2.4. Laboratory experiment for determining the isotopic fractionation
In order to evaluate the food sources for A. japonicus, the isotopic fractionation
between the diet and animal tissue was determined by a laboratory
feeding experiment. The experimental diet was prepared by
mixing LIVIC-BW with fine sediments (b125 μm in diameter), which
were collected from an intertidal flat in Gokasho Bay and combusted in
amuffle furnace at 400 °C for 2 h, at a ratio of 1:4 by weight. The experiment
started on 9 August. Twenty juveniles (range of wet weight=31
to 40 mg, mean±SD=36±3 mg) were cultured in an aquarium filled
with 7 L of seawater (salinity=~34) for 83 days. Sand-filtered seawater
was supplied at a rate of 500 ml min−1.Water temperature ranged from
21 °C to 26 °C. The culture water was aerated using an electric air pump.
Fecal matter and unconsumed feed were cleaned before feeding. Ten
grams of the diet was added to the aquarium once a day. During 2 h
after feeding, water supply was stopped. At the end of the experiment,
the sea cucumbers were kept overnight in filtered seawater to allow