THE CYTOTOXIC EFFECT OF VOLATILE COMPOUNDS ISOLATED FROM LUNARIA ANNUA SEEDS VARIOUS CELL LINES OF HUMAN CANCERS MEASURED BY THE MTT ASSAY

Cilj istraživanja: Cilj istraživanja je ispitati potencijalno citotoksično djelovanje hlapljivih spojeva izoliranih iz sjemena biljne vrste Lunaria annua na humane karcinomske stanice dojke (MDA-MB-231) te pluća (A549). Pretpostavka je da će se broj karcinomskih stanica, nakon izlaganja hlapljivim spojevima, smanjiti u odnosu na kontrolnu skupinu. Materijali i metode: Citotoksičnost se ispitivala MTT metodom kojom se određuje postotak metabolički aktivnih stanica nakon izlaganja Clevenger destilatu, mikrovalnom ekstraktu te ekstraktu biljne vrste Lunaria annua. Uzorci biljke su pripravljeni u pet različitih koncentracija (1 µg/mL, 5 µg/mL, 10 µg/mL, 50 µg/mL i 100 µg/mL), a učinak je gledan nakon 4, 24, 48 i 72 sata. Djelotvornost ekstrakata određivana je spektrofotometrijski mjerenjem apsorbancije pri 570 nm. Rezultati: Rezultati su prikazani grafički u odnosu vremena inkubacije i postotka metabolički aktivnih stanica. •Kod stanica karcinoma dojke, hlapljivi spojevi izolirani iz sjemena Lunaria annua (uzorci Clevenger destilat, mikrovalni ekstrakt i ekstrakt) ostvaruju bolji učinak pri višim koncentracijama, pri različitim vremenima inkubacije. Najznačajniji učinak na smanjenje metabolički aktivnih stanica imaju uzorci mikrovalni ekstrakt i ekstrakt pri koncentraciji od 100 µg/mL, nakon 72h inkubacije. •Kod stanica karcinoma pluća, hlapljivi spojevi izolirani iz sjemena Lunaria annua (uzorci mikrovalni ekstrakt i ekstrakt) također ostvaruju bolji učinak pri višim koncentracijama, pri različitim vremenima inkubacije. Kod uzorka Clevenger destilata dugoročno gledano svaka od koncentracija daje otprilike jednak (konstantan) učinak. Najznačajniji učinak na smanjenje metabolički aktivnih stanica imaju uzorci mikrovalni ekstrakt i ekstrakt pri koncentraciji od 100 µg/mL, nakon 72h inkubacije. Zaključak: In vitro izlaganje stanica karcinoma dojke (MDA-MB-231) i pluća (A549) hlapljivim spojevima izoliranih iz sjemena biljne vrste Lunaria annua dovodi do smanjenja preživljenja tih stanica. Ispitivani hlapljivi spojevi pokazuju citotoksični učinak ovisan o koncentraciji i vremenu inkubacije. Djelovanje izoliranih hlapljivih spojeva nije uvijek razmjerno povećanju koncentracije i vremenu inkubacije, te u pojedinim slučajevima dolazi do oporavka stanica. Citotoksični učinak hlapljivih spojeva izoliranih iz sjemena Lunaria annua, što je ujedno i hipoteza našeg ispitivanja, je potvrđen, a idući korak je potvrđivanje tih učinaka in vivo ispitivanjem na modelima karcinoma dojke i pluća kod životinja.The aim of the research: The aim of the research is to examine the potential cytotoxic effects of volatile compounds isolated from seeds of the plant species Lunaria annua on human carcinoma cells of breast (MDA-MB-231) and lungs (A549). The assumption is that after the exposure of carcinoma cells to volatile compounds their number will be reduced in comparison to the control group. Material and methods: The cytotoxicity was tested by the MTT method used to determine the percentage of metabolically active cells after exposure to Clevenger distillate, microwave extract, and Lunaria annua plant extract. Plant samples were prepared at five different concentrations (1 µg/mL, 5 µg/mL, 10 µg/mL, 50 µg/mL i 100 µg/mL), and the effect is seen after 4, 24, 48 and 72 hours. The efficacy of the extracts was observed spectrophotometrically after measurement of absorbance at 570 nm. Results: The results are graphically presented in relation to incubation times and the percentages of metabolically active cells. •In breast carcinoma cells, volatile compounds isolated from Lunaria annua seeds (Clevenger distillate, microwave extract and extract samples) have a better effect at higher concentrations, at different incubation times. The most significant effect on the reduction of metabolically active cells is achieved by microwave extract and extract samples at a concentration of 100 µg /mL after 72-hour incubation time. •In lung cancer cells, volatile compounds isolated from Lunaria annua seeds (microwave extract and extract samples) also have a better effect at higher concentrations, at different incubation times. In the Clevenger distillate sample, in the long run, each of the concentrations has approximately the same (constant) effect. The most significant effect on the reduction of metabolically active cells is achieved by the microwave extract and extract samples at the concentration of 100 µg/mL after 72-hour incubation time. Conclusion: In vitro exposure of human carcinoma cells of breast (MDA-MB-231) and lungs (A549) to volatile compounds isolated from seeds of the plant species Lunaria annua leads to reduced cell survival rate. Cytotoxic effects of volatile compounds depend on concentration and incubation time. Effect of isolated volatile compounds is not always proportional to the increase in concentration and incubation time and in some cases cell recovery occurs. The cytotoxic effect of volatile compounds isolated from Lunaria annua seeds is confirmed, which was the hypothesis of the research, and the next step is the confirmation of these effects by in vivo studies on the models of animal breast and lung cancers