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Extracts from Silybum marianum seeds have high industrial potential due to their application as phytopharmaceuticals and food. In order to optimize its production, conventional extraction (in Soxhlet apparatus using solvents ethanol and n-hexane) and supercritical fluid extraction (using CO2 at the pressure of 30 MPa and temperature of 40 °C with and without co-solvent ethanol, from oil-rich and defatted seeds) were compared. In addition, the effect of plant cultivation (years 2011 and 2019) and the effect of seeds and extract storage on the yield and quality of extracts were assessed. It was shown that unsaturated fatty acids (linoleic and oleic acids being most dominant) constituted 64–87% of obtained extracts. Content of α-tocopherol varied from 0.01 to 13 g/100 g, while total phenolic content was in the range from 13.2 to 104.2 gGAE/100 gextract. In vitro cytotoxic activity analysis confirmed that extracts obtained from defatted seeds showed activity against cancer cells. This study pointed out the important role of the cultivation year, selection of extraction technique and solvent, as well as storage on the extraction yield, chemical profile, and cytotoxic activity of extracts.

V doktorski dizertaciji smo raziskovali uporabo ogljikovega dioksida pri visokih tlakih za impregnacijo maščobo-topnih vitaminov. Za impregnacijo smo izbrali vitamina K3 in D3. Predvsem vitamin D3 je izjemno občutljiva substanca, zato zahteva izjemno pozornost. Za impregnacijo obeh vitaminov smo uporabili polisaharidne arogele. Aerogeli so porozni materiali z izjemnimi lastnostmi. Zaradi visokih poroznosti imajo velike specifične površine in nizke gostote. Če te lastnosti združimo še z biokomatibilnostjo ter biorazgradljivostjo polisaharidov, dobimo idealne materiale za farmacevtske aplikacije. Cilj doktorske dizertacije je bil impregnacija vitaminov v polisaharidne aerogele, da bi jih s tem dodatno zaščitili ter tako povišali njihovo stabilnost. Delo je razdeljeno v dva večja dela. V prvem delu smo uporabili superkritični ogljikov dioksid za impregnacijo vitamina K3 in vitamina D3. Vitamin K3 smo uporabili kot modelno substanco, naš glavni cilj pa je bil impregnacija vitamina D3. Raziskali smo termodinamske in kinetične lastnosti obeh vitaminov. Dobljene vrednosti smo razložili z uporabo že razvitih kinetičnih modelov. V prvem delu dizertacije smo določili tudi strukturne lastnosti vzorcev ter preverili sproščanje vitamina in vitro. V drugem delu doktorske dizertacije smo preiskovali impregnacijo vitamina D3 z ogljikovim dioksidom. Tokrat smo uporabili ogljikov dioksid pri različnih pogojih, od superkritičnega do tekočega ogljikovega dioksida. Preiskovali smo kinetično obnašanje pri različnih pogojih ter določili stopnjo degradacije vitamina glede na izbrane pogoje. Z različnimi karakterizacijskimi metodami smo določil vpliv tako superkritičnega kot tekočega ogljikovega dioksida na končne strukturne lastnosti aerogelov. V tem delu dizertacije smo prav tako izvedli sproščanje vitamina D3 in vitro, izvedli pa smo tudi stabilnostno študijo za potrditev povišane stabilnosti vitamina D3 ob impregnaciji v aerogele. This doctoral dissertation examines utilization of carbon dioxide as solvent for high pressure impregnation of fat-soluble vitamins, in particular vitamin K3 and vitamin D3. Chosen vitamins, especially vitamin D3, are handling challenging since they are highly sensitive substances. Polysaccharide alginate aerogels were used as vitamins’ carriers. Aerogels are airy materials with outstanding properties such as high porosity, high specific surface area and low density. Combined with the biocompatible and biodegradable polysaccharides, they fulfil pharmaceutical demands for ideal candidates. General idea of this dissertation was to achieve protection and prolonged stability of sensitive vitamins against environments by impregnating them into polysaccharide aerogels. The work is divided in main two parts. In the first part, carbon dioxide was used in its supercritical state. The impregnation experiments were conducted with both vitamin K3 and vitamin D3. Vitamin K3 had the role of a model substance while the main focus was on vitamin D3, as an active substance. The thermodynamic and kinetic behaviour of both vitamins was examined and fitted to appropriate models. Furthermore, detailed characterisation of the samples was applied and finally, drug release experiments were performed to examine possible application of developed products. In the second part, the study was extended from the area of supercritical carbon dioxide to the area of subcritical liquid carbon dioxide. Up now, this field was unexplored and new. The experiments were performed this time only with vitamin D3. The kinetic behaviour under various operating conditions was examined and associated with the degradation of vitamin in the system. Detailed influence of both supercritical and subcritical liquid carbon dioxide on the end structure of aerogels was investigated. Moreover, characterisation of the samples as well as drug release experiments were performed as part of the study. Finally, stability tests were carried out to investigate the protection of impregnated vitamin D3.

Results of numerous studies indicate that young people have improper eating habits. The aim of this study was to determine whether daily intake of vitamins, minerals and energy satisfies nutrition requirements for graduating classes of elementary and high school students. An epidemiological cross-sectional study was conducted on 84 students, including 42 students from graduating class of an elementary school and 42 students from graduating class of a high school. The students were asked to write the amount and composition of food consumed over a week, their age, body weight and height. 'USDA National Nutrient Database for Standard Reference, Release 25' was used to calculate daily intake of vitamins, minerals and energy. Comparison was made with respect to recommendations of Dietary Reference Intake of the Food and Nutrition Board of the Institute of Medicine, US National Academy of Science. The mean, standard deviation and percentiles of daily intake of certain nutrients were determined. For nutrients for which there was defined value of Estimated Average Requirement the results are presented as percentages of individuals with intakes less than the Estimated Average Requirement as estimates of the prevalence of inadequacy, whereas for nutrients for which only Adequate Intake was defined the results are presented as percentages of individuals with intakes at or greater than the Adeqate Intake. Assessment of adequacy of energy intake was done with respect to calculated Estimated Energy Requirement for each participant in the study. It was shown that 47.6% of elementary and 38.1% of high school students have adequate energy intake. Daily intake of students from both elementary and high school does not meet the dietary recommendations for magnesium, calcium, vitamin C and liposoluble vitamins A, D, E and K. The findings of this study emphasize the importance of proper nutrition. There is a need for increased intake of milk, dairy products, fresh fruits and vegetables among the elementary and high school students in order to satisfy daily nutritional requirements.