Application of deep eutectic solvents as a green alternative for extraction of active substances from milk thistle (silybum marianum)

Magistrsko delo prikazuje študijo priprave globoko evtektičnih topil kot nadomestnega medija škodljivim organskim topilom za ekstrakcijo aktivnih učinkovin iz pegastega badlja. V okviru magistrske naloge smo pripravili pet globoko evtektičnih topil različne sestave ter dodatno 80 % etanol kot konvencionalno organsko topilo in preverjali, katero od njih je najbolj ugodno za ekstrakcijo silimarinov iz pegastega badlja. Za sintezo globoko evtektičnih topil smo uporabili naslednje spojine: holin klorid, mentol, 1,4-butandiol, 1,2-propandiol, mlečno kislino in sečnino. Po HPLC analizi smo iz kromatogramov in enačbe umeritvene premice izračunali, kolikšna je bila koncentracija skupnih silimarinov, ki smo jih ekstrahirali iz posameznega vzorca. Vsa DES topila, razen DES2 (mentol : 1,4-butandiol v razmerju 1 : 2), so bila redčena z vodo, in sicer v razmerju 75 % DES-25 % H2O. Ugotovili smo, da se je največ silimarinov sprostilo v 80 % etanolu, ki je polarno protično topilo. Sledili sta globoko evtektični topili iz holin klorida in 1,2-propandiola ter holin klorida in 1,4-butandiola. Njima sta sledili globoko evtektični topili iz holin klorida in mlečne kisline ter holin klorida in uree (sečnine), najmanj silimarinov pa se je sprostilo v globoko evtektično topilo iz mentola in 1,4-butandiola, ki je bilo tudi edino topilo, ki ni vsebovalo vode, torej hidrofobno. Iz rezultatov je razvidno, da je prisotnost vode v topilu pozitivno vplivala na izkoristek ekstrakcije silimarinov iz pegastega badlja, vendar so bile koncentracije skupnih silimarinov, ekstrahiranih iz pegastega badlja, kljub temu izredno nizke, zaradi česar bo v prihodnje potrebnih še veliko raziskav, da bi to lahko izboljšali.This masters thesis presents a study of the preparation of deep eutectic solvents as a substitute medium for harmful organic solvents during the extraction of active substances from milk thistle. As part of this thesis, we prepared five deep eutectic solvents and an additional solvent of 80 % ethanol and studied which is more suitable for the extraction of silymarins from milk thistle. The following compounds were used for synthesis of the deep eutectic solvents: choline chloride, menthol, 1.4-butanediol, 1.2-propanediol, lactic acid and urea. After HPLC analysis, the concentration of total silymarins extracted from each sample was calculated using the chromatograms and the calibration line equation. All deep eutectic solvents except DES2 (menthol : 1.4-butanediol in a ratio of 1 : 2) were diluted with water in a ratio of 75 % DES to 25 % H2O. We determined that the 80 % ethanol, which is a polar protic solvent, released the most silymarins. This was followed by deep eutectic solvents made of choline chloride with 1.2-propanediol and then choline chloride with 1.4-butanediol. Those were followed by deep eutectic solvents combining choline chloride with lactic acid and choline chloride with urea. The fewest silymarins were released into the deep eutectic solvent made of menthol and 1.4-butanediol, which was also the only used hydrophobic solvent. The results show that the presence of water in the solvent had a positive effect on the efficiency of silymarin extraction from milk thistle. However, concentrations of total silymarins extracted from the milk thistle using all solvents remained extremely low. Further research will be required to improve upon these results