1. Effect of green food processing technology on the enzyme activity in spelt flourMaja Leitgeb, Željko Knez, Gordana Hojnik Podrepšek, 2022, izvirni znanstveni članek Opis: In this research, a new approach to enzyme inactivation in flour was presented by supercritical
technology, considered a sustainable technology with lower energy consumption compared
to other technologies that use ultra-high temperature processing. Total protein concentration and
the activity of enzymes -amylase, lipase, peroxidase, polyphenol oxidase, and protease were determined
in flour pre-treated with scCO2. During the study, it was observed that the activity of enzymes
such as lipase and polyphenol oxidase, was significantly reduced under certain conditions of scCO2
treatment, while the enzymes -amylase and protease show better stability. In particular, polyphenol
oxidase was effectively inactivated below the 60% of preserved activity at 200 bar and 3 h, whereas
-amylase under the same conditions retained its activity. Additionally, the moisture content of the
scCO2-treated spelt flour was reduced by 5%, and the fat content was reduced by 58%, while the
quality of scCO2-treated flour was maintained. In this regard, the sustainable scCO2 process could be
a valuable tool for controlling the enzymatic activity of spelt flour since the use of scCO2 technology
has a positive effect on the quality of flour, which was verified by the baking performance of spelt
flour with the baked spelt bread as an indicator of quality. Ključne besede: food treatment, spelt flour, proteins, supercritical carbon dioxide, enzyme inactivation, defatted flour, carbon dioxide solubility Objavljeno v DKUM: 17.04.2025; Ogledov: 0; Prenosov: 0
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2. Enzyme activity and physiochemical properties of flour after supercritical carbon dioxide processingMaja Leitgeb, Željko Knez, Gordana Hojnik Podrepšek, 2022, izvirni znanstveni članek Opis: The objectives of this study were to inactivate the enzymes α-amylase, lipase, protease,
and peroxidase in flour with supercritical carbon dioxide (scCO2), and to optimize the enzymatic
treatment conditions. Enzyme inactivation is important, due to the undesirability of certain flour
enzymes that cause adverse reactions during storage as unpleasant rancidity of flour, and, at the
same time, reduce the shelf life of flour. Therefore, crude enzymes and flour were initially exposed
to scCO2 to determine the effect on specific enzyme activity under appropriate conditions. The
activity of the unwanted enzymes lipase and peroxidase decreased under optimal process conditions
of scCO2 exposure, lipase by 30%, and peroxidase by 12%, respectively. It was discovered that the
inactivation of enzymes in wheat flour occurred, where, at the same time, this sustainable method
allows the regulation of enzyme activity in the baking process. Afterwards, the effect of scCO2 on the
physicochemical properties of flour, morphological changes on starch granules, and content of total
lipids was studied. In scCO2-treated white wheat flour, the fat content decreased by 46.15 ± 0.5%,
the grain structure was not damaged, and the bread as the final product had a lower specific surface
volume. Therefore, this could be a promising technology for flour pretreatment, potentially impacting
the prolonging of its shelf-life. Ključne besede: enzymes, enzyme activity, proteins, supercritical fluids, wheat flour Objavljeno v DKUM: 10.04.2025; Ogledov: 0; Prenosov: 1
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3. Supercritical fluid and conventional extractions of high value-added compounds from pomegranate peels waste: production, quantification and antimicrobial activity of bioactive constituentsKaja Kupnik, Maja Leitgeb, Mateja Primožič, Vesna Postružnik, Petra Kotnik, Nika Kučuk, Željko Knez, Maša Knez Marevci, 2022, izvirni znanstveni članek Opis: This study is focused on different extractions (Cold Maceration (CM), Ultrasonic Extraction
(UE), Soxhlet Extraction (SE) and Supercritical Fluid Extraction (SFE)) of bioactive compounds from
pomegranate (Punica Granatum L.) fruit peels using methanol, ethanol, and acetone as solvents in
conventional extractions and changing operating pressure (10, 15, 20, 25 MPa) in SFE, respectively.
The extraction yields, total phenols (TP) and proanthocyanidins (PAC) contents, and antioxidant
activity of different extracts are revealed. TP and PAC recovered by extracts ranged from 24.22 to
42.92 mg gallic acid equivalents (GAE)/g and 2.01 to 5.82 mg PAC/g, respectively. The antioxidant
activity of extracts ranged from 84.70% to 94.35%. The phenolic compound identification and
quantification in selective extracts was done using the LC-MS/MS method. The contents of different
flavonoids and phenolic acids have been determined. SFE extract, obtained at 20 MPa, contained
the highest content (11,561.84 µg/g) of analyzed total polyphenols, with predominant ellagic acid
(7492.53 µg/g). For the first time, Microbial Growth Inhibition Rates (MGIRs) were determined at
five different concentrations of pomegranate SFE extract against seven microorganisms. Minimal
Inhibitory Concentration (MIC90) was determined as 2.7 mg/mL of SFE pomegranate peel extract in
the case of five different Gram-negative and Gram-positive bacteria. Ključne besede: antimicrobial activity, antioxidants, bioactive compounds, extraction, LC-MS/MS, phenolics, phytochemistry, pomegranate, Punica Granatum L., secondary metabolites Objavljeno v DKUM: 24.03.2025; Ogledov: 0; Prenosov: 3
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4. Comparison of conventional and supercritical isolation methods from bee pollen and their effects on the concentrations of bioactive compoundsTaja Žitek Makoter, Vesna Postružnik, Anastasia Kyriakoudi, Elli Pantazi, Natasa Kalogiouri, Ioannis Mourtzinos, Željko Knez, Maša Knez Marevci, 2025, izvirni znanstveni članek Opis: The present work represents a profound investigation of material pretreatment, comparison of different isolation methods (conventional and supercritical), and characterization of phenolic compounds and amino acids from bee pollen. The determination of amino acids was carried out after developing a novel hydrophilic interaction chromatography combined with quadrupole time-of-flight tandem mass spectrometric method (HILIC-QTOF-MS/MS) with limits of detection over the range 1 mg/kg -5 mg/kg and recoveries over the range 87.6% - 97.1%. The optimal conditions for the isolation of amino acids are in the range of temperatures between 25°C to 55°C and at pressures between 300 bar and 350 bar. It can be concluded that the extraction temperature and the pressure affect antioxidant activity. The favored conditions are lower temperatures and higher pressures. Ključne besede: bee pollen, in vitro antioxidant activity, HILIC-LC-QTOF-MS.MS, amino acids, WM-266-4 cells Objavljeno v DKUM: 20.03.2025; Ogledov: 0; Prenosov: 2
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5. αP-x,y equilibrium data of the binary systems of 1-Butanol and 2-Butanol with argon at 313.15 K and 333.15 K – measurements and modelingDragana Borjan, Mariusz Nowak, Irena T. Žižović, Željko Knez, Maša Knez Marevci, 2024, izvirni znanstveni članek Ključne besede: high-pressure phase equilibrium Argon, 1-butanol, 2-butanol, Peng-Robinson equation of state, Boston-Mathias alpha function Objavljeno v DKUM: 19.03.2025; Ogledov: 0; Prenosov: 2
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6. Ketoprofen-loaded PLGA-based bioactive coating prepared by supercritical foaming on a TiAl6V4 substrate for local drug delivery in orthopedic applicationsKatja Andrina Kravanja, Klodian Xhanari, Maša Knez Marevci, Uroš Maver, Matjaž Finšgar, 2024, izvirni znanstveni članek Opis: In this study, a novel biodegradable poly(lactic-co-glycolic) acid coating containing a model anti-inflammatory drug (ketoprofen) was prepared on TiAl6V4 substrate for use in orthopedic medicine by a two-step process combining drop casting and supercritical CO2-assisted foaming. The prepared coating was first investigated by surface analysis techniques using time-of-flight secondary ion mass spectrometry and X-ray photoelectron spectroscopy. The combined results confirmed the loading of ketoprofen and its homogeneous spatial distribution in the coating. 3D profilometry revealed increased surface roughness of the coating compared to the bare TiAl6V4 substrate, which is favorable for cell adhesion. Furthermore, the electrochemical measurements (i.e., electrochemical impedance spectroscopy and potentiodynamic curve measurements) demonstrated that the coating application significantly mitigated corrosion compared to the bare TiAl6V4 substrate. In vitro drug release tests revealed extended drug release in simulated body fluids with zero-order release kinetics. Finally, the promising cell testing results using adipose-derived mesenchymal stem cells and osteoblasts confirmed the applicability of the coating for implants. Overall, the results of this study highlight the significant potential of the developed bioactive coating for future orthopedic applications. Ključne besede: TiAlV, bioactive coating, implants, PLGA, supercritical foaming, ketoprofen Objavljeno v DKUM: 14.03.2025; Ogledov: 0; Prenosov: 4
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7. Inovacijske tehnologije pridobivanja eteričnih olj in antioksidantov iz suhih listov rožmarina : magistrsko deloNina Kugl, 2025, magistrsko delo Opis: Namen magistrske naloge je bil pridobiti podatke o vplivu različnih tehnik obdelave suhih listov rožmarina na vsebnosti antioksidantov in na koncentracijo eteričnih olj (EO) v rožmarinu. Iskali smo metode, s katerimi lahko uspešno pridobivamo antioksidanta rožmarinsko kislino (RK) in karnozolno kislino (KK) ter EO iz mletih, suhih listov tunizijskega rožmarina. Preverili smo, kako uporabljene metode vplivajo na vsebnost antioksidantov in koncentracijo EO (γEO) v rožmarinu, tako da smo vsebnost antioksidantov in γEO v obdelanem rožmarinu primerjali z vsebnosti antioksidantov in γEO, pridobljenih iz rožmarina, ki ga nismo predhodno obdelali.
Primerjali smo tehnike mehanskega peletiranja, izpostavitve rožmarina v avtoklavu pri 70°C in različno visokih tlakih plinov N2 in CO2 ter mikrovalovno ekstrakcijo brez topil (SFME). Iz tako obdelanega rožmarina smo nato z hidrodestilacijo s Clevengerjevim aparatom destilirali EO, z acetonsko ekstrakcijo pa smo pridobili suhi praškast produkt z visoko vsebnostjo naravnih antioksidantov. Obdelane liste rožmarina in praškast ekstrakcijski produkt smo analizirali s tehnikama tekočinske kromatografije visoke ločljivosti z detektorjem UV-VIS in plinsko kromatografijo s plamensko ionizacijskim detektorjem.
Ugotovili smo, da smo s SFME najbolj znižali γEO v rožmarinu, kjer se po 10 min obdelavi koncentracija EO znižala za 80,52%. γEO v rožmarinu pa se je najmanj znižala pri mehanskem peletiranju. Najvišje vsebnosti antioksidantov RK in KK v rožmarinu smo določili v rožmarinu, ki je bil obdelan z mehanskim peletiranjem. V rožmarinu, obdelanem z mehanskim peletiranjem smo prav tako določili najvišje izkoristke ekstrakcije z acetonom. Ključne besede: rožmarin, antioksidanti, eterična olja, SFME, peletiranje, hidrodestilacija, avtoklaviranje Objavljeno v DKUM: 11.03.2025; Ogledov: 0; Prenosov: 7
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8. Izboljšanje stabilnosti fermentacijske brozge klavulanske kisline pred izolacijo z optimiranjem procesnih parametrov : magistrsko deloSimon Zadravec, 2025, magistrsko delo Opis: Bakterija Streptomyces clavuligerus med fermentacijskim procesom sintetizira klavulansko kislino (KK), ki učinkovito deluje kot inhibitor β-laktamaz in se uporablja v kombinaciji z antibiotiki za premagovanje bakterijske odpornosti. Nestabilnost KK med proizvodnim procesom predstavlja velik izziv, saj hitro razpada, kar zmanjšuje izkoristek in povečuje stroške proizvodnje. Namen te študije je izboljšati biosintezo KK z osredotočanjem na preučevanje vpliva procesnih parametrov na njeno stabilnost in produkcijo.
V magistrskem delu smo razvili kvantitativni spektrometrični model za merjenje koncentracij KK in glicerola. Razvit model omogoča hitro, učinkovito in robustno analizo teh dveh spojin med eksperimentalnimi poskusi, kar predstavlja izboljšanje v primerjavi s klasičnimi kromatografskimi tehnikami, saj pospeši pridobivanje rezultatov.
Model smo nato uporabili za spremljanje koncentracij KKin glicerola med eksperimentalnimi poskusi, v katerih smo spreminjali različne procesne parametre, kot so mešanje, zračenje, nadtlak, temperatura, redčenje ter kombinirali brozge iz različnih faz produkcijskega cikla S. clavuligerus. Poleg tega smo merili tudi koncentracijo raztopljenega kisika, viskoznost, pH in prisotnost nečistoče A, ki je razgradni produkt klavulanske kisline.
Rezultati so pokazali, da lahko pravilna nastavitev procesnih parametrov bistveno izboljša stabilnost in produktivnost klavulanske kisline. Povečanje zračenja in mešanja ter zvišanje tlaka je prineslo do 5 % izboljšanje izkoristka po 6 urah zadrževalnega časa ter 17 % izboljšanje izkoristka po 27 urah procesa, hkrati pa je koncentracija nečistoče A ostala stabilna, kar kaže na manjšo razgradnjo KK. Posamezno povečanje mešanja, zračenja ali tlaka ni dalo enako učinkovitih rezultatov, čeprav je prispevalo k zmanjšanju razgradnje KK od 11 % do 26 %. Ugotovitve prispevajo k optimizaciji industrijskega procesa proizvodnje KK, saj zmanjšujejo razgradnjo produkta in povečujejo izkoristek, kar vodi v bolj učinkovit proces in nižje stroške proizvodnje. Ključne besede: klavulanska kislina, fermentacijski proces, stabilnost, optimizacija procesnih parametrov, spektrometrija Objavljeno v DKUM: 28.02.2025; Ogledov: 0; Prenosov: 16
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9. Kromatografske metode za določevanje biooznačevalcev lipidne peroksidacije iz fizioloških vzorcev : magistrsko deloAmadeja Prša, 2025, magistrsko delo Opis: V magistrskem delu smo razvili in validirali kromatografske metode za določanje biomarkerjev lipidne peroksidacije v fizioloških vzorcih, s poudarkom na malondialdehidu (MDA), ki je ključni kazalec oksidativnega stresa. Uporabili smo tehnike plinske kromatografije z masno spektrometrijo (GC-MS) in tekočinske kromatografije z masno spektrometrijo (LC-MS/MS), ki omogočajo visoko občutljivost in selektivnost pri analizi kompleksnih bioloških vzorcev, kot je serum. Validacija metod je vključevala oceno linearnosti, ponovljivosti, točnosti in mej zaznave (LOD) za MDA in sorodne spojine. Poleg tega smo izvajali teste antioksidativne kapacitete, kot so DPPH, FRAP, CUPRAC, ORAC in TAC, da bi ocenili sposobnost seruma in drugih vzorcev pri nevtralizaciji prostih radikalov. Rezultati kažejo, da so razvite metode natančne, občutljive in primerne za analizo MDA v fizioloških vzorcih. Ugotovili smo, da koncentracije MDA korelirajo z intenzivnostjo oksidativnega stresa, kar potrjuje uporabnost teh biomarkerjev pri spremljanju patoloških stanj. Poleg tega smo pokazali, da ima serum pomembno antioksidativno kapaciteto, ki jo je mogoče učinkovito ovrednotiti z uporabo kombinacije različnih testov. Magistrsko delo prispeva k razvoju naprednih analitskih orodij za raziskave oksidativnega stresa, ki so ključnega pomena za razumevanje vpliva lipidne peroksidacije na zdravje in bolezni ter za razvoj novih terapevtskih strategij. Ključne besede: Oksidativni stres, lipidna peroksidacija, malondialdehid, testi antioksidativnosti, kromatografske metode (GC-MS, LC-MS/MS). Objavljeno v DKUM: 27.02.2025; Ogledov: 0; Prenosov: 20
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10. An overview of the current state of cell viability assessment methods using OECD classificationEneko Madorran, Miha Ambrož, Jure Knez, Monika Sobočan, 2025, pregledni znanstveni članek Opis: Over the past century, numerous methods for assessing cell viability have been
developed, and there are many different ways to categorize these methods accordingly. We
have chosen to use the Organisation for Economic Co-operation and Development (OECD)
classification due to its regulatory importance. The OECD categorizes these methods
into four groups: non-invasive cell structure damage, invasive cell structure damage, cell
growth, and cellular metabolism. Despite the variety of cell viability methods available,
they can all be categorized within these four groups, except for two novel methods based
on the cell membrane potential, which we added to the list. Each method operates on
different principles and has its own advantages and disadvantages, making it essential
for researchers to choose the method that best fits their experimental design. This review
aims to assist researchers in making this decision by describing these methods regarding
their potential use and providing direct references to the cell viability assessment methods.
Additionally, we use the OECD classification to facilitate potential regulatory use and to
highlight the need for adding a new category to their list. Ključne besede: cell viability, cell-based methods, in vitro toxicology, OECD cell viability classification Objavljeno v DKUM: 13.02.2025; Ogledov: 0; Prenosov: 6
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