TOPOISOMERASE INHIBITIONS ACTIVITY OF SEVERAL INDONESIAN PLANTS OF APOCYNACEAE, SIMAROUBACEAE, MAGNOLIACEAE USING MECHANISM-BASED YEAST BIOASSAY AND ACTIVE COMPOUND ISOLATION OF SELECTED PLANT

• Main Author: Zuhrotun, Ade
• Format: Dissertations
• Language: Indonesia
• Subjects: Farmakologi dan terapeutik
• Online Access: https://digilib.itb.ac.id/gdl/view/32844
• Institution: Institut Teknologi Bandung

Cancers is characterized by unregulated growth and spread of cells through lymphatic system and bloodstream to other parts of the body. These cells growth will continue and damage vital organs or eventually death. In the world wide, cancer incidences increases from 12.7 million in 2008 to 14.1 million in 2012 and it was predicted to be 25 million over the next two decades. In Indonesia, cancer prevalence in 2013 was 1.4 ‰or 347,792 people. Combination of surgery, radiation therapy, and chemotherapy are the most chosen therapy for cancer. Variation of cancer type for each patient, diseases stadium, toxicity risks, side effects and drug resistances made cancer treatments often difficult. So these conditions become opportunities and motivated the research for new anticancer agents. Recently, 74 of 488 anticancer drugs that are released by National Cancer Institute (NCI) of USA derived from plants. Whereas in Indonesia 16 of 74 anticancer drugs that are currently used derived from plants. The NCI has reported Families of Special Interest (FOSI) or plant families that are active as anticancer or contain anticancer agents, such as Apocynaceae, Simaroubaceae and Magnoliaceae. In Indonesia, about 63 species of Apocynaceae, 11 species of Simaroubaceae and 18 species of Magnoliaceae are grown and distributed in many areas. Mechanism-based yeast bioassay is one of the in vitro assay that is used to search anticancer agents from natural products. This assay’s result had correlation to a mechanism of anticancer agents i.e topoisomerase enzyme inhibitor. Activity of this inhibitor of Apocynaceae, Simaroubaceae and Magnoliaceae has not been reported yet, so this research was aimed to screen some Indonesian plants of those families and further to isolate active compound of selected plant. Screening activity was tested on methanol bark extract of 15 species of Apocynaceae, 3 species of Simaroubaceae and 2 species of Magnoliaceae against S. cerevisiae mutant (DNA repair- or recombination deficient). The results showed that extract of Kibatalia arborea (Blume) G. Don. and Michelia champaca L. were active as topoisomerase I and II inhibitors. Extract of Plumeria alba L., Tabernaemontana macrocarpa Jack., Wrightia pubescens Blume., ii Picrasma javanica Blume., Picrodendron baccatum Krug. & Urb. Ex. Urb., and Quassia indica (L) Nooteboom were active as topoisomerase I inhibitors, while extract of Ochrosia citrodora Lauterb & K. Schum and Michelia alba DC. were active as topoisomerase II inhibitors. Based on the screening, M. champaca extract was selected as the best choice that showed as topoisomerase I and II inhibitors. It had toxic effect based on brine shrimp lethality bioassay (LC50<1000 ?g/mL) and was activity as antifungi. Those data showed that the plant had potential active as anticancer agent. So, the plant was selected for its active compound isolation. The extract of M. champaca was fractionated by liquid-liquid extraction (LLE). Based on activity test results of topoisomerase inhibitor, using mechanism-based yeast bioassay and bioautography TLC, showed that the target compound from the fraction was spot number 5 (Rf 0.68) with chloroform-methanol (9:1) as development system. The ethyl acetate fraction was further separated by combination of fast column chromatography, followed by classical column chromatography. At the end, purification process was done by recrystallization using chloroform. The yellow needles crystal with melting point of 271.5-272.6 ?C, was named as MCET51 isolate. Topoisomerase inhibitor activity of MCET51 then was also tested by mechanismbased yeast bioassay methods and enzymatic reaction to human topoisomerase I. The result showed that the isolate was active as topoisomerase I and II inhibitors. Characterization of MCET51 was done by UV-Vis, infrared spectrophotometric, mass spectrometric and nuclear magnetic resonance spectroscopy of proton ( 1 H), carbon ( 13 C), Heteronuclear Single Quantum Coherence (HSQC) and Heteronuclear multiple-bond correlation spectroscopy (HMBC) methods. Based on characterization data compared to literatures, it can be concluded that MCET51 was liriodenine (C17H9NO3). The activity of M. champaca extract and liriodenine as topoisomerase I and II inhibitors using mechanism-based yeast bioassay were the first time reported by this research. And the activity of liriodenine as topoisomerase I inhibitor using enzymatic reaction to human topoisomerase I was the first time reported by this research, whereas other research only reported liriodenin as topoisomerase II inhibitor by different methods. The results of this research was in line with NCI, that reported Apocynaceae, Simaroubaceae and Magnoliaceae as FOSI or plant families that contained anticancer agents. Isolated compound from selected plant Michelia champaca L., i.e liriodenine was proven as topoisomerase I and II inhibitors. These results can be used as scientific evidence for further research to develop Indonesian plant as herbal anticancer.