Effectiveness of Methanol Solvent Extraction for Red Macroalgae Acanthophora spicifera Antitumoric Activity

Abstract

Introduction: Red macroalgae Acanthophora spicifera is commonly found in most parts of Indonesia coastal areas. However, information regarding their potential pharmaceutical uses remains largely unexplored. This study evaluates the antiproliferative activity of Acanthophora spicifera collected from Indonesian extracted with different solvents of ethanol and methanol. Methods: Cytotoxicity and antiproliferative activity of A. spicifera crude ethanol extract (AS-EtOH) and A. spicifera crude methanol extract (AM-MetOH) were evaluated with Lactate dehidrogenase (LDH) and Trypan blue exclusion assay in Human cervical cancer (HeLa) cells for 72h. Concentration of extracts treated in HeLa cells were 6.25 to 200 µg/mL. Viability staining with fluorescence stain Hoechst33342/PI was conducted to investigate apoptotic activity. Potential apoptotic activity of treatments was confirmed by DNA fragmentation assay. Results: AS-MetOH demonstrated high cytotoxicity in HeLa cells with IC 50 of 127.3 ± 33.13 µg/mL. However, AS-EtOH IC 50 values could not be determined in this study. Antiproliferative activity was highly significant in AS-MetOH treated cells as cell density was suppressed to 20 x 10 4 cells/mL compared to AS-EtOH (73 x 10 4 cells/mL) and untreated cells (83 x 10 4 cells/mL). Finally, apoptotic activity could be determined in AS-MetOH treated cells with increase PI fluorescence emitting cells and DNA fragmentation. On the other hand, apoptotic activity was not observable in AS-EtOH treated cells based on fluorescence viability staining and DNA fragmentation assay. Conclusion: Current results show methanol solvent as an effective solvent to promote potential pharmaceutical properties of macroalgae A. spicifera. Further advanced studies in a compound level from A. spicifera methanol fraction would be reasonable for development of macroalgae based anticancer agent.

A Simple Method for In-Depth Proteome Analysis of Mammalian Cell Culture Conditioned Media Containing Fetal Bovine Serum

“…This is actually the case because we found the presence of approximately 1% of dead cells (measured by a trypan blue staining assay; about 2–3% of apoptotic cells measured by fluorescence flow cytometry) under the culture conditions of the HeLa cells in this study ( Supplementary Figure S1 ). Thus, we consider that the considerable number of cytoplasmic and nuclear proteins in the conditioned medium in our study as well as previous reports [ 7 , 8 , 9 ] might be released from dead cells although we did not experimentally confirm the origin of these proteins in this study. It is interesting to note that proteins with signal peptide, which are regarded as authentic secretory proteins through the classical secretory pathway, occupied only 42.3% and 32.9% of the total proteins detected in the conditioned medium before and after the albumin depletion in this study, respectively ( Figure 1 C and Supplementary Table S1 ).…”