*Article* **Preliminary Examination of the Toxicity of Spalting Fungal Pigments: A Comparison between Extraction Methods**

**Badria H. Almurshidi 1, R.C. Van Court <sup>1</sup> , Sarath M. Vega Gutierrez <sup>1</sup> , Stacey Harper <sup>2</sup> , Bryan Harper <sup>2</sup> and Seri C. Robinson 1,\***


Stacey.Harper@oregonstate.edu (S.H.); Bryan.Harper@oregonstate.edu (B.H.)

**\*** Correspondence: seri.robinson@oregonstate.edu

**Abstract:** Spalting fungal pigments have shown potential in technologies ranging from green energy generation to natural colorants. However, their unknown toxicity has been a barrier to industrial adoption. In order to gain an understanding of the safety of the pigments, zebrafish embryos were exposed to multiple forms of liquid media and solvent-extracted pigments with concentrations of purified pigment ranging from 0 to 50 mM from *Chlorociboria aeruginosa, Chlorociboria aeruginascens,* and *Scytalidium cuboideum.* Purified xylindein from *Chlorociboria sp*. did not show toxicity at any tested concentration, while the red pigment dramada from *S. cuboideum* was only associated with significant toxicity above 23.2 uM. However, liquid cultures and pigment extracted into dichloromethane (DCM) showed toxicity, suggesting the co-production of bioactive secondary metabolites. Future research on purification and the bioavailability of the red dramada pigment will be important to identify appropriate use; however, purified forms of the blue-green pigment xylindein are likely safe for use across industries. This opens the door to the adoption of green technologies based on these pigments, with potential to replace synthetic colorants and less stable natural pigments.

**Keywords:** spalting; fungal pigment; xylindein; dramada; *Chlorociboria aeruginosa*; *Chlorociboria aeruginascens*; *Scytalidium cuboideum*; natural pigment; natural colorant
