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1.
Synergistic effect of supercritical and ultrasound-assisted ginger (Zingiber officinale Roscoe) extracts
Taja 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 characterisation
Taja 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 flour
Maja 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 processing
Maja 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 constituents
Kaja 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 compounds
Taja 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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Isolation and characterization of nanocellulose from Polypodiophyta fern using chemo-mechanical method
Katja 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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10.
Optimization of the supercritical extraction and decarboxylation process of industrial hemp
Taja 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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