Bioprospecting of Selected Species of Polypore Fungi from the Western Balkans.

Growing mushrooms means meeting challenges while aiming for sustainability and circularity. Wherever the producer is located, commercial strains are the same originating from several producers. Customized strains adapted to local conditions are urgently needed. Before introducing new species to the strain development pipeline, the chemical characterization and biological activity of wild ones need to be assessed. Accordingly, the mycoceutical potential of five polypore mushroom species from Serbia was evaluated including: secondary metabolite composition, oxidative damage prevention, anti-tyrosinase, and anti-angiotensin converting enzyme (ACE). The phenolic pattern was comparable in all samples, but the amounts of specific chemicals varied. Hydroxybenzoic acids were the primary components. All samples had varying quantities of ascorbic acid, carotene, and lycopene, and showed a pronounced inhibition of lipid peroxidation (LPx) and ability to scavenge HO•. Extracts were more potent tyrosinase inhibitors but unsuccessful when faced with ACE. Fomitopsis pinicola had the strongest anti-tumor efficacy while Ganoderma lucidum demonstrated strong selectivity in anti-tumor effect in comparison to normal cells. The evaluated species provided a solid foundation for commercial development while keeping local ecology in mind.

Is malting an absolute must? Native triticale as a stand-in for barley malt in the brewing process.

BACKGROUND: To remain competitive, brewers must innovate by incorporating novel elements beyond traditional styles. Thus, exploring triticale as a modern substitute for barley malt is promising, especially given its higher amylolytic activity compared to barley. This study aimed to assess the impact of substituting up to 50% of barley malt with unmalted triticale on green beer quality, encompassing multiple stages from wort production to primary fermentation at a laboratory scale. RESULTS: Triticale-based worts (ratios 10-50%) had lower extract content than 100% barley malt. However, incorporating 10% of triticale led to only a 1% decrease in extract content compared to the all-malt wort. Shearzyme® 500L, an endo-1,4-ß-xylanase with ß-glucanase side activity, effectively addressed wort viscosity by breaking down arabinoxylans and ß-glucans in triticale cell walls. All triticale-based beers exhibited lower ethanol content compared to reference beer, as is typical when using adjuncts. In green beer, a 50% triticale ratio lowered ethanol content by 16% (without enzyme) and 19% (with enzyme) compared to 100% malt beer. However, green beer with 10% triticale had satisfactory levels of total polyphenol and vicinal diketone content, among other parameters. CONCLUSION: Commercial enzyme application significantly enhanced proteolytic activity within the grain. Fermentations of enzyme-treated worts showed higher amino acid levels, further confirming the increased proteolytic activity facilitated by the chosen enzyme. Overall, this study provides a comprehensive analysis of the brewing process using native triticale. Building on this foundation, future studies will focus on optimizing mashing conditions to enhance the fermentation profile of the wort. © 2024 Society of Chemical Industry.

Selection of Non-Mycotoxigenic Inulinase Producers in the Group of Black Aspergilli for Use in Food Processing.

Research background: Inulinases are used for fructooligosaccharide production and they are of interest for both scientific community and industry. Black aspergilli represent a diverse group of species that has use for enzyme production, in particular some species are known as potent inulinase producers. Finding new potential producers from the environment is as important as improving the production with known strains. Safe use of enzymes produced by aspergilli in food industry is placed ahead of their benefit for inulinase production. Experimental approach: Here we show a specific approach to finding/screening of newly isolated fungal inulinase producers that combines a newly developed screening method and an equally important assessment of the toxigenic potential of the fungus. In this study 39 black aspergilli collected from different substrates in Serbia were identified and assessed for inulinase production. Results and conclusions: The most common species were Aspergillus tubingensis (51.2%), followed by A. niger (23.1%), A. welwitschiae (23.1%) and A. uvarum (2.6%). The isolates for inulinase production were selected using a cheap and easy, fast and non-hazardous alternative inulinase screening test developed in this work. Enzymatic activity of selected inulinase-producing strains was confirmed spectrophotometrically. Since some A. niger and A. welwitschiae strains are able to produce mycotoxins ochratoxin A (OTA) and fumonisins (FB), the toxigenic potential of selected inulinase producers was assessed analytically and genetically. Fungal enzyme producer can be considered safe for use in food industry only after comparing the results of both approaches for investigating toxic potential, the direct presence of mycotoxins in the enzyme preparation (analytically) and the presence of mycotoxin gene clusters (genetically). In some strains the absence of OTA and FB production capability was molecularly confirmed by the absence of complete or critical parts of biosynthetic gene clusters, respectively. The two best inulinase producers and mycotoxin non-producers (without mycotoxin production capability as additional safety) were selected as potential candidates for further development of enzyme production. Novelty and scientific contribution: The presented innovative approach for the selection of potential fungal enzyme producer shows that only non-toxigenic fungi could be considered as useful in food industry. Although this study was done on local isolates, the approach is applicable globally.

Micro/trace/toxic elements and insecticide residues level in monofloral bee-collected sunflower pollen- health risk assessment.

The aim of the current research was to determine the content of (potentially) toxic elements and insecticide residues in monofloral sunflower bee-collected pollen. For micro- and trace elements determination Inductively Coupled Plasma Optical Emission (ICP-OES) analytical method was used while insecticide residue content was monitored by applying Liquid Chromatography-Mass Spectrometry (LC-MS/MS) technique. In total, seventeen micro/trace elements were quantified. None of the twenty four examined insecticides were detected above the limit of detection (LOD) which makes studied sunflower bee-collected pollen eco-friendly both to bees and humans. Based on presence of several toxic as well as potentially toxic elements calculations for estimated weekly intakes (EWI), and oral intakes (OI) were made and used for health risk assessment based on the computation of two different health risk quotients (HQ)- acute (HQA) and long-term (HQL). The obtained results proved that all HQ values for adults were negligible or low except in case of HQL value for arsenic (0.32) which can be characterized as medium. However, in case of children much more precaution is needed due to significant HQL risk for arsenic (1.511). The attained data can help to make additional linkage between bee-collected pollen as food ingredients and potential benefits/risks for human health.

Effect of Ripening in Brine and in a Vacuum on Protein, Fatty Acid and Mineral Profiles, and Antioxidant Potential of Reduced-Fat White Cheese.

RESEARCH BACKGROUND: Numerous factors affect the ripening of reduced-fat white cheese. The aim of this study is to investigate the influence of ripening environment (brine or vacuum plastic bags without brine) on the chemical composition, protein, fatty acid profile and mineral content as well as antioxidant properties of industrially produced reduced-fat white cheese. EXPERIMENTAL APPROACH: A low-fat white cheese was manufactured on an industrial scale from milk that remained after the production of kajmak and ripened for 60 days at 4 °C after packaging in a polystyrene container with brine containing 6% salt or in vacuum-sealed polyethylene bags. The influence of ripening environment on proteolysis was monitored by the change of soluble nitrogen fractions as well as by sodium dodecyl sulphate-polyacrylamide gel electrophoresis of tris(hydroxymethyl) aminomethane-HCl extracts of cheese proteins under non-reducing conditions and water-soluble fractions under reducing conditions. An effect that ripening environment had on fatty acid and mineral content was also monitored. The change of antioxidant potential of the investigated cheese during ripening led to the change of iron(II) chelating ability, reducing power and free-radical scavenging activity. RESULTS AND CONCLUSIONS: The ripening environment differently affected proteolysis, fatty acid composition, mineral profile and antioxidant properties of reduced-fat white cheese. White cheese ripened in brine had more intensive proteolytic changes than the cheese ripened in a vacuum, but also more intensive diffusion processes, especially between the 40th and 60th day of ripening. The brine-ripened cheese had higher values of water-soluble nitrogen content, but lower contents of trichloroacetic acid-soluble and phosphotungstic acid-soluble nitrogen than the vacuum-ripened cheese. Cheese ripened in brine had a lower content of almost all investigated macro- and microelements. After 60 days of ripening, in cheese ripened in brine only myristic (C14:0) and palmitic acid (C16:0) were detected, whereas in the vacuum-ripened cheese C10:0-C16:0 fatty acids dominated. Vacuum-ripened reduced-fat cheese had more favourable reducing power, while white brined reduced-fat cheese had better radical scavenging activity and iron(II) chelating activity. NOVELTY AND SCIENTIFIC CONTRIBUTION: These results suggest significant influence of ripening conditions (immersion in brine or in vacuum-sealed polyethylene bags) on nutritive and functional properties of reduced-fat white cheese. Ripening in a vacuum has become a useful method for obtaining high-value reduced-fat white cheese.

Cumulative risk assessment of dietary exposure of the adult population in Serbia to pesticides that have chronic effects on the thyroid gland through fresh fruits and vegetables.

In contrast to the traditional approach to risk assessment, which focuses on a single chemical, cumulative exposure and risk assessment considers the consequences of exposure to multiple chemical combinations. A cumulative risk assessment of dietary exposure of adult females and adult males to pesticides with chronic effects on the thyroid was conducted by estimation of the Total Margin of Exposure (MOET). Exposure to each active substance was estimated using a second-order Monte Carlo simulation. Input values for the simulation were based on over 2300 conventionally produced fruit and vegetable samples analysed from 2021 to 2023 and consumption data collected using the Food Frequency Questionnaire (FFQ) and 24h recall method. MOET values, for both populations assessed, did not exceed thresholds for regulatory consideration established by risk managers. Considering that MOETs values from consumption of fresh fruits and vegetables were relatively close to the threshold value, total exposure to the pesticides could likely reach risk-associated MOET levels, through the consumption of other food products that may be contaminated.

Monofloral Corn Poppy Bee-Collected Pollen-A Detailed Insight into Its Phytochemical Composition and Antioxidant Properties.

The aim of this study was to compile a detailed phytochemical profile and assess the antioxidant properties of bee-collected pollen (PBP) obtained from corn poppy (Papaver rhoeas L.) plants. To achieve this, a lipid fraction was prepared for quantifying fatty acids using GC-FID. Extractable and alkaline-hydrolysable PBP fractions (obtained from a defatted sample) were used to determine the qualitative and quantitative profiles of phenolic compounds, phenylamides and alkaloids using UHPLC/Q-ToF-MS. Additionally, various spectrophotometric assays (TAC, FRP, CUPRAC, DPPH⦁) were conducted to evaluate the antioxidant properties. Phenolic compounds were more present in the extractable fraction than in the alkaline-hydrolysable fraction. Luteolin was the predominant compound in the extractable fraction, followed by tricetin and various derivatives of kaempferol. This study presents one of the first reports on the quantification of tricetin aglycone outside the Myrtaceae plant family. The alkaline-hydrolysable fraction exhibited a different phenolic profile, with a significantly lower amount of phenolics. Kaempferol/derivatives, specific compounds like ferulic and 5-carboxyvanillic acids, and (epi)catechin 3-O-gallate were the predominant compounds in this fraction. Regarding phenylamides, the extractable fraction demonstrated a diverse range of these bioactive compounds, with a notable abundance of different spermine derivatives. In contrast, the hydrolysable fraction contained six spermine derivatives and one spermidine derivative. The examined fractions also revealed the presence of seventeen different alkaloids, belonging to the benzylisoquinoline, berberine and isoquinoline classes. The fatty-acid profile confirmed the prevalence of unsaturated fatty acids. Furthermore, both fractions exhibited significant antioxidant activity, with the extractable fraction showing particularly high activity. Among the assays conducted, the CUPRAC assay highlighted the exceptional ability of PBP's bioactive compounds to reduce cupric ions.

Atropine and Scopolamine in Maize Products from the Retail Stores in the Republic of Serbia.

The cereal grains, which represent the cultivated grasses fruits, supply almost half of the total caloric requirements for humans and provide more nourishment compared with any other class of the food. Out of many cereals used for food, maize, rice, and wheat are the most important food resources for humans, representing 94% of the total cereals consumption. According to the data of the Republic Institute of Statistics for the year 2018, the harvested areas of corn amount to 906,753 hectares. The production of about 7 million tons was achieved with an average yield of 7.7 t/ha according to the Ministry of Agriculture of the Republic of Serbia. Serbia is still among the ten largest exporters of wheat and corn in the world for the period of 2014/15-2017/18. More precisely, it ranks seventh in the export of corn. Utilization of maize products for food animal nutrition (1000 t) is 491,48, and for industrial processing (1000 t) 278,862 expressed as the total consumption (1000 t) is 769,910. Therefore, a total of 103 samples of maize products were analyzed for the presence of toxins, i.e., tropane alkaloids (TAs). The samples were collected from the retail stores in the Republic of Serbia in 2021 and analyzed for the presence of atropine and scopolamine (33 corn grits, 39 polenta, and 31 semolina samples). Therefore, the Recommendation 2015/976/EU on the monitoring of TAs in food was adopted by the EU Commission to obtain more occurrence data on TAs in food. The monitoring extent, however, is restricted because reliable analytical methods and appropriate sensitivity are limited. There was a limit of 1 g/kg for each atropine and scopolamine in cereals containing millet, sorghum, buckwheat, or their derivatives. All the samples were analyzed by the LC-MS/MS. The LOQ was set at 1.0 µg/kg. Out of the total 103 tested samples, 32 samples (31.1%) were contaminated with atropine and scopolamine in concentrations above the LOQ. The highest concentrations of the studied TAs were observed in a semolina sample-atropine: 58.80 µg/kg, scopolamine: 10.20 µg/kg. The obtained results indicate that the TAs concentrations are above the LOQ which can be considered potential human and animal health hazards.

Inhibitory Effects of Brassicaceae Cover Crop on Ambrosia artemisiifolia Germination and Early Growth.

Several cover crops (CCs) exert allelopathic effects that suppress weed growth. The aim of the present study was to evaluate the effects of aqueous extracts containing different concentrations [0, 0.5, 1, 2.5, 5, 7.5 and 10% (w/v)] of Brassicaceae CCs (Sinapis alba, Raphanus sativus, Camellina sativa) and of the CCs Fagopyrum esculentum and Guizotia abyssinica on germination and early growth of Ambrosia artemisiifolia L. The allelopathic effects were species and concentration-dependent. C. sativa, for example, caused the greatest potential to inhibit germination, shoot, radicle length and fresh seedling weight, whereas S. alba and R. sativus inhibited germination and early growth of A. artemisiifolia only at concentrations ≥7.5%. In contrast, no inhibition was observed when aqueous extracts of F. escultneum and G. abyssinica were added at any of tested concentration. Liquid chromatography-tandem mass spectrometry detected 15 phenolic compounds in Brassicaceae CCs with the highest content (µg/g) of vanillin (48.8), chlorogenic acid (1057), vanilic acid (79), caffeic acid (102.5) and syringic acid (27.3) in C. sativa. Our results suggest that C. sativa is the most allelopathic CCs and that the fruits of C. sativa are the plant organs richest in allelochemicals.

Phytochemical Profile and Antioxidant Properties of Bee-Collected Artichoke (Cynara scolymus) Pollen.

The current study intended to determine, for the first time, phenolic and fatty acid profile, antioxidant and certain nutritional properties of monofloral bee-collected artichoke (Cynara scolymus) pollen. Based on UHPLC-DAD MS-MS analysis the main phenolics in extractable fraction were different flavonol glycosides (in particular Isorhamnetin-3-O-glucoside, 49.2 mg/kg of dry weight) while ferulic acid was the predominant phenolic compound (39.4 mg/kg of dry weight) in the alkaline hydrolyzable fraction. Among fatty acids (FAs), results of GC-FID analysis revealed prevalence of unsaturated FAs with cis-5,8,11,14,17-eicosapentaenoic acid (EPA) and oleic acid as the main ones- 28.4% and 24.9%, respectively. Based on the FA composition, nutritional analysis proved that artichoke bee-collected pollen had balanced ω-6 and ω-3 FAs content. To determine the antioxidant properties of pollen, five different assays were applied. It was proved that bioactive compounds in artichoke pollen possessed significant ability to quench DPPH radical as well as ABTS radical cation. In addition, in vitro phosphomolybdenum assay confirmed that artichoke pollen is an excellent source of different antioxidants. Pollen extracts exhibited moderate ferric reducing power as well as low ferrous chelating ability. Some further antioxidant studies (preferably in vivo) should be performed to confirm the observed results.

In vitro assessment of pesticide residues bioaccessibility in conventionally grown blueberries as affected by complex food matrix.

The aim of this study was to investigate the bioaccessibility of pesticide residues in blueberries (commercial and sample from controlled field trial) from Serbia, involving the presence of a complex food matrix and to assess the potential risk to human health. The presence of nine active substances (azoxystrobin, boscalid, fludioxonil, cyprodinil, pyrimethanil, pyridaben, pyriproxyfen, acetamiprid and thiametoxam) in initial blueberry samples was determined in concentration range from 5.15 µg/kg for thiametoxam to 187 µg/kg for azoxystrobin. Clothianidin, metabolite of thiametoxam, was not detected in any blueberry sample. However, after in vitro digestion, the content of initially detected pesticides residues was significantly decreased or it was below limit of quantification resulting in the total bioaccessibility of about 15%. Azoxystrobin, pyrimethanil and fludioxonil was quantified in digestive juice at concentrations which were about 81%, 37% and 10% less than the inital concentration, respectively. The presence of food matrix during digestion of blueberries even more severely reduced concentration of pesticide residues (total bioaccessibility was about 7%) compared to digestion without the food matrix. Only azoxystrobin was quantified after digestion with food matrix in concentration of 27 µg/kg in sample from controlled field trial and detected in two commercial samples but below the limit of quantification. Furthermore, chronic risk assessment indicated that risk is acceptable for the health of different human subpopulation groups. The current study on pesticides residues, most commonly applied on blueberries, provides for the first time an insight into their bioaccessibility under conditions that mimic physiological environment of human digestive tract.