1. Synergistic effect of supercritical and ultrasound-assisted ginger (Zingiber officinale Roscoe) extractsTaja Žitek Makoter, Nika Kučuk, Vesna Postružnik, Maja Leitgeb, Željko Knez, Mateja Primožič, Maša Knez Marevci, 2022, izvirni znanstveni članek Opis: Proper processing of natural material is crucial to obtain an extract with high content of
biologically active components. Dried, grinded ginger roots were extracted by ultrasonic method
and supercritical extraction with CO2
. The aim of the study was to determine if a mixture of the two
types of extracts attained by different methods and solvents exhibits better bioavailability than each
extract itself. Therefore, both extracts were analytically evaluated and then mixed in a ratio of 1:1.
The supercritical extract (SCG extract) and the mixed extract (mixG extract) had high antioxidant
activity (78% and 73%) and total phenols (827 mg/g ext. and 1455 mg/g ext.), which is also consistent
with the levels of gingerol (303 mg/g ext. and 271 g/g ext.) and shogaol (111 mg/g ext. and 100 g/g
ext.) in the extracts. In comparison to both pure extracts higher levels of total phenols were found
in the extract mixG. This could be the reason for the significant inhibition of melanoma cells and
antimicrobial potential (against Staphylococcus aureus, Escherichia coli, and Candida albicans). The
combination of the extracts resulted in a significant increase in the inhibition of selected microbial
and melanoma cells WM-266-4 compared to the control. Cell viability decreased below 60% when
mixG extract was applied. Antimicrobial activity has been confirmed.
Ključne besede: ginger, supercritical fluid extraction, ultrasound-assisted extraction, antimicrobial activity
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2. Magnetne lastnosti magnetnih nanodelcev (mnps) z različnimi polimernimi prevlekamiKarmen Zorič, 2025, magistrsko delo Opis: Nanodelci imajo čedalje večji potencial v različnih industrijah zaradi svojih izjemnih fizikalnih in kemijskih lastnosti. Prilagajanje magnetnih lastnosti nanodelcev je ključno za oblikovanje novih večnamenskih materialov.
V magistrskem delu smo proučevali vpliv različnih razmerij ionov Fe2+ in Fe3+ na strukturne in magnetne lastnosti nanodelcev. MNPs smo sintetizirali z metodo koprecipitacije. Izvedli smo tudi študijo vpliva različnih polimernih prevlek (aminosilan ali arabinogalaktan) na lastnosti modificiranih MNPs. Prevleka preprečuje aglomeracijo MNPs in stabilizira MNPs. MNPs, prevlečeni s polimernimi prevlekami, so po navadi tudi bolj biokompatibilni.
Sintetizirane delce smo okarakterizirali z analiznimi metodami, kot so FTIR, TGA in VSM.
Z analizami smo potrdili prisotnost polimernih prevlek na površini MNPs. Določili smo termične stabilnosti modificiranih MNPs. Prav tako so vsi modificirani MNPs izkazovali superparamagnetno naravo. MNPs z arabinogalaktansko prevleko v razmerju Fe²⁺ : Fe³⁺ = 2 : 1 celo izkazujejo protibakterijsko učinkovitost na E. coli in S. aureus.
Ključne besede: magnetni nanodelci, polimerne prevleke
Objavljeno v DKUM: 07.05.2025; Ogledov: 0; Prenosov: 8
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3. Izolacija encimov in proteinov iz celic črne kvasovke wallemia ichthyophaga z uporabo mehanskega postopkaMarko Kresnik, 2025, diplomsko delo Opis: V svetu s hitro naraščajočo populacijo, onesnaženostjo in z izčrpanjem številnih naravnih virov, encimska tehnologija ponuja velik potencial za številne industrijske aplikacije. Nekatera področja, kjer se uporablja encimska tehnologija, so prehrambena industrija (encimi se uporabljajo za izboljšanje proizvodnje hrane, na primer pri predelavi žit, proizvodnji sladkorja, sira, sokov..), farmacevtska industrija (encimi se uporabljajo v sintezi zdravil, proizvodnji antibiotikov in pripravi cepiv), tekstilna industrija, proizvodnja biogoriv, kozmetična industrija in še druge panoge. Mikrobni encimi lahko na ekonomičen in okolju prijazen način delujejo kot biokatalizatorji za izvajanje reakcij v bioprocesih. Vsak živ organizem vsebuje biološko aktivne snovi, ki jih je mogoče ekstrahirati. Mikrobne celice proizvajajo intracelularne in ekstracelularne encime. Za sprostitev intracelularnih komponent iz celice je potrebno celično steno mikrobov razbiti. Za to smo uporabili postopek mehanske homogenizacije. Postopek homogeniziranja se običajno izvaja s pomočjo homogenizacijskih naprav, ki lahko delujejo s hidravličnim tlakom, ultrazvokom, visokim tlakom ali drugimi mehanskimi silami. V našem primeru bomo uporabili rotor-stator homogenizator, kjer vzorec potuje skozi ozek prostor med vrtečim rotorjem in statorjem, kar povzroči razbijanje delcev in razlitje celic v celično suspenzijo. Namen te diplomske naloge je bil ugotoviti kako čas homogenizacije vpliva na preživelost črnih kvasovk W.Ichthyophaga v celični suspenziji, preostalo aktivnost encimov (amilaza, β- glukozidaza, proteaza, celulaza) in preostalo koncentracijo proteinov. Celično suspenzijo smo homogenizirali v času od 0 – 90 min pri konstantni temperaturi 35 ºC. Rezultati so pokazali, da je preživelost W.Ichthyophaga s podaljševanjem časa homogenizacije upadala, preostala koncentracija proteinov pa je naraščala. Zaznali smo povečanje preostale aktivnosti encimov α-amilaze, celulaze in proteaze. Na preostalo aktivnost β-glukozidaze pa je imela homogenizacija negativen vpliv, saj smo dokazali takojšen upad preostale aktivnosti.
Ključne besede: W.Ichthyophaga, črne glive, homogenizacija, encimi, proteini.
Objavljeno v DKUM: 06.05.2025; Ogledov: 0; Prenosov: 6
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4. 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: 1
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5. 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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6. 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
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7. Isolation and characterization of nanocellulose from Polypodiophyta fern using chemo-mechanical methodKatja Vasić, Monika Dokl, Željko Knez, Maja Leitgeb, 2024, izvirni znanstveni članek Opis: Nanocellulose is considered a promising and sustainable biomaterial, with excellent properties of biorenewability with improved mechanical properties. As a unique natural biopolymer, it has been applied to many different industries, where efficient and environmentally friendly productions are in demand. For the first time, ferns from the class Polypodiopsida were used for the isolation of cellulose fibers, which was performed using a chemo-mechanical method. As chemical treatment plays a crucial role in the isolation of nanocellulose, it affects the efficiency of the extraction process, as well as the properties of the resulting nanocellulose. Therefore, mechanical fibrillation was performed via grinding, while the chemical process consisted of three different treatments: alkali treatment, bleaching, and acid hydrolysis. In three different experiments, each treatment was separately prolonged to investigate the differing properties of isolated nanocellulose. Structural analysis and morphological analysis were investigated by SEM, EDS, FT-IR, and DLS. The thermal stability of cellulose fibers was investigated by TGA/DSC. The morphology of obtained nanocellulose was confirmed via SEM analysis for all samples, with particles ranging from 20 nm up to 600 nm, while the most consistent sizes were observed for NC3, ranging from 20 to 60 nm. FT-IR spectra showed prominent absorption peaks corresponding to cellulose, as well as the absence of absorption peaks, corresponding to lignin and hemicellulose. The EDS confirmed the elemental purity of nanocellulose, while TGA/DSC indicated higher thermal stability of nanocellulose, compared to untreated fern, which started to degrade earlier than nanocellulose. Such characteristics with unique properties make nanocellulose a versatile biomaterial for the industrial production of cellulosic materials.
Ključne besede: nanocellulose, isolation, Polypodiophyta fern, chemo-mechanical method, biomaterial, biopolymers
Objavljeno v DKUM: 31.01.2025; Ogledov: 0; Prenosov: 2
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8. Optimizacija procesnih parametrov za proizvodnjo sekundarnih metabolitov v šaržnem bioreaktorju : magistrsko deloMelani Dolgov, 2024, magistrsko delo Opis: Sekundarni metaboliti so naravne spojine, ki jih sintetizirajo mikroorganizmi, vključno z bakterijami, glivami, kvasovkami in rastlinami. Čeprav ti produkti niso nujno potrebni za osnovne biološke funkcije, kot so sinteza lipidov, aminokislin, ogljikovih hidratov in nukleinskih kislin, imajo pomembno vlogo v različnih industrijskih aplikacijah. Kvasovka Saccharomyces cerevisiae je ključni mikroorganizem v biotehnologiji zaradi svoje sposobnosti pretvorbe sladkorjev v etanol in ogljikov dioksid ter njene prilagodljivosti na različne okoljske pogoje.
Cilj magistrskega dela je bil optimizirati proizvodnjo sekundarnih metabolitov s pomočjo S. cerevisiae v šaržnem bioreaktorju. Raziskava je obsegala preučevanje vpliva različnih vrst mešal, začetnih koncentracij inokuluma ter hitrosti mešanja na rast celične mase in proizvodnjo sekundarnih metabolitov. Med optimizacijo smo spremljali spremembe pH med fermentacijo ter izvajali UV-VIS spektrofotometrične analize za pridobitev rezultatov o količini celokupnih proteinov, antioksidativnem potencialu in aktivnosti encima alkohol dehidrogenaze (ADH). Antioksidativni potencial proizvedenih sekundarnih metabolitov smo določili z DPPH metodo , za določanje koncentracije celokupnih proteinov je bila uporabljena Bradfordova metoda, aktivnost znotrajceličnega encima ADH pa je bila določena s pomočjo encimskega testa.
Optimalni parametri za doseganje najboljših rezultatov vključujejo hitrost mešanja 200 rpm, uporabo dvojnega mešala in začetno koncentracijo inokuluma 6,62 ∙ 107 CFU/mL. Ti parametri so bili potrjeni v zaključnem eksperimentu, kjer smo ponovno gojili S. cerevisiae pod enakimi pogoji, kar je potrdilo ponovljivost gojenja in učinkovitost izbranih parametrov. Prav tako so rezultati analitskih metod pokazali, da so permeabilizirani vzorci dosegli najvišjo antioksidativno aktivnost, najvišjo koncentracijo proteinov in največjo aktivnost ADH. Na podlagi teh ugotovitev smo zaključili, da višja hitrost mešanja, uporaba dvojnega mešala in večja začetna koncentracija inokuluma zagotavljajo optimalne rezultate v proizvodnji sekundarnih metabolitov.
Ključne besede: bioreaktor, procesni parametri, optimizacija, S. cerevisiae, alkohol dehidrogenaza, sekundarni metaboliti
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9. Imobilizacija l-asparaginaze na magnetne nanodelce : magistrsko deloMaja Verdev, 2024, magistrsko delo Opis: V okviru zaključnega dela smo izvedli optimizacijo imobilizacije l-asparaginaze (l-ASNaze) na aminoksilanske magnetne nanodelce (AMN-MNPs). Postopek imobilizacije sestoji iz dveh korakov, funkcionalizacije AMN-MNPs in imobilizacije encima. V prvem koraku smo funkcionalizirali AMN-MNPs, v drugem koraku smo nanje imobilizirali l-ASNazo.
Najprej smo v teoretičnem delu opisali lastnosti l-ASNaze, prikazali njeno strukturo, mehanizem delovanja in uporabo v različnih panogah. V nadaljnje smo opisali imobilizacijo na magnetne nanodelce (MNP), pri čemer smo se osredotočili na AMN-MNPs. Podan je tudi pregled literature o imobilizaciji l-ASNaze na MNPs.
Pri eksperimentalnem delu smo proučevali vpliv različnih parametrov, kot so koncentracija mrežnega povezovalca glutaraldehida (GA), vrtilna hitrost, čas in temperatura imobilizacije, z namenom doseganje čim višje aktivnosti imobiliziranega encima in učinkovitost imobilizacije. Zanimalo nas je kakšno aktivnost encima dosežemo ob dodatku ogrodnih proteinov, ki delujejo kot stabilizatorji. Uporabili smo goveji serumski albumin (BSA) in albumin iz jajčnih beljakov (EA). V nadaljnje smo proučili termično stabilnost proste in imobilizirane l-ASNaze pri različnih časih inkubacije in pri različnih temperaturah. Prosto in imobilizirano l-ASNazo smo vzpostavili na magnetno polje in primerjali aktivnosti l-ASNaze pred in po vzpostavitvi.
Ključne besede: l-asparaginaza, magnetni nanodelci, stabilnost, kovalentna vezava, funkcionalizacija, imobilizacija
Objavljeno v DKUM: 02.10.2024; Ogledov: 0; Prenosov: 32
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10. Funkcionalizirana metalo-organska ogrodja za razgradnjo penicilina : magistrsko deloLucija Dolinšek, 2024, magistrsko delo Opis: Onesnaževanje zaradi zlorabe antibiotikov resno ogroža okolje in zdravje ljudi, zato je nujno potreben razvoj učinkovitih strategij za razgradnjo in odstranjevanje ostankov antibiotikov. V magistrskem delu smo imobilizirali encim v metalo-organsko ogrodje (MOFs). V zeolitno imidazolatno ogrodje-8 (ZIF-8) smo z metodo samosestavljanja imobilizirali β-laktamazo (β-laktamaza@ZIF-8), katalitično učinkovitost imobiliziranih poroznih materialov (β-laktamaze@ZIF-8) za razgradnjo penicilinov (PEN) pa smo določevali z uporabo visoko zmogljivostne tekočinske kromatografije HPLC. Morfologijo in kemijsko strukturo β-laktamaze@ZIF-8 smo opredelili z vrstično elektronsko mikroskopijo (SEM). Rezultati so pokazali, da se je katalitična aktivnost enkapsuliranega encima v primerjavi s prostim encimom znatno povečala. Ugotovili smo tudi, da je ZIF-8 zaradi svoje velike površine in adsorpcijskih lastnosti že sam po sebi odlično orodje za odstranjevanje PEN iz vode. Katalitični mehanizem razgradnje PEN z β-laktamazo@ZIF-8 temelji na tvorbi kompleksa med ionom Zn (II) na ogrodju ZIF-8 in ciljno molekulo, kar oslabi vez štiričlenskega β-laktamskega obroča v molekuli PEN ter tako poveča učinkovitost encimske razgradnje PEN. Magistrsko delo je zagotovilo obetavno strategijo za razgradnjo antibiotika PEN v vodnem okolju.
Ključne besede: β-laktamaza, kovinsko-organska ogrodja (MOFs), ZIF-8, penicilin (PEN), imobilizacija, razgradnja
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