2.4. Determination of digestive enzyme activities
The activities of trypsin, lipase and amylase and protein concentration
in the liver and the intestine of grass carp were determined
by spectrophotometry using kits from Nanjing Jiancheng Bioengineering
Institute (Nanjing, China). The units of the digestive enzymes
were defined based on the manufacturer's recommendations. Briefly,
1 U of trypsin activity was expressed as the equivalent enzyme activity
required to generate an optical density (OD) change of 0.003 at
pH 8.0 and 37 °C. Amylase activity was calculated as the activity
required to hydrolyze 10 mg of starch in 30 min at 37 °C. One unit
of lipase activity was defined as the μmol of substrate hydrolyzed per
minute at 37 °C. Enzyme activities were expressed as specific activity
(U mg protein−1
). A subset of the fish (n = 26) was used for determination
of enzyme activities. Data are presented as mean ± standard
error (n = 3).
2.5. Statistical analysis
Statistical analysis was performed with SPSS13.0 software. Significant
differences were found using one-way analysis of variance
(ANOVA), followed by the post hoc test, after confirming for data
normality and homogeneity of variances. Differences were considered
to be significant if P b 0.05.