An enzymatically hydrolyzed protein composition as an alternative to fish meal and its preparation method

Abstract

This invention relates to an enzymatically hydrolyzed protein composition as a substitute for fishmeal and its preparation method, belonging to the field of feed technology. The enzymatically hydrolyzed protein composition comprises the following raw materials: animal protease hydrolysate, brewer's yeast extract, bile acids, sodium butyrate, sodium selenite, and fructooligosaccharides. This application selects cricket powder and defatted mealworm powder as animal protein sources. Simultaneously, chitinase, alkaline protease, and neutral protease are used sequentially to enzymatically hydrolyze the cricket and mealworm complex. The cricket powder and defatted mealworm powder after enzymatic hydrolysis effectively eliminate anti-nutritional factors in the original insect feed, and have high protein utilization, effectively releasing nutrients and improving nutritional value. Based on this, brewer's yeast extract and bile acids are added for compounding, which synergistically promotes weight gain and growth in aquatic animals. Furthermore, this application uses sodium butyrate, sodium selenite, and fructooligosaccharides in combination, which effectively gives the prepared composition significant antioxidant effects from different dimensions.

Description

Technical Field

[0001] This invention belongs to the field of feed technology, specifically, it relates to an enzymatically hydrolyzed protein composition that can replace fishmeal and its preparation method. Background Technology

[0002] In aquaculture, animal protein is a key nutrient for fish growth and development. Fishmeal, as an important component of traditional animal feed, is not only a high-quality source of animal protein but also rich in essential amino acids and growth factors, significantly impacting the growth and feed utilization of aquatic animals. However, with the rapid development of aquaculture in recent years, the demand for fishmeal has continued to rise, making it difficult to meet market demand. Since animal protein sources are similar to fishmeal in nutritional composition and have the advantages of wide availability and low price, they have the potential to become fishmeal substitutes in aquatic feed. However, existing animal protein substitutes for fishmeal often lack good antioxidant properties, making it difficult to improve the immunity and stress resistance of aquatic animals. This makes them susceptible to oxidative stress, reducing growth rate and survival rate. Furthermore, although many manufacturers are currently dedicated to researching growth-promoting feed compositions, their functional characteristics are limited, making it difficult to provide comprehensive nutrition.

[0003] In recent years, enzymatic protein hydrolysis technology has attracted much attention due to its ability to effectively release small-molecule bioactive peptides, free amino acids, and functional factors. Therefore, applying it to the preparation of enzymatically hydrolyzed protein compositions for feed to create a composition with good growth-promoting and antioxidant effects can not only alleviate the pressure caused by fishmeal resource shortages and reduce aquaculture costs, but also improve the growth performance and health of aquatic animals, thus promoting the sustainable development of the aquaculture industry. Summary of the Invention

[0004] The purpose of this invention is to provide an enzymatically hydrolyzed protein composition that can replace fish meal and its preparation method, which solves the problem of poor growth promotion and antioxidant effects in the prior art.

[0005] The objective of this invention can be achieved through the following technical solutions:

[0006] The first aspect of this application provides an enzymatically hydrolyzed protein composition as a substitute for fish meal, comprising the following raw materials:

[0007] Animal protein hydrolysate, brewer's yeast extract, bile acids, sodium butyrate, sodium selenite, fructooligosaccharides;

[0008] Saccharomyces cerevisiae extract is made from fresh yeast through autolysis, enzymatic hydrolysis, separation, concentration, and drying. It retains complete cytoplasmic nutrition and contains abundant functional nutrients such as small peptides, nucleic acids, high protein, and soluble immunopolysaccharides. It is nutritionally comprehensive and can be quickly absorbed by the body, resulting in good nutrient absorption and utilization.

[0009] Bile acids are amphoteric sterol compounds synthesized from cholesterol metabolism in the liver. They have a special molecular structure that can effectively emulsify lipids, accelerate fat digestion, and improve the growth performance of aquatic animals.

[0010] As a preferred embodiment of the present invention, the enzymatically hydrolyzed protein composition that replaces fishmeal comprises 68-88 parts by weight of animal protease hydrolysate, 5-8 parts by weight of Saccharomyces cerevisiae extract, 0.008-0.01 parts by weight of bile acids, 0.01-0.04 parts by weight of sodium butyrate, 0.002-0.005 parts by weight of sodium selenite, and 1-2 parts by weight of fructooligosaccharides.

[0011] As a preferred embodiment of the present invention, the animal protein hydrolysate includes cricket powder and defatted yellow mealworm powder;

[0012] Cricket powder is a product obtained by crushing and drying crickets. Cricket powder contains a large number of highly nutritious components, such as amino acids, proteins, minerals and vitamins, which can positively affect the growth performance of aquatic animals and regulate the intestinal microbiota and immune response.

[0013] Yellow mealworm powder is a product obtained by crushing and drying yellow mealworms. Yellow mealworm powder has a high protein content, a balanced amino acid composition, and balanced nutrition, which has a positive effect on promoting body growth. However, yellow mealworm powder has a high fat content. Therefore, this application selects defatted yellow mealworm powder to make it more effective.

[0014] Furthermore, the mass ratio of the cricket powder to the defatted mealworm powder is 3-5:4-6.

[0015] As a preferred embodiment of the present invention, the animal protease hydrolysate further includes a complex enzyme.

[0016] Furthermore, the complex enzyme components include chitinase, alkaline protease, and neutral protease.

[0017] Furthermore, the amount of chitinase added is 1-3% of the total mass of cricket powder and defatted mealworm powder, the amount of alkaline protease added is 2-3% of the total mass of cricket powder and defatted mealworm powder, and the amount of neutral protease added is 1-3% of the total mass of cricket powder and defatted mealworm powder.

[0018] The second aspect of this application provides a method for preparing an enzymatically hydrolyzed protein composition that can replace fish meal, comprising the following steps:

[0019] (1) After mixing cricket powder and defatted yellow mealworms evenly, chitinase, alkaline protease and neutral protease are added in sequence, and the enzymes are hydrolyzed under controlled temperature. After hydrolysis, the enzymes are dried under controlled temperature to obtain animal protein hydrolysate.

[0020] (2) Mix and stir the animal protein hydrolysate, brewer's yeast extract, bile acid, sodium butyrate, sodium selenite and fructooligosaccharide, extrude and granulate, and dry to obtain the finished product of the composition.

[0021] As a preferred embodiment of the present invention, in step (1), the temperature-controlled enzymatic hydrolysis temperature of the chitinase is 40-45℃ and the time is 8-10h; the temperature-controlled enzymatic hydrolysis temperature of the alkaline protease is 45-50℃ and the time is 3-5h; the temperature-controlled enzymatic hydrolysis temperature of the neutral protease is 45-50℃ and the time is 4-6h; and the temperature-controlled drying temperature is 45-50℃ and the time is 20-24h.

[0022] As a preferred technical solution of the present invention, the stirring time in step (2) is 30-40 min; the extrusion speed of the extrusion granulation is 60-65 r / min, and the granulation template is 0.25 mm; the drying is to control the moisture content of the final product to 6-10%.

[0023] The third aspect of this application provides the application of an enzymatically hydrolyzed protein composition that can replace fishmeal in the preparation of feed, wherein the composition is added at 2-3% of the total mass of the feed.

[0024] The beneficial effects of this invention are:

[0025] (1) This application selects cricket powder and defatted mealworm powder as animal protein sources. At the same time, chitinase, alkaline protease and neutral protease are used in sequence to enzymatically hydrolyze the cricket and mealworm complex. Under the enzymatic hydrolysis sequence set in this application, chitinase can effectively solve the large amount of insoluble chitin contained in the insect shell and epidermis, effectively improve the protein utilization rate of insects, while alkaline protease can effectively improve the degree of hydrolysis, and neutral protease can degrade bitter peptides, increase the proportion of small peptides and improve palatability to a certain extent. Therefore, the cricket powder and defatted mealworm powder after enzymatic hydrolysis can effectively eliminate the anti-nutritional factors in the original insect feed, and its protein utilization rate is high, effectively releasing nutrients, improving nutritional value, and playing a positive role in the growth and development of aquatic animals.

[0026] In addition, brewer's yeast extract contains a variety of functional nutrients, which are comprehensive and can be quickly absorbed by the body. Bile acids have a special molecular structure that can accelerate fat digestion and improve the growth performance of aquatic animals. Therefore, this application combines compound enzymatic hydrolysate, brewer's yeast extract and bile acids to synergistically promote the weight gain and growth of aquatic animals.

[0027] (2) Sodium butyrate can effectively reduce the content of protein hydroxyl groups and endogenous reactive oxygen species to reduce free radicals in the body. At the same time, it can also activate the AMPK / mitochondrial autophagy pathway induced by H2O2 in IPEC-J2 cells, thereby increasing the mRNA expression level of enzymes such as GST. Sodium selenite can activate the antioxidant enzyme system, increase the resistance to hydroxyl free radicals and superoxide anions, thereby increasing the activity of antioxidant enzymes. Fructooligosaccharides can enhance the body's antioxidant capacity, regulate the balance between oxidation and antioxidation in the body, reduce the content of lipid peroxidation products, and prevent damage to the body. Therefore, this application combines sodium butyrate, sodium selenite and fructooligosaccharides to effectively make the prepared composition have significant antioxidant effects from different dimensions. Detailed Implementation

[0028] The technical solutions of the present invention will be clearly and completely described below with reference to the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative effort are within the scope of protection of the present invention.

[0029] The cricket powder and defatted mealworm powder used in the embodiments, comparative examples and test examples of this application were all purchased from Hunan Aozhou Agricultural Technology Co., Ltd.; the chitinase and alkaline protease used were all purchased from Shanghai Kanglang Biotechnology Co., Ltd., and the neutral protease was purchased from Shandong Pingju Biotechnology Co., Ltd.; the above will not be repeated hereafter.

[0030] Example 1

[0031] An enzymatically hydrolyzed protein composition as an alternative to fishmeal comprises the following raw materials in parts by weight:

[0032] Animal protein hydrolysate 78 parts by weight, brewer's yeast extract 5 parts by weight, bile acids 0.008 parts by weight, sodium butyrate 0.01 parts by weight, sodium selenite 0.003 parts by weight, fructooligosaccharides 2 parts by weight;

[0033] The animal protease hydrolysate comprises cricket powder and defatted yellow mealworm powder in a mass ratio of 5:4; the animal protease hydrolysate also includes a complex enzyme, the components of which include chitinase, alkaline protease and neutral protease.

[0034] The amounts of chitinase, alkaline protease, and neutral protease added are 1%, 2.5%, and 1% of the total mass of cricket powder and defatted yellow mealworm powder, respectively.

[0035] The method for preparing the enzymatically hydrolyzed protein composition that replaces fish meal includes the following steps:

[0036] (1) After mixing cricket powder and defatted yellow mealworms evenly, chitinase, alkaline protease and neutral protease are added in sequence, and the enzymes are hydrolyzed under controlled temperature. After hydrolysis, the enzymes are dried at 50℃ for 20 hours to obtain animal protein hydrolysate.

[0037] The temperature-controlled hydrolysis temperature of the chitinase is 40℃, and the time is 9 hours; the temperature-controlled hydrolysis temperature of the alkaline protease is 47.5℃, and the time is 4 hours; the temperature-controlled hydrolysis temperature of the neutral protease is 45℃, and the time is 6 hours.

[0038] (2) Mix the animal protein hydrolysate, brewer's yeast extract, bile acid, sodium butyrate, sodium selenite and fructooligosaccharide and stir for 35 minutes. Extrude and granulate, and dry to obtain the finished product of the composition.

[0039] The extrusion granulation process uses an extrusion speed of 60 r / min and a granulation template of 0.25 mm; the drying process controls the moisture content of the final product to be 8%.

[0040] Example 2

[0041] An enzymatically hydrolyzed protein composition as an alternative to fishmeal comprises the following raw materials in parts by weight:

[0042] Animal protein hydrolysate 68 parts by weight, brewer's yeast extract 8 parts by weight, bile acids 0.009 parts by weight, sodium butyrate 0.04 parts by weight, sodium selenite 0.002 parts by weight, fructooligosaccharides 1.5 parts by weight;

[0043] The animal protease hydrolysate comprises cricket powder and defatted yellow mealworm powder in a mass ratio of 4:6; the animal protease hydrolysate also includes a complex enzyme, the components of which include chitinase, alkaline protease and neutral protease.

[0044] The amounts of chitinase, alkaline protease, and neutral protease added are 2%, 2%, and 3% of the total mass of cricket powder and defatted yellow mealworm powder, respectively.

[0045] The method for preparing the enzymatically hydrolyzed protein composition that replaces fish meal includes the following steps:

[0046] (1) After mixing cricket powder and defatted yellow mealworms evenly, chitinase, alkaline protease and neutral protease are added in sequence, and the enzymes are hydrolyzed under controlled temperature. After hydrolysis, the mixture is dried at 47.5℃ for 22 hours to obtain animal protein hydrolysate.

[0047] The temperature-controlled hydrolysis temperature of the chitinase is 45℃, and the time is 10h; the temperature-controlled hydrolysis temperature of the alkaline protease is 50℃, and the time is 3h; the temperature-controlled hydrolysis temperature of the neutral protease is 47.5℃, and the time is 4h.

[0048] (2) Mix the animal protein hydrolysate, brewer's yeast extract, bile acid, sodium butyrate, sodium selenite and fructooligosaccharide and stir for 30 minutes. Extrude and granulate, and dry to obtain the finished product of the composition.

[0049] The extrusion granulation process uses an extrusion speed of 63 r / min and a granulation template of 0.25 mm; the drying process controls the moisture content of the final product to 6%.

[0050] Example 3

[0051] An enzymatically hydrolyzed protein composition as an alternative to fishmeal comprises the following raw materials in parts by weight:

[0052] Animal protein hydrolysate 82 parts by weight, brewer's yeast extract 6.5 parts by weight, bile acids 0.01 parts by weight, sodium butyrate 0.03 parts by weight, sodium selenite 0.005 parts by weight, fructooligosaccharides 1 part by weight;

[0053] The animal protease hydrolysate comprises cricket powder and defatted yellow mealworm powder in a mass ratio of 3:5; the animal protease hydrolysate also includes a complex enzyme, the components of which include chitinase, alkaline protease and neutral protease.

[0054] The amounts of chitinase, alkaline protease, and neutral protease added are 3%, 3%, and 2% of the total mass of cricket powder and defatted yellow mealworm powder, respectively.

[0055] The method for preparing the enzymatically hydrolyzed protein composition that replaces fish meal includes the following steps:

[0056] (1) After mixing cricket powder and defatted yellow mealworms evenly, chitinase, alkaline protease and neutral protease are added in sequence, and the enzymes are hydrolyzed at controlled temperature. After hydrolysis, the enzymes are dried at controlled temperature of 45℃ for 24 hours to obtain animal protein hydrolysate.

[0057] The temperature-controlled hydrolysis temperature of the chitinase is 43℃, and the time is 8 hours; the temperature-controlled hydrolysis temperature of the alkaline protease is 45℃, and the time is 5 hours; the temperature-controlled hydrolysis temperature of the neutral protease is 50℃, and the time is 5 hours.

[0058] (2) Mix the animal protein hydrolysate, brewer's yeast extract, bile acid, sodium butyrate, sodium selenite and fructooligosaccharide and stir for 40 minutes. Extrude and granulate, and dry to obtain the finished product of the composition.

[0059] The extrusion granulation process uses an extrusion speed of 65 r / min and a granulation template of 0.25 mm; the drying process controls the moisture content of the final product to 10%.

[0060] Comparative Example 1

[0061] Compared with Example 3, the difference is that no cricket powder was added in step (1) of Comparative Example 1, and the missing weight was made up with defatted yellow mealworms. The other operation steps and parameters remained unchanged.

[0062] Comparative Example 2

[0063] Compared with Example 3, the difference is that defatted mealworms are not added in step (1) of Comparative Example 2, and the missing weight is made up with cricket powder. The other operation steps and parameters remain unchanged.

[0064] Comparative Example 3

[0065] Compared with Example 3, the difference is that no Saccharomyces cerevisiae extract was added in Comparative Example 3, and the missing weight was made up with animal protease hydrolysis and bile acids in a mass ratio of 82:0.01. All other operation steps and parameters remained unchanged.

[0066] Comparative Example 4

[0067] Compared with Example 3, the difference is that bile acids were not added in Comparative Example 4, and the missing weight was made up with animal proteolytic enzymes and Saccharomyces cerevisiae extract in a mass ratio of 82:6.5. All other operation steps and parameters remained unchanged.

[0068] Comparative Example 5

[0069] Compared with Example 3, the difference is that in Comparative Example 5, chitinase, alkaline protease, and neutral protease were mixed for complex enzymatic hydrolysis, specifically:

[0070] The method for preparing the enzymatically hydrolyzed protein composition that replaces fish meal includes the following steps:

[0071] (1) After mixing cricket powder and defatted yellow mealworms evenly, add chitinase, alkaline protease and neutral protease and stir evenly. Then, control the temperature at 45℃ for 12 hours for enzymatic hydrolysis. After enzymatic hydrolysis, control the temperature at 45℃ for 24 hours for drying to obtain animal protease hydrolysate.

[0072] (2) Mix the animal protein hydrolysate, brewer's yeast extract, bile acid, sodium butyrate, sodium selenite and fructooligosaccharide and stir for 40 minutes. Extrude and granulate, and dry to obtain the finished product of the composition.

[0073] The extrusion granulation process uses an extrusion speed of 65 r / min and a granulation template of 0.25 mm; the drying process controls the moisture content of the final product to be 10%.

[0074] All other operating steps and parameters remain unchanged.

[0075] Comparative Example 6

[0076] Compared with Example 3, the difference is that sodium butyrate was not added in Comparative Example 6, and the missing weight was made up with sodium selenite and fructooligosaccharides in a mass ratio of 0.005:1. All other operation steps and parameters remained unchanged.

[0077] Comparative Example 7

[0078] Compared with Example 3, the difference is that sodium selenite was not added in Comparative Example 7, and the missing weight was made up with sodium butyrate and fructooligosaccharides in a mass ratio of 0.03:1. All other operation steps and parameters remained unchanged.

[0079] Comparative Example 8

[0080] Compared with Example 3, the difference is that no fructooligosaccharides were added in Comparative Example 8, and the missing weight was supplemented with sodium butyrate and sodium selenite in a mass ratio of 0.03:0.005. All other operating steps and parameters remained unchanged.

[0081] Test case

[0082] Yellow catfish (initial weight 6.5±0.1g) from the same batch, exhibiting good vitality and apparent health, were selected and randomly assigned to rearing tanks (1.2×1.2m, effective water volume: 1000L) after a 24-hour starvation period. Three tanks were placed in each group, with 50 fish per tank. During the rearing period, fish were fed twice daily (8:00 AM and 5:00 PM), and feed intake was recorded daily. The experiment was conducted under natural light, with water temperature maintained at 24.0℃, dissolved oxygen maintained above 6.0 mg / L, and total ammonia nitrogen maintained below 0.1 mg / L. Feed products prepared by adding the enzymatically hydrolyzed protein compositions obtained in Examples 1-3 and Comparative Examples 1-8 were fed into the breeding tanks respectively (the feed composition of each group is: 35wt% soybean meal, 30wt% wheat bran, 20wt% rapeseed meal, 12wt% cottonseed meal, 0.5wt% premix (the premix is ​​from Guangdong Xintun Biotechnology Co., Ltd., and the product name is meat and fish compound premixed feed) and 0.5wt% calcium dihydrogen phosphate, and 2wt% enzymatically hydrolyzed protein composition). Half of the daily feed amount was fed at 8:00 and 17:00. At the same time, the feces were cleaned up daily and the uneaten feed was removed before feeding. The entire test period was 4 weeks. After the test, the following tests (1)-(2) were carried out:

[0083] (1) After the experiment, record the final body weight of the yellow catfish in each culture tank of Examples 1-3 and Comparative Examples 1-5, and calculate the weight gain rate according to the following formula:

[0084] Weight gain rate (%) = [(final weight - initial weight) / initial weight] × 100%;

[0085] The final results of each group are averaged, and the results are shown in Table 1.

[0086] (2)(a) Sample collection: After the experiment, 18 fish (6 fish per breeding tank in each group of Examples 1-3 and Comparative Examples 6-8) were randomly selected. After anesthetizing with MS-222 at 150 mg / L, blood was collected from the tail vein. The serum was obtained by centrifugation at 4°C for 15 min and then stored at -80°C for analysis of antioxidant capacity.

[0087] (b) Determination of antioxidant indicators: The antioxidant indicators of serum were strictly determined in accordance with the instructions of the Total Antioxidant Capacity (T-AOC) Test Kit and the Malondialdehyde (MDA) Test Kit (both T-AOC and MDA kits were purchased from Nanjing Jiancheng Bioengineering Research Institute Co., Ltd.). The final results of each group were averaged, and the results are shown in Table 2.

[0088] Table 1

[0089]

[0090]

[0091] As can be seen from Table 1, the composition product obtained in this application has excellent growth-promoting effect.

[0092] Table 2

[0093] T-AOC (nmol / mL) MDA (nmol / mL) Example 1 702.68 12.76 Example 2 704.09 12.51 Example 3 705.21 12.13 Comparative Example 6 680.10 17.34 Comparative Example 7 693.92 14.07 Comparative Example 8 688.52 15.22

[0094] As can be seen from Table 2, the finished product of the composition prepared in this application has significant antioxidant effect.

[0095] In the description of this specification, the references to terms such as "an embodiment," "example," "specific example," etc., indicate that a specific feature, structure, material, or characteristic described in connection with that embodiment or example is included in at least one embodiment or example of the present invention. In this specification, the illustrative expressions of the above terms do not necessarily refer to the same embodiment or example. Furthermore, the specific features, structures, materials, or characteristics described may be combined in any suitable manner in one or more embodiments or examples.

[0096] The above description is merely an example and illustration of the concept of the present invention. Those skilled in the art can make various modifications or additions to the specific embodiments described or use similar methods to replace them, as long as they do not deviate from the concept of the invention or exceed the scope defined in the claims, they should all fall within the protection scope of the present invention.

Claims

1. An enzymatically hydrolyzed protein composition as a substitute for fishmeal, characterized in that, Including the following parts by weight of raw materials: 68-88 parts by weight of animal protein hydrolysate, 5-8 parts by weight of brewer's yeast extract, 0.008-0.01 parts by weight of bile acids, 0.01-0.04 parts by weight of sodium butyrate, 0.002-0.005 parts by weight of sodium selenite, and 1-2 parts by weight of fructooligosaccharides. The animal proteolytic feed includes cricket powder and defatted yellow mealworm powder, and also includes compound enzymes; The mass ratio of cricket powder to defatted yellow mealworm powder is 3-5:4-6; The complex enzyme components include chitinase, alkaline protease, and neutral protease.

2. The enzymatically hydrolyzed protein composition as a substitute for fish meal according to claim 1, characterized in that, The amount of chitinase added is 1-3% of the total mass of cricket powder and defatted mealworm powder, the amount of alkaline protease added is 2-3% of the total mass of cricket powder and defatted mealworm powder, and the amount of neutral protease added is 1-3% of the total mass of cricket powder and defatted mealworm powder.

3. A method for preparing an enzymatically hydrolyzed protein composition as a substitute for fish meal according to any one of claims 1-2, characterized in that, Includes the following steps: (1) After mixing cricket powder and defatted yellow mealworms evenly, chitinase, alkaline protease and neutral protease are added in sequence, and the enzymes are hydrolyzed under controlled temperature. After hydrolysis, the enzymes are dried under controlled temperature to obtain animal protein hydrolysate. (2) Mix and stir the animal protein hydrolysate, brewer's yeast extract, bile acid, sodium butyrate, sodium selenite and fructooligosaccharide, extrude and granulate, and dry to obtain the finished product of the composition.

4. The method for preparing the enzymatically hydrolyzed protein composition as a substitute for fish meal according to claim 3, characterized in that, In step (1), the temperature-controlled enzymatic hydrolysis of the chitinase is 40-45℃ for 8-10 hours; the temperature-controlled enzymatic hydrolysis of the alkaline protease is 45-50℃ for 3-5 hours; the temperature-controlled enzymatic hydrolysis of the neutral protease is 45-50℃ for 4-6 hours; and the temperature-controlled drying is 45-50℃ for 20-24 hours.

5. The method for preparing the enzymatically hydrolyzed protein composition as a substitute for fish meal according to claim 3, characterized in that, The stirring time in step (2) is 30-40 min; the extrusion speed of the extrusion granulation is 60-65 r / min, and the granulation template is 0.25 mm; the drying is to control the moisture content of the final product to 6-10%.

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