Fractionation of aqueous extract of ficus deltoidea var. Kunstleri leaves using solid phase extraction method for anticancer activity on du145 cell line

Abstract

Ficus deltoidea var. kunstleri or also known as 'Mas Cotek' belongs to the Moraceae family. It is widely used for postnatal treatment, anti-diabetic, anti-breast cancer, anti-nociceptive and anti-hypertension. In this study, the anticancer effects of flavonoid compounds from aqueous ethanolic fractions of F. deltoidea var. kunstleri leaves against prostate cancer cell lines, DU145 were investigated. The aqueous extract of F. deltoidea var. kunstleri was subjected for fractionation by solid phase extraction method by using various concentration of aqueous - ethanol ranging 20-99.8%. The fractions (fraction I-IV) were screened to obtain the most active fraction through total phenolic and flavonoid contents and MTT assays. The induction of apoptosis was evaluated using Annexin V-FITC/PI while the morphological changes of DU145 cells was assessed using acridine orange/propidium iodide (AO/PI) staining method. HPLC, FTIR-ATR and LC-MS/MS analysis were conducted for identification and confirmation of bioactive compounds. Fraction I which was fractionated using 20% of ethanol exhibited the highest polyphenol contents and highest inhibition against DU145 at 100 μg/mL (45.45%). Furthermore, cell viability of DU145 cells treated with crude extract of F. deltoidea var. kunstleri and its active fraction (I) were also compared using MTT assay. It was revealed that the crude and its active fraction I caused cell reduction through apoptotic machinery which was evidenced by the formation of apoptotic bodies, fragmentation, cell blebbing and cell shrinkage. These findings were confirmed by AO/PI staining method and Annexin V-FITC/PI. The presence of bioactive compounds of vitexin and isovitexin were identified by HPLC, FTIR-ATR and LC-MS/MS analysis. Positive correlations had been established between total phenolic and flavonoid content and anti prostate cancer activities. Therefore, these results suggest that flavonoids content of F. deltoidea var. kunstleri and its fraction I could potentially can be developed as an anti-cancer agent for prostate cancer.