Kristina Vlahoviček-Kahlina

Halyomorpha halys is a highly polyphagous invasive pest whose management still relies heavily on broad-spectrum insecticides, creating a need for more sustainable behaviour-based alternatives. This study evaluated clove leaf and lemongrass essential oils (EOs) as potential repellents against adult H. halys by integrating behavioural bioassays, controlled-release formulations, molecular modelling and in silico ecotoxicological assessment. Male and female adults were tested in a two-choice Y-tube olfactometer; the individual EOs were encapsulated in calcium-alginate microspheres, and selected constituents were assessed using molecular docking and computational ecotoxicity models. Clove EO, lemongrass EO and their binary combination induced strong avoidance responses in both sexes, with pooled avoidance ranging from 90.0% to 96.7% and repellency indices from 0.80 to 0.93. No significant differences were detected among the three EO treatments when responses were pooled across sexes (χ² = 2.14, p = 0.343). Both EOs were successfully incorporated into calcium-alginate microspheres, with encapsulation efficiencies exceeding 90% (91.57 ± 0.67% for clove EO and 96.42 ± 0.29% for lemongrass EO). Lemongrass EO showed higher encapsulation efficiency and loading capacity and reached near-complete cumulative release, whereas clove EO release plateaued at approximately 70%. Phylogenetic, structural and physicochemical analyses supported the selection of H. halys HhOBP21 for comparative molecular modelling with the structurally characterised Drosophila melanogaster DmOBP28a. Molecular docking indicated that selected EO constituents could be accommodated within the binding cavities of these OBPs, with β-caryophyllene, α-humulene and limonene among the most favourably scored ligands, suggesting potential interactions with these proteins. Predicted acute toxicity values for the standard non-target test organism Tetrahymena pyriformis, Daphnia magna and Pimephales promelas remained below the applied thresholds for all evaluated compounds, although several constituents showed elevated predicted bioconcentration potential and limited biodegradability. Overall, the results demonstrate strong avoidance responses to the tested EOs under laboratory Y-tube conditions and support their further evaluation in semi-field and field studies to determine whether these responses translate into spatial repellency under practical conditions. Experimental validation of predicted molecular interactions and environmental fate is also needed.

Phytopathogenic fungi are living organisms that cause plant diseases and great damage to agricultural products. Despite the wide range of commercial fungicide products in use, there is a clear need for new and environmentally friendly fungicides. Here we propose a new ecological fungicide, copper alginate microspheres prepared by ionic gelation. The microspheres were characterized (morphology and topography, encapsulation efficiency, loading capacity, swelling behavior, rheology, kinetics and mechanism of copper ions release) and their in vitro antifungal potential against selected genera of phytopathogenic fungi was evaluated. Copper alginate microspheres inhibited spore germination of Botrytis cinerea. Compared to the control, the inhibition of B. cinerea spore germination (48%) was greater than that of the commercial fungicide Neoram^® (22%). The mycelial growth of Cercospora beticola and Phytophthora ramorum was also significantly inhibited by the addition of copper alginate microspheres. Novel fungicide offer effective disease control while minimizing environmental impact and promoting sustainable agriculture practices.

Phytopathogenic fungi are living organisms that cause plant diseases and great damage to agri-cultural products. Despite the wide range of commercial fungicide products in use, there is a clear need for new and environmentally friendly fungicides. Here we propose a new ecological fun-gicide, copper alginate microspheres prepared by ionic gelation. The microspheres were charac-terized (morphology and topography, encapsulation efficiency, load capacity, swelling behavior, rheology, kinetics and mechanism of copper ions release) and their in vitro antifungal potential against selected genera of phytopathogenic fungi was evaluated. Novel copper alginate micro-spheres inhibited spore germination of Botrytis cinerea and mycelial growth of Cercospora beticola and Phytophthora ramorum. They offer effective disease control while minimizing environmental impact and promoting sustainable agriculture practices.

Addressing the growing need for environmentally friendly fungicides in agriculture, this study explored the potential of biopolymer microparticles loaded with metal ions as a novel approach to combat fungal pathogens. Novel alginate microspheres and chitosan/alginate microcapsules loaded with zinc or with zinc and silver ions were prepared and characterized (microparticle size, morphology, topography, encapsulation efficiency, loading capacity, and swelling behavior). Investigation of molecular interactions in microparticles using FTIR-ATR spectroscopy exhibited complex interactions between all constituents. Fitting to the simple Korsmeyer–Peppas empirical model revealed the rate-controlling mechanism of metal ions release from microparticles is Fickian diffusion. Lower values of the release constant k imply a slower release rate of Zn²⁺ or Ag⁺ ions from microcapsules compared to that of microspheres. The antimicrobial potential of the new formulations against the fungus Botrytis cinerea was evaluated. When subjected to tests against the fungus, microspheres exhibited superior antifungal activity especially those loaded with both zinc and silver ions, reducing fungal growth up to 98.9% and altering the hyphal structures. Due to the slower release of metal ions, the microcapsule formulations seem suitable for plant protection throughout the growing season. The results showed the potential of these novel microparticles as powerful fungicides in agriculture.

Addressing the growing need for effective and environmentally friendly fungicides in agricul-ture, this study explored the potential of alginate microparticles loaded with metal ions as a novel approach to combat fungal pathogens. Novel alginate microparticles (microspheres and microcapsules) loaded with zinc or simultaneously with zinc and silver ions were prepared and physicochemically characterized (microparticle size, morphology, topography, encapsulation efficiency, loading capacity, and swelling behavior). Investigation of molecular interactions in microparticles using FTIR-ATR spectroscopy exhibited complex interactions between all con-stituents. Fitting to the simple Korsmeyer-Peppas empirical model revealed the rate-controlling mechanism of zinc and silver ions release from microparticles is Fickian diffusion. Lower values of the release constant k imply a slower release rate of Zn or Ag ions from microcapsules com-pared to that of microspheres. The antimicrobial potential of the new formulations against the fungus Botrytis cinerea was evaluated. When subjected to tests against the fungus, microspheres and microparticles loaded with both zinc and silver ions effectively reduced its growth and al-tered the fungal hyphal structures well as exhibited superior antifungal activity. The results showed the potential of these novel microparticles as potent fungicides in agriculture.

The increased demand for functional food with added health benefits is directing industrial procedures toward more sustainable production of naturally added bioactive compounds. The objective of this research was to investigate the potential of bioactive compounds from rosemary extract obtained using high-voltage electrical discharge as a green extraction method, for microencapsulation as a protective method for future application in functional food. Four types of microparticles were made via the ionic gelation method using alginate (Alg), zein (Z), and hydroxypropyl methylcellulose (HPMC) biopolymers and were analyzed considering the physicochemical properties. The diameter of dry microparticles ranged from 651.29 to 1087.37 μm. The shape and morphology analysis of microparticles showed that the obtained microparticles were quite spherical with a granular surface. The high encapsulation efficiency was obtained with a loading capacity of polyphenols up to 11.31 ± 1.47 mg GAE/g (Alg/Z microparticles). The microencapsulation method showed protective effects for rosemary polyphenols against pH changes during digestion. Specifically, the addition of both zein and HPMC to calcium-alginate resulted in microparticles with a prolonged release for better availability of polyphenols in the intestine. This research background indicates that the release of rosemary extract is highly dependent on the initial biopolymer composition with high potential for further functional food applications.

Mandarins are the second most farmed citrus. Consumers demand a mandarin fruit that is both tasty and excellent in quality. The fruit quality, flavonoid profile, antioxidant capacity and total carotenoid content of five varieties (‘Zorica’, ‘Chahara’, ‘Kawano Wase’, ‘Owari’ and ‘Saigon’) of Satsuma mandarins grown in Neretva valley (Croatia) were determined. In this research, the distribution of bioactive compounds was different for mandarin juice, dry pulp residue and dry peels. Dry peels showed higher levels of polyphenolic compounds and antioxidant activity. Total carotenoids were found to be in a greater concentration in dry pulp residue than in the dry peel. The highest levels of total carotenoids (543 μg β-carotene/g) were found in the dry pulp residue of ‘Owari’ and ‘Saigon’, while the lowest levels were in ‘Chahara’ (227.87 μg β-carotene/g). In dry mandarin peels, the highest levels of total carotenoids were in ‘Kawano Wase’ (227.58 μg β-carotene/g), and the lowest levels were in ‘Chahara’ (52.24 μg β-carotene/g). The most abundant component of polymethoxyflavones (PMFs) in mandarin dry peel was nobiletin, ranging from 0.204 mg/g (‘Chahara’) to 0.608 mg/g (‘Saigon’), followed by tangeretin, ranging from 0.133 mg/g (‘Chahara’) to 0.251 mg/g (‘Saigon’), and sinesestin (‘Zorica’), ranging from 0.091 mg/g to 0.353 mg/g (‘Saigon’).

Silver (Ag) and zinc (Zn) are very powerful antimicrobial metals. Therefore, in this research, a high-throughput, sensitive, and rapid method was developed for the determination of Ag and Zn in microcapsules using inductively coupled plasma mass spectrometry (ICP-MS). The sample preparation procedure employed simple microwave digestion of the microcapsules with 55.55% v/v HNO₃ and 44.45% v/v H₂O₂. The method was applied to determine Ag and Zn in microcapsule samples of different sizes (120 and 450 μm) after their preparation with and without chitosan. Prepared microcapsules, after characterization, were bonded to a polymer carrier by sol-gel procedure and the materials were characterized by FTIR spectroscopy and high-resolution optical microscopy. Significant differences were found in Ag and Zn levels between microcapsules samples prepared with and without chitosan. The results have shown that samples with chitosan had up to 20% higher levels of Zn than Ag: 120 μm microcapsules contained 351.50 μg/g of Ag and 85.51 μg/g of Zn, respectively. In contrast, samples prepared without chitosan showed larger overall variability: In microcapsules with a diameter of 120 μm, the amounts of antimicrobial metals were 98.32 μg/g of Ag and 106.75 μg of Zn, respectively. Moreover, 450 μm microcapsules contained 190.98 μg/g of Ag and 121.35 μg/g of Zn. Those quantities are high enough for efficient antimicrobial activity of newly prepared microcapsules, enabling the application of microcapsules in different antimicrobial coatings.

Alternatives and replacements for synthetic chemical-based plant protectants are required. In this study biopolymeric microspheres containing arthropod-derived apitoxin are explored as a possible novel environmentally friendly formulation for plant protection. Here we document the optimization process for microencapsulation of apitoxin into a stable formulation, for ready use in agricultural applications. Efficacy trials were carried out on three different beetle species at various developmental stages (Leptinotarsa decemlineata (Say, 1824.), Tenebrio molitor (Linnaeus, 1758.), Sitophilus granarius (Linnaeus, 1758.)). The encapsulated apitoxin has a steady initial and long residual effect, due to the slow release of apitoxin which is one of its main advantages over other conventional control methods. Microspheres loaded with apitoxin have a detrimental effect on insects, of which it is significantly better gastric compared to contact action (due to pH). The results showed that the highest and fastest mortality was obtained when the highest concentrations (0.6%) were applied, chosen to be economically acceptable. These important findings contribute to knowledge on the application and development of encapsulated apitoxin formulations, and their effectiveness, as an alternative eco-friendly control method in agricultural production.