*3.3. Analysis of Carotenoids*

The extraction and identification of carotenoids were carried out according to our routine methods [19]. Carotenoids were extracted from living or fresh animal specimens with acetone. The acetone extract was transferred to ether-hexane (1:1) layer after the addition of water. The total carotenoid contents were calculated 

employing an extinction coefficient of E1%cm = 2100 (astaxanthin) [20] for the starfish *A.* 

*<sup>p</sup>lanci* and E1%cm = 1350 (peridinin) [20] for *A. japonica*, *T. squamosa*, and *D. fragum* at <sup>Ώ</sup>

max. The ether-hexane solution was evaporated. The residue was subjected to HPLC on silica gel. Carotenoid compositions were estimated by the peak area of the HPLC on silica gel with 

acetone-hexane (3:7) monitored at 470 nm.

 Individual carotenoids were identified by UV-VIS (ether), FAB MS, and partial 1H NMR (500 MHz, CDCl3). 
