饲料中还原型谷胱甘肽、α-硫辛酸及二者分别与硒的交互作用对牙鲆（Paralichthys olivaceus）生长和抗氧化的影响
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摘要
本文以海水肉食性鱼类牙鲆(Paralichthys olivaceus, Temminck et Schlegel)作为研究对象,分别研究了还原型谷胱甘肽、α-硫辛酸及二者分别与硒的交互作用对牙鲆生长和抗氧化力的影响,主要研究结果如下：
实验一：利用单因素实验设计研究了在精制饲料中添加α-硫辛酸(LA)不同梯度(0、200、400、800、1600和3200 mg/Kg),投喂给牙鲆(P. olivaceus)幼鱼(初始体重9.39±0.17 g),养殖8周后,计算和测定牙鲆生长和存活、肝脏还原型谷胱甘肽(GSH)和丙二醛(MDA)含量、肝脏谷胱甘肽过氧化物酶(GPx)、谷胱甘肽S-转移酶(GST)、谷胱甘肽还原酶(GR)、超氧化物歧化酶(SOD)和总抗氧化力(T-AOC),来探讨饲料中LA添加量对牙鲆生长和抗氧化的影响。
实验结果表明：饲料中添加外源的LA对牙鲆存活率没有显著影响(P＞0.05)。牙鲆增重率(WGR)在饲料LA添加量为400和800 mg/Kg时达到最大值,显著高于其它各组(P＜0.05)。各处理间肝脏GSH积累量和SOD活性没有显著差异(P＞0.05)。各处理组肝脏GST和GR活性随着饲料中LA添加量含量升高而先降低后上升的趋势(P＜0.05),且当饲料中LA添加量为400和800 mg/Kg时GST活性显著低于其他各处理组(P＜0.05),LA添加量为800 mg/Kg处理组的GR活性显著高于除LA添加量为1600 mg/Kg的其它各处理组(P＜0.05)。各处理组肝脏中MDA的含量随着饲料中LA添加量的增加而显著下降(P＜0.05),且饲料中LA添加量为1600和3200 mg/Kg两组的肝脏MDA含量显著低于对照组(P＜0.05)。各处理组肝脏中GPx活性随着饲料中LA添加量的增加而呈现先升高后降低的趋势(P＜0.05),且饲料中LA添加量为200、400和800 mg/Kg三组的GPx活性显著高于对照组和LA饲料添加量为3200 mg/Kg两个处理组(P<0.05)。各处理组肝脏中T-AOC随着饲料中LA添加量的增加而呈现先升高后降低的趋势(P<0.05),且饲料中LA添加量为800 mg/Kg处理组的T-AOC显著高于对照组和LA饲料添加量为3200 mg/Kg两个处理组(P＜0.05)。根据WGR的实验数据,利用折线模型得到牙鲆幼鱼饲料中LA的适宜添加量为745.05 mg/Kg,过高或过低都会对生长造成负面影响。
实验二：利用单因素实验设计研究了在精制饲料中添加不同梯度(0、97.96、189.92、371.84、688.72及1063.56 mg/Kg)还原型谷胱甘肽(GSH),投喂给牙鲆(P. olivaceus)幼鱼(初始体重9.49±0.22 g),养殖8周后,计算和测定牙鲆生长和存活、肝脏GSH和MDA含量、肝脏GPx、GST、GR、SOD和T-AOC,以探讨饲料中GSH含量对牙鲆生长和抗氧化的影响。
实验结果表明：饲料中添加外源的GSH对牙鲆存活率没有显著影响(P＞0.05)。WGR在饲料GSH含量为371.84 mg/Kg时达到最大值,显著高于其它各组(P＜0.05)。饲料GSH添加组牙鲆肝脏中GSH的含量显著高于零添加组(P＜0.05)。各处理组肝脏中MDA的含量没有显著差异(P>0.05)。肝脏中GR的活力随饲料中GSH含量的升高呈现先降低后上升的趋势,并在饲料中GSH含量为371.84 mg/Kg时达到最低值。肝脏中SOD和GPx活力未受到饲料中GSH含量的显著影响(P＞0.05)。饲料中GSH含量为371.84 mg/Kg时,肝脏中T-AOC显著高于1063.56 mg/Kg组(P＜0.05),GST活力显著低于其它各处理组(P＜0.05)。根据WGR的实验数据,利用折线模型得到牙鲆幼鱼饲料中GSH的适宜含量为368.92 mg/Kg,过高或过低都会对生长造成负面影响。
实验三：利用3×3实验设计,研究了在精制饲料中联合添加不同梯度LA(0、800和3200 mg/Kg)和硒(0、1.07和40 mg/Kg),配制9种实验饲料,投喂牙鲆(P. olivaceus)幼鱼(初始体重9.42±0.16 g),养殖8周后,计算和测定牙鲆生长和存活、肝脏GSH和MDA含量、肝脏GPx、GST、GR、SOD和T-AOC,以探讨饲料中LA和硒的交互作用对牙鲆生长和抗氧化的影响。
实验结果表明：LA与硒二者均过量添加组(LA:3200 mg/Kg;硒：40 mg/Kg)牙鲆幼鱼的存活率显著低于其它各处理组(P＜0.05),其余各组间存活率没有显著差异(均高于96.43%)(P＞0.05)。WGR受饲料中LA和硒的交互作用影响显著(P＞0.05),当饲料中缺硒(0 mg/Kg)或正常添加硒(1.07 mg/Kg)时,LA的适量添加(800 mg/Kg)能够使WGR有上升的趋势,特别在硒添加1.07 mg/Kg组,LA添加800 mg/Kg能够显著提高WGR(P＜0.05)。然而当饲料中硒含量过高(40 mg/Kg)时,饲料中LA的加入反而使WGR显著下降(P＜0.05)。肝脏中GSH含量受饲料中LA和硒交互作用影响显著(P＜0.05),饲料中缺硒(0 mg/Kg)或正常添加硒(1.07 mg/Kg)时,LA添加组的肝脏GSH积累量显著低于不添加LA各组(P＜0.05),然而当饲料中硒含量过高(40 mg/Kg)时,LA的添加能够使肝脏GSH的积累量有上升趋势(P＞0.05)。肝脏中MDA含量受到饲料中LA和硒的交互作用影响显著(P＜0.05),饲料缺硒时LA的过量添加(3200 mg/Kg)组的肝脏MDA含量显著高于其它两组(P＜0.05),饲料中硒含量为1.07 mg/Kg时LA的添加能够显著降低肝脏MDA积累量(P＜0.05),饲料中硒过量添加(40 mg/Kg)时LA添加3200 mg/Kg组的肝脏MDA积累量显著高于其它两组(P＜0.05)。肝脏中GPx活性变化受到饲料中LA和硒的影响显著(P＜0.05),当饲料中LA添加800 mg/Kg、硒含量为1.07 mg/Kg时,肝脏GPx活性显著高于其它各组(P＜0.05),饲料中硒含量一定时LA在饲料中添加量为800 mg/Kg各组肝脏GPx活性显著高于LA不添加(0 mg/Kg)和过量添加(3200 mg/Kg)各组(P<0.05),饲料中LA添加量一定时硒在饲料中含量为1.07 mg/Kg各组肝脏GPx活性显著高于硒不添加(0 mg/Kg)和过量添加(40 mg/Kg)各组(P<0.05)。肝脏中GST活性变化受到饲料中LA和硒的影响显著(P<0.05),当饲料中硒含量一定时LA添加800 mg/Kg各组GST活性显著最高(P<0.05),LA添加3200 mg/Kg各组的GST活性显著最低(P<0.05),当饲料中LA添加量一定时硒正常添加(1.07 mg/Kg)和过量添加(40 mg/Kg)各组肝脏GST活性显著高于硒零添加(0 mg/Kg)各组(P<0.05)。饲料中LA和硒的添加对肝脏GR活性均有显著影响(P<0.05),饲料中硒含量为1.07 mg/Kg、LA添加量为800 mg/Kg组的GR活性显著高于其它各组(P<0.05)。饲料中硒和LA对肝脏中T-AOC影响显著(P＜0.05),硒含量为1.07 mg/Kg、LA添加量为800 mg/Kg组的T-AOC显著高于其它各组(P＜0.05)。饲料中LA添加量对肝脏SOD影响并不显著(P＞0.05),饲料中LA添加量一定时,硒含量为40 mg/Kg各组SOD活性显著高于硒含量为0 mg/Kg和1.07 mg/Kg各组(P<0.05)。
实验四：利用3×3实验设计,研究了在精制饲料中联合添加不同梯度GSH(0、371.84和1063.56 mg/Kg)和硒(0、1.07和40 mg/Kg),配制9种实验饲料,投喂给牙鲆(P. olivaceus)幼鱼(初始体重9.49±0.17 g),养殖8周后,计算和测定牙鲆生长和存活、肝脏GSH和MDA含量、肝脏GPx、GST, GR、SOD和T-AOC,以探讨饲料中LA和硒的交互作用对牙鲆生长和抗氧化的影响。
实验结果表明：饲料中硒和GSH的交互作用对牙鲆存活率没有显著影响(P＜0.05)。在饲料中硒含量0 mg/Kg组的WGR显著低于饲料硒含量为1.07和40 mg/Kg组(P＜0.05)。饲料中硒和GSH的含量对牙鲆肝脏GSH积累量影响显著(P＜0.05),当饲料中硒含量一定时,饲料GSH含量为371.84 mg/Kg各组的肝脏GSH积累量显著高于其它各组(P＜0.05),或饲料中GSH含量一定时,饲料硒含量为1.07 mg/Kg各组的肝脏GSH积累量显著高于其它各组(P＜0.05)。饲料中硒和GSH的交互作用对肝脏MDA含量和SOD活性没有显著影响(P＞0.05)。饲料中添加GSH能够显著提高肝脏GPx活性(P＜0.05)。饲料中硒和GSH的含量对肝脏GST、GR活性和T-AOC均有显著影响(P＜0.05)。饲料中缺硒时(0 mg/Kg)添加371.84 mg/Kg GSH组的肝脏GST活性和T-AOC显著高于GSH零添加组(0 mg/Kg)和过量添加组(1063.56 mg/Kg) (P<0.05),而GR活性显著低于其它两组。饲料中硒含量合适(1.07 mg/Kg)时添加371.84 mg/Kg GSH组的肝脏GST和GR活性显著低于GSH零添加组(0 mg/Kg)和过量添加组(1063.56mg/Kg) (P<0.05),而T-AOC显著高于其它两组(P＜0.05)。饲料中硒过量添加(40 mg/Kg)时,肝脏GST活性随着饲料中GSH含量的升高而下降,GR活性在GSH添加量为371,84-mg/Kg组的活性显著高于其他两组(P＜0.05)。
A series of experiments were conducted to determine the effects of dietary reduced glutathione (GSH), a-lipoic acid (LA) and the interaction between selenium (Se) and GSH, Se and LA on growth and antioxidation in Japanese Flounder (Paralichthys olivaceus), a marine carnivorous fish. The results of the present study were summarized as follows:
Experiment 1:A one-factorial experiment was conducted to determine the effects of gradients of diatary LA on growth and antioxidation in juvenile Japanese flounder (initial weight was 9.39±0.17 g). The juveniles were divided into six triplicate groups, and fed one of the six purified diets (containing 0,200,400,800,1600 or 3200 LA mg/Kg dry diet respectively) for 8 weeks in a flow-through system. A series of indexes were conducted, such as weight gain rate (WGR), survival rate, the concentration of hepatic GSH and malonaldehyde (MDA), the hepatic total antioxidant capacity (T-AOC) and the activities of hepatic glutathione peroxidase (GPx), glutathione S-transferase (GST), glutathione reductase (GR) and superoxide dismutase (SOD).
The results showed that there were no significant difference in the survival of Japanese Flounder among six dietary treatments(P>0.05). The average WGR reached the significantly highest value in the groups fed the diets with 400 or 800 mg LA, which were significantly higher than those of the other groups (P<0.05). No significant difference was found between groups in the concentrations of hepatic GSH and activities of SOD (P>0.05). The activities of hepatic GST and GR decreased then increased with the increasing of supplied dietary LA (P<0.05). The lowest values of GST were found in the treatments with 400 or 800 mg LA/Kg diet (P<0.05), and the GR value of the treatment with 800 mg LA/Kg diet was significantly higher than the values in other treatments, except for the treatment with 1600 mg LA/Kg diet (P<0.05). The concentrations of hepatic MDA were significantly decreased with rising content of dietary LA (P<0.05), and the concentrations of hepatic MDA with 1600 and 3200 mg LA/Kg diet were significantly lower than the control group with 0 mg/Kg dietary LA (P<0.05). The activities of hepatic GPx and T-AOC increased then decreased with the increasing of supplied dietary LA (P<0.05). The values of GPx in the three treatments with 200,400 and 800 mg LA/Kg diet were significantly higher than the two treatments with 0 arid 3200 mg/Kg dietary LA (P<0.05). The highest value of T-AOC was found in the treatment with 800 mg LA/Kg diet, which was significantly higher than the two treatments with 0 and 3200 mg LA/Kg diet (P<0.05). According to the average WGR data, a broken-line model was employed to calculate the optimal levels of dietary LA, which was 745.05 mg/Kg diet. Deficient or overdose of dietary LA could cause negative effects on the growth and antioxidative capacity of Paralichthys olivaceus.
Experiment 2:A one-factorial experiment was conducted to determine the effects of gradients of diatary GSH on growth and antioxidation in juvenile Japanese flounder (initial weight was 9.49±0.22 g). The juveniles were divided into six triplicate groups, and fed on one of the six purified diets (containing 0,97.96,189.92,371.84,688.72 or 1063.56 GSH mg/Kg dry diet respectively) for 8 weeks in a flow-through system. A series of indexes were conducted, such as WGR, survival rate, the concentration of hepatic GSH and MDA, the hepatic T-AOC and the activities of hepatic GPx, GST, GR and SOD.
The results showed that survival was not significantly affected by dietary GSH levels (P>0.05). The average WGR in the treatment with 371.84 mg/Kg dietary GSH was significantly higher than those in other treatments (P<0.05). The hepatic GSH concentration in dietary GSH added groups was significantly higher than that in dietary GSH deprived group (control group) (P<0.05). No significant differences in hepatic MDA concentration were found between GSH deprived and supplemented groups (P>0.05). The lowest value of hepatic GR activities was found in treatment with 371.84 mg/Kg dietary GSH (P<0.05). There were no significant differences in the activities of hepatic SOD and GPx among the treatments (P>0.05). In the group with 371.84 mg/Kg dietary GSH, the T-AOC was significantly higher than that in the group with 1063.56 mg/Kg dietary GSH, however, the GST activity was significantly lower than those in the other groups (P<0.05). Based on the WGR data in the present experiment, a broken-line model was employed to calculate the optimal levels of dietary GSH, which was 368.92 mg/Kg diet. Deficient or overdose of dietary GSH could cause negative effects on the growth and anti-oxidative capacity of Paralichthys olivaceus.
Experiment 3:A two-factorial experiment was conducted to determine the Se and LA interaction on growth and antioxidation in juvenile Japanese flounder (initial weight was 9.42±0.16 g). Totally nine purified diets were made to provide graded levels of Se (0,1.07,40 mg/Kg, supplied as Na2Se03) and LA (0,800,3200 mg/Kg). The juveniles were divided into nine triplicate groups, and fed on one of the nine purified diets respectively, for 8 weeks in a flow-through system. A series of indexes were conducted, such as WGR, survival rate, the concentration of hepatic GSH and MDA, the hepatic T-AOC and the activities of hepatic GPx, GST, GR and SOD.
The results showed that the value of survival in the treatment with both 3200 mg LA/Kg diet and 40 mg Se/Kg diet was significantly lower than other treatments (P<0.05), and there was no significant difference between other groups (all higher than 96.43) (P>0.05). WGR was significantly affected by the interaction between LA and Se (P<0.05). When the diets were provided 0 or 1.07 mg Se/Kg diet, WGR presented a trend to rise in the treatment with 800 mg LA/Kg diet, and WGR reached the highest value in the treatment with both 800 mg LA and 1.07 mg Se/Kg diet, which was significant higher than other groups (P<0.05). Oppositely, in the diets with high selenium level (40 mg/Kg), values of WGR in the LA provided treatments was significantly lower than the control group (0 mg/Kg LA). Hepatic GSH concentration was significantly affected by the interaction between dietary LA and selenium (P<0.05). When dietary Se was deprived or provided 1.07 mg/Kg, the values of hepatic GSH concentration in the treatments provided LA were significantly lower than LA deprived treatments (P<0.05). Nevertheless, between the treatments of high Se level (40 mg/Kg), hepatic GSH concentration of the treatments with LA provided (800 or 3200 mg/Kg) had a rising trend compared with control group (P>0.05). Hepatic MDA concentration was significantly affected by the interaction between dietary LA and Se (P<0.05). The value of hepatic MDA concentration in the treatment with 3200 mg LA/Kg diet was significantly higher than the other two treatments when dietary Se was deprived (P<0.05). The values of hepatic MDA concentration in the treatments provided LA (800 or 3200 mg/Kg) were significantly lower than the control group (0 mg LA/Kg diet) when dietary Se was 1.07 mg/Kg (P<0.05). In the treatments of high Se level (40 mg/Kg), the values of hepatic MDA concentration of the treatment with 3200 mg LA/Kg diet was significantly higher than the other two treatments (P<0.05). The activities of hepatic GPx and GST were significantly affected by the interaction between dietary LA and Se (P<0.05). The highest value of hepatic GPx activity was found in the treatment with both 800 mg LA and 1.07 mg Se/Kg diet, and this value was significantly higher than those in other treatments (P<0.05). The values of hepatic GPx and GST of the treatments with 800 mg LA/Kg diet was significantly higher than those of the treatments with 0 or 3200 mg LA/Kg diet when dietary Se level was invariable (P<0.05). Likewise, the values of hepatic GPx of the treatments with 1.07 mg Se/Kg diet was significantly higher than those of the treatments with 0 or 40 mg Se/Kg diet when dietary LA level was invariable (P<0.05). The lowest values of hepatic GST activity was found in the treatments with 3200 mg LA/Kg diet (P<0.05). The values of hepatic GST of the treatments with 1.07 or 40 mg Se/Kg diet were significantly higher than the control group (0 mg Se/Kg diet) when dietary LA level was invariable (P<0.05). Both T-AOC and the activities of hepatic GR were significantly affected by both dietary LA and Se (P<0.05). The highest values were found in the treatment with both 800 mg LA and 1.07 mg Se/Kg diet, and the two values were significantly higher than those of other treatments, separately (P<.05). No significant difference was found between the diets with grads of LA (P<0.05). SOD activities in the treatments with 40 mg Se/Kg diet were significantly higher than SOD in both the control groups (dietary selenium 0 mg/Kg) and dietary Se 1.07 mg/Kg groups when dietary LA level was invariable (P<0.05).
Experiment 4:A two-factorial experiment was conducted to determine Se and GSH interaction on growth and antioxidation in juvenile Japanese flounder (initial weight was 9.49±0.17 g). Totally nine purified diets were made to provide graded levels of Se (0,1.07,40mg/Kg, supplied as Na2Se03) and GSH (0,371.84,1063.56 mg/Kg). The juveniles were divided into nine triplicate groups, and fed on one of the nine purified diets respectively, for 8 weeks in a flow-through system. A series of indexes were conducted, such as WGR, survival rate, the concentration of hepatic GSH and MDA, the hepatic T-AOC and the activities of hepatic GPx, GST, GR and SOD.
The results showed that survival was not significantly affected by the interaction between Se and GSH (P>0.05). The values of WGR in the Se provided treatments (1.07 or 40 mg/Kg) were significantly higher than the Se deprived treatments (0 mg/Kg) (P<0.05). The hepatic GSH concentrations were significantly affected by the interaction between GSH and Se (P<0.05). The hepatic GSH concentrations of the treatments with 371.84 mg GSH/Kg diet were significantly higher than those in other treatments when dietary Se level was invariable (P<0.05). Likewise, the hepatic GSH concentrations of the treatments with 1.07 mg Se/Kg diet were significantly higher than those in other treatments when dietary GSH level was invariable (P<0.05). There was no significant difference in hepatic MDA concentrations among nine dietary treatments, and so was the SOD activities (P>0.05). Hepatic GPx activities were significantly affected by dietary GSH levels (P<0.05), moreover hepatic T-AOC and GST, GR activities were significantly affected by the interaction between dietary Se and GSH (P<.05). Both hepatic T-AOC and GST activity in the treatment with 371.84 mg GSH/Kg diet were significantly higher than those in the treatments dealed with 0 or 1063.56 mg GSH/Kg diet when dietary Se was deprived (P<0.05). When dietary Se was 1.07 mg/Kg, the hepatic GST and GR in the treatment with 371.84 mg/Kg was significantly lower compared with the other two treatments with 0 or 1063.56 mg GSH/Kg diet, but T-AOC in 371.84 mg GSH/Kg diet treatment was significantly higher than the other two treatments (P<0.05). When dietary Se was 40 mg/Kg, hepatic GST activity decreased with dietary GSH level increased, and GR activity in dietary GSH 371.84 mg/Kg group was significantly higher than the other two groups (P<0.05).
实验一：利用单因素实验设计研究了在精制饲料中添加α-硫辛酸(LA)不同梯度(0、200、400、800、1600和3200 mg/Kg),投喂给牙鲆(P. olivaceus)幼鱼(初始体重9.39±0.17 g),养殖8周后,计算和测定牙鲆生长和存活、肝脏还原型谷胱甘肽(GSH)和丙二醛(MDA)含量、肝脏谷胱甘肽过氧化物酶(GPx)、谷胱甘肽S-转移酶(GST)、谷胱甘肽还原酶(GR)、超氧化物歧化酶(SOD)和总抗氧化力(T-AOC),来探讨饲料中LA添加量对牙鲆生长和抗氧化的影响。
实验结果表明：饲料中添加外源的LA对牙鲆存活率没有显著影响(P＞0.05)。牙鲆增重率(WGR)在饲料LA添加量为400和800 mg/Kg时达到最大值,显著高于其它各组(P＜0.05)。各处理间肝脏GSH积累量和SOD活性没有显著差异(P＞0.05)。各处理组肝脏GST和GR活性随着饲料中LA添加量含量升高而先降低后上升的趋势(P＜0.05),且当饲料中LA添加量为400和800 mg/Kg时GST活性显著低于其他各处理组(P＜0.05),LA添加量为800 mg/Kg处理组的GR活性显著高于除LA添加量为1600 mg/Kg的其它各处理组(P＜0.05)。各处理组肝脏中MDA的含量随着饲料中LA添加量的增加而显著下降(P＜0.05),且饲料中LA添加量为1600和3200 mg/Kg两组的肝脏MDA含量显著低于对照组(P＜0.05)。各处理组肝脏中GPx活性随着饲料中LA添加量的增加而呈现先升高后降低的趋势(P＜0.05),且饲料中LA添加量为200、400和800 mg/Kg三组的GPx活性显著高于对照组和LA饲料添加量为3200 mg/Kg两个处理组(P<0.05)。各处理组肝脏中T-AOC随着饲料中LA添加量的增加而呈现先升高后降低的趋势(P<0.05),且饲料中LA添加量为800 mg/Kg处理组的T-AOC显著高于对照组和LA饲料添加量为3200 mg/Kg两个处理组(P＜0.05)。根据WGR的实验数据,利用折线模型得到牙鲆幼鱼饲料中LA的适宜添加量为745.05 mg/Kg,过高或过低都会对生长造成负面影响。
实验二：利用单因素实验设计研究了在精制饲料中添加不同梯度(0、97.96、189.92、371.84、688.72及1063.56 mg/Kg)还原型谷胱甘肽(GSH),投喂给牙鲆(P. olivaceus)幼鱼(初始体重9.49±0.22 g),养殖8周后,计算和测定牙鲆生长和存活、肝脏GSH和MDA含量、肝脏GPx、GST、GR、SOD和T-AOC,以探讨饲料中GSH含量对牙鲆生长和抗氧化的影响。
实验结果表明：饲料中添加外源的GSH对牙鲆存活率没有显著影响(P＞0.05)。WGR在饲料GSH含量为371.84 mg/Kg时达到最大值,显著高于其它各组(P＜0.05)。饲料GSH添加组牙鲆肝脏中GSH的含量显著高于零添加组(P＜0.05)。各处理组肝脏中MDA的含量没有显著差异(P>0.05)。肝脏中GR的活力随饲料中GSH含量的升高呈现先降低后上升的趋势,并在饲料中GSH含量为371.84 mg/Kg时达到最低值。肝脏中SOD和GPx活力未受到饲料中GSH含量的显著影响(P＞0.05)。饲料中GSH含量为371.84 mg/Kg时,肝脏中T-AOC显著高于1063.56 mg/Kg组(P＜0.05),GST活力显著低于其它各处理组(P＜0.05)。根据WGR的实验数据,利用折线模型得到牙鲆幼鱼饲料中GSH的适宜含量为368.92 mg/Kg,过高或过低都会对生长造成负面影响。
实验三：利用3×3实验设计,研究了在精制饲料中联合添加不同梯度LA(0、800和3200 mg/Kg)和硒(0、1.07和40 mg/Kg),配制9种实验饲料,投喂牙鲆(P. olivaceus)幼鱼(初始体重9.42±0.16 g),养殖8周后,计算和测定牙鲆生长和存活、肝脏GSH和MDA含量、肝脏GPx、GST、GR、SOD和T-AOC,以探讨饲料中LA和硒的交互作用对牙鲆生长和抗氧化的影响。
实验结果表明：LA与硒二者均过量添加组(LA:3200 mg/Kg;硒：40 mg/Kg)牙鲆幼鱼的存活率显著低于其它各处理组(P＜0.05),其余各组间存活率没有显著差异(均高于96.43%)(P＞0.05)。WGR受饲料中LA和硒的交互作用影响显著(P＞0.05),当饲料中缺硒(0 mg/Kg)或正常添加硒(1.07 mg/Kg)时,LA的适量添加(800 mg/Kg)能够使WGR有上升的趋势,特别在硒添加1.07 mg/Kg组,LA添加800 mg/Kg能够显著提高WGR(P＜0.05)。然而当饲料中硒含量过高(40 mg/Kg)时,饲料中LA的加入反而使WGR显著下降(P＜0.05)。肝脏中GSH含量受饲料中LA和硒交互作用影响显著(P＜0.05),饲料中缺硒(0 mg/Kg)或正常添加硒(1.07 mg/Kg)时,LA添加组的肝脏GSH积累量显著低于不添加LA各组(P＜0.05),然而当饲料中硒含量过高(40 mg/Kg)时,LA的添加能够使肝脏GSH的积累量有上升趋势(P＞0.05)。肝脏中MDA含量受到饲料中LA和硒的交互作用影响显著(P＜0.05),饲料缺硒时LA的过量添加(3200 mg/Kg)组的肝脏MDA含量显著高于其它两组(P＜0.05),饲料中硒含量为1.07 mg/Kg时LA的添加能够显著降低肝脏MDA积累量(P＜0.05),饲料中硒过量添加(40 mg/Kg)时LA添加3200 mg/Kg组的肝脏MDA积累量显著高于其它两组(P＜0.05)。肝脏中GPx活性变化受到饲料中LA和硒的影响显著(P＜0.05),当饲料中LA添加800 mg/Kg、硒含量为1.07 mg/Kg时,肝脏GPx活性显著高于其它各组(P＜0.05),饲料中硒含量一定时LA在饲料中添加量为800 mg/Kg各组肝脏GPx活性显著高于LA不添加(0 mg/Kg)和过量添加(3200 mg/Kg)各组(P<0.05),饲料中LA添加量一定时硒在饲料中含量为1.07 mg/Kg各组肝脏GPx活性显著高于硒不添加(0 mg/Kg)和过量添加(40 mg/Kg)各组(P<0.05)。肝脏中GST活性变化受到饲料中LA和硒的影响显著(P<0.05),当饲料中硒含量一定时LA添加800 mg/Kg各组GST活性显著最高(P<0.05),LA添加3200 mg/Kg各组的GST活性显著最低(P<0.05),当饲料中LA添加量一定时硒正常添加(1.07 mg/Kg)和过量添加(40 mg/Kg)各组肝脏GST活性显著高于硒零添加(0 mg/Kg)各组(P<0.05)。饲料中LA和硒的添加对肝脏GR活性均有显著影响(P<0.05),饲料中硒含量为1.07 mg/Kg、LA添加量为800 mg/Kg组的GR活性显著高于其它各组(P<0.05)。饲料中硒和LA对肝脏中T-AOC影响显著(P＜0.05),硒含量为1.07 mg/Kg、LA添加量为800 mg/Kg组的T-AOC显著高于其它各组(P＜0.05)。饲料中LA添加量对肝脏SOD影响并不显著(P＞0.05),饲料中LA添加量一定时,硒含量为40 mg/Kg各组SOD活性显著高于硒含量为0 mg/Kg和1.07 mg/Kg各组(P<0.05)。
实验四：利用3×3实验设计,研究了在精制饲料中联合添加不同梯度GSH(0、371.84和1063.56 mg/Kg)和硒(0、1.07和40 mg/Kg),配制9种实验饲料,投喂给牙鲆(P. olivaceus)幼鱼(初始体重9.49±0.17 g),养殖8周后,计算和测定牙鲆生长和存活、肝脏GSH和MDA含量、肝脏GPx、GST, GR、SOD和T-AOC,以探讨饲料中LA和硒的交互作用对牙鲆生长和抗氧化的影响。
实验结果表明：饲料中硒和GSH的交互作用对牙鲆存活率没有显著影响(P＜0.05)。在饲料中硒含量0 mg/Kg组的WGR显著低于饲料硒含量为1.07和40 mg/Kg组(P＜0.05)。饲料中硒和GSH的含量对牙鲆肝脏GSH积累量影响显著(P＜0.05),当饲料中硒含量一定时,饲料GSH含量为371.84 mg/Kg各组的肝脏GSH积累量显著高于其它各组(P＜0.05),或饲料中GSH含量一定时,饲料硒含量为1.07 mg/Kg各组的肝脏GSH积累量显著高于其它各组(P＜0.05)。饲料中硒和GSH的交互作用对肝脏MDA含量和SOD活性没有显著影响(P＞0.05)。饲料中添加GSH能够显著提高肝脏GPx活性(P＜0.05)。饲料中硒和GSH的含量对肝脏GST、GR活性和T-AOC均有显著影响(P＜0.05)。饲料中缺硒时(0 mg/Kg)添加371.84 mg/Kg GSH组的肝脏GST活性和T-AOC显著高于GSH零添加组(0 mg/Kg)和过量添加组(1063.56 mg/Kg) (P<0.05),而GR活性显著低于其它两组。饲料中硒含量合适(1.07 mg/Kg)时添加371.84 mg/Kg GSH组的肝脏GST和GR活性显著低于GSH零添加组(0 mg/Kg)和过量添加组(1063.56mg/Kg) (P<0.05),而T-AOC显著高于其它两组(P＜0.05)。饲料中硒过量添加(40 mg/Kg)时,肝脏GST活性随着饲料中GSH含量的升高而下降,GR活性在GSH添加量为371,84-mg/Kg组的活性显著高于其他两组(P＜0.05)。
A series of experiments were conducted to determine the effects of dietary reduced glutathione (GSH), a-lipoic acid (LA) and the interaction between selenium (Se) and GSH, Se and LA on growth and antioxidation in Japanese Flounder (Paralichthys olivaceus), a marine carnivorous fish. The results of the present study were summarized as follows:
Experiment 1:A one-factorial experiment was conducted to determine the effects of gradients of diatary LA on growth and antioxidation in juvenile Japanese flounder (initial weight was 9.39±0.17 g). The juveniles were divided into six triplicate groups, and fed one of the six purified diets (containing 0,200,400,800,1600 or 3200 LA mg/Kg dry diet respectively) for 8 weeks in a flow-through system. A series of indexes were conducted, such as weight gain rate (WGR), survival rate, the concentration of hepatic GSH and malonaldehyde (MDA), the hepatic total antioxidant capacity (T-AOC) and the activities of hepatic glutathione peroxidase (GPx), glutathione S-transferase (GST), glutathione reductase (GR) and superoxide dismutase (SOD).
The results showed that there were no significant difference in the survival of Japanese Flounder among six dietary treatments(P>0.05). The average WGR reached the significantly highest value in the groups fed the diets with 400 or 800 mg LA, which were significantly higher than those of the other groups (P<0.05). No significant difference was found between groups in the concentrations of hepatic GSH and activities of SOD (P>0.05). The activities of hepatic GST and GR decreased then increased with the increasing of supplied dietary LA (P<0.05). The lowest values of GST were found in the treatments with 400 or 800 mg LA/Kg diet (P<0.05), and the GR value of the treatment with 800 mg LA/Kg diet was significantly higher than the values in other treatments, except for the treatment with 1600 mg LA/Kg diet (P<0.05). The concentrations of hepatic MDA were significantly decreased with rising content of dietary LA (P<0.05), and the concentrations of hepatic MDA with 1600 and 3200 mg LA/Kg diet were significantly lower than the control group with 0 mg/Kg dietary LA (P<0.05). The activities of hepatic GPx and T-AOC increased then decreased with the increasing of supplied dietary LA (P<0.05). The values of GPx in the three treatments with 200,400 and 800 mg LA/Kg diet were significantly higher than the two treatments with 0 arid 3200 mg/Kg dietary LA (P<0.05). The highest value of T-AOC was found in the treatment with 800 mg LA/Kg diet, which was significantly higher than the two treatments with 0 and 3200 mg LA/Kg diet (P<0.05). According to the average WGR data, a broken-line model was employed to calculate the optimal levels of dietary LA, which was 745.05 mg/Kg diet. Deficient or overdose of dietary LA could cause negative effects on the growth and antioxidative capacity of Paralichthys olivaceus.
Experiment 2:A one-factorial experiment was conducted to determine the effects of gradients of diatary GSH on growth and antioxidation in juvenile Japanese flounder (initial weight was 9.49±0.22 g). The juveniles were divided into six triplicate groups, and fed on one of the six purified diets (containing 0,97.96,189.92,371.84,688.72 or 1063.56 GSH mg/Kg dry diet respectively) for 8 weeks in a flow-through system. A series of indexes were conducted, such as WGR, survival rate, the concentration of hepatic GSH and MDA, the hepatic T-AOC and the activities of hepatic GPx, GST, GR and SOD.
The results showed that survival was not significantly affected by dietary GSH levels (P>0.05). The average WGR in the treatment with 371.84 mg/Kg dietary GSH was significantly higher than those in other treatments (P<0.05). The hepatic GSH concentration in dietary GSH added groups was significantly higher than that in dietary GSH deprived group (control group) (P<0.05). No significant differences in hepatic MDA concentration were found between GSH deprived and supplemented groups (P>0.05). The lowest value of hepatic GR activities was found in treatment with 371.84 mg/Kg dietary GSH (P<0.05). There were no significant differences in the activities of hepatic SOD and GPx among the treatments (P>0.05). In the group with 371.84 mg/Kg dietary GSH, the T-AOC was significantly higher than that in the group with 1063.56 mg/Kg dietary GSH, however, the GST activity was significantly lower than those in the other groups (P<0.05). Based on the WGR data in the present experiment, a broken-line model was employed to calculate the optimal levels of dietary GSH, which was 368.92 mg/Kg diet. Deficient or overdose of dietary GSH could cause negative effects on the growth and anti-oxidative capacity of Paralichthys olivaceus.
Experiment 3:A two-factorial experiment was conducted to determine the Se and LA interaction on growth and antioxidation in juvenile Japanese flounder (initial weight was 9.42±0.16 g). Totally nine purified diets were made to provide graded levels of Se (0,1.07,40 mg/Kg, supplied as Na2Se03) and LA (0,800,3200 mg/Kg). The juveniles were divided into nine triplicate groups, and fed on one of the nine purified diets respectively, for 8 weeks in a flow-through system. A series of indexes were conducted, such as WGR, survival rate, the concentration of hepatic GSH and MDA, the hepatic T-AOC and the activities of hepatic GPx, GST, GR and SOD.
The results showed that the value of survival in the treatment with both 3200 mg LA/Kg diet and 40 mg Se/Kg diet was significantly lower than other treatments (P<0.05), and there was no significant difference between other groups (all higher than 96.43) (P>0.05). WGR was significantly affected by the interaction between LA and Se (P<0.05). When the diets were provided 0 or 1.07 mg Se/Kg diet, WGR presented a trend to rise in the treatment with 800 mg LA/Kg diet, and WGR reached the highest value in the treatment with both 800 mg LA and 1.07 mg Se/Kg diet, which was significant higher than other groups (P<0.05). Oppositely, in the diets with high selenium level (40 mg/Kg), values of WGR in the LA provided treatments was significantly lower than the control group (0 mg/Kg LA). Hepatic GSH concentration was significantly affected by the interaction between dietary LA and selenium (P<0.05). When dietary Se was deprived or provided 1.07 mg/Kg, the values of hepatic GSH concentration in the treatments provided LA were significantly lower than LA deprived treatments (P<0.05). Nevertheless, between the treatments of high Se level (40 mg/Kg), hepatic GSH concentration of the treatments with LA provided (800 or 3200 mg/Kg) had a rising trend compared with control group (P>0.05). Hepatic MDA concentration was significantly affected by the interaction between dietary LA and Se (P<0.05). The value of hepatic MDA concentration in the treatment with 3200 mg LA/Kg diet was significantly higher than the other two treatments when dietary Se was deprived (P<0.05). The values of hepatic MDA concentration in the treatments provided LA (800 or 3200 mg/Kg) were significantly lower than the control group (0 mg LA/Kg diet) when dietary Se was 1.07 mg/Kg (P<0.05). In the treatments of high Se level (40 mg/Kg), the values of hepatic MDA concentration of the treatment with 3200 mg LA/Kg diet was significantly higher than the other two treatments (P<0.05). The activities of hepatic GPx and GST were significantly affected by the interaction between dietary LA and Se (P<0.05). The highest value of hepatic GPx activity was found in the treatment with both 800 mg LA and 1.07 mg Se/Kg diet, and this value was significantly higher than those in other treatments (P<0.05). The values of hepatic GPx and GST of the treatments with 800 mg LA/Kg diet was significantly higher than those of the treatments with 0 or 3200 mg LA/Kg diet when dietary Se level was invariable (P<0.05). Likewise, the values of hepatic GPx of the treatments with 1.07 mg Se/Kg diet was significantly higher than those of the treatments with 0 or 40 mg Se/Kg diet when dietary LA level was invariable (P<0.05). The lowest values of hepatic GST activity was found in the treatments with 3200 mg LA/Kg diet (P<0.05). The values of hepatic GST of the treatments with 1.07 or 40 mg Se/Kg diet were significantly higher than the control group (0 mg Se/Kg diet) when dietary LA level was invariable (P<0.05). Both T-AOC and the activities of hepatic GR were significantly affected by both dietary LA and Se (P<0.05). The highest values were found in the treatment with both 800 mg LA and 1.07 mg Se/Kg diet, and the two values were significantly higher than those of other treatments, separately (P<.05). No significant difference was found between the diets with grads of LA (P<0.05). SOD activities in the treatments with 40 mg Se/Kg diet were significantly higher than SOD in both the control groups (dietary selenium 0 mg/Kg) and dietary Se 1.07 mg/Kg groups when dietary LA level was invariable (P<0.05).
Experiment 4:A two-factorial experiment was conducted to determine Se and GSH interaction on growth and antioxidation in juvenile Japanese flounder (initial weight was 9.49±0.17 g). Totally nine purified diets were made to provide graded levels of Se (0,1.07,40mg/Kg, supplied as Na2Se03) and GSH (0,371.84,1063.56 mg/Kg). The juveniles were divided into nine triplicate groups, and fed on one of the nine purified diets respectively, for 8 weeks in a flow-through system. A series of indexes were conducted, such as WGR, survival rate, the concentration of hepatic GSH and MDA, the hepatic T-AOC and the activities of hepatic GPx, GST, GR and SOD.
The results showed that survival was not significantly affected by the interaction between Se and GSH (P>0.05). The values of WGR in the Se provided treatments (1.07 or 40 mg/Kg) were significantly higher than the Se deprived treatments (0 mg/Kg) (P<0.05). The hepatic GSH concentrations were significantly affected by the interaction between GSH and Se (P<0.05). The hepatic GSH concentrations of the treatments with 371.84 mg GSH/Kg diet were significantly higher than those in other treatments when dietary Se level was invariable (P<0.05). Likewise, the hepatic GSH concentrations of the treatments with 1.07 mg Se/Kg diet were significantly higher than those in other treatments when dietary GSH level was invariable (P<0.05). There was no significant difference in hepatic MDA concentrations among nine dietary treatments, and so was the SOD activities (P>0.05). Hepatic GPx activities were significantly affected by dietary GSH levels (P<0.05), moreover hepatic T-AOC and GST, GR activities were significantly affected by the interaction between dietary Se and GSH (P<.05). Both hepatic T-AOC and GST activity in the treatment with 371.84 mg GSH/Kg diet were significantly higher than those in the treatments dealed with 0 or 1063.56 mg GSH/Kg diet when dietary Se was deprived (P<0.05). When dietary Se was 1.07 mg/Kg, the hepatic GST and GR in the treatment with 371.84 mg/Kg was significantly lower compared with the other two treatments with 0 or 1063.56 mg GSH/Kg diet, but T-AOC in 371.84 mg GSH/Kg diet treatment was significantly higher than the other two treatments (P<0.05). When dietary Se was 40 mg/Kg, hepatic GST activity decreased with dietary GSH level increased, and GR activity in dietary GSH 371.84 mg/Kg group was significantly higher than the other two groups (P<0.05).
引文
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