渔业科学进展  2022, Vol. 43 Issue (2): 80-88  DOI: 10.19663/j.issn2095-9869.20210713001
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引用本文 

贲玲芝, 史雪莹, 郭金龙, 陈秀玲, 张配瑜, 刘海燕. 全脂黑水虻幼虫粉替代鱼粉对大菱鲆养殖性能、生理代谢及体色的影响[J]. 渔业科学进展, 2022, 43(2): 80-88. DOI: 10.19663/j.issn2095-9869.20210713001.
BEN Lingzhi, SHI Xueying, GUO Jinlong, CHEN Xiuling, ZHANG Peiyu, LIU Haiyan. Effects of Replacement of Fish Meal with Full-Fat Hermetia illucens Larvae on Culture Performance, Physiological Metabolism, and Skin Color in Turbot[J]. Progress in Fishery Sciences, 2022, 43(2): 80-88. DOI: 10.19663/j.issn2095-9869.20210713001.

基金项目

河北省省级科技计划(19226705D)、河北省现代农业产业技术体系专项(HBCT2018170205)和河北省农业农村厅科技计划(冀农科19033)共同资助

作者简介

贲玲芝，E-mail: 1741885806@qq.com

通讯作者

刘海燕，教授，E-mail: liuhaiyan@hebtu.edu.cn

文章历史

收稿日期：2021-07-13
收修改稿日期：2021-09-22
Contents              Abstract              Full text              Figures/Tables              PDF
全脂黑水虻幼虫粉替代鱼粉对大菱鲆养殖性能、生理代谢及体色的影响
贲玲芝 1, 史雪莹 1, 郭金龙 2, 陈秀玲 3, 张配瑜 1, 刘海燕 1     
1. 河北师范大学生命科学学院 河北 石家庄 050024;
2. 河北省海洋与水产科学研究院 河北 秦皇岛 066002;
3. 北戴河新区水产技术推广站 河北 秦皇岛 066600
收稿日期：2021-07-13；收修改稿日期：2021-09-22
基金项目：河北省省级科技计划(19226705D)、河北省现代农业产业技术体系专项(HBCT2018170205)和河北省农业农村厅科技计划(冀农科19033)共同资助
作者简介：贲玲芝，E-mail: 1741885806@qq.com.
通讯作者：刘海燕，教授，E-mail: liuhaiyan@hebtu.edu.cn.
摘要：以大菱鲆(Scophthalmus maximus)为研究对象，探讨其配合饲料中以全脂黑水虻(Hermetia illucens)幼虫粉替代鱼粉后对大菱鲆生长性能、饲料利用、血清生理生化指标及体色的影响。本研究设计4个处理组，分别为全脂黑水虻幼虫粉替代0(FM)(对照组)、20%(HI20)、40%(HI40)的鱼粉及40%替代后添加晶体赖氨酸(Lys)和蛋氨酸(Met)(HI40AA)形成4种等氮、等能的实验饲料。将1200尾大菱鲆幼鱼[(26.57±0.54) g]随机分为4组，每组3个重复，每个重复100尾鱼，实验时间为56 d。结果显示，黑水虻幼虫粉替代鱼粉显著影响了大菱鲆的相对增重率(WGR)和特定生长率(SGR)，HI40和HI40AA组显著低于HI20和FM组(P < 0.05)，对大菱鲆的摄食率(SR)、饲料系数(FCR)、肝体比(HSI)、脏体比(VSI)、肥满度(CF)及体成分中的粗蛋白、粗灰分含量均无显著影响(P > 0.05)，FM和HI20组大菱鲆血清总胆固醇(TCHO)及高密度脂蛋白(HDL)含量显著高于HI40和HI40AA组(P < 0.05)，FM组低密度脂蛋白(LDL)含量显著高于各替代组(P < 0.05)；HI40和HI40AA组大菱鲆背部体色L*值、a*值显著低于FM和HI20组(P < 0.05)，FM组背部体色b*值显著低于其他各实验组(P < 0.05)，HI40组的b*值显著高于HI20和HI40AA组(P < 0.05)；大菱鲆腹部的体色参数各实验组相比无显著差异(P > 0.05)。研究表明，本研究条件下，全脂黑水虻幼虫粉替代大菱鲆饲料中鱼粉为20%时，不会影响大菱鲆的生长与生理功能，其在大菱鲆幼鱼饲料中的鱼粉替代推荐量为20%。
关键词：大菱鲆    黑水虻虫    生长    血清生化指标    体色    
Effects of Replacement of Fish Meal with Full-Fat Hermetia illucens Larvae on Culture Performance, Physiological Metabolism, and Skin Color in Turbot
BEN Lingzhi 1, SHI Xueying 1, GUO Jinlong 2, CHEN Xiuling 3, ZHANG Peiyu 1, LIU Haiyan 1     
1. College of Life Sciences, Hebei Normal University, Shijiazhuang Hebei 050024, China;
2. Hebei Ocean and Fisheries Science Research Institute, Qinhuangdao Hebei 066002, China;
3. Beidaihe New District Aquatic Technology Promotion Station, Qinhuangdao Hebei 066600, China
Corresponding author: LIU Haiyan, E-mail: liuhaiyan@hebtu.edu.cn.
Fund: This work was supported by Science and Technology Program of Hebei (19226705D), Hebei MATRT (HBCT2018170205), and Sciense and Technology Program of Apartment of Agriculture and Rural affairs of Hebei (19033)
Abstract: Turbot (Scophthalmus maximus) is an important mariculture species in northern China because of its high economic value. As a cold-water carnivorous fish, its rapid growth relies on high dietary protein in the form of fish meal. Fish meal is a major component of aquatic feed because of its highly digestible protein, balanced amino acids, and good palatability. However, global fish meal production is insufficient, and its price has soared due to overfishing and climate change. There is an urgent need to find high-quality protein sources to replace fish meal in the diet. Many experimental studies on replacing fish meal with protein sources have identified shortcomings. Plant protein sources contain anti-nutritional factors and defects in the amino acid profile, and have poor palatability; animal protein sources are unstable in their nutrient composition. Much work remains to be done to identify new protein sources. Insects are the largest organism community in ecosystems, and offer a new kind of protein source with great potential. The European Commission has recently approved the use of protein derived from insects in aquatic feed. Hermetia illucens is a saprophytic insect that can consume livestock manure and domestic waste to produce high-value animal feedstuff. Its larvae can be used as an alternative protein source to fish meal because they are rich in amino acids and polyunsaturated fatty acids. In addition, H. illucens is fed on decaying organic matter such as food waste, animal excrement, and animal and plant carcasses; the breeding cost is low, and the adults do not disturb humans. It can be added to the feed of sea bass (Dicentrarchus labrax), rainbow trout (Oncorhynchus mykiss), catfish (Pelteobagrus fulvidraco), large yellow croaker (Larimichthys crocea), grass carp (Ctenopharyngodon idellus), california bass (Micropterus salmoides), and Scophthalmus maximus (turbot). Most research has investigated the defatted dry powder of H. illucens. However, there are no reports on the replacement of fish meal with full-fat H. illucens in turbot. Therefore, in this study, we aimed to evaluate the application of H. illucens larvae as turbot feed. This experiment was conducted to investigate the effects of replacement of fish meal protein with full-fat H. illucens larvae meal on the growth performance, feed utilization, serum biochemical indexes, and skin color of turbot, in order to provide important information for turbot culture. Four treatments were designed for this study. Four isonitrogenous and isoenergetic diets were formulated with H. illucens larvae meal by substituting it into fish meal at 0% (FM), 20% (HI20), and 40% (HI40); the fourth experimental diet (HI40AA) was HI40 supplemented with methionine and lysine. A total of 1200 turbot [initial body weight: (26.57±0.54) g] were randomly distributed into four experimental diets, each group with three replicates, and each replicate with 100 juvenile turbot. The fish were fed twice daily to apparent satiation. The water temperature was kept at 11.0℃~14.4℃ and the water was changed twice daily. After the 56 d feeding trial, the results showed that H. illucens larvae meal significantly influenced weight gain and specific growth rate, which were significantly lower in the HI40 and HI40AA groups than in the HI20 and FM groups (P < 0.05); no significant differences were observed in the feeding rate, feed conversion ratio, hepatosomatic index, viscerosomatic index, condition factor, or body composition, including crude protein and ash, among all groups (P > 0.05). The serum cholesterol and high-density lipoprotein in the HI40 and HI40AA groups were significantly lower than those in the HI20 and FM groups (P < 0.05), and the content of serum low-density lipoprotein in the FM group was significantly higher than that in the other groups (P < 0.05). The values of L* (lightness) and a* (redness) of turbot dorsal skin color in the HI40 and HI40AA groups were significantly lower than those in the FM and HI20 groups (P < 0.05). The value of b* (yellowness) of turbot dorsal skin color in the FM group was significantly lower than those in the other groups (P < 0.05); it was higher in the HI40 group than in the HI20 and HI40AA groups (P < 0.05). No significant differences were observed in skin color parameters in the ventral area of the turbot (P > 0.05). In conclusion, the replacement of dietary fish meal with H. illucens did not affect the growth performance and physiological parameters of turbot when the replacement level was no more than 20%, which is therefore the recommended substitution level of fish meal by full-fat H. illucens larvae meal in the diet of juvenile turbot.
Key words: Turbot    Hermetia illucens    Growth performance    Serum biochemical indexes    Skin color    

大菱鲆(Scophthalmus maximus)，又名多宝鱼，是我国北方三省重要的海水养殖动物。作为冷水肉食性鱼类，其对饲料的蛋白质需求较高，且主要依赖于鱼粉。然而，由于过度捕捞及不良气候的影响，全球鱼粉资源已明显不足且价格飙升(杜玉雯等, 2016)，亟需展开替代鱼粉的研究，并开发适合大菱鲆的优质蛋白源。昆虫是一类新型动物蛋白源，其有望在未来替代鱼粉方面发挥重要作用。黑水虻(Hermetia illucens)为腐生性昆虫，能取食禽畜粪便和生活垃圾并生产高价值的动物蛋白饲料。其幼虫氨基酸含量丰富，中链脂肪酸与多不饱和脂肪酸含量占脂肪酸总量的60%以上，其中，月桂酸(Lauric acid)含量高达40%，可作为鱼粉的替代蛋白源(刘世胜, 2016; Oteri et al, 2021)。目前，已经在海鲈(Dicentrarchus labrax) (Rui et al, 2017)、虹鳟(Oncorhynchus mykiss)(Renna et al, 2017)、黄颡鱼(Pelteobagrus fulvidraco)(Xiao et al, 2018; 陈晓瑛等, 2019)、大黄鱼(Larimichthys crocea)(韩星星, 2019)、草鱼(Ctenopharyngodon idellus)(Lu et al, 2020)、加州鲈(Micropterus salmoides)(彭凯等, 2021)及大菱鲆(Kroeckel et al, 2012)中进行了应用研究。这些研究结果多是基于黑水虻虫的脱脂干粉进行的，但在实际生产中，市场上常见的是全脂黑水虻虫粉。全脂黑水虻虫粉对大菱鲆的影响尚未见报道。因此，本研究拟探讨全脂黑水虻虫粉替代鱼粉对大菱鲆幼鱼的生长、饲料利用、血清生理生化指标及体色的影响，为全脂黑水虻虫粉在大菱鲆饲料中的应用提供理论依据。

1 材料与方法 1.1 实验设计

本实验以大菱鲆为研究对象，共分4个处理组：分别采用全脂黑水虻幼虫粉替代基础饲料中0、20%和40%的鱼粉，及在40%替代组添加晶体氨基酸[赖氨酸(Lys)和蛋氨酸(Met)]，分别标记为FM (对照组)、HI20、HI40和HI40AA组，每组3个重复，每个重复100尾大菱鲆，共1200尾，养殖时间为56 d。

1.2 实验鱼及实验饲料

大菱鲆幼苗购于天津盛亿水产公司，从中选出1200尾大小均匀、健康的幼鱼，随机分到12个养殖缸内，大菱鲆幼鱼的初始体重为(26.57±0.54) g。实验所用黑水虻幼虫粉购自广州飞禧特生物科技有限公司，其粗蛋白及粗脂肪的含量分别为38.81%和29.94%。以白鱼粉、乌贼肝粉、豆粕、谷朊粉为主要蛋白源，鱼油、豆油为主要脂肪源配置实验饲料。饲料组成和营养成分见表 1。使用颗粒机将饲料制成直径为3.0 mm的软颗粒，存放至–20℃冰箱备用。

表 1 实验饲料配方及营养组成 Tab.1 Formulation and composition of experimental diet nutrient content
1.3 实验条件

养殖实验在河北省秦皇岛天合水产良种有限公司进行，所用鱼缸为长方体玻璃纤维缸(2 m×1 m×1 m)，缸内水深为40 cm。每个玻璃纤维缸均匀放置2个充氧气石，24 h增氧，保证溶解氧(DO)充足。DO > 6.0 mg/L，氨氮(NH4+-N)浓度＜0.05 mg/L。养殖用水为砂滤处理后的地下海水(盐度为25~28)，缸内水温为11.0℃~14.4℃。光照采用自然光照加人工光照，光照周期为14L∶10D。

1.4 养殖管理

养殖实验正式开始前，对大菱鲆幼鱼进行为期14 d的驯化，与正式实验养殖环境及投喂时间相同，使其行为安逸，摄食正常。结束后，禁食24 h，称重，选择体质健壮、规格整齐的个体作为实验用鱼，根据动物分组的随机化原则，分组称重。每日饱食投喂2次(08:00、18:00)，换水2次，定时记录水温；每日观察大菱鲆幼鱼的摄食和活动状态，如有死亡，及时取出并称重记录。

1.5 实验样品的采集和处理

养殖实验结束后，禁食24 h，对每缸实验鱼进行称重并记录。每缸随机取3尾大菱鲆，称重记录后，装入塑封袋保存至–20℃冰箱中，用于全鱼常规营养成分指标检测；每缸随机取6尾鱼尾静脉抽血，静置2 h，使用D1008E离心机(美国)于3000 r/min离心11 min，取血清，保存至–80℃冰箱待测血清生理生化指标；每缸随机取3尾鱼，称重、记录，使用游标卡尺测量并记录每条鱼的体重、体长，再迅速于冰盘上解剖，取完整内脏团，采用生理盐水冲洗，滤纸吸干，称重、记录，再剥离肝脏，称重、记录，放入液氮中速冻，用于肝脏抗氧化能力的测定；每缸取3尾鱼在其背、腹面各设置3个固定点进行色差的测定。

1.6 养殖性能指标

存活率(survival rate, SR, %)=100×终末鱼数量/初始鱼数量；

增重率(weight gain rate, WGR, %)=100×(Wt– W0)/W0；

饲料系数(feed efficiency ratio, FCR)=It/(Wt–W0)；

特定生长率(specific growth rate, SGR, %/d)=100× [lnWt–lnW0]/t；

摄食率(feeding rate, FR, %/d)=100×It/[t×(Wt+ W0)/2]；

式中，Wt(g)为终末体重；W0(g)为初始体重；t为实验天数；It(g)为摄入饲料干重。

肥满度(condition factor, CF) =100×体重/体长3；

脏体比(viscerosomatic index, VSI, %)=100×内脏重/体重；

肝体比(hepatosomatic index, HSI, %)=100×肝脏重/体重。

1.7 样品分析

水分含量采用直接干燥法进行测定(GB/T6435)；粗蛋白含量采用FOSS全自动凯氏定氮仪测定(GB/T6432)；粗脂肪含量采用索氏提取法测定(GB/T6433)；粗灰分含量采用马弗炉灼烧法测定(GB/T6438)；能量采用Parr6300 (Parr, 美国)全自动氧弹式热量计测定。血清生理生化指标在河北师范大学采用Mindary BS-180全自动分析仪测定。肝脏组织样本的超氧化物歧化酶(SOD)活性、丙二醛(MDA)含量以及总蛋白浓度的测定分别采用BCA(bicinchoninic acid)法、黄嘌呤氧化酶法、TBA法，即硫代巴比妥酸(Thibabituric Acid)法，所用试剂盒购自南京建成生物工程研究所。体色采用美能达CR–400色彩色差仪(日本)进行测定。

1.8 数据处理及统计分析

各实验数据以平均值±标准差(mean±SD)表示，使用Statistica 10.0软件首先对各项实验数据进行单因素方差分析(one-way ANOVA)，差异显著时进行Duncan’s多重比较检验，P < 0.05为差异显著。

2 结果 2.1 黑水虻幼虫粉替代鱼粉对大菱鲆养殖性能的影响

从表 2可以看出，饲料中黑水虻幼虫粉替代鱼粉对大菱鲆的终末均体重(FBW)、WGR及SGR的影响均差异显著(P < 0.05)。其中，FM和HI20组显著高于HI40和HI40AA组，FM和HI20组、HI40和HI40AA组相比无显著差异(P > 0.05)；黑水虻幼虫粉替代鱼粉对大菱鲆的FR无显著影响(P > 0.05)；FCR在各组之间相比也无显著差异(P > 0.05)；大菱鲆的SR、VSI、HSI和CF各组相比均无显著差异(P > 0.05)。

表 2 黑水虻幼虫粉替代鱼粉对大菱鲆生长、饲料利用及形体指标的影响(平均数±标准差) Tab.2 Effects of replacement of fish meal with H. illucens in diets on the growth performance, feed utilization and body index of turbot (Mean±SD)
2.2 黑水虻幼虫粉替代鱼粉对大菱鲆鱼体成分的影响

从表 3可以看出，黑水虻幼虫粉替代鱼粉对大菱鲆体成分中的水分、粗蛋白及粗灰分含量均无显著影响(P > 0.05)。HI40AA组大菱鲆粗脂肪含量显著低于其他3组(P < 0.05)。

表 3 黑水虻幼虫粉替代鱼粉对大菱鲆体成分的影响(平均数±标准差) Tab.3 Effects of replacement of fish meal with H. illucens in diets on the body composition of turbot (Mean±SD)
2.3 黑水虻幼虫粉替代鱼粉对大菱鲆血清生化指标的影响

从表 4可以看出，FM和HI20组的总胆固醇(TCHO)和高密度脂蛋白(HDL)含量均显著高于HI40和HI40AA组(P < 0.05)。FM组血清中低密度脂蛋白(LDL)含量显著高于各实验组(P < 0.05)。除此之外，甘油三酯(TG)、血糖(GLU)、谷丙转氨酶(ALT)、谷草转氨酶(AST)与碱性磷酸酶(ALP)等指标各实验组相比无显著差异(P > 0.05)。

表 4 黑水虻幼虫粉替代鱼粉对大菱鲆血清生化指标的影响(平均数±标准差) Tab.4 Effects of replacement of fish meal with H. illucens in diets on the serum biochemical indexes of turbot (Mean±SD)
2.4 黑水虻幼虫粉对大菱鲆肝脏抗氧化能力的影响

从表 5可以看出，黑水虻幼虫粉替代鱼粉后，各实验组SOD活力及MDA含量相比均无显著差异(P > 0.05)。但随替代比例的增加，各实验组呈先升高再降低的趋势，各黑水虻幼虫添加组SOD活力均高于FM组。

表 5 黑水虻幼虫粉替代鱼粉对大菱鲆肝脏抗氧化能力的影响(平均数±标准差) Tab.5 Effects of replacement of fish meal with H. illucens in diets on the hepatic antioxidant capacity of turbot (Mean±SD)
2.5 黑水虻幼虫粉替代鱼粉对大菱鲆体色的影响

从表 6可以看出，HI40和HI40AA组大菱鲆幼鱼的体色特征表现为背部体色L*值、a*值显著低于FM和HI20组(P < 0.05)。HI40组的b*值显著高于其他各组(P < 0.05)，HI20和HI40AA组的b*值也显著高于FM组(P < 0.05)；大菱鲆幼鱼腹部体色L*值、a*值和b*值相比均无显著差异(P > 0.05)。

表 6 黑水虻幼虫粉替代鱼粉对大菱鲆体色的影响(平均数±标准差) Tab.6 Effects of replacement of fish meal with H. illucens in diets on the skin color of turbot (Mean±SD)
3 讨论

黑水虻虫粉替代饲料中鱼粉蛋白质的研究在一些鱼类中已有报道。Caimi等(2020)在含70%鱼粉的饲料中，以脱脂黑水虻虫粉替代了鱼粉用量的25%时，西伯利亚鲟鱼(Acipenser baerii)的生长不受影响，当替代比例高达50%时，生长出现显著下降；Rui等(2017)研究发现，饲料中脱脂黑水虻幼虫粉的添加量为19.5%时，可改善海鲈的生长和摄食，但未达到显著水平；Xiao等(2018)研究表明，黄颡鱼饲料中含黑水虻虫粉为22.3%时，其WGR和SGR有所提高，当添加水平高于34.3%时，生长性能下降。其他昆虫蛋白在鱼类饲料研究中也有相似的结果，梅琳等(2015)在含鱼粉为65%的基础饲料中，采用蛹肽蛋白粉替代鱼粉用量的15%时，不会对大菱鲆幼鱼的生长和饲料利用造成影响，若高于此比例，幼鱼的生长性能随替代比例的增加而显著降低；饶远等(2019)在含鱼粉40%的基础饲料中，以蚕粉替代17.90%~22.00%的鱼粉蛋白，加州鲈幼鱼的生长性能不受影响。与以上研究结果相似，本研究中，随着饲料中全脂黑水虻虫粉添加量的增加，大菱鲆的生长呈下降趋势，当黑水虻幼虫粉替代鱼粉蛋白20%时，对大菱鲆的生长性能无显著影响，然而，当替代水平达到40%(在饲料中的添加比例为27.39%)时，生长性能出现显著降低。此研究结果与Kroeckel等(2012)研究结果有所不同。Kroeckel等(2012)研究发现，当脱脂黑水虻虫粉替代鱼粉比例超过17%时，大菱鲆的SGR出现显著降低。这说明脱脂工艺可能也引起黑水虻幼虫粉中某些营养物质的流失，降低了黑水虻幼虫粉的营养价值。脱脂黑水虻虫粉与全脂黑水虻虫粉相比，其在饲料中替代鱼粉时的水平要略低。当替代鱼粉比例过高时，2种黑水虻虫粉均导致大菱鲆的生长性能降低。黑水虻虫粉高水平替代带来的不良影响可能与黑水虻虫粉中的氨基酸模式及高含量的几丁质有关。黑水虻幼虫粉中的必需氨基酸模式略逊于鱼粉，赖氨酸和蛋氨酸等的含量低于鱼粉(Li et al, 2021)。本研究在HI40AA组额外补充了赖氨酸和蛋氨酸后，仍未达到提高大菱鲆生长的目的，这间接说明本研究中高含量黑水虻虫粉降低大菱鲆生长的主要原因可能不在于其氨基酸模式，而是由于其所含有的几丁质成分。黑水虻虫具有高含量的几丁质，Kroeckel等(2012)并没有在大菱鲆的肠中检测到几丁质酶的活性，高含量的几丁质能抑制消化道中营养物质的吸收，从而降低了大菱鲆的生长。

本研究中，大菱鲆各实验组VSI、HSI、CF相比无显著差异。但Kroeckel等(2012)研究表明，饲料中脱脂黑水虻虫粉含量升高使大菱鲆的CF和HSI含量显著降低，与本研究结果略有不同，这可能与实验所使用大菱鲆的大小及黑水虻幼虫品质存在的差异有关，不同的养殖基质饲养黑水虻幼虫，对幼虫的营养成分影响很大。本研究中，各实验组大菱鲆体成分中的粗蛋白、水分及粗灰分含量相比均无显著影响，而随着替代比例的增加，粗脂肪含量下降，这与黑水虻幼虫制品替代鱼粉在虹鳟、罗非鱼(Oreochromis mossambicus)中的研究结果相似(St-hilair et al, 2007；胡俊茹等, 2018)。在一些鱼类的研究中发现，随着黑水虻替代鱼粉的比例增加，鱼体脂肪呈现出不同的变化规律。Zhang等(2008)和Li等(2016)研究表明，几丁质及其衍生物能干扰脂肪酸的合成以及促进肝脏中脂蛋白和TG的水解。因此，脂肪含量的降低可能是由于几丁质影响了鱼体肝脏脂肪酸的合成(Li et al, 2017)。饲料中脂肪酸的比例不同，会对鱼体脂质消化率以及能量变化产生影响，进而影响鱼体脂肪的含量(邝哲师等, 2021)。但大菱鲆脂肪代谢受黑水虻虫粉影响的调控机制还有待进一步研究。

鱼类的血清生理生化指标可以体现机体的营养代谢特点及生理健康状况。本研究中，替代比例为40%时，TCHO含量显著降低，与黑水虻虫制品在幼建鲤(Cyprinus carpiovar Jian)、海鲈、鲈鱼(Lateolabrax japonicus)等饲料中添加的研究结果相似(Li et al, 2017; Rui et al, 2017; 胡俊茹等, 2018)，可能与饲料中黑水虻虫与饲料中添加的油脂组成有关。HDL将机体中的胆固醇运送回肝脏组织中进行代谢，LDL作用则相反，且易于沉积从而引起疾病的发生。本研究中，大菱鲆饲料中黑水虻幼虫粉替代鱼粉超过40%时，HDL含量显著降低，FM组的LDL含量显著高于其他3个实验组，也进一步说明在此替代比例下，大菱鲆幼鱼血清中胆固醇在肝脏和其他组织的输送过程出现了异常。这与韩星星(2019)使用脱脂黑水虻虫粉替代鱼粉在大黄鱼的研究结果相一致。郝甜甜等(2019)使用复合蛋白源替代大菱鲆幼鱼饲料中的鱼粉时发现，大菱鲆脂质代谢出现异常，主要表现为血清胆固醇、HDL及LDL降低。ALT和AST在正常情况下分布于肝细胞中，当细胞受到损伤时，这2种酶会从肝细胞释放到血清中，从而含量升高，其升高程度表明肝细胞的受损程度。本研究中，以黑水虻幼虫粉替代大菱鲆饲料中的鱼粉未对血清中的ALT及AST含量造成显著影响，但40%组的AST比对照组高，表明黑水虻幼虫粉较高水平的替代可能对大菱鲆幼鱼的肝脏产生一定的应激。

SOD具有清除活性氧，能阻断脂质过氧化作用，其活力的变化可以反映鱼机体健康及免疫状态(Campa-Cordova et al, 2002)；MDA是脂质发生过氧化反应后的产物之一，不仅能表明机体脂质过氧化程度，还可以评价细胞氧化损伤程度(白冬清等, 2011)。刘兴等(2017)研究发现，随着黑水虻替代鱼粉比例的增加，锦鲤(Cyprinus carpio)肝胰脏内SOD活力升高，MDA含量降低；黑水虻幼虫替代鱼粉组锦鲤肝胰脏SOD活力均显著高于鱼粉组(石洪玥等, 2019)；适量的脱脂黑水虻幼虫粉替代鱼粉对大黄鱼幼鱼的抗氧化能力有一定的增强作用，但是高比例的替代则可能引发机体的氧化应激反应(韩星星, 2019)。黑水虻幼虫的体内含有生物活性物质，如抗菌肽、甲壳素等，可以作为免疫增强剂提高动物机体非特异免疫能力和抗氧化能力(徐齐云等, 2012)。尽管本研究中各实验组SOD活力与MDA含量相比无显著影响，但与上述研究结果类似的是，各实验组大菱鲆幼鱼肝脏SOD活力随着替代比例的增加，呈先升高后降低的趋势，MDA含量也呈此趋势。究其原因，可能是高水平的替代加快了大菱鲆幼鱼正常的代谢，从而产生了过量的O2–，机体处于一种氧化应激状态，所以使MDA含量有所增加，为清除过量MDA过氧化产物，SOD及相关酶活性也相应增强。

鱼类的体色在评价鱼类品质等方面不可或缺，体色不仅能反应外界环境对鱼体的影响，也能代表其健康状态，体色异常则意味着品质和价格的降低(Breithaupt, 2007)。鱼类的体色一方面直接受到饲料源中色素物质的影响，另一方面也会间接受到外界环境、饲料营养与机体健康状况的影响。本研究发现，全脂黑水虻幼虫粉替代鱼粉比例达到40%，大菱鲆幼鱼背部体色亮度减弱。Hearing(2005)研究表明，体表颜色较暗是因为其中含有较多的黑色素。而酪氨酸酶作为限速酶控制着黑色素合成，酪氨酸酶活力与黑色素的含量呈正比关系(Koga et al, 1999)。本研究未对大菱鲆幼鱼体内酪氨酸酶的活力进行测定，推测黑水虻幼虫粉在饲料中添加比例较高时，影响了酪氨酸酶活力。类胡萝卜素是鱼体红色素细胞和黄色素细胞的成色物质之一，黑水虻幼虫粉替代鱼粉比例超过40%时，大菱鲆幼鱼背部的红度和黄度增强，原因可能为黑水虻幼虫中含有大菱鲆幼鱼能够代谢的类胡萝卜素，对鱼体呈色起到了一定作用，类胡萝卜素的着色效果受饲料中脂肪的添加量和来源的影响(刘晓东等, 2018)，也可能是饲料脂肪酸改变引起了此变化。目前，蛋白替代对鱼类体色的影响研究较少，本研究中，大菱鲆体色改变的内在机制还需今后进一步探讨。

4 结论

本研究条件下，全脂黑水虻幼虫粉可以替代20%的鱼粉蛋白而不影响大菱鲆幼鱼生长及健康状态，因此，全脂黑水虻幼虫粉在大菱鲆幼鱼饲料中替代鱼粉蛋白的推荐量为20%。

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