A compound functional additive for relieving oxidative stress and endoplasmic reticulum stress of protocobitis tiemanni
By using a compound functional additive composed of sodium caprylate, sodium butyrate, and rice husk powder, the problems of liver oxidative stress and endoplasmic reticulum stress in mandarin fish were solved, the antioxidant capacity of mandarin fish was improved, and the health of fish and ecological safety were promoted.
Patent Information
- Application Number
- CN202311811545.6
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- Filing Date
- 2023-12-26
- Publication Date
- 2026-02-27
- Estimated Expiration
- 2043-12-26
AI Technical Summary
In existing technologies, mandarin fish that have been fed artificial formulated feed suffer from severe oxidative stress and endoplasmic reticulum stress in the liver, leading to liver cell damage and affecting the fish's health.
A compound functional additive is provided, consisting of sodium octanoate, sodium butyrate and rice husk powder, for use in feed preparation. Through simple mixing, it can alleviate liver oxidative stress and endoplasmic reticulum stress in mandarin fish.
It effectively enhances the antioxidant capacity of mandarin fish, reduces liver oxidative damage, improves fish health, and is suitable for ecological and healthy aquaculture, reducing environmental pollution.
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Abstract
Description
TECHNICAL FIELD
[0001] The present application belongs to the technical field of feed additives, and particularly relates to a compound functional additive for relieving liver oxidative stress and endoplasmic reticulum stress of mandarin fish. BACKGROUND
[0002] Mandarin fish is a freshwater rare meat fish originally from China. Mandarin fish is rich in nutrition and delicious, with high content of amino acids and unsaturated fatty acids. The fish meat is snow-white, compact, elastic, in the shape of garlic cloves, and without intermuscular spines, and is known as the "King of Freshwater Fish". The total production of mandarin fish in China in 2021 was 374,000 tons. Mandarin fish is one of the most potential special breeding varieties in China.
[0003] In recent years, with the gradual maturity of mandarin fish seed domestication technology, mandarin fish has made some breakthroughs in feeding with compound feed. Although there are many successful cases of mandarin fish feeding with artificial compound feed, the majority of mandarin fish still fail to be bred, and the main reason is that feeding with artificial compound feed causes excessive burden on the liver of mandarin fish, leading to intensified liver oxidative stress and endoplasmic reticulum stress, and other adverse problems. Imbalance of oxidation and antioxidant homeostasis in the liver leads to the inability of enzyme antioxidant system (superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), and catalase (CAT)) and non-enzyme antioxidant system (vitamin C / E, alpha-lipoic acid, carotenoids, etc.) to remove excess free radicals (reactive oxygen species (ROS) (O 2- , H2O2, and hydroxyl radicals, etc.) and reactive nitrogen species (RNS) (NO, NO2, and ONOO - , etc.), resulting in a large accumulation of free radicals, inducing oxidative stress, and having a toxic effect on liver cells. When the body is under too strong or too long endoplasmic reticulum stress, a large number of unfolded and misfolded proteins are retained in liver cells, which exceeds the self-regulation capacity of the cells, causing cell metabolic disorder and apoptosis, and endangering the health of fish.
[0004] Therefore, it is urgent to develop functional additives to relieve liver oxidative stress and endoplasmic reticulum stress caused by artificial compound feed. SUMMARY
[0005] The present application aims to at least solve one of the above-mentioned status and technical problems in the prior art, and provides a compound functional additive and its application in preparing a product for relieving liver oxidative stress and endoplasmic reticulum stress of mandarin fish. The compound functional additive can effectively improve the oxidative stress resistance of mandarin fish and relieve liver endoplasmic reticulum stress.
[0006] The first aspect of the present application aims to provide an additive.
[0007] The second aspect of the present application aims to provide use of the additive of the first aspect of the present application in the preparation of a feed.
[0008] The third aspect of the present application aims to provide a feed.
[0009] The fourth aspect of the present application aims to provide use of the additive of the first aspect of the present application or the feed of the third aspect of the present application in the preparation of a product for alleviating oxidative stress and / or endoplasmic reticulum stress in fish.
[0010] To achieve the above-mentioned objects, the technical solutions adopted by the present application are as follows:
[0011] The first aspect of the present application provides an additive comprising the following components: sodium octanoate, sodium butyrate and rice husk powder.
[0012] In some embodiments of the present application, the additive comprises the following components in mass fraction: 2-50 parts of sodium octanoate, 2-50 parts of sodium butyrate and 10-100 parts of rice husk powder.
[0013] In some preferred embodiments of the present application, the additive comprises the following components in mass fraction: 5-40 parts of sodium octanoate, 5-40 parts of sodium butyrate and 20-90 parts of rice husk powder.
[0014] In some more preferred embodiments of the present application, the additive comprises the following components in mass fraction: 5-20 parts of sodium octanoate, 5-20 parts of sodium butyrate and 60-90 parts of rice husk powder.
[0015] In some most preferred embodiments of the present application, the additive comprises the following components in mass fraction: 10 parts of sodium octanoate, 10 parts of sodium butyrate and 80 parts of rice husk powder.
[0016] In some embodiments of the present application, the preparation method of the additive comprises simply mixing the components.
[0017] The second aspect of the present application provides use of the additive of the first aspect of the present application in the preparation of a feed.
[0018] In some embodiments of the present application, the feed is an aquaculture feed.
[0019] In some embodiments of the present application, the aquatic product includes but is not limited to fish, shrimp and the like.
[0020] In some embodiments of the present application, the fish includes mandarin fish.
[0021] The third aspect of the present application provides a feed comprising the additive of the first aspect of the present application.
[0022] In some embodiments of the application, the feed comprises the following mass parts of the mass raw materials: 95-100 parts of the basic feed and 1-5 parts of the additive.
[0023] In some preferred embodiments of the application, the feed comprises the following mass parts of the mass raw materials: 95-98 parts of the basic feed and 1-3 parts of the additive.
[0024] In some more preferred embodiments of the application, the feed comprises the following mass parts of the mass raw materials: 98 parts of the basic feed and 2 parts of the additive.
[0025] In some embodiments of the application, the protein source, the oil, the mineral and the vitamins.
[0026] In some embodiments of the application, the protein source comprises white fish meal, krill meal and corn gluten meal.
[0027] In some embodiments of the application, the oil comprises fish oil and soybean oil.
[0028] In some embodiments of the application, the vitamins comprise vitamin Bl, vitamin B2, vitamin B6, vitamin B 12 3, vitamin E, vitamin K, folic acid, biotin, inositol, vitamin C, niacinamide and choline.
[0029] In some embodiments of the application, vitamin Bl is 25 mg / kg, vitamin B2 is 15 mg / kg, vitamin B6 is 30 mg / kg, vitamin B 12 0.1 mg / kg, vitamin D3 is 5 mg / kg, vitamin E is 200 mg / kg, vitamin K is 15 mg / kg, folic acid is 20 mg / kg, biotin is 60 mg / kg, inositol is 600 mg / kg, vitamin C is 250 mg / kg, niacinamide is 200 mg / kg and choline is 1000 mg / kg.
[0030] In some embodiments of the application, the mineral comprises MnSO4, MgSO4, FeSO4, CoSO4, Na2SeO3, CuSO4, KCl, NaCl, CuSO4, ZnSO4, H2CaIO4 and CoCl2.
[0031] In some embodiments of the present application, the minerals include MnSO4 45mg / kg, MgSO4 800mg / kg, FeSO4 100mg / kg, CoSO4 0.02mg / kg, Na2SeO3 1mg / kg, CuSO4 10mg / kg, KCl 3000mg / kg, NaCl 1500mg / kg, CuSO4 10mg / kg, ZnSO4 50mg / kg, H2CaIO4 50mg / kg, CoCl2 50mg / kg.
[0032] In some embodiments of the present application, the method for preparing the aquaculture feed comprises the step of mixing the additive with the base feed.
[0033] In a fourth aspect of the present application, the additive of the first aspect of the present application or the feed of the third aspect of the present application is used in (1) or (2):
[0034] (1) preparing a product for alleviating oxidative stress and / or endoplasmic reticulum stress in the liver of fish;
[0035] (2) alleviating oxidative stress and / or endoplasmic reticulum stress in the liver of fish.
[0036] In some embodiments of the present application, the fish includes Siniperca duensis.
[0037] The present application has the following beneficial effects:
[0038] The additive provided by the present application can effectively alleviate oxidative stress and endoplasmic reticulum stress in the liver of Siniperca duensis, reduce the content of malondialdehyde, improve the ability of antioxidant enzyme activity of Siniperca duensis, and is beneficial to the health of fish.
[0039] The additive provided by the present application does not contain antibiotics, is suitable for ecological and healthy breeding, can promote the safety of aquaculture, and reduce environmental pollution.
[0040] The feed provided by the present application has stable raw material sources and low price, and the production process of the feed is simple and convenient to use. BRIEF DESCRIPTION OF DRAWINGS
[0041] Figure 1 The figure is a comparison chart of the effect of the composite functional additive on the malondialdehyde content in the liver tissue of Siniperca duensis, wherein the same letter represents no significant difference, and different letters represent significant difference.
[0042] Figure 2 The figure is a comparison chart of the effect of the composite functional additive on the total antioxidant capacity in the liver tissue of Siniperca duensis, wherein the same letter represents no significant difference, and different letters represent significant difference.
[0043] Figure 3Figure for the effect of the composite functional additive on the total superoxide dismutase activity of the liver tissue of the golden mandarin fish, in which the same letter represents no significant difference, and different letters represent significant difference.
[0044] Figure 4 Figure for the effect of the composite functional additive on the reduced glutathione content of the liver tissue of the golden mandarin fish, in which the same letter represents no significant difference, and different letters represent significant difference.
[0045] Figure 5 Figure for the effect of the composite functional additive on the catalase activity of the liver tissue of the golden mandarin fish, in which the same letter represents no significant difference, and different letters represent significant difference.
[0046] Figure 6 Figure for the effect of the composite functional additive on the antioxidant-related genes of the liver tissue of the golden mandarin fish, in which the same letter represents no significant difference, and different letters represent significant difference.
[0047] Figure 7 Figure for the effect of the composite functional additive on the endoplasmic reticulum stress-related genes of the liver tissue of the golden mandarin fish, in which the same letter represents no significant difference, and different letters represent significant difference. DETAILED DESCRIPTION
[0048] The content of the present application will be further described in detail through specific examples.
[0049] It should be understood that these examples are only used to illustrate the present application and not used to limit the scope of the present application.
[0050] In order to make the purpose, technical solutions and advantages of the embodiments of the present application more clear, the technical solutions in the embodiments of the present application will be clearly and completely described below. The specific conditions not mentioned in the embodiments are carried out according to the conventional conditions or the conditions suggested by the manufacturers. The reagents or instruments not mentioned by the manufacturers are all conventional products that can be purchased in the market.
[0051] The features and performances of the present application will be further described in detail below in combination with the examples.
[0052] Example 1
[0053] A composite functional additive for relieving the oxidative stress and endoplasmic reticulum stress of the liver of the golden mandarin fish, comprising the following components in parts by mass: sodium octanoate 5 parts, sodium butyrate 5 parts, and rice husk powder 90 parts.
[0054] Example 2
[0055] A composite functional additive for relieving liver oxidative stress and endoplasmic reticulum stress of Procyprinus, comprising the following components in parts by mass: sodium octanoate 10 parts, sodium butyrate 10 parts, and rice husk powder 80 parts.
[0056] Example 3
[0057] A composite functional additive for relieving liver oxidative stress and endoplasmic reticulum stress of Procyprinus, comprising the following components in parts by mass: sodium octanoate 20 parts, sodium butyrate 20 parts, and rice husk powder 60 parts.
[0058] Example 4
[0059] A composite functional additive for relieving liver oxidative stress and endoplasmic reticulum stress of Procyprinus, comprising the following components in parts by mass: sodium octanoate 40 parts, sodium butyrate 40 parts, and rice husk powder 20 parts.
[0060] Comparative Example 1
[0061] A composite functional additive, comprising the following components in parts by mass: sodium octanoate 5 parts and rice husk powder 95 parts.
[0062] Comparative Example 2
[0063] A composite functional additive, comprising the following components in parts by mass: sodium butyrate 5 parts and rice husk powder 95 parts.
[0064] Effect Example
[0065] This example investigates the effects of adding the composite functional additives of Examples 1-4 and Comparative Examples 1-2 on liver oxidative stress and endoplasmic reticulum stress of Procyprinus
[0066] Test fish and culture management:
[0067] The culture test was carried out at the experimental base of Guangdong Haide Group, and the test fish Procyprinus were purchased from a local fry farm in Qingyuan. The test fish were first temporarily raised for 2 weeks to adapt to the test conditions. Before formal culture, the test fish were starved for 24 h, and 1680 Procyprinus with uniform size and an average weight of (25.0 ± 0.1 g) were randomly selected and divided into 28 300L circulating water aquariums, randomly divided into 7 groups (Table 1), with 4 aquariums in each group and 60 fish in each aquarium. The corresponding feed (the feed was prepared by Procyprinus juvenile basic feed + the corresponding composite functional additive of the examples and comparative examples, wherein white fish meal, krill meal and corn gluten meal were the protein source, and fish oil and soybean oil were the oil source. The specific composition of the groups and the corresponding feed is shown in Table 1) was fed according to the grouping, twice a day (06:00 and 18:00). The culture test period was 8 weeks.
[0068] Table 1 Grouping and feed formula
[0069]
[0070] Table, multi-dimensional 1 : Vitamin B1 25mg / kg, Vitamin B2 15mg / kg, Vitamin B6 30mg / kg, Vitamin B 12 0.1mg / kg, Vitamin D3 5mg / kg, Vitamin E 200mg / kg, Vitamin K 15mg / kg, Folic acid 20mg / kg, Biotin 60mg / kg, Myo-inositol 600mg / kg, Vitamin C(35%) 250mg / kg, Nicotinamide 200mg / kg, Choline 1000mg / kg.
[0071] Multi-mineral 2 : MnSO4 45mg / kg, MgSO4 800mg / kg, FeSO4 100mg / kg, CoSO4 0.02mg / kg, Na2SeO3 1mg / kg, CuSO4 10mg / kg, KCl 3000mg / kg, NaCl 1500mg / kg, CuSO4 10mg / kg, ZnSO450mg / kg, H2CaIO4 50mg / kg, CoCl2 50mg / kg.
[0072] Sample collection and index analysis:
[0073] After the end of the breeding test, 24h starvation was carried out. 4 fish were taken from each aquarium, and the liver tissue was dissected into a frozen tube and immediately frozen in liquid nitrogen for later use in the detection of each index.
[0074] Malondialdehyde (MDA) content determination: according to the MDA kit (Nanjing Jiancheng Biological Engineering Research Co., Ltd., item number A003-1-2) instruction. The general process is as follows: take an appropriate amount of sample, add working solution, and water bath at 95℃ for 2h, and cool with running water. Centrifuge at 4000rpm for 10min. Then take the supernatant, and use the full-automatic enzyme marker to measure the OD value at 530nm wavelength.
[0075] Total superoxide dismutase (T-SOD): according to the T-SOD kit (Nanjing Jiancheng Biological Engineering Research Co., Ltd., item number A001-3-2) instruction. The general process is as follows: before the formal test, the appropriate sample concentration is determined, and the sample concentration with about 50% inhibition rate is selected. After diluting the sample to the appropriate concentration, take an appropriate amount of sample in the 96-well enzyme plate, add enzyme working solution and substrate application liquid, and react at 37℃ for 20min. Then use the full-automatic enzyme marker to measure the OD value at 450nm wavelength.
[0076] Catalase (CAT): Determined according to the CAT kit (Nanjing Jiancheng Bioengineering Institute, item number A007-1-1) instructions. Roughly the procedure is as follows: mix the appropriate amount of sample with reagent one, reagent two, under the condition of 37°C, react for 1 min, then add reagent three and reagent four, mix well, then use the full-automatic enzyme marker to determine the OD value at 405 nm wavelength.
[0077] Total antioxidant capacity (T-AOC): Determined according to the T-AOC kit (Nanjing Jiancheng Bioengineering Institute, item number A015-1-2) instructions. Roughly the procedure is as follows: take the appropriate amount of sample in the 96-well enzyme plate, add reagent four and the prepared ABTS working solution, under the condition of room temperature, react for 6 min, then use the full-automatic enzyme marker to determine the OD value at 405 nm wavelength.
[0078] Reduced glutathione (GSH) content: Determined according to the GSH kit (Nanjing Jiancheng Bioengineering Institute, item number A006-2-1) instructions. Roughly the procedure is as follows: first add 0.1 mL reagent one to the sample, under the condition of 4°C, 3500 rpm, centrifuge for 10 min, take the supernatant. Add reagent two and reagent three, stand at room temperature for 5 min, then use the full-automatic enzyme marker to determine the OD value at 405 nm wavelength.
[0079] The expression levels of antioxidant-related genes and endoplasmic reticulum stress-related genes were detected and analyzed by q-PCR. The specific genes included were: Nuclear factor erythroid2-related factor 2 (nrf2), Kelch-like ECH-associated protein 1 (keap1), superoxide dismutase (sod), glutathione peroxidase (gpx), catalase (cat), GRP78 immunoglobulin heavy chain-binding protein (bip), Inositol-requiring protein-1 (ire), Eukaryotic translation initiation factor 2-alpha kinase 3 (perk), X-box-binding protein 1 (xbp1), DNA-damage-inducible transcript 3 (chop), and Activating transcription factor 4 (atf4), with beta-actin (β-actin) as the internal reference gene. q-PCR primers were designed based on the nucleotide sequence of Mandarin fish (Table 2).
[0080] Table 2 q-PCR primer sequences
[0081]
[0082]
[0083] Experimental data were statistically analyzed using one-way ANOVA in SPSS 20.0. After significant differences were identified, Tukey's multiple comparisons were performed, with a significance level of P < 0.05. The experimental data are expressed as mean ± standard deviation (n = 4).
[0084] Experimental results:
[0085] Malondialdehyde content: such as Figure 1 As shown, the malondialdehyde (MDA) content of feeds 1, 2, 3, 4, 5, 6, and 7 were 2.63, 2.48, 2.58, 1.88, 1.70, 1.90, and 2.00 U / mg protein, respectively. The results indicate that compared to feeds 1-3, feeds 4, 5, 6, and 7 significantly reduced the MDA content in the liver tissue of juvenile mandarin fish. Adding appropriate amounts of the compound functional additives from Examples 1-4 can reduce oxidative damage to the liver tissue of juvenile mandarin fish and alleviate oxidative stress.
[0086] Total antioxidant capacity: such as Figure 2 As shown, the total antioxidant capacities of feeds 1, 2, 3, 4, 5, 6, and 7 were 1.31, 1.34, 1.30, 1.93, 1.93, 1.86, and 1.84 U / mg protein, respectively. The results indicate that compared to feeds 1-3, feeds 4, 5, 6, and 7 significantly improved the total antioxidant capacity of the liver tissue of juvenile mandarin fish, demonstrating that the addition of the compound functional additives from Examples 1-4 can significantly improve the total antioxidant capacity of the liver tissue of juvenile mandarin fish.
[0087] Total superoxide dismutase activity: such as Figure 3 As shown, the total superoxide dismutase activities of feeds 1, 2, 3, 4, 5, 6, and 7 were 59.75, 60.25, 60.50, 72.25, 76.50, 76.25, and 68.00 U / mg protein, respectively. The results indicate that, compared with feed 1, feeds 5 and 6 significantly increased the total superoxide dismutase activity in the liver tissue of juvenile mandarin fish.
[0088] Catalase activity: such as Figure 4 As shown, the catalase activities of feeds 1, 2, 3, 4, 5, 6, and 7 were 82.75, 81.75, 87.25, 103.50, 102.00, 88.50, and 85.00 U / mg protein, respectively. The results indicate that, compared with feeds 1–3, feeds 4 and 5 significantly increased the catalase activity in the liver tissue of juvenile mandarin fish.
[0089] Reduced glutathione content: such as Figure 5 As shown, the reduced glutathione contents of feeds 1, 2, 3, 4, 5, 6, and 7 were 4.98, 4.83, 5.08, 5.68, 6.65, 6.80, and 6.20 U / mg protein, respectively. The results indicate that compared with feeds 1–3, feeds 4, 5, 6, and 7 significantly increased the reduced glutathione content in the liver tissue of juvenile mandarin fish.
[0090] Expression of antioxidant-related genes: such as Figure 6 As shown, compared with feed 1, there were no significant differences in the expression of nrf2 and keap1 genes in feed groups 2, 3, 4, 5, 6 and 7. The expression of sod gene was significantly increased in feed groups 4, 5 and 6, the expression of gpx gene was significantly increased in feed groups 4, 5 and 6, and the expression of cat gene was significantly increased in feed groups 4, 5 and 6.
[0091] Endoplasmic reticulum stress-related gene expression: such asFigure 7 Compared with feed 1, the expression of ire, perk and atf4 genes in feed 2, feed 3, feed 4, feed 5, feed 6 and feed 7 groups had no significant difference, the expression of bip and xbp1 genes in feed 4, feed 5, feed 6 and feed 7 groups was significantly reduced, and the expression of chop gene in feed 4, feed 5 and feed 6 groups was significantly reduced.
[0092] The above results can draw the conclusion that the complex functional additive containing sodium caprylate and sodium butyrate (Examples 1-4) provided by the present application can effectively improve the antioxidant stress resistance and relieve endoplasmic reticulum stress of juvenile mandarin fish, and improve the health of fish body.
[0093] The above embodiments of the present application are described in detail, but the present application is not limited to the above embodiments, and various changes can be made within the knowledge range possessed by those skilled in the art without departing from the purpose of the present application. In addition, the embodiments of the present application and the features in the embodiments can be combined with each other without conflict.
Claims
1. A composite functional additive for alleviating oxidative stress and endoplasmic reticulum stress in the liver of mandarin fish, said composite functional additive comprising the following components in parts by weight: Sodium octanoate 5-40 parts, sodium butyrate 5-40 parts, and rice husk powder 20-90 parts.
2. The composite functional additive according to claim 1, characterized in that, The composite functional additive comprises the following components in parts by weight: 5-20 parts sodium octanoate, 5-20 parts sodium butyrate, and 60-90 parts rice husk powder.
3. The application of the compound functional additive according to any one of claims 1 to 2 in the preparation of feed.
4. A feed comprising the compound functional additive as described in any one of claims 1 to 2.
5. The feed according to claim 4, characterized in that, The feed comprises the following raw materials in parts by weight: 95-100 parts of basic feed and 1-5 parts of the compound functional additive.
6. The feed according to claim 5, characterized in that, The basic feed includes protein sources, oils, minerals, and vitamins.
7. The feed according to claim 6, characterized in that, The protein sources include white fish meal, krill meal, and corn gluten meal; the oils include fish oil and soybean oil.
8. The use of the compound functional additive of any one of claims 1 to 2 or the feed of any one of claims 4 to 7 in the preparation of products that alleviate oxidative stress and / or endoplasmic reticulum stress in fish livers.