Application of copper methionine in preparing feed for relieving yellowing of channel catfish

Abstract

The application provides application of methionine copper in preparation of feed for relieving yellowing of Ictalurus punctatus, and belongs to the technical field of aquatic animal breeding. The methionine copper has an important role in regulating the body color of fish, improves the antioxidant and anti-inflammatory capacity of the body of Ictalurus punctatus by activating the Keap1 / Nrf2 / ARE pathway, promotes melanin synthesis and inhibits yellow pigment deposition, and relieves the yellowing phenomenon of Ictalurus punctatus. The feed is added with 14 mg / kg to 21 mg / kg of the methionine copper level (the actual copper level in the feed is 14.25 mg / kg to 15.58 mg / kg), and the regulation effect is better.

Description

Technical Field

[0001] This invention belongs to the field of aquaculture technology, and in particular relates to the application of copper methionine in the preparation of feed to alleviate yellowing of spotted catfish. Background Technology

[0002] Copper is one of the most abundant metallic elements in living organisms and an essential trace element for optimal growth and health in aquatic animals (Watanabe et al., 1997). As a cofactor for redox enzymes such as cytochrome C oxidase, monoamine oxidase, and copper-zinc superoxide dismutase (Cu, Zn-SOD), copper participates in the formation of copper chelates, antioxidant enzymes, cytochrome oxidases, and various metabolism-related proteins, playing a crucial role in redox reactions, antioxidant defense, and neuropeptide synthesis (Scott et al., 2018; Bonham et al., 2002). Furthermore, the antifungal properties of copper have been extensively studied, showing promising applications in aquaculture (Ghuglot et al., 2021; Chari et al., 2017).

[0003] Fish body color, as a direct reflection of species characteristics and health status, has a complex formation and regulation mechanism, mainly relying on the synergistic action of pigment cells such as melanocytes and xanthocyanins, as well as the synthesis and deposition of pigment substances such as melanin and carotenoids (Luo et al., 2021). Copper plays an irreplaceable role in the regulation of fish body color. As a key component of tyrosinase, copper directly participates in the biosynthesis of melanin. Insufficient copper supply will reduce tyrosinase levels, inhibit melanin synthesis, and thus cause abnormal phenomena such as paleness, yellowing, or whitening of fish body color (Wu et al., 2020). At the same time, copper maintains body color stability by regulating the function of the antioxidant system, reducing free radical damage to pigment cells (Vutukuru et al., 2006). Conversely, an imbalance in copper supply will disrupt the body's metabolic balance, leading to abnormal body color by inhibiting melanin synthesis, and at the same time causing oxidative stress that aggravates the symptoms of abnormal body color. Therefore, adding copper to feed is an important way to regulate fish body color. Traditional copper preparations used in aquaculture are mostly inorganic copper (such as copper sulfate and copper chloride). These copper sources are widely available and inexpensive, but they suffer from low bioavailability and are prone to accumulating in fish by combining with other substances to form insoluble complexes. This not only makes it difficult to accurately meet the nutritional needs of fish for copper, but also easily leads to supply imbalances due to improper dosage control, exacerbating oxidative stress and the risk of abnormal body color (Azevedo AC et al., 2021). Therefore, finding new copper sources or developing copper preparations with high bioavailability is an important way to promote better copper utilization in fish.

[0004] Compared to inorganic copper, organic copper chelates formed with amino acids, peptides, or other organic compounds achieve higher bioavailability and absorption rates (Chen et al., 2020). Methionine copper refers to copper ions (Cu²⁺). + Copper methionine forms organometallic chelates with methionine via coordination bonds. Their molecular structure typically involves copper ions coordinating with 1-2 methionine molecules in a 1:1 or 1:2 molar ratio, forming stable five- or six-membered ring structures (Yang et al., 2023). Current research on the application of copper methionine in aquaculture has been extensive, focusing primarily on promoting growth and improving feed conversion ratios (Wang et al., 2018; Zhou et al., 2010). However, research on its ability to improve abnormal fish coloration through regulation of pigment synthesis pathways and alleviating oxidative stress is still lacking, especially in its application in specifically alleviating yellowing of the skin.

[0005] The references are as follows: Watanabe T,Kiron V,Satoh S.Trace minerals in fish nutrition[J].Aquaculture,1997,151(1-4):185-207. Scott A, Vadalasetty KP, Chwalibog A, et al. Copper nanoparticles as analternative feed additive in poultry diet: a review [J]. Nanotechnology Reviews, 2018, 7(1): 69-93. Bonham M,O'Connor JM,Hannigan BM,et al.The immune system as aphysiological indicator of marginal copper status[J].British Journal ofNutrition,2002,87(5):393-403. Ghuglot R,Titus W,Agnihotri A S,et al.Stable copper nanoparticles aspotential antibacterial agent against aquaculture pathogens and humanfibroblast cell viability[J].Biocatalysis and Agricultural Biotechnology,2021,32:101932. Chari N,Felix L O,Davoodbasha M A,et al.In vitro and in vivoantibiofilm effect of copper nanoparticles against aquaculture pathogens[J].Biocatalysis&Agricultural Biotechnology,2017,10:336-341. Luo M,Lu G,Yin H,et al.Fish pigmentation and coloration:Molecularmechanisms and aquaculture perspectives[J].Reviews in Aquaculture,2021,13(4):2395-2440. Wu M,Chen X,Cui K,et al.Pigmentation formation and expressionanalysis of tyrosinase in Siniperca chuatsi[J].Fish Physiology andBiochemistry,2020,46(4):1279-1293. Vutukuru S,Chintada S,Radha Madhavi K,et al.Acute effects of copperon superoxide dismutase,catalase and lipid peroxidation in the freshwaterteleost fish,Esomus danricus[J].Fish Physiology and Biochemistry,2006,32(3):221-229. Azevedo A C B,Bozza D A,Doria H B,et al.Low levels of inorganiccopper impair reproduction parameters in Oreochromis niloticus after chronicexposure[J].Aquaculture,2021,545:737186. Chen K,Yamamoto F Y,Gatlin III D M.Effects of inorganic and organicdietary copper supplementation on growth performance and tissue compositionfor juvenile red drum(Sciaenops ocellatus L.)[J].Aquaculture Nutrition,2020,26(3):820-827. Yang J,Zhang Z,Lin G,et al.Organic copper promoted copperaccumulation and transport,enhanced low temperature tolerance andphysiological health of white shrimp(Litopenaeus vannamei Boone,1931)[J].Fish&Shellfish Immunology,2023,132:108459. Wang H, Zhu H, Wang Zhou Meng, Wang Anli, Cao Junming. Effects of different forms of copper in feed and their addition levels on growth performance, serum ceruloplasmin and growth hormone levels in Litopenaeus vannamei [J]. Marine and Limnological Journal, 2010, 41(4):577-582. Summary of the Invention

[0006] In view of this, the purpose of this invention is to provide an application of copper methionine in the preparation of feed to alleviate yellowing in channel catfish. Based on the important function of copper in regulating fish body color and the higher efficiency and safety of copper methionine's bioavailability, this invention focuses on the restorative effect of copper methionine on the body color of yellowed channel catfish. By adding different concentrations of copper methionine to the feed, the effects on body color parameters, melanin and yellow pigment content, tyrosinase, and antioxidant-related indicators of yellowed channel catfish are investigated, clarifying the appropriate addition level and potential mechanism of action of copper methionine in alleviating yellowing. The aim is to provide a scientific basis for the development of safe and efficient body color regulating feed additives, solve the yellowing problem in channel catfish farming, and improve the theoretical system of fish micronutrient nutrition and body color regulation, thus contributing to the green and healthy development of the industry.

[0007] To achieve the above-mentioned objectives, the present invention provides the following technical solution: This invention provides the application of copper methionine in the preparation of feed to alleviate yellowing in spotted catfish.

[0008] Preferably, the amount of copper methionine added is 7~27 mg / kg.

[0009] Compared with the prior art, the present invention has the following beneficial effects: The copper methionine of this invention plays an important role in regulating fish body color. By activating the Keap1 / Nrf2 / ARE pathway, it enhances the antioxidant and anti-inflammatory capabilities of channel catfish, promotes melanin synthesis, and inhibits xanthophyll deposition, thus alleviating yellowing in channel catfish. Adding copper methionine at a level of 14 mg / kg to 21 mg / kg to the feed (the actual measured copper levels in the feed were 14.25 mg / kg to 15.58 mg / kg) resulted in a superior regulatory effect. Attached Figure Description

[0010] Figure 1 These are the skin and muscle color values ​​(data expressed as mean ± standard deviation; different superscript values ​​indicate significant differences). P<0.05 ), b Yellowness value); Figure 2 These are plasma AST and ALT levels (data are expressed as mean ± standard deviation; different superscript values ​​indicate significant differences). P <0.05)); Figure 3 These are antioxidant levels in the skin and muscles (data are expressed as mean ± standard deviation; different superscript values ​​indicate significant differences). P <0.05)); Figure 4 This refers to pigment deposition and tyrosinase content in the skin and muscles (data are expressed as mean ± standard deviation; different superscript values ​​indicate significant differences). P <0.05)); Figure 5 This represents the expression of genes related to pigment synthesis in the skin and muscles (data are expressed as mean ± standard deviation; different superscript values ​​indicate significant differences). P <0.05)); Figure 6 This represents the expression of antioxidant and immune-related genes in the skin and muscles (data are expressed as mean ± standard deviation; different superscript values ​​indicate significant differences). P <0.05). Detailed Implementation

[0011] This invention provides the application of copper methionine in the preparation of feed to alleviate yellowing in channel catfish. In this invention, the amount of copper methionine added is 7-27 mg / kg, preferably 14 mg / kg-21 mg / kg.

[0012] The technical solutions provided by the present invention will be described in detail below with reference to the embodiments, but they should not be construed as limiting the scope of protection of the present invention.

[0013] Example 1

[0014] 1. Experimental Materials and Methods

[0015] 1.1 Experimental Feed

[0016] Commercial feed was used as the control group in this experiment. Five experimental feeds were prepared by adding 0 mg / kg (Cu1), 7 mg / kg (Cu2), 14 mg / kg (Cu3), 21 mg / kg (Cu4), and 27 mg / kg (Cu5) copper methionine to the control group feeds. The measured copper content in the five experimental feeds were 10.82 mg / kg, 12.63 mg / kg, 14.35 mg / kg, 15.58 mg / kg, and 17.40 mg / kg, respectively. The copper methionine was purchased from Shanghai Maclean Biochemical Technology Co., Ltd., with a purity of 98%.

[0017] Table 1. Experimental feed formulation and nutrient composition levels (% dry weight)

[0018] Note: 66% protein imported fishmeal: crude protein 66.5%, crude fat 7.8%; pork meal: crude protein 67.3%, crude fat 12.1%; soybean meal: crude protein 45.7%, crude fat 1.2%; rapeseed meal: crude protein 36.8%, crude fat 2.1%; wheat flour: crude protein 13.2%, crude fat 1.7%; high-fat rice bran: crude protein 13.5%, crude fat 16.5%. All raw materials were purchased from Wuxi Tongwei Feed Co., Ltd. (Wuxi, China).

[0019] Multivitamin and mineral premix, including vitamin premix and mineral premix. Both were purchased from Wuxi Huanoway Animal Health Products Co., Ltd. (Wuxi, China). Vitamin premix (IU or mg per kg of feed): Vitamin A: 900,000 IU; Vitamin D: 250,000 IU; Vitamin E: 4,500 mg; Vitamin K: 3,220 mg; Vitamin B1: 320 mg; Vitamin B2: 1,090 mg; Vitamin B5: 2,000 mg; Vitamin B6: 5,000 mg; Vitamin B12: 116 mg; Pantothenic acid: 1,000 mg; Folic acid: 165 mg; Biotin: 50 mg; Niacin: 2,500 mg. Mineral premix (grams per kilogram of feed): Calcium phosphate: 20 g; Sodium chloride: 2.6 g; Potassium chloride: 5 g; Magnesium sulfate: 2 g; Ferrous sulfate: 0.9 g; Zinc sulfate: 0.06 g; Copper sulfate: 0.02 g; Manganese sulfate: 0.03 g; Sodium selenite: 0.02 g; Cobalt chloride: 0.05 g; Potassium iodide: 0.004 g.

[0020] 1.2 Experimental Fish and Feeding Management

[0021] First, channel catfish were subjected to a one-month yellowing treatment using feed containing 66.8 mg / kg MDA. The yellowness of the skin of the yellowed channel catfish was measured using a colorimeter. Uninjured channel catfish of similar size and yellowness were selected as experimental fish and fed the control group feed for two weeks to acclimatize them to the culture environment. The culture experiment was conducted in a pond cage (2 m²) at the Nanquan Experimental Base of the Freshwater Fisheries Research Center, Chinese Academy of Fishery Sciences. 2 m The experiment was conducted in 15 net cages (2 m). 180 channel catfish with an initial weight of 830±5 g were randomly divided into 5 groups, with 3 replicates per group and 20 fish per replicate, for a total of 15 net cages. The fish were cultured for 8 weeks. During the experiment, five different experimental feeds were fed in equal amounts daily (7:00 and 17:00), at 3% of the fish's initial weight, with adjustments made according to weather conditions. The activity and health status of the fish were observed daily, and the number of deaths and the amount of feed were recorded. Water quality parameters in the ponds were monitored regularly. Environmental parameters throughout the culture period included dissolved oxygen (>6 mg / L), water temperature (24-29℃), ammonia nitrogen (≤0.02 mg / L), and natural light cycle conditions.

[0022] 1.3 Sample Collection and Colorimetric Analysis

[0023] After the aquaculture experiment concluded, feeding was stopped for 24 hours. Three fish were randomly selected from each aquaculture cage. They were first anesthetized with MS-222 (200 mg / L), and then the skin and muscle at the same location on the back of the fish were measured using a colorimeter (CR-400 model) to obtain color values ​​(b...). The following steps were performed: Blood was collected from the tail vein of the experimental fish into anticoagulant tubes and centrifuged at 3000 rpm for 10 min at 4°C to obtain plasma, which was then stored. Finally, skin and muscle tissue from the back were collected. After sampling, all samples were stored in an ultra-low temperature freezer at -80°C for subsequent experimental analysis.

[0024] Experimental results: such as Figure 1 As shown. In skin tissue, compared with the control group (Cu1), the Cu3-Cu5 group significantly reduced b... ( P <0.05). In muscle tissue, compared with the control group (Cu1), the Cu2-Cu5 group significantly reduced b ( P <0.05).

[0025] 1.4 Experimental Measurement Methods

[0026] 1.4.1 Analysis of plasma transaminase activity and tissue antioxidant indicators

[0027] The activities of AST and ALT in plasma were determined using commercial kits from Nanjing Jiancheng Biotechnology Institute (Nanjing, China) via the microplate method. Kit models (C010-2-1 and C009-2-1) were used. Specific methods were described in their respective instructions. The levels of antioxidant markers in skin and muscle were determined using commercial kits from Nanjing Jiancheng Biotechnology Institute (Nanjing, China), including total antioxidant capacity (T-AOC), total superoxide dismutase (T-SOD), glutathione peroxidase (GPx), catalase (CAT), and MDA. The methods and kit models used were ABTS method (A015-2-1), WST-1 method (A001-3-2), colorimetric method (A005-1-2), ammonium molybdate method (A007-1-1), and TBA method (A003-1-2), respectively. Specific methods were described in their respective instructions.

[0028] Experimental results: such as Figure 2 As shown, no significant differences were observed in ALT and AST levels among the groups. P >0.05).

[0029] like Figure 3 As shown. In skin tissue, compared with the control group (Cu1), the GSH level in the Cu2-Cu5 group and the GPx activity in the Cu2-Cu4 group were significantly increased ( P <0.05. No significant differences were observed in T-AOC, T-SOD, CAT, and MDA levels among the groups. P >0.05). In muscle tissue, compared with the control group (Cu1), the CAT activity in the Cu3-Cu5 group and the GSH level in the Cu4-Cu5 group were significantly increased ( P <0.05), while no significant differences were observed in T-AOC, T-SOD, GPx, and MDA indicators ( P >0.05).

[0030] 1.4.2 ELISA analysis of pigment-related indicators in skin and muscle

[0031] The contents of xanthophyll, melanin and tyrosinase in skin and muscle were detected using an ELISA kit from Shanghai Enzyme-Link Biotechnology Co., Ltd. (Shanghai, China). The methods are as follows: (1) The content of xanthophyll in the sample was determined by a competitive method (kit model YJ270190). Briefly, the purified xanthophyll antibody was coated onto a microplate to form a solid phase antibody. The xanthophyll sample was added and competitively bound to the horseradish peroxidase (HRP)-labeled xanthophyll antigen. After thorough washing, tetramethylbenzidine (TMB) substrate was added for color development. The color intensity of the sample was negatively correlated with the xanthophyll content. The absorbance (OD value) was measured at 450 nm using an enzyme-linked immunosorbent assay (ELISA) reader, and the xanthophyll content was calculated by the standard curve method; (2) The content of melanin in the sample was determined by a competitive method (kit model YJ234125). Briefly, the sample, standard and HRP-labeled detection antibody were added sequentially to a microplate coated with melanin antibody. After incubation and thorough washing, the substrate TMB was added for color development. TMB turns blue under the catalysis of peroxidase, and turns yellow after acid termination. The color intensity of the sample is positively correlated with the melanin concentration. The OD value is measured at a wavelength of 450 nm to calculate the melanin content; (3) The content of tyrosinase in the sample is determined by competitive method (kit model H310-1-2). First, biotin-labeled antigen is added to the enzyme-labeled plate coated with antibody and incubated at 37°C for 30 min to form immune complexes. Then, after thoroughly washing with phosphate buffer solution Tween 20 (PBST) to remove unbound biotin antigen, avidin-HRP is added and incubated at 37°C for another 30 min to bind avidin-HRP to biotin antigen. After washing, the bound HRP catalyzes the color development of TMB, which turns from blue to yellow after acid termination. The OD value is measured at a wavelength of 450 nm. The OD value is negatively correlated with the antigen concentration in the sample. The analysis and calculation of all detection indicators are performed according to the instructions of the relevant kit.

[0032] Experimental results: such as Figure 4 As shown. In skin tissue, compared with the control group (Cu1), the melanin content of Cu2 and Cu3 groups was significantly increased ( P <0.05%, the content of yellow pigment in the Cu3-Cu5 group was significantly reduced ( P <0.05%, the tyrosinase content in the Cu3 and Cu4 groups was significantly increased ( P <0.05%. In muscle tissue, compared with the control group (Cu1), the Cu3-Cu5 group showed significantly increased tyrosinase and melanin content ( P <0.05%, the yellow pigment content in the Cu4 and Cu5 groups was significantly reduced ( P <0.05).

[0033] 1.4.3 qRT-PCR analysis

[0034] RNA was extracted from skin and muscle using the RNAiso Plus kit (Nanjing Novizan Biotechnology Co., Ltd., Nanjing, China). RNA concentration and quality were measured using a NanoDrop 2000 spectrophotometer (Thermo Fisher Scientific, Wilmington, USA), ensuring an A260 / 280 value between 1.8 and 2.0. qRT-PCR was performed using the OneStep qRT-PCR SYBR Green kit (Nanjing Novizan Biotechnology Co., Ltd., Nanjing, China) on a CFX96Touch (Bio-Rad, California, USA). The glyceraldehyde-3-phosphate dehydrogenase (gapdh) gene was selected as an internal control gene due to its stability in this study. Primers used are shown in Table 2 and were synthesized by Sangon Biotech (Shanghai) Co., Ltd. (Shanghai, China). Relative gene expression was calculated using the 2-ΔΔCt method.

[0035] Table 2 Primer sequences for real-time quantitative PCR analysis

[0036] Experimental results: such as Figure 5 As shown. In skin tissue, compared with the control group (Cu1), the Cu5 group... mitf Cu4 group tyr Groups Cu3 and Cu4 etb and Cu3-Cu5 group camk2 Gene expression was significantly increased ( P <0.05). Cu2-Cu5 group xdh Gene expression was significantly reduced ( P <0.05). α-msh There was no significant difference in expression among the groups. P >0.05). In muscle tissue, compared with the control group (Cu1), the Cu4 group... mitf and tyr Cu3-Cu5 group etb and Cu4 and Cu5 groups camk2 The relative expression levels were significantly increased ( P <0.05). Cu3-Cu5 group xdh The relative expression level was significantly reduced ( P <0.05). α- msh There was no significant difference in relative expression among the groups. P >0.05).

[0037] like Figure 6 As shown. In skin tissue, compared with the control group (Cu1), the Cu2-Cu4 group... keap1 and Cu2-Cu5 group nfκb The relative expression level was significantly reduced ( P <0.05), Cu3-Cu5 group nrf2 and Cu4 group il-10 The relative expression levels were significantly increased ( P <0.05), Ifn-γ and gpx There was no significant difference in relative expression levels among the groups. P >0.05). In muscle tissue, compared with the control group (Cu1), the Cu3-Cu5 group keap1 The relative expression level was significantly reduced ( P <0.05), Cu4 group nrf2 The relative expression level was significantly increased ( P <0.05). Ifn-γ , gpx , nfκb and il-10 There was no significant difference in relative expression levels among the groups. P >0.05).

[0038] 1.5 Data Analysis

[0039] Before conducting statistical analysis, the Shapiro-Wilk test was used to assess the normality of the data, and the Levene test was used to verify homogeneity of variance. When the data met the requirements of normality and homogeneity of variance, one-way ANOVA (Tukey's test) was used for comparisons between groups. Data are expressed as mean ± standard deviation, and the significance threshold was set to [value missing]. P <0.05.

[0040] As demonstrated in the above examples, the addition of appropriate amounts of copper methionine to the feed significantly increased the GSH content in the skin and muscle, the CAT activity in the muscle, and the GPx activity in the skin. This indicates that the intervention of exogenous copper methionine greatly enriches the body's non-enzymatic antioxidant reserves and enhances the activity of key antioxidant enzymes. With the increase of copper methionine addition in the feed, the MDA content in each group decreased slightly, but there was no significant difference between the groups. This indicates that the addition of 7-27 mg / kg of copper methionine in the feed not only did not cause oxidative damage to the skin and muscle tissue of channel catfish, but also maintained the integrity of the cell membrane by enhancing the activity of antioxidant enzymes and the level of non-enzymatic antioxidants.

[0041] With the addition of copper methionine to the feed, the skin and muscle tissue of the spotted catfish showed increased activity. keap1 Expression was significantly downregulated. nrf2Significant upregulation of expression indicates that copper methionine can efficiently promote the dissociation of Nrf2 from Keap1, thereby enhancing the body's antioxidant defense capabilities by inhibiting the ubiquitination and degradation of Nrf2 by Keap1. Pro-inflammatory factors in the skin of spotted catfish in the Cu2-Cu4 group. nfκb The relative expression levels of [a specific substance] were significantly lower than those in the Cu1 group, while the expression levels of anti-inflammatory factors [were lower]. il-10 The expression of methionine copper was significantly upregulated in the Cu4 group, indicating that while enhancing antioxidant capacity, methionine copper effectively inhibited the inflammatory response in fish, thus providing a non-inflammatory microenvironment conducive to the survival and development of pigment cells and the normal synthesis of pigments.

[0042] With increasing addition of copper methionine to feed, the skin and muscle of spotted catfish showed increased beta-methionine levels. The color value was significantly reduced. This change in color value indicates that copper methionine can effectively alleviate the yellowing phenomenon in fish. After adding an appropriate concentration of copper methionine, the melanin content in the skin and muscle increased significantly, while the yellow pigment content decreased significantly. Specifically, the tyrosinase content in the skin and muscle of the Cu3 and Cu4 groups increased significantly, and this trend was highly consistent with the increase in melanin content, indicating that the addition of copper methionine to the feed promotes melanin production by increasing tyrosinase content. By supplementing the feed with an appropriate amount of copper methionine, the spotted catfish can obtain sufficient enzyme cofactors, thereby restarting the blocked melanin synthesis pathway. This not only promotes the large-scale synthesis and deposition of melanin but also inhibits the synthesis of yellow pigment to a certain extent, ultimately manifesting as a yellow pigment in the fish's macroscopic body color. The decrease in color and the improvement in body color.

[0043] In skin and muscle tissue, appropriate levels of copper methionine in the feed significantly upregulated [the condition]. mitf and tyr The relative expression level of mRNA in the skin and muscles of the spotted catfish. mitf and tyr The simultaneous upregulation of methionine copper reveals that it does not merely participate in the reaction passively as a cofactor, but may also indirectly regulate methionine copper by affecting copper ion homeostasis and transport. mitf and tyr It actively activates the entire melanin production pathway at the transcriptional level.

[0044] etb and camk2 The expression of these genes significantly increased after the addition of appropriate levels of copper methionine. This indicates that copper methionine not only promotes melanin production but may also improve the body color of yellow-spotted channel catfish by promoting the development and distribution of melanocytes. In stark contrast to the upregulation of genes related to melanin synthesis, xdh Gene expression was significantly downregulated, while xdhThe expression of [a specific pigment] is closely related to the synthesis of xanthophyll. Since fish body color is a comprehensive manifestation of multiple pigment cells, xdh The downregulation explains, at the molecular level, the significant reduction in flavonoid content in the group with appropriate levels of copper methionine, and b The significant decrease in melanin production, combined with the inhibition of yellow pigment synthesis, constitutes the core molecular mechanism by which copper methionine efficiently restores the yellow body color of spotted catfish.

[0045] The ALT and AST levels in the plasma of channel catfish in each experimental group did not show significant statistical differences due to the addition of copper methionine, suggesting that the addition of 7-27 mg / kg of copper methionine to the feed did not cause significant substantial damage or toxicity to the liver of channel catfish.

[0046] The above description is only a preferred embodiment of the present invention. It should be noted that for those skilled in the art, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. Application of copper methionine in the preparation of feed to alleviate yellowing in spotted catfish.

2. The application according to claim 1, characterized in that, The amount of copper methionine added is 7~27 mg / kg.

Images