IN VITRO CYTOTOXIC ACTIVITY OF ETHYL ACETATE FRACTIONS OF MARINE SPONGE Calthropella sp. COLLECTED FROM KRAKAL BEACH GUNUNGKIDUL

FITRIA SUSILOWATI, Dr. Respati Tri Swasono, M.Phil.; Dr. Winarto Haryadi, M.Si.

2016 | Tesis | S2 Ilmu Kimia

Telah dilakukan penelitian tentang isolasi, uji sitotoksik in vitro, dan identifikasi struktur dari fraksi etil asetat spons Caltropella sp. asal Pantai Krakal Gunungkidul. Penelitian ini bertujuan untuk mengevaluasi aktivitas sitotoksik fraksi etil asetat hasil isolasi dari sponge Calthropella sp. asal Pantai Krakal Gunung Kidul. Isolasi dilakukan dengan metode ekstraksi menggunakan diklorometana:metanol (2:1, v/v) sebagai pelarut. Ekstrak diklorometana selanjutnya dilakukan partisi dalam pelarut etil asetat:air (3:2, v/v). Ekstrak etil asetat yang diperoleh dipisahkan dengan kromatografi kolom. Uji toksisitas ekstrak etil asetat dilakukan dengan metode Brine Shrimp Lethality Test (BSLT) dan uji sitotoksik in vitro dilakukan dengan metode MTT terhadap kultur sel kanker payudara (MCF-7), sel kanker paru-paru (H-460), dan sel kanker hati (HepG-2). Analisis senyawa dari fraksi aktif dilakukan dengan High Resolution Liquid Chromatography-Mass Spectrometry (HR LC-MS) untuk menentukan senyawa penyusunnya berdasarkan massa molekul. Hasil ekstraksi diperoleh ekstrak kasar etil asetat berwarna kuning kecoklatan sebesar 217,5 mg. Hasil pemisahan diperoleh 8 fraksi dan uji toksisitas ekstrak etil asetat menunjukkan nilai LC50 105,38 mikrogram/mL, memberikan indikasi toksik dan memiliki aktivitas farmakologi. Berdasarkan hasil uji sitotoksik in vitro dengan metode MTT, fraksi 6 menunjukkan aktivitas sitotoksik kuat terhadap sel MCF-7 (IC50 22,84 mikrogram/mL), tetapi sitotoksiknya melemah terhadap sel H-460 dan HepG-2 dengan nilai IC50 323,85 dan 335,46 mikrogram/mL berturut-turut. Fraksi 7 menunjukkan aktivitas sitotoksik kuat terhadap sel MCF-7 (IC50 1,92 mikrogram/mL) dan sitotoksik sedang terhadap sel H-460 dan HepG-2 dengan nilai IC50 165,13 dan 53,01 mikrogram/mL berturut-turut. Fraksi 6 dan 7 disimpulkan menunjukkan potensi sitotoksik terhadap sel MCF-7, dengan memiliki nilai IC50< 30 mikrogram/mL. Analisis LC-MS mengindikasi kuat bahwa senyawa hasil isolasi dari fraksi aktif 6 terdiri dari 4'-N-metil-5'-hidroksistaurosporin, 24-metileneergos-4-en-3-on, dan satu senyawa baru. Fraksi 7 diprediksi kuat mengandung bengamida Q, klafepiktin A, 4'-N-metil-5'-hidroksistaurosporin, asam biemnik, karteriofenon A, dan satu senyawa baru.

A research on isolation, in vitro cytotoxic assay, and structure identification of ethyl acetate fractions of sponge Calthropella sp. from Krakal Beach Gunungkidul had been carried out. This study is aimed to investigate the cytotoxic activity of the ethyl acetate fractions isolated from Calthropella sp. marine sponge collected from Krakal Beach Gunung Kidul. The extraction was done by using dichloromethane:methanol (2:1, v/v). The dichloromethane extract was partitioned in ethyl acetate:water (3:2, v/v). The interesting ethyl acetate extract was then separated by column chromatography. Toxicity test from ethyl acetate extract was carried out by Brine Shrimp Lethality Test (BSLT) method, while the in vitro cytotoxic assay was performed by MTT assay against breast adenocarcinoma cancer (MCF-7), lung cancer (H-460), and hepatocarcinoma liver cancer cell lines (HepG-2). The active fractions were then analyzed by High Resolution Liquid Chromatography-Mass Spectrometry (HR LC-MS). The ethyl acetate extract was obtained as yellowish brown (217.50 mg). This extract gave LC50 value 105.38 microgram/mL from BSLT, which is considered to be toxic and has pharmacological activity. In MTT in vitro cytotoxic assay, fraction 6 has strong activity against MCF-7 cell lines (IC50 22.84 microgram/mL), but showed weak activity against H-460 and HepG-2 cell lines IC50 value 323.85 and 335.46 microgram/mL respectively. Fraction 7 has strong cytotoxic activity against MCF-7 cell lines (IC50 1.92 microgram/mL), but showed moderate cytotoxicity against H-460 and HepG-2 cell lines with IC50 value 165.13 and 53.01 microgram/mL respectively. Fraction 6 and 7 showed promising cytotoxicity on MCF-7 cell lines with IC50< 30 microgram/mL. Based on LC-MS analyses, it is strongly predicted that active fraction 6 consist of 4'-Nmethyl-5'-hydroxystaurosporine, 24-methyleneergost-4-ene-3-one, and one new compound. Fraction 7 is predicted to consist bengamide Q, clavepictine A, 4'-Nmethyl-5'-hydroxystaurosporine, biemnic acid, carteriofenone A, and one new compound.

Kata Kunci : Calthropella sp., MTT assay, MCF-7, H-460, HepG-2