Spartina alterniflora crude polysaccharide as well as preparation method and application thereof
By using coarse polysaccharides of Eels as feed additives, the problems of easy stress and high incidence in eel farming were solved, and its antioxidant performance and breeding success rate were significantly improved.
Patent Information
- Application Number
- CN202411942510.0
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2024-12-27
- Publication Date
- 2025-05-13
AI Technical Summary
Eels are prone to stress and have a high incidence rate during breeding, resulting in large-scale deaths. The existing technology is difficult to effectively improve its antioxidant performance.
By extracting coarse polysaccharides from the Eel and using it as an eel feed additive, the serum and intestinal antioxidant capacity of eels are improved.
It effectively improves the serum and intestinal antioxidant capacity of eels, significantly reduces the mortality rate during breeding, and increases the value of feed.
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Abstract
Description
Technical Field
[0001] The invention relates to the field of aquaculture feed additives, and in particular to a crude polysaccharide of Spartina alterniflora and a preparation method and application thereof. Technical Background
[0002] Spartina alterniflora is a perennial herbaceous plant of the genus Spartina of the Poaceae family, originating in the Atlantic coast, and was widely introduced into my country in the 1980s to protect the shore beaches. Due to its strong adaptability and reproductive ability, it occupies large tracts of mudflats, compresses the living space of local plants to a great extent, and changes the biodiversity of benthic organisms, birds, etc. The harm of Spartina alterniflora is diverse, including destroying the habitat of marine organisms, destroying the coastal wetland ecosystem, polluting water quality to block rivers, threatening offshore aquaculture, etc. In order to prevent and control the large-scale expansion of Spartina alterniflora, the main prevention and control means in China at present is physical mowing technology, which has the characteristics of wide application range, low cost of governance, good governance effect, and less damage to the environment than chemical control methods. And how to use the Spartina alterniflora straw obtained by harvesting, it is still mainly concentrated on fertilizer and fuel at present, and the development of its feed is still less.
[0003] Yellow eel (Monopterus albus) is widely sought after by consumers for its delicious meat and rich nutrition, and the current production is in short supply. One of the important reasons restricting the development of yellow eel farming industry is that it is easy to be stressed and has a high incidence rate, which easily causes large-scale deaths during the farming process.
[0004] Therefore, it is particularly important to find a plant-derived feed additive to improve the antioxidant properties of eels. Summary of the invention
[0005] The present invention first provides a method for extracting crude polysaccharides from Spartina alterniflora, comprising the following steps:
[0006] 1) drying the Spartina alterniflora, and crushing the Spartina alterniflora through a 60-mesh sieve to obtain Spartina alterniflora coarse powder;
[0007] 2) Degreasing by Soxhlet extraction, the degreasing conditions are reflux petroleum ether at 50°C for 7-8 hours; after degreasing, air-dry in a fume hood to evaporate the residual petroleum ether;
[0008] 3) Weigh the coarse powder of Spartina alterniflora, add water, the solid-liquid ratio is 1g:10-30mL, extract at 80-100°C for 2-4h, cool to room temperature and centrifuge at 4000rpm for 10min, collect the supernatant, repeat the extraction of the residue, combine the extracts and concentrate them by rotation;
[0009] 4) Add anhydrous ethanol to the concentrated extract to make the final ethanol concentration of 60%, stir thoroughly and place in a 4°C refrigerator to precipitate for 12-36 hours, centrifuge (the centrifugal condition for ethanol precipitation is 6000 rpm for 15 minutes), discard the supernatant and collect the precipitate to obtain the crude polysaccharide.
[0010] Preferably, the material-liquid ratio of water extraction in step 3) is 1 g:20 mL, and extraction is carried out in a constant temperature water bath at 90° C. for 3 h.
[0011] An appropriate amount of distilled water was added to the collected crude polysaccharide of Spartina alterniflora to dissolve it, and the residual impurities were removed again. The crude polysaccharide of Spartina alterniflora was freeze-dried using a vacuum freeze dryer to obtain a purer crude polysaccharide of Spartina alterniflora. The centrifugal condition for freeze-drying to remove impurities was centrifugation at a speed of 8000 rpm for 10 minutes.
[0012] The present invention also provides the crude polysaccharide of Spartina alterniflora prepared by the method.
[0013] The present invention further provides the application of the crude polysaccharide of Spartina alterniflora, which is added as an additive to the feed of rice field eel;
[0014] The addition ratio of crude polysaccharide of Spartina alterniflora to the eel feed is 0.5-1.5% (weight ratio), and the eel feed added with crude polysaccharide of Spartina alterniflora is fed daily according to 3-5% of the total weight of the eel.
[0015] Compared with the prior art, the present invention has the following advantages and effects:
[0016] (1) The large amount of Spartina alterniflora harvested by preventing the expansion of Spartina alterniflora can be effectively utilized as a resource, which not only protects the local ecological environment balance, but also increases its value as feed;
[0017] (2) The crude polysaccharides in Spartina alterniflora can be effectively extracted by hot water extraction, and the operation method is simple and effective;
[0018] (3) By using the extracted crude polysaccharide of Spartina alterniflora as an eel feed additive, the antioxidant capacity of the eel serum and intestine can be effectively improved. DETAILED DESCRIPTION
[0019] The following examples are only used to further illustrate the content of the present invention, but should not be construed as limiting the present invention. Without departing from the spirit and essence of the present invention, modifications or substitutions made to the method, steps or conditions of the present invention all belong to the scope of the present invention. The experimental methods and reagents without specifying the specific conditions in the examples are all in accordance with the conventional conditions in the art.
[0020] The Spartina alterniflora used below were all harvested from the coastal beaches of Chongming District, Shanghai. The eels used were all farmed by the Zhuangxing Comprehensive Experimental Station of Shanghai Academy of Agricultural Sciences. The eel powder feed used was fish meal purchased from the market.
[0021] Example 1: Extraction of crude polysaccharides from Spartina alterniflora
[0022] (1) Drying and crushing
[0023] Rinse the Spartina alterniflora with distilled water, dry in an oven at 55°C, and grind with a wall-breaking machine to pass through a 60-mesh sieve to obtain coarse Spartina alterniflora powder.
[0024] (2) Degreasing
[0025] The degreasing was carried out by Soxhlet extraction at 50°C with petroleum ether reflux for 7-8 hours. After degreasing, the samples were air-dried in a fume hood to evaporate the residual petroleum ether.
[0026] (3) Hot water extraction
[0027] Weigh a certain amount of coarse powder and transfer it to a beaker. Add a certain volume of distilled water to make the solid-liquid ratio 1g:20mL and stir thoroughly. Extract in a 90℃ constant temperature water bath for 3h, then take out and cool to room temperature. Centrifuge at 4000rpm for 10min, collect the supernatant by filtering with gauze, repeat the extraction of the filter residue 3 times, combine the extracts, and concentrate to 1 / 3 of the original volume using a rotary evaporator.
[0028] (4) Ethanol precipitation
[0029] Anhydrous ethanol was added to the concentrated extract to make the final ethanol volume concentration of 60%, and the mixture was stirred thoroughly to ensure full contact between the ethanol solution and the extract. The mixture was placed in a 4°C refrigerator for 24 hours to allow the polysaccharide to fully precipitate. The mixture was centrifuged at 6000 rpm for 15 minutes to separate the precipitate and the solution. The supernatant was discarded and the precipitate was collected as the crude polysaccharide. The precipitate was spread on a plate and the residual ethanol was completely evaporated in a ventilated place.
[0030] (5) Freeze drying
[0031] An appropriate amount of distilled water was added to the polysaccharide to dissolve it, and the remaining impurities were removed by centrifugation at a speed of 8000 rpm for 10 minutes. The supernatant was collected and freeze-dried using a vacuum freeze dryer to obtain the crude polysaccharide of Spartina alterniflora.
[0032] Extraction rate of crude polysaccharide of Spartina alterniflora = weight of crude polysaccharide of Spartina alterniflora / weight of crude powder of Spartina alterniflora ╳ 100%
[0033] In this example, the extraction rate of crude polysaccharide from Spartina alterniflora was 20%.
[0034] The crude Spartina alterniflora polysaccharide prepared in this example was detected by phenol method, wherein the content of active polysaccharide was 25%.
[0035] Example 2: Utilization of Spartina alterniflora crude polysaccharide
[0036] A total of 120 eels were randomly divided into 4 groups, with 3 replicates in each group and 10 eels in each replicate. The average body weight of the eels was 27.15±0.44g.
[0037] The crude polysaccharide of Spartina alterniflora extracted in Example 1 was added to fish meal (control group was 0%) according to the weight ratio of 0.5%, 1% and 1.5%, respectively, and a paste feed was formed after adding water and stirring to feed the eel every day. The daily feeding amount during the experiment was 4-5% of the body weight of the eel, the water temperature during the experiment was 27±2°C, pH 7.3±0.2, dissolved oxygen ≥5.8mg / L, 1 / 2 of the water was replaced every day, and the experimental period was set to 6 weeks. After the experiment, fasting for 24h, 2 eels were randomly selected in each repetition to take serum and intestinal tissue for detection using enzyme activity reagent detection kit (purchased from Nanjing Jiancheng Bioengineering Institute).
[0038] Data were calculated using Microsoft Excel and analyzed using SPSS 22.0 software. Significant differences between groups were assessed using one-way analysis of variance and Duncan's multiple comparisons. All results are presented as mean ± standard deviation, and P values < 0.05 were considered statistically significant. Specific results are shown in Tables 1 and 2 below.
[0039] Table 1 Antioxidant capacity of eel serum after adding crude polysaccharide of Spartina alterniflora for 6 weeks
[0040]
[0041] Table 2 Antioxidant capacity of the intestine of the eel after adding crude polysaccharide of Spartina alterniflora for 6 weeks
[0042]
[0043] The test results are shown in Tables 1 and 2. Adding crude polysaccharides of Spartina alterniflora to feed can effectively improve the antioxidant capacity in serum and intestines, and the best effect is shown at an addition ratio of 1.5%.
Claims
1. A method for extracting crude polysaccharides from Spartina alterniflora, characterized in that The steps include: 1) drying the Spartina alterniflora, and crushing the Spartina alterniflora through a 60-mesh sieve to obtain Spartina alterniflora coarse powder; 2) Degreasing by Soxhlet extraction, the degreasing conditions are reflux of petroleum ether at 50°C for 7 to 8 hours; 3) Weigh the coarse powder of Spartina alterniflora, add water, the solid-liquid ratio is 1g:10-30mL, extract at 80-100°C for 2-4h, cool to room temperature and centrifuge at 4000rpm for 10min, collect the supernatant, repeat the extraction of the residue, combine the extracts and concentrate them by rotation; 4) adding anhydrous ethanol to the concentrated extract to make the final ethanol volume concentration be 50-70%, stirring thoroughly and placing in a 4°C refrigerator to precipitate for 12-36 hours, and discarding the supernatant after centrifugation to collect the precipitate as the crude polysaccharide.
2. The method for extracting crude polysaccharides from Spartina alterniflora according to claim 1, wherein the solid-liquid ratio of the water extraction in step 3) is 1 g:20 mL, and the extraction is carried out in a constant temperature water bath at 90°C for 3 hours.
3. The crude polysaccharide of Spartina alterniflora prepared by any method of claim 1 or 2.
4. The use of the crude polysaccharide of Spartina alterniflora according to claim 3, characterized in that Add it as an additive to eel feed.
5. The use of the crude polysaccharide of Spartina alterniflora according to claim 4, wherein the addition ratio of the crude polysaccharide of Spartina alterniflora to the eel feed is 0.5-1.5% by weight.