Secondary Metabolites from Fungal Endophytes of Echinacea purpurea Suppress Cytokine Secretion by Macrophage-Type Cells

Abstract

Botanical extracts of Echinacea purpurea have been widely used for the treatment of upper respiratory infections. We sought to chemically examine fungal endophytes inhabiting E. purpurea, and to identify compounds produced by these endophytes with in vitro cytokine-suppressive activity. Twelve isolates from surface sterilized seeds of E. purpurea were subjected to fractionation and major components were isolated. Sixteen secondary metabolites belonging to different structural classes were identified from these isolates based on NMR and mass spectrometry data. The compounds were tested for their influence on cytokine secretion by murine macrophage-type cells. Alternariol (1), O-prenylporriolide (4), porritoxin (10) β-zearalenol (13), and (S)-zearalenone (14) inhibited production of TNF-α from RAW 264.7 macrophages stimulated with LPS in the absence of any significant cytotoxicity. This is the first report of a cytokine-suppressive effect for 4. The results of this study are particularly interesting given that they show the presence of compounds with cytokine-suppressive activity in endophytes from a botanical used to treat inflammation. Future investigations into the role of fungal endophytes in the biological activity of E. purpurea dietary supplements may be warranted.

Keywords: Asteraceae; Echinacea purpurea; TNF-α; fungal endophytes; inflammation; seeds.

Figure 1

Secondary metabolites isolated from fungal endophytes of Echinacea purpurea.

Figure 2

Influence of fungal metabolites on RAW 264.7 viability and secretion of TNF-α. Cells were treated with fungal compounds (numbered via Table 2) at 2.5 and 25 µg/mL alone (A) or in the presence of 10 ng/mL LPS (B). Cytotoxicity was measured using a rhodamine 123 assay (A). In B, culture supernatants were collected and assayed for TNF-α levels using ELISA. Values shown are means of two independent experiments, each of which included duplicate biological replicates (total of four biological replicates). Positive controls (+) include 100 µg/mL cycloheximide for cytotoxicity (A) [31] and dodeca-2E,4E-dienoic acid isobutylamide for inhibition of TNF-α production [30] (B). Data was analyzed using a one way ANOVA with Tukey’s post hoc test; *=p<0.05, **=p<0.01, and ***=p<0.001. Compound 2 was not soluble at 25 µg/mL and was tested at 1.25 and 12.5 µg/mL.