2.4 Data analysis
All statistical analyses were conducted in SPSS 17.0. Specifically, for the short-term depuration experiments, a paired t-test was used to compare the mean values of the food carbon AE measured under the different depuration times (24 h and 36 h), and an independent t-test was used to compare mean values among different depuration times in the same experiment or among experiments with different food rations at the same time. For the long-term depuration experiments, analysis of covariance (ANCOVA) was used to compare the release rates of fish body carbon under different daily food rations. Bivariate correlation with the Pearson correlation coefficient was used to examine the correlation of the proportions of DOC, CO2, and PC with depuration time in the long-term depuration experiments.
3 Results
3.1 Recovery of 14C released in seawater
The results showed that the 14C released from the food and fish body sources could be sufficiently recovered. For the food carbon,F14Crelease/F14C releasewere 105%, 108% and 112% at food rations of 2.2%, 3.2% and 5.7% of the fish DW, respectively. For the fish body carbon,B14Crelease/B14C releasewere 89% and 105% at the daily food rations of 2.2% and 4.3% of the fish DW, respectively.
3.2 Food carbon assimilation efficiency and release
The amount of ingested carbon retained in the marine medaka decreased quickly (0.07–0.15 h-1) during the first 4 h, and then the decrease slowed to lower rates (0.02–0.03 h-1) until the end of the depuration (Figure 3). During the depuration from 24 h to 36 h, there were no significant changes in the carbon retained in fish at any of the three food rations (t-test,p > 0.10) (Figure 3). The food carbon AE decreased with increasing food rations (Figure S1). Based on the experiments with different food rations, the carbon AEs in the fish were 30%–49% (38.9% on average) and 25%–47% (33.7% on average) corresponding to depuration times of 24 h and 36 h, respectively; the difference was not statistically significant (paired t-test, p > 0.05).
Figure 3 . Retention of ingested carbon in marine medaka during the 36-h depuration. Ingested food rations are expressed in percentages of fish dry weight (DW). Data are the mean ± SD (n = 3–5). The error bars represent the standard deviations. Note the natural logarithm scale in the left subfigure.
The proportions of carbon released as DOC, CO2, and PC varied during the depuration (Figure 4). During the first 2 h, most of the released carbon was DOC (55%–60%), and the proportion of DOC decreased during the first 16 h (to 39%–55%). In contrast, the proportion of CO2 increased (from 25%–32% to 40%–54%) during the first 16 h. The contribution of PC peaked during the first 4 h (up to 15%–25%); thereafter, it decreased quickly to the lowest values (to less than 8%) during the depuration from 8–16 h, and remained constant until the end of the depuration (Figure 4a, b, c).
The release rates of DOC, CO2, and PC from ingested food decreased with the depuration time. With the food ration of 2.2% fish DW, the release rates of DOC, CO2, and PC decreased from 0.61, 0.32, and 0.13 µg C/mg DW/h at the beginning to 0.03, 0.05 and < 0.01 µg C/mg DW/h during the depuration from 8–16 h, respectively. With the increase in food rations, all release rates of DOC, CO2, and PC were increased proportionally (Figure 4d, e, f). However, no significant differences in the release rates of PC among the different food rations were observed after 8 h of depuration (t-test, p > 0.1), and the release rates of PC at all three food rations remained constant (< 0.01 µg C/mg DW/h) after 16 h of depuration (Figure 4d, e, f) (t-test, p> 0.1).
Figure 4 . Relative contributions (a, b, c) and release rates (d, e, f ) of different forms of carbon at different stages of the 36-h depuration under different food rations. Data are the mean ± SD (n =3–5). The error bars represent the standard deviations. DOC, dissolved organic carbon; CO2, carbon dioxide; PC, particulate carbon. Ingested food rations are expressed in percentages of fish dry weight (DW)
During the whole depuration period, most of the unassimilated food carbon was released into seawater in the forms of DOC (48%–59%) and CO2 (30%–40%), and only 11%–13% of the released carbon was PC in fecal pellets (Figure 5a).
Taking the ingested food carbon as 100%, for the 36-h depuration, 25%–45% (34.8% on average), 18%–29% (22.9% on average), and 7%–8% (7.6% on average) of the food carbon were released as DOC, CO2, and PC, respectively (Figure 5b). For the 24-h depuration, the released DOC, CO2, and PC accounted for 26%–42% (32.7% on average), 18%–25% (20.9% on average), and 7%–8% (7.5% on average) of the food carbon, respectively.
A substantial proportion (46%–49%) of the released DOC was COC. The ratio did not vary significantly at different stages of the 36-h depuration, or with different food rations (Figure S2).