1. How to establish a novel liver cell culture system that resembles the in vivo liver microenvironmentEneko Madorran, 2025, original scientific article Abstract: Subtle physiological patterns within the human organism, such as molecular fluctuations or systemic adaptations, often remain hidden from direct observation due to the inherent variability and noise within biological samples. The liver, a vital organ essential for systemic regulation and toxicological assessment, presents this challenge due to its heightened activity, which can influence enzyme dynamics and metabolic processes. Unlike direct observation, in vitro liver models offer a more precise means of understanding these trends, providing insights into the organ's (patho)physiology, and serving as a platform for toxicity evaluation. However, current liver models lack essential features required to faithfully replicate the liver's microenvironment, resulting in reduced accuracy in toxicity assessments. Furthermore, while researchers emphasize mechanistic insights, such as the molecular processes governing glucose metabolism and cellular energy production, clinicians focus on tangible health outcomes, like blood glucose levels in patients. This disconnect between the objectives and methodologies of basic researchers and clinicians amplifies this gap, hindering effective translational research. Addressing these challenges, a novel liver cell culture system that resembles the in vivo liver microenvironment with clinical instrumentation has been proposed to enhance current liver models, improving their capacity to emulate in vivo conditions. This study introduces a novel liver cell culture system, utilizing a 96-well plate format incorporating hepatic sinusoidal endothelial cells, hepatic stellate cells, Kupffer cells, and hepatocytes to replicate the liver microenvironment. The model integrates clinical instrumentation, specifically a biochemical analyzer to ensure biomarkers closely align with those observed in clinical diagnostics. This design enables researchers to fine-tune conditions that mimic the liver's microanatomy and physiological responses, enhancing its translational potential for toxicity assessments. The approach involves primary cell culture preparation, supernatant analysis through a clinical biochemistry analyzer, and cell viability assessment using the Membrane Potential Cell Viability Assay (MPCVA) method. Additionally, advanced imaging techniques and data analysis tools are incorporated to refine the model's capabilities and ensure greater translatability to clinical applications.
Keywords: in vitro toxicity, liver (patho)physiology, liver in vitro model, membrane potential cell viability assay, translational research
Published in DKUM: 14.08.2025; Views: 0; Downloads: 8
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2. Energy intake models for intermittent operation of dead-end microfiltration filling lineJure Ravnik, Gorazd Bombek, Aleš Hribernik, Timi Gomboc, Matej Zadravec, Aleks Kapun, Grega Hrovat, Jure Gradišek, Matjaž Hriberšek, 2022, original scientific article Abstract: In filling lines equipped with membrane separation devices in the form of filters energy,
consumption is only one of the important working parameters, the other being sustainable filter
performance in terms of separation efficiency. As the filling line is typically equipped with a valve,
intermittent operation of the filter is an important form of its use. Whereas the overall energy
consumption of the filtration process is governed by the continuous operation mode, the intermittent
mode, characterised by opening/closing of the valve, contributes most to problems of filter failure,
i.e., the breakthrough of filtered particles through the membrane. A model for determination of the
energy intake of a microfiltration membrane during the opening and closing of a valve is presented
in this work. The model is based on computational analysis of the pressure wave signals recorded
during the opening/closing of the valve using Fourier transform, and expressed in a nondimensional
filter area specific energy intake form. The model is applied to a case of constant pressure dead-end
microfiltration with three filter types: a single membrane filter, a stacked filter and a pleated filter
with filtration surface areas ranging from 17.7 cm2
to 2000 cm2. Both clean filters, as well as partially clogged filter cases are taken into account. Second order polynomial models of the energy intake are developed and evaluated based on extensive analysis of the experimental data. The analysis of energy intake results show that the largest energy intake was observed for the clean filter case. When membrane fouling occurs at the constant flow rate values it leads to larger energy intake, however,
due to a decreasing specific flow rate during fouling these values do not exceed the clean filter case.
Keywords: membrane filtration, water hammer effect, membrane energy intake, filter clogging
Published in DKUM: 28.03.2025; Views: 0; Downloads: 2
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3. Structural and chromatographic characterization of cation‑exchange membranes based on carboxymethyl/ nanofbrillated cellulose using lysozymeVanja Kokol, Tina Simčič, Urh Černigoj, 2025, original scientific article Abstract: Bio-based membranes are becoming highly-desired low-cost, environmentally friendly, and readily available supports for the separation and purification of biomacromolecules. In this work, weak cation-exchange and highly (> 95%) microporous (> 80 μm) cellulose-based membranes were prepared from different weight ratios of carboxymethyl cellulose (CMC) as anionic polymer and cellulose nanofibrils (CNFs) as a stabilizing and structural filler, by the freeze-casting process and citric-acid (CA) mediated in situ cross-linking (esterification). It was ascertained that mono-esterified/grafted CA also contributes to the total carboxylic groups (1.7–2.6 mmol/g), while the CMC-induced CNF orientation affected the membrane’s morphology and lysozyme (Lys) binding capacity. A static binding capacity (SBC) between 370 and 1080 mg/g, and equilibrium within 3.3 h for 1 g/mL Lys was thus achieved with increasing the total solid and CMC content by forming more isotropic microporous structures. The selected membranes were then packed in a chromatographic housing, analyzed for pressure drop, and evaluated for dynamic binding capacity (DBC), depending on the process performance (flow rates, Lys concentration). A DBC in the 165–417 mg/g range was determined at a throughput of 0.5 mL/min, and elution yield of 78–99% with > 95% recovery. The Lys adsorption and transfer were reduced by the increasing flow rate and membrane density due to compressibility issues, resulting in smaller and irregularly distributed pores and the unavailability of carboxylic groups. Although the DBC was still comparable with the commercial CIM® monoliths, the convection-based transport of molecules inside the membrane and the membrane stiffness needs to be improved in further research.
Keywords: cation-exchange membrane, cellulose nanofibrils, carboxymethyl cellulose, citric acid, lysozyme, static and dynamic binding capacity
Published in DKUM: 17.03.2025; Views: 0; Downloads: 7
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4. Napredni biološko aktivni transportni sistemi ekstraktov manga (Mangifera indica) za terapevtske aplikacije in vitro : doctoral dissertationNika Kučuk, 2024, doctoral dissertation Abstract: Doktorska disertacija je razdeljena na štiri dele, v katerih sta obravnavana študija biološke aktivnosti ekstraktov olupkov manga (MPE) ter optimiranje sinteznih metod transportnih sistemov (alginatne kroglice, liposomi in micelarne membrane mikrobnega izvora), vključno z njihovo funkcionalizacijo z MPE za različne terapevtske aplikacije (in vitro).
V prvem delu smo s pomočjo različnih ekstrakcijskih metod iz svežih in suhih olupkov manga pridobili MPE z visoko vsebnostjo totalnih fenolov, proantocianidinov, posameznih fenolnih spojin, proteinov in nekaterih encimov v aktivnih oblikah ter z visoko antioksidativno aktivnostjo. Nadalje smo potrdili protibakterijske lastnosti testiranih MPE na rast Gram-negativnih (Escherichia coli, Pseudomonas aeruginosa) ter Gram-pozitivnih bakterij (Bacillus cereus, Staphylococcus aureus). Z uporabo nizkocenovnega postopka sušenja olupkov manga na zraku smo ohranili biološko aktivnost MPE. Na vsebnost bioaktivnih spojin in biološko aktivnost pa sta vplivala tudi uporabljena ekstrakcijska metoda in topilo. Ultrazvočni etanolni MPE iz suhih olupkov je zagotovil dobro protibakterijsko in antioksidativno delovanje, zato smo ga v nadaljevanju vgradili v različne transportne sisteme.
V drugem delu smo MPE enkapsulirali v alginatne kroglice, proizvedene z metodo ionotropske gelacije. Sintetizirani napredni biološko aktivni sistem izkazuje dobro termično stabilnost s temperaturo degradacije nad 270 °C. Prav tako ni bilo prisotnih razpok na površini sintetiziranih liofiliziranih alginatnih kroglic. Alginatne kroglice so se izkazale kot odličen transportni sistem za enkapsuliran MPE zaradi stabilnosti pri kislih želodčnih pogojih in posledično doseganje tarčnega mesta (črevesje), kjer smo dosegli postopno sproščanje MPE, kar pomembno vpliva na uspešno absorpcijo v telesu. S protibakterijsko študijo smo dokazali pomemben inhibitorni učinek alginatnih kroglic z MPE na rast patogenih črevesnih bakterij E. coli in S. aureus.
V tretjem delu smo optimirali sintezo liposomov za uspešno enkapsulacijo MPE. Za formulacijo liposomov smo uporabili metodo hidracije tankega lipidnega filma z dodatkom 5 mm steklenih kroglic in 24-h stresanjem. Zelo stabilno in monodisperzno formulacijo liposomov z velikostjo delcev <300 nm smo dosegli z uporabo polarnega organskega topila etanola, z uporabo nepolarnih topil pa so najboljšo stabilnost izkazovali liposomi, proizvedeni s kloroformom. Uspešno smo enkapsulirali MPE v liposome in potrdili profil enakomernega sproščanja MPE, vključno z inhibitorno aktivnostjo na rast E. coli in S. aureus. Sintetizirani liposomi z enkapsuliranim MPE se med postopkom liofilizacije niso degradirali, prav tako so bili termično stabilni in ni bilo zaznati signifikantnih kemijskih interakcij med enkapsulirano bioaktivno spojino ter lipidi fosfolipidnega dvosloja.
V četrtem delu smo optimirali gojenje terapevtskih gob Ganoderma lucidum in Pleurotus ostreatus s submerzno kultivacijo za doseganje najvišjega prirasta in posledično najintenzivnejše tvorbe micelarnih membran s sposobnostjo visoke absorpcije vode. Sestava in pH rastnega medija sta imela ključen vpliv na tvorbo membran ter njihove karakteristike. Najustreznejši rastni pogoji so bili zagotovljeni pri bazičnem pH v mediju, ki je vseboval sladni ekstrakt za G. lucidum, in v mediju z dodatkom glukoze za P. ostreatus. Zaradi hidrofilne spodnje površine micelarnih membran so te izkazovale visoko sposobnost absorpcije vode, in sicer več kot 600 % za P. ostreatus in do 500 % za G. lucidum. Potrdili smo prisotnost širokega spektra funkcionalnih skupin, termično stabilnost s temperaturo degradacije nad 260 °C ter vlaknasto in porozno strukturo nastalih micelarnih membran. Micelarne membrane obeh gob smo nadalje uspešno funkcionalizirali z MPE ter dosegli enakomeren in postopen profil sproščanja MPE, vključno z uspešnim protibakterijskim delovanjem funkcionaliziranih membran na rast E. coli in S. aureus.
Keywords: mango, ekstrakcija, polifenolne spojine, protibakterijska aktivnost, transportni sistemi, alginatne kroglice, liposomi, micelarne membrane, funkcionalizacija, enkapsulacija, vgrajevanje, in vitro sproščanje, karakterizacija
Published in DKUM: 02.10.2024; Views: 0; Downloads: 65
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5. Antibacterial Komagataeibacter hansenii nanocellulose membranes with avocado seed bioactive compoundsKaja Kupnik, Mateja Primožič, Vanja Kokol, Željko Knez, Maja Leitgeb, 2024, original scientific article Abstract: Biocompatible, mechanically stable, highly hydrophilic/swellable and safe antibacterial biomaterials are crucial for wound dressing and other applications in the health sector. Therefore, this study was conducted for the development of bacterial nanocellulose membranes, which were, for the frst time, enriched with bacteriostatic and bactericidal efective avocado seed extracts prepared by diferent extraction techniques (ultrasonic, Soxhlet, high pressure with supercritical carbon dioxide). First, the production process of bacterial nanocellulose membranes from Komagataeibacter hansenii bacteria was optimized related to the fermentation media composition and
culture conditions, resulting in bacterial nanocellulose membranes with up to 83% crystallinity and 54.5 g/L
yield. The morphological structure of the membranes was varied further by using air- and freeze-drying
processes. The Soxhlet and high pressure with supercritical carbon dioxide avocado seed extracts with the
most charge negative surface (-33 mV) and smallest hydrodynamic size (0.1 µm) thus resulted in 100%
reduction of both Gram-negative Escherichia coli and Gram-positive Staphylococcus aureus with up to log
reduction of 2.56 and up to 100% bactericidal efficacy after 24 h of exposure, and at 14 mg/g of avocado
seed extracts integrated in the bacterial nanocellulose membranes homogeneously. The high swelling (up
to 600%) and water retention ability of avocado seed extracts enriched bacterial nanocellulose membranes,
with a biocidal release up to 2.71 mg/mL, shows potential for antibacterial applications in the biomedicine, cosmetics, and pharmaceutical industries.
Keywords: bacterial nanocellulose membrane, avocado seed extracts, extraction, swelling, release, antibacterial activity
Published in DKUM: 23.08.2024; Views: 70; Downloads: 15
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6. pH-dependence of glucose- dependent activity of beta cell networks in acute mouse pancreatic tissue sliceSandra Postić, Marko Gosak, Wen-Hao Tsai, Johannes Pfabe, Srdjan Sarikas, Andraž Stožer, Dean Korošak, Shi-Bing Yang, Marjan Rupnik, 2022, original scientific article Abstract: Extracellular pH has the potential to affect various aspects of the pancreatic beta cell function. To explain this effect, a number of mechanisms was proposed involving both extracellular and intracellular targets and pathways. Here, we focus on reassessing the influence of extracellular pH on glucose-dependent beta cell activation and collective activity in physiological conditions. To this end we employed mouse pancreatic tissue slices to perform high-temporally resolved functional imaging of cytosolic Ca2+ oscillations. We investigated the effect of either physiological H+ excess or depletion on the activation properties as well as on the collective activity of beta cell in an islet. Our results indicate that lowered pH invokes activation of a subset of beta cells in substimulatory glucose concentrations, enhances the average activity of beta cells, and alters the beta cell network properties in an islet. The enhanced average activity of beta cells was determined indirectly utilizing cytosolic Ca2+ imaging, while direct measuring of insulin secretion confirmed that this enhanced activity is accompanied by a higher insulin release. Furthermore, reduced functional connectivity and higher functional segregation at lower pH, both signs of a reduced intercellular communication, do not necessary result in an impaired insulin release.
Keywords: insulin secretion, membrane excitability, potassium channels, beta cell network, collective activity, calcium waves, pancreatic islets, pH-dependence
Published in DKUM: 01.07.2024; Views: 149; Downloads: 7
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7. Efficiency of neat and quaternized-cellulose nanofibril fillers in chitosan membranes for direct ethanol fuel cellsMaša Hren, Damjan Makuc, Janez Plavec, Michaela Roschger, Viktor Hacker, Boštjan Genorio, Mojca Božič, Selestina Gorgieva, 2023, original scientific article Keywords: chitosan, cellulose nanofibrils, anion exchange membrane, direct alkaline alcohol fuel cells
Published in DKUM: 10.04.2024; Views: 222; Downloads: 26
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8. Cationised fibre-based cellulose multi-layer membranes for sterile and high-flow bacteria retention and inactivationVanja Kokol, Monika Kos, Vera Vivod, Nina Gunde-Cimerman, 2023, original scientific article Abstract: Low-cost, readily available, or even disposable membranes in water purification or downstream biopharma processes are becoming attractive alternatives to expensive polymeric columns or filters. In this article, the potential of microfiltration membranes prepared from differently orientated viscose fibre slivers, infused with ultrafine quaternised (qCNF) and amino-hydrophobised (aCNF) cellulose nanofibrils, were investigated for capturing and deactivating the bacteria from water during vacuum filtration. The morphology and capturing mechanism of the single- and multi-layer structured membranes were evaluated using microscopic imaging and colloidal particles. They were assessed for antibacterial efficacy and the retention of selected bacterial species (Escherichia coli, Staphylococcus aureus, Micrococcus luteus), differing in the cell envelope structure, hydrodynamic biovolume (shape and size) and their clustering. The aCNF increased biocidal efficacy significantly when compared to qCNF-integrated membrane, although the latter retained bacteria equally effectively by a thicker multi-layer structured membrane. The retention of bacterial cells occurred through electrostatic and hydrophobic interactions, as well as via interfibrous pore diffusion, depending on their physicochemical properties. For all bacterial strains, the highest retention (up to 100% or log 6 reduction) at >50 L/h∗bar∗m2 flow rate was achieved with a 4-layer gradient-structured membrane containing different aCNF content, thereby matching the performance of industrial polymeric filters used for removing bacteria.
Keywords: fibrous membrane, cationised cellulose nanofibrils, amino-hydrophobised cellulose nanofibrils, antibacterial activity, multi-layer structure, flux, bacteria retention
Published in DKUM: 28.03.2024; Views: 220; Downloads: 12
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9. Uravnavanje hidrofilnosti površine poli(akrilatov) : diplomsko delo univerzitetnega študijskega programa I. stopnjeKaja Pleša, 2022, undergraduate thesis Abstract: Cilj diplomskega dela je bil pripraviti poli(metakrilaten) material s kopolimerizacijo kombinacije hidrofilnih in hidrofobnih metakrilatov za povečanje hidrofilnost materiala na eni strani in izboljšanje selektivne vezave določenih beljakovinskih molekul na drugi strani. Polimerizirali smo monolite in membrane z različnim razmerjem metakrilatov butil metakrilat, 2-etilheksil metakrilat, lavril metakrilat in zamreževalom etilenglikol dimetakrilatom. Monomer BMA je najbolj hidrofilen LMA pa najmanj, zato smo povečevali delež BMA in zmanjševali delež LMA. Za polimerizacijo smo uporabili termični iniciator AIBN in foto iniciator I819. Ugotovili smo, da prisotnost kisika, uporaba iniciatorja in količina zamreževala vplivajo na potek proste radikalske polimerizacije. Dobljene produkte smo okarakterizirali z različnimi analiznimi metodami. S FT-IR spektroskopijo smo potrdili prisotnost glavnih funkcionalnih skupin v sintetiziranih polimerih. Z adsorpcijo/desorpcijo dušika z uporabo Brunauer-Emmett-Teller metode smo izmerili specifične površine materialov, ki so bile manjše od 0,5 m2/g. Z analizo za določanje stičnega kota pa smo ugotovili, da imajo pripravljeni materiali hidrofilno površino. Meritve stičnega kota so se zaradi spreminjanja molskega razmerja metakrilatov gibale med 57,8 ° in 92,7 °.
Keywords: polimeri, polimerizacija v masi, metakrilati, monoliti, polimerne membrane, stični kot
Published in DKUM: 29.08.2022; Views: 594; Downloads: 96
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