The fish immune and digestive enzyme activities were determined
using commercial assay kits (Nanjing Jiancheng Institute, Nanjing,
China) according to the instructions of the manufacturer.
The total superoxide dismutase (SOD) activity in the serum was
measured according to the method of Beauchamp and Fridovich
(1971). One unit of SOD activity was determined by the amount of
superoxide dismutase used to inhibit the reduction of nitroblue tetra-
zolium at 50%. The level of glutathione peroxidase (GPX) activity was
measured using the method of Flohé and Günzler (1984).Theamount
of GPX needed to oxidize 1 μmol of NADPH per min was defined
as one unit. The activity of serum lysozyme (LSZ) was determined
following the turbidimetric method (Boman et al., 1974; Hultmark
et al., 1980), which involved the lysis of lysozyme-sensitive Gram-
positive bacteria by LSZ. The serum total protein concentration of fish,
whichwas determined using the biuret protein assay kit,was calculated
using bovine serum albumin as the protein standard and expressed as
mg mL−1
of fish serum.