Application of oleic acid in improving disease resistance and meat quality of prawns
Patent Information
- Application Number
- CN202511491632.7
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-10-19
- Publication Date
- 2025-12-12
Smart Images
Figure CN121101080A_ABST
Abstract
Description
TECHNICAL FIELD
[0001] The application belongs to the technical field of disease prevention and control and aquaculture, and particularly relates to application of oleic acid in improving disease resistance and meat quality of prawns. BACKGROUND
[0002] P. vannamei is one of the most important aquaculture species in the world, and is favored by consumers for its delicious taste and rich nutrition. However, with the development of intensive farming mode, disease problems caused by multiple pathogens seriously threaten the sustainable development of prawn aquaculture. Among them, the bacterial disease caused by Vibrio parahaemolyticus (Vp) is particularly prominent, which not only leads to the decline of prawn growth performance and the deterioration of product quality, but also can cause large-scale death events and cause huge economic losses. More seriously, contaminated prawn products may cause acute gastroenteritis when consumed by humans, posing a significant food safety hazard. Vibrio parahaemolyticus
[0003] Currently, antibiotics are commonly used in aquaculture practice to prevent and treat bacterial diseases. However, long-term and improper use of antibiotics can lead to drug residues in biological tissues and the environment, harming the health of prawns, and even causing serious food safety problems. Therefore, developing new green feed that can replace antibiotics while improving the health level and product quality of prawns has become a key problem to be solved in prawn aquaculture.
[0004] Oleic acid (OA) is a monounsaturated fatty acid and is the main bioactive component of various medicinal plant extracts. Existing studies have shown that oleic acid, as an important nutrient, has multiple physiological functions in mammals, such as regulating energy homeostasis, improving lipid metabolism, anti-inflammatory, and enhancing insulin sensitivity. However, current research on oleic acid mainly focuses on mammalian systems, and is mostly based on cell models or direct injection methods. The application effect of dietary supplementation in aquaculture is not clear. SUMMARY
[0005] The purpose of the present application is to overcome the shortcomings of the prior art and provide a feed additive based on oleic acid and its application method to solve the disease prevention and control problem in prawn aquaculture and achieve significant improvement in farming quality.
[0006] Application of oleic acid in improving disease resistance and meat quality of prawns.
[0007] Adding 1% oleic acid in prawn feed can activate prawn defensive lipid droplets and enhance the in vitro antibacterial activity of lipid droplet proteins.
[0008] Adding 1% oleic acid in prawn feed can reduce the load of Vibrio parahaemolyticus in prawn hepatopancreas and blood lymph.
[0009] The addition of 1% oleic acid in the feed of prawns can reduce the tissue damage of hepatopancreas and intestinal tract caused by infection.
[0010] The addition of 1% oleic acid in the feed of prawns can improve the muscle texture of prawns, enhance the gumminess, chewiness and hardness, and make the muscle fiber arrangement more compact and the density higher.
[0011] A prawn feed added with 1% oleic acid.
[0012] The addition of 1% oleic acid in the feed of prawns can activate the defensive lipid droplets of prawns, enhance the in vitro antibacterial activity of lipid droplet protein, reduce the load of Vibrio parahaemolyticus in hepatopancreas and hemolymph, and reduce the tissue damage of hepatopancreas and intestinal tract caused by infection, thereby significantly improving the disease resistance of prawns. At the same time, the addition of 1% oleic acid can improve the muscle texture of prawns, enhance the gumminess, chewiness and hardness, and make the muscle fiber arrangement more compact and the density higher, effectively optimizing the eating quality of prawns. The present application provides a basis for the application of oleic acid as a green feed additive in the health breeding and quality improvement of prawns, and has important production practice value. BRIEF DESCRIPTION OF DRAWINGS
[0013] Figure 1 The result graph of the optimal additive amount of OA in Example 1 of the present application. After feeding prawns with feed containing different concentrations of OA for 42 days, the prawns were infected with Vibrio parahaemolyticus, and the survival rate after infection was calculated.
[0014] Figure 2 The result graph of the significant improvement of the antibacterial activity of lipid droplet protein by the addition of 1% OA in Example 2 of the present application; A is a representative plate image in the antibacterial experiment; B is a bacterial colony count graph.
[0015] Figure 3 The result graph of the removal of Vibrio parahaemolyticus in the hepatopancreas and hemolymph of prawns supplemented with 1% OA in Example 3 of the present application; A is the bacterial load in the hepatopancreas, and B is the bacterial load in the hemolymph.
[0016] Figure 4 The result graph of the reduction of tissue damage in the hepatopancreas and intestinal tract of prawns induced by Vibrio parahaemolyticus by the addition of 1% OA in Example 4 of the present application; A is a representative H&E stained section of hepatopancreas, with inflammatory cell infiltration (a↑), lumen dilation (b↑), and connective tissue damage (c↑); B is a representative H&E stained section of intestinal tract.
[0017] Figure 5 The result graph of the improvement of muscle texture of prawns by the addition of 1% OA in Example 5 of the present application; A is gumminess, B is chewiness, C is hardness, D is adhesiveness, E is elasticity, and F is the use of radar chart to present muscle texture parameters.
[0018] Figure 6 For Example 6 of the present application, the results of adding 1% OA to make prawn muscle fibers dense; A is the H&E staining image of muscle tissue (longitudinal section), B-D are the statistical graphs of muscle fiber diameter, muscle fiber area and muscle fiber density, respectively. DETAILED DESCRIPTION
[0019] In order to facilitate the understanding of the present application, the present application will be described more fully below. However, the present application can be realized in many different forms and is not limited to the embodiments described herein. On the contrary, the purpose of providing these embodiments is to make the disclosure of the present application more thorough and comprehensive.
[0020] Example 1 Preparation of feed and determination of optimal concentration Commercial feed uses commercially available products, specifically, South American white prawn compound feed (model: 8831) produced by Tongwei Co., Ltd.
[0021] 0.5%, 1% and 1.5% oleic acid (OA, purity ≥ 99%, from Beijing Solarbio Co., Ltd.) were added to the commercial feed to prepare three test feeds. The control feed uses standard commercial feed (without OA). The commercial prawn feed is fully ground into powder by an electric blender, then different proportions of oleic acid are added, and distilled water is added to make the mixture completely homogeneous. The feed is prepared by a pelletizer, then sealed and dried at 25°C for 12 hours, i.e. the preparation of the feed is completed, and stored at 4°C until use.
[0022] South American white prawns (4-6 g each) were randomly divided into 4 groups and fed with control feed without OA and test feed containing 0.5%, 1%, and 1.5% OA, respectively, for 42 consecutive days. The prawns were fed twice a day (8:00, 17:00) with a feeding amount of 5% of body weight. After the end of the feeding experiment, each group of prawns was injected with Vibrio parahaemolyticus (1 × 10 6 CFU) intraperitoneally, and the survival rate of each group of prawns was recorded every 12 hours.
[0023] The results of the detection are shown in Figure 1 The survival rate of all OA added groups was significantly higher than that of the control group, among which the survival rate of the 1% OA group (50%) and the 1.5% OA group (46.7%) was the highest. Considering the safety and cost-effectiveness of the feed, 1% OA was chosen as the optimal concentration for further study in the subsequent feeding experiment.
[0024] Example 2 Addition of 1% OA significantly improves the antibacterial activity of lipid droplet protein Next, the inventors investigated how the lipid droplets provide effective protection against invading bacteria. The key immune organ of shrimp, hepatopancreas, which has high abundance of lipid droplets, was chosen for isolation of lipid droplet proteins. The hepatopancreas tissue (1 g) was homogenized with 3 ml of homogenization buffer (25 mM Tris-HCl, pH 7.5, 100 mM KCl, 1 mM EDTA and 5 mM EGTA). The hepatopancreas homogenate was centrifuged at low speed (500 x g, 10 min, 4°C) and 2.5 ml of supernatant was mixed with equal volume of 2.5 M sucrose solution. The sample-sucrose mixture was loaded at the bottom of an ultracentrifuge tube, which was sequentially filled with 25%, 15%, 10% and 5% sucrose solutions from bottom to top, and a top layer solution (25 mM Tris-HCl pH 7.5, 1 mM EDTA and 5 mM EGTA) was added on top. The tube was centrifuged at 12,000 x g for 1 h at 4°C. The fraction enriched with lipid droplets at the top of the gradient was collected and centrifuged at 16,000 x g for 10 min at 4°C. Then, the lower phase was removed to purify the lipid droplets. For extraction of lipid droplet proteins, the purified lipid droplets were mixed with 1 ml of ice-cold acetone and stored at -20°C for 48 h. After three washes with cold acetone, the pellet was dissolved in 10 mM Tris-HCl (pH 7.5) to obtain LD proteins.
[0025] Subsequently, the antibacterial experiment was performed by incubating 50 μl of overnight bacterial culture (1 x 10 5 CFU / ml) with 20 μl of protein (1 mg / ml) at 37°C for 12 h. Subsequently, the mixture was diluted 10-fold and 100 μl of the dilution was spread on LB plates. After incubation for 12 h, the colonies were counted and the average value was calculated.
[0026] The results of the detection are shown in Figure 2 As shown, the lipid droplet proteins exhibited significant bactericidal ability against V. parahaemolyticus in vitro, and this ability was significantly enhanced after the addition of 1% OA in the feed. This result indicates that the addition of 1% OA in the feed enhances the antibacterial ability of the lipid droplets in shrimp, thereby providing effective antibacterial defense during the infection of Vibrio.
[0027] Example 3 Elimination of V. parahaemolyticus in the hepatopancreas and hemolymph of shrimp by the addition of 1% OA After the end of the feeding experiment, the shrimp fed with the control diet and the 1% OA diet were injected intraperitoneally with V. parahaemolyticus (1 x 10 6 CFU). After 1 h of injection, the hepatopancreas tissue and hemolymph samples were collected aseptically (6 per group). Subsequently, the hepatopancreas tissue was homogenized, and the homogenized hepatopancreas and hemolymph samples were sequentially diluted and spread on LB plates, which were incubated at 37°C until bacterial colonies appeared.
[0028] The results of the detection are shown in Figure 3As shown, the bacterial load in the hepatopancreas and hemolymph of shrimp in the 1% OA diet group was significantly reduced compared to shrimp on the control diet.
[0029] Example 4. Addition of 1% OA alleviates Vibrio parahaemolyticus infection-induced hepatopancreas and intestinal damage in shrimp Shrimp were injected with Vibrio parahaemolyticus as described in Example 2, and the hepatopancreas and intestinal tissues were collected 12 h post-injection. The tissue samples were fixed in 4% paraformaldehyde for 24 h, followed by ethanol dehydration, paraffin embedding, and sectioning. Hematoxylin and eosin (H&E) staining was performed, and morphological changes among groups were observed under a microscope (Olympus).
[0030] The results are shown in Figure 6. Figure 4 As shown, Vibrio parahaemolyticus infection induced significant pathological changes in the hepatopancreas of shrimp, including inflammatory cell infiltration, lumen dilation, and connective tissue damage, while the addition of 1% OA in the feed effectively alleviated the infection-induced hepatopancreas lesions. In addition, Vibrio parahaemolyticus infection also caused significant pathological changes in the intestinal tissue of shrimp, including intestinal wall thickening and inflammatory cell infiltration, while the addition of 1% OA effectively alleviated or even reversed this change.
[0031] Example 5. Addition of 1% OA improves shrimp muscle texture After 42 days of feeding, texture analysis of muscle was performed using a texture analyzer (XT Plus, Stable Micro Systems, UK). Muscle samples (12 per group) were processed into standardized test pieces (1 cm x 1 cm x 0.5 cm) and compressed with a cylindrical probe of 8 mm diameter (25 N range). TPA settings were: down speed 30 mm / min, compression level 50%, interval 2 s. Parameters such as adhesiveness, chewiness, hardness, cohesiveness, and springiness were calculated from the TPA force-time curve.
[0032] The results are shown in Figure 7. Figure 5 As shown, the addition of 1% OA in the feed significantly optimized the adhesiveness, chewiness, and hardness of shrimp meat, effectively improving the eating quality of shrimp.
[0033] Example 6. Addition of 1% OA makes shrimp muscle microstructure more compact After 42 days of feeding, muscle histology analysis was performed to further evaluate the effect of 1% OA added in the feed on the muscle quality of shrimp. Muscle tissue of shrimp was collected and fixed in 4% paraformaldehyde for 24 hours, then dehydrated in graded ethanol. Paraffin-embedded, sections of 7 μm thickness were prepared, followed by hematoxylin and eosin (H&E) staining. Optical microscopy (Olympus) was used to observe the muscle microstructure, and ImageJ software was used to quantify muscle fiber characteristics, including muscle fiber diameter, area, and density.
[0034] The detection results are shown in Table 1. Figure 6 As shown in Table 1, after adding 1% OA in the feed, the muscle fibers of the shrimp are arranged more closely, the muscle fiber diameter and area are obviously reduced, and the muscle fiber density is also higher. The above-described embodiments only express several embodiments of the present application, and the description is more specific and detailed, but it should not be understood as a limitation on the scope of the patent. It should be noted that for ordinary skilled persons in the art, without departing from the concept of the present application, several modifications and improvements can be made, which are within the scope of protection of the present application. Therefore, the protection scope of the patent of the present application should be subject to the appended claims.
Claims
1. Application of oleic acid in improving disease resistance and meat quality of shrimp.
2. The application according to claim 1, characterized in that, Adding 1% oleic acid to shrimp feed can activate shrimp defensive lipid droplets and enhance the in vitro antibacterial activity of lipid droplet proteins.
3. The application according to claim 1, characterized in that, Adding 1% oleic acid to shrimp feed can reduce the viral load of Vibrio parahaemolyticus in the hepatopancreas and hemolymph of shrimp.
4. The application according to claim 1, characterized in that, Adding 1% oleic acid to shrimp feed can reduce damage to the hepatopancreas and intestinal tissues caused by shrimp infection.
5. The application according to claim 1, characterized in that, Adding 1% oleic acid to shrimp feed can improve shrimp muscle texture, enhance adhesiveness, chewiness, and firmness, and make muscle fibers more tightly packed and denser.
6. A shrimp feed, characterized in that, The shrimp feed contains 1% oleic acid.