydroxychromenones via α,ß-unsaturated acyl azoliums.

A novel synthetic method for enantioselective C2-functionalization of 3-hydroxychromenones promoted by N-heterocyclic carbenes via the formation of α,ß-unsaturated acyl azolium intermediates, which occurs with Coates-Claisen rearrangement is established. This synthetic strategy enabled the rapid assembly of enantiomerically enriched δ-hydroxychromenone-derived esters/amides under mild conditions with good to excellent yields and broad substrate scope.

Comparison Study of Cytotoxicity of Bare and Functionalized Zinc Oxide Nanoparticles.

In this paper, a study of the cytotoxicity of bare and functionalized zinc oxide nanoparticles (ZnO NPs) is presented. The functionalized ZnO NPs were obtained by various types of biological methods including microbiological (intra- and extracellular with Lactobacillus paracasei strain), phytochemical (Medicago sativa plant extract) and biochemical (ovalbumin from egg white protein) synthesis. As a control, the bare ZnO NPs gained by chemical synthesis (commercially available) were tested. The cytotoxicity was measured through the use of (3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) dye as well as lactate dehydrogenase (LDH) assays against murine fibroblast L929 and Caco-2 cell lines. As a complementary method, scanning electron microscopy (SEM) was performed to assess the morphology of the tested cells after treatment with ZnO NPs. The microscopic data confirmed the occurrence of apoptotic blebbing and loss of membrane permeability after the administration of all ZnO NPs. The reactive oxygen species (ROS) concentration during the cell lines' exposure to ZnO NPs was measured fluorometrically. Additionally, the photocatalytic degradation of methylene blue (MB) dye in the different light conditions, as well as the antioxidant activity of bare and functionalized ZnO NPs, is also reported. The addition of all types of tested ZnO NPs to methylene blue resulted in enhanced rates of photo-degradation in the presence of both types of irradiation, but the application of UV light resulted in higher photocatalytic activity of ZnO NPs. Furthermore, bare (chemically synthetized) NPs have been recognized as the strongest photocatalysts. In the context of the obtained results, a mechanism underlying the toxicity of bio-ZnO NPs, including (a) the generation of reactive oxygen species and (b) the induction of apoptosis, is proposed.

Comparative Studies of Selected Criteria Enabling Optimization of the Extraction of Polar Biologically Active Compounds from Alfalfa with Supercritical Carbon Dioxide.

The aim of this research was to provide crucial and useful data about the selection of the optimization criteria of supercritical carbon dioxide extraction of alfalfa at a quarter-technical plant. The correlation between more general output, including total phenolics and flavonoids content, and a more specified composition of polar constituents was extensively studied. In all alfalfa extracts, polar bioactive constituents were analyzed by both spectrometric (general output) and chromatographic (detailed output) analyses. Eight specific phenolic acids and nine flavonoids were determined. The most dominant were salicylic acid (221.41 µg g-1), ferulic acid (119.73 µg g-1), quercetin (2.23 µg g-1), and apigenin (2.60 µg g-1). For all seventeen analyzed compounds, response surface methodology and analysis of variance were used to provide the optimal conditions of supercritical fluid extraction for each individual constituent. The obtained data have shown that eight of those compounds have a similar range of optimal process parameters, being significantly analogous for optimization based on total flavonoid content.

Promising Green Technology in Obtaining Functional Plant Preparations: Combined Enzyme-Assisted Supercritical Fluid Extraction of Flavonoids Isolation from Medicago Sativa Leaves.

To elaborate a complete extraction protocol for the enhanced release of biologically active compounds from plant cells, this study aimed to optimize together the parameters of the supercritical fluid extraction (SFE) process (temperature, pressure, and percentage of cosolvent) and enzymatic treatment of plant material (pH, enzyme concentration, time, and temperature) by response surface methodology (RSM). Medicago sativa L. was selected as a plant material due to its richness in phenolics and flavonoids. HPLC-MS/MS analysis allowed evaluating the content of individual bioactive compounds in obtained extracts. The total content of polyphenolic compounds in the extract obtained after two-step optimization was much higher (546 ± 21 µg/g) than in the extract obtained from non-hydrolyzed material (275 ± 23 µg/g) and in the extract obtained by maceration (162 ± 20 µg/g). Furthermore, it was evidenced that extract with the highest content of polyphenolic compounds can support the cellular antioxidant system both as a free radical scavenger and by stimulating the antioxidant enzyme system.

Spatial and Temporal Distribution of Arabinogalactan Proteins during Larix decidua Mill. Male Gametophyte and Ovule Interaction.

The role of ArabinoGalactan Proteins (AGPs) in the sexual reproduction of gymnosperms is not as well documented as that of angiosperms. In earlier studies, we demonstrated that AGPs play important roles during ovule differentiation in Larix decidua Mill. The presented results encouraged us to carry out further studies focused on the functions of these unique glycoproteins during pollen/pollen tube and ovule interactions in Larix. We identified and analyzed the localization of AGPs epitopes by JIM4, JIM8, JIM13 and LM2 antibodies (Abs) in male gametophytes and ovule tissue during pollination, the progamic phase, and after fertilization and in vitro growing pollen tubes. Our results indicated that (1) AGPs recognized by JIM4 Abs play an essential role in the interaction of male gametophytes and ovules because their appearance in ovule cells is induced by physical contact between reproductive partners; (2) after pollination, AGPs are secreted from the pollen cytoplasm into the pollen wall and contact the extracellular matrix of stigmatic tip cells followed by micropylar canal cells; (3) AGPs synthesized in nucellus cells before pollen grain germination are secreted during pollen tube growth into the extracellular matrix, where they can directly interact with male gametophytes; (4) in vitro cultured pollen tube AGPs labeled with LM2 Abs participate in the germination of pollen grain, while AGPs recognized by JIM8 Abs are essential for pollen tube tip growth.