CN112194520B

CN112194520B - Black soldier fly compound amino acid fertilizer and preparation method thereof

Info

Publication number: CN112194520B
Application number: CN202011196860.9A
Authority: CN
Inventors: 张文飞; 赵新晨; 高雨; 赵梦秋; 王澜馨; 何佳利
Original assignee: Haikou Haisenyuan Biotechnology Co ltd; Hainan Normal University
Current assignee: Haikou Haisenyuan Biotechnology Co ltd; Hainan Normal University
Priority date: 2020-10-30
Filing date: 2020-10-30
Publication date: 2022-08-30
Anticipated expiration: 2040-10-30
Also published as: CN112194520A

Abstract

The invention provides a black soldier fly compound amino acid fertilizer and a preparation method thereof, wherein the black soldier fly compound amino acid fertilizer comprises the following steps of (1) crushing and drying sun-dried black soldier fly bodies, and stirring and mixing the crushed black soldier fly bodies and dried grape skin powder residues according to a mass ratio of (50-80): 1 under hot air; (2) inoculating bacillus amyloliquefaciens Bt521 into a seed culture medium for fermentation to obtain a seed solution; (3) adding 50-60% of the composite hermetia illucens powder into a fermentation culture medium; (4) inoculating 1-3% of the seed solution to a fermentation culture medium, and performing oscillation fermentation at the temperature of 26-30 ℃ and the rpm of 150-250 for 72-78 h to obtain the black soldier fly compound amino acid fertilizer; the prepared compound amino acid fertilizer has the advantages of rich protein in raw materials, high black soldier fly degradation rate, short fermentation period, rich types of amino acids and amino acid content of 0.32 mg/mL.

Description

Black soldier fly compound amino acid fertilizer and preparation method thereof

Technical Field

The invention relates to the technical field of microbial fermentation engineering, in particular to a black soldier fly compound amino acid fertilizer and a preparation method thereof.

Background

The amino acid can promote the growth of plant root system, strengthen seedling, strengthen plant, strengthen photosynthesis of leaf and disease and pest resistance of crop, and has obvious yield increasing effect on various crops. However, the content of amino acids in natural soil is low, and the intake of amino acids in soil by microorganisms, soil animals and the like in soil often makes it difficult for plants to gain advantages in the process of competing for amino acids in soil. Therefore, many crops now require artificial fertilization to supplement the amino acids required for crop growth. In the process of artificial fertilization, in order to realize high efficiency and no pollution of the fertilizer, high protein substances are generally decomposed into amino acids to be used as biological fertilizers, and for producing the amino acid fertilizer by hydrolyzing the high protein substances at present, protease or microbial degraded fish meal is generally adopted to prepare the fish protein fertilizer, the main components of the fish protein fertilizer are a mixture formed by amino acids, polypeptides and partial proteins, and a small amount of trace elements, lipids, vitamins and other components, the amino acids contained in the fish protein fertilizer are limited in types and low in content, and the fish protein serving as a nutritional agent has slow fertilizer effect and higher price.

The black soldier fly (Hermetia illucens L.) is a rotten hermetidae insect, can take and eat livestock excrement and domestic garbage, is usually used for treating garbage problems such as production of high-value animal protein feed, livestock excrement and the like due to the characteristics of rapid propagation, large biomass, wide feeding range, high absorption and conversion rate, easiness in management, low feeding cost, good animal palatability and the like, so that the black soldier fly can decompose and utilize the garbage excrement to achieve the purpose of environmental protection and resource utilization. The black soldier fly is rich in 45-48% of crude protein and 34-35% of fat, and the crude protein content of the black soldier fly is similar to or slightly higher than that of fresh fish, fish meal and meat and bone meal. The black soldier fly also has comprehensive nutritional ingredients, contains various amino acids required by animals, the content of each amino acid is higher than that of fish meal, the total amount of the essential amino acids is 2.3 times of that of the fish meal, the content of methionine is 2.7 times of that of the fish meal, and the content of lysine is 2.6 times of that of the fish meal.

However, due to the fact that the hermetia illucens propagate quickly, the problems that in the existing process of taking hermetia illucens as a high-protein substance and obtaining an amino acid fertilizer by using a microbial fermentation technology, protein hydrolysis is slow and unstable, undecomposed living bodies are prone to occurring, the content of hydrolyzed amino acid is low, the fermentation period is long and the like exist. Therefore, the preparation process of the black soldier fly compound amino acid fertilizer with short fermentation period and more thorough and stable decomposition is found, and has important significance for realizing the wide application and production of the amino acid liquid fertilizer mainly containing black soldier fly protein.

Disclosure of Invention

Therefore, the invention provides a black soldier fly compound amino acid fertilizer and a preparation method thereof.

The technical scheme of the invention is realized as follows:

a preparation method of a black soldier fly compound amino acid fertilizer comprises the following steps:

(1) pretreating the hermetia illucens: crushing the dried hermetia illucens until the water content is less than or equal to 10%, and stirring and mixing the crushed hermetia illucens and the dried grape skin powder residues according to the mass ratio of (50-80): 1 at 40-50 ℃ to obtain pretreated composite hermetia illucens powder;

(2) preparing a seed solution: inoculating bacillus amyloliquefaciens Bt521 into a seed culture medium for fermentation to obtain a seed solution;

(3) preparation of a fermentation medium: adding the preprocessed composite hermetia illucens powder into a fermentation culture medium according to the addition amount of 50-60% of the mass percentage to obtain the fermentation culture medium added with the composite hermetia illucens powder;

(4) and (3) composite fermentation: inoculating the seed solution into a fermentation culture medium according to the inoculation amount of 1-3% of the volume ratio, and performing oscillation fermentation at the temperature of 26-30 ℃ and the rpm of 150-250 for 72-78 h to obtain the black soldier fly compound amino acid fertilizer. The invention provides a preparation method of a black soldier fly compound amino acid fertilizer, which is characterized in that a microbial fermentation technology is adopted, a specific bacillus amyloliquefaciens Bt521 is inoculated into a fermentation culture medium added with compound black soldier fly powder and is fermented to be complete under the fermentation conditions of 26-30 ℃ and 150-250 rpm, and protein in the black soldier fly body is fully decomposed into amino acid, so that the compound amino acid fertilizer containing multiple amino acids is prepared. According to the invention, through pretreatment of hermetia illucens, the dried hermetia illucens and a certain amount of dried grape skin powder are mixed to obtain composite hermetia illucens powder, and the composite hermetia illucens powder is added into a fermentation culture medium in a certain amount, so that the fermentation decomposition effect of bacillus amyloliquefaciens Bt521 on the composite hermetia illucens powder is further induced, the degradation rate of hermetia illucens is obviously improved, the decomposition is more thorough, the fermentation period is obviously shortened, and the composite amino acid fertilizer mainly containing hermetia illucens protein is obtained.

Compared with the microbial fermentation process in the prior art, the fermentation medium used in the invention has the characteristics of simple components, low cost and wide raw material source, except a small amount of inorganic salt and no additional carbon and nitrogen source; according to the invention, under the optimization of the extraction of strains, the black soldier fly pretreatment technology and the optimization of the fermentation process are combined, so that the addition amount of the black soldier fly fermented by a culture medium reaches 60%, the rapid and sufficient decomposition of the black soldier fly body protein is effectively realized, the fermentation period is obviously shortened to 72h, the amino acid types of the prepared compound amino acid fertilizer are rich, the amino acid content can reach 0.32mg/mL, and the compound amino acid fertilizer has the advantages of rich protein in raw materials, short fermentation period, rich amino acid types, high black soldier fly degradation rate and the like, so that the compound amino acid fertilizer production technology taking the black soldier fly as a high-protein substance is established, and the industrial production and the wider application of the amino acid liquid fertilizer taking the black soldier fly protein as a main material are favorably realized.

Further, the bacillus amyloliquefaciens Bt521 is preserved in China center for type culture Collection in 2020, 9 and 21 months, with the preservation numbers as follows: CCTCC NO. M2020525. The preservation address is as follows: wuhan university, Wuhan, China.

Bacillus amyloliquefaciens Bt521 is separated from a Hainan peak natural protective region, and the 16S rDNA sequence of the Bacillus amyloliquefaciens Bt521 has the highest similarity with Bacillus amyloliquefaciens Strain KJ-16.

Further, the mass ratio of the hermetia illucens to the grape skin powder residue in the composite hermetia illucens powder is 60: 1.

Further, the formula of the seed culture medium is as follows: each liter of water comprises 8-12 g of peptone, 4-5 g of yeast extract and 9-11 g of sodium chloride.

Further, in the step (2), the seed culture medium is subjected to shaking fermentation culture at the temperature of 32-35 ℃ and the rpm of 100-120 for 48-54 h to obtain a seed solution.

Further, in the step (4), the inoculation amount of the seed solution is 2%.

Further, the formula of the fermentation medium is as follows: each liter of water comprises 10-20 g of NaCl and 15-24 g of composite hermetia illucens powder.

Further, in the step (4), the black soldier fly compound amino acid fertilizer is obtained by oscillating and fermenting the black soldier fly in the fermentation medium at the temperature of 28 ℃ and at the rpm of 200 for 72 hours.

The black soldier fly compound amino acid fertilizer prepared by the preparation method of the black soldier fly compound amino acid fertilizer.

Compared with the prior art, the invention has the beneficial effects that:

(1) the fermentation medium used in the invention has the characteristics of simple components, low price and wide raw material source, except a small amount of inorganic salt and no additional carbon and nitrogen source;

(2) according to the invention, through the selection of strains and the optimization of a fermentation process, the composite fermentation period of the hermetia illucens is effectively shortened to 72 hours;

(3) according to the invention, through optimization of the black soldier fly pretreatment technology and the fermentation process, the combination of the dried grape skin residue and the dried black soldier fly is utilized, and the fermentation decomposition of the composite black soldier fly powder by the strain is induced under a certain fermentation condition, so that the fermentation period is obviously shortened, the addition amount of the black soldier fly fermented by the culture medium reaches 60%, the rapid and sufficient decomposition of black soldier fly body protein is effectively realized, the variety of amino acids of the prepared composite amino acid fertilizer is rich, and the amino acid content reaches 0.32 mg/mL;

(4) the production technology of the compound amino acid fertilizer taking the hermetia illucens as a high-protein substance is established, and the industrial production and the wider application of the amino acid liquid fertilizer taking the hermetia illucens as the main protein are facilitated.

Drawings

FIG. 1 is a colony morphology of Bacillus amyloliquefaciens Bt521 according to the present invention;

FIG. 2 is a diagram showing the cell morphology (Coomassie blue staining) of a Bacillus amyloliquefaciens Bt521 under an optical microscope;

FIG. 3 is a 16S rRNA gene PCR amplified fragment agarose electrophoresis picture of Bacillus amyloliquefaciens Bt521 of the present invention;

FIG. 4 is an amino acid map of free states of black soldier flies fermented by Bacillus amyloliquefaciens Bt521 according to the present invention.

Detailed Description

In order that the technical contents of the invention may be better understood, specific examples are provided below to further illustrate the invention.

The experimental methods used in the examples of the present invention are all conventional methods unless otherwise specified.

The materials, reagents and the like used in the examples of the present invention can be obtained commercially without specific description.

Example 1-a method for preparing a black soldier fly compound amino acid fertilizer, comprising the steps of:

(1) b, black soldier fly pretreatment: crushing the sun-dried hermetia illucens, drying until the water content is less than or equal to 10%, and stirring and mixing the crushed hermetia illucens and the dried grape skin powder residues at 40 ℃ according to the mass ratio of 50:1 to obtain pretreated composite hermetia illucens powder;

(2) preparing a seed solution: inoculating Bacillus amyloliquefaciens Bt521 into a seed culture medium, and performing shake fermentation culture at 32 ℃ and 100rpm for 48h to obtain a seed solution; the formula of the seed culture medium is as follows: each liter of water comprises 8g of peptone, 4g of yeast extract and 9g of sodium chloride;

(3) preparation of a fermentation medium: adding the pretreated composite hermetia illucens powder into a fermentation culture medium according to the addition amount of 60% by mass to obtain the fermentation culture medium added with the composite hermetia illucens powder; the formula of the fermentation medium is as follows: each liter of water comprises 10g of NaCl and 15g of composite hermetia illucens powder;

(4) and (3) composite fermentation: inoculating the seed solution into a fermentation culture medium according to the inoculation amount of 1% of the volume ratio, and performing oscillation fermentation for 78h at 26 ℃ and 150rpm to obtain the liquid black soldier fly compound amino acid fertilizer.

Example 2-a method for preparing a black soldier fly compound amino acid fertilizer, comprising the steps of:

(1) pretreating the hermetia illucens: crushing the sun-dried hermetia illucens, drying until the water content is less than or equal to 10%, and stirring and mixing the crushed hermetia illucens and the dried grape skin powder residues at 50 ℃ in a mass ratio of 80:1 to obtain pretreated composite hermetia illucens powder;

(2) preparing a seed solution: inoculating Bacillus amyloliquefaciens Bt521 into a seed culture medium, and performing shaking fermentation culture at 35 ℃ and 120rpm for 54h to obtain a seed solution; the formula of the seed culture medium is as follows: each liter of water comprises 12g of peptone, 5g of yeast extract and 11g of sodium chloride;

(3) preparation of a fermentation medium: adding the pretreated composite hermetia illucens powder into a fermentation culture medium according to the addition amount of 50% by mass to obtain the fermentation culture medium added with the composite hermetia illucens powder; the formula of the fermentation medium is as follows: each liter of water comprises 20g of NaCl and 20g of composite hermetia illucens powder;

(4) and (3) composite fermentation: inoculating the seed solution into a fermentation culture medium according to the inoculation amount of 3% of the volume ratio, and performing oscillation fermentation at 30 ℃ and 250rpm for 72h to obtain the liquid black soldier fly compound amino acid fertilizer.

Example 3-a method for preparing a black soldier fly compound amino acid fertilizer, comprising the steps of:

(1) pretreating the hermetia illucens: crushing the sun-dried hermetia illucens, drying until the water content is less than or equal to 10%, and stirring and mixing the crushed hermetia illucens and the dried grape skin powder residues at 45 ℃ according to the mass ratio of 60:1 to obtain pretreated composite hermetia illucens powder;

(2) preparing a seed solution: inoculating Bacillus amyloliquefaciens Bt521 into a seed culture medium, and performing shake fermentation culture at 33 ℃ and 110rpm for 54h to obtain a seed solution; the formula of the seed culture medium is as follows: each liter of water comprises 10g of peptone, 4.5g of yeast extract and 10g of sodium chloride

(3) Preparation of a fermentation medium: adding the pretreated composite hermetia illucens powder into a fermentation culture medium according to the addition amount of 60% by mass to obtain the fermentation culture medium added with the composite hermetia illucens powder; the formula of the fermentation medium is as follows: each liter of water comprises 16g of NaCl and 24g of composite hermetia illucens powder;

(4) and (3) composite fermentation: inoculating the seed solution into a fermentation culture medium according to the inoculation amount of 2% of the volume ratio, and performing oscillation fermentation for 72h at the temperature of 28 ℃ and the speed of 200rpm to obtain the liquid black soldier fly compound amino acid fertilizer.

The free amino acids of the liquid hermetia illucens compound amino acid fertilizer obtained after the fermentation for 72 hours in the above example 3 are detected, and the results are shown in the following table:

TABLE 1 free amino acids of liquid Hermetia illucens compound amino acid fertilizer

Name of Compound	Retention time/(min)	Response to	content/(mg/mL)	Content/(%)
					Phosphoserine	3.245	2226.535	0.01018	3.2
Taurine	4.819	1929.914	0.00668	2.1
					Urea	7.360	1006.334	0.00648	2.0
Aspartic acid	17.821	482.234	0.00149	0.5
					Threonine	24.512	604.561	0.00247	0.8
Serine	26.763	506.997	0.00140	0.4
					Asparagine	30.152	105.248	0.00106	0.3
Glutamic acid	32.344	6189.885	0.08427	26.2
					Glycine	46.648	1650.650	0.00467	1.5
Alanine	48.845	3287.848	0.00768	2.4
					Alpha-amino butyric acid	52.816	756.425	0.00212	0.7
Valine	56.189	1510.780	0.00473	1.5
					Methionine	61.917	730.484	0.00406	1.3
Isoleucine	64.981	763.454	0.00340	1.1
					Leucine	66.315	1658.978	0.00540	1.7
Tyrosine	69.139	2589.137	0.01237	3.8
					Phenylalanine	72.568	6476.995	0.09001	28.0
Beta alanine	74.867	83.405	0.00074	0.2
					Beta-aminoisobutyric acid	75.579	163.513	0.00168	0.5
Gamma amino butyric acid	77.621	132.978	0.00039	0.1
					Histidine	82.536	8819.363	0.03088	9.6
3 methyl histidine	83.733	126.660	0.00055	0.2
					Carnosine	88.528	169.437	0.00180	0.6
Tryptophan	89.907	1468.286	0.01875	5.8
					Ornithine	93.272	267.169	0.00099	0.3
Lysine	94.789	1218.771	0.00721	2.2
					Arginine	102.952	1978.556	0.01052	3.3
Proline	43.408	109.927	0.00201	0.0
					Total up to	--	--	0.32199	100

As can be seen from the table above, the compound amino acid fertilizer prepared by the preparation method of the black soldier fly compound amino acid fertilizer has rich amino acid types, and the amino acid content can reach 0.32 mg/mL. Wherein, the free amino acid content map is shown in FIG. 4.

The colony morphology and the cell morphology of the bacillus amyloliquefaciens Bt521 are shown in a figure 1 and a figure 2; the DNA sequence of Bacillus amyloliquefaciens Bt521 is as follows:

TGCAAGTCGAGCGGACAGATGGGAGCTTGCTCCCTGATGTCAGCGGCGGACGGGTGAGTAACACGTGGGTAACCTGCCTGTAAGACTGGGATAACTCCGGGAAACCGGGGCTAATACCGGATGCTTGATTGAACCGCATGGTTCAATTATAAAAGGTGGCTTTTAGCTACCACTTACAGATGGACCCGCGGCGCATTAGCTAGTTGGTGAGGTAACGGCTCACCAAGGCAACGATGCGTACCCGACCTGAGAGGGTGATCGGCCACACTGGGACTGAGACACGGCCCAAACTCCTACGGGAGGCAGCAGTAGGGAATCTTCCGCAATGGACAAAAGTCTGACGGACCAACGCCGCGTGAGTGATGAAGGTTTTCGGATCGTAAAACTTCTGTTGTTAGGGAAGAACAAGTACCGTTCGAATAGGGCGGTACCTTGACGGTACCTAACCAAAAAGCCACGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGTGGCAAGCGTTGTCCGGAATTATTGGGCGTAAAGCGCGCGCAGGCGGTTTCTTAAGTCTGATGTGAAAGCCCCCGGCTCAACCGGGGAGGGTCATTGGAAACTGGGGAACTTGAGTGCAGAAGAGGAGAGTGGAATTCCACGTGTAGCGGTGAAATGCGTAGAGATGTGGAGGAACACCAGTGGCGAAGGCGACTCTCTGGTCTGTAACTGACGCTGAGGCGCGAAAGCGTGGGGAGCGAACAGGATTAGATACCCTGGTAGTCCACGCCGTAAACGATGAGTGCTAAGTGTTAGAGGGTTTCCGCCCTTTAGTGCTGCAGCAAACGCATTAAGCACTCCGCCTGGGGAGTACGGTCGCAAGACTGAAACTCAAAGGAATTGACGGGGGCCCGCACAAGCGGTGGAGCATGTGGTTTAATTCGAAGCAACGCGAAGAACCTTACCAGGTCTTGACATCCTCTGACAACCCTAGAGATAGGGCTTCCCCTTCGGGGGCAGAGTGACAGGTGGTGCATGGTTGTCGTCAGCTCGTGTCGTGAGATGTTGGGTTAAGTCCCGCAACGAGCGCAACCCTTGATCTTAGTTGCCAGCATTCAGTTGGGCACTCTAAGGTGACTGCCGGTGACAAACCGGAGGAAGGTGGGGATGACGTCAAATCATCATGCCCCTTATGACCTGGGCTACACACGTGCTACAATGGGCAGAACAAAGGGCAGCGAAGCCGCGAGGCTAAGCCAATCCCACAAATCTGTTCTCAGTTCGGATCGCAGTCTGCAACTCGACTGCGTGAAGCTGGAATCGCTAGTAATCGCGGATCAGCATGCCGCGGTGAATACGTTCCCGGGCCTTGTACACACCGCCCGTCACACCACGAGAGTTTGTAACACCCGAAGTCGGTGAGGTAACCTTTTGGAGCCAGCCGCCGAA

the invention carries out agarose electrophoresis on the 16S rRNA gene PCR amplified fragment of Bacillus amyloliquefaciens Bt521 as shown in figure 3;

comparative example 1-method of preparing a black soldier fly compound amino acid fertilizer according to example 3, with the difference that: in the step (1), the sun-dried hermetia illucens are crushed, dried until the water content is less than or equal to 10%, and then are not mixed with the dried grape skin powder residues, and are directly added into a fermentation culture medium according to the addition amount of 60% by mass; the rest is the same as in example 3.

Comparative example 2-method for preparing a black soldier fly compound amino acid fertilizer according to example 3, with the difference that: in the step (1), the fresh hermetia illucens larvae are directly crushed and then directly mixed with the undried grape skin powder residue at normal temperature according to the mass ratio of 60:1 to obtain the composite hermetia illucens powder, and the rest is the same as that in the example 3.

Comparative example 3-method for preparing a black soldier fly compound amino acid fertilizer according to example 3, with the difference that: in the step (2), the seed solution was inoculated into the fermentation medium in an amount of 5% by volume, and the rest was the same as in the example.

Comparative example 4-method of preparing a black soldier fly compound amino acid fertilizer according to example 3, with the difference that: in the step (2), commercial bacillus amyloliquefaciens is adopted to replace bacillus amyloliquefaciens Bt521 to be inoculated into a seed culture medium for fermentation, so as to obtain a seed solution, and the rest is the same as the step in the embodiment 3.

The method for preparing the black soldier fly compound amino acid fertilizer according to the example 3 and the comparative examples 1 to 4 is set as an experimental group 1 and a control group 1 to 4.

(1) The degradation rates of the composite hermetia illucens powder at different periods are respectively measured, wherein the degradation rate of the composite hermetia illucens powder is (dry weight of the composite hermetia illucens powder added before fermentation-dry weight of filter residue after fermentation)/dry weight of the composite hermetia illucens powder added before fermentation multiplied by 100%, and the results are as follows:

TABLE 2 degradation rates of the composite hermetia illucens powder at different periods

As can be seen from the above table, the degradation rates of the hermetia illucens powder in the comparison group 1 and the composite hermetia illucens powder in the comparison group 2 are both reduced obviously, which indicates that the mixing of a certain amount of dried grape skin powder and dried hermetia illucens can be beneficial to inducing the fermentation decomposition effect of bacillus amyloliquefaciens Bt521 on the composite hermetia illucens powder, so that the degradation rate of the hermetia illucens is improved obviously, and the fermentation period is shortened obviously; meanwhile, the hermetia illucens larvae are dried, the moisture content of the composite hermetia illucens powder is controlled, the protein decomposition efficiency is improved, and the decomposition is more thorough and stable. The degradation rate of the composite hermetia illucens powder in the comparison group 4 and the comparison group 3 is relatively low compared with that in the embodiment 3, which shows that the degradation rate of the composite hermetia illucens powder is accelerated by selecting strains, and under certain fermentation conditions, when the inoculation amount of the bacillus amyloliquefaciens Bt521 seed liquid is controlled to be too much, the conditions of too fast fermentation rate in the early stage of thallus fermentation, insufficient oxygen dissolution in the later stage, premature senility, autolysis and the like of the thallus can be caused, and finally the degradation effect of the composite hermetia illucens powder is obviously reduced.

(2) Respectively and equivalently sampling the liquid hermetia illucens compound amino acid fertilizer, and determining the amino acid content under different fermentation periods, wherein the determination results are as follows:

TABLE 3 amino acid content at different fermentation stages

As can be seen from the above table, in the same fermentation period, the total content of free amino acids in the black soldier fly compound amino acid fertilizer obtained in the experimental group 1 is obviously increased, while the total content of free amino acids after fermentation for 72 hours in the comparative groups 1 to 4 is lower, the decomposition degree of the compound black soldier fly powder is lower, and the proteolysis is slow and unstable. In the comparison group 3, although the decomposition speed of the amino acid at the early stage is higher, the composite hermetia illucens powder cannot be sufficiently decomposed after fermentation for 72 hours; the method has the advantages that under the optimization of the extraction of the strains, the black soldier fly pretreatment technology and the optimization of the fermentation process are combined, so that the addition amount of the black soldier fly in the fermentation medium reaches 60%, the degradation efficiency is improved, the rapid degradation of the black soldier fly body protein is realized, the black soldier fly body protein is more sufficiently decomposed, after the black soldier fly body protein is decomposed for 72 hours, the variety of amino acids in the prepared compound amino acid fertilizer is rich, the amino acid content can reach 0.32mg/mL, and the amino acid content of the liquid black soldier fly compound amino acid fertilizer is remarkably improved.

The above description is only for the purpose of illustrating the preferred embodiments of the present invention and is not to be construed as limiting the invention, and any modifications, equivalents, improvements and the like that fall within the spirit and principle of the present invention are intended to be included therein.

Sequence listing

<110> Haikou Heisenyurt Biotech Ltd

HAINAN NORMAL University

<120> hermetia illucens compound amino acid fertilizer and preparation method thereof

<160> 1

<170> SIPOSequenceListing 1.0

<210> 1

<211> 1423

<212> DNA

<213> Artificial Sequence (Artificial Sequence)

<400> 1

tgcaagtcga gcggacagat gggagcttgc tccctgatgt cagcggcgga cgggtgagta 60

acacgtgggt aacctgcctg taagactggg ataactccgg gaaaccgggg ctaataccgg 120

atgcttgatt gaaccgcatg gttcaattat aaaaggtggc ttttagctac cacttacaga 180

tggacccgcg gcgcattagc tagttggtga ggtaacggct caccaaggca acgatgcgta 240

cccgacctga gagggtgatc ggccacactg ggactgagac acggcccaaa ctcctacggg 300

aggcagcagt agggaatctt ccgcaatgga caaaagtctg acggaccaac gccgcgtgag 360

tgatgaaggt tttcggatcg taaaacttct gttgttaggg aagaacaagt accgttcgaa 420

tagggcggta ccttgacggt acctaaccaa aaagccacgg ctaactacgt gccagcagcc 480

gcggtaatac gtaggtggca agcgttgtcc ggaattattg ggcgtaaagc gcgcgcaggc 540

ggtttcttaa gtctgatgtg aaagcccccg gctcaaccgg ggagggtcat tggaaactgg 600

ggaacttgag tgcagaagag gagagtggaa ttccacgtgt agcggtgaaa tgcgtagaga 660

tgtggaggaa caccagtggc gaaggcgact ctctggtctg taactgacgc tgaggcgcga 720

aagcgtgggg agcgaacagg attagatacc ctggtagtcc acgccgtaaa cgatgagtgc 780

taagtgttag agggtttccg ccctttagtg ctgcagcaaa cgcattaagc actccgcctg 840

gggagtacgg tcgcaagact gaaactcaaa ggaattgacg ggggcccgca caagcggtgg 900

agcatgtggt ttaattcgaa gcaacgcgaa gaaccttacc aggtcttgac atcctctgac 960

aaccctagag atagggcttc cccttcgggg gcagagtgac aggtggtgca tggttgtcgt 1020

cagctcgtgt cgtgagatgt tgggttaagt cccgcaacga gcgcaaccct tgatcttagt 1080

tgccagcatt cagttgggca ctctaaggtg actgccggtg acaaaccgga ggaaggtggg 1140

gatgacgtca aatcatcatg ccccttatga cctgggctac acacgtgcta caatgggcag 1200

aacaaagggc agcgaagccg cgaggctaag ccaatcccac aaatctgttc tcagttcgga 1260

tcgcagtctg caactcgact gcgtgaagct ggaatcgcta gtaatcgcgg atcagcatgc 1320

cgcggtgaat acgttcccgg gccttgtaca caccgcccgt cacaccacga gagtttgtaa 1380

cacccgaagt cggtgaggta accttttgga gccagccgcc gaa 1423

Claims

1. A preparation method of a black soldier fly compound amino acid fertilizer is characterized by comprising the following steps: the method comprises the following steps:

(2) preparing a seed solution: inoculating Bacillus amyloliquefaciens Bt521 into a seed culture medium, and performing shaking fermentation culture at the temperature of 32-35 ℃ and the rpm of 100-120 for 48-54 h to obtain seed liquid;

the formula of the seed culture medium is as follows: each liter of water comprises 8-12 g of peptone, 4-5 g of yeast extract and 9-11 g of sodium chloride;

(4) performing composite fermentation: inoculating the seed solution into a fermentation medium according to the inoculation amount of 1-3% of the volume ratio, wherein the formula of the fermentation medium is as follows: each liter of water comprises 10-20 g of NaCl and 15-24 g of composite hermetia illucens powder; and carrying out oscillatory fermentation at 26-30 ℃ and 150-250 rpm for 72-78 h to obtain the black soldier fly compound amino acid fertilizer.

2. The method for preparing the black soldier fly compound amino acid fertilizer according to claim 1, wherein the method comprises the following steps: the bacillus amyloliquefaciens Bt521 is preserved in the China center for type culture Collection with the preservation number as follows: CCTCC NO. M2020525.

3. The method for preparing the black soldier fly compound amino acid fertilizer according to claim 1, wherein the method comprises the following steps: the mass ratio of the hermetia illucens to the grape skin powder residue in the composite hermetia illucens powder is 60: 1.

4. The method for preparing the black soldier fly compound amino acid fertilizer according to claim 1, wherein the method comprises the following steps: in the step (4), the inoculation amount of the seed liquid is 2%.

5. The method for preparing the black soldier fly compound amino acid fertilizer according to claim 1, wherein the method comprises the following steps: in the step (4), oscillating and fermenting the black soldier fly in a fermentation culture medium for 72 hours at the temperature of 28 ℃ and at the rpm of 200 to obtain the black soldier fly compound amino acid fertilizer.

6. The black soldier fly compound amino acid fertilizer prepared by the method for preparing the black soldier fly compound amino acid fertilizer according to any one of claims 1 to 5.