Application of acebutolol hydrochloride as or in preparation of preparation for relieving acute stress of pomfret
By adding acetoluol hydrochloride to the silver pompeo feed, the problem of central muscle injury in the silver pompeo stress response was solved, and the effect of significantly reducing mortality and improving anti-stress ability was achieved.
Patent Information
- Application Number
- CN202510241536.0
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-03-03
- Publication Date
- 2025-05-30
AI Technical Summary
The prior art is difficult to effectively alleviate myocardial damage in stress responses, and there is a lack of effective drugs or treatment options for cardiac protection in fish.
By adding acetobutol hydrochloride to the silver pomfen feed, it uses its properties as an adrenaline β1 receptor antagonist to relieve myocardial damage caused by the silver pomfen stress response.
It significantly alleviates myocardial injury under the stress state of silver pompa, reduces the mortality rate of fish, improves anti-stress ability, and delays myocardial injury by reducing the apoptosis rate of cardiomyocytes.
Smart Images
Figure CN120052471A_ABST
Abstract
Description
Technical Field
[0001] The present invention relates to the technical field of fish physiology, and particularly relates to the application of acebutolol hydrochloride in the preparation of a preparation for alleviating acute stress of silver pomfret or as such a preparation. Background Art
[0002] Silver pomfret is an important economic fish in the East China Sea, but it has a super strong stress response, even reaching the level of "dying immediately out of water". At present, the preparations for silver pomfret stress mainly focus on reducing oxidative stress, such as the antioxidant effects of vitamin C, plant extracts, etc. However, there is currently a technical blank in improving the heart damage of fish caused by stress and formulating corresponding stress alleviation techniques. In particular, there are still large blanks in the exploration of stress response alleviation and myocardial protection of silver pomfret.
[0003] Although existing studies have shown that excessive activation of the sympathetic nervous system and surges in catecholamines can trigger various heart diseases, and certain curative effects have been achieved through methods such as sympathetic nervous system inhibitory drugs and adrenergic receptor blockers, most of these treatment methods are limited to the research of mammalian models or human diseases, and the research on the stress response and myocardial protection mechanism of aquatic animals, especially fish such as silver pomfret, is still insufficient. Therefore, there is currently a lack of effective drugs or treatment regimens for fish heart protection, especially for heart damage and dysfunction caused by stress. To address this problem, it is particularly important to develop drugs that can effectively alleviate the stress response of silver pomfret and have a myocardial protection effect. Summary of the Invention
[0004] The purpose of the present invention is to provide an application that can alleviate myocardial damage caused by stress response during the breeding process of silver pomfret by adding acebutolol hydrochloride, improve its stress resistance, reduce myocardial cell death, thereby reducing the mortality rate of fish and enhancing the growth and health level.
[0005] To achieve the above purpose, the specific technical solutions adopted by the present invention are as follows:
[0006] In the first aspect, the present invention provides the application of acebutolol hydrochloride in the preparation of a preparation for alleviating acute stress of silver pomfret or as such a preparation.
[0007] Preferably, the preparation for alleviating acute stress of silver pomfret includes an active ingredient and a carrier. The active ingredient is acebutolol hydrochloride, and the carrier is silver pomfret feed, which can be commercially available (such as the fish feed produced by Hayashi Kane Industry Co., Ltd. of Japan) or self-made feed suitable for silver pomfret. By adding acebutolol hydrochloride to the silver pomfret feed and then feeding the silver pomfret, it can effectively alleviate myocardial damage of silver pomfret in a stress state, reduce its mortality rate after stress, and enhance its stress resistance.
[0008] Preferably, the preparation method of the preparation for alleviating the acute stress of silver pomfret is as follows: Dissolve acebutolol hydrochloride powder in absolute ethanol, and then evenly spray the solution containing acebutolol hydrochloride on the silver pomfret feed, and dry it for standby. Further, the concentration of acebutolol hydrochloride on the silver pomfret feed reaches 40 mg / kg.
[0009] Preferably, the usage method of the preparation for alleviating the acute stress of silver pomfret is as follows: Feed the silver pomfret with the feed added with acebutolol hydrochloride, and feed it regularly and quantitatively every day. Further, the feeding amount of the feed is calculated according to the weight of the fish to ensure that the daily food intake is 2%-3% of the fish weight; Feed it twice a day, respectively in the morning and evening, and continuously feed for 14 days.
[0010] In a second aspect, the present invention provides a preparation for alleviating the acute stress of silver pomfret, which comprises an active ingredient and a carrier. The active ingredient is acebutolol hydrochloride, and the carrier is silver pomfret feed.
[0011] The present invention has the following beneficial effects:
[0012] 1. The present invention innovatively proposes the application of acebutolol hydrochloride as a myocardial protectant. As an adrenergic β1 receptor antagonist, it can significantly alleviate myocardial damage in silver pomfret under stress conditions, providing a new and effective solution for dealing with myocardial damage in fish after stress.
[0013] 2. The present invention successfully regulates the stress response of silver pomfret by adding acebutolol hydrochloride to the silver pomfret feed, reduces the negative impact of stress on the heart, thereby improving the stress resistance of fish, reducing the mortality rate, and improving the growth and health level.
[0014] 3. The present invention proves through TUNEL experiments that acebutolol hydrochloride can effectively reduce the apoptosis rate of fish myocardial cells. This technical solution provides a reliable way to reduce the mortality rate of fish myocardial cells and delay myocardial damage.
[0015] 4. The technical solution of the present invention has high operability and practical application value through a simple feed addition method. Breeders only need to add acebutolol hydrochloride to the feed in a certain proportion to effectively improve the health level of silver pomfret, enhance the stress resistance, and is suitable for large-scale aquaculture.
[0016] 5. The present invention not only helps to reduce the health problems of silver pomfret under stress response and reduce the mortality rate, but also reduces environmental pollution in aquaculture by reducing drug use, improves the aquaculture efficiency, and has high social and economic value.
[0017] 6. The technical solution of the present invention is applicable to different species of fish, especially marine fish that are easily affected by environmental changes, and has broad application prospects. By standardizing the application of acebutolol hydrochloride, the universal promotion of fish stress management can be achieved. Description of the Drawings
[0018] Figure 1 : HE staining map of the heart section of the control group after stress in Example 1. In the figure, a. HE staining map of the ventricle of silver pomfret without stress; b. HE staining map of the ventricle of silver pomfret after 0.5 h of stress; c. HE staining map of the ventricle of silver pomfret after 12 h of stress; d. HE staining map of the ventricle of silver pomfret after 24 h of stress.
[0019] Figure 2 : HE staining map of the heart section of the experimental group after stress in Example 1. a. HE staining map of the ventricle of silver pomfret without stress; b. HE staining map of the ventricle of silver pomfret after 0.5 h of stress with drug administration; c. HE staining map of the ventricle of silver pomfret after 12 h of stress with drug administration; d. HE staining map of the ventricle of silver pomfret after 24 h of stress with drug administration.
[0020] Note: Figure 1-2 The area indicated by the arrow in is the myocardial injury area, which is used to indicate the injury of the ventricular myocardial tissue of silver pomfret at different stress time points.
[0021] Figure 3 : TUNEL map of the heart section of the control group after stress in Example 1. In the figure, a. DAPI staining of cell nuclei; b. Green fluorescence labeling of apoptotic cells; c. Composite map of DAPI and green fluorescence.
[0022] Figure 4 : TUNEL map of the heart section of the experimental group after stress in Example 1. In the figure, a. DAPI staining of cell nuclei; b. Green fluorescence labeling of apoptotic cells; c. Composite map of DAPI and green fluorescence.
[0023] Figure 5 : Statistical chart of the difference in mortality between the control group and the experimental group in Example 1. Detailed Embodiments
[0024] The present invention will be further described below in conjunction with the drawings and specific embodiments.
[0025] Example 1: Application of acebutolol hydrochloride in silver pomfret feed and its effect evaluation
[0026] 1. Temporary rearing of fish
[0027] Select healthy silver pomfret fry with similar weights and randomly divide them into eight groups. Allocate the experimental fish into eight fish tanks with the same environmental conditions, ensuring that the temperature, pH value, dissolved oxygen, water quality, etc. in each fish tank are consistent. Among the eight fish tanks, four are control groups and four are experimental groups. During the acclimation period, all fish in the fish tanks are fed with standard feed to ensure that they are fully adapted to the environment without drug interference for two weeks. During the acclimation process, each fish tank is equipped with a lid to avoid interference from the external environment and ensure stable experimental conditions.
[0028] 2. Feed preparation
[0029] Dissolve acebutolol hydrochloride powder in absolute ethanol to ensure complete dissolution. Then use a spray bottle to evenly spray the solution containing acebutolol hydrochloride on the feed (in this embodiment, the feed is specifically the fish feed produced by Hayashi-Kan Sangyo Co., Ltd. of Japan), ensuring that the drug concentration on the feed reaches 40 mg / kg. No drugs are added to the feed of the control group. After the feed is naturally air-dried, store it in a refrigerator at 4°C for subsequent use.
[0030] 3. Feed feeding
[0031] The acclimation period is 14 days. After 14 days, the experiment officially begins. The control group continues to be fed with conventional feed, while the experimental group is fed with feed added with acebutolol hydrochloride. The feeding amount of the feed needs to be calculated according to the weight of the fish to ensure that the daily food intake is 2%-3% of the fish weight. To ensure the freshness and effectiveness of the feed, the feed needs to be fed twice a day, in the morning and evening respectively, and fed regularly and quantitatively every day for 14 days. All feeding activities are carried out carefully to avoid stress reactions in the fish caused by human operation.
[0032] 4. Water environment management
[0033] To ensure the consistency and stability of the experimental environment for all fish, the water quality, temperature, dissolved oxygen, and pH value of all fish tanks need to be strictly monitored and kept stable during the experiment. The pH value of the water body should be controlled between 7.5 and 8.5, the dissolved oxygen content should be kept above 10 mg / L, and the water temperature should be maintained at 21±1°C. Regularly treat pollutants to avoid the accumulation of pollutants affecting the experimental results.
[0034] 5. Fish stress treatment and sampling
[0035] ① Stress treatment: On the last day of the experiment, all fish are subjected to fishing stress treatment. Use a fishing net to lift the fish out of the water for 10 s and then put them back into the water for 20 s, repeating this three times, and finally put them back into the water to simulate the fishing stress that may be encountered in aquaculture.
[0036] ② Sampling:
[0037] Anesthesia treatment of fish after stress: Before sampling, prepare an appropriate amount of eugenol solution (usually with a concentration of 100 - 200 mg / L), add it to an appropriate amount of water to ensure that the concentration is sufficient to quickly put the fish into an anesthetic state. Gently place the fish into the water containing eugenol, ensuring that the fish remains stationary during the anesthesia process. Observe its anesthetic reaction, and immediately proceed to the next step after it loses the ability to swim and shows an anesthetic state. Avoid over-anesthetizing and causing tissue damage.
[0038] Fish body dissection and heart sampling: Use sterile scissors or scalpels to cut open the abdomen of the fish to expose the internal organs. Use sterile forceps to remove the fish heart, ensuring that it is not under external pressure or damaged.
[0039] Heart preservation method: Immediately place the removed heart into a test tube containing 1 ml of 4% paraformaldehyde fixative. If long-term preservation or transportation to a laboratory is required, dry ice preservation can be selected. At this time, place the removed heart into a sterile empty tube, ensure it is completely sealed, and immediately place it in dry ice for freezing to avoid sample deterioration.
[0040] Sampling time and grouping:
[0041] Control group (fed with conventional feed): Sampling was carried out at the time points of before stress, 0.5 hour, 12 hours, and 24 hours after stress respectively. At least three fish samples were used for the experiment at each time point to increase the reliability of the data.
[0042] Experimental group (fed with feed added with acebutolol hydrochloride): The same operations were also carried out to ensure consistent sampling conditions for later comparison.
[0043] 6. Tissue processing
[0044] Place the heart tissue in paraformaldehyde for fixation for two weeks, and then start dehydration, clearing, embedding, and sectioning. Perform HE staining on the sections to observe the morphological changes of the myocardial tissue. Use a TUNEL kit to detect apoptotic cells in the heart sections, operate according to the instructions, and observe and record the number of apoptotic cells.
[0045] 7. Evaluation of stress response
[0046] Mortality rate evaluation: During the entire experiment, regularly check the survival status of the fish and record the mortality rates of different groups at different time points. By statistically analyzing the differences in mortality rates between the experimental group and the control group, further verify the alleviating effect of the drug on the stress response of fish.
[0047] 8. Result analysis
[0048] Results of HE staining: Figure 1The results of the control group showed that the heart sections of non-stressed silver pomfrets fed with conventional feed were structurally intact (a), while the heart sections of stressed silver pomfrets (b-d) showed obvious signs of damage such as broken myocardial fibers and enlarged cell gaps; and Figure 2 The results of the experimental group showed that the heart sections of non-stressed silver pomfrets (a) and stressed silver pomfrets (b-d) fed with feed supplemented with acebutolol hydrochloride were relatively intact, with neatly arranged myocardial fibers and significantly reduced damage.
[0049] TUNEL results: Figure 3 It showed that there were a large number of TUNEL-positive cells (green fluorescence) in the heart sections of the control group fed with conventional feed after stress, indicating significant apoptosis of myocardial cells; and Figure 4 It showed that the number of TUNEL-positive cells in the heart sections of the experimental group fed with feed supplemented with acebutolol hydrochloride was significantly reduced, and the apoptosis rate was significantly decreased.
[0050] Mortality assessment results: Figure 5 It showed that after the experimental group was fed with feed containing acebutolol hydrochloride, its mortality was significantly lower than that of the control group.
[0051] This specific implementation manner is only an interpretation of the present invention and not a limitation thereof. Any changes made by those skilled in the art after reading the specification of the present invention will be protected by the patent law as long as they are within the scope of the claims of the present invention.
Claims
1. Use of acebutolol hydrochloride as or in the preparation of a preparation for relieving acute stress in silver pomfret.
2. The use according to claim 1, characterized in that: The preparation for alleviating acute stress of silver pomfret comprises an active ingredient and a carrier, wherein the active ingredient is acebutolol hydrochloride, and the carrier is silver pomfret feed.
3. The use according to claim 2, characterized in that: The preparation method of the preparation for relieving acute stress of silver pomfret is as follows: dissolving acebutolol hydrochloride powder in anhydrous ethanol, then spraying the solution containing acebutolol hydrochloride evenly on silver pomfret feed, and drying for later use.
4. The use according to claim 3, characterized in that: The concentration of acebutolol hydrochloride in the silver pomfret feed reaches 40 mg / kg.
5. The use according to any one of claims 2 to 4, characterized in that: The method for using the preparation for relieving acute stress of silver pomfret is as follows: feed added with acebutolol hydrochloride is fed to the silver pomfret at regular intervals and in fixed quantities every day.
6. The use according to claim 5, characterized in that: The amount of feed to be fed is calculated based on the weight of the fish, ensuring that the daily feed intake is 2%-3% of the fish's body weight.
7. The use according to claim 5, characterized in that: Feed was given twice a day, in the morning and evening, for 14 days.
8. A preparation for relieving acute stress of silver pomfret, characterized in that: The invention comprises an active ingredient and a carrier, wherein the active ingredient is acebutolol hydrochloride and the carrier is silver pomfret feed.