Enteromorpha feed additive, and preparation method and application thereof

Abstract

This application relates to the field of feed additive technology, specifically disclosing a seaweed feed additive, its preparation method, and its application. The preparation method of the seaweed feed additive provided by this application includes the following steps: seaweed pretreatment, high-temperature fermentation, and functional fermentation; specifically, freshly harvested seaweed is washed sequentially with seawater and fresh water, then dehydrated to a moisture content of 75-85%; the seaweed is then cut into sections, crushed, and pulped, and auxiliary materials are added to obtain a mixed seaweed slurry; then, the mixed seaweed slurry is subjected to high-temperature fermentation at 65-80℃ using high-temperature compound bacteria; subsequently, the mixed slurry is subjected to functional fermentation at 25-35℃ using functional compound bacteria. The preparation method of the seaweed feed additive provided by this application has advantages such as low processing cost and simple operation, and the effective components of the obtained seaweed feed additive have a high utilization rate, which has a good promoting effect on animal growth.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of feed additives, in particular to a Enteromorpha prolifera feed additive and a preparation method and application thereof. BACKGROUND

[0002] Enteromorpha prolifera, also known as "strip of moss" and "moss vegetables", is a kind of algae plant of green algae class and Ulvaceae, which grows in clusters, with obvious main branches and slender branches, and the height can reach 1 meter. Enteromorpha prolifera is a kind of large green algae with high protein, high dietary fiber, polysaccharide, rich in mineral elements and low fat, which grows widely in the sea of the world, and has a particularly strong natural reproduction capacity. Every time it is in the flourishing season, a large amount of Enteromorpha prolifera will float and gather on the shore, which not only blocks the waterway, but also destroys the marine ecosystem, causing great threat to the coastal fishery, tourism and ecological environment.

[0003] There is a shortage of feed protein in China, and it is a hot research topic to expand the resource source of feed protein substitution in all directions. Enteromorpha prolifera is rich in carbohydrates, protein, crude fiber, amino acids, fat, vitamins and various minerals, and has the potential to become a high-quality feed additive and feed protein substitute. In the current research progress of Enteromorpha prolifera feed, most of them are dehydrated and dried, and then fermented by microbial technology to prepare feed additives. However, due to the problems of Enteromorpha prolifera such as concentrated outbreak period, high water content after salvage, high salt content, short storage period, easy to rot and stink, on the one hand, the treatment cost of the related pretreatment process is high, the operation is complex, and secondary pollution is easy to cause; on the other hand, the drying process is easy to cause the loss of nutritional components in Enteromorpha prolifera, which makes the utilization rate of Enteromorpha prolifera poor, and further leads to the low nutritional components in Enteromorpha prolifera feed additive, and the promotion of Enteromorpha prolifera feed additive is limited. SUMMARY

[0004] In order to reduce the preparation cost of Enteromorpha prolifera feed additive and improve the utilization rate of Enteromorpha prolifera, the present application provides an Enteromorpha prolifera feed additive and a preparation method and application thereof.

[0005] In the first aspect, the present application provides a preparation method of Enteromorpha prolifera additive, which adopts the following technical scheme:

[0006] A preparation method of Enteromorpha prolifera additive, comprising the following steps:

[0007] Enteromorpha prolifera pretreatment: the collected fresh Enteromorpha prolifera is washed with seawater and fresh water in sequence, and then dehydrated to have a water content of 75-85%; then the Enteromorpha prolifera is cut into sections, crushed and pulped, and then auxiliary materials are added to obtain Enteromorpha prolifera mixed slurry;

[0008] High-temperature fermentation: high-temperature fermentation of the Enteromorpha mixed slurry at 65-80°C using high-temperature composite bacteria; the high-temperature composite bacteria are composed of Bacillus subtilis, Bacillus thermophilus, Bacillus licheniformis, yeast and lactic acid bacteria with a ratio of viable bacterial count of (1-1.2):(0.8-1.1):(1.2-1.5):(1.2-1.5):(0.8-1.2).

[0009] The present application provides a low-cost and rapid preparation process of Enteromorpha feed additive. In the Enteromorpha pretreatment step of the preparation process, fresh Enteromorpha is first washed with offshore seawater to remove impurities such as sand, and then washed with fresh water to remove salt in the Enteromorpha. The clean water after washing is not polluted and can be directly discharged into the sea. Therefore, the above pretreatment process can save the cost of transporting Enteromorpha from collection to processing plant, reduce the use of fresh water resources and reduce sewage discharge. In addition, in the preparation process of the present application, high-temperature fermentation of Enteromorpha mixed slurry at 65-80°C using high-temperature composite bacteria composed of Bacillus subtilis, Bacillus thermophilus, Bacillus licheniformis, yeast and lactic acid bacteria can rapidly degrade the macromolecular substances such as polysaccharides, proteins and unsaturated fatty acids in Enteromorpha into small molecular substances, release the nutrients inside Enteromorpha, and effectively improve the utilization rate of nutrients in Enteromorpha; on the other hand, it can kill other bacteria other than high-temperature composite bacteria.

[0010] In a specific embodiment, the ratio of viable bacterial count of Bacillus subtilis, Bacillus thermophilus, Bacillus licheniformis, yeast and lactic acid bacteria in the high-temperature composite bacteria can be 1:1:1.2:1.2:1, 1:0.8:1.5:1.2:0.8, 1.2:1:1.2:1.2:1.2, 1:0.8:1.2:1.5:1 or 1:1.1:1.2:1.5:0.8.

[0011] Optionally, the addition amount of the high-temperature composite bacteria is 0.02-0.05% of the mass of the Enteromorpha mixed slurry, and the effective viable bacterial count of the high-temperature composite bacteria is ≥100 million / g.

[0012] In a specific embodiment, the addition amount of the high-temperature composite bacteria is 0.02%, 0.03%, 0.05% or 0.07% of the mass of the Enteromorpha mixed slurry.

[0013] In a specific embodiment, the effective viable bacterial count of the high-temperature composite bacteria is 150 million / g.

[0014] Further, the addition amount of the high-temperature composite bacteria is 0.03-0.05% of the mass of the Enteromorpha mixed slurry.

[0015] In the present application, the addition amount of the high-temperature composite bacteria affects the degradation rate and degree of macromolecular substances in Enteromorpha and the flavor of Enteromorpha. It is found through experiments that, when the addition amount of the high-temperature composite bacteria is controlled in the range of 0.03-0.05% by mass of the Enteromorpha mixed slurry, the nutritional value of the obtained Enteromorpha feed additive is higher, and the growth performance and immune function of animals are more significantly promoted. In addition, the Enteromorpha feed additive obtained under the above addition amount has a stronger aroma and better flavor, and can play a good feeding role when added to feed.

[0016] Optionally, the preparation method further comprises functional fermentation, and the specific steps are as follows: cooling the mixture obtained through high-temperature fermentation to 25-35℃, then adding functional composite bacteria and fermenting at the above temperature;

[0017] The functional composite bacteria are composed of Lactobacillus plantarum, Candida utilis and Aspergillus niger with a ratio of 1: (0.8-1.5) : (0.3-0.6) in terms of viable cell count.

[0018] The effective viable cell count of the functional composite bacteria is ≥2.0 billion / g.

[0019] In the present application, the functional composite bacteria are further inoculated into the mixture obtained through high-temperature fermentation, which can further degrade macromolecular substances in the mixture, improve the palatability of the feed additive, and convert Enteromorpha polysaccharides into protein, acid-soluble protein and other nutrients such as protease activity in the feed additive under the action of microorganisms. On the other hand, the growth performance and immune capacity of animals consuming the feed can be improved through the reproduction and metabolic products of the functional composite bacteria.

[0020] The functional composite bacteria of the present application, Lactobacillus plantarum, can promote the absorption of nutrients, improve the balance of intestinal microbial flora, and enhance the immune capacity of the body by producing various bacteriostatic substances such as organic acids, bacteriocins, hydrogen peroxide and diacetyl. Candida utilis can produce edible protein from macromolecular Enteromorpha polysaccharides and other substances in the mixture, thereby increasing the protein content of the additive. In addition, Candida utilis has rich enzymes in the body, which can degrade cellulose and other substances in Enteromorpha into small molecular substances, facilitating absorption and improving palatability. The cellulase, hemicellulase and pectinase produced by Aspergillus niger can decompose cellulose and pectin in Enteromorpha, release nutrients therein, and convert relatively large molecular compounds into small molecular compounds, thereby fully exerting the nutritional value of the feed and facilitating absorption by animals. The cellulase, hemicellulase and pectinase produced by Aspergillus niger can decompose cellulose and pectin in Enteromorpha, release nutrients therein, and convert relatively large molecular compounds into small molecular compounds, thereby fully exerting the nutritional value of the feed and facilitating absorption by animals. The cellulase, hemicellulase and pectinase produced by Aspergillus niger can decompose cellulose and pectin in Enteromorpha, release nutrients therein, and convert relatively large molecular compounds into small molecular compounds, thereby fully exerting the nutritional value of the feed and facilitating absorption by animals.

[0021] In a specific embodiment, the ratio of viable cell numbers of Lactobacillus plantarum, Candida utilis and Aspergillus niger in the functional complex bacteria can be 1:1:0.5, 1:1.5:0.5, 1:0.8:0.6 or 1:1:1.

[0022] In a specific embodiment, the effective viable cell number of the functional complex bacteria is 200 million / g.

[0023] In the present application, the addition amount of the functional complex bacteria is 0.1-0.3% of the mixed Enteromorpha slurry.

[0024] Optionally, the high-temperature fermentation time is 8-12h, and the functional fermentation time is 12-24h.

[0025] Optionally, the Enteromorpha is cut and crushed to obtain Enteromorpha slurry, and auxiliary materials are added to prepare mixed Enteromorpha slurry, and the water content of the mixed Enteromorpha slurry is 50-65%; the mixed Enteromorpha slurry is composed of Enteromorpha slurry and auxiliary materials in a weight ratio of 1:(0.2-0.4); and the auxiliary materials are selected from one or more of the group consisting of wheat bran, corn husk and flammulina velutipes residue.

[0026] Optionally, the preparation method further comprises adjusting the moisture content, crushing and sieving, and the specific steps are as follows: after the functional fermentation is completed, the temperature of the mixed material is increased to 55-60℃, so that the water content of the mixed material is ≤15%; then the mixed material is cooled, crushed and sieved through a 5-20 mesh sieve to obtain the Enteromorpha feed additive.

[0027] In the present application, the water content in the Enteromorpha feed additive at the above-mentioned temperature can ensure the activity of beneficial bacteria in Enteromorpha, and the Enteromorpha feed additive with the above-mentioned water content can be stored for a long time, is not easy to deteriorate, and is convenient to use.

[0028] The water content in the Enteromorpha feed additive is controlled in the above-mentioned range,

[0029] In a second aspect, the present application provides an Enteromorpha feed additive prepared by the preparation method of the Enteromorpha feed additive.

[0030] The present application uses the preparation method of the Enteromorpha feed additive to prepare an Enteromorpha feed additive, which contains a large amount of small molecule Enteromorpha polysaccharide, vitamins, small molecule protein, polypeptide, amino acid, etc., can improve the growth performance of animal body, promote the development of intestinal tract of animal body, improve the intestinal immune response, and regulate the intestinal microecology.

[0031] In a third aspect, the present application provides the application of the Enteromorpha feed additive in poultry feed, aquatic feed and pig feed.

[0032] Optionally, the added amount of the Enteromorpha feed additive in the poultry feed is 1-3% by mass.

[0033] In a specific embodiment, the added amount of the Enteromorpha feed additive in the poultry feed is 2% by mass.

[0034] Optionally, the added amount of the Enteromorpha feed additive in the aquatic feed is 2-5% by mass.

[0035] In summary, the present application has the following beneficial effects:

[0036] 1. In the Enteromorpha pretreatment process of the present application, the new Enteromorpha is first preliminarily cleaned with offshore seawater, and then cleaned with fresh water, which can save the cost of transporting the Enteromorpha from collection to the processing plant, and also reduce the use of fresh water resources and sewage discharge.

[0037] 2. The high-temperature composite bacteria are first used to ferment the Enteromorpha mixed slurry, which can quickly degrade the macromolecular substances such as polysaccharides, proteins, and unsaturated fatty acids in the Enteromorpha into small molecular substances, release the nutrients inside the Enteromorpha, and effectively improve the utilization rate of the nutrients in the Enteromorpha; then the functional composite bacteria are used for functional fermentation, which further improves the activity of acid-soluble protein, crude protein, and protease in the feed additive, and the growth performance and immune capacity of the animal body through the reproduction and metabolic products of the functional composite bacteria.

[0038] 3. The Enteromorpha feed additive is added to the poultry feed in an amount of 1-3% by mass, which can improve the appetite of broilers, provide sufficient nutrients for the broilers, promote the development of the intestinal tract of the animal body, and enhance the growth performance and immune function of the animal, so that the broilers can fully convert the feed into chicken meat, thereby increasing the breeding benefit. BRIEF DESCRIPTION OF DRAWINGS

[0039] Figure 1 is a preparation method flowchart of the Enteromorpha feed additive provided by the present application. DETAILED DESCRIPTION

[0040] The present application provides an Enteromorpha feed additive, and the preparation method thereof comprises the following steps:

[0041] (1) Enteromorpha pretreatment: The collected fresh Enteromorpha was washed with seawater for 2-3 times and fresh water for 1-2 times, and then dehydrated by a mechanical centrifugal device (purchased from Jiangsu Younaite Biological Technology Co., Ltd., model LWS450) to reduce the water content of Enteromorpha to 75-85%; then the Enteromorpha was cut, crushed, and pulped using a pulping machine (purchased from Ningbo Kaicheng Ecological Technology Co., Ltd., model KCPS-15) to obtain Enteromorpha slurry; the Enteromorpha slurry and the auxiliary material were mixed in a weight ratio of 1:(0.2-0.4) and added to a biochemical treatment machine (BGB-SCZ-7500 biochemical treatment machine developed by Beijing Jiaboxin Biological Technology Co., Ltd.), and the water content was adjusted to 50-65% to obtain Enteromorpha mixed slurry; wherein the auxiliary material is selected from one or more of wheat bran, corn husk, and shiitake mushroom residue, and the fineness of the auxiliary material is 5-20 mesh.

[0042] (2) High-temperature fermentation: Start the solid fermentation device, turn on the heating device, air circulation system and stirring system, gradually heat the mixture to 65-80°C, keep constant temperature and stirring, inoculate high-temperature composite bacteria, and keep constant temperature and stirring for 8-12h, during which the air circulation system continuously sends fresh air into the solid fermentation bin to ensure sufficient oxygen during fermentation; wherein the high-temperature composite bacteria are added in an amount of 0.02-0.05% by mass of the Enteromorpha mixed slurry, the effective viable count of the high-temperature composite bacteria is ≥1.0 billion / g, and the high-temperature composite bacteria are composed of Bacillus subtilis, Bacillus thermophilus, Bacillus licheniformis, yeast and lactic acid bacteria in a ratio of (1-1.2):(0.8-1.1):(1.2-1.5):(1.2-1.5):(0.8-1.2) by viable count.

[0043] (3) Functional fermentation: After high-temperature fermentation, control the water content in the fermentation bin to 35-45%, gradually cool to 25-35°C, close the air circulation system, inoculate functional composite bacteria for the second time, and keep constant temperature and stirring for 12-24h; the functional composite bacteria are composed of Lactobacillus plantarum, Candida utilis and Aspergillus niger in a ratio of 1:(0.8-1.5):(0.3-0.6) by viable count; the effective viable count of the functional composite bacteria is ≥2.0 billion / g.

[0044] (4) Adjusting the moisture content, crushing and screening: After functional fermentation, increase the ventilation volume to further reduce the water content of the material to ≤15%, then stop heating and gradually reduce the material in the fermentation bin to room temperature by air circulation system; then the material is crushed and transferred to a roller screen with a mesh size of 5-20 mesh to obtain a feed additive.

[0045] In the present application, the preservation number of Bacillus subtilis is CCTCC NO: M 2020296; the preservation number of Bacillus thermophilus is CCTCC NO: M 2013537; the preservation number of Bacillus licheniformis is CGMCC No. 8718; the preservation number of Saccharomyces is CCTCC NO: M 2011108; the preservation number of Lactobacillus is CGMCC NO: 4483; the preservation number of Lactobacillus plantarum is CGMCC NO. 16436; the preservation number of Candida utilis is CGMCC No. 7691; the preservation number of Aspergillus niger is CGMCC No. 2715; and the rest of the raw materials and the like can be obtained by commercial purchase.

[0046] The present application will be further described in detail in the following combined with the embodiments, performance detection tests and the description of the accompanying drawings.

[0047] Example 1

[0048] Example 1 provides a Enteromorpha feed additive, and the preparation method thereof comprises the following steps:

[0049] (1) Enteromorpha pretreatment: the collected fresh Enteromorpha (Enteromorpha linza) is washed with seawater for 3 times and fresh water for 2 times near the sea, and then is dewatered by a mechanical centrifugal device to make the water content of the Enteromorpha be about 75%; then the Enteromorpha is cut into sections, crushed and pulped to obtain Enteromorpha slurry; the Enteromorpha slurry and the pine mushroom residue are mixed according to a weight ratio of 1:0.3 and are added into a solid fermentation device, and the water content is adjusted to 60% to obtain Enteromorpha mixed slurry for standby.

[0050] (2) High-temperature fermentation: the solid fermentation device is started, and the heating device, the air circulation system and the stirring system are turned on; the mixed material is gradually heated to 70℃, and the constant temperature and stirring are maintained; the high-temperature compound bacteria are inoculated in an amount of 0.03% of the mass of the Enteromorpha mixed slurry, and the constant temperature stirring is maintained for 10 hours; during the process, the air circulation system continuously sends fresh air into the solid fermentation bin to make there be sufficient oxygen in the fermentation process; wherein the effective viable count of the high-temperature compound bacteria is 1.5 billion / g, and the high-temperature compound bacteria are composed of Bacillus subtilis, Bacillus thermophilus, Bacillus licheniformis, Saccharomyces and Lactobacillus with a ratio of 1:1:1.2:1.2:1.

[0051] (3) Functional fermentation: after the high-temperature fermentation is completed, the water content in the fermentation bin is controlled to be about 40%, and the temperature is gradually lowered to 30℃, and the air circulation system is closed; the functional compound bacteria are inoculated in an amount of 0.2% of the mass of the Enteromorpha mixed slurry, and the constant temperature stirring is maintained for 20 hours; the functional compound bacteria are composed of Lactobacillus plantarum, Candida utilis and Aspergillus niger with a ratio of 1:1:0.5; and the effective viable count of the functional compound bacteria is 2.0 billion / g.

[0052] (4) Adjusting moisture, crushing, and screening: after the functional fermentation is completed, the temperature of the bin is increased to 60℃, and the ventilation volume is increased to further reduce the moisture content of the material to 13%, and then the heating is stopped and the material in the fermentation bin is gradually reduced to normal temperature through the air circulation system; then the material is crushed and transferred to a roller screen, the roller screen has a mesh size of 5 meshes, and a feed additive is obtained.

[0053] Index detection

[0054] The crude protein content, acid-soluble protein content, crude fiber content, crude fat content, and other indexes of the material before and after the fermentation of Example 1 are detected, and the detection results are as shown in Table 1:

[0055] The detection before the fermentation is the conversion of each nutritional index of Enteromorpha prolifera mixed pulp to 15% moisture content; the detection after the fermentation is the fermented material obtained after the functional fermentation.

[0056] The detection method of the crude protein content refers to GB / T 6432-2018; the detection method of the acid-soluble protein content refers to NY / T3801-2020; the detection method of the crude fiber content refers to GB / T 6434-2006; the detection method of the crude fat content refers to GB / T6433-2006; and the detection method of the crude ash content refers to GB / T 6438-2007.

[0057] Table 1: Index evaluation results of the material before and after the fermentation of Example 1

[0058] Indicators Before fermentation After fermentation Crude protein content (%) 19.32 23.14 Acid soluble protein content (%) 2.15 4.67 Crude fat content (%) 1.42 1.12 Crude fiber content (%) 16.83 13.2 Crude ash 9.8 8.6

[0059] As can be seen from Table 1, after the high-temperature fermentation and the functional fermentation of the Enteromorpha prolifera mixed pulp, the crude protein and the acid-soluble protein in the fermented material are obviously improved, and in particular, the proportion of the acid-soluble protein in the crude protein is increased from 11.1% before the fermentation to 20.2%. In addition, the crude fiber and ash indexes are also reduced to different degrees. Therefore, it is proved that the process of high-temperature fermentation + functional fermentation provided by the present application has obvious effects on improving the nutritional value and palatability of the feed additive.

[0060] Example 2

[0061] Example 2 provides an Enteromorpha prolifera feed additive, and a preparation method thereof includes the following steps:

[0062] (1) Enteromorpha prolifera pretreatment: the collected fresh Enteromorpha prolifera is washed with seawater for 3 times and fresh water for 2 times, and then dehydrated by a mechanical centrifugal device to make the water content of the Enteromorpha prolifera about 75%; then the Enteromorpha prolifera is cut, crushed, and pulped to obtain Enteromorpha prolifera slurry; the Enteromorpha prolifera slurry and the golden needle mushroom residue are mixed according to a weight ratio of 1:0.2 and added to a solid fermentation device, and the water content is adjusted to 64% to obtain Enteromorpha prolifera mixed slurry for standby.

[0063] (2) High-temperature fermentation: start the solid fermentation device, turn on the heating device, air circulation system and stirring system, gradually heat the mixture to 75°C, keep constant temperature and stirring, inoculate high-temperature composite bacteria in an amount of 0.03% of the mixed Enteromorpha prolifera slurry, keep constant temperature and stirring for 8 hours, during which the air circulation system continuously sends fresh air into the solid fermentation bin to ensure sufficient oxygen during fermentation; wherein the effective viable count of the high-temperature composite bacteria is 150 million / g, and the high-temperature composite bacteria are composed of Bacillus subtilis, Bacillus thermophilus, Bacillus licheniformis, yeast and lactic acid bacteria with a viable count ratio of 1:1:1.2:1.2:1.

[0064] (3) Functional fermentation: after the high-temperature fermentation is completed, control the moisture in the fermentation bin at about 40%, gradually cool to 30°C, turn off the air circulation system, inoculate functional composite bacteria in an amount of 0.2% of the mixed Enteromorpha prolifera slurry, and keep constant temperature and stirring for 18 hours; the functional composite bacteria are composed of Lactobacillus plantarum, Candida utilis and Aspergillus niger with a viable count ratio of 1:1:0.5; the effective viable count of the functional composite bacteria is 200 million / g.

[0065] (4) Adjusting moisture, crushing and screening: after the functional fermentation is completed, increase the ventilation volume to further reduce the moisture content of the material to 13%, then stop heating and gradually reduce the material in the fermentation bin to room temperature through the air circulation system; then crush the material and transfer it to a roller screen with a screen size of 10 meshes to obtain the feed additive.

[0066] Examples 3-6

[0067] Examples 3-6 respectively provide an Enteromorpha prolifera feed additive.

[0068] The difference between the above examples and Example 1 is that the high-temperature composite bacteria used in the preparation method of the Enteromorpha prolifera feed additive are as follows:

[0069] In the high-temperature composite bacteria of Example 3, the viable count ratio of Bacillus subtilis, Bacillus thermophilus, Bacillus licheniformis, yeast and lactic acid bacteria is 1:0.8:1.5:1.2:0.8.

[0070] In the high-temperature composite bacteria of Example 4, the viable count ratio of Bacillus subtilis, Bacillus thermophilus, Bacillus licheniformis, yeast and lactic acid bacteria is 1.2:1:1.2:1.2:1.2.

[0071] In the high-temperature composite bacteria of Example 5, the viable count ratio of Bacillus subtilis, Bacillus thermophilus, Bacillus licheniformis, yeast and lactic acid bacteria is 1:0.8:1.2:1.5:1.

[0072] In the high-temperature composite bacteria of Example 6, the ratio of viable bacteria of Bacillus subtilis, Bacillus thermophilus, Bacillus licheniformis, yeast and lactic acid bacteria is 1:1.1:1.2:1.5:0.8.

[0073] Example 7

[0074] Example 7 provides a seaweed feed additive.

[0075] The difference between the above embodiment and Embodiment 1 is that the amount of high-temperature compound bacteria added is 0.02% of the mass of the seaweed mixed slurry.

[0076] Example 8

[0077] Example 8 provides a seaweed feed additive.

[0078] The difference between the above embodiment and Embodiment 1 is that the amount of high-temperature compound bacteria added is 0.05% of the mass of the seaweed mixed slurry.

[0079] Example 9

[0080] Example 9 provides a seaweed feed additive.

[0081] The difference between the above embodiment and Embodiment 1 is that the amount of high-temperature compound bacteria added is 0.07% of the mass of the seaweed mixed slurry.

[0082] Examples 10-12

[0083] Examples 10-12 provide a seaweed feed additive.

[0084] The difference between the above embodiments and Embodiment 1 lies in the high-temperature compound bacteria used in the preparation method of the *Ulva prolifera* feed additive, as detailed below:

[0085] In the functional compound bacteria of Example 10, the ratio of viable bacteria of Lactobacillus plantarum, Candida utilis and Aspergillus niger is 1:1.5:0.5.

[0086] In the functional compound bacteria of Example 11, the ratio of viable bacteria of Lactobacillus plantarum, Candida utilis and Aspergillus niger is 1:0.8:0.6.

[0087] In the functional compound bacteria of Example 12, the ratio of viable bacteria of Lactobacillus plantarum, Candida utilis and Aspergillus niger is 1:1:1.

[0088] Comparative Examples 1-3

[0089] Comparative Examples 1-3 each provide a Ulva prolifera feed additive.

[0090] The difference between the above comparative example and Example 1 lies in the high-temperature compound bacteria used in the preparation method of the *Ulva prolifera* feed additive, as detailed below:

[0091] In the high-temperature complex bacteria of Comparative Example 1, the ratio of viable bacteria of Bacillus subtilis, Bacillus thermophilus, Bacillus licheniformis, yeast and lactic acid bacteria was 1:1.5:1.2:1.2:1.

[0092] In the high-temperature complex bacteria of Comparative Example 2, the ratio of viable bacteria of Bacillus subtilis, Bacillus thermophilus, Bacillus licheniformis, yeast and lactic acid bacteria was 1:1:0.8:0.8:1.

[0093] In the high-temperature complex bacteria of Comparative Example 3, the ratio of viable bacteria of Bacillus subtilis, Bacillus thermophilus, Bacillus licheniformis, yeast and lactic acid bacteria was 0.8:1:1.2:1.2:1.

[0094] Comparative Example 4

[0095] Comparative Example 4 provides a seaweed feed additive.

[0096] The difference between the above comparative example and Example 1 is that the preparation method of the *Ulva prolifera* feed additive does not involve high-temperature fermentation. The specific steps are as follows:

[0097] (1) Pretreatment of seaweed: The fresh seaweed harvested was rinsed three times with seawater and twice with fresh water in the near sea. Then it was dehydrated by a mechanized centrifuge to make the water content of the seaweed about 75%. The seaweed was then cut into sections, crushed and pulped to obtain seaweed slurry. The seaweed slurry was mixed with enoki mushroom residue at a weight ratio of 1:0.3 and added to a solid fermentation device. The water content was adjusted to 60% to obtain seaweed mixed slurry for later use.

[0098] (2) Heat and ventilate the mixed slurry of seaweed to 70°C and stir for 10 hours.

[0099] (3) Functional fermentation: The moisture content in the fermentation silo is controlled at about 40%, the temperature is controlled at 30℃, the air circulation system is turned off, and functional compound bacteria are inoculated at 0.2% by mass of the seaweed mixture slurry, and the mixture is stirred at a constant temperature for 20 hours; the functional compound bacteria are composed of Lactobacillus plantarum, Candida utilis and Aspergillus niger with a live bacteria ratio of 1:1:0.5; the effective live bacteria count of the functional compound bacteria is 200 million / g.

[0100] (4) Adjusting moisture, crushing and screening: After the functional fermentation is completed, the temperature of the silo is raised to 60°C and the ventilation is increased to further reduce the moisture content of the material to 13%. Then, heating is stopped and the material in the fermentation chamber is gradually reduced to room temperature through the air circulation system. The material is then crushed and transferred to the drum screen with a 5-mesh aperture to obtain feed additives.

[0101] Comparative Example 5

[0102] Comparative Example 5 provides a seaweed feed additive.

[0103] The difference between the above comparative example and Example 1 is that the preparation method of the *Ulva prolifera* feed additive does not involve functional fermentation. The specific steps are as follows:

[0104] (1) Pretreatment of seaweed: The fresh seaweed harvested was rinsed three times with seawater and twice with fresh water in the near sea. Then it was dehydrated by a mechanized centrifuge to make the water content of the seaweed about 75%. The seaweed was then cut into sections, crushed and pulped to obtain seaweed slurry. The seaweed slurry was mixed with enoki mushroom residue at a weight ratio of 1:0.3 and added to a solid fermentation device. The water content was adjusted to 60% to obtain seaweed mixed slurry for later use.

[0105] (2) High-temperature fermentation: Start the solid fermentation device, turn on the heating device, air circulation system and stirring system, and gradually heat the mixture to 70°C. Maintain constant temperature and stir. Inoculate with high-temperature compound bacteria at a rate of 0.03% by mass of the seaweed mixture slurry, and maintain constant temperature and stir for 10 hours. During this period, the air circulation system continuously sends fresh air into the solid fermentation chamber to ensure sufficient oxygen during the fermentation process. The effective viable count of the high-temperature compound bacteria is 150 million / g. The high-temperature compound bacteria consists of Bacillus subtilis, Bacillus thermophilus, Bacillus licheniformis, yeast and lactic acid bacteria in a viable count ratio of 1:1:1.2:1.2:1.

[0106] (3) Adjusting moisture, crushing and screening: After the high-temperature fermentation is completed, the temperature of the silo is reduced to 60°C and the ventilation is increased to further reduce the moisture content of the material to 13%. Then, heating is stopped and the material in the fermentation chamber is gradually reduced to room temperature through the air circulation system. The material is then crushed and transferred to a drum screen with a 5-mesh aperture to obtain feed additives.

[0107] Broiler feeding trial

[0108] Feeding trials of *Ulva prolifera* feed additives provided in Examples 1-12 and Comparative Examples 1-5 were conducted on broiler chickens. The experimental methods are as follows:

[0109] (1) Preparation of broiler feed: The *Ulva prolifera* feed additives provided in Examples 1-12 and Comparative Examples 1-5 of this application were added to commercially available feed (purchased from Hangzhou Baoji Biotechnology Co., Ltd.). The addition amount of the *Ulva prolifera* feed additive in the commercially available feed was 2% by mass, resulting in broiler feeds A1-A12 and B1-B5. In addition, the *Ulva prolifera* feed additive of Example 1 was added to commercially available feed at a dosage of 4% by mass, resulting in broiler feed B6.

[0110] (2) 190 one-day-old AA+ white-feathered broilers were selected for the experiment, half male and half female, and randomly divided into 19 groups with 10 birds in each group; the experimental period was 42 days.

[0111] (3) Eighteen groups were fed one type of broiler feed (A1-A12 and B1-B6) prepared above; one group was fed commercially available feed. During the experiment, all conditions were basically the same. Immunization and deworming were carried out according to the routine procedure. The chickens were fed twice a day, with free access to feed and water. The feces were cleaned once a day, and the water troughs were scrubbed once a day.

[0112] (4) Record the feed consumption of each group of broilers over 42 days and calculate the average daily feed intake of each broiler; weigh the fasting weight of the broilers before the experiment begins; weigh the fasting weight of the broilers after the experiment ends and calculate the average daily weight gain of each broiler; and calculate the feed conversion ratio. The results are shown in Table 2 below.

[0113] Average daily feed intake per broiler chicken = Feed consumption of each group of broilers over 42 days / 420

[0114] Average daily weight gain per broiler = (Average weight at the end of the trial for each group of broilers - Average initial weight for each group of broilers) / 420.

[0115] Feed conversion ratio = Average daily feed intake per broiler / Average daily weight gain per broiler

[0116] Table 2. Results of the test on average daily feed intake and average daily weight gain of broiler chickens.

[0117]

[0118]

[0119] As shown in the table above, the average daily weight gain of broilers fed broiler feeds A1-A12 was significantly higher than that of broilers fed broiler feeds B1-B5 and commercially available feeds. Furthermore, the average daily feed intake of broilers fed broiler feeds A1-A12 was >110g / bird, and the feed conversion ratio (FCR) was ≤1.80, while the average daily feed intake of broilers fed broiler feeds B1-B5 and commercially available feeds was ≤110g / bird, and the FCR was >1.80. Therefore, this indicates that the *Ulva prolifera* feed additive provided in this application can improve the appetite of broilers and provide them with sufficient nutrients, thereby enabling broilers to fully convert the feed into chicken meat and increase breeding efficiency.

[0120] Immune function testing of broilers

[0121] The immune organ indices (thymus index, bursa of Fabricius index, and spleen index) of the broilers in the above broiler feeding experiment were tested, and the results are shown in Table 3 below.

[0122] After the broiler feeding trial concluded, three broilers were randomly selected from each group. Each broiler was weighed, dissected, and its thymus, bursa of Fabricius, and spleen were collected and weighed. The immune organ index was then calculated, and the average value of the immune organ index for each group was determined. The formula for calculating the immune organ index is as follows:

[0123] Immune organ index (mg / g) = Weight of immune organs / Total weight of chicken

[0124] Table 3 Results of Immune Organ Index Detection in Broilers

[0125]

[0126]

[0127] As shown in the table above, the immune organ indices of broilers fed with broiler feeds A1-A12 are all higher than those of broilers fed with broiler feeds B1-B5 and commercially available feeds. This indicates that the preparation method of the *Ulva prolifera* feed additive provided in this application can maximize the conversion and retention of nutrients in *Ulva prolifera*, achieving efficient utilization of *Ulva prolifera* and obtaining a *Ulva prolifera* feed additive with high nutrient content. Adding this *Ulva prolifera* feed additive to poultry feed for broilers can significantly enhance their growth performance and immune function, increase breeding efficiency, and has good economic value and application prospects.

[0128] Furthermore, due to its high water and salt content, short shelf life, susceptibility to decay and odor, and concentrated outbreak period, existing technologies struggle to process Ulva prolifera quickly and efficiently. If not treated promptly after being harvested, Ulva prolifera often becomes a source of secondary pollution. However, the pretreatment process of the Ulva prolifera feed additive preparation method provided in this application directly uses near-shore seawater to wash the Ulva prolifera. The wastewater after washing is free of pollutants and can be directly discharged into the sea, effectively avoiding secondary environmental pollution. In addition, washing directly in near-shore waters eliminates the costs associated with collecting and transporting the Ulva prolifera to the processing plant, effectively reducing production costs.

[0129] In summary, the preparation method of the seaweed feed additive provided in this application can significantly reduce production costs, reduce environmental pollution, and achieve efficient resource utilization of seaweed. The obtained seaweed feed additive has high nutritional value, can promote intestinal development in animals, enhance animal growth performance and immune function, and has good application prospects.

[0130] Although the present invention has been described in detail above with general descriptions and specific embodiments, modifications or improvements can be made to it, which will be obvious to those skilled in the art. Therefore, all such modifications or improvements made without departing from the spirit of the present invention fall within the scope of protection claimed by the present invention.

Claims

1. A method for preparing a feed additive of Enteromorpha, characterized by, It comprises the following steps: Enteromorpha pretreatment: the collected fresh Enteromorpha is washed with seawater and fresh water in turn, then dehydrated to make the water content 75-85%; then the Enteromorpha is cut into sections, crushed, and pulped, and then auxiliary materials are added to obtain Enteromorpha mixed slurry; High-temperature fermentation: high-temperature fermentation of the Enteromorpha mixed slurry is carried out at 65-80℃ by using high-temperature composite bacteria; the high-temperature composite bacteria are composed of Bacillus subtilis, Bacillus caldolyticus, Bacillus licheniformis, yeast and lactic acid bacteria with a ratio of live bacteria number of (1-1.2):(0.8-1.1):(1.2-1.5):(1.2-1.5):(0.8-1.2); the addition amount of the high-temperature composite bacteria is 0.03-0.05% of the mass of the Enteromorpha mixed slurry, and the effective live bacteria number of the high-temperature composite bacteria is ≥1.0 billion / g; Functional fermentation: the mixture obtained by the high-temperature fermentation is cooled to 25-35℃, and then functional composite bacteria are added and fermented at the above-mentioned temperature; the functional composite bacteria are composed of Lactobacillus plantarum, Candida utilis and Aspergillus niger with a ratio of live bacteria number of 1:(0.8-1.5):(0.3-0.6); the addition amount of the functional composite bacteria is 0.1-0.3% of the mass of the Enteromorpha mixed slurry, and the effective live bacteria number of the functional composite bacteria is ≥2.0 billion / g.

2. The method of claim 1, wherein the Enteromorpha feed additive is prepared by the steps of: The time of the high-temperature fermentation is 8-12h, and the time of the functional fermentation is 12-24h.

3. The method of claim 1, wherein the Enteromorpha feed additive is prepared by the steps of: The Enteromorpha is cut into sections and crushed to obtain Enteromorpha slurry, and auxiliary materials are added to prepare Enteromorpha mixed slurry, and the water content of the Enteromorpha mixed slurry is 50-65%; The Enteromorpha mixed slurry is composed of Enteromorpha slurry and auxiliary materials with a weight ratio of 1:(0.2-0.4); The auxiliary materials are selected from one or more of wheat bran, corn husk and flammulina velutipes residue.

4. The method of claim 1, wherein the Enteromorpha feed additive is prepared by the steps of: The preparation method further comprises adjusting the moisture, crushing and sieving, and the specific steps are as follows: after the functional fermentation is completed, the temperature of the mixture is increased to 55-60℃ to make the water content of the mixture ≤15%; then the mixture is cooled, crushed and sieved through a 5-20 mesh sieve to obtain Enteromorpha feed additive.

5. An Enteromorpha feed additive obtained by the preparation method of the Enteromorpha feed additive according to any one of claims 1-4.

6. Application of the Enteromorpha feed additive according to claim 5 in poultry feed, aquatic feed and pig feed.

7. The use of Enteromorpha sp. feed additive according to claim 6, characterized in that, The addition amount of the Enteromorpha feed additive in poultry feed is 1-3% by mass.

8. The use of Enteromorpha sp. feed additive according to claim 7, characterized in that, The addition amount of the Enteromorpha feed additive in aquatic feed is 2-5% by mass.

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