This study explored the effects of butylated hydroxytoluene (BHT) and ethoxyquin (EQ) and ethyl ether extracts, ethyl acetate extracts (EAE), acetone extracts, ethanol extracts and aqueous extracts of Ginkgo biloba leaves (EGbs) on lipid oxidation in a linoleic acid emulsion, fish flesh and fish feed and in hydroxyl radical (·OH)-treated carp erythrocytes. The linoleic acid, fish flesh and fish feed were incubated with BHT, EQ and EGbs at 45°C for 8 d, respectively, except for the control group. The lipid oxidation in the linoleic acid emulsion, fish flesh and fish feed was then measured by the ferric thiocyanate method or thiobarbituric acid method. The carp erythrocytes were treated with BHT, EQ or EGbs in the presence of 40 μmol/L FeSO4 and 20 μmol/L H2O2 at 37°C for 6 h, except for the control group. Oxidative stress and apoptosis parameters in carp erythrocytes were then evaluated by the commercial kit. The results showed that BHT, EQ and EGbs inhibited lipid oxidation in the linoleic acid emulsion, fish flesh and fish feed and ·OH-induced phosphatidylserine exposure and DNA fragmentation (the biomarkers of apoptosis) in carp erythrocytes. Furthermore, BHT, EQ and EGbs decreased the generation of reactive oxygen species (ROS), inhibited the oxidation of cellular components and restored the activities of enzymatic antioxidants in ·OH-treated carp erythrocytes. Of all examined EGbs, EAE showed the strongest effects. The effects of EAE on lipid oxidation in the linoleic acid emulsion and on superoxide anion and malonaldehyde levels, catalase activity and apoptosis in ·OH-treated carp erythrocytes were equivalent to or stronger than those of BHT. Moreover, these results indicated that the inhibition order of EGbs on the generation of ROS and oxidation of cellular components in fish erythrocytes approximately agreed with that for the food and feed materials tested above. And, the antioxidative and anti-apoptotic effects of EGbs were positively correlated with their flavonoid content. Taken together, these results revealed that the fish erythrocyte system can be used as an experimental model to evaluate lipid oxidation in food and feed ingredients. The EAE can be used as a potential natural antioxidant or apoptosis inhibitor. The inhibition effects of EGbs on lipid oxidation and apoptosis may be due to the presence of flavonoid compounds.

Erythrocytes play an essential role in transporting O2 and CO2 for respiration in fish. However, erythrocytes continuously suffer from reactive oxygen species (ROS) -induced oxidative stress and apoptosis. Thus, it is essential to expand our knowledge of how to protect erythrocytes against ROS-induced oxidative stress and apoptosis in fish. In this study, we explored the cytotoxicity and the effects of butylated hydroxyanisole (BHA), ethyl ether extracts, ethyl acetate extracts, acetone extracts (AE), ethanol extracts, and aqueous extracts of Astragalus membranaceus (EAm) on hydroxyl radical (•OH)-induced apoptosis in carp erythrocytes. The rat hepatocytes and carp erythrocytes were incubated with different concentrations of BHA or EAm(0.125 to 1 mg/mL). The toxicity in rat hepatocytes and carp erythrocytes was then measured using a 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay and a haemolysis assay, respectively. The carp erythrocytes were treated with BHA or EAm in the presence of 40 μmol/L FeSO4 and 20 μmol/L H2O2 at 37 °C, except for the control group. Oxidative stress and apoptosis parameters in the carp erythrocytes were then evaluated using the commercial kit. The results indicated that at high concentrations, BHA and EAm could induce toxicity in rat hepatocytes and fish erythrocytes. However, BHA was more toxic than EAm at the same concentrations. Moreover, the toxicity order of BHA and EAm in the fish erythrocytes approximately agreed with that for the rat hepatocytes. Butylated hydroxyanisole and EAm suppressed the •OH-induced phosphatidylserine exposure and DNA fragmentation (the biomarkers of apoptosis) by decreasing the generation of ROS, inhibiting the oxidation of cellular components, and restoring the activities of antioxidants in carp erythrocytes. Of all of the examined EAm, the AE showed the strongest effects. The effects of AE on superoxide anion, H2O2, met-haemoglobin and reduced glutathione levels, as well as glutathione reductase activity and apoptosis were equivalent to or stronger than those of BHA. These results revealed that the AE of Astragalus membranaceus could be used as a potential natural antioxidant or apoptosis inhibitor in fish erythrocytes.

In sustainable aquaculture systems, copper sulfate (CuSO₄) is widely applied as a disinfectant to control parasitic infections and algal growth. However, aquatic organisms may suffer from exposure to excessive concentrations of Cu. Elevated Cu concentrations could activate damage to the respiratory functions of aquatic animals. Thus, this study explored the effects exerted by ferulic acid (FA) on respiratory metabolism, oxidation-related lesions, and the apoptosis parameters of the gills and red blood cells in copper sulfate pentahydrate (CuSO₄·5H₂O)-treated carp (Cyprinus carpio var. Jian). When the 30-day feeding experiment was completed, the carp were exposed to 12.5 μM of Cu for 4 days. The results indicated that the Cu decreased the oxygen consumption and ammonia excretion rates in the carp, reduced the metabolic- and antioxidant-related enzyme activities and glutathione levels in the carp, and enhanced the caspase activities and reactive oxygen species and malondialdehyde levels in the gills of the carp. Moreover, in addition to the changes in the above parameters, the Cu decreased the cell numbers and hemoglobin concentrations and increased the phosphatidylserine exposure and cytochrome c levels in the red blood cells of the carp. These results demonstrate that Cu is capable of decreasing respiratory metabolism and increasing oxidation-related lesions and apoptosis inside the gills and red blood cells of the fish. However, dietary FA quenched the Cu-induced apoptosis and oxidative lesions by reversing the same biomarker indicators, thereby suppressing the Cu-induced decrease in respiratory metabolism. Thus, FA can be used as a suppressor of Cu stress in fish.

The study was to explore the effect of the extracts of Angelica sinensis (EAs) on lipid oxidation in fish feeds compared with ethoxyquin (EQ) and the effect of dietary EAs on growth performance of carp (Cyprinus carpio var. Jian). Firstly, fish feeds were respectively added with EQ, and ethyl ether extract, ethyl acetate extract (EAE), acetone extract, ethanol extract (EE) and aqueous extract (AQE) of Angelica sinensis, except for the control. The results showed that EAs and EQ inhibited lipid oxidation in fish feeds (P < 0.05). Of all of the examined EAs, EAE showed the strongest protective effects against lipid oxidation (P < 0.05). Moreover, EAE at high concentrations showed a stronger effect on lipid oxidation compared with EQ (P < 0.05). Then, 7 experimental diets respectively supplemented with 0.0, 1.0, 2.0, 3.0, 4.0, 5.0 and 6.0 g/kg of EAE were fed to the respective treatment groups for 30 d. Four replicates were performed for each treatment group; 20 carp (mean weight: 12.10 ± 0.13 g) were in each replicate. The results indicated that dietary EAE improved the growth performance in carp (P < 0.05). The appropriate concentration of EAE for carp growth was estimated to be 3.643 g/kg diet. Thus, EAE could be used as a natural antioxidant in feeds for Jian carp.