Improvement of fruit juice quality: novel endo-polygalacturonase II from Aspergillus tubingensis FAT 43 for enhanced liquefaction, clarification, and antioxidant potential.

This study focuses on the isolation, purification, and characterisation of endo-polygalacturonase II from Aspergillus tubingensis FAT43, particularly emphasising its potential applications in the fruit juice industry. A comprehensive screening test revealed the temporal dynamics of endo-polygalacturonase production during a 96-hour fermentation process. The purification process, involving ammonium sulfate and ethanol precipitation followed by ion-exchange chromatography, resulted in a 3.3-fold purification of PG II with a yield of 16% and a specific activity of 6001.67 U mg-1. Molecular analysis confirmed the identity of PG II, its gene (pgaII), and a high degree of sequence identity with Aspergillus tubingensis in the SWISS-PROT database. The optimal pH for PG II activity was 3.5-4.5, with robust stability across a broad pH spectrum (3-7). The enzyme exhibited optimal temperature activity at 45 °C, with a retention of 90% activity at 50 °C. The calculated activation energy for PG II was 62.1 kJ mol-1, indicating good stability. Inactivation kinetics revealed a half-life of 13.7 h at 40 °C, 5.4 h at 50 °C, and 0.85 h at 60 °C, with an activation energy of denaturation of 32.8 kJ mol-1. Compared to literature-reported PGs, PG II from A. tubingensis FAT43 demonstrated superior thermal stability. Hydrolysis experiments on different pectins revealed the highest specificity for non-methylated substrates (polygalacturonic acid). In fruit juice processing, PG II significantly increased juice yield and clarity, with the highest impact observed in strawberry juice. Antioxidant activity assays indicated enhanced antioxidant potential in enzyme-treated juices, especially strawberry, quince, and apple juices. The study highlights PG II's potential as an industrially valuable enzyme for fruit juice processing, offering improved thermostability and versatility across various fruit types.

Optimization of solid-state fermentation for enhanced production of pectinolytic complex by Aspergillus tubingensis FAT43 and its application in fruit juice processing.

The main goal of this study was to examine the efficiency of a newly isolated fungus from quince, Aspergillus tubingensis FAT43, to produce the pectinolytic complex using agricultural and industrial waste as the substrate for solid state fermentation. Sugar beet pulp was the most effective substrate inducer of pectinolytic complex synthesis out of all the waste residues examined. For endo-pectinolytic and total pectinolytic activity, respectively, statistical optimization using Placked-Burman Design and Optimal (Custom) Design increased production by 2.22 and 2.15-fold, respectively. Liquification, clarification, and an increase in the amount of reducing sugar in fruit juices (apple, banana, apricot, orange, and quince) processed with pectinolytic complex were identified. Enzymatic pre-treatment considerably increases yield (14%-22%) and clarification (90%). After enzymatic treatment, the best liquefaction was observed in orange juice, whereas the best clarification was obtained in apricot juice. Additionally, the pectinolytic treatment of apricot juice resulted in the highest increase in reducing sugar concentration (11%) compared to all other enzymatically treated juices. Optimizing the production of a highly active pectinolytic complex and its efficient utilization in the processing of fruit juices, including the generation of an increasing amount of waste, are the significant outcomes of this research.

BrainLine: An Open Pipeline for Connectivity Analysis of Heterogeneous Whole-Brain Fluorescence Volumes.

Whole-brain fluorescence images require several stages of computational processing to fully reveal the neuron morphology and connectivity information they contain. However, these computational tools are rarely part of an integrated pipeline. Here we present BrainLine, an open-source pipeline that interfaces with existing software to provide registration, axon segmentation, soma detection, visualization and analysis of results. By implementing a feedback based training paradigm with BrainLine, we were able to use a single learning algorithm to accurately process a diverse set of whole-brain images generated by light-sheet microscopy. BrainLine is available as part of our Python package brainlit: http://brainlit.neurodata.io/ .

Chemical Fractionation, Environmental, and Human Health Risk Assessment of Potentially Toxic Elements in Soil of Industrialised Urban Areas in Serbia.

The primary focus of this research was the chemical fractionation of potentially toxic elements (PTEs) and their presence in several industrialised cities in Serbia. Furthermore, their origin, contamination levels, and environmental and human health risks were assessed. The results indicated that the examined soils were characterised by slightly higher Cu, Ni, Pb, and Zn levels than those set by European and national regulations. These elevated Cu, Pb, and Zn concentrations were caused by intensive traffic and proximity to industry, whereas the higher Ni levels were a result of the specific geological substrate of the soil in the study area. The environmental risk was found to be low and there was no enrichment/contamination of the soil with these elements, except in the case of Pb, for which moderate to significant enrichment was found. Lead also poses a potential non-carcinogenic risk to children through ingestion and requires special attention due to the fact that a significant proportion of this element was present in the tested soil samples in a potentially available form. Analysis of the health risks showed that children are more at risk than adults from contaminants and that ingestion is the riskiest exposure route. The carcinogenic risk was within the acceptable limits.

Fractionation of Potentially Toxic Elements (PTEs) in Urban Soils from Salzburg, Thessaloniki and Belgrade: An Insight into Source Identification and Human Health Risk Assessment.

Concentrations of potentially toxic elements (PTEs) (Al, As, Cd, Cr, Cu, Ni, Pb, and Zn) were measured in topsoil samples collected from parks in the cities of Salzburg (Austria), Thessaloniki (Greece), and Belgrade (Serbia) in order to assess the distribution of PTEs in the urban environment, discriminate natural (lithogenic) and anthropogenic contributions, identify possible sources of pollution, and compare levels of pollution between the cities. An assessment of the health risks caused by exposure to PTEs through different pathways was also conducted. The study revealed that, with the exception of Pb in Salzburg, levels of PTEs in the soils in polluted urban parks were higher than in unpolluted ones, but still lower than those recorded in other European soils. Results of sequential analyses showed that Al, Cr, and Ni were found in residual phases, proving their predominantly lithogenic origin and their low mobility. In contrast, the influence of anthropogenic factors on Cu, Pb, and Zn was evident. Site-dependent variations showed that the highest concentrations of As, Cu, Pb, and Zn of anthropogenic origin were recorded in Salzburg, while the highest levels of Al, Cr, and Ni of lithogenic origin were recorded in Belgrade and Thessaloniki, which reflects the specificity of the geological substrates. Results obtained for the health risk assessment showed that no human health risk was found for either children or adults.

Evaluation of Salix alba, Juglans regia and Populus nigra as biomonitors of PTEs in the riparian soils of the Sava River.

A large number of human activities result in the release of potentially toxic elements (PTEs) into the environment, which could lead to the degradation of riparian areas. This study aimed to evaluate the potential of Salix alba, Juglans regia and Populus nigra for the biomonitoring of PTEs in the riparian soils of the Sava River. Levels of seven PTEs (As, Cd, Cr, Cu, Ni, Pb and Zn) were measured in the soils, roots and leaves of plants at selected sampling sites and evaluated according to bioaccumulation and translocation factors. The obtained results showed that in riparian soils, As, Cr, Cu, Ni and Zn were at levels considered to be critical for plants. The levels of As, Cd, Cr, Ni and Zn measured in roots of Salix alba and As, Cr, Ni and Zn in its leaves were toxic for plant tissue. Toxic levels of Cr were also measured in the roots of Juglans regia and As in its leaves, as well as As and Cr in the roots of Populus nigra, and Zn in its leaves. Bioconcentration and translocation factors showed that S. alba and P. nigra have potential for the phytoextraction of Zn and Cd, while J. regia has potential for the phytoextraction of As. In terms of phytostabilization potential, S. alba proved to be good for the phytostabilization of Cd and Cu, and J. regia for the phytostabilization of Cr, As, Ni and Pb, while P. nigra showed potential for the phytostabilization of Cr, Ni, Pb and Cu.

Sources and a Health Risk Assessment of Potentially Toxic Elements in Dust at Children's Playgrounds with Artificial Surfaces: A Case Study in Belgrade.

The focus of this research on children's playgrounds with artificial surfaces aimed to establish levels of potentially toxic elements (PTEs) in dust, their origin, and impact on children at 15 playgrounds: 9 on school grounds and 6 on day nurseries in Belgrade (Serbia). Soil samples were taken from the immediate vicinity of the playgrounds to establish the origin of PTEs in the dust samples. Soil analyses revealed the lithogenic origin of Co, Cr, Ni, Fe, Mn, As, Cd, Cu, Pb, and Zn and the anthropogenic origin of As, Cd, Cu, Pb, and Zn. However, in the dust samples, the origin of the elements was different with As, Co, Fe, and Mn originating from the surrounding soil; Cr and Ni levels affected by both atmospheric deposition and the surrounding soil; Cd, Pb, and Zn concentrations impacted by atmospheric deposition; and Cu levels affected by factors of a local character. No noncancer risk was found for any of the individual elements investigated, nor for any of the playgrounds being studied, while a minimal cancer risk was found from As with values greater than 1E-6 at almost all the sites. Based on the results obtained for the spatial distribution of individual PTE levels, it was determined that the surrounding soil and atmospheric deposition have an almost equal impact on noncancer risk values.

Influence of breed on selected quality parameters of fresh goat meat.

The potential of goats to produce a high-quality meat is mainly reflected in their healthy fats, low calorie intramuscular fats, saturated fats, and, especially, their high ratios of unsaturated (UFA) and saturated (SFA) fatty acids, as well as hypocholesterolemic and hypercholesterolemic fatty acids. The aim of this study was to collect and compare meat quality parameters for domestic Balkan, Alpine and Saanen goats of the same age. Samples for all tests were taken from musculus gluteus superficialis. Chemical composition, pH value, fatty acid composition, content of volatile compounds, color and overall sensory quality (appearance, texture and smell) were determined. In chemical composition, moisture, fat, protein and ash varied significantly between each of the examined groups as opposed to pH values. Furthermore, among all the examined groups a significant difference was found for fatty acids and volatile compounds. Determined ratio of polyunsaturated fatty acids (PUFAs) to SFAs was 0.089, 0.085 and 0.071 for Balkan, Alpine and Saanen goat meats, respectively. Regarding that ratio, Saanen goat meat had the most favorable characteristics. Saanen goat meat showed the highest nutritional value. On the other hand, Balkan goat meat had the lowest intramuscular fat content. Measurements of the meat color from all three groups, as well as overall acceptability, showed significant differences between breeds. Obtained results point to the impact of breed on chemical composition and fatty acid profile of goat meat.

Fractionation, Mobility, and Contamination Assessment of Potentially Toxic Metals in Urban Soils in Four Industrial Serbian Cities.

The main soil properties, concentrations of selected elements (As, Cd, Cr, Cu, Fe, Mn, Ni, Pb, and Zn), and the chemical speciation of each element were determined in urban soil samples taken from urban parks in four Serbian cities (Belgrade, Pancevo, Obrenovac, and Smederevo) exposed to different sources of pollution. Pollution indices (PI, PIN) and factors (MF, ICF, GCF) also were evaluated. The study revealed As and Cd concentrations below the detection limit, whereas the content of Cr, Cu, Fe, Mn, Ni, Pb, and Zn at some sites exceeded the limits established by local regulations, as well as the background values, which may represent an environmental threat. Sequential extraction results show that Fe, Cr, Cu, and Ni were predominantly in the residual fraction at most sites; however, Ni from Pancevo and Smederevo also was bound to the reducible fraction. The presence of Pb at all sites and Zn in Smederevo and Belgrade was mainly associated with the reducible and residual fractions. The highest Mn content was found in the reducible fraction, followed by the acid soluble/exchangeable and residual fractions. Based on the obtained indices and factors, the overall soil status at the selected sampling sites was found to range from the warning limit to slightly polluted, whereby Smederevo had the highest risk, and Pancevo and the control site the lowest risk of contamination by toxic metals.

Traditional wound-healing plants used in the Balkan region (Southeast Europe).

ETHNOPHARMACOLOGICAL RELEVANCE: The geographical and ecological specificity of the Balkan Peninsula has resulted in the development of a distinct diversity of medicinal plants. In the traditional culture of the Balkan peoples, plants have medicinal, economic and anthropological/cultural importance, which is reflected in the sound knowledge of their diversity and use. This study analyses the traditional use of medicinal plants in the treatment of wounds and the pharmacological characteristics of the most frequently used species. MATERIALS AND METHODS: A detailed analysis of the literature related to ethnobhe uses of medicinal plants in the Balkan region was carried out. Twenty-five studies were analysed and those plants used for the treatment of wounds were singled out. RESULT: An ethnobotanical analysis showed that 128 plant species (105 wild, 22 cultivated and 1 wild/cultivated) are used in the treatment of wounds. Their application is external, in the form of infusions, decoctions, tinctures, syrups, oils, ointments, and balms, or direct to the skin. Among those plants recorded, the most commonly used are Plantago major, Hypericum perforatum, Plantago lanceolata, Achillea millefolium, Calendula officinalis, Sambucus nigra, Tussilago farfara and Prunus domestica. The study showed that the traditional use of plants in wound healing is confirmed by in vitro and/or in vivo studies for P. major and P. lanceolata (3 laboratory studies for P. major and 2 for P. lanceolata), H. perforatum (5 laboratory studies and 3 clinical trials), A. millefolium (3 laboratory studies and one clinical trial), C. officinalis (6 laboratory studies and 1 clinical trial), S. nigra (3 laboratory studies) and T. farfara (one laboratory study). CONCLUSION: The beneficial effects of using medicinal plants from the Balkan region to heal wounds according to traditional practices have been proven in many scientific studies. However, information on the quantitative benefits to human health of using herbal medicines to heal wounds is still scarce or fragmented, hindering a proper evaluation. Therefore, further studies should be aimed at isolating and identifying specific active substances from plant extracts, which could also reveal compounds with more valuable therapeutic properties. Furthermore, additional reliable clinical trials are needed to confirm those experiences encountered when using traditional medicines. A combination of traditional and modern knowledge could result in new wound-healing drugs with a significant reduction in unwanted side effects.

Customizing the spent coffee for Trichoderma reesei cellulase immobilization by modification with activating agents.

This study has explored the possibility to reuse the waste, spent coffee material for the cellulase enzyme immobilization. By the coffee surface modification with different activating agents, it was attempted to develop the convenient method for creation of a capable porous carrier for this purpose. Among the most common activating agents, glutaraldehyde, chlorine dioxide and hydrogen peroxide provided the most acceptable choice for the coffee surface modification. The changes that occurred on the coffee surface due to agents' treatment exposure were recorded by using of the FTIR spectra and SEM micrographs. The highest immobilization yield (55%) and immobilization efficiency (45%) were attained during 30min of the treatment time, by employing of 30% chlorine dioxide aqueous solution within 6mL/g activator/carrier ratio. The kinetic process was found to be predicted by the pseudo-second-order model. The cellulase immobilization onto the coffee surface provides an excellent base for increasing the enzyme availability to the substrate and enhancing the enzyme productivity, by offering the new perspectives to the industrial sector.

Plant growth-promoting bacteria elevate the nutritional and functional properties of black cumin and flaxseed fixed oil.

BACKGROUND: In order to study the influence of plant growth-promoting bacteria (PGPB) belonging to Streptomyces sp., Paenibacillus sp., and Hymenobacter sp. on fixed oil content of flaxseed and black cumin, 2-year field experiments were conducted. PGPB was applied during seedtime of plants. The extraction of oil from seeds was performed using supercritical CO2 . RESULTS: The addition of PGPB significantly increases the content of C18:1 (from 16.06 ± 0.03% to 16.97 ± 0.03%) and C18:3 (from 42.97 ± 0.2% to 45.42 ± 0.5%) in flaxseed oil and C18:2 (from 52.68 ± 0.50% to 57.11 ± 0.40%) and C20:2 (from 4.34 ± 0.02% to 4.54 ± 0.03%) in black cumin seed oil. The contents of total polyphenols, flavonoids, and carotenoids, as well as antioxidant activity measured by ferric-reducing ability of plasma assay, were found to be greater in the oil from the seeds of plants treated with the PGPB, compared with the respective non-treated samples. CONCLUSION: The use of PGPB enhances plant nutritive properties; these represent a great source for obtaining valuable functional food ingredients. © 2017 Society of Chemical Industry.

Utilization of spent coffee grounds for isolation and stabilization of Paenibacillus chitinolyticus CKS1 cellulase by immobilization.

This study has explored the feasibility of using spent coffee grounds as a good supporting material for the Paenibacillus chitinolyticus CKS1 cellulase immobilization. An optimal operational conditions in a batch-adsorption system were found to be: carrier mass of 12 g/L, under the temperature of 45 °C and no pH adjustments. The immobilization yield reached about 71%. An equilibrium establishment between the cellulase and the carrier surface occurred within 45 min, whereas the process kinetics may be predicted by the pseudo-second-order model. An immobilized cellulase preparation expressed very good avicelase activity, this reached up to 2.67 U/g, and revealed an improved storage stability property, compared to free enzyme sample counterpart. The addition of metal ions, such as K(+) and Mg(2+) did not affect positively immobilization yield results, but on the contrary, contributed to an improved bio-activities of the immobilized cellulase, thus may be employed before each enzyme application. The method developed in this study offers a cheap and effective alternative for immediate enzyme isolation from the production medium and its stabilization, compared to other carriers used for the immobilization.

Identification and characterization of antioxidant peptides obtained by gastrointestinal digestion of amaranth proteins.

The objective of the present work was to separate and identify antioxidant peptides from a simulated gastrointestinal digest (Id) from Amaranthus mantegazzianus proteins (I), which has previously been demonstrated to have this activity. I and Id were separated by preparative RP-HPLC. Fractions were evaluated by the ORAC method and the more active ones were analyzed by LC-MS/MS. Each fraction presented diverse peptides from different proteins, most of them from the 11S globulin. After grouping the peptides from 11S globulin according to their overlapping sequences, and based on previous information about structure-activity relationships, ten sequences were synthesized, in order to evaluate their antioxidant activity. Four peptides presented interesting activity: AWEEREQGSR>YLAGKPQQEH∼IYIEQGNGITGM∼TEVWDSNEQ. They exhibited some of the structural characteristics already known to demonstrate this activity, all of them containing at least one bulky aromatic residue. All belonged to little structured, internal or exposed regions of the acid subunit of the 11S globulin.

Understanding the Remodelling of Cell Walls during Brachypodium distachyon Grain Development through a Sub-Cellular Quantitative Proteomic Approach.

Brachypodiumdistachyon is a suitable plant model for studying temperate cereal crops, such as wheat, barley or rice, and helpful in the study of the grain cell wall. Indeed, the most abundant hemicelluloses that are in the B. distachyon cell wall of grain are (1-3)(1-4)-ß-glucans and arabinoxylans, in a ratio similar to those of cereals such as barley or oat. Conversely, these cell walls contain few pectins and xyloglucans. Cell walls play an important role in grain physiology. The modifications of cell wall polysaccharides that occur during grain development and filling are key in the determination of the size and weight of the cereal grains. The mechanisms required for cell wall assembly and remodelling are poorly understood, especially in cereals. To provide a better understanding of these processes, we purified the cell wall at three developmental stages of the B. distachyon grain. The proteins were then extracted, and a quantitative and comparative LC-MS/MS analysis was performed to investigate the protein profile changes during grain development. Over 466 cell wall proteins (CWPs) were identified and classified according to their predicted functions. This work highlights the different proteome profiles that we could relate to the main phases of grain development and to the reorganization of cell wall polysaccharides that occurs during these different developmental stages. These results provide a good springboard to pursue functional validation to better understand the role of CWPs in the assembly and remodelling of the grain cell wall of cereals.

Recombinant pediocin in Lactococcus lactis: increased production by propeptide fusion and improved potency by co-production with PedC.

We describe the impact of two propeptides and PedC on the production yield and the potency of recombinant pediocins produced in Lactococcus lactis. On the one hand, the sequences encoding the propeptides SD or LEISSTCDA were inserted between the sequence encoding the signal peptide of Usp45 and the structural gene of the mature pediocin PA-1. On the other hand, the putative thiol-disulfide oxidoreductase PedC was coexpressed with pediocin. The concentration of recombinant pediocins produced in supernatants was determined by enzyme-linked immunosorbent assay. The potency of recombinant pediocins was investigated by measuring the minimal inhibitory concentration by agar well diffusion assay. The results show that propeptides SD or LEISSTCDA lead to an improved secretion of recombinant pediocins with apparently no effect on the antibacterial potency and that PedC increases the potency of recombinant pediocin. To our knowledge, this study reveals for the first time that pediocin tolerates fusions at the N-terminal end. Furthermore, it reveals that only expressing the pediocin structural gene in a heterologous host is not sufficient to get an optimal potency and requires the accessory protein PedC. In addition, it can be speculated that PedC catalyses the correct formation of disulfide bonds in pediocin.

Type 1 Diabetes Prone NOD Mice Have Diminished Cxcr1 mRNA Expression in Polymorphonuclear Neutrophils and CD4+ T Lymphocytes.

In humans, CXCR1 and CXCR2 are two homologous proteins that bind ELR+ chemokines. Both receptors play fundamental roles in neutrophil functions such as migration and reactive oxygen species production. Mouse Cxcr1 and Cxcr2 genes are located in an insulin-dependent diabetes genetic susceptibility locus. The non obese diabetic (NOD) mouse is a spontaneous well-described animal model for insulin-dependent type 1 diabetes. In this disease, insulin deficiency results from the destruction of insulin-producing beta cells by autoreactive T lymphocytes. This slow-progressing disease is dependent on both environmental and genetic factors. Here, we report descriptive data about the Cxcr1 gene in NOD mice. We demonstrate decreased expression of mRNA for Cxcr1 in neutrophils and CD4+ lymphocytes isolated from NOD mice compared to other strains, related to reduced NOD Cxcr1 gene promoter activity. Looking for Cxcr1 protein, we next analyze the membrane proteome of murine neutrophils by mass spectrometry. Although Cxcr2 protein is clearly found in murine neutrophils, we did not find evidence of Cxcr1 peptides using this method. Nevertheless, in view of recently-published experimental data obtained in NOD mice, we argue for possible Cxcr1 involvement in type 1 diabetes pathogenesis.

Cell wall proteomic of Brachypodium distachyon grains: A focus on cell wall remodeling proteins.

Cell walls play key roles during plant development. Following their deposition into the cell wall, polysaccharides are continually remodeled according to the growth stage and stress environment to accommodate cell growth and differentiation. To date, little is known concerning the enzymes involved in cell wall remodeling, especially in gramineous and particularly in the grain during development. Here, we investigated the cell wall proteome of the grain of Brachypodium distachyon. This plant is a suitable model for temperate cereal crops. Among the 601 proteins identified, 299 were predicted to be secreted. These proteins were distributed into eight functional classes; the class of proteins that act on carbohydrates was the most highly represented. Among these proteins, numerous glycoside hydrolases were found. Expansins and peroxidases, which are assumed to be involved in cell wall polysaccharide remodeling, were also identified. Approximately half of the proteins identified in this study were newly discovered in grain and were not identified in the previous proteome analysis conducted using the culms and leaves of B. distachyon. Therefore, the data obtained from all organs of B. distachyon infer a global cell wall proteome consisting of 460 proteins. At present, this is the most extensive cell wall proteome of a monocot species.

Allergen relative abundance in several wheat varieties as revealed via a targeted quantitative approach using MS.

Food allergy has become a major health issue in developed countries, therefore there is an urgent need to develop analytical methods able to detect and quantify with a good sensitivity and reliability some specific allergens in complex food matrices. In this paper, we present a targeted MS/MS approach to compare the relative abundance of the major recognized wheat allergens in the salt-soluble (albumin/globulin) fraction of wheat grains. Twelve allergens were quantified in seven wheat varieties, selected from three Triticum species: T. aestivum (bread wheat), T. durum (durum wheat), and T. monococcum. The allergens were monitored from one or two proteotypic peptides and their relative abundance was deduced from the intensity of one fragment measured in MS/MS. Whereas the abundance of some of the targeted allergens was quite stable across the genotypes, others like alpha-amylase inhibitors showed clear differences according to the wheat species, in accordance with the results of earlier functional studies. This study enriches the scarce knowledge available on allergens content in wheat genotypes, and brings new perspectives for food safety and plant breeding.