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Plasma technologies today are taking attention in the development of emerging advanced materials of new generation due of its unique properties, broadness of applications and effectiveness of treatment. Plasma processing of materials is increasingly used in industrial, technological and medical applications. This work is based on innovative laser synthesis of colloidal nanoparticle solutions and their applications. Laser synthesis of nanoparticles is based on a process of laser ablation of metallic or metal-oxide targets immersed in liquids and it is known as ‘green’ and versatile synthesis route (no chemicals involved in the synthesis) allowing synthesis of pure nanoparticles of wide variety of materials. Nanoparticle applications to be discussed: - Metal oxide nanoparticles as solid contact in ion-selective electrodes sensitive to potassium ions - Controlling the composition of plasma-activated water by Cu ions - Bacteria Exposed to Silver Nanoparticles Synthesized by Laser Ablation in Water: Modelling E. Coli Growth and Inactivation - Photodegradation of Methylene Blue and Rhodamine B Using Laser-Synthesized ZnO Nanoparticles

In nanoscience and nanotechnology, nanoparticles have a significant role due to their unique magnetic, electrical, mechanical, optical, and electronic properties with respect to the bulk materials [1]. Zinc oxide (ZnO) is one of the most widely used materials with unique physicochemical properties and direct band-gap energy of 3.37 eV [2]. Introducing a new element in the crystal structure of ZnO leads to enhancement of the electrical and optical properties and broadens the area of its application [3]. Pulsed laser ablation in liquid (PLAL) gained a lot of attention due to the simplicity, effectiveness, high purity, and fast production of nanoparticles [4], while a wide range of parameters can be optimized [5, 6]. In this work, a simple two-step method for the synthesis of silver (Ag), gold (Au), and platinum (Pt) doped ZnO nanoparticles have been developed. The method is based on the fabrication of targets using pulsed laser deposition (PLD) technique where thin layers of metals (Ag, Pt, Au) have been deposited on a ZnO substrate. Such formed two-layer structures were used as a target for the production of doped nanoparticles (ZnO: Ag, ZnO: Au, and ZnO: Pt) by pulsed laser ablation in water. The optical properties, crystalline structure, elemental composition, morphology, and nanoparticle size distribution were studied using UV-VIS spectrophotometer, Xray diffraction (XRD), X-ray spectroscopy (XPS), scanning electron microscope (SEM), and atomic force microscope (AFM), respectively.

Listeriosis is a serious disease caused by food contaminated with Listeria monocytogenes. This bacterium is found in soil and surface water samples, but can also be found in dairy products, meat and seafood. Membrane-active antimicrobials such as quaternary ammonium salts (QASs), represent an important class of bacteriostatic and biocidal amphiphilic agents that are widely used in daily life. Given the widespread use of QASs, there have been reports of L. monocyogenes isolates resistant to these compounds. Here, we present the synthesis of a new series of quaternary amidoquinuclidine salts with potent antibacterial activity against L. monocytogenes. The antibacterial activity was determined for bacteria in the form of suspension and biofilm in the low micromolar range. The newly synthesized 3- aminoquinuclidines (C10, C12, C14, and C16) were quaternized with allyl or methyl substituents. We found that regardless of the type of substituent, the derivatives with longer alkyl chains had minimal inhibitory concentrations against L. monocytogenes that were almost identical to those of the standards (BAB, CPC, and cefotaxime). The identified candidates had low toxicity to healthy human cell lines with potent genotoxic activity against bacterial cells. In addition to the mechanism of action, we demonstrated that these agents target the cell membrane and cause membrane perforation and cell death. Parallel artificial membrane permeability assays mimicking human skin showed that candidates with longer alkyl chains did not penetrate the membrane, highlighting their potential application as disinfectants against this and other highly pathogenic bacteria.

Polymer surface modification has gained a lot of interest over the last few decades. The surface modification offers new physical and chemical properties to polymers by changing their surface properties or incorporating metal or metal oxide nanoparticles [1, 2]. Nanoparticles have a significant role due to their unique magnetic, electrical, mechanical, optical, and electronic properties with respect to the bulk materials. Zinc oxide (ZnO) is one of the most widely used materials with unique physicochemical properties and direct band-gap energy of 3.37 eV, making him a good UV absorber [3]. Polymer/nanoparticle composites gain attention due to their excellent properties for applications in the food industry, biomedicine, biotechnology, optoelectronics, etc. In this work, PVC (polyvinyl chloride) and HDPE (high-density polyethylene) polymers were treated with an atmospheric pressure plasma jet using helium as a working gas. The ZnO colloidal solution was synthesized using pulsed laser ablation in water. After plasma treatment, ZnO colloidal solution was dropped on the polymer surface, and, finally, the polymer was treated again with helium plasma. Optical emission spectroscopy (OES) was used to determine the elemental composition of plasma. All polymer samples (treated and untreated) were characterized using Fourier-transform infrared spectroscopy (FTIR), atomic force microscopy (AFM), and scanning electron microscopy (SEM). The ZnO optical properties, crystalline structure, elemental composition, morphology, and nanoparticle size distribution were studied using UV-VIS spectrophotometer, X-ray diffraction (XRD), X-ray spectroscopy (XPS), and scanning electron microscope (SEM), respectively.

Quaternary ammonium salts (QASs) are irreplaceable membrane- active antimicrobial agents that have been widely used for almost a century. Cetylpyridinium chloride (CPC) is one of the most potent QASs, however, recent data from the literature indicate a 2- to 4-fold decrease in CPC activity against resistant bacterial strains. Given the growing demand for effective antimicrobials, especially in times of current and future spread of infectious diseases, the number of resistant isolates is expected to increase. One plausible approach to address this problem is to structurally modify the CPC structure by adding other biologically active functional groups. Here, a series of QASs based on pyridine-4-aldoxime were synthesized, characterized, and tested for antimicrobial activity in vitro. Although we obtained several potent antiviral candidates, Py- C12Br, Py-C12, and Py-C14, these candidates had lower antibacterial activity than commercial CPC. AFM images showed damage to the cell membrane and no viable cells after the bacteria were exposed to 4xMIC of Py-C12 for 3 hours. We found that the addition of an oxime group to the pyridine backbone resulted in an unfavorable electron density distribution and cLogP values and disrupted the interaction with the QacR dimer that regulates efflux pump expression. MD simulations showed that binding of Py-C16 to QacR leads to dissociation of the dimer within 50 ns, whereas the same was not observed in the case of the QacR dimer and the QacR dimer bound to CPC. This explains the lower bioactivity of our compounds, as they are likely to induce premature expression of the efflux pumps.

In this paper a model for quantitative and calibration-free determination of colloidal metal nanoparticle diameter and concentration synthesized by laser ablation in water is developed. Model is valid under assumption that total ablated material is transferred into nanoparticles which is proved by Mie theory under dipole approximation and Beer Lambert law using experimentally determined ablated crater volumes, size distributions and photoabsorbances. The novelty of the model manifests in the possibility of determination of nanoparticle diameter and concentration using only volume (or mass) of ablated crater and UV-Vis photoabsorbance spectrum. It allows fast, accurate and simple colloidal metal nanoparticle quantitative analysis without use of electron microscopy, mass spectrometry or relevant experimental techniques.

Introduction Silver nanoparticles are one of the most commercialized types of nanoparticles, with a wide range of applications owing to their antimicrobial activity. However, there is a lack of data on cumulative toxicity of silver nanoparticles used with polymeric materials, especially polymer nanoparticles. In this study, toxicity of PVP-coated silver nanoparticles (AgNP) combined with polystyrene nanoparticles (PSNP) was tested on Jurkat cells. Results Silver and polystyrene nanoparticles reduced cell viability through generation of reactive oxygen species (ROS), depletion of cellular GSH, and damage to cellular structures and mitochondria. Nanoparticle treatment also reduced cell stiffness, measured by Atomic Force Microscope (AFM), which is indicative of changes in cytoskeletal structure. Cellular treatment with combinations of non-toxic AgNP and PSNP concentrations resulted in significant induction of oxidative stress and reduction of cell viability. Conclusions AgNP share the mechanism of toxicity with PSNP and results for combined AgNP/PSNP treatments can be understood as compounded effect of overloading cellular mechanisms for managing oxidative stress. These results have important implications for use of these materials in medical contexts, particularly for long-term medical implants.

Predstavljamo primjer nastavnog sata s temom Stefan- Boltzmannovog zakona kojeg započinjemo motivacijskim pitanjima: Koliko bi iznosila prosječna temperatura Zemlje kada ne bi postojala atmosfera? Koja je uloga atmosfere? Kako nastaje globalno zatopljenje? Kroz odgovore na postavljena pitanja dolazimo do Stefan- Boltzmannovog zakona i njegove primjene. U laboratorijskom eksperimentu, koji se zasniva na mjerenju I-V karakteristike žaruljice sa žarnom niti, pokazujemo proporcionalnost snage žaruljice i četvrte potencije temperature žarne niti. PHET simulacijom pokazujemo utjecaj atmosfere na temperaturu Zemlje te uspoređujemo temperaturu Zemlje iz simulacije s temperaturom Zemlje izračunatom u okviru jednostavnog modela zasnovanog na Stefan- Boltzmannovom zakonu. Staklenički učinak atmosfere objašnjavamo pomoću analognog modela temeljenog na punjenju i pražnjenju rezervoara s vodom, pri čemu je voda energija, a razina vode u rezervoaru temperatura Zemlje. Na kraju sata objašnjavamo primjene zakona zračenja u svakodnevnom životu.

Introduction: Tropaeolum majus L. altum is a plant that belongs to Tropaeolaceae family and is known for its ornamental and medicinal properties [1]. It contains benzyl glucosinolate (glucotropaeolin) whose degradation product benzyl isothiocyanate shows various biological activities such as anticancer, antimicrobial and anti-inflammatory [2, 3, 4]. Objectives: The aim of this study was to investigate the anticancer activity of T. majus essential oil against human bladder cancer cell line TCCSUP and to determine if membrane alterations are a part of the anticancer mechanism of T. majus essential oil. Materials & methods: The essential oil of T. majus seeds was obtained by microwave-assisted distillation. Gas chromatograph equipped with mass spectrometer was used to identify volatile compounds. The half maximal inhibitory concentration (IC50) of T. majus essential oil was determined against TCCSUP cell line via the MTT assay. Atomic force and optical microscopy were used to detect morphology and membrane changes of cells treated with sub-IC50 concentration. Results: T. majus essential oil has shown anticancer activity against TCCSUP cell line due to high amount of benzyl isothiocyanate. Bright-field microscopy of sub-IC50 concentration treated cells reveals the change in cell morphology and possible membrane blebbing of some cells. Atomic force microscopy analysis suggests notable cell surface alterations/membrane bleb formation as a part of the anticancer mechanism. Conclusion: The T. majus essential oil exhibits anticancer activity against TCCSUP cancer cells by altering cell morphology after exposure with sub-IC50 concentration.

Silver nanoparticles are one of the most commercialized types of nanoparticles, with a wide range of applications owing to their antimicrobial activity.[1] However, there is a lack of data on cumulative toxicity of silver nanoparticles used with polymeric materials, especially polymer nanoparticles. In this study, toxicity of PVP-coated silver nanoparticles (AgNP) combined with polystyrene nanoparticles (PSNP) was tested on Jurkat cells. Silver and polystyrene nanoparticles reduced cell viability through generation of reactive oxygen species (ROS), depletion of cellular GSH, and damage to cellular structures and mitochondria. Cellular treatment with combinations of non-toxic AgNP and PSNP concentrations resulted in significant induction of oxidative stress and reduction of cell viability. Results for combined AgNP/PSNP treatments can be understood as compounded effect of overloading cellular mechanisms for managing oxidative stress. These results have important implications for use of tested nanomaterials in medical contexts.

The aim of this thesis is to elaborate on the mode of antibacterial action by colloidal silver nanoparticles against Escherichia coli as a model organism. Two approaches were used to investigate the bactericidal mechanism of silver nanoparticles (AgNPs): experimental probing and modelling of the treated bacterial cell growth. AgNPs were synthesized by laser ablation in water and the obtained colloid was thoroughly characterized. An upgradable interaction-based growth model was developed to understand the effect of AgNPs on bacterial cell dynamics. The model explains untreated and AgNP-treated Escherichia coli growth in batch culture. Subsequently, fluorescence probing and atomic force microscopy and spectroscopy were used to experimentally study the AgNP bactericidal mechanism. This study shows that the mode of antibacterial action of silver nanoparticles includes penetration of AgNPs inside the cell/membrane, membrane permeabilization, reduction of the cell’s Young modulus and possible cell leakage while undistinguished reactive oxygen species level was detected inside lethally treated cells., Cilj ove teze je razjasniti način antibakterijskog djelovanja koloidnih nanočestica srebra na stanice Escherichia coli kao modelnog organizma. Istraživanje baktericidnog mehanizma srebrnih nanočestica provedeno je eksperimentalnim i teorijskim metodama kako bi se čim bolje opisao rast tretiranih bakterijskih stanica. Srebrne nanočestice su sintetizirane laserskom ablacijom u vodi, a dobiveni koloid je temeljito karakteriziran. Uveden je model rasta temeljen na interakcijama kako bi se razumio učinak nanočestica na dinamiku rasta bakterijskih stanica. Model objašnjava rast netretiranih i tretiranih bakterijskih stanica u tekućoj kulturi. Za eksperimentalno proučavanje baktericidnog mehanizma nanočestica korištene su fluorescentna spektroskopija te mikroskopija i spektroskopija atomskom silom. Istraživanje pokazuje da antibakterijsko djelovanje srebrnih nanočestica na bakterijske stanice uključuje njihov prodor unutar stanice/membrane, permeabilizaciju membrane, smanjenje Youngovog modula s vremenom te moguće istjecanje unutarstaničnog materijala, dok je inducirana razina reaktivnih kisikovih vrsta u smrtonosno tretiranim stanicama zanemariva.

Quaternary ammonium compounds (QACs) have long been praised for their powerful antimicrobial potential. With a wide range of application, from different industries to pharmacy and medicine, these compounds are omnipresent ingredients of many commercial products. However, recent studies show that as much as 83% of MRSA isolates are resistant toward most of the commonly used QACs. Therefore, elucidation of resistance mechanism(s) and development of new potent QACs are in the centre of further research in the field. Quinuclidine is a bicyclic part of alkaloids isolated from a bark of the Cinchona trees. Even though these alkaloids have long been used in a traditional medicine, the biological potential of quinuclidine derivatives is still underexplored. Apart from its anticholinergic, antiparasitic, antioxidative and antitumor activity, some quinuclidine derivatives have been shown to act on FitZ protein, known to be essential for bacterial cell division. All this motivated us to synthesize new quinuclidine containing QACs in order to further explore its antimicrobial potential and to develop new and powerful naturally derived QACs. Our studies have shown that quaternisation of substituted quinuclidine improves its antimicrobial activity by several hundred folds and that this activity depends on the type of substituent used for quaternisation. Aryl- quinuclidines typically had lower antimicrobial activity than their alkyl counterparts. Also, somewhat better antimicrobial activity was observed toward Gram-positive bacteria, suggesting that bacterial membrane might be involved in the mode of action mechanism. Therefore, the PI- uptake measurements were performed along with atomic force microscopy (AFM) and indeed, obtained results indicated bacterial membrane perforation. Moreover, alkyl-quinuclidines, more specifically an identified candidate with the longest alkyl chain, exhibited the best antimicrobial activity with the lowest MIC values across selected panel of the bacteria and good activity toward S. aureus biofilms. In addition, this compound had lower toxicity toward healthy human cell lines than the referent QACs, suggesting that quinuclidine could serve as a new possible core molecule for future QACs development.

Study of bactericidal effect of silver nanoparticles (AgNPs), free of chemical byproducts, could be of great importance in understanding the underlying antibacterial mechanism(s). In this work, we present the production and characterization of AgNPs, synthesized by laser ablation of silver in water, and optical density (OD) growth curves of E. coli cells treated at sub MIC (minimal inhibitory concentration) and MIC concentrations. Besides the increase of the baseline and reduction of the maximal OD, the main impact of the treatment is the increase of the lag time, which becomes infinitive for the MIC treatment. The obtained bacterial growth curves are described by a novel growth and inactivation model given by the set of three differential equations. Our model fits the OD data very well and explains the apparent lag phase, which we found to be a dynamic state in which the bacterial growth and death rates are close in value.

There are many growth and inactivation models in the literature which are purely empirical and do not poster a mechanistic insight [1, 2]. Here, we present a novel, intuitive and mathematically relatively simple model for bacterial growth and inactivation. The proposed model well describes E. coli batch culture growth curves obtained for different inoculum sizes and different initial nutrients concentration. The model is, furthermore, expanded in order to fit and explain growth curves obtained for E. coli treated with different concentrations of laser produced silver nanoparticles. The model is described by three nonlinear mutually dependent di erential equations that give time evolution of bacterial, nutrients and nanoparticle concentrations. These equations are of general nature and can be used to predict growth of different bacteria in batch culture since the equations do not include a parameter that is characteristic for E. coli. As our model can be linked to the logistic model, it is possible that the equations could be applicable to any single species growth. [1] M. Peleg and M. G. Corradini, Crit. Rev. Food Sci. Nutr. 51, 917 (2011) [2] K. D. Dolan and D. K. Mishra, Annu. Rev. Food Sci. Technol. 4, 401 (2013)

Antimicrobial peptides (AMPs) are endogenous antibiotics present in all organisms acting directly towards pathogens and also showing immunomodulatory properties. Skin secretions of many tested anuran species have been shown to contain a variety of bioactive peptides that often exhibit antibacterial activity, also against multidrug resistant isolates. We have selectively amplified transcripts likely to encode for AMPs, thus providing their sequences. After RNA extraction from frog skin tissue samples, cDNA synthesis followed by PCR amplification was performed. For this purpose, forward degenerate primers were designed based on highly conserved signal peptide regions from Ranidae, Hylidae and Bombinatoridae together with a reverse primer designed on the poly-A tail of mRNA. Signal peptide regions themselves were derived from sequences already available in DADP database and transcriptome data in SRA database. Resulting amplicons were size-selected and processed by ion semiconductor sequencing, obtaining several thousand sequencing reads which were then assembled into contigs representing nearly full-length AMP-encoding transcripts. Analysis of the assembled sequencing output allowed to identify more than a hundred full-length mature peptides from 5 different specimens belonging to 5 different frog species, mostly from Ranidae species. Based on appropriate biophysical properties (e.g. charge, hydrophobicity, amphipathicity) six of the most promising candidates were chosen for chemical synthesis and extensive characterization. All peptides were tested against a panel of Gram-negative and Gram-positive bacteria, as well on tumour cell lines. One peptide, identified in Rana arvalis, proved to be active against both reference ATCC strains and cancer cells, while non-toxic for human MEC-1 cells in a range of concentrations several folds higher then minimal inhibitory concentration (MIC) and IC50 values. Furthermore, the peptide was found to disrupt the bacterial membrane even at sub-MIC concentrations, as observed by flow cytometry and visualised by atomic force microscopy (AFM).

Skin secretions of many tested anuran species have been shown to contain a variety of antimicrobial peptides (AMPs) acting directly towards pathogens, including multidrug resistant isolates, and also showing immunomodulatory properties [1]. We have selectively amplified transcripts likely to encode for AMPs, thus providing their sequences. After RNA extraction from frog skin tissue samples, cDNA synthesis followed by PCR amplification was performed. For this purpose, forward degenerate primers were designed based on highly conserved signal peptide regions [2], together with a reverse primer designed on the poly-A tail of mRNA. Signal peptide regions were derived from sequences deposited in DADP database [3] and transcriptome data in SRA database [4]. Resulting amplicons were size-selected and processed by ion semiconductor sequencing, obtaining several thousand sequencing reads. Those were then assembled into contigs representing nearly full-length AMP-encoding transcripts. Analysis of the assembled sequencing output allowed to identify more than a hundred full-length mature peptides from 5 different specimens belonging to 5 different frog species, mostly from Ranidae species. Based on appropriate biophysical properties (e.g. charge, hydrophobicity, amphipathicity) six of the most promising candidates were chosen for chemical synthesis and extensive characterization. All peptides were tested against a panel of Gram-negative and Gram-positive bacteria, as well on tumour cell lines. One peptide, identified in Rana arvalis, proved to be active against both reference ATCC strains and epithelial cancer cell lines, while less toxic for circulating MEC-1 cells in a range of concentrations several fold higher then minimal inhibitory concentration (MIC) and IC50 values. Furthermore, the peptide was found to disrupt the bacterial membrane even at sub-MIC concentrations, as observed by flow cytometry and visualised by atomic force microscopy (AFM). [1] A. Nijnik and R. Hancock, Emerg. Health Threats J. 2, (2009). [2] V. Tessera, F. Guida, D. Juretić, and A. Tossi, FEBS J. 279, 724 (2012). [3] M. Novković, J. Simunić, V. Bojović, A. Tossi, and D. Juretić, Bioinformatics 28, 1406 (2012). [4] Y. Kodama, M. Shumway, and R. Leinonen, Nucleic Acids Res. 40, D54 (2012).

There are numerous reports of antibacterial activity of the silver nanoparticles (AgNPs) produced by chemical methods [1, 2], but the laser ablation in liquid has been proven to be the most convenient technique for the production of nanoparticles free of chemical byproducts. Here we present the study of AgNPs and the influence these nanoparticles have on the bacteria and their growth kinetics. AgNPs were produced by laser ablation in water and characterized by different experimental methods. In the present literature, there are only few reports on attempts to model the bacterial growth in the presence of nanoparticles [3]. To explain the obtained growth curves of untreated and treated bacterial cells, we have designed the novel microscopic growth model described by three differential equations. Our model gives a good description of the experimentally obtained data.

There are numerous reports of antimicrobial activity of the silver nanoparticles (AgNPs) produced by different methods (1-2), but the laser ablation in liquid has been proven to be the most convenient technique for the production of nanoparticles free of chemical byproducts. Here we present the study of AgNPs and the influence these nanoparticles have on the bacteria and their growth kinetics. AgNPs were produced by laser ablation in water and characterized by different experimental methods. Their antibacterial activity was observed by different microscopic techniques. In the present literature, there are only few reports on attempts to model the bacterial growth in the presence of nanoparticles. The reported model (3) is just a modification of the empirical Gompertz model. To explain the obtained growth curves of treated and untreated bacterial cells, we have designed the novel microscopic model described by three differential equations. Our model gives a good description of the experimentally obtained data. References: 1. I. Sondi and B. Salopek- Sondi, J. Colloid Interface Sci. 275 (2004) 177-182. 2. G. A. Martinez-Castanon et al., J Nanopart Res 10 (2008) 1343–1348. 3. T. Chatterjee, et al., Biochim. Biophys. Acta 1850 (2014) 299–306.

Given the current global problem of multidrug resistance in human opportunistic pathogens, antimicrobial peptides (AMPs) show significant potential as novel anti-infective agents1. Notably, they show broad spectrum antibacterial activity and their most common mode of action, namely the disruption of bacterial membrane and/or the formation of pores or lesions, makes it hard for bacteria to gender permanent resistance. The toxicity towards host cells is however a key limitation factor, which imposes the necessity to design artificial AMPs de novo, or redesigning/mutating known natural AMPs to have an increased selectivity index (SI) and reduce side effects. We report four novel Gly and Lys rich peptides which were computationally designed based on a set of training natural peptides with low reported MIC values against E.coli and taking into account a defined set of parameters ; net charge, hydrophobicity, number of Lysines, etc. One additional peptide was designed by creating the specular image of a previously designed sequence. All peptides were named kiadins, from the first three residues in the ‘parent’ peptide sequence1. Susceptibility testing was performed on clinically relevant, drug- resistant bacteria chosen among the ESKAPE pathogens ; carbapenem-resistant Acinetobacter baumannii and Pseudomonas aeruginosa, 3rd generation cephalosporin-resistant E. coli and Klebsiella pneumoniae, as well as meticillin- resistant Staphylococcus aureus (MRSA). The results show different range of antibacterial activity with respect to Gly substitutions, where the peptides with higher proportion of Gly showed significantly weaker bactericidal effects. The best antibacterial activity was observed for Kiadin-2, which exhibited MIC values from 0.25 to 8 µM, a promising SI values and low cytogenotoxicity on HPBLs, making it a suitable candidate for further development.

Predstavljamo projekt izgradnje istraživačkih kapaciteta u MEMS tehnologiji u Hrvatskoj kroz nedavno odobreni projekt financiran iz Europskih strukturnih fondova, zasnovan na suradnji između Sveučilišta i tvrtke u kategoriji malog i srednjeg poduzetništva.

Laser ablation in liquids is a very convenient technique for production of nanoparticles free of chemical byproducts. Silver nanoparticles made in this way could show antimicrobial activity. For the purpose of this work we produced colloidal silver nanoparticles in water using a 1064 nm ns Nd:YAG laser at 100 mJ output energy and 5 Hz repetition rate. Size- distribution of nanoparticles was obtained from atomic force microscopy (AFM) images. We found the average diameter of silver nanoparticles to be around 10 nm with a relatively narrow size distribution. Obtained colloidal solution is stable on a scale of few months, which implies that nanoparticles are well dispersed and/or charged. We use this colloidal solution to investigate antimicrobial effects of silver nanoparticles on different strains of E. coli bacteria where the changes on bacterial membrane are monitored by AFM. Here we present our preliminary results.

We present an initiative to build research capacities in MEMS technology in Croatia through recently granted project financed by the European structural funds, based on the co- operation between the public university and SME.

Laserska ablacija u tekućini je jedinstvena tehnika za proizvodnju nanočestica bez kemijskih nusprodukata koji mogu utjecati na rezultate istraživanja. Srebrne nano- čestice, dobivene na ovaj način, trebale bi također pokazati antimikrobnu aktivnost. U našem laboratoriju proizveli smo srebrne koloidne nanočestice u vodi koristeći nanosekundni Nd:YAG laser valne duljine 1064 nanometara, izlazne energije 100 mJ te frekvencije 5 Hz. Distribuciju promjera nanočestica dobili smo iz snimaka napravljenih pretražnikom atomskih sila (AFM). Prosječni promjer srebrnih nanočestica iznosio je oko 10 nanometara uz relativno usku distribuciju. Stabilnost koloida ispitivali smo analizom apsorbcijskog spektra. Dobiveni koloid pokazao se stabilnim tokom nekoliko mjeseci iz čega možemo zaključiti da su dobivene nanočestice dobro raspršene. Koristeći ovaj koloid istraživali smo antimikrobne efekte srebrnih nanočestica na sojevima E. coli bakterija promatrajući promjene bakterijske membrane također pomoću AFM-a. U ovom radu predstavljamo inicijalne rezultate istraživanja.

Nanoparticles of various materials are today implemented in wide variety of industrial, scientific and medical applications. Biocompatible nanoparticles are used for cell treatment ; as nano-biomarkers, for therapy and diagnostics. Nanoparticles can be synthesized conventionally either using wet chemistry methods or gas phase processes. Such nanoparticles are often characterized by impurities which are reaction products of additives and precursors. Laser ablation in liquids appeared to be a solution for that drawback. It is recognised as simple and versatile technique and there are almost no limitations in a selection of materials for the nanoparticle synthesis. The remarkable advantages of this technique over other techniques are absence of impurities in the final product (free of contaminating processes, chemical precursors not required), possibility of preparation of multicomponent nanoparticles, weak agglomeration, narrow size distribution of nanoparticles, etc. In our work we studied laser synthesized silver nanoparticles produced in water by ns Nd:YAG laser operating at 1064 nm with 100 mJ of output energy and 5 Hz of repetition rate. From AFM figures size- distribution of nanoparticles is obtained. Laser ablation is monitored by optical emission spectroscopy while colloidal solution is characterized by photospectrometry and photoluminescence. Our results show that average diameter of laser synthesized silver nanoparticles is around 10 nm and size distribution is relatively narrow. Our colloidal solution is very stable in time (few months) implying that nanoparticles are well dispersed and/or charged.

Giant unilamellar vesicles (GUVs) are widely used as artificial cell membrane models. Electroformation is the most commonly used method for their production. The method depends on multiple parameters, so reproducible production of high quality GUVs can be challenging, especially when working with lipid mixtures containing cholesterol concentrations reaching and surpassing the bilayer saturation threshold. After analyzing existing electroformation protocols, we identified two key issues for such mixtures. The first issue is related to experiment reproducibility. Most protocols use the drop-deposition method which produces lipid films of nonuniform thickness, resulting in lower experiment reproducibility and efficiency of GUV formation. The second is cholesterol demixing in the form of anhydrous crystals during lipid film drying, resulting in an artifactual decrease of cholesterol concentration in GUVs compared to the initial lipid mixture. To deal with the first problem, we first tested electroformation from such mixtures with replacement of the drop-deposition step by spin-coating. Optimizing this process improved the GUV size and yield, but the cholesterol demixing issue remained. To address this issue, we developed a new protocol which combines the rapid solvent exchange, plasma cleaning and spin-coating techniques to produce GUVs by electroformation from damp lipid films. We believe that this new improved electroformation protocol will allow us to successfully study models of eye lens fiber cell membranes with their very high Chol content., Divovske unilamelarne vezikule (DUV) naširoko se koriste kao modeli staničnih membrana. Elektroformacija je najčešće korištena metoda za njihovu proizvodnju. Metoda ovisi o više parametara, tako da ponovljiva proizvodnja visokokvalitetnih DUV-ova može biti izazovna, osobito pri radu s lipidnim smjesama koje sadrže koncentracije kolesterola koje dosežu i premašuju prag zasićenja membranskog dvosloja. Nakon analize postojećih protokola elektroformacije, identificirali smo dva ključna problema za takve smjese. Prvi se odnosi na ponovljivost eksperimenata. Većina protokola koristi metodu nakapavanja koja proizvodi lipidne filmove nejednake debljine, što rezultira smanjenom ponovljivošću eksperimenata i učinkovitošću stvaranja DUV-ova. Drugi je izlučivanje kolesterola u obliku dehidriranih kristala tijekom sušenja lipidnog filma, što dovodi do artefaktualnog smanjenja koncentracije kolesterola u DUV-ovima u usporedbi s početnom lipidnom smjesom. Kako bismo umanjili prvi problem, testirali smo elektroformaciju iz takvih smjesa uz zamjenu koraka nakapavanja tehnikom brzog razmazivanja. Optimizacija ovog procesa poboljšala je veličinu i prinos DUV-ova, ali i dalje je preostalo pitanje izlučivanja kolesterola. Potaknuti time, razvili smo novi protokol koji kombinira tehnike brze izmjene otapala, čišćenja plazmom i brzog razmazivanja za proizvodnju DUV-ova elektroformacijom iz vlažnih lipidnih filmova. Smatramo da će nam ovaj novi protokol pomoći u daljnjem istraživanju svojstava membrana s vrlo visokim udjelom kolesterola, poput vlaknastih stanica leće oka.

Neobnovljivi izvori energije će se iscrpiti te je nužno prijeći na obnovljive izvore energije. Sunce je neiscrpan izvor energije koji se ni približno dovoljno ne iskorištava što je velika prednost solarnim ćelijama općenito. Bojom senzibilizirane solarne ćelije pripadaju trećoj generaciji solarnih ćelija, a ističu se zbog niske cijene, širokog raspona mogućnosti primjene, jednostavne strukture i načina rada. S obzirom na ogroman napredak u efikasnosti u zadnjih nekoliko desetljeća, te većoj zainteresiranosti znanstvenika, očekivano je povećanje efikasnosti u bližoj budućnosti. Dakle, imaju brojne prednosti, a nedostaci se konstantno poboljšavaju te je neupitan njihov ogroman potencijal, pa je očekivano da će u budućnosti značajan udio iskorištene energije biti proizveden ovim ćelijama., Non-renewable energy sources will be exhausted, so it is necessary to switch to renewable energy sources. The sun is an inexhaustible source of energy that is not used nearly enough, which is a big advantage for solar cells in general. Dye sensitized solar cells belong to the third generation of solar cells, and they stand out due to their low price, various application possibilities, simple structure and working principle. Considering the enormous progress in efficiency in the last few decades, and the greater interest of scientists, an increase in efficiency is expected in the near future. So, they have numerous advantages, and the disadvantages are constantly improving, and their great potential is unquestionable, so it is expected that in the future a significant share of the used energy will be produced by these cells.

Antimicrobial resistance, which can be partially attributed to irresponsible and improper use of antibiotics, remains a major concern worldwide in recent decades. One of the potential alternatives to classical antibiotic treatment are antimicrobial peptides (AMPs) - structurally different (macro)molecules with direct antimicrobial and/or immunomodulatory properties. Helminths, a group of parasitic organisms, successfully co-exist in bacteria-rich surroundings during their life cycle, and among the possible mechanisms of defense of these organisms are likely be AMPs. By exploring the genomes of several helminth species, 31 AMPs were obtained, after which a bioinformatics analysis was performed and 3 were selected for further characterization. Moreover, several N- and C-terminal fragments of a peptide identified in Taenia solium were synthesized to conduct structure-activity studies. All selected peptides, as well as the fragments, were characterized by structural, antimicrobial, and cytotoxic tests, and modeled by molecular dynamics simulations. Most of the peptides showed broad-spectrum antimicrobial activity, with (mostly) acceptable levels of toxicity, which warrants further mode-of-action studies, and development for possible biomedical applications., Rezistencija bakterija na antibiotike, za koju je djelomično zaslužna neodgovorna i pretjerana uporaba, gorući je problem diljem svijeta u zadnjim desetljećima. Kao potencijalna alternativa klasičnim antibioticima nameću se antimikrobni peptidi (AMP), strukturno različite makromolekule koje imaju direktnu antimikrobnu i/ili imunomodulatornu aktivnost. Plošnjaci, skupina nametničkih organizma, uspješno ko-egzistira u okolini (najčešće) bogatoj bakterijama, a jedan od mogućih mehanizama obrane ovih organizama su AMP-i. Analizom genoma većeg broja plošnjaka, identificirali smo 31 AMP, te nakon detaljne bioinformatičke analize odabrali tri peptida za daljine istraživanje. Istovremeno, u cilju ispitivanja utjecaja pojedinih segmenata peptidne sekvence na aktivnost i mehanizam djelovanja, sintetizirani su N- i C-fragmenti peptida identificiranog u Taenia solium. Svi odabrani peptidi, kao i fragmenti, karakterizirani su putem strukturalnih, antimikrobnih i citotoksičnih ispitivanja, te istraženi na molekularnoj razini korištenjem simulacija molekularne dinamike. Većina peptidi pokazuje antimikrobnu aktivnost širokog spektra, dok je toksičnost često u prihvatljivim okvirima, što otvara vrata daljnjim ispitivanjima mehanizama djelovanja, kao i potencijalnoj biomedicinskoj uporabi.

U radu je prikazan eksperimentalni uređaj za mjerenje frekvencije srca koji funkcionira na principu transmisijske i refleksijske fotopletizmografije u spektru vidljive svjetlosti. Glavne komponente uređaja korištenog u istraživanju su platforma Arduino Uno i senzor KY-018 Fotowiderstand Modul. U eksperimentu su korištene različite konfiguracije uređaja, a za potrebe njegovog rada napisan je programski kod u programskom jeziku C za softverski integrirano razvojno okruženje Arduino. Broj otkucaja srca određen je upotrebom različitih izvora svjetlosti. Podaci su iz uređaja Arudino Uno očitani bez i uz upotrebu algoritma. Valjanost dobivenih podataka provjerena je pomoću pametnog mobitela sa senzorom za mjerenje otkucaja srca. Zaključeno je da su najbolji rezultati mjerenja dobiveni u slučaju svjetlosti koja se najmanje apsorbira u tkivu i koja je jačeg intenziteta. Na kraju rada ukazano je na neka ograničenja i mogućnosti poboljšanja uređaja., The paper presents an experimental device for measuring heart rate that works on the principle of transmission- and reflectance-mode photoplethysmography in the spectrum of visible light. The main components of the device used in the research are the Arduino Uno platform and the KY-018 Fotowiderstand Module sensor. Different configurations of the device were used in the experiment and, for the purpose of its functioning, the program code was written in the C programming language for the integrated development environment Arduino. The heart rate was determined using different light sources. Data were taken from the Arudino Uno device without and with the use of the algorithm. The validity of the obtained data was checked using a smartphone with a heart rate sensor. It was concluded that the best measurement results were obtained in the case of light that is least absorbed in the tissue and which is of stronger intensity. At the end of the paper, some limitations and possibilities for improving the device were pointed out.

U ovom završnom radu izgrađuje se multimetar pomoću Arduino platforme. Arduino je softverska i hardverska platforma koja korisniku nudi provedbu raznih projekata. To se postiže uz spajanje raznih električnih komponenti, senzora i aktuatora s mikrokontrolornom pločom te programiranjem njihovih funkcija na računalu. Ovdje se Arduino spaja i programira tako da mjeri napon i struju nekog strujnog kruga te nepoznati otpor i kapacitet. Dobivaju se poprilično točni rezultati, a neke greške se javljaju radi nedostatka elektroničkih komponenti te velikog dometa provjeravanih komponenti. Cilj rada je prikazati mogućnosti Arduino platforme. Arduino široj znanstvenoj zajednici nudi moćan instrument koji je dostupan po vrlo pristupačnoj cijeni i time je odličan za školstvo i edukaciju mnogih., In this bachelor thesis, a multimeter is built using the Arduino platform. Arduino is a software and hardware platform that offers users the implementation of various projects. This is achieved by connection of various electrical components, sensors and actuators to the microcontroller board and with programming their functions on the computer. Here the Arduino is connected and programmed to measure voltage and current of a circuit and the unknown resistance and capacitance. The results obtained are quite accurate, some error occur due to the lack of electronic components and a large range of the tested components. The aim of this thesis is to present the capabilities of the Arduino. The Arduino offers the wider scientific community a powerful instrument that is available at a very affordable price and thus is great for the schooling and education of many.

U ovom radu dan je primjer istraživački usmjerene online nastave za učenike drugog razreda srednje škole na temu Rad i snaga električne struje. Lekcija je pripremljena uz pomoć virtualnih laboratorija na Go-Lab portalu te Google obrasca koji je, uz prenošenje sadržaja, dao uvid u statistiku uspješnosti nastave. Napravljeni su i odgovarajući web sadržaji koji su uključivali pred-kviz za ponavljanje gradiva te tekst teorijskog objašnjenja lekcije. Istraživački pristup ostvaren je pomoću virtualnog laboratorija u kojemu se snaga grijača određivala mjerenjem brzine promjene temperature vode u kalorimetru. U drugom virtualnom laboratoriju mjerila se ovisnost snage žarulje o njezinom otporu uz konstantni napon te o naponu uz konstantni otpor. Analizom i sintezom rezultata vlastitih mjerenja učenici su trebali formulirati matematički izraz za snagu električne struje. Nakon toga su ispunjavali upitnik s konceptualnim pitanjima te zadacima vezanim uz svakodnevni život, koji provjeravaju sposobnost primjene usvojenog znanja. Rezultati istraživanja ukazuju na učeničke poteškoće, najčešće u crtanju i interpretaciji grafova. Sadržaj lekcije prilagodili smo mogućnostima Go-Lab sjedišta kreiranjem vlastitog okruženja za učenje istraživanjem (ILS-a). Također smo oblikovali odgovarajuće upute za izradu ILS-a, kako bismo s ovom metodom rada upoznali i druge nastavnike., This thesis we present an example of a research-based online lesson for second grade high school students on the topic of Work and Power of Electric Current. The lesson was prepared with the help of virtual laboratories on the Go-Lab portal and the Google form which, in addition to providing content, also provided insights into the statistics of the success of the lesson. Related web content was also created, including a pre-quiz for the subject repetition and the text of the theoretical explanation of the lesson. The research approach was implemented using a virtual laboratory in which the power of the heater was determined by measuring the rate of change of water temperature in the calorimeter. Another virtual lab was used to measure the dependence of the lamp's output on its resistance at constant voltage and on the voltage at constant resistance. By analysing and interpretating the results of their own measurements, the students were able to formulate a mathematical expression for the power of electric current. Finally, they completed a questionnaire with conceptual questions and tasks related to everyday life, that tested their ability to apply the knowledge they had acquired. The results of the investigation indicate student difficulties, most often in drawing and interpreting graphs. We adapted the lesson content to the Go-Lab platform by creating our own ILS. We have also designed a suitable guide for developing ILS to introduce other teachers to this method.

Predstavljamo primjer metodički oblikovane online nastavne lekcije iz fizike kreirane uz pomoć virtualnog laboratorija i Google obrasca koji omogućuje jednostavnu transakciju sadržaja, kao i statistički uvid u povratne informacije učenika. U radu je prikazana metoda poučavanja vođenim istraživanjem koja je napravljena u skladu s edukacijskim preporukama utemeljenim na rezultatima istraživanja. Sagledali smo u kojoj mjeri online nastavna lekcija omogućuje samostalno učenje učenika prvih razreda srednje škole na temu Elastična potencijalna energija te pregledali internetske resurse s ciljem pronalaska lako dostupnih materijala i alata svim nastavnicima. Učenje se sastoji od tri dijela. Prvi dio započinje opažanjem pojava unutar virtualnog laboratorija i postavljanjem hipoteza. Ispituje se ovisnost elastične potencijalne energije o pojedinim varijablama. Koristeći se analizom rezultata mjerenja, učenici provjeravaju ispravnost postavljenih hipoteza te izraz za elastičnu potencijalnu energiju opruge sintetiziraju empirijski. Drugi dio posvećen je teorijskom razmatranju u kojem učenici do istog izraza trebaju doći teorijskim razmatranjem. U posljednjem dijelu nastavnik provjerava znanje učenika u vidu različitih konceptualnih i numeričkih zadataka koji predstavljaju problemske situacije iz svakidašnjeg života. Vrednovanje nastavne lekcije zamišljeno je kroz vrednovanje ishoda učenja danih u Nacionalnom kurikulumu vezanih uz istraživanje fizičkih pojava. U istraživanju je sudjelovalo 30 učenika prvog razreda gimnazije. Analiza učeničkih odgovora pokazala je da je približno polovica učenika mogla samostalno provesti istraživanje i u potpunosti savladati ishode učenja., We present an example of a methodologically designed online physics lesson created by using a virtual lab and a Google form that allows both easy transaction of content and statistical insight into student feedback. The paper presents a method of instructionally guided research created in accordance with pedagogical recommendations based on research findings. We explored the extent to which online lessons facilitate first grade students' independent learning on the topic of Elastic Potential Energy and reviewed online resources with the goal of finding easily accessible materials and tools for all teachers. The lessons consist of three parts. The first part begins with observing phenomena in a virtual laboratory and making hypotheses. The dependence of elastic potential energy on individual variables is investigated. By analyzing the results of measurements, students verify the correctness of the hypotheses made and empirically compile the expression for the elastic potential energy of the spring. The second part is devoted to theoretical consideration, in which students arrive at the same expression through theoretical consideration. In the last part, the teacher checks the knowledge of the students in the form of various conceptual and numerical tasks that represent problem situations from everyday life. The evaluation of the lesson was conceived on the basis of the assessment of the learning outcomes related to the study of research of physical phenomena given in the National Curriculum. Thirty first grade high school students participated in the study. Analysis of student responses showed that about half of the students were able to conduct independent research and fully master the learning outcomes.