Anti-inflammatory activity of edible brown alga Eisenia bicyclis and its constituents fucosterol and phlorotannins in LPS-stimulated RAW264.7 macrophages

Abstract

Although individual phlorotannins contained in the edible brown algae have been reported to possess strong anti-inflammatory activity, the responsible components of Eisenia bicyclis have yet to be fully studied. Thus, we evaluated their anti-inflammatory activity via inhibition against production of lipopolysaccharide (LPS)-induced nitric oxide (NO) and tert-butylhydroperoxide (t-BHP)-induced reactive oxygen species (ROS), along with suppression against expression of inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX-2), in RAW 264.7 cells. The anti-inflammatory activity potential of the methanolic extract and its fractions of E. bicyclis was in the order of dichloromethane>methanol>ethyl acetate>n-butanol. The strong anti-inflammatory dichloromethane fraction was further purified to yield fucosterol. From the ethyl acetate fraction, six known phlorotannins were isolated: phloroglucinol, eckol, dieckol, 7-phloroeckol, phlorofucofuroeckol A and dioxinodehydroeckol. We found that these compounds, at non-toxic concentrations, dose-dependently inhibited LPS-induced NO production. Fucosterol also inhibited t-BHP-induced ROS generation and suppressed the expression of iNOS and COX-2. These results indicate that E. bicyclis and its constituents exhibited anti-inflammatory activity which might attribute to inhibition of NO and ROS generation and suppression of the NF-κB pathway and can therefore be considered as a useful therapeutic and preventive approach to various inflammatory and oxidative stress-related diseases.

Keywords: 2-amino-5,6-dihydro-6-methyl-4H-1,3-thiazine hydrochloride; 2′,7′-dichlorodihydrofluorescein; 2′,7′-dichlorodihydrofluorescein diacetate; 2′,7′-dichlorofluorescein; 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide; 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid; AMT; Anti-inflammation; COX-2; DCF; DCFH; DCFH-DA; DMEM; DMSO-d(6); Dulbecco’s Modified Eagle’s Medium; Eisenia bicyclis; FBS; Fucosterol; HRP; LPS; MTT; NO; NOS; PVDF; Phlorotannin; ROS; TLC; Trolox; cyclooxygenase-2; deuterated dimethylsulfoxide; fetal bovine serum; horseradish peroxidase; iNOS; inducible nitric oxide synthase; lipopolysaccharide; nitric oxide; nitric oxide synthase; polyvinylidene fluoride; reactive oxygen species; t-BHP; tert-butylhydroperoxide; thin layer chromatography.