SYNTHESIS OF FOLIC ACID CONJUGATED CHITOSAN NANOPARTICLES AS A CARRIER IN DRUG DELIVERY SYSTEM OF PROPOLIS COMPOUND AND ITS ACTIVITIES TOWARD MCF-7 CANCER CELL LINE

• Main Author: Khristiani Rahayu, Adelina
• Format: Theses
• Language: Indonesia
• Online Access: https://digilib.itb.ac.id/gdl/view/37076
• Institution: Institut Teknologi Bandung

Breast cancer is the leading cause of cancer deaths among women . As many as 2,081,200 women in the world are predicted to have breast cancer in 2018. The use of natural compounds such as propolis has been widely used and proven to have the ability as an anticancer compound. One of the problems that arise in delivering natural compounds which are given directly is that the compound is suspected not to reach the target cell maximally because it has a low bioavailability. Developments in the field of biotechnology provide an alternative delivery of natural compounds to be more effective, namely using nanoparticles as a drug delivery system. Chitosan as a polymer is often used as an ingredient in synthesis nanoparticles because it is biodegradable, non-toxic, and inexpensive. The high expression of folic acid receptors in breast cancer cells can be utilized in targeting nanoparticles, so the addition of folic acid ligands in nanoparticles is expected to increase the delivery of targeted nanoparticles to breast cancer cells. Therefore the aim of this study was to synthesize folic acid conjugated chitosan nanoparticles which encapsulated propolis, and examine their activity against the MCF-7 cancer cell line. Both folic acid conjugated chitosan nanoparticles containing propolis (NPKF- P) and not containing propolis (NP-KF-blank) were synthesized with ionic gelation method. The NP-KF-P and NP-KF blank that were successfully synthesized were characterize including diameter, PDI, potential zeta, nanoparticle morphology, and percentage of propolis encapsulation in chitosan nanoparticles. Cytotoxicity test of NP-KF-P, NP-KF and propolis on the MCF-7 cell line was carried out using the MTT method. The internalization of NP-KF-P by MCF-7 cell line was observed using confocal microscopy and TEM. In order to be observed on a confocal microscope, NP-KF-P was labeled with fluorescence Rhodamin B dye. Analysis of the relative expression of the Ki-67 gene at the mRNA level after the MCF-7 cell line was treated with NP-KF-blank (11 ?g/ml), NP-KF-propolis IC25 (3,7 ?g/ml), IC50 (11 ?g/ml) , IC75 (33 ?g/ml) and control was done by qPCR method. The diameter of the nanoparticles that were synthesized was 129 ± 3.4 nm (NP-KF-blank) and 153.9 ± 1.3 (NP-KF-P), with the efficiency of propolis encapsulation of 30.37 - 73.36%. The forms of NP-KF-blank and NP-KF-P are spherical and not-aggregated. Zeta potential of NP-KF-blank is 30.5 ± 1.04 mV and NP-KF-P is 29.7 ± 0.82 mV, show that NP-KF-P and NP-KF-blank were stable and not easy to aggregate. Cytotoxicity test showed that IC50 NP-KF-blank value was 51.4 ± 6.7 ?g/ml, NP-KF-P was 11.0 ± 1.1 ?g/ml, and free propolis was 22.2 ± 1.2 ?g/ml. This shows that the use of chitosan-folic acid nanoparticles as a carrier in delivery system of propolis can increase the cytotoxicity of propolis in compare to propolis it self. The results of the internalization test showed that NP-KF-P was successfully internalized by the MCF-7 cell started from the 3rd hour to 6th hour of treatment indicated by the presence of red color (from rhodamin-B) within the MCF-7 cell line. The results of internalization using TEM also showed that at the 6th hour, NP-KF-P had been internalized and was found in both the membrane and the MCF-7 cell line cytoplasm. Provision of free folic acid (0.05 and 0.25 mg/ml) before administration of NP-KF-P in the internalization test caused a decrease in targeting of nanoparticles, so that internalization of NP-KF-P by MCF-7 cell line decreased. These results indicated that folic acid as a ligand in NP-KF-P could increase the effectiveness of targeting nanoparticles. Thus, folic acid conjugated chitosan nanoparticles ware effectively used as carriers in the drug delivery system of propolis compounds, so as to increase the cytotoxicity response of the MCF-7 cancer cell line. The relative expression of the Ki-67 level mRNA gene in the MCF- 7 cell line after IC 25 NP-KF-propolis treatment was shown to decline, which was relatively expressed 0.77 lower than the control group. These Ki-67 relative expressions decreased and differed significantly after the MCF-7 cell line treated by NP-KF-P with the concentration of IC50 and IC75 to 0.55 time and 0.49 respectively, lower than control treatment. This shows that the relative expression of the Ki-67 level mRNA gene can be suppressed by administering NP-KF-P either at low doses (IC25) or IC50 to high doses (IC75). Based on this study, it could be concluded that NP-KF-P was successfully synthesized and able to increase targeting directed to the MCF-7 cell line, so that NP-KF-P could be internalized and increased the value of cytotoxicity by decreasing the relative expression of Ki-67 genes at the mRNA level. This explains that chitosan nanoparticles conjugated with folic acid are effectively used as carriers in the drug delivery system of propolis compounds.