Separation Of The Antitumor Component From Nereis Virens And It’s Effect Of Anticancer

Since the 1980s, an upsurge of blue pharmaceutical ascendant in the global scope, many scientific research institutions at home and abroad in the field of marine drugs was carried out by the distinctive research. Due to the special marine environment, biological variety and large amount of creature, more and more compound with new structure and unique features have been discovered. In this work, marine annelid Nereis virens catched in the coast of Fujian province was used to separate anticancer components, and the mechanism of anticancer was further discussed.The powder of freeze-dried Nereis virens were extracted by 95% ethyl alcohol,and then extracted successively with organic solvent:petroleum ether, ethyl acetate and two kinds of extract A1, A2, were harvested. The anticancer activity was assayed against AGS, BGC, A375, HeLa and SW579 cancer cells, by MTT method. The extract A2 was found to have better antitumor activity. Then it was separated on a silicon column and the eluted fractions were collected(B1、B2...、B8). The component B4 were found to have obvious antitumor activity. Subsequently, the component B4 was separated by preparative HPLC and the eluted component C44 was tested to possess the high significant antitumor activity which was specific to A375 cells, use HPLC-MS to identify the structure of component C44 in order to investigate the structure-activity relationship.The cytotoxicity of component C44 to A375 cells was assayed by MTT method. MTT assay demonstrated that the component C44 had significant anticancer activity with IC50 values against A375 cells for 48 h was 52.67±4.26μg/mL. Fluorescence morphological observation of A375 cells stained by AO/EB was performed after treating with different concentration of component C44. The nuclei appeared chromatin condensation, nuclear fragmentation, cellmembrane vesicles and apoptotic bodies, which were typical characteristics of apoptosis. Flow cytometry indicated that the cell cycle was blocked at the stage of G0/G1 and (or) S. Also the Sub-G0/G1 phase was accumulated with the increasing of the drug concentration. The analysis of flow cytometry by Annexin V/PI staining showed that the apoptosis ratio was related to treating drug concentration. After treatment with different concentration of component C44, total caspase 3 activity in vitro of A375 was up-regulated remarkably.This work premilinarily studied the effect of C44 to A375 cell differentiation. The melanin content and total activity of tyrosinase of A375 cells were assessed after the treatment by different concentration of component C44. The experimental data showed that the component C44 could promote A375 cells to biosynthetize melanin and to up-regulate it’s total tyrosinase activity, which illustrated that component C44 could promote A375 differentiation to normal morphology. The antitumor mechanism of component C44 to A375 mainly induces cell differentiation.This work regard mice as experimental object to test the effect of alcohol extract and water extract for fatigue resistance, immune regulation, anti-inflammatory and anti-asthmatic effect, providing the basis for further research of biological active component.