A composition preparation and fermentation method for palm meal functional solid-state fermentation material

Abstract

The present invention belongs to the field of additives and discloses a composition for palm meal functional solid-state fermentation feed, comprising palm meal, wheat bran, nutrient solution, concentrated fish paste hydrolyzate, soy molasses, composite bacterial seed liquid, barley worm larvae intestinal culture, and black soldier fly intestinal culture; the composite bacterial species in the composite bacterial seed liquid are composed of Bacillus, yeast, Aspergillus, and Lactobacillus plantarum. The composition uses the composite bacteria for anaerobic fermentation. At the same time, the concentrated fish paste hydrolyzate and soy molasses are used to enhance bacterial activity, providing nutritional support for the efficient function of the bacterial species. The composition of the present invention can effectively reduce the crude fiber and mannan content of palm meal and effectively decompose the crude fiber and mannan. Another object of the present invention is to provide a fermentation method for the palm meal.

Description

Technical Field

[0001] The invention belongs to the biological field, and particularly relates to a composition preparation and fermentation method for palm meal functional solid-state fermentation material. Background Art

[0002] Palm meal is a byproduct of palm kernel shelling and oil extraction. It contains 14%-21% crude protein, 21%-23% crude fiber, and 8%-17% crude fat. When fed directly to animals, palm meal suffers from high fiber content, poor palatability, and low energy and protein utilization rates. This results in extremely low inclusion rates in feed (e.g., 1%-7% for poultry and 3%-9% for pigs), making this inexpensive meal ineffective in the animal feed industry.

[0003] CN111642618A discloses a ruminant fermented feed and a preparation method thereof, belonging to the field of feed technology. The feed comprises the following raw materials in parts by weight: 22-30 parts of corn, 2-10 parts of sprayed corn husks, 4-12 parts of DDGS, 6-14 parts of corn germ meal, 10-20 parts of wheat bran, 1.5-5.5 parts of cottonseed meal, 2-8 parts of palm meal, 2-8 parts of sesame meal, 1.2-3.2 parts of glutamic acid residue, 0.5-1.5 parts of molasses, 0.05-0.15 parts of live bacteria fermentation agent, 0.05-0.15 parts of complex enzyme preparation, and 18-26 parts of water. The live bacteria fermentation agent comprises brewer's yeast, lactic acid bacteria, and bacillus, with the weight ratio of brewer's yeast, lactic acid bacteria, and bacillus being (2.5-3.5):(3.8-4.2):(1.7-2.3).

[0004] CN110463828A discloses a method for producing biofeed from palm meal. The method uses palm meal as raw material and adopts a combination of enzymatic hydrolysis and microbial fermentation to obtain a palm meal fermented feed with good palatability, high energy and protein utilization, and rich in probiotics, small molecule peptides, lactic acid, biological enzymes, and oligosaccharides. The palm meal fermented feed obtained by the method has a pH of 4.2-5.0, an NDF of 33%-40%, a small molecule peptide content of 27%-33%, an amino acid nitrogen content of 1.89-2.10%, a crude protein content of 13.5-17.0%, which is slightly higher than the crude protein content of the starting material, and a lactic acid colony count of 5.1-7.0×10 8 cfu / g, the number of Bacillus subtilis spores is 6.2-8×10 9 cfu / g, protease activity 120-210u / g, saccharifying enzyme activity 310-620u / g, mannanase activity 1.5-2.8u / g, adding a certain amount of vitamins, trace elements and lysine, methionine, can replace 20%-40% (1:1 equivalent replacement) of animal complete diet.

[0005] It can be seen that fermentation is a common method to reduce the fiber content of palm meal and increase the protein content.

[0006] Palm meal, due to its rich nutritional content, has considerable application value. Numerous studies, both domestically and internationally, have confirmed its suitability in ruminant diets. However, studies have also examined its use in pigs, rabbits, poultry, and aquatic animals, establishing limits on its use in diets: no more than 20% in pig feed, 10% in chicken feed, and 30% in aquatic feed. Due to its high crude fiber and mannan-oligosaccharide content, as well as its low energy and crude protein utilization rates, palm meal is not suitable for young and growing animals. Excessive consumption by some monogastric species can also negatively impact their growth performance, limiting its widespread use.

[0007] However, for palm meal, there are several components that require special attention to controlling the amount of material used, such as crude fat and mannan. Taking mannan as an example, its content is as high as 360 mg / g; mannan has physical and chemical properties such as low heat, stability, safety and non-toxicity, and also has the functions of protecting the intestines and improving immunity. Many scholars at home and abroad have reported that palm meal is more suitable for ruminant diets, but there are also relevant reports on its application in pigs, rabbits, poultry and aquatic animals, which have determined that the use of palm meal in diets is limited, such as the amount used in pig feed should not exceed 20%, the amount used in chicken feed should not exceed 10%, and the amount used in aquatic feed should not exceed 30%. Due to the high crude fiber content, high mannan oligosaccharide content, and low energy and crude protein utilization, palm meal is not suitable for young and growing animals. Excessive feeding of some monogastric animals will also affect their growth performance, which limits the widespread use of palm meal. Therefore, controlling the content of crude fiber and mannan in palm meal is a key point of this application.

[0008] Therefore, the technical problem to be solved in this case is: how to increase the crude protein content of palm meal and reduce the fiber, fat and mannan content of palm meal. Summary of the Invention

[0009] In view of the shortcomings and deficiencies of the current existing technology, the primary purpose of the present invention is to provide a composition for palm meal functional solid-state fermentation material, which uses composite bacteria for anaerobic fermentation. At the same time, the activity of the bacteria is improved by concentrating fish paste hydrolyzate and soy molasses, providing nutritional support for the bacteria to efficiently exert their efficacy. The composition of the present invention can effectively reduce the fiber, fat and mannan content of palm meal.

[0010] Another object of the present invention is to provide a fermentation method of the palm meal.

[0011] The specific plan is as follows:

[0012] A composition for palm meal functional solid-state fermentation feed, comprising palm meal, wheat bran, nutrient solution, concentrated fish paste hydrolyzate, soybean molasses, composite bacterial seed liquid, mealworm larvae intestinal culture, and black soldier fly intestinal culture;

[0013] The composite bacterial species in the composite bacterial seed liquid are composed of Bacillus, yeast, Aspergillus and Lactobacillus plantarum;

[0014] Each 100kg composition contains 3-5kg of composite bacterial seed liquid; the concentration of each bacteria in the composite bacterial seed liquid is Bacillus content: 1-1.5×10 10 cfu / ml, yeast content: 3.5-5.0×10 9 cfu / ml, Aspergillus content: 3.5-5.5×10 9 cfu / ml, Lactobacillus plantarum content: 5.0-10×10 8 cfu / ml.

[0015] After the above treatment, the technical parameters of the composition are:

[0016] Morphology: Electron microscopy shows a loose microporous structure with uniform fiber breakage, and significantly improved sensory quality.

[0017] Color: dark coffee or dark brown with special aroma such as wine and coffee.

[0018] The crude protein content shall not be less than 20.0%;

[0019] Crude fat content 2.5-3.5%;

[0020] Crude fiber should not exceed 10.0%;

[0021] The crude ash content shall not exceed 5.0%;

[0022] Mannan (mg / g) shall not exceed 100mg / g;

[0023] Aflatoxins should comply with: GB 31656-2021;

[0024] Escherichia coli should comply with: GB 4789.3-2016;

[0025] Salmonella should comply with: GB 4789.4-2016.

[0026] In the above-mentioned composition for palm meal functional solid-state fermentation material, the weight ratio of the palm meal, wheat bran, nutrient solution, concentrated fish paste hydrolyzate, soybean molasses, composite bacterial seed liquid, barley worm larvae intestinal culture, and black soldier fly intestinal culture is: 55-65:12-17:1:4-6:12-17:3-5:1-5:1-5.

[0027] In the above-mentioned composition for palm meal functional solid-state fermentation material, the weight ratio of the palm meal, wheat bran, nutrient solution, concentrated fish paste hydrolyzate, soybean molasses, composite bacterial seed liquid, mealworm larvae intestinal culture, and black soldier fly intestinal culture is: 60:15:1:5:15:4:3:3.

[0028] In the above-mentioned composition for palm meal functional solid-state fermentation material, the ingredients of the nutrient solution are: 4.0wt% (NH4)2SO4, 0.4wt% MgSO4·7H2O, 0.2wt% KH2PO4, and the balance water.

[0029] In the above-mentioned composition for palm meal functional solid-state fermentation feed, the solid content of the concentrated fish paste hydrolyzate is 35%.

[0030] In the above-mentioned composition for palm meal functional solid-state fermentation material, the preparation method of the concentrated fish paste enzymatic hydrolysis product is: after the fish paste is concentrated, it is transferred into an enzymatic hydrolysis tank, and Bacillus subtilis, actinomycetes, Aspergillus niger fermentation products and Aspergillus oryzae fermentation products are added, stirred evenly, and bacterial hydrolysis and enzymatic hydrolysis are carried out at room temperature (≧20°C). The concentrated fish paste enzymatic hydrolysis product can be obtained by synergistic treatment.

[0031] In the above-mentioned composition for palm meal functional solid-state fermentation material, during the fermentation, an appropriate amount of water is added to adjust the moisture content of the composition to 38%.

[0032] In the above-mentioned composition for palm meal functional solid-state fermentation feed, the preparation method of the intestinal culture of the barleyworm larvae and the intestinal culture of the black soldier fly is:

[0033] The corresponding intestinal tract of the worm body is homogenized to obtain a slurry, which is inoculated into a mixed liquid culture medium at an inoculum rate of 3 to 5 wt%, anaerobically cultured for 36 to 48 hours, and freeze-dried to obtain a slurry having a solid content of 45% to 65%;

[0034] The mixed liquid culture medium contains the following components in a weight ratio: NA:TSA:LB:MRS=4:2:2:2.

[0035] At the same time, the present invention also discloses a palm meal fermentation method, which uses any of the above compositions, adjusts the moisture content of the composition to 35% to 45%, places the composition in an anaerobic fermentation bag and seals it for fermentation at a temperature of 25°C to 35°C, a fermentation time of 3 to 5 days, a gas release frequency of 3 to 5 times, and a material pH value of 5.5 to 6.5.

[0036] In the above-mentioned palm meal fermentation method, the palm meal has a mesh size of 40-60.

[0037] Compared with the prior art, the present invention has the following beneficial effects:

[0038] The composite bacterial strains of the present invention contain Bacillus, yeast, Aspergillus and Lactobacillus plantarum; Bacillus has the function of producing protease and; yeast and Aspergillus have the function of producing cellulase; the metabolites of Lactobacillus plantarum are organic acids and fatty acids; the bacterial strains contained in the intestinal culture of barley worm larvae have the function of producing mannanase, and the bacterial strains contained in the intestinal culture of black soldier flies have the function of producing lipase. The above bacteria and cultures work together to reduce fat content and improve the efficiency of converting cellulose and mannan into protein.

[0039] The composition of the invention adopts concentrated fish paste hydrolyzate and soybean molasses as culture medium to provide nutrients for bacteria, thereby improving the activity of the bacteria. BRIEF DESCRIPTION OF THE DRAWINGS

[0040] Figure 1 Photograph of culture medium activated for gut culture of mealworm larvae;

[0041] Figure 2 This is a photo of the transparent circle showing the cellulase production capacity of the intestinal culture of the mealworm larvae;

[0042] Figure 3 Photograph of culture medium activated for black soldier fly gut culture;

[0043] Figure 4 This is a photo of the transparent circle showing the ability of the black soldier fly intestinal culture to produce lipase;

[0044] Figure 5 are microscopic photographs before and after fermentation, wherein UFO represents microscopic photographs of palm meal before fermentation (2 photos, the same sample, different samplings), and FO represents microscopic photographs of palm meal after fermentation of Example 1 (2 photos, the same sample, different samplings);

[0045] Figure 6 These are electron microscopic images of palm meal cork cells before and after fermentation (two images, the same sample, different sampling, the left is before fermentation; the right is after fermentation);

[0046] Figure 7These are electron microscope scans of palm meal wood fiber before and after fermentation (2 images, same sample, different sampling, left is before fermentation; right is after fermentation). DETAILED DESCRIPTION

[0047] The present invention will be further described in detail below with reference to examples, but the embodiments of the present invention are not limited thereto.

[0048] Before describing the embodiments and comparative examples of the present invention, the detailed preparation process of the intestinal cultures of barleyworm larvae, the intestinal cultures of black soldier flies, the concentrated fish paste hydrolyzate, and the raw materials is first introduced:

[0049] Preparation of intestinal cultures of barleyworm larvae (refer to CN113493288A, a composition for fermentation and deodorization of livestock and poultry manure and its preparation method):

[0050] Under aseptic conditions, a 4 wt% homogenate of the intestinal tract of the mealworm larvae was added to a fermenter containing 50 L of mixed liquid culture medium I (NA:TSA:LB:MRS = 4:2:2:2) and cultured anaerobically for 36-48 hours. After fermentation, the culture medium was removed and freeze-dried under vacuum for 6-8 hours to obtain the intestinal cavity and intestinal culture of the mealworm larvae. Plate colony counts showed that the total bacterial count of the mealworm larval intestinal culture was ≥4.5×10 10 cfu / g. The enzyme production capacity was qualitatively evaluated using Congo red medium, and the maximum transparent zone diameter was ≥40.24 mm.

[0051] Figure 1 Photograph of culture medium activated for gut culture of mealworm larvae;

[0052] Figure 2 This is a photo of the transparent circle showing the cellulase production capacity of the intestinal culture of the mealworm larvae.

[0053] Preparation of black soldier fly intestinal culture (refer to CN113493288A, a composition for fermentation and deodorization of livestock and poultry manure and its preparation method):

[0054] Under aseptic conditions, a 4 wt% intestinal homogenate of black soldier fly larvae was added to a fermenter containing 50 L of mixed liquid culture medium I (NA:TSA:LB:MRS = 4:2:2:2) and cultured anaerobically for 36-48 hours. After fermentation, the culture medium was removed and freeze-dried for 6-8 hours to obtain the intestinal cavity and intestinal culture of the mealworm larvae. Plate colony counts showed that the total bacterial count of the mealworm larval intestinal culture was ≥3.5×10 10 cfu / g. The enzyme production capacity was qualitatively evaluated using WTL medium, and the maximum transparent zone diameter was ≥41.18 mm.

[0055] Enzyme production capacity assessment medium (WTL): 2.5 ml of olive oil, 1.0 g of peptone, 0.5 g of beef extract, 0.3 g of glucose, 1.0 g of PVA, 0.5 g of sodium chloride, 0.5 ml of Tween 80, 1.5 g of agar, 0.016% bromocresol purple, dilute to 100 ml with distilled water, adjust the pH to 7.5, and sterilize at 121°C for 15-20 min.

[0056] Figure 3 Photograph of culture medium activated for black soldier fly gut culture;

[0057] Figure 4 This is a photo of the transparent circle showing the ability of the black soldier fly intestinal culture to produce lipase;

[0058] Preparation of concentrated fish paste hydrolysate: After fish paste (produced by Guangdong Yuechuang Ecological Agriculture Technology Co., Ltd.) is concentrated (dry matter content is about 60%), 1000 g of the concentrated fish paste is transferred to an enzymolysis tank, 5 g of Bacillus subtilis (10 billion / g), 2.5 g of actinomycetes (10 billion / g), 10 g of Aspergillus niger fermentation product, and 10 g of Aspergillus oryzae fermentation product are added, and the mixture is stirred evenly. Bacterial hydrolysis and enzymatic hydrolysis are carried out at room temperature (≧20° C.), and the concentrated fish paste enzymatic hydrolysate is obtained by synergistic treatment.

[0059] The composition of the nutrient solution is: 4.0 wt% of (NH4)2SO4, 0.4 wt% of MgSO4·7H2O, 0.2 wt% of KH2PO4, and the balance of water.

[0060] Bacillus subtilis, actinomycetes, fermentation products of Aspergillus niger, fermentation products of Aspergillus oryzae, yeast, Aspergillus fumigatus, Lactobacillus plantarum, and commercially available complex enzymes are all from Shandong Hezhong Kangyuan Biotechnology Co., Ltd.

[0061] Examples 1-7

[0062] The palm meal is controlled to have a mesh size of 40-60, the moisture content of the composition is adjusted to 40%, the mixture is placed in an anaerobic fermentation bag and sealed for fermentation at room temperature (25°C-35°C), for 5 days, with three deflation times (once every 18-24 hours), and a pH value of the material of 5.5-6.5.

[0063] The dosage of each component of the composition is shown in Table 1.

[0064] Table 1

[0065]

[0066] The concentration of each bacteria in the composite bacterial seed solution is Bacillus content: 1.2×10 10 cfu / ml, yeast content: 4×10 9cfu / ml, Aspergillus content: 4×10 9 cfu / ml, Lactobacillus plantarum content: 8×10 8 cfu / ml.

[0067] Comparative Examples 1-3

[0068] The palm meal is controlled to have a mesh size of 40-60, the moisture content of the composition is adjusted to 40%, the mixture is placed in an anaerobic fermentation bag and sealed for fermentation at room temperature (25° C. to 35° C.), for 5 days, with 5 deflation times (deflation per day), and a pH value of 5.5 to 6.5.

[0069] The dosage of each component of the composition is shown in Table 2;

[0070] Table 2

[0071]

[0072]

[0073] The concentration of each bacteria in the composite bacterial seed solution is Bacillus content: 1.2×10 10 cfu / ml, yeast content: 4×10 9 cfu / ml, Aspergillus content: 4×10 9 cfu / ml, Lactobacillus plantarum content: 8×10 8 cfu / ml.

[0074] Comparative Example 4

[0075] The same as Example 1, except that the composite bacterial seed solution only contains yeast, Aspergillus, and Lactobacillus plantarum; the concentration of each bacteria is: yeast content: 4×10 9 cfu / ml, Aspergillus content: 8×10 9 cfu / ml, Lactobacillus plantarum content: 8×10 8 cfu / ml.

[0076] Comparative Example 5

[0077] The same as Example 1, except that the composite bacterial seed solution only contains Bacillus, yeast, and Lactobacillus plantarum; the concentration of each bacteria in the composite bacterial seed solution is: Bacillus content: 1.2×10 10 cfu / ml, yeast content: 8×10 9 cfu / ml, Lactobacillus plantarum content: 8×10 8 cfu / ml.

[0078] Comparative Example 6

[0079] The same as Example 1, except that the composite bacterial seed solution only contains Bacillus, yeast, and Aspergillus; the concentration of each bacteria in the composite bacterial seed solution is: Bacillus content: 1.2×10 10 cfu / ml, yeast content: 4.5×10 9 cfu / ml, Aspergillus content: 4.5×10 9 cfu / ml.

[0080] Comparative Example 7

[0081] The method is substantially the same as Example 1, except that: the concentrated fish paste enzymatic hydrolysis product is prepared by transferring the concentrated fish paste into an enzymatic hydrolysis tank, adding a commercially available complex enzyme, stirring evenly, performing enzymatic hydrolysis at 50-60° C. for a certain period of time, adding a preservative, and stirring evenly to obtain a concentrated fish paste enzymatic hydrolysis product.

[0082] Performance Testing

[0083] The fermentation products of Examples 1-7 and Comparative Examples 1-7 were tested for content. Note: “—” indicates the lower limit of detection.

[0084] The specific results can be seen in Tables 3 and 4;

[0085] Figure 5 Microscopic photographs before and after fermentation are shown, wherein UFO represents a microscopic photograph of palm meal before fermentation (2 photos, the same sample, different samplings), and FO represents a microscopic photograph of palm meal after fermentation of Example 1 (2 photos, the same sample, different samplings). Figure 6 and Figure 7 These are electron microscope scans of palm meal before and after fermentation (2 images, same sample, different sampling times).

[0086] Table 3 Test results of substance content of fermentation products of Examples 1-7

[0087]

[0088]

[0089] Table 4 Test results of substance content of fermentation products of Comparative Examples 1-7

[0090]

[0091]

[0092] Result analysis:

[0093] 1. Comparison of Examples 1-7 with Comparative Examples 1-7 shows that the Examples are generally superior to the Comparative Examples, indicating that the concentrated fish paste hydrolyzate, the composite bacterial seed solution, the intestinal culture of the mealworm larvae, and the intestinal culture of the black soldier fly are all crucial to the final form and nutritional content of the palm meal, and none of them can be dispensable.

[0094] 2. As shown in Examples 1-7, among the four products, concentrated fish paste hydrolysate, composite bacterial seed solution, mealworm larvae intestinal culture, and black soldier fly intestinal culture, the performance trends show that composite bacterial seed solution and black soldier fly intestinal culture are more important, followed by the other two.

[0095] 3. Comparative Examples 1-3 show that the composite bacterial seed solution, the intestinal culture of the mealworm larvae, and the intestinal culture of the black soldier fly are indispensable;

[0096] 4. As shown in Comparative Examples 4-6, yeast, Aspergillus, and Lactobacillus plantarum are indispensable in the present invention;

[0097] 5. Comparative Example 7 shows that the enzyme used in the concentrated fish paste enzymatic hydrolysis product and whether bacterial hydrolysis and enzymatic hydrolysis are carried out simultaneously are also crucial to the product performance.

[0098] The above embodiments are preferred implementations of the present invention, but the implementations of the present invention are not limited to the above embodiments. Any other changes, modifications, substitutions, combinations, and simplifications that do not deviate from the spirit and principles of the present invention should be considered as equivalent replacement methods and are included in the scope of protection of the present invention.

Claims

1. A composition for palm meal functional solid-state fermentation material, characterized in that: Including palm meal, wheat bran, nutrient solution, concentrated fish paste hydrolyzate, soy molasses, composite bacterial seed liquid, mealworm larvae intestinal culture, black soldier fly intestinal culture; The composite bacterial species in the composite bacterial species seed liquid are composed of Bacillus, yeast, Aspergillus and Lactobacillus plantarum; Each 100kg composition contains 3-5kg of composite bacterial seed liquid; the concentration of each bacteria in the composite bacterial seed liquid is Bacillus content: 1-1.5×10 10 cfu / ml, yeast content: 3.5-5.0×10 9 cfu / ml, Aspergillus content: 3.5-5.5×10 9 cfu / ml, Lactobacillus plantarum content: 5.0-10×10 8 cfu / ml; The weight ratio of the palm meal, wheat bran, nutrient solution, concentrated fish paste hydrolyzate, soybean molasses, composite bacterial seed solution, barley worm larvae intestinal culture, and black soldier fly intestinal culture is: 55-65:12-17:1:4-6:12-17:3-5:1-5:1-5; The composition of the nutrient solution is: 4.0 wt% (NH4)2SO4, 0.4 wt% MgSO4·7H2O, 0.2 wt% KH2PO4, and the balance water; The preparation method of the intestinal culture of the barley worm larvae and the intestinal culture of the black soldier fly is as follows: The corresponding intestinal tract of the worm body is homogenized to obtain a slurry, which is inoculated into a mixed liquid culture medium at an inoculum rate of 3 to 5 wt%, anaerobically cultured for 36 to 48 hours, and freeze-dried to obtain a slurry having a solid content of 45% to 65%; The mixed liquid culture medium contained the following components in a weight ratio: NA:TSA:LB:MRS=4:2:2:2; The preparation method of the concentrated fish paste hydrolysate is as follows: After the fish paste is concentrated, it is transferred to an enzymatic hydrolysis tank, and Bacillus subtilis, actinomycetes, fermentation products of Aspergillus niger and Aspergillus oryzae are added, stirred evenly, and subjected to bacterial hydrolysis and enzymatic hydrolysis at room temperature to obtain concentrated fish paste enzymatic hydrolysis products.

2. The composition for palm meal functional solid-state fermentation material according to claim 1, characterized in that: The weight ratio of the palm meal, wheat bran, nutrient solution, concentrated fish paste hydrolyzate, soybean molasses, composite bacterial seed liquid, barley worm larvae intestinal culture, and black soldier fly intestinal culture is 60:15:1:5:15:4:3:

3.

3. The composition for palm meal functional solid-state fermentation material according to claim 1, characterized in that: The solid content of the concentrated fish paste hydrolysate is 35%.

4. The composition for palm meal functional solid-state fermentation material according to claim 1, characterized in that: During the fermentation of the composition, an appropriate amount of water is added to adjust the moisture content of the composition to 38%.

5. A palm meal fermentation method, characterized in that: The composition according to any one of claims 1 to 4 is used, the moisture content of the composition is adjusted to 35% to 45%, and the composition is placed in an anaerobic fermentation bag and sealed for fermentation at a temperature of 25° C. to 35° C., a fermentation time of 3 to 5 days, 3 to 5 gas release times, and a material pH value of 5.5 to 6.

5.

6. The palm meal fermentation method according to claim 5, wherein: The palm meal has a mesh size of 40-60.

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