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, original scientific article Abstract: 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. Keywords: ginger, supercritical fluid extraction, ultrasound-assisted extraction, antimicrobial activity Published in DKUM: 09.05.2025; Views: 0; Downloads: 0
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2. Optimisation of the green process of industrial hemp - preparation and its extract characterisationTaja Žitek Makoter, Petra Kotnik, Teo Makoter, Vesna Postružnik, Željko Knez, Maša Knez Marevci, 2022, original scientific article Abstract: Natural medicines and products are becoming increasingly important in the pharmaceutical
and food industries. The most important step in obtaining a natural remedy is the processing of
the natural material. This study offers the separation of the industrial hemp plant into fractions by
mechanical treatment, which has a significant impact on the selectivity of the obtained fractions. This
study also offers a solution to reduce waste by fractionating industrial hemp, focusing on the fraction
with the highest cannabinoid content (49.5% of CBD). The study confirmed the anticancer potential
of the extract, which prevents further division of WM-266-4 melanoma cells at a concentration of
10−3 mg/mL. However, application of the extract (c = 10−3 mg/mL) to normal human epidermal
melanocytes proved to be insignificant, as the metabolic activity of the cells was the same as in the
control cell group. Keywords: supercritical fluid extraction, ultrasonic extraction, melanoma cells, extraction, WM-266-4 Published in DKUM: 09.05.2025; Views: 0; Downloads: 1
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3. Effect of green food processing technology on the enzyme activity in spelt flourMaja Leitgeb, Željko Knez, Gordana Hojnik Podrepšek, 2022, original scientific article Abstract: 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. Keywords: food treatment, spelt flour, proteins, supercritical carbon dioxide, enzyme inactivation, defatted flour, carbon dioxide solubility Published in DKUM: 17.04.2025; Views: 0; Downloads: 1
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4. Enzyme activity and physiochemical properties of flour after supercritical carbon dioxide processingMaja Leitgeb, Željko Knez, Gordana Hojnik Podrepšek, 2022, original scientific article Abstract: 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. Keywords: enzymes, enzyme activity, proteins, supercritical fluids, wheat flour Published in DKUM: 10.04.2025; Views: 0; Downloads: 1
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5. 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, original scientific article Abstract: 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. Keywords: antimicrobial activity, antioxidants, bioactive compounds, extraction, LC-MS/MS, phenolics, phytochemistry, pomegranate, Punica Granatum L., secondary metabolites Published in DKUM: 24.03.2025; Views: 0; Downloads: 3
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6. 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, original scientific article Abstract: 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. Keywords: bee pollen, in vitro antioxidant activity, HILIC-LC-QTOF-MS.MS, amino acids, WM-266-4 cells Published in DKUM: 20.03.2025; Views: 0; Downloads: 2
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7. 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, original scientific article Keywords: high-pressure phase equilibrium Argon, 1-butanol, 2-butanol, Peng-Robinson equation of state, Boston-Mathias alpha function Published in DKUM: 19.03.2025; Views: 0; Downloads: 2
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8. Isolation and characterization of nanocellulose from Polypodiophyta fern using chemo-mechanical methodKatja Vasić, Monika Dokl, Željko Knez, Maja Leitgeb, 2024, original scientific article Abstract: 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. Keywords: nanocellulose, isolation, Polypodiophyta fern, chemo-mechanical method, biomaterial, biopolymers Published in DKUM: 31.01.2025; Views: 0; Downloads: 3
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9. Enhancing the properties and morphology of starch aerogels with nanocelluloseMilica Pantić, Mariusz Nowak, Gregor Lavrič, Željko Knez, Zoran Novak, Irena T. Žižović, 2024, original scientific article Keywords: starch aerogels, cellulose, supercritical impregnation, xanthohumol, usnic acid Published in DKUM: 15.01.2025; Views: 0; Downloads: 4
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10. Optimization of the supercritical extraction and decarboxylation process of industrial hempTaja Žitek Makoter, Petra Kotnik, Teo Makoter, Željko Knez, Maša Knez Marevci, 2025, original scientific article Abstract: Cannabinoids are the main active ingredients of Cannabis sativa L., obtained by a suitable extraction method and decarboxylation, in which the acid forms of the cannabinoids are converted into active forms. Supercritical CO2 extraction method and decarboxylation process were optimized for 7 cannabinoids (CBD, CBDA, THC, THCA, CBGA, CBN and CBC). The optimal extraction conditions for all cannabinoids were determined at a temperature of 60 °C and a pressure of 300 bar to 550 bar. At higher temperatures and reaction times, a significant depletion of neutral cannabinoids was observed. The optimal conditions of the decarboxylation process varied depending on the component. For the CBD component, which could be measured in the highest concentrations (560 mg/g), the optimal conditions were 140 °C and 10 min. The study can be an important model for predicting the behavior of cannabinoids under certain parameters. Keywords: supercritical fluids, extraction, cannabinoids, decarboxilation, Cannabis sativa I Published in DKUM: 08.01.2025; Views: 0; Downloads: 7
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