Vpliv s subkritično vodo pridobljenih izvlečkov iz bele jelke (Abies alba) na viabilnost in migracijo celic Caco-2 in HaCaT

Beguš Mihelič, Rok (author), Pečar Fonović, Urša (mentor), Schoss, Katja (comentor)

Bela jelka (Abies alba Mill.) je vednozeleni iglavec. Razširjena je na različnih območjih osrednje, južne ter vzhodne Evrope, kjer v ugodnih razmerah doseže starost tudi do 600 let. Izvlečki iz bele jelke vsebujejo različne polifenole, med katerimi so najbolj zastopani lignani. S svojim antioksidativnim in protivnetnim učinkom imajo ugoden vpliv pri različnih zdravstvenih težavah. Med najbolj raziskanimi učinki so vplivi na kožo, kardiovaskularni sistem, črevesno mikrobioto, hiperglikemijo in obrabo sklepnega hrustanca. Poznamo več klasičnih metod za pridobivanje izvlečkov iz zdravilnih rastlin, v ospredje pa prihajajo tudi modernejše tehnike, med katere štejemo tudi ekstrakcijo s subkritično vodo (ESV). Želeli smo preveriti, ali ima različna priprava izvlečkov iz bele jelke vpliv na vsebnost lignanov, viabilnost ter migracijo celic HaCaT in Caco-2. Devet izvlečkov pridobljenih z ESV, ki so se med seboj razlikovali po postopku priprave in izbiri dela rastline, smo liofilizirali, raztopili v vodi oziroma metanolu ter v njih s pomočjo metode HPLC določili koncentracijo lignanov. Rezultate smo primerjali z dvema industrijsko pridobljena izvlečkoma iz vej bele jelke s tržnim imenom Belinal®. Ugotovili smo, da način priprave izvlečka in izbira rastlinskega dela vplivata na koncentracijo lignanov v izvlečkih. Izvlečki, ki so bili pripravljeni iz vej in lubja ter postopkom ESV, so vsebovali višje koncentracije lignanov napram izvlečkom, ki so bili pridobljeni samo iz vej. Pripravili smo ustrezne raztopine izvlečkov in s pomočjo testa MTS določili viabilnost celic HaCaT in Caco-2. Ugotovili smo, da noben izvleček ni pomembno zmanjšal viabilnosti celic. S testom migracije smo ugotavljali, ali ima način priprave in sestava izvlečkov vpliv na zapiranje vrzeli na celicah HaCaT in Caco-2. Klasično pripravljen izvleček z vodo (lubje, 70 ␃) zraven industrijskega na obeh celičnih linijah pozitivno vpliva na zapiranje vrzeli napram kontroli. Skoraj vsi izvlečki, ki so bili pridobljeni s postopkom ESV, pa nimajo pomembnega vpliva na potek migracije celic. Izstopa le izvleček, pridobljen z ESV (veje in lubje; 100 ␃; 100 bar), ki zaradi svoje visoke polifenolne sestave opazno zavira migracijo celičnih linij.

• Language: Slovenian
• Keywords: Abies alba, Caco-2, ESV, HaCaT, lignani, migracija, MTS
• Work type: Master's thesis/paper
• Organization: FFA - Faculty of Pharmacy
• Year: 2023
• PID: 20.500.12556/RUL-149991
• Publication date in RUL: 13.09.2023

Secondary language

• Language: English
• Title: Effect of silver fir (Abies alba) extracts obtained with subcritical water on the viability and migration of Caco-2 and HaCaT cells

Abstract

The Silver fir (Abies alba Mill.) is an evergreen conifer. It is widespread in various areas of central, southern, and eastern Europe, where, under favorable conditions, it can live up to 600 years. Extracts from the Silver Fir contain various polyphenols, among which lignans are the most represented. With their antioxidant and anti-inflammatory effects, they have a beneficial impact on various health issues. The most researched effects include impacts on the skin, cardiovascular system, gut microbiota, hyperglycemia, and joint cartilage wear. Several traditional methods are known for obtaining extracts from medicinal plants, but modern techniques are also emerging, including subcritical water extraction (SWE). We wanted to check whether different preparations of Silver fir extracts affect the content of lignans, the viability, and migration of HaCaT and Caco-2 cells. Nine extracts obtained with SWE, which differed in preparation procedure and choice of plant part, were lyophilized, dissolved in water or methanol, and their lignan concentration was determined using the HPLC method. The results were compared with two industrially obtained extracts from Silver fir branches with the trade name Belinal®. We found that the method of extract preparation and the choice of plant part affect the concentration of lignans in the extracts. Extracts prepared from branches and bark using the SWE method contained higher concentrations of lignans compared to extracts obtained only from branches. We prepared appropriate extract solutions and determined the viability of HaCaT and Caco-2 cells using the MTS test. We found that none of the extracts significantly reduced cell viability. With the migration test, we determined whether the preparation method and composition of the extracts affect the closure of gaps in HaCaT and Caco-2 cells. The traditionally prepared extract with water (bark, 70 ␃) alongside the industrial one positively affects the closure of gaps compared to the control on both cell lines. Almost all extracts obtained with the SWE procedure do not have a significant impact on cell migration. Only the extract obtained with SWE (branches and bark; 100 ␃; 100 bar) stands out, which, due to its high polyphenolic composition, noticeably inhibits the migration of cell lines.

• Keywords: Abies alba, Caco-2, SWE, HaCaT, lignans, migration, MTS