黄条仔稚幼鱼消化酶活性变化研究*

张正荣 柳学周 于 毅 史 宝 张言祥 徐永江 王 滨 姜 燕 孙冉冉

张正荣1,2柳学周1,2①于 毅3史 宝2张言祥3徐永江2王 滨2姜 燕2孙冉冉2

(1. 上海海洋大学水产与生命学院 上海 201306；2. 中国水产科学研究院黄海水产研究所 农业农村部海洋渔业可持续发展重点实验室 青岛海洋科学与技术试点国家实验室海洋渔业科学与食物产出过程功能实验室 青岛 266071；3. 大连富谷水产有限公司 大连 116400)

在鱼类规模化育苗生产中，鱼苗死亡率较高。Lauff等(1984)研究发现，消化酶是影响仔稚幼鱼死亡率的重要因素。消化酶活力高低直接影响鱼类的消化吸收，间接影响鱼类的生长发育。因此，对仔稚幼鱼消化酶活性的研究利于了解其消化生理特征，也有利于探索早期发育中鱼苗大量死亡的原因。目前，研究的鱼类消化酶主要包括脂肪酶、淀粉酶、胰蛋白酶和碱性磷酸酶等，脂肪酶主要由鱼类的肝胰脏分泌，起分解脂肪的作用；胰蛋白酶主要由胰脏分泌，鱼类早期发育蛋白消化主要靠胰蛋白酶完成；鱼类各消化器官都有淀粉酶存在，分解碳水化合物；碱性磷酸酶主要存在于鱼类肠道上皮细胞，与葡萄糖、Ca和P等的吸收有关，这些消化酶在鱼类早期发育过程的消化生理中起重要作用。目前，对仔稚幼鱼消化酶的研究日益增多，国内外学者对大菱鲆()、半滑舌鳎()、金头鲷()、古巴雀鳝()和日本黄姑鱼()等仔稚幼鱼消化酶的分泌规律已有研究(陈慕雁等, 2005; 常青等, 2005; Moyano, 1996; Comabella, 2006; 孙敏等, 2012)。

1 材料与方法

1.1 实验材料

1.2 实验方法

1.2.2 消化酶活性测定 使用碧云天的BCA蛋白浓度测定试剂盒，测量酶液中的蛋白浓度。脂肪酶用南京建成的脂肪酶试剂盒测定，活力单位定义：在37℃条件下，每毫升酶液在反应体系中与底物反应 1 min，消耗1 mmol底物为1个酶活力单位。淀粉酶用南京建成淀粉酶试剂盒测定，其活力单位定义：组织中每毫克蛋白37℃、pH 7.0条件下与底物作用 30 min，水解10 mg淀粉定义为1个淀粉酶活力单位。胰蛋白酶用南京建成胰蛋白酶试剂盒测定，其活力单位定义：在37℃、pH 8.0条件下，每毫克蛋白中含有的胰蛋白酶，每分钟使吸光度变化0.003，即为 1个酶活力单位。碱性磷酸酶用南京建成碱性磷酸酶试剂盒测定，其活力单位定义：在37℃条件下，每克组织蛋白与基质作用15 min产生1 mg酚为1个酶活力单位。

1.3 数据处理

采用SPSS 22.0软件对实验数据进行分析，运用单因子方差分析(One-way ANOVA)和Duncan氏检验法对各组数据进行显著性差异分析和多重比较，显著性水平为0.05，所有数值均采用平均值±标准差(Mean±SD)表示。

全长的平均日增长计算公式：

G=(2–1)/(2–1)

用指数函数分析全长和日龄的关系，公式：

=aebt

式中，为全长(mm)，为日龄(d)，、为常数。

2 结果

2.1 黄条仔稚幼鱼的全长

=3.95e0.0358t，R=0.9788

孵化后60 d内的仔稚幼鱼全长随着日龄的增加而加快增长，25 d后全长大幅增加。

图1 黄条仔稚幼鱼生长曲线和饵料投喂

2.2 黄条胚胎、仔鱼消化酶比活力变化

图2 黄条胚胎、仔鱼脂肪酶的比活力变化

胚胎阶段记为孵化后0 d。不同小写字母表示消化酶比活力存在显著差异(<0.05,=3)，下同

The embryonic stage is recorded as 0 d after hatching. Different lowercase letters mean significant differences in specific activities of digestive enzymes (<0.05,=3), the same as below

图3 黄条胚胎、仔鱼淀粉酶的比活力变化

图4 黄条胚胎、仔鱼胰蛋白酶的比活力变化

Fig.5 Specific activities of alkaline phosphatase during embryo and larval development of

2.3 黄条稚幼鱼消化酶比活力变化

图6 黄条稚幼鱼脂肪酶的比活力变化

图7 黄条稚幼鱼淀粉酶的比活力变化

图8 黄条稚幼鱼胰蛋白酶的比活力变化

图9 黄条稚幼鱼碱性磷酸酶的比活力变化

2.4 不同发育阶段4种消化酶比活力值的特征

图10 不同发育阶段黄条仔稚幼鱼消化酶的比活力变化

不同小写字母表示同一日龄不同消化酶存在差异显著

Different lowercase letters mean significant differences at same age in different digestive enzymes

3 讨论

3.1 早期阶段消化酶的出现

3.2 黄条仔稚幼鱼4种消化酶活力的变化

3.3 不同发育阶段4种消化酶比活力值特征分析

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The Variations of Digestive Enzymes in Larval and Juvenile

ZHANG Zhengrong1,2, LIU Xuezhou1,2①, YU Yi3, SHI Bao2, ZHANG Yanxiang3, XU Yongjiang2, WANG Bin2, JIANG Yan2, SUN Ranran2

(1. College of Fisheries and Life Science, Shanghai Ocean University, Shanghai 201306; 2. Yellow Sea Fisheries Research Institute, Chinese Academy of Fishery Sciences, Key Laboratory of Sustainable Development of Marine Fisheries, Ministry of Agriculture and Rural Affairs, Laboratory for Marine Fisheries Science and Food Production Processes, Pilot National Laboratory for Marine Science and Technology (Qingdao), Qingdao 266071; 3. Dalian Fugu Fishery Co., Ltd., Dalian 116400)

To clarify the digestive and physiological characteristics in the early developmental stage of, the activity of different digestive enzymes (lipase, amylase, trypsin, and alkaline phosphatase) were studiedFertilized eggs and 1- to 60-day-old larvae and juveniles ofwere selected as the research objects. The relationship between total length and days after hatching was also studied. The results indicated that the total length of larvae and juveniles increased with age within 60 days after hatching and increased significantly after 25 days. Lipase, amylase, and alkaline phosphatase were detectable during the embryonic stage while trypsin was detected in newly hatched larvae, which indicated thatcan digest lipid, carbohydrate, and proteins before feeding. The specific activities of lipase and alkaline phosphatase increased rapidly after hatching (<0.05), reached a peak at (5.88±0.45) and (5.56±0.41) U/mg, respectively at first feeding. The amylase activity reached its peak at (1.59±0.02) U/mg on day 7. The specific activity of trypsin increased slowly after hatching, and reached the highest value ((1098.67±24.03) U/mg on day 15 (<0.05). Lipase, alkaline phosphatase, and trypsin were basically stable in the visceral mass of juvenile fish. However, theactivity of lipase, alkaline phosphatase, and trypsin increased with the development ofThe amylase activity in the visceral mass of juvenile fish showed a downward trend and was basically stable at a lower specific activity level. The results of this study showed significant changes in various digestive enzyme activities during the development of larvae and juveniles, which were closely related to the development stage and feeding habits. The digestive enzymes were detected in early larvae that had not yet eaten food which was considered that the digestive enzymes were maternal in origin and not from exogenous food. The lipase, alkaline phosphatase, and trypsin activities in juveniles significantly increased, which reflect the improvement of intestinal structure and digestive function and the increasing demand for fat and protein.

; Embryo; larvae and juvenile; digestive enzymes; Specific activities

S965.399

A

2095-9869(2020)02-0061-08

柳学周，研究员，E-mail: liuxz@ysfri.ac.cn

2018-12-11,

2019-02-14

*中国水产科学研究院中央级公益性科研院所基本科研业务费专项资金(2018GH17; 2019GH15; 2019CY0204)、青岛市民生科技计划项目(17-3-3-61-nsh)、国家重点研发计划项目(2018YFD0901204; 2019YFD0900901)、青岛海洋科学与技术国家实验室海洋渔业科学与食物产出过程功能实验室开放课题(2017-3A01)和现代农业产业技术体系专项经费(CARS-47)共同资助[This work was supported by Special Scientific Research Funds for Central Non-Profit Institutes, Chinese Academy of Fishery Sciences (2018GH17; 2019GH15; 2019CY0204), Qingdao People's Livelihood Science and Technology Project(17-3-3-61-nsh), National Key Research and Development Program(2018YFD0901204; 2019YFD0900901), Opening Topic of Qingdao National Laboratory of Marine Science and Technology Functional Laboratory of Marine Fisheries Science and Food Output Process(2017-3A01), and China Agriculture Research System (CRAS-47)]. 张正荣, E-mail: 997197691@qq.com

10.19663/j.issn2095-9869.20181211002

http://www.yykxjz.cn/

Zhang ZR, Liu XZ, Yu Y, Shi B, Zhang YX, Xu YJ, Wang B, Jiang Y, Sun RR. The variations of digestive enzymes in larval and juvenile. Progress in Fishery Sciences, 2020, 41(2): 61–68

LIU Xuezhou, E-mail: liuxz@ysfri.ac.cn

(编辑 陈 辉)