*4.4. Experimental Design*

### 4.4.1. Assays with *Mytilus* sp. Gills

Ex vivo bioassays using *Mytilus* sp. gills were performed to investigate DNA damage resulting from exposure to pigments under light or dark conditions. For the purpose, valves of live mussels (ca. 40 mm in length) were carefully separated to retain integrity of gills and the visceral mass swiftly removed. Assays began immediately after excision. One valve being used as a test sample (pigment extract, diluted in PBS as described above) and the other as its respective control (PBS only). For both crude extracts, proboscis (P) and skin (S), the test valve was exposed to 1 mL of D1 or D2 dilutions, while the control was treated with 1 mL of PBS. Test valves and controls were subjected to either ambient light (L), i.e., indirect daylight, or dark (D) conditions for 1 h and processed immediately afterwards. All assays were made in triplicate (*n* = 3).
