Triploid rainbow trout juvenile fish feed and application thereof

Abstract

The invention discloses a triploid rainbow trout juvenile fish feed and application thereof. The formula of the feed comprises 300.0 g / kg of fish meal, 120.0 g / kg of krill meal, 100.0 g / kg of soybean meal, 80.0 g / kg of soybean protein concentrate, 80.0 g / kg of corn protein, 20.0 g / kg of wheat bran powder, 165.4 g / kg of wheat flour and a mixture of different proportions of fish oil and dephenolized cottonseed oil, the proportion of the dephenolized cottonseed oil replacing the fish oil in the mixture is 25%-100%, and 10.0 g / kg of premix, 0.2 g / kg of astaxanthin, 3.0 g / kg of choline chloride, 1.0 g / kg of EDTA, 0.3 g / kg of betaine and 0.1 g / kg of calcium propionate are added. The feed disclosed by the invention is applied to culture of juvenile triploid rainbow trout, and can be used for remarkably regulating fatty acid composition of a fish body, improving muscle texture and enhancing oxidation resistance and heat stress resistance of the liver on the premise of not influencing growth performance, so that dependence on fish oil is reduced, and green, efficient and sustainable development of aquaculture is promoted.

Description

Technical Field

[0001] This invention relates to the field of aquatic feed technology, specifically to a triploid rainbow trout juvenile feed that uses dephenolized cottonseed oil as a fish oil substitute for fat source and its application in triploid rainbow trout juvenile farming. Background Technology

[0002] Triploid rainbow trout (Oncorhynchus mykiss (triploid)) is an important species in cold-water fish farming, holding a significant position in global intensive aquaculture due to its rapid growth, strong adaptability, and non-reproductive characteristics. In recent years, with the continuous expansion of rainbow trout farming, the demand for fish oil in feed has been increasing. However, fish oil resources are highly dependent on marine fisheries, and their supply is severely constrained by fluctuations in natural resources, rising prices, and sustainability issues. There is an urgent need to develop stable, economical, and environmentally friendly alternative lipid sources to reduce dependence on fish oil.

[0003] Degossypolized cottonseed oil (CSO) is a vegetable oil extracted from cottonseed after detoxification treatment. This process significantly improves the safety and bioavailability of the oil by removing anti-nutritional factors such as gossypol. Due to its relatively balanced fatty acid composition, low price, and stable supply, degossypolized cottonseed oil has been widely used in poultry and livestock feed as a potential source of plant-based fats. However, research on the application of degossypolized cottonseed oil in aquatic animal feed, especially its effects on the growth, health, and lipid metabolism of triploid rainbow trout juveniles, remains limited. Existing aquatic feed research only includes a few studies on the use of ordinary cottonseed oil or partially degossypolized oil in warm-water fish, and the results often vary depending on the fish species, substitution ratio, and processing method. Summary of the Invention

[0004] The purpose of this invention is to solve the problems of high dependence on fish oil, high cost and lack of sustainability in existing rainbow trout farming, and to provide a triploid rainbow trout juvenile feed and its application. This feed can effectively improve the growth performance, muscle texture and stress resistance of triploid rainbow trout juveniles.

[0005] The technical solution adopted in this invention is as follows:

[0006] A triploid rainbow trout juvenile fish feed, the feed formula being: 300.0 g / kg fish meal, 120.0 g / kg krill meal, 100.0 g / kg soybean meal, 80.0 g / kg soybean protein concentrate, 80.0 g / kg corn protein, 20.0 g / kg wheat bran, 165.4 g / kg wheat flour, a mixture of fish oil and dephenolized cottonseed oil with a total addition of 120.0 g / kg, 10.0 g / kg premix, 0.2 g / kg astaxanthin, 3.0 g / kg choline chloride, 1.0 g / kg EDTA, 0.3 g / kg betaine, and 0.1 g / kg calcium propionate; wherein the dephenolized cottonseed oil replaces 25%~100% of the fish oil in the mixture of fish oil and dephenolized cottonseed oil.

[0007] This invention also provides the application of the above-mentioned feed in the culture of triploid rainbow trout juveniles. Triploid rainbow trout juveniles with an initial weight of 51.65 ± 1.56g are stocked in the culture pond for culture. The feed is given three times a day at 6:00, 12:00 and 18:00, and the daily feed amount is 3% of the body weight.

[0008] Preferably, during the breeding process, the water is changed twice a day, and the amount of water changed each time is 50% of the total volume.

[0009] Preferably, the breeding cycle is 56 days.

[0010] Preferably, during the aquaculture process, the water temperature is controlled at 13~15℃, dissolved oxygen is 6.63 ± 0.5 mg / L, pH is 8.3 ± 0.1, ammonia nitrogen is ≤ 0.05mg / L, and nitrite is ≤ 0.04mg / L.

[0011] The beneficial effects of this invention are as follows: By using dephenolized cottonseed oil to replace 25%-100% of fish oil in the feed, this invention produces isonitrogenous and isolipid rainbow trout feed. When this feed is used in the breeding of triploid rainbow trout juveniles, it can significantly optimize the composition of muscle fatty acids, improve muscle texture, and enhance the liver's antioxidant capacity and heat stress resistance while maintaining growth performance and survival rate. This effectively reduces dependence on fish oil and promotes the development of aquatic feed towards a more economical and sustainable direction. Attached Figure Description

[0012] Figure 1 The effect of dephenolized cottonseed oil as a substitute for fish oil on the survival rate, growth performance and body mass index of triploid rainbow trout juveniles; Note: The same letter in the same row indicates no significant difference (P > 0.05, n = 8) was used for Turkey multiple test.

[0013] Figure 2The effect of dephenolized cottonseed oil as a substitute for fish oil on the crude composition of triploid rainbow trout juveniles; Note: (1) According to the Turkey multiple test, the same letter in the same row indicates no significant difference (P > 0.05, n = 4). (2) ww, wet weight.

[0014] Figure 3 The effect of dephenolized cottonseed oil as a substitute for fish oil on the fatty acid composition of triploid rainbow trout juvenile muscle; Note: (1) n = 3. (2) SFA, saturated fatty acids; MUFA, monounsaturated fatty acids; PUFA, total polyunsaturated fatty acids.

[0015] Figure 4 The effect of dephenolized cottonseed oil as a substitute for fish oil on the muscle texture of triploid rainbow trout juveniles; Note: According to the Turkey multiple test, the same letter in the same row indicates no significant difference (P > 0.05, n = 4).

[0016] Figure 5 The effect of dephenolized cottonseed oil as a substitute for fish oil on the antioxidant capacity of triploid rainbow trout livers was investigated. The Turkey multiple test was performed, and the same letter in the same row indicated no significant difference (P > 0.05, n = 6).

[0017] Figure 6 This study investigated the effects of different levels of dephenolized cottonseed oil substitution for fish oil on the expression of genes related to lipid metabolism in the liver of triploid rainbow trout juveniles. Note: pparα, peroxisome proliferator-activated receptor α; srebp1, sterol regulatory element-binding protein 1; fasn, fatty acid synthase; fads2, fatty acid desaturase 2 (Δ6 desaturase); elovl2, very long chain fatty acid elongase 2; cpt1, carnitine palmitoyltransferase 1; fabp1, fatty acid-binding protein 1 (liver type). Data are presented as mean ± standard error (n = 4). Turkey's multiple test was used; identical letters in the bars indicate no significant difference (P > 0.05).

[0018] Figure 7 The effect of different levels of dephenolized cottonseed oil substitution for fish oil on the heat stress resistance of triploid rainbow trout juveniles; Note: (A) serum cortisol level; (B) hepatic heat shock protein (Hsp70) content; (C, D) expression of hepatic stress-related genes. Note: hspa8, member 8 of heat shock protein family A (Hsp70); nrf2, nuclear factor E2-related factor 2; cat, catalase; pdia4, protein disulfide isomerase A4. Data are presented as mean ± standard error (n = 4). Turkey's multiple test was used; identical letters in the bar charts indicate no significant difference (P > 0.05). Detailed Implementation

[0019] The present invention will be further described below with reference to the accompanying drawings and specific embodiments.

[0020] Example 1

[0021] 1. Feed and grouping

[0022] The basic feed formula for triploid rainbow trout juveniles is as follows: 300.0 g / kg fish meal, 120.0 g / kg krill meal, 100.0 g / kg soybean meal, 80.0 g / kg soybean protein concentrate, 80.0 g / kg corn protein, 20.0 g / kg wheat bran, 165.4 g / kg wheat flour, 10.0 g / kg premix, 0.2 g / kg astaxanthin, 3.0 g / kg choline chloride, 1.0 g / kg EDTA, 0.3 g / kg betaine, 0.1 g / kg calcium propionate, and the total addition of the mixture of fish oil and dephenolized cottonseed oil is 120.0 g / kg.

[0023] In a mixture of fish oil and dephenolized cottonseed oil, dephenolized cottonseed oil was used to replace 25% (CSO25), 50% (CSO50), 75% (CSO75), and 100% (CSO100) of fish oil, respectively, to produce four kinds of isonitrogenous and isolipid feeds (the feed formula and conventional ingredients are shown in Table 1, and the fatty acid composition is shown in Table 2).

[0024] A control group was set up, which was fed a whole fish oil diet (FO); four experimental groups were set up, which were fed isonitrogenous and isolithic diets (CSO25, CSO50, CSO75, and CSO100) after the above four types of dephenolized cottonseed oil replaced fish oil.

[0025] 2. Rainbow trout farming and feeding management

[0026] Triploid rainbow trout juveniles with an initial body weight of 51.65 ± 1.56 g were randomly assigned to 20 500 L culture tanks, with 4 replicates per treatment group and 35 fish per tank. The experimental period was 56 days, with a 7-day acclimatization period before the experiment. Fish were fed three times daily at 6:00, 12:00, and 18:00, with a daily feed amount of 3% of body weight and a feed particle size of 5 mm. The culture water was freshwater, with a temperature controlled at 13–15℃, dissolved oxygen 6.63 ± 0.5 mg / L, pH 8.3 ± 0.1, ammonia nitrogen ≤ 0.05 mg / L, and nitrite ≤ 0.04 mg / L. 50% of the water was changed twice daily. A thermostat was used to control the water temperature during the culture period.

[0027] Table 1. Feed formulation and typical nutrient composition (% of feed dry weight)

[0028]

[0029] 1) Using FO as the control group, experimental groups were formed by replacing 25%, 50%, 75%, and 100% of the fish oil with dephenolized cottonseed oil, respectively. The experimental groups were named CSO25, CSO50, CSO75, and CSO100, respectively.

[0030] 2) Purchased from Qingdao Saigelin Marine Biological Feed Co., Ltd.

[0031] 3) Purchased from Shandong Liaocheng Yijiafu Grain and Oil Co., Ltd.

[0032] 4) The premix is ​​provided by Qingdao Master Biotechnology Co., Ltd. (kg) -1 (Feed dry weight): Vitamin A acetate 800,000 IU; Riboflavin 1.5 g; Folic acid 0.8 g; Magnesium 10 g; Copper 0.3 g; Vitamin D3 200,000 IU; Pyridoxine hydrochloride 1.3 g; D-Biotin 0.01 g; Iron 10 g; Cobalt 0.05 g; Ethyl d1-α-tocopherol 7 g; Cyanocobalamin 0.005 g; L-Ascorbic acid-2-phosphate 30 g; D-Calcium pantothenate 3.4 g; Menadione 2 g; Zinc 56 g; Selenium 0.01 g; Manganese 3 g; Iodine 0.1 g; Thiamine nitrate 1 g; Nicotinamide 5 g; Inositol 10 g.

[0033] 5) Purchased from Beijing Leigen Biotechnology Co., Ltd.

[0034] Table 2 Fatty acid composition of each group of feeds (% total fatty acids)

[0035]

[0036]

[0037] 1) n = 3.

[0038] 2) SFA, saturated fatty acids; MUFA, monounsaturated fatty acids; PUFA, total polyunsaturated fatty acids.

[0039] 3. Indicator Measurement

[0040] After the experiment, fish were fasted for 24 hours. Eight fish from each treatment group were taken to measure body length, body weight, liver weight, and visceral mass weight. Weight gain rate (WGR), specific growth rate (SGR), protein efficiency (PER), feed conversion ratio (FCR), liver-to-body ratio (HSI), and visceral mass ratio (VSI) were calculated.

[0041] Four whole fish and four muscle and liver samples were taken from each treatment group to determine crude fat, crude protein, moisture and ash content.

[0042] Blood was collected from the vena cava of 6 fish in each treatment group. The serum was separated and stored at -80℃ for subsequent biochemical analysis. At the same time, muscle and liver samples were collected and stored at -80℃ to determine fatty acid composition, antioxidant indicators, heat shock protein Hsp70 content and related gene expression (qRT-PCR, primer sequences are shown in Table 3).

[0043] Four fish from each treatment group were taken on-site to determine the texture of the back muscles (hardness, elasticity, adhesiveness, etc.).

[0044] In addition, a 24-hour heat stress experiment was conducted. The control group was maintained at 14°C, while the heat stress group was raised by 1°C per hour to 24°C and maintained for 24 hours. Immediately after the experiment, samples were taken to detect serum cortisol, liver heat shock protein Hsp70, and the expression of other stress genes.

[0045] All the above indicators were measured using common aquatic experimental methods and corresponding kits. The primers used in the qRT-PCR experiment were based on target genes screened from sequenced transcriptome data and designed using Primer Primer 5.0 software.

[0046] Table 3 Primer sequences for real-time quantitative PCR

[0047]

[0048] 1) pparα, peroxisome proliferator-activated receptor α; hspa8, heat shock protein family A (Hsp70) member 8; nrf2, nuclear factor E2-related factor 2; srebp1, sterol regulatory element-binding protein 1; fabp1, fatty acid-binding protein 1 (liver type); fads2, fatty acid desaturase 2 (Δ6 desaturase); elovl2, very long chain fatty acid elongase 2; cpt1, carnitine palmitoyltransferase 1; pdia4, protein disulfide isomerase A4; cat, catalase; fasn, fatty acid synthase.

[0049] 4. Test results as follows Figures 1-7 As shown.

[0050] (1) Growth performance and body shape index

[0051] Depend on Figure 1 It was observed that the substitution of fish oil with dephenolized cottonseed oil did not exhibit a significant linear or quadratic effect on final body weight, weight gain rate, or specific growth rate (P > 0.05), and there were no significant differences among the groups. Survival rate decreased with increasing substitution ratio, with the CSO100 group significantly lower than the FO group (P < 0.05). The liver-to-body ratio increased with increasing substitution ratio, with the CSO100 group significantly higher than the FO group (P < 0.05). There was no significant difference in the organ-to-body ratio among the groups (P > 0.05).

[0052] (2) Crude composition of fish body and tissue

[0053] Depend on Figure 2 It was observed that there were no significant differences in crude fat, crude protein, moisture, and ash content among the groups in whole fish and muscle (P > 0.05). Crude fat in the liver increased significantly with the increase of the proportion of cottonseed oil replaced by dephenolized oil (P < 0.05), with the CSO75 and CSO100 groups significantly higher than the FO group; liver moisture, on the other hand, showed a decreasing trend with the increase of the replacement proportion, with the CSO100 group significantly lower than the FO group (P < 0.05).

[0054] (3) Composition of muscle fatty acids

[0055] Depend on Figure 3 It is evident that with the increase in the substitution ratio of dephenolized cottonseed oil, the saturated fatty acid (ΣSFA) content in muscle significantly increased, the monounsaturated fatty acid (ΣMUFA) significantly decreased, and the total polyunsaturated fatty acid (ΣPUFA) and n-3 series polyunsaturated fatty acid (Σn-3PUFA) significantly decreased (P < 0.05). The contents of C20:5n3 (EPA) and C22:6n3 (DHA) decreased significantly with the increase in the substitution ratio, while the Σn-6 / Σn-3 ratio and the DHA / EPA ratio increased significantly (P < 0.05).

[0056] (4) Muscle texture characteristics

[0057] Depend on Figure 4 It is evident that the substitution of fish oil with dephenolized cottonseed oil has a significant effect on muscle hardness, adhesiveness, chewiness, and tearing force (P < 0.05). Among them, the hardness and tearing force of the CSO25 and CSO100 groups are significantly higher than those of the FO group; the elasticity is the lowest in the CSO25 group, and there is no significant difference among the other groups (P > 0.05).

[0058] (5) Liver antioxidant capacity

[0059] Depend on Figure 5 It can be seen that the total antioxidant capacity (T-AOC) was the highest in the CSO25 group and the lowest in the CSO100 group; the catalase (CAT) activity was the highest in the CSO50 group and was significantly higher than other groups (P < 0.05); the malondialdehyde (MDA) content was the lowest in the CSO25 and CSO50 groups; and the reduced glutathione (GSH) was the highest in the CSO50 group (P < 0.05).

[0060] (6) Expression of lipid metabolism and stress-related genes

[0061] Depend on Figure 6It is evident that as the proportion of cottonseed oil replaced by dephenolized cottonseed oil increases, the expression of lipid metabolism-related genes in the liver, such as pparα, fasn, fads2, and elovl2, is downregulated to varying degrees, while cpt1 shows the highest expression in the CSO100 group.

[0062] Depend on Figure 7 It is evident that after heat stress, the increase in serum cortisol in the CSO25 and CSO50 groups was lower than that in the FO group; the liver Hsp70 content and the expression of related genes hspa8, nrf2, cat, and pdia4 remained at a high level in the CSO25 and CSO50 groups, demonstrating better heat stress resistance.

[0063] In summary, this invention uses dephenolized cottonseed oil to replace fish oil in feed for the breeding of triploid rainbow trout juveniles. While maintaining growth performance, it can significantly regulate the fatty acid composition of the fish, improve muscle texture, enhance liver antioxidant capacity, and improve their tolerance to heat stress, thus providing a feasible solution for the efficient replacement of fish oil in aquatic feed.

[0064] The specification and drawings of this invention are intended to be illustrative rather than restrictive. Based on this invention, those skilled in the art can make substitutions and modifications to some of the technical features without creative effort, and all such modifications are within the scope of protection of this invention.

Claims

1. A feed for triploid rainbow trout juveniles, characterized in that, The feed formulation is as follows: 300.0 g / kg fish meal, 120.0 g / kg krill meal, 100.0 g / kg soybean meal, 80.0 g / kg soybean protein concentrate, 80.0 g / kg corn protein, 20.0 g / kg wheat bran, 165.4 g / kg wheat flour, a mixture of fish oil and dephenolized cottonseed oil totaling 120.0 g / kg, 10.0 g / kg premix, 0.2 g / kg astaxanthin, 3.0 g / kg choline chloride, 1.0 g / kg EDTA, 0.3 g / kg betaine, and 0.1 g / kg calcium propionate; in the mixture of fish oil and dephenolized cottonseed oil, the dephenolized cottonseed oil replaces 25%~100% of the fish oil.

2. The application of the feed as described in claim 1 in the rearing of triploid rainbow trout juveniles, characterized in that, Triploid rainbow trout juveniles with an initial weight of 51.65 ± 1.56 g were stocked in the breeding pond for cultivation. They were fed three times a day at 6:00, 12:00 and 18:00, with a daily feed amount of 3% of their body weight.

3. The application according to claim 2, characterized in that, During the breeding process, the water is changed twice a day, with each change replacing 50% of the total volume.

4. The application according to claim 2, characterized in that, The breeding cycle is 56 days.

5. The application according to claim 2, characterized in that, During the aquaculture process, the water temperature is controlled at 13~15℃, dissolved oxygen is 6.63 ± 0.5mg / L, pH is 8.3 ± 0.1, ammonia nitrogen is ≤ 0.05mg / L, and nitrite is ≤ 0.04mg / L.

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