Bioactive-Loaded Hydrogels Based on Bacterial Nanocellulose, Chitosan, and Poloxamer for Rebalancing Vaginal Microbiota.

Biocompatible drug-delivery systems for soft tissue applications are of high interest for the medical and pharmaceutical fields. The subject of this research is the development of hydrogels loaded with bioactive compounds (inulin, thyme essential oil, hydro-glycero-alcoholic extract of Vitis vinifera, Opuntia ficus-indica powder, lactic acid, citric acid) in order to support the vaginal microbiota homeostasis. The nanofibrillar phyto-hydrogel systems developed using the biocompatible polymers chitosan (CS), never-dried bacterial nanocellulose (NDBNC), and Poloxamer 407 (PX) incorporated the water-soluble bioactive components in the NDBNC hydrophilic fraction and the hydrophobic components in the hydrophobic core of the PX fraction. Two NDBNC-PX hydrogels and one NDBNC-PX-CS hydrogel were structurally and physical-chemically characterized using Fourier-transform infrared (FTIR) spectroscopy, X-ray diffraction (XRD), transmission electron microscopy (TEM), and rheology. The hydrogels were also evaluated in terms of thermo-responsive properties, mucoadhesion, biocompatibility, and prebiotic and antimicrobial effects. The mucin binding efficiency of hydrogel base systems was determined by the periodic acid/Schiff base (PAS) assay. Biocompatibility of hydrogel systems was determined by the MTT test using mouse fibroblasts. The prebiotic activity was determined using the probiotic strains Limosilactobacillus reuteri and Lactiplantibacillus plantarum subsp. plantarum. Antimicrobial activity was also assessed using relevant microbial strains, respectively, E. coli and C. albicans. TEM evidenced PX micelles of around 20 nm on NDBNC nanofibrils. The FTIR and XRD analyses revealed that the binary hydrogels are dominated by PX signals, and that the ternary hydrogel is dominated by CS, with additional particular fingerprints for the biocompounds and the hydrogel interaction with mucin. Rheology evidenced the gel transition temperatures of 18-22 °C for the binary hydrogels with thixotropic behavior and, respectively, no gel transition, with rheopectic behavior for the ternary hydrogel. The adhesion energies of the binary and ternary hydrogels were evaluated to be around 1.2 J/m2 and 9.1 J/m2, respectively. The hydrogels exhibited a high degree of biocompatibility, with the potential to support cell proliferation and also to promote the growth of lactobacilli. The hydrogel systems also presented significant antimicrobial and antibiofilm activity.

Sustainable Recovery of Anthocyanins and Other Polyphenols from Red Cabbage Byproducts.

The objective of this work was to develop a sustainable process for the extraction of anthocyanins from red cabbage byproducts using, for the first time, apple vinegar in extractant composition. Our results showed that the mixture 50% (v/v) ethanol-water, acidified with apple vinegar, used in the proportion of 25 g of red cabbage by-products per 100 mL of solvent, was the best solvent for the preparation of an anthocyanin extract with good stability for food applications. The chemical characterization of this extract was performed by FTIR, UV-VIS, HPLC-DAD, and ICP-OES. The stability was evaluated by determining the dynamics of the total polyphenol content (TPC) and the total monomeric anthocyanin pigment content (TAC) during storage. On the basis of the statistical method for analysis of variance (ANOVA), the standard deviation between subsamples and the repeatability standard deviation were determined. The detection limit of the stability test of TPC was 3.68 mg GAE/100 g DW and that of TAC was 0.79 mg Cyd-3-Glu/100 g DW. The red cabbage extract has high TPC and TAC, good stability, and significant application potential. The extracted residues, depleted of anthocyanins and polyphenols with potential allelopathic risks, fulfill the requirements for a fertilizing product and could be used for soil treatment.

Novel Strigolactone Mimics That Modulate Photosynthesis and Biomass Accumulation in Chlorella sorokiniana.

In terrestrial plants, strigolactones act as multifunctional endo- and exo-signals. On microalgae, the strigolactones determine akin effects: induce symbiosis formation with fungi and bacteria and enhance photosynthesis efficiency and accumulation of biomass. This work aims to synthesize and identify strigolactone mimics that promote photosynthesis and biomass accumulation in microalgae with biotechnological potential. Novel strigolactone mimics easily accessible in significant amounts were prepared and fully characterized. The first two novel compounds contain 3,5-disubstituted aryloxy moieties connected to the bioactive furan-2-one ring. In the second group of compounds, a benzothiazole ring is connected directly through the cyclic nitrogen atom to the bioactive furan-2-one ring. The novel strigolactone mimics were tested on Chlorella sorokiniana NIVA-CHL 176. All tested strigolactones increased the accumulation of chlorophyll b in microalgae biomass. The SL-F3 mimic, 3-(4-methyl-5-oxo-2,5-dihydrofuran-2-yl)-3H-benzothiazol-2-one (7), proved the most efficient. This compound, applied at a concentration of 10-7 M, determined a significant biomass accumulation, higher by more than 15% compared to untreated control, and improved the quantum yield efficiency of photosystem II. SL-F2 mimic, 5-(3,5-dibromophenoxy)-3-methyl-5H-furan-2-one (4), applied at a concentration of 10-9 M, improved protein production and slightly stimulated biomass accumulation. Potential utilization of the new strigolactone mimics as microalgae biostimulants is discussed.

The Antioxidant and Prebiotic Activities of Mixtures Honey/Biomimetic NaDES and Polyphenols Show Differences between Honeysuckle and Raspberry Extracts.

In our previous research, we demonstrated that honey and its biomimetic natural deep eutectic solvent (NaDES) modulate the antioxidant activity (AOA) of the raspberry extract (RE). In this study, we evaluated the AOA behaviour of the mixture honey/NaDES-honeysuckle (Lonicera caprifolium, LFL) extract and compared it with the mixture honey/NaDES-RE. These two extracts have similar major flavonoids and hydroxycinnamic acid compounds but differ in their total content and the presence of anthocyanins in RE. Therefore, it was of interest to see if the modulation of the LFL polyphenols by honey/NaDES was similar to that of RE. We also evaluated the prebiotic activity of these mixtures and individual components on Limosilactobacillus reuteri DSM 20016. Although honey/NaDES modulated the AOA of both extracts, from synergism to antagonism, the modulation was different between the two extracts for some AOA activities. Honey/NaDES mixtures enriched with LFL and RE did not show significant differences in bacterial growth stimulation. However, at a concentration of 45 mg/mL, the honey -LFL mixture exhibited a higher effect compared to the honey-RE mixture. The antioxidant and prebiotic properties of mixtures between honey and polyphenol-rich extracts are determined by multiple interactions in complex chemical systems.

Selenium-Fortified Kombucha-Pollen Beverage by In Situ Biosynthesized Selenium Nanoparticles with High Biocompatibility and Antioxidant Activity.

Biogenic selenium nanoparticles (SeNPs) have been shown to exhibit increased bioavailability. Fermentation of pollen by a symbiotic culture of bacteria and yeasts (SCOBY/Kombucha) leads to the release of pollen content and enhances the prebiotic and probiotic effects of Kombucha. The aim of this study was to fortify Kombucha beverage with SeNPs formed in situ by Kombucha fermentation with pollen. Response Surface Methodology (RSM) was used to optimize the biosynthesis of SeNPs and the pollen-fermented Kombucha beverage. SeNPs were characterized by Transmission electron microscopy energy-dispersive X-ray spectroscopy (TEM-EDX), Fourier-transform infrared spectroscopy (FTIR), Dynamic light scattering (DLS), and Zeta potential. The pollen-fermented Kombucha beverage enriched with SeNPs was characterized by measuring the total phenolic content, antioxidant activity, soluble silicon, saccharides, lactic acid, and the total content of Se0. The polyphenols were identified by liquid chromatography-mass spectrometry (LC-MS). The pollen and the bacterial (nano)cellulose were characterized by scanning electron microscopy-energy dispersive X-ray spectroscopy (SEM-EDX), FTIR, and X-Ray diffraction (XRD). We also assessed the in vitro biocompatibility in terms of gingival fibroblast viability and proliferation, as well as the antioxidant activity of SeNPs and the pollen-fermented Kombucha beverage enriched with SeNPs. The results highlight their increased biological performance in this regard.

Spectroscopic Analyses Highlight Plant Biostimulant Effects of Baker's Yeast Vinasse and Selenium on Cabbage through Foliar Fertilization.

The main aim of this study is to find relevant analytic fingerprints for plants' structural characterization using spectroscopic techniques and thermogravimetric analyses (TGAs) as alternative methods, particularized on cabbage treated with selenium-baker's yeast vinasse formulation (Se-VF) included in a foliar fertilizer formula. The hypothesis investigated is that Se-VF will induce significant structural changes compared with the control, analytically confirming the biofortification of selenium-enriched cabbage as a nutritive vegetable, and particularly the plant biostimulant effects of the applied Se-VF formulation on cabbage grown in the field. The TGA evidenced a structural transformation of the molecular building blocks in the treated cabbage leaves. The ash residues increased after treatment, suggesting increased mineral accumulation in leaves. X-ray diffraction (XRD) and Fourier-transform infrared spectroscopy (FTIR) evidenced a pectin-Iα-cellulose structure of cabbage that correlated with each other in terms of leaf crystallinity. FTIR analysis suggested the accumulation of unesterified pectin and possibly (seleno) glucosinolates and an increased network of hydrogen bonds. The treatment with Se-VF formulation induced a significant increase in the soluble fibers of the inner leaves, accompanied by a decrease in the insoluble fibers. The ratio of soluble/insoluble fibers correlated with the crystallinity determined by XRD and with the FTIR data. The employed analytic techniques can find practical applications as fast methods in studies of the effects of new agrotechnical practices, while in our particular case study, they revealed effects specific to plant biostimulants of the Se-VF formulation treatment: enhanced mineral utilization and improved quality traits.

Closing the Nutrient Loop-The New Approaches to Recovering Biomass Minerals during the Biorefinery Processes.

The recovery of plant mineral nutrients from the bio-based value chains is essential for a sustainable, circular bioeconomy, wherein resources are (re)used sustainably. The widest used approach is to recover plant nutrients on the last stage of biomass utilization processes-e.g., from ash, wastewater, or anaerobic digestate. The best approach is to recover mineral nutrients from the initial stages of biomass biorefinery, especially during biomass pre-treatments. Our paper aims to evaluate the nutrient recovery solutions from a trans-sectorial perspective, including biomass processing and the agricultural use of recovered nutrients. Several solutions integrated with the biomass pre-treatment stage, such as leaching/bioleaching, recovery from pre-treatment neoteric solvents, ionic liquids (ILs), and deep eutectic solvents (DESs) or integrated with hydrothermal treatments are discussed. Reducing mineral contents on silicon, phosphorus, and nitrogen biomass before the core biorefinery processes improves processability and yield and reduces corrosion and fouling effects. The recovered minerals are used as bio-based fertilizers or as silica-based plant biostimulants, with economic and environmental benefits.

Cytocompatibility, Antimicrobial and Antioxidant Activity of a Mucoadhesive Biopolymeric Hydrogel Embedding Selenium Nanoparticles Phytosynthesized by Sea Buckthorn Leaf Extract.

Phytosynthesized selenium nanoparticles (SeNPs) are less toxic than the inorganic salts of selenium and show high antioxidant and antibacterial activity. Chitosan prevents microbial biofilm formation and can also determine microbial biofilm dispersal. Never-dried bacterial nanocellulose (NDBNC) is an efficient carrier of bioactive compounds and a flexible nanofibrillar hydrophilic biopolymer. This study aimed to develop a selenium-enriched hydrogel nanoformulation (Se-HNF) based on NDBNC from kombucha fermentation and fungal chitosan with embedded biogenic SeNPs phytosynthesized by an aqueous extract of sea buckthorn leaves (SbLEx)-SeNPsSb-in order to both disperse gingival dysbiotic biofilm and prevent its development. We determined the total phenolic content and antioxidant activity of SbLEx. Liquid chromatography-mass spectrometry (LC-MS) and high-performance liquid chromatography (HPLC) were used for the identification of polyphenols from SbLEx. SeNPsSb were characterized by transmission electron microscopy-energy-dispersive X-ray spectroscopy (TEM-EDX), dynamic light scattering (DLS), zeta potential, Fourier transform infrared spectroscopy (FTIR) and X-ray diffraction (XRD) in small- and wide-angle X-ray scattering (SAXS and WAXS). The hydrogel nanoformulation with embedded SeNPsSb was characterized by SEM, FTIR, XRD, rheology, mucin binding efficiency, contact angle and interfacial tension measurements. We also assessed the in vitro biocompatibility, antioxidant activity and antimicrobial and antibiofilm potential of SeNPsSb and Se-HNF. TEM, DLS and SAXS evidenced polydisperse SeNPsSb, whereas FTIR highlighted a heterogeneous biocorona with various biocompounds. The contact angle on the polar surface was smaller (52.82 ± 1.23°) than that obtained on the non-polar surface (73.85 ± 0.39°). The interfacial tension was 97.6 ± 0.47 mN/m. The mucin binding efficiency of Se-HNF decreased as the amount of hydrogel decreased, and the SEM analysis showed a relatively compact structure upon mucin contact. FTIR and XRD analyses of Se-HNF evidenced an interaction between BNC and CS through characteristic peak shifting, and the rheological measurements highlighted a pseudoplastic behavior, 0.186 N adhesion force and 0.386 adhesion energy. The results showed a high degree of cytocompatibility and the significant antioxidant and antimicrobial efficiency of SeNPsSb and Se-HNF.

Honey and Its Biomimetic Deep Eutectic Solvent Modulate the Antioxidant Activity of Polyphenols.

Honey is a highly valued natural product with antioxidant, antimicrobial and anti-inflammatory properties. However, its antioxidant activity (AOA) is not as high as that of other honeybee products, such as propolis. Several polyphenol-honey formulations have been proposed up to now, most of them using maceration of biomass in honey or mixtures with liquid extracts, which either limit polyphenols bioavailability or destroy the characteristics of honey. To improve the health benefits of honey by increasing AOA and keeping its structural and sensory properties, we propose its enrichment in a polyphenol extract of raspberry after solvent evaporation. A honey-biomimetic natural deep eutectic solvent (NaDES) was prepared and compared with honey. The main polyphenols found in the raspberry extract were tested in combination with honey and NaDES, respectively. The AOA was determined by DPPH, ABTS, CUPRAC, and FRAP methods. The AOA behaviour of honey-polyphenol mixtures varied from synergism to antagonism, being influenced by the AOA method, polyphenol type, and/or mixture concentration. The honey-biomimetic NaDES resulted in similar AOA behaviour as with honey mixed with polyphenols. Honey seems to have additional properties that increase synergism or reduce antagonism in some cases. Honey and its biomimetic NaDES modulate AOA of polyphenols extract.

Humic Substances as Microalgal Biostimulants-Implications for Microalgal Biotechnology.

Humic substances (HS) act as biostimulants for terrestrial photosynthetic organisms. Their effects on plants are related to specific HS features: pH and redox buffering activities, (pseudo)emulsifying and surfactant characteristics, capacity to bind metallic ions and to encapsulate labile hydrophobic molecules, ability to adsorb to the wall structures of cells. The specific properties of HS result from the complexity of their supramolecular structure. This structure is more dynamic in aqueous solutions/suspensions than in soil, which enhances the specific characteristics of HS. Therefore, HS effects on microalgae are more pronounced than on terrestrial plants. The reported HS effects on microalgae include increased ionic nutrient availability, improved protection against abiotic stress, including against various chemical pollutants and ionic species of potentially toxic elements, higher accumulation of value-added ingredients, and enhanced bio-flocculation. These HS effects are similar to those on terrestrial plants and could be considered microalgal biostimulant effects. Such biostimulant effects are underutilized in current microalgal biotechnology. This review presents knowledge related to interactions between microalgae and humic substances and analyzes the potential of HS to enhance the productivity and profitability of microalgal biotechnology.

Effects of Siliceous Natural Nanomaterials Applied in Combination with Foliar Fertilizers on Physiology, Yield and Fruit Quality of the Apricot and Peach Trees.

Siliceous natural nanomaterials (SNNMs), i.e., diatomaceous earth and natural zeolites, have a nanoporous structure with large active surfaces that adsorb cations or polarized molecules. Such nanoporous feature determines the effects related to SNNM utilization as low-risk plant protectants and soil improvers. This work used SNNMs from Romanian quarries as carriers for foliar fertilizers applied to stone-fruit trees, apricot and peach. We determined the effects of SNNMs on the physiology, yield and fruit quality of the treated stone-fruit trees. SNNM application determined impacts specific to the formation of particle films on leaves: reduced leaf temperature (up to 4.5 °C) and enhanced water use efficiency (up to 30%). Foliar fertilizers' effects on yield are amplified by their application with SNNMs. Yield is increased up to 8.1% by the utilization of SNNMs with foliar fertilizers, compared to applying foliar fertilizer alone. Diatomaceous earth and natural zeolites promote the accumulation of polyphenols in apricot and peach fruits. The combined application of SNNMs and foliar fertilizer enhance the performance of peach and apricot trees.

The Scavenging Effect of Myoglobin from Meat Extracts toward Peroxynitrite Studied with a Flow Injection System Based on Electrochemical Reduction over a Screen-Printed Carbon Electrode Modified with Cobalt Phthalocyanine: Quantification and Kinetics.

The scavenging activity of myoglobin toward peroxynitrite (PON) was studied in meat extracts, using a new developed electrochemical method (based on cobalt phthalocyanine-modified screen-printed carbon electrode, SPCE/CoPc) and calculating kinetic parameters of PON decay (such as half-time and apparent rate constants). As reactive oxygen/nitrogen species (ROS/RNS) affect the food quality, the consumers can be negatively influenced. The discoloration, rancidity, and flavor of meat are altered in the presence of these species, such as PON. Our new highly thermically stable, cost-effective, rapid, and simple electrocatalytical method was combined with a flow injection analysis system to achieve high sensitivity (10.843 nA µM-1) at a nanomolar level LoD (400 nM), within a linear range of 3-180 µM. The proposed biosensor was fully characterized using SEM, FTIR, Raman spectroscopy, Cyclic Voltammetry (CV), Differential Pulse Voltammetry (DPV), and Linear Sweep Voltammetry (LSV). These achievements were obtained due to the CoPc-mediated reduction of PON at very low potentials (around 0.1 V vs. Ag/AgCl pseudoreference). We also proposed a redox mechanism involving two electrons in the reduction of peroxynitrite to nitrite and studied some important interfering species (nitrite, nitrate, hydrogen peroxide, dopamine, ascorbic acid), which showed that our method is highly selective. These features make our work relevant, as it could be further applied to study the kinetics of important oxidative processes in vivo or in vitro, as PON is usually present in the nanomolar or micromolar range in physiological conditions, and our method is sensitive enough to be applied.

Effects of Foliar Treatment with a Trichoderma Plant Biostimulant Consortium on Passiflora caerulea L. Yield and Quality.

The influence of spore concentration on the ability of a Trichoderma consortium to colonize the Passiflora caerulea phyllosphere was evaluated by determining the effects of foliar treatments with two spore concentrations, in two repeated treatments, on the morphological, physiological, and ultrastructural characteristics, and on the yield and quality of P. caerulea. The studied crop quality features were related to its nutraceutical use: the accumulation of polyphenols and flavonoids, antioxidant activity, and effects on mouse fibroblast L929 cells. The Trichoderma consortium consisted of two strains, T. asperellum T36b and T. harzianum Td50b, and the concentrations used were 106 colony forming units (cfu)/mL and 108 cfu/mL. As a reference treatment, a commercial product that was based on herbs and algal extracts was used. As compared to the negative control, the treatment with the Trichoderma consortium at 108 cfu/mL concentration determines the accumulation of higher level of polyphenols and flavonoids and increased antioxidant activity. This enhancement of P. caerulea quality characteristics after treatment with the higher concentration of Trichoderma consortium was associated with larger leaves, increased number and size of chloroplasts, improved plant physiology characteristics, and an increased yield. The treatment with high concentration of Trichoderma consortium spores promotes phyllosphere colonization and benefits both crop yield and quality.

Effect of betaine and selenium on the growth and photosynthetic pigment production in Dunaliella salina as biostimulants.

The aim of our study was to establish the effect of selenium and betaine on the growth of D. salina, accumulation of photosynthetic pigments and antioxidant activity of the hydrophobic fraction. This approach was an attempt to demonstrate 'microalgae biostimulant' effects, similar to 'plant biostimulant' effects, i.e. increased tolerance to abiotic stress and enhanced accumulation of bioactive compounds. A high-throughput assay was done in 24-well microplates, at 15% NaCl and different concentrations of sodium selenite (0, 0.5, 2 and 8 µM) or betaine (0, 5, 50 and 500 µM). Both selenium and betaine induced a slight delay in algae growth during the actively growing stage but the final density reached similar values to the control. Betaine significantly enhanced (50%-100%) carotenoids and chlorophyll a accumulation, in a concentration depending manner. Antioxidant activity increased almost 3-fold in extracts of algae treated with 50 µM betaine. Selenium had a much more discrete effect than betaine on pigments biosynthesis. The antioxidant activity of the extracts increased 2-fold in the presence of Se compared to the control. Our work proves that it is possible to enhance production and activity of bioactive compounds from microalgae by using ingredients, which already proved to act as plant biostimulants.

Selenium Analysis and Speciation in Dietary Supplements Based on Next-Generation Selenium Ingredients.

Selenium is essential for humans and the deficit of Se requires supplementation. In addition to traditional forms such as Se salts, amino acids, or selenium-enriched yeast supplements, next-generation selenium supplements, with lower risk for excess supplementation, are emerging. These are based on selenium forms with lower toxicity, higher bioavailability, and controlled release, such as zerovalent selenium nanoparticles (SeNPs) and selenized polysaccharides (SPs). This article aims to focus on the existing analytical systems for the next-generation Se dietary supplement, providing, at the same time, an overview of the analytical methods available for the traditional forms. The next-generation dietary supplements are evaluated in comparison with the conventional/traditional ones, as well as the analysis and speciation methods that are suitable to reveal which Se forms and species are present in a dietary supplement. Knowledge gaps and further research potential in this field are highlighted. The review indicates that the methods of analysis of next-generation selenium supplements should include a step related to chemical species separation. Such a step would allow a proper characterization of the selenium forms/species, including molecular mass/dimension, and substantiates the marketing claims related to the main advantages of these new selenium ingredients.

Bee Collected Pollen with Enhanced Health Benefits, Produced by Fermentation with a Kombucha Consortium.

The bioavailability of pollen bioactive compounds for humans is limited. In this study, our aim was to enhance the health-related benefits of pollen by fermentation with a Kombucha/SCOBY (symbiotic culture of bacteria and yeasts) consortium. We performed the fermentation of pollen suspended from the beginning with SCOBY on sweetened green tea or on Kombucha vinegar, by adding pollen after 20 days of Kombucha fermentation. We analyzed: formation of bioactive compounds (anti-oxidant polyphenols, soluble silicon, hydroxy-acids, short chain fatty acids-SCFA); parameters related to Kombucha fermentation (dynamics of lactic acid bacteria-LAB, formation of organic acids, soluble sugar evolution on Kombucha vinegar); the influence of Kombucha fermentation on pollen morphology and ultrastructure; in vitro cytotoxic and antitumoral effects of the Kombucha fermented pollen. The pollen addition increases LAB proportion in the total number of SCOBY microbial strains. SEM images highlight the adhesion of the SCOBY bacteria to pollen. Ultrastructural analysis reveals the release of the pollen content. The content of bioactive compounds (polyphenols, soluble silicon species and SCFA) is higher in the fermented pollen and the product shows a moderate antitumoral effect on Caco-2 cells. The health benefits of pollen are enhanced by fermentation with a Kombucha consortium.

Degradation of keratin substrates by keratinolytic fungi

Background: The hydrolysis of keratin wastes by microorganisms is considered a biotechnological alternative for recycling and valorization through keratinolytic microorganisms. Despite their resistant structure, keratin wastes can be efficiently degraded by various microorganisms through the secretion of keratinases, which are promising enzymes for several applications, including detergents, fertilizers, and leather and textile industry. In an attempt to isolate keratinolytic microorganisms that can reach commercial exploitation as keratinase producers, the current work assesses the dynamics of keratin biodegradation by several keratinolytic fungal strains isolated from soil. The activity of fungal strains to degrade keratin substrates was evaluated by SEM, FTRIR-ATR spectra and TGA analysis. Results: SEM observations offered relevant information on interactions between microorganism and structural elements of hair strands. FTIR spectra of the bands at 1035­1075 cm-1 assigned to sulfoxide bond appeared because of S­S bond breaking, which demonstrated the initiation of keratin biodegradation. According to TGA, in the second zone of thermal denaturation, where keratin degradation occurs, the highest weight loss of 71.10% was obtained for sample incubated with Fusarium sp. 1A. Conclusions: Among the tested strains, Fusarium sp. 1A was the most active organism in the degradation process with the strongest denaturation of polypeptide chains. Because keratinolytic microorganisms and their enzymes keratinases represent a subject of scientific and economic interest because of their capability to hydrolyze keratin, Fusarium sp. 1A was selected for further studies.

Kinetic characterization of apoptotic Ras signaling through Nore1-MST1 complex formation.

Ras-mediated apoptotic signaling is expected to be mediated via Rassf-MST complexes, but the system has been poorly characterized in vitro until now. Here we demonstrate that active H-Ras, Nore1A and MST1 form a stable ternary complex in vitro without other external factors, Nore1A interacting simultaneously with H-Ras and MST1 via its RBD and SARAH domain, respectively. Moreover, our data show for the first time that the SARAH domain of Nore1A plays a role in the Nore1A binding to H-Ras. Finally, we analyze the relation between the electrostatic and hydrophobic forces and kinetic constants of the Nore1A - H-Ras complex.

The temperature dependence of the Hofmeister series: thermodynamic fingerprints of cosolute-protein interactions.

The Hofmeister series is a universal homologous series to rank ion-specific effects on biomolecular properties such as protein stability or aggregation propensity. Although this ranking is widely used, outliers and exceptions are discussed controversially and a molecular level understanding is still lacking. Studying the thermal unfolding equilibrium of RNase A, we here show that this ambiguity arises from the oversimplified approach to determine the ion rankings. Instead of measuring salt effects on a single point of the protein folding stability curve (e.g. the melting point Tm), we here consider the salt induced shifts of the entire protein 'stability curve' (the temperature dependence of the unfolding free energy change, ΔGu(T)). We found multiple intersections of these curves, pinpointing a widely ignored fact: the Hofmeister cation and anion rankings are temperature dependent. We further developed a novel classification scheme of cosolute effects based on their thermodynamic fingerprints, reaching beyond salt effects to non-electrolytes.

Peroxynitrite Sensor Based on a Screen Printed Carbon Electrode Modified with a Poly(2,6-dihydroxynaphthalene) Film.

For the first time the electropolymerization of 2,6-dihydroxynaphthalene (2,6-DHN) on a screen printed carbon electrode (SPCE) was investigated and evaluated for peroxynitrite (PON) detection. Cyclic voltammetry was used to electrodeposit the poly(2,6-DHN) on the carbon electrode surface. The surface morphology and structure of poly(2,6-DHN) film were investigated by SEM and FTIR analysis, and the electrochemical features by cyclic voltammetry. The poly(2,6-DHN)/SPCE sensor showed excellent electrocatalytic activity for PON oxidation in alkaline solutions at very low potentials (0-100 mV vs. Ag/AgCl pseudoreference). An amperometric FIA (flow injection analysis) system based on the developed sensor was optimized for PON measurements and a linear concentration range from 2 to 300 µM PON, with a LOD of 0.2 µM, was achieved. The optimized sensor inserted in the FIA system exhibited good sensitivity (4.12 nA·µM-1), selectivity, stability and intra-/inter-electrode reproducibility for PON determination.