Research background: Although there are many studies of the bioimprinting of lipases, there is no study comparing the strategies of bioimprinting prior to immobilization (pre-immobilization) and bioimprinting after immobilization (post-immobilization). Likewise, there is no study that compares bioimprinting of lipases immobilized from a pure lipase preparation and lipases immobilized from a crude extract. We therefore investigated these strategies, using the metagenomic lipase LipC12.
Experimental approach: We immobilized LipC12 covalently on the commercial support Immobead 150P and treated it with various bioimprinting agents, either pre-immobilization or post-immobilization. We also compared immobilization from a pure LipC12 preparation and immobilization from a crude cell-free extract.
Results and conclusions: The best improvements in triolein-hydrolyzing activity in n-hexane, compared to a non-bioimprinted control, were obtained with post-immobilization bioimprinting, using oleic acid dissolved in t-butanol: a 12-fold improvement for immobilization from a pure LipC12 preparation and an almost 14-fold improvement for immobilization from the crude cell-free extract. This bioimprinting agent also gave a 3.5-fold increase in activity for the synthesis of ethyl oleate in n-hexane, this result being obtained for pre-immobilization bioimprinting and immobilization from the cell-free extract.
Novelty and scientific contribution: This study is the first to compare pre-immobilization and post-immobilization bioimprinting strategies, as well as bioimprinting of enzymes immobilized from both pure enzyme preparations and crude cell-free extracts. These results encourage further investigation into bioimprinting strategies.
Research background: Anthocyanins, the most abundant pigments in red wine, play an important role in the visual aspect of wine sensory properties. However, due to their unstable nature, their ability to polymerise with tannins is important for colour stability. Their content varies with grapevine variety, growing conditions, viticultural and winemaking practices. Leaf removal, a common viticultural practice, enhances anthocyanin accumulation in red grapevines, and partial fruit zone leaf removal at different phenological stages can significantly influence the anthocyanin content of grapes and wine. This two-year study examined how two different times of fruit zone leaf removal at different phenological stages affect the initial anthocyanin content in wine and their stability during ageing in Merlot, Syrah and Cabernet Sauvignon wines grown in a Mediterranean climate.
Experimental approach: Partial leaf removal was applied during flowering and during vérasion and compared with untreated control. The wines obtained from all treatments and varieties were bottled two months after the end of fermentation, and then stored and matured under the cellar conditions for one year. To determine the influence of different times of leaf removal on the concentration of anthocyanins and their stability in the wine, the wines were analysed immediately after bottling and again after 6 and 12 months of storage. For the determination of all phenolic compounds, high-performance liquid chromatography (HPLC) was used.
Results and conclusions: Leaf removal treatments increased the concentration of anthocyanins in all three cultivars. The obtained results showed that malvidin-3-O-glucoside (Mal-3-Glc) was the most abundant individual anthocyanin, while the most unstable anthocyanin was petunidin-3-O-coumaroyl glucoside (Pet-3-Coum-Glc). Initial concentration of total anthocyanins in all wine samples was significantly affected by different conditions in the two years of study, but with a significant effect of the defoliation treatments. Anthocyanin concentration decreased during the ageing of the wine, and the degradation of anthocyanins ranged from 36 to 90 %. The stability of anthocyanins in wine was most influenced by ageing time, while year and treatment had no influence. The concentration of total phenolic acids increased during wine ageing, while the concentration of total flavonol glycosides (TFG) decreased in all wine samples except Merlot from 2016.
Novelty and scientific contribution: The results of this study contribute to a better understanding of the stability of increased concentrations of anthocyanins in wines during ageing obtained by the practice of grapevine leaf removal in the vineyard.
Research background: Nowadays, there is still no portable electroanalytical device suitable for the monitoring concentration of chlorophenols in technologically used water, especially in the brewing industry. This problem could be solved by developing an electroanalytical screening method based on chlorophenol anodic oxidation.
Experimental approach: The electrochemical behaviour of the target chlorophenols was investigated to find the optimum working conditions for their selective electrochemical detection in beer.
Results and conclusions: Electrochemical oxidation pathways were proposed for each investigated chlorophenol. The sum of all chlorophenols present in the brewing water, expressed as the concentration equivalent of 2,6-dichlorophenol, can be determined electrochemically, so that in future real-time monitoring of chlorophenols in the individual stages of the beer production process will be possible. Moreover, the cathodic reduction of their oxidation products proved to be a suitable electroanalytical tool for the selective detection of their presence in beer.
Novelty and scientific contribution: The research shows that an electroanalytical approach could be useful in the control of beer biotechnology to prevent sensory changes caused by the chlorophenols formed.
Research background: Recently, natural ingredients have come to the fore instead of synthetic additives in meat and meat products. In this context, the use of propolis extracts, a natural bee product, prepared with different solvents is quite widespread. From this perspective, this study investigates the contribution of ethanolic extract of propolis on the shelf life of beef meatballs and the extent to which it maintains the quality characteristics of the samples during storage at 4 °C.
Experimental approach: In this study, an ethanolic extract of 0.1, 0.3, 0.5 and 1 % of propolis was added to meatball samples. After preparation, the samples were packaged and stored at 4 °C. During storage, the meatball samples were subjected to physicochemical, microbiological and sensory analysis.
Results and conclusions: The addition of propolis did not affect significantly the water activity values. However, significantly lower pH values were observed during prolonged storage, especially in the samples containing higher amounts of propolis. The addition of propolis also effectively delayed oxidation and there was an amount-dependent decrease in TBARS values. The use of propolis in the preparation of meatballs did not have a significant effect on the initial CIELab parameters of the samples, but changes in a* and b* values were observed at the end of storage compared to the control sample. A significant increase in the total phenolic content as well as the DPPH˙ and ABTS+ radical scavenging activities of the meatballs was observed depending on the propolis amount. Considering the results of microbiological analysis, it was found that propolis could increase the microbiological quality of the meatballs, but the addition of more than 0.5 % propolis affected the overall acceptability of the samples.
Novelty and scientific contribution: As a result, the addition of certain amounts of propolis was found to be a potential alternative to synthetic counterparts that could be used to preserve refrigerated meatballs to delay oxidation and microbial spoilage without affecting the sensory properties of the samples.
Research background: Ectoine is a desirable molecule with high application potential, particularly in the cosmetics and pharmaceutical industries. The current production method uses microorganisms that require high salinity. Therefore, purification of the product is expensive, complex and requires appropriate equipment. To overcome these obstacles, we were looking for new moderately halophilic, ectoine-producing bacteria.
Experimental approach: The bacteria were isolated from high-salinity environments: in the vicinity of Tyrawa spring, in Złockie near the Na Mokradłach spring and in Rajcza, all in Poland. Their ability to biosynthesise ectoine and additionally hydroxyectoine in a 10 % premixed seawater environment was assessed semiquantitatively using mass spectrometry (MS). The growth of the bacteria was also compared under these conditions. The most promising strains were then identified based on 16S rDNA sequence and their morphological, biochemical and physiological properties were described. The ectoine was biosynthesised based on the collected data and the preferences of individual strains. The concentrations of the final product were determined by HPLC. After the screening process, the most suitable strain was identified.
Results and conclusions: Fifty-six bacterial strains were isolated. Most strains produced insignificant amounts of ectoine or hydroxyectoine in the presence of 10 % salt. However, ten strains, all isolated from the Tyrawa spring, showed promising properties and were used in further studies. Based on the 16S rDNA sequence, four were identified as Halobacillus sp., two as Virgibacillus sp. and one as Bacillus sp., Pseudalkalibacillus sp., Salimicrobium sp. and Thalassobacillus sp. The basic biochemical and physiological properties as well as the ability to grow in the presence of NaCl, KCl, (NH4)2SO4 and MgSO4 were described. Ectoine was biosynthesised following the best parameters estimated for each strain. Based on the results, Virgibacillus salarius BHTA19 was identified as a new potential producer of ectoine.
Novelty and scientific contribution: We isolated a promising ectoine producer, moderately halophilic bacterium - Virgibacillus salarius BHTA19. BHTA19 is a wild-type strain that produces significant amounts of ectoine in environments with moderate salt concentrations. It has great potential and the possibility of industrial application.
Research background: Gluten contamination is the main concern of those who follow a gluten-free diet. Although rice is naturally gluten-free, previous studies have identified gluten contamination of rice that can occur during processing, storage, handling or cooking. As a result, consumer confidence may be affected, emphasising the need to examine how these concerns shape their risk perceptions and influence their subsequent decisions. This study aims to evaluate: (i) the perceived risk of gluten contamination among gluten-free diet followers and (ii) the actual presence of gluten contamination in commercially available rice on the Croatian market.
Experimental approach: This cross-sectional study combined survey methods and laboratory analysis. An online questionnaire was used to assess the perceived risk of gluten contamination in rice among individuals following a gluten-free diet (N=66). The presence of gluten in forty-one samples of white, brown and parboiled rice from six producers on the Croatian market was then analysed using an enzyme-linked immunosorbent assay (ELISA).
Results and conclusions: Laboratory assays failed to detect gluten in any of the rice samples (limit of quantification 5 mg/kg), yet 54 (82 %) respondents expressed high uncertainty about the risk of gluten contamination in rice. These results show a significant discrepancy between consumer perception and scientific evidence. They emphasise the need for improved communication and clearer labelling to build consumer trust and support informed dietary choices.
Novelty and scientific contribution: This study highlights the gap between the perceived and actual risk of gluten contamination in naturally gluten-free food and emphasises the importance of addressing consumer concerns through better education and transparent product information.
Research background: Cactus pear (Opuntia ficus-indica) is an excellent source of polysaccharides and bioactive compounds with remarkable health benefits. The mucilage of the cactus pear, which consists mainly of water and complex carbohydrates, has properties similar to gum due to its unique physiological properties. Recently, plant-derived mucilage has gained significant attention in the dairy industry for its potential as a natural thickening and colloidal stabilizing agent.
Experimental approach: This study investigates the use of freeze-dried cactus pear cladode pulp from Opuntia ficus-indica L. Miller as a source of mucilage and its interaction with a commercial stabilizer on the physical properties of low-fat cocoa ice cream (3.0 % fat). The study evaluates the influence of cactus pear cladode pulp on the physicochemical properties and technological parameters of the ice cream. Ice cream samples containing 1.0, 1.5 and 2.0 % cactus pear cladode pulp were compared with a control sample (without cactus pear cladode pulp).
Results and conclusions: The results show that cactus pear cladode pulp is rich in fiber and phenolic compounds and has significant technological potential due to its water absorption capacity (WAC), water solubility index (WSI) and oil absorption capacity (OAC). The addition of cactus pear cladode pulp lowered the pH of the ice cream, improved its darkness and yellowness, increased the overrun and delayed the melting process. These results suggest that cactus pear cladode pulp works synergistically with the commercial stabilizer, highlighting its potential as a natural fat substitute and stabilizer for low-fat ice cream formulations.
Novelty and scientific contribution: This study represents pioneering research into the use of freeze-dried Opuntia ficus-indica cladode pulp in the production of ice cream. The results offer valuable insights for the ice cream industry and provide a natural alternative for stabilizers and fat substitutes.
Research background: Chronic, unregulated inflammation is a crucial factor in the development of numerous diseases and is closely linked to the increased expression of cyclooxygenase-2 (COX-2). While various bioactive peptides from marine organisms have shown COX-2 inhibitory effects, peptides derived from the collagen of the peanut worm (Siphonosoma australe) have not yet been demonstrated. The aim of this study is to investigate the potential COX-2 inhibitory activity of peanut worm collagen by simulated digestion in vitro with pepsin-pancreatin followed by molecular docking.
Experimental approach: During simulated in vitro digestion, commercial pepsin (at pH=3) and pancreatin (at pH=7.5) were applied for 240 min at 37 °C to evaluate the degree of hydrolysis, peptide concentration and COX-2 inhibitory activity. The samples with the most significant COX-2 inhibitory activity were then separated into fractions and identified.
Results and conclusions: The 210-minute simulated digestion in vitro showed the highest COX-2 inhibitory activity (64.31 %). This result was confirmed by the increased degree of hydrolysis (DH) and peptide concentrations observed during the simulated in vitro digestion. The peptide fraction of <1 kDa had the highest inhibitory activity (89.05 %) and was subsequently subjected to sequencing analysis. Three novel peptides, ADIAGQAAQVLR, LNNEITTLR and VGTVEK, were identified and confirmed to contain crucial amino acids and therefore verified as COX-2 inhibitors. VGTVEK has the most potent interaction, as shown by the lowest binding energy (-4.41 kcal/mol). The molecular docking revealed that VGTVEK (631.35 Da) binds to the active site of COX-2 and forms hydrogen bonds with Gln178, Leu338, Ser339, Tyr371, Ile503, Phe504, Val509 and Ser516 and hydrophobic interactions with Met99, Val102, Val330, Ile331, Tyr334, Val335, Leu345, Trp373, Leu517 and Leu520. Other biological activities of the produced peptides included ACE inhibitors, dipeptidyl peptidase-IV (DPP-IV) inhibitors and α-glucosidase inhibitors. After toxicity prediction, the peptides were classified as non-toxic.
Novelty and scientific contribution: The study found that peptides derived from peanut worm collagen have the potential to be novel, natural agents for anti-inflammatory therapy. Their broader application in functional foods, nutraceuticals and pharmaceuticals could provide new options for people suffering from inflammation and support both treatment and maintenance of overall health.
Research background: Dandelion flowers have a very short shelf life. The canning process is known not only to stabilise food and preserve its nutritional content at a high level, but also to significantly extend its shelf life. For this reason, canned dandelion flowers are believed to be beneficial for both consumers and the gastronomy sector.
Experimental approach: In this study, fresh dandelion (Taraxacum officinale L.) flowers were canned using sucrose syrup with different (20 and 30) degrees of Brix (°Bx) as filling medium and stored at 25 °C for 30 days. A total of 56 phytochemicals were identified using liquid chromatography-tandem mass spectrometry (LC-MS/MS), while the in vitro antioxidant activity of 2,2-diphenyl-1-picrylhydrazyl (DPPH) and cupric reducing antioxidant capacity (CUPRAC) and the total phenolic content (TPC) were analysed in both the canned flowers and the syrup at different storage times (on days 10, 20 and 30).
Results and conclusions: The antioxidant activities of fresh dandelion flowers were 89.6 % and 0.8 mmol Trolox equivalents (TE) per gram, respectively. The lowest DPPH (41.4 %) and CUPRAC expressed as TE (0.3 mmol/g) activities were observed on day 20 in samples stored in the 30 °Bx syrup. The TPC in fresh flowers, expressed as gallic acid equivalents (GAE) per g of extract, was 367.4 mg/g. The highest TPC in canned flowers was determined on day 10 in the samples in syrup with both °Bx. LC-MS/MS analysis identified 24 phytochemicals in fresh flowers, including quinic acid, luteolin, siranoside, chlorogenic acid, fumaric acid, caffeic acid, protocatechuic acid, quercetin, cosmosiin, isoquercitrin and apigenin. A decrease in the polyphenol content of canned flowers was observed during storage. The results indicate that canning dandelion flowers in a 30 °Bx syrup and storing them for 20 days preserved their phenolic content and antioxidant capacity.
Novelty and scientific contribution: Numerous studies in the literature focus on extending the shelf life of fruit and vegetables by the canning method. However, this study fills a gap in the literature by successfully applying the canning technique to edible flowers for the first time. Furthermore, the results of this study contribute to future research on the potential commercialisation of canned dandelion flowers as a food product.
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