Isolation and characterization of sesquiterpenes from Arecophila saccharicola YMJ96022401 with NO production inhibitory activity

Sesquiterpenes, arecoic acids A-F and arecolactone, were isolated from the ethyl acetate extracts of the fermented broth of Arecophila saccharicola YMJ96022401 along with two known analogues 1,7 alpha,10 alpha-trihydroxyeremophil-11(13)-en-12,8-olide and 1,10 alpha,13-trihydroxyeremophil-7(11)-en-12,8-olide. Their structures were elucidated on the basis of spectroscopic data analyses. The inhibitory effects of all of these compounds on nitric oxide (NO) production in lipopolysaccharide (LPS, 200 mu g/mL)-activated murine macrophage RAW264.7 cells were also evaluated. Among these compounds, 1,7 alpha,10 alpha-trihydroxyeremophil-11(13)-en-12,8-olide significantly inhibited NO production without any cytotoxicity, and its average maximum inhibition (E-max) at 100 p,M and median inhibitory concentration (IC50) were 85.7% +/- 0.8% and 16.5 +/- 1.0 mu M, respectively. Arecolactone was the most potent, with the Emax at 12.5 mu M and IC50 being 94.7% +/- 0.8% and 1.32 +/- 0.1 mu M, respectively, but displayed cytotoxicity at considerable higher concentrations than 25 mu M. Analyses of Western blotting indicated that arecolactone (0.8-12.5 p,M) inhibited induction of inducible NO synthase (iNOS) by LPS, which involved suppression of NE-kappa B activation and the phosphorylation of extracellular signal-regulated kinase (ERK), c-Jun NH(2)-terminal kinase (INK) and p38 mitogen-activated protein kinases (MAPKs) in activated RAW 264.7 cells. In addition, arecolactone concentration-dependently prevented the vascular hyporeactivity to phenylephrine induced by LPS (300 ng/mL) through iNOS pathway in isolated rat thoracic aortic rings. These results indicated that both of these naturally occurring iNOS inhibitors may provide a rationale for the potential anti-inflammatory effect of A. saccharicola YMJ96022401. (C) 2012 Elsevier Ltd. All rights reserved.