Chinese Journal of Animal Nutrition >

2025 , Vol. 37 >Issue 11: 7807 - 7819

DOI: https://doi.org/10.12418/CJAN2025.634

RESEARCH PAPER

Effects of Shrimp Meal Freshness on Growth Performance, Antioxidant Capacity and Hepatopancreas Health of Macrobrachium rosenbergii

  • WU Yuheng , 1 ,
  • XU Jie 1 ,
  • DENG Kangyu 2 ,
  • WANG Anran 1 ,
  • CHANG Enhui 1 ,
  • LUO Dongxu 1 ,
  • DONG Xiaojing 1 ,
  • DENG Deng 2 ,
  • MIAO Shuyan , 1, *
Expand
  • 1 College of Animal Science and Technology, Yangzhou university, Yangzhou 225009, China
  • 2 Shenzhen Alpha Feed Co., Ltd., Shenzhen 518054, China
*professor, E-mail:

Received date: 2025-04-10

  Online published: 2025-11-14

Fold

Abstract

This experiment was conducted to investigate the effect of shrimp meal freshness on growth performance, antioxidant capacity and hepatopancreas health of Macrobrachium rosenbergii. Three experimental diets were formulated by adding shrimp meal with different freshness levels [fresh: total volatile basic nitrogen (TVB-N) content of 104 mg/100 g; medium freshness: TVB-N content of 200 mg/100 g; low freshness: TVB-N content of 300 mg/100 g] into the basal diet at a proportion of 9%, resulting in dietary TVB-N contents of 55 mg/100 g (ST55 group, added with fresh shrimp meal), 70 mg/100 g (ST70 group, added with medium freshness shrimp meal) and 78 mg/100 g (ST78 group, added with low-freshness shrimp meal), respectively. A total of 552 healthy Macrobrachium rosenbergii juveniles with uniform size and an initial body weight of (1.07±0.01) g were selected and randomly divided into 3 groups, with 4 replicates per group and 46 juveniles per replicate. The feeding trial lasted for 73 days. The results showed as follows: 1) the final body weight (FBW), survival rate (SR), weight gain rate (WGR), specific growth rate (SGR), hepatosomatic index (HSI), and condition factor (CF) of ST70 group and ST78 group were significantly lower than those of ST55 group (P<0.05), while the feed conversion ratio (FCR) was significantly higher than that of ST55 group (P<0.05). 2) The activities of lipase (LPS), catalase (CAT), glutathione S-transferase (GST), glutathione peroxidase (GPx) and total superoxide dismutase (T-SOD) in hepatopancreas, as well as the contents of nitric oxide (NO), total protein (TP) and albumin (ALB) in serum of ST70 group and ST78 group were significantly lower than those of ST55 group (P<0.05), while the serum alanine aminotransferase (ALT) activity was significantly higher than that of ST55 group (P<0.05). 3) The mRNA relative expression levels of Relish, immune deficiency protein (IMD), GPx, copper-zinc superoxide dismutase (Cu/Zn SOD) and inhibitor of apoptosis protein (IAP) in hepatopancreas of ST70 group and ST78 group were significantly lower than those of ST55 group (P<0.05), while the mRNA relative expression levels of cysteine aspartate-specific protease 3 (Caspase3), cysteine aspartate-specific protease 9 (Caspase9), B-cell lymphoma-2 associated ovarian killer protein (Bok), and nuclear factor-κB (NF-κB) in hepatopancreas were significantly higher than those of ST55 group (P<0.05). 4) The results of ammonia-nitrogen stress and high-temperature stress tests showed that the cumulative survival rate of Macrobrachium rosenbergii in ST70 group and ST78 group was significantly lower than that in ST55 group (P<0.05). In conclusion, when the dietary TVB-N content exceeds 70 mg/100 g, the growth performance, antioxidant capacity, and stress resistance of Macrobrachium rosenbergii are significantly reduced, and hepatopancreas cell apoptosis is induced.

Key words： Macrobrachium rosenbergii; shrimp meal freshness; total volatile basic nitrogen; growth performance; antioxidant capacity; hepatopancreas health

Cite this article

WU Yuheng , XU Jie , DENG Kangyu , WANG Anran , CHANG Enhui , LUO Dongxu , DONG Xiaojing , DENG Deng , MIAO Shuyan . Effects of Shrimp Meal Freshness on Growth Performance, Antioxidant Capacity and Hepatopancreas Health of Macrobrachium rosenbergii[J]. Chinese Journal of Animal Nutrition, 2025 , 37(11) : 7807 -7819 . DOI: 10.12418/CJAN2025.634

虾粉是虾类经冷冻、干燥等工艺加工制成的蛋白质原料,富含虾青素、不饱和脂肪酸和磷脂,是水产饲料中常用的优质蛋白质源之一[1-2]。虾粉的新鲜度是评价其养殖效果与安全性的重要指标,主要通过蛋白质腐败程度和油脂氧化程度进行评估。蛋白质腐败可产生三甲胺、二甲胺、甲胺、氨氮及生物胺等物质,油脂氧化则生成丙二醛、低级脂肪酸、酮类和醇类等产物[3]。这些产物会显著降低虾粉的营养价值,并对养殖鱼、虾、蟹类的胃肠道及肝胰脏健康造成较严重的损伤[4-5]。因此,虾粉的新鲜度已成为制约饲料蛋白质源功能发挥的重要因素之一。
罗氏沼虾(Macrobrachium rosenbergii)作为我国重要的淡水养殖品种,凭借生长速度快、食性范围广、肉质鲜美及营养价值高等优势,在水产养殖领域备受关注。据统计,2023年我国罗氏沼虾养殖总量达19.64万t[6]。研究与生产实践表明,罗氏沼虾配合饲料中蛋白质含量通常为30%~45%[7]。虾粉作为常用蛋白质源,其新鲜度对罗氏沼虾生长与健康的影响尚未有系统研究。为此,本研究以虾粉蛋白质新鲜度为核心,通过将经腐败处理的虾粉添加至饲料中,配制不同总挥发性盐基氮(total volatile basic nitrogen,TVB-N)含量的试验饲料并投喂罗氏沼虾。通过测定其生长性能、形态学指标、肌肉常规营养成分、血清生化指标、肝胰腺消化酶活性、抗氧化相关指标和细胞凋亡相关基因的表达及抗应激能力,综合评估虾粉新鲜度对罗氏沼虾生长性能、抗氧化能力及肝胰腺健康的影响,以期为罗氏沼虾配合饲料的科学配制与质量评价提供理论依据。

1 材料与方法

1.1 虾粉腐败处理

在配制试验饲料前,对初始TVB-N含量为104 mg/100 g的新鲜虾粉进行腐败处理。将适量水与虾粉混合均匀后,置于23 ℃恒温培养箱中培养,每间隔4 h测定1次TVB-N含量,测定方法参考《食品安全国家标准 食品中挥发性盐基氮的测定》(GB 5009.228—2016)[8]。分别于处理后40和48 h获得TVB-N含量为200(中新鲜度)和300 mg/100 g(低新鲜度)的腐败虾粉样品各1份。

1.2 试验饲料

以鱼粉和鸡肉粉为主要蛋白质源,豆油和鱼油为主要脂肪源配制基础饲料。在基础饲料中按9%的比例分别添加不同新鲜度的虾粉,配制3种试验饲料,分别命名为ST55(添加新鲜虾粉,饲料TVB-N含量为55 mg/100 g)、ST70(添加中新鲜度虾粉,饲料TVB-N含量为70 mg/100 g)和ST78(添加低新鲜度虾粉,饲料TVB-N含量为78 mg/100 g)。试验饲料组成及营养水平见表1。
表1 试验饲料组成及营养水平(风干基础)

Table 1 Composition and nutrient levels of experimental diets (air-dry basis)%

项目
Items		 饲料Diets
ST55 ST70 ST78
原料Ingredients
豆粕Soybean meal 20.00 20.00 20.00
鱼粉(进口日本级) Fish meal (imported Japanese grade) 19.00 19.00 19.00
鱼粉(国产半脱脂) Fish meal (domestic semi degreased) 13.00 13.00 13.00
新鲜虾粉Fresh shrimp meal 9.00
中新鲜度虾粉Medium-freshness shrimp meal 9.00
低新鲜度虾粉Low-freshness shrimp meal 9.00
花生饼Peanut meal 5.00 5.00 5.00
家禽副产品粉Poultry by-product meal 2.50 2.50 2.50
虾膏Shrimp paste 2.00 2.00 2.00
面粉Wheat flour 19.00 19.00 19.00
乌贼膏Squid paste 2.00 2.00 2.00
大豆磷脂油Soybean lecithin oil 2.50 2.50 2.50
磷酸二氢钙Ca(H2PO4)2 1.50 1.50 1.50
大豆油Soybean oil 0.45 0.45 0.45
鱼油Fish oil 0.45 0.45 0.45
氯化胆碱Choline chloride (60%) 0.25 0.25 0.25
维生素预混料Vitamin premix1) 0.20 0.20 0.20
矿物质预混料Mineral premix2) 0.20 0.20 0.20
维生素C-2-磷脂酸Vitamin C-2-phosphate (35%) 0.10 0.10 0.10
有机酸Organic acid 0.05 0.05 0.05
膨润土Bentonite 2.60 2.60 2.60
瓜尔豆胶Guar gum 0.20 0.20 0.20
合计Total 100.00 100.00 100.00
营养水平Nutrient levels3)
水分Moisture 7.88 7.67 7.63
粗蛋白质CP 44.81 44.81 44.09
粗脂肪EE 7.79 7.50 7.39
粗灰分Ash 13.69 13.42 13.27
总挥发性盐基氮TVB-N/(mg/100 g) 55 70 78

1)每千克维生素预混料含有 One kilogram of vitamin premix contained the following:VA 10 000 IU,VD 2 500 IU,VK 64 mg,硫氨素 thiamine 60 mg,VB6 60 mg,泛酸钙 calcium pantothenate 240 mg,叶酸 folic acid 12 mg,生物素 biotin 50 mg,VB12 4 mg。

3)实测值 Measured values。

2)每千克矿物质预混料含有 One kilogram of mineral premix contained the following:Ca(H2PO4)2 10 g,MgSO4·7H2O 2.4 g,KCl 4.5 g,NaCl 2.1 g,FeSO4·H2O 155 mg,CuSO4·5H2O 40 mg,ZnSO4·H2O 80 mg,MnSO4·H2O 30 mg,KI 11.7 mg,CoCl2·6H2O 4.8 mg,Na2SeO3 2.4 mg。

饲料制备流程如下:所有固体原料均经微型研磨机(HK820,广州市旭朗机械制造有限公司)研磨后过100目筛,充分混合均匀;向混合后的固体原料中加入液体原料及适量水,再次搅拌混匀;将混匀的物料通过双螺杆挤条机(F-26,广州华工光机电科技有限公司)制成直径1.5 mm的颗粒饲料,充分晾干后置于-20 ℃冰箱中保存备用。

1.3 试验设计与饲养管理

动物试验已通过扬州大学实验动物伦理委员会批准(批准号:YZUDWSY 2017-09-06)。试验所用罗氏沼虾虾苗购自江苏省扬州市某虾苗厂,在室内循环养殖系统中驯化1周后,选取552尾健康、规格均匀、初始体重为(1.07±0.01) g的虾苗,随机分为3组,每组4个重复,每个重复46尾。以重复为单位将虾苗饲养于室内养殖池内的尼龙网箱(0.8 m×0.8 m×0.7 m)中,试验期73 d。每日06:30和18:30各饱食投喂1次,投喂后1.5 h捞出残饵,晾干后置于-20 ℃冰箱保存,并每日记录投喂量与残饵量。养殖期间,采用增氧系统24 h持续供氧,每日换水量控制在30%~50%,每周使用水质多参数监测仪(YSI,美国)检测1次水质,控制水体溶解氧浓度≥6 mg/L、亚硝酸盐氮浓度≤0.02 mg/L、总氨氮浓度≤0.1 mg/L、温度为25~31 ℃、pH为6.5~8.5。

1.4 样品采集

养殖试验结束后,试验虾禁食24 h,记录每个网箱中存活虾数量及终末重量,以计算存活率(SR)、增重率(WGR)、特定生长率(SGR)和饲料系数(FCR)。从每个网箱中随机选取10尾虾,测量其体长和体重,以计算肥满度(CF);使用无菌注射器从虾围心腔抽取0.5 mL血淋巴加入装有等量抗凝剂的离心管中,1 790×g离心10 min后收集血清,置于-80 ℃冰箱保存;完整取出肝胰腺并称重,以计算肝体指数(HSI),将每个网箱10尾虾的肝胰腺混匀后分装至冻存管中,置于-80 ℃冰箱保存;收集每尾虾完整的肌肉组织,置于-20 ℃冰箱保存。

1.5 氨氮应激及高温应激试验

样品采集完成后,从每个网箱剩余虾中随机选取10尾活力良好、无残肢的健康个体,分别置于10 L圆柱形水箱中进行氨氮应激试验。试验条件参考Yang等[9]的48 h半致死浓度(LC50)试验及本研究初步试验结果设定。将试验虾置于离子态氨氮浓度为150 mg/L的水环境中持续应激48 h,氨氮浓度通过添加10 mg/mL的氯化铵溶液进行调节。试验期间禁食,保持水体溶解氧充足,控制水温为(30±1) ℃、pH为7.0±0.5,每2 h记录1次各水箱中存活虾数量。
高温应激试验采用与氨氮应激试验一致的虾只选取标准,同样于10 L圆柱形水箱中进行。使用加热棒控温,将试验虾置于38.5 ℃条件下持续应激12 h,期间每2 h记录1次各水箱中存活虾数量。

1.6 指标测定及方法

1.6.1 生长性能和形态学指标

生长性能和形态学指标计算公式如下:
SR(%)=100×Nf/Ni;
WGR(%)=100×(Wf-Wi)/Wi;
SGR(%/d)=100×(lnWf-lnWi)/T;
FCR=I/(Wf-Wi);
HSI(%)=100×Wh/W;
CF(g/cm3)=100×W/L3。
式中:Nf为终末存活虾数量(尾);Ni为初始虾数量(尾);Wi为初始体重(g);Wf为终末体重(g);T为养殖天数(d);I为摄入饲料质量(g);Wh为肝胰腺重(g);W为虾体重(g);L为虾体长(cm)。

1.6.2 饲料和肌肉营养成分

参考AOAC(1995)[10]的方法测定饲料及试验虾肌肉的粗蛋白质、粗脂肪、粗灰分和水分含量。将样品在65 ℃下烘至恒重,并在自然条件下回潮24 h,然后使用粉碎机粉碎,获得分析样品。采用105 ℃烘干恒重法测定水分含量,采用高温灼烧法测定粗灰分含量,采用凯氏定氮法测定粗蛋白质含量,采用索氏提取法测定粗脂肪含量。饲料TVB-N含量参考《食品安全国家标准 食品中挥发性盐基氮的测定》(GB 5009.228—2016)的方法进行测定。

1.6.3 肝胰腺消化酶活性和抗氧化指标

肝胰腺α-淀粉酶(α-AMS)、脂肪酶(LPS)、胰蛋白酶(TPS)活性;总超氧化物歧化酶(T-SOD)、谷胱甘肽硫转移酶(GST)、谷胱甘肽过氧化物酶(GPx)、过氧化氢酶(CAT)活性、总抗氧化能力(T-AOC)及丙二醛(MDA)含量均采用南京建成生物工程研究所生产的试剂盒进行测定。

1.6.4 血清生化指标

血清谷丙转氨酶(ALT)、谷草转氨酶(AST)活性及一氧化氮(NO)、总蛋白(TP)、白蛋白(ALB)含量均采用南京建成生物工程研究所生产的试剂盒进行测定。

1.6.5 肝胰腺抗氧化及细胞凋亡相关基因mRNA相对表达量

使用Trizol法提取肝胰腺总RNA,经反转录后使用Quant Studio 3荧光定量分析仪(Thermo Fisher Scientific,美国)进行实时荧光定量PCR分析。基因引物序列如表2所示,所有引物序列均通过NCBI在线引物设计工具设计。以β-肌动蛋白(β-actin)为内参基因,采用2-△△Ct法计算目的基因的mRNA相对表达量。
表2 引物序列

Table 2 Primer sequences

基因Genes 引物序列Primer sequences (5'—3')
半胱氨酸天冬氨酸蛋白酶3 Caspase3 F:CGGATTCAAACGCGATGACC
R:GACGACAACGTGGTCTGACT
半胱氨酸天冬氨酸蛋白酶8 Caspase8 F:GCGAAAGAACTACTCGGCCG
R:AGCAGCAGCCAGGAACTTGT
半胱氨酸天冬氨酸蛋白酶9 Caspase9 F:ACCTTCTTTCATATTTCGTCC
R:ACTGTGCTCGCTGTCCTA
B细胞淋巴瘤-2相关卵巢杀伤蛋白Bok F:CGCCACAGTAGGAGAGAAGG
R:TGAAAACGGCAATGGACATA
核因子-κB NF-κB F:GTGGCTCACTTACGACTC
R:AAGGTCCATACTCTTTGC
凋亡抑制蛋白IAP F:ACCGTTTGGCAGCTTGCACCA
R:CTCCGGCCTGCAGTGTGTCC
Relish F:GATGAGCCTTCAGTGCCAGA
R:CCAGGTGACGCCATGTATCA
免疫缺陷蛋白IMD F:CGACCACATTCTCCTCCTCCC
R:TTCAGTGCATCCACGTCCCTC
谷胱甘肽过氧化物酶GPx F:AGATGAGCGCAATCAAAAGC
R:GCCGTGTTCTCCACAAGAAT
铜锌超氧化物歧化酶Cu/Zn SOD F:GGAGCTAGATTGGCTTGCTG
R:AGAAAATCACTGCCCCTCCT
β-肌动蛋白β-actin F:TCCGTAAGGACCTGTATGCC
R:TCGGGAGGTGCGATGATTTT

1.7 数据统计与分析

使用SPSS 27.0软件对试验数据进行单因素方差分析(one-way ANOVA),并采用Duncan氏法进行多重比较,结果以平均值±标准差表示,P<0.05为差异显著。

2 结 果

2.1 虾粉新鲜度对罗氏沼虾生长性能、形态学指标和肌肉营养成分的影响

由表3可知,ST70组和ST78组终末体重、SR、WGR、SGR、HSI和CF显著低于ST55组(P<0.05),FCR显著高于ST55组(P<0.05);ST70组与ST78组之间SR和HSI无显著差异(P>0.05)。由表4可知,3组间各肌肉营养成分指标均无显著差异(P>0.05)。
表3 虾粉新鲜度对罗氏沼虾生长性能及和形态学指标的影响

Table 3 Effects of shrimp meal freshness on growth performance and morphological indices of Macrobrachium rosenbergii

项目
Items		 组别Groups P值
P-value
ST55 ST70 ST78
初始体重IBW/g 1.06±0.01 1.07±0.01 1.07±0.01 0.114
终末体重FBW/g 12.74±0.27c 11.43±0.39b 10.53±0.23a <0.001
存活率SR/% 81.67±1.11b 77.23±1.11a 76.67±1.28a <0.001
增重率WGR/% 1 110.60±8.98c 954.01±31.18b 872.76±17.70a <0.001
特定生长率SGR/(%/d) 3.40±0.03c 3.24±0.06b 3.13±0.04a <0.001
饲料系数FCR 4.84±0.06a 5.56±0.07b 6.46±0.20c <0.001
肝体比HSI/% 3.90±0.13b 3.46±0.08a 3.39±0.22a 0.002
肥满度CF/(g/cm3) 1.06±0.01b 1.04±0.00a 1.04±0.01a <0.001

同行数据肩标无字母或相同小写字母表示差异不显著(P>0.05),不同小写字母表示差异显著(P<0.05)。下表同。

In the same row, values with no letter or the same small letter superscripts mean no significant difference (P>0.05), while with different small letter superscripts mean significant difference (P<0.05). The same as below.

表4 虾粉新鲜度对罗氏沼虾肌肉营养成分的影响

Table 4 Effects of shrimp meal freshness on muscle nutrient of Macrobrachium rosenbergii%

项目
Items		 组别Groups P值
P-value
ST55 ST70 ST78
水分Moisture 77.72±0.72 77.74±0.35 77.65±0.29 0.961
粗蛋白质CP 21.21±0.51 21.12±0.19 21.25±0.26 0.870
粗脂肪EE 0.65±0.01 0.62±0.04 0.68±0.05 0.119
粗灰分Ash 1.48±0.27 1.60±0.09 1.67±0.05 0.336

水分含量为湿物质基础,粗蛋白质、粗脂肪和粗灰分含量为风干基础。

Moisture content was wet matter basis, while the contents of CP, EE and Ash were air-dry basis.

2.2 虾粉新鲜度对罗氏沼虾肝胰腺消化酶活性的影响

由表5可知,ST70组和ST78组肝胰腺LPS活性显著低于ST55组(P<0.05),且ST70组与ST78组之间无显著差异(P>0.05)。3组之间肝胰腺α-AMS和TPS活性均无显著差异(P>0.05)。
表5 虾粉新鲜度对罗氏沼虾肝胰腺消化酶活性的影响

Table 5 Effects of shrimp meal freshness on hepatopancreas digestive enzyme activities of Macrobrachium rosenbergii U/(mg prot)

项目
Items		 组别Groups P值
P-value
ST55 ST70 ST78
α-淀粉酶α-AMS 64.56±9.46 55.05±2.84 54.99±1.70 0.070
胰蛋白酶TPS 3 056.21±458.86 3 044.48±173.48 2 926.70±439.84 0.870
脂肪酶LPS 18.73±1.94b 13.32±1.79a 12.16±1.16a <0.001

2.3 虾粉新鲜度对罗氏沼虾肝胰腺抗氧化能力的影响

由表6和图1可知,虾粉新鲜度显著影响肝胰腺GPx、GST、T-SOD、CAT活性、T-AOC及MDA含量(P<0.05),同时也显著影响肝胰腺Relish、免疫缺陷蛋白(IMD)、GPx和铜锌超氧化物歧化酶(Cu/Zn SOD)的mRNA相对表达量(P<0.05)。肝胰腺GST、T-SOD和CAT活性随着虾粉TVB-N含量的增加逐渐降低,ST70组显著低于ST55组(P<0.05),ST78组显著低于ST70组(P<0.05);肝胰腺Cu/Zn SOD的mRNA相对表达量也呈现相同趋势。ST70组和ST78组肝胰腺GPx活性及Relish、IMD的mRNA相对表达量显著低于ST55组(P<0.05),且ST70组与ST78组之间无显著差异(P>0.05)。此外,ST78组肝胰腺T-AOC及GPx的mRNA相对表达量显著低于ST55组和ST70组(P<0.05),肝胰腺MDA含量显著高于ST55组和ST70组(P<0.05),但ST55组与ST70组之间上述指标均无显著差异(P>0.05)。
表6 虾粉新鲜度对罗氏沼虾肝胰腺抗氧化指标的影响

Table 6 Effects of shrimp meal freshness on hepatopancreas antioxidant indices of Macrobrachium rosenbergii

项目
Items		 组别Groups P值
P-value
ST55 ST70 ST78
谷胱甘肽过氧化物酶GPx/(U/mg prot) 249.63±20.70b 195.42±28.48a 193.33±0.46a 0.005
谷胱甘肽硫转移酶GST/(U/mg prot) 103.30±0.78c 85.74±2.18b 64.75±2.60a <0.001
总抗氧化能力T-AOC/(mmol/g prot) 0.14±0.01b 0.13±0.02b 0.11±0.01a 0.016
总超氧化物歧化酶T-SOD/(U/mg prot) 157.14±0.76c 127.48±9.56b 117.01±1.66a <0.001
丙二醛MDA/(nmol/mg prot) 2.62±0.36a 2.87±0.67a 7.20±0.96b <0.001
过氧化氢酶CAT/(U/mg prot) 1.24±0.11c 0.72±0.12b 0.39±0.07a <0.001
图1 虾粉新鲜度对罗氏沼虾肝胰腺抗氧化相关基因表达的影响

数据柱形标注不同小写字母表示差异显著(P<0.05)。下图同。

Fig.1 Effects of shrimp meal freshness on expression of genes related to antioxidant in hepatopancreas of Macrobrachium rosenbergii

Value columns with different small letters mean significant difference (P<0.05). The same as below.

2.4 虾粉新鲜度对罗氏沼虾血清生化指标的影响

由表7可知,虾粉新鲜度对血清AST活性无显著影响(P>0.05)。血清TP含量随着虾粉TVB-N含量的增加逐渐降低,ST70组显著低于ST55组(P<0.05),ST78组显著低于ST70组(P<0.05)。ST70组和ST78组血清ALT活性显著高于ST55组(P<0.05),血清NO和ALB含量显著低于ST55组(P<0.05),且ST70组与ST78组之间上述指标均无显著差异(P>0.05)。
表7 虾粉新鲜度对罗氏沼虾血清生化指标的影响

Table 7 Effects of shrimp meal freshness on serum biochemical indices of Macrobrachium rosenbergii

项目
Items		 组别Groups P值
P-value
ST55 ST70 ST78
谷丙转氨酶ALT/(U/L) 8.47±0.03a 15.69±1.84b 15.39±1.52b <0.001
谷草转氨酶AST/(U/L) 108.61±13.07 120.18±12.35 119.57±15.49 0.439
白蛋白ALB/(g/L) 58.64±6.26b 47.14±1.49a 47.86±1.90a 0.004
总蛋白TP/(g/L) 187.43±19.83c 105.47±5.24b 85.16±0.29a <0.001
一氧化氮NO/(μmol/L) 104.15±2.18b 85.62±4.55a 83.43±10.77a 0.004

2.5 虾粉新鲜度对罗氏沼虾肝胰腺细胞凋亡相关基因表达的影响

由图2可知,虾粉新鲜度显著影响肝胰腺半胱氨酸天冬氨酸蛋白酶3(Caspase3)、半胱氨酸天冬氨酸蛋白酶8(Caspase8)、半胱氨酸天冬氨酸蛋白酶9(Caspase9)、B细胞淋巴瘤-2相关卵巢杀伤蛋白(Bok)、核因子-κB(NF-κB)和凋亡抑制蛋白(IAP)的mRNA相对表达量(P<0.05)。肝胰腺Bok和NF-κB的mRNA相对表达量随着虾粉TVB-N含量的增加逐渐升高,ST70组显著高于ST55组(P<0.05),ST78组显著高于ST70组(P<0.05)。ST70组和ST78组肝胰腺Caspase3和Caspase9的mRNA相对表达量显著高于ST55组(P<0.05),肝胰腺IAP的mRNA相对表达量显著低于ST55组(P<0.05),且ST70组与ST78组之间上述指标均无显著差异(P>0.05)。
图2 虾粉新鲜度对罗氏沼虾肝胰腺细胞凋亡相关基因表达的影响

Fig.2 Effects of shrimp meal freshness on expression of genes related to apoptosis in hepatopancreas of Macrobrachium rosenbergii

2.6 虾粉新鲜度对罗氏沼虾抗应激能力的影响

氨氮应激试验结果如图3和表8所示,应激12 h时,ST70组累积存活率与ST55组接近,而24 h后其累积存活率显著低于ST55组(P<0.05);ST78组则从应激12 h开始,累积存活率便显著低于ST55组(P<0.05),且随着应激时间的延长,该组累积存活率持续下降,始终维持在最低水平。
图3 氨氮应激下罗氏沼虾的累积存活率变化趋势

Fig.3 Trend of cumulative survival rate of Macrobrachium rosenbergii under ammonia-nitrogen stress

表8 虾粉新鲜度对罗氏沼虾抗氨氮应激能力的影响

Table 8 Effects of shrimp meal freshness on ammonia-nitrogen stress resistance of Macrobrachium rosenbergii%

项目
Items		 组别Groups P值
P-value
ST55 ST70 ST78
12 h累积存活率Cumulative survival rate at 12 h 93.75±7.22b 87.50±10.21b 75.00±0.00a 0.015
24 h累积存活率Cumulative survival rate at 24 h 71.88±6.25b 50.00±10.21a 37.50±10.21a 0.001
36 h累积存活率Cumulative survival rate at 36 h 62.50±10.21c 40.63±11.97b 21.88±11.97a 0.002
48 h累积存活率Cumulative survival rate at 48 h 46.88±6.25c 28.13±11.97b 6.25±7.22a <0.001
高温应激试验结果如图4和表9所示,应激2~12 h,ST78组累积存活率始终显著低于ST70组(P<0.05),ST70组累积存活率始终显著低于ST55组(P<0.05)。
图4 高温应激下罗氏沼虾的累积存活率变化趋势

Fig.4 Trend of cumulative survival rate of Macrobrachium rosenbergii under high-temperature stress

表9 虾粉新鲜度对罗氏沼虾抗高温应激能力的影响

Table 9 Effects of shrimp meal freshness on high-temperature stress resistance of Macrobrachium rosenbergii%

项目
Items		 组别Groups P值
P-value
ST55 ST70 ST78
2 h累积存活率Cumulative survival rate at 2 h 93.75±7.22c 81.25±7.22b 62.50±0.00a <0.001
4 h累积存活率Cumulative survival rate at 4 h 81.25±7.22c 59.38±6.25b 40.63±6.25a <0.001
6 h累积存活率Cumulative survival rate at 6 h 68.75±7.22c 43.75±7.22b 25.00±10.21a <0.001
8 h累积存活率Cumulative survival rate at 8 h 56.25±7.22c 31.25±7.22b 6.25±7.22a <0.001
10 h累积存活率Cumulative survival rate at 10 h 40.63±6.25c 18.75±12.50b 0.00±0.00a <0.001
12 h累积存活率Cumulative survival rate at 12 h 37.50±0.00c 12.50±10.21b 0.00±0.00a <0.001

3 讨论

罗氏沼虾的生长性能与其营养状况密切相关。虾粉作为饲料中常用的高蛋白质原料,在高温、高湿及氧化条件下易发生变质,导致蛋白质及其他含氮化合物降解,产生有机胺类物质并增加TVB-N含量[11],进而降低水产饲料的营养价值和适口性。在本试验中,当饲料所用虾粉的TVB-N含量超过200 mg/100 g(即饲料TVB-N含量超过70 mg/100 g)时,罗氏沼虾的生长受到明显抑制,SR显著降低,FCR显著提高。Ricque-Marie等[12]研究发现,与摄食对照组饲料(所用鱼粉TVB-N含量为14 mg/100 g)相比,摄食试验组饲料(所用鱼粉TVB-N含量分别为30和50 mg/100 g)使得凡纳滨对虾(Penaeus vannamei)和蓝对虾(Penaeus stylirostris)幼体的生长和摄食量受到了明显抑制。Hu等[13]研究表明,摄食低TVB-N含量(100 mg/100 g)鱼粉饲料组的日本海鲈(Lateolabrax japonicus)生长性能和饲料利用率优于摄食中国本地鱼粉(TVB-N含量为130 mg/100 g)饲料组。上述研究结果均与本试验结果相符。消化酶活性能够部分反映动物基本的消化生理特征及其对饲料营养物质的利用能力。在本试验中,ST70组和ST78组罗氏沼虾肝胰腺LPS活性较ST55组显著降低,这可能与虾粉腐败分解产生的胺类物质对罗氏沼虾消化能力产生的不利影响有关[14]。
机体的抗氧化能力可通过T-AOC及T-SOD、GST、GPx和CAT活性进行评估。这些抗氧化物质在抵御氧化应激、维持机体氧化还原平衡的过程中发挥着至关重要的作用[15-17]。本试验结果表明,随着饲料中虾粉TVB-N含量的升高,罗氏沼虾肝胰腺GST、GPx、CAT、T-SOD活性及T-AOC均显著降低。这可能是由于饲料中虾粉腐败产生了有害的胺类或酮类、醛类等物质,导致罗氏沼虾摄食后对外源产物刺激的代偿能力和对机体自由基的清除能力减弱[4,18]。MDA是机体内膜脂质过氧化反应的终产物,由自由基氧化脂质生成。它不仅能与蛋白质、核酸等生物大分子发生反应,导致这些分子丧失正常生理功能,还具有一定的细胞毒性,可干扰细胞膜完整性、破坏能量代谢过程及信号传导通路,进而影响细胞正常生理功能[19-20]。本试验中,ST78组罗氏沼虾肝胰腺MDA含量较ST70组和ST50组显著提高,一方面可能是由于虾粉自身脂肪氧化产生的MDA随饲料摄入后,在肝胰腺中积累[21];另一方面可能与虾体抗氧化能力下降导致活性氧大量积累,引发脂质过氧化反应加剧,间接引起细胞膜选择透过性降低、胞内物质外流及肝胰腺组织受损有关,从而使MDA含量进一步升高[22-23]。此外,Relish、IMD、GPx及Cu/Zn SOD均为机体抗氧化相关基因[24-25],其中GPx和Cu/Zn SOD常被用作评估机体氧化应激状态的分子标志物[26-27]。本试验结果表明,随着饲料中虾粉TVB-N含量的升高,罗氏沼虾肝胰腺Relish、IMD、GPx及Cu/Zn SOD的表达下调,这与肝胰腺抗氧化酶活性的测定结果一致。以上结果表明,摄食较低新鲜度虾粉饲料会引起罗氏沼虾肝胰腺抗氧化功能异常。
ALT、AST活性及TP、NO含量也常被用于评估罗氏沼虾的抗氧化能力与免疫功能。其中,AST与ALT作为肝细胞内的重要转氨酶,对鱼类和虾类的代谢活动具有重要意义[28],其在血清中的活性可反映罗氏沼虾的肝胰腺损伤程度[29]。TP主要包括ALB和球蛋白(GLB),参与蛋白质代谢与肝功能调节;NO则由一氧化氮合酶催化产生,是一种重要的活性氮分子,在免疫防御中发挥关键作用[30]。本试验结果表明,随着饲料中虾粉TVB-N含量的升高,罗氏沼虾血清ALT活性显著提高,提示肝胰腺出现明显损伤;同时,血清TP、ALB和NO含量均显著降低,表明摄食较低新鲜度虾粉饲料削弱了罗氏沼虾的免疫功能。
当机体面临的氧化应激强度超出自身固有抗氧化系统的清除能力时,细胞会发生凋亡反应[31]。肝胰腺细胞凋亡作为一种高度有序的程序性过程,在生物体自我稳态调节、维持消化代谢等多种生理功能正常运行中发挥着基础性作用[32-33]。在细胞凋亡的分子调控机制中,半胱氨酸天冬氨酸蛋白酶(Caspase)基因家族既是介导凋亡过程的关键因子,也是凋亡信号传导通路中的核心共同途径[34];此外,Bok、IAP与NF-κB同样参与细胞凋亡的调控过程。本试验结果表明,随着饲料中虾粉TVB-N含量的升高,罗氏沼虾肝胰腺中Bok、NF-κB、Caspase3、Caspase8及Caspase9的表达均显著上调,而具有凋亡抑制功能的IAP的表达则显著下调。该结果表明,摄食较低新鲜度虾粉饲料可诱导罗氏沼虾肝胰腺细胞发生凋亡,进而引发肝胰腺损伤,这一结果也通过血清ALT和AST活性的提高得到了证实。
应激试验常用来综合评价动物的健康状态和免疫能力[35]。在罗氏沼虾池塘养殖中,水环境中的高氨氮胁迫、异常天气引发的高温胁迫等环境压力,已成为制约养殖产量与生产效益的主要因素[36-37]。因此,本研究采用氨氮应激和高温应激试验来综合评价虾粉新鲜度对罗氏沼虾抗应激能力的影响。结果表明,投喂较低新鲜度虾粉饲料显著降低了罗氏沼虾的抗应激能力,这也与在罗氏沼虾的生长性能和肝胰腺健康状况中所观察到的趋势一致。

4 结论

在本试验中,TVB-N含量(200和300 mg/100 g)较高的虾粉对罗氏沼虾的生长性能、抗氧化能力、肝胰腺健康和抗应激能力产生了不利影响,并导致了肝胰腺的细胞凋亡。生产中,罗氏沼虾配合饲料的TVB-N含量不应超过70 mg/100 g。

References

Publishing order | Descend order by publishing year | Descend order by cited within
[1]
韩萌萌, 陈迪, 杨颖. 海洋动物蛋白及其产品应用的研究进展[J]. 现代食品, 2024, 30(14):220-222.

HAN M M, CHEN D, YANG Y. Research progress in the application of marine animal protein and its products[J]. Modern Food, 2024, 30(14):220-222. (in Chinese)

[2]
CARVALHO M, MONTERO D, BETANCOR M, et al. Benefits of dietary krill meal inclusion towards better utilization of nutrients,and response to oxidative stress in gilthead seabream (Sparus aurata) juveniles[J]. Aquaculture, 2025,598:741957.

[3]
曹小华, 李小銮, 蔡懋成, 等. 饲用虾粉的营养特性与新鲜度测定[J]. 粮食与饲料工业, 2017(7):46-50.

CAO X H, LI X L, CAI M C, et al. Nutritional characteristics and freshness determination of feed shrimp meal[J]. Cereal & Feed Industry, 2017(7):46-50. (in Chinese)

[4]
刘晓蕊, 马喜波, 孔祎頔, 等. 组胺对水产动物影响的研究进展[J]. 饲料工业, 2025, 46(4):107-111.

LIU X R, MA X B, KONG Y D, et al. Research progress on effects of histamine on aquatic animals[J]. Feed Industry, 2025, 46(4):107-111. (in Chinese)

[5]
杨保和, 袁明凤, 唐精. 氧化油脂对水产动物的危害[J]. 饲料博览, 2013(4):35-39.

YANG B H, YUAN M F, TANG J. Hazards of oxidized oils in aquatic animals[J]. Feed Review, 2013(4):35-39. (in Chinese)

[6]
中华人民共和国农业农村部渔业渔政管理局, 全国水产技术推广总站,中国水产学会.2024中国渔业统计年鉴[M]. 北京: 中国农业出版社, 2024.

Ministry of Agriculture and Rural Affairs of the People's Republic of China,National Fisheries Technology Extension Center, China Society of Fisheries.China fishery statistical yearbook in 2024[M]. Beijing: China Agriculture Press, 2024. (in Chinese)

[7]
刘敏, 孙广文, 王卓铎, 等. 罗氏沼虾营养需求与功能性饲料研究进展(一)[J]. 广东饲料, 2022, 31(2):35-40.

LIU M, SUN G W, WANG Z D, et al. Research progress of nutritional requirement and functional feed of Macrobrachium rosenbergii[J]. Guangdong Feed, 2022, 31(2):35-40. (in Chinese)

[8]
中华人民共和国国家卫生和计划生育委员会.食品安全国家标准食品中挥发性盐基氮的测定:GB 5009.228—2016[S]. 北京: 中国标准出版社, 2017.

National Health and Family Planning Commission of the People's Republic of China.National food safety standard for determination of volatile base nitrogen in food:GB 5009.228—2016[S]. Beijing: Standards Press of China, 2017. (in Chinese)

[9]
YANG X Z, SHI A Y, SONG Y M, et al. The effects of ammonia-N stress on immune parameters,antioxidant capacity,digestive function,and intestinal microflora of Chinese mitten crab,Eriocheir sinensis,and the protective effect of dietary supplement of melatonin[J]. Comparative Biochemistry and Physiology Part C:Toxicology & Pharmacology, 2021,250:109127.

[10]
AOAC. Official methods of analysis of official analytical chemists international[M]. 16th ed. Arlington: Association of Official Analytical Chemists,1995.

[11]
BEKHIT A E D A, HOLMAN B W B, GITERU S G, et al. Total volatile basic nitrogen (TVB-N) and its role in meat spoilage:a review[J]. Trends in Food Science & Technology, 2021,109:280-302.

[12]
RICQUE-MARIE D, ABDO-DE LA PARRA M I, CRUZ-SUAREZ L E, et al. Raw material freshness,a quality criterion for fish meal fed to shrimp[J]. Aquaculture, 1998, 165(1/2):95-109.

[13]
HU L, YUN B, XUE M, et al. Effects of fish meal quality and fish meal substitution by animal protein blend on growth performance,flesh quality and liver histology of Japanese seabass (Lateolabrax japonicus)[J]. Aquaculture, 2013,372:52-61.

[14]
张金彪. 常见野杂鱼中的生物胺分析及其对中华绒螯蟹(Eriocheir sinensis)存活、生长和体内累积的影响[D]. 硕士学位论文. 上海: 上海海洋大学, 2013.

ZHANG J B. Survival,growth and tissue histamine accumulation of the Eriocheir sinensis,fed biogenic amies from several common trash fish[D]. Master's Thesis. Shanghai: Shanghai Ocean University, 2013. (in Chinese)

[15]
彭士明, 陈立侨, 侯俊利, 等. 氧化鱼油词料中添加VE对黑鲷幼鱼体脂含量及肝脏抗氧化酶活性的影响[J]. 上海水产大学学报, 2008, 17(3):298-304.

PENG S M, CHEN L Q, HOU J L, et al. Effect of dietary vitamin E on lipid content and hepatic antioxidant enzyme activities of juvenile black seabream, Acanthopagrus schlegeli fed oxidized fish oil[J]. Journal of Shanghai Ocean University, 2008, 17(3):298-304. (in Chinese)

[16]
PATI S G, PANDA F, PAITAL B, et al. Oxidative stress physiology in Scylla serrata for environmental health assessment[J]. Frontiers in Environmental Science, 2023,11:1142495.

[17]
WU P, ZHANG L, JIANG W D, et al. Dietary vitamin A improved the flesh quality of grass carp (Ctenopharyngodon idella) in relation to the enhanced antioxidant capacity through Nrf2/Keap 1a signaling pathway[J].Antioxidants, 2022, 11(1):148.

[18]
梁高杨, 李小勤, 高启平, 等. 氧化油脂对斑点叉尾鮰生长、体色和抗氧化能力的影响[J]. 水生生物学报, 2019, 43(1):60-68.

LIANG G Y, LI X Q, GAO Q P, et al. Effect of oxidized oils on growth,body color and antioxidant capacity of channel catfish,Ictalurus punctatus[J]. Acta Hydrobiologica Sinica, 2019, 43(1):60-68. (in Chinese)

[19]
吴倩, 刘雪婷, 胡俊茹, 等. 壳寡糖对甲壳动物的生长性能、免疫力和肠道健康的影响[J]. 饲料工业, 2025, 46(8):42-48.

WU Q, LIU X T, HU J R, et al. Effects of chitosan on growth performance,immunity,and gut health of crustaceans[J]. Feed Industry, 2025, 46(8):42-48. (in Chinese)

[20]
宋嘉欣, 简永慧, 陈致远, 等. 饲料中不同脂肪水平对罗氏沼虾蚤状幼体生长、抗氧化、免疫和抗应激能力的影响[J]. 水生生物学报, 2025, 49(7):124-134.

SONG J X, JIAN Y H, CHEN Z Y, et al. Dietary lipid levels on growth,antioxidant capacity,immunity,and stress resistance in zoeae larvae of Macrobrachium rosenbergii[J]. Acta Hydrobiologica Sinica, 2025, 49(7):124-134. (in Chinese)

[21]
林秀秀, 叶元土, 蔡春芳, 等. 丙二醛引起草鱼肠道、肝胰脏谷胱甘肽/谷胱甘肽转移酶通路抗氧化应激[J]. 动物营养学报, 2015, 27(11):3604-3612.

LIN X X, YE Y T, CAI C F, et al. Malondialdehyde causes glutathione/glutathione transferase pathway oxidative stress in intestine and hepatopancreas of grass carp (Ctenopharyngodon idellus)[J]. Chinese Journal of Animal Nutrition, 2015, 27(11):3604-3612. (in Chinese)

[22]
YIN H Y, XU L B, PORTER N A. Free radical lipid peroxidation:mechanisms and analysis[J]. Chemical Reviews, 2011, 111(10):5944-5972.

[23]
SLATTER D A, BOLTON C H, BAILEY A J. The importance of lipid-derived malondialdehyde in diabetes mellitus[J]. Diabetologia, 2000, 43(5):550-557.

[24]
杨密, 宋宇朵, 陈瑞, 等. 饲粮甜菜碱水平对雪山草鸡腿肌肉品质、糖酵解水平和抗氧化能力的影响[J]. 南京农业大学学报, 2022, 45(3):578-586.

YANG M, SONG Y D, CHEN R, et al. Effects of dietary betaine levels on meat quality,glycolysis level,antioxidant ability of Xueshan chicken[J]. Journal of Nanjing Agricultural University, 2022, 45(3):578-586. (in Chinese)

[25]
WANG H, LIU P P, WEI Z, et al. PcEiger links the Imd/Relish pathway to ROS production in the intestine of the red swamp crayfish[J]. EMBO Reports, 2023, 24(5):55903.

[26]
朱嘉宁, 孙万合, 雷元曦, 等. 内源RAS启动子提高草菇cat2基因表达效率的研究[J]. 菌物学报, 2025, 44(1):240183.

ZHU J N, SUN W H, LEI Y X, et al. Endogenous RAS promoter enhancing the expression efficiency of cat2 gene in Volvariella volvacea[J]. Mycosystema, 2025, 44(1):240183. (in Chinese)

[27]
DONG X X, LIU Q G, ZHAO W H, et al. Effects of ammonia-N stress on the antioxidant enzymes,heat shock proteins,and apoptosis-related genes of Macrobrachium rosenbergii[J]. Italian Journal of Animal Science, 2021, 20(1):453-464.

[28]
SHEIKHZADEH N, TAYEFI-NASRABADI H, OUSHANI A K, et al. Effects of haematococcus pluvialis supplementation on antioxidant system and metabolism in rainbow trout (Oncorhynchus mykiss)[J]. Fish Physiology and Biochemistry, 2012, 38(2):413-419.

[29]
WANGARI M R, GAO Q, SUN C X, et al. Effect of dietary Clostridium butyricum and different feeding patterns on growth performance,antioxidant and immune capacity in freshwater prawn (Macrobrachium rosenbergii)[J]. Aquaculture Research, 2021, 52(1):12-22.

[30]
LAI Q N, YU C, ZHAO F R, et al. Protection of pacific white shrimp (Penaeus vannamei) against white spot syndrome virus infection by nitric oxide-generating compound s-nitrosoglutathione[J]. Aquaculture, 2024,579:740204.

[31]
ZHENG X, FU D Y, CHENG J H, et al. Effects of hypoxic stress and recovery on oxidative stress,apoptosis,and intestinal microorganisms in Pelteobagrus vachelli[J]. Aquaculture, 2021,543:736945.

[32]
SHUANG L, CHEN S L, REN C, et al. Effects of hypoxia and reoxygenation on oxidative stress,histological structure,and apoptosis in a new hypoxia-tolerant variety of blunt snout bream (Megalobrama amblycephala)[J]. Comparative Biochemistry and Physiology Part A:Molecular & Integrative Physiology, 2023,278:111358.

[33]
LU Z J, TANG M Z, ZHANG M L, et al. Expression and functional analysis of the BCL2-associated agonist of cell death (BAD) gene in grass carp (Ctenopharyngodon idella) during bacterial infection[J]. Developmental & Comparative Immunology, 2021,123:104160.

[34]
SMOLEWSKI P, DARZYNKIEWICZ Z, ROBAK T. Caspase-mediated cell death in hematological malignancies:theoretical considerations,methods of assessment,and clinical implications[J]. Leukemia & Lymphoma, 2003, 44(7):1089-1104.

[35]
LIU T, LU Y, SUN M Y, et al. Effects of acute hypoxia and reoxygenation on histological structure,antioxidant response,and apoptosis in razor clam Sinonovacula constricta[J]. Fish & Shellfish Immunology, 2024,145:109310.

[36]
BIR J, SARKER H, MITA F S, et al. The impact of salinity and temperature stress on survival,behaviour,immune response,and proximate composition of giant freshwater prawn Macrobrachium rosenbergii[J]. Aquaculture International, 2024, 32(5):6333-6352.

[37]
成良峰, 戴习林. 慢性氨氮胁迫对罗氏沼虾生长与存活及能量代谢的影响[J]. 南方农业学报, 2023, 54(9):2752-2761.

CHENG L F, DAI X L. Effects of chronic ammonia nitrogen stress on growth,survival and energy metabolism of Macrobrachium rosenbergii[J]. Journal of Southern Agriculture, 2023, 54(9):2752-2761. (in Chinese)

Options
Outlines

/

〈 〉

京ICP备17050989号-2
Copyright © Chinese Journal of Animal Nutrition, All Rights Reserved.
Tel: 010－62817823　E-mail: cjan@vip.163.com
Powered by Beijing Magtech Co., Ltd.