CN110496849B - Efficient processing method of shrimp waste - Google Patents
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K10/00—Animal feeding-stuffs
- A23K10/10—Animal feeding-stuffs obtained by microbiological or biochemical processes
- A23K10/12—Animal feeding-stuffs obtained by microbiological or biochemical processes by fermentation of natural products, e.g. of vegetable material, animal waste material or biomass
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- A—HUMAN NECESSITIES
- A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
- A23K—FODDER
- A23K10/00—Animal feeding-stuffs
- A23K10/20—Animal feeding-stuffs from material of animal origin
- A23K10/26—Animal feeding-stuffs from material of animal origin from waste material, e.g. feathers, bones or skin
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B09—DISPOSAL OF SOLID WASTE; RECLAMATION OF CONTAMINATED SOIL
- B09B—DISPOSAL OF SOLID WASTE NOT OTHERWISE PROVIDED FOR
- B09B3/00—Destroying solid waste or transforming solid waste into something useful or harmless
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B09—DISPOSAL OF SOLID WASTE; RECLAMATION OF CONTAMINATED SOIL
- B09B—DISPOSAL OF SOLID WASTE NOT OTHERWISE PROVIDED FOR
- B09B5/00—Operations not covered by a single other subclass or by a single other group in this subclass
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Abstract
The invention relates to a high-efficiency processing method of shrimp wastes. The high-efficiency processing method comprises the following steps: s1: crushing the shrimp waste, sterilizing, adding MRS culture and water to prepare a culture medium; s2: inoculating Lactococcus garvieae LGHK2, and fermenting under shaking for at least 36 h. The treatment method provided by the invention utilizes Lactococcus garvieae LGHK2 to ferment, the strain has strong acid production capability and is used for fermenting fresh shrimp shells, and the decalcification rate can reach 69.47% -77.75%; when the shrimp heads are used for fermenting fresh shrimp heads, the pH value of fermentation liquor can be as low as 3.3, the apparent hydrolysis degree can reach 40.74 percent at most, the protein removal rate reaches 97.0 percent, and the shrimp heads have high-efficiency calcium and protein dissolving capacity; the processing method provided by the invention has higher utilization rate and better environmental compatibility.
Description
Technical Field
The invention belongs to the field of microorganisms, and particularly relates to a high-efficiency treatment method of shrimp waste.
Background
China has wide ocean resources, is a large shrimp breeding country, and has a plurality of shrimp product processing factories. At present, the processing of shrimps in China mainly takes shrimp meat as main raw material, and a large amount of waste byproducts such as shrimp shells and shrimp heads which account for about 30-40% of the whole shrimps are generated while the shrimp meat is processed. The byproducts contain abundant proteins, chitin, chitosan, astaxanthin, unsaturated fatty acid and essential amino acid, and have wide development and utilization prospects. However, due to the limitation of technical conditions, the utilization rate of the byproducts is not high, and a large amount of land is occupied, thereby causing environmental pollution. In fact, the shrimp heads and shells contain a large amount of valuable resources such as chitin, astaxanthin, protein, high-quality calcium and the like. Only taking chitin as an example, the chitin is nontoxic, tasteless and free of side effects, and has wide application in the fields of medicine, paper making, printing and dyeing, plant growth hormone, daily chemicals, food industry, fruit preservation, textile industry, environmental protection and the like; in addition, the important derivative obtained by the chitosan and the phthalidyl is chitosan, which is a fine chemical with high added value, the industrial grade chitosan is quite expensive internationally, and the high-quality chitosan is imported domestically.
At present, the large-scale utilization of the shrimp heads and the shrimp shells mainly has two ways, one is that the shrimp heads and the shrimp shells are dried and crushed to be used as raw materials of mineral additives such as animal feed calcium and the like; secondly, chitin is produced. The former has low output value, which causes great resource loss; in addition, the chitin and mineral substances in the shrimp shells cannot be fully recovered due to insufficient calcium dissolution, and the added value of the final product is low. There are also some scholars and companies who recover proteins, astaxanthin and the like by treating shrimp waste using an enzymatic hydrolysis method. The enzymolysis method has the advantages that the raw materials are convenient and easy to obtain, the operation is simple and convenient, the defects are that the enzymolysis solution and the recovered protein have heavy fishy and astringent taste, the direct utilization is difficult, and additives or reprocessing is often needed to remove the fishy and astringent taste, for example, activated carbon and yeast powder are adopted to remove fishy and bitter taste of the enzymolysis solution of shrimp heads and shrimp shells; some monomer amino acid and reducing sugar are added to generate Maillard reaction to remove fishy and astringent taste, and the processing methods make the processing technology complicated and increase the cost.
In order to solve the problems, the research and report of fermenting the shrimp heads and the shrimp shells by using the lactic acid coccus are reported at home and abroad, but the lactic acid coccus can not be put into large-scale production at present. Because different microorganisms have different abilities to produce acid and produce protease enzymes themselves, there is also a large difference in the growth and effectiveness of the microorganisms under different culture conditions. Most of the results of microbial fermentation studies have a common problem that insufficient acid and protease are generated, which results in insufficient desalting and deproteinization, and the production speed of acid is slow, which cannot effectively prevent the putrefaction of shrimp shell waste. Therefore, how to effectively utilize shrimp wastes to produce bioactive substances with high added values is a hot point of research and development at home and abroad at present, and especially how to research and develop a microbial fermentation method with production and application values besides the traditional acid-base method is a core problem in the field.
Disclosure of Invention
The invention aims to overcome the defects and shortcomings that the existing technology for fermenting the shrimp heads and the shrimp shells by using the lactococcus lactis is insufficient in desalination and deproteinization, the production speed of acid is low, and the waste of the shrimp shells cannot be effectively prevented from being corrupted, so that large-scale production and application cannot be realized, and provides an efficient treatment method for shrimp wastes. The treatment method provided by the invention utilizes Lactococcus garvieae LGHK2 to ferment, is simple to operate and easy for large-scale production, has high growth speed, easy culture, strong activity and strong acid production capacity, is used for fermenting fresh shrimp shells, and has decalcification rate of 69.47-77.75%; when the shrimp heads are used for fermenting fresh shrimp heads, the pH value of fermentation liquor can be as low as 3.3, the apparent hydrolysis degree can reach 40.74 percent at most, the protein removal rate reaches 97.0 percent, and the shrimp heads have high-efficiency calcium and protein dissolving capacity; the processing method provided by the invention has the advantages of high utilization rate of the shrimp waste and better environmental compatibility.
In order to achieve the purpose, the invention adopts the following technical scheme:
a high-efficiency processing method of shrimp wastes comprises the following steps:
s1: crushing the shrimp waste, sterilizing, adding an MRS culture medium and water to prepare a culture medium with a solid-to-liquid ratio of 1: 1-20, and adjusting the pH to 5.5-7.5;
s2: inoculating Lactococcus garvieae LGHK2, and performing shaking fermentation for at least 36 h; the Lactococcus lactis garvieae LGHK2 is preserved in Guangdong province microbial strain preservation center in 2018, 9 and 5, with the preservation number being GDMCC No.60443 and the preservation address being No. 59 floor 5 of Michelia Tokyo 100, Guangzhou, Guangdong province.
The invention discloses a strain separated and purified from Penaeus vannamei Boone shells, which is identified as Lactococcus garvieae LGHK2 and is preserved in Guangdong province microorganism strain collection center in 2018, 9 and 5 days, and the preservation number is GDMCC No. 60443. The storage address is No. 59 building of No. 59 Dazhou school of Jiuli Zhonglu 100, Guangzhou, Guangdong province.
The treatment method provided by the invention utilizes Lactococcus garvieae LGHK2 to ferment, is simple to operate and is easy for large-scale production; the lactococcus lactis has the advantages of high growth speed, easiness in culture, strong activity and strong acid production capacity, is used for fermenting fresh shrimp shells, and can achieve the decalcification rate of 69.47-77.75%; when the shrimp heads are used for fermenting fresh shrimp heads, the pH value of fermentation liquor can be as low as 3.3, the apparent hydrolysis degree can reach 40.74 percent at most, the protein removal rate reaches 97.0 percent, and the shrimp heads have high-efficiency calcium and protein dissolving capacity; the processing method provided by the invention has the advantages of high utilization rate of the shrimp waste and better environmental compatibility.
The Lactococcus garvieae LGHK2 provided by the invention was identified, and the results are as follows.
First, preliminary identification of Lactococcus garvieae LGHK2
(1) Morphological characteristics
Lactococcus garvieae LGHK2 did not form a transparent ring on MRS agar medium containing 2% calcium carbonate, and the colony was small, milky white or pale yellow, and had circular protrusions on the surface (FIG. 1A). Gram staining was positive (+), cells were spherical or oval, often paired, and had no spores (FIG. 1B).
(2) Physiological and biochemical characteristics
Physiological and biochemical characteristics of strain LGHK2 are shown in FIG. 2 and Table 1, physiological and biochemical characteristics of strain LGHK2 are substantially similar to those of lactococcus garvieae, and strain LGHK2 was preliminarily determined to be lactococcus garvieae.
TABLE 1 results of physiological and biochemical tests of Strain LGHK2
Note: +: positive; -: and (4) negativity.
Second, molecular identification results and analysis
(1) Genomic DNA extraction results
Total DNA of Lactococcus garvieae LGHK2 was extracted using the bacterial genomic DNA rapid extraction kit, and the results are shown in FIG. 3, M: DL5000 Marker; 1: lactococcus garvieae LGHK 2.
(2) PCR amplification results
The electrophoretic band of the 16S rDNA PCR product is 1500bp, the electrophoretic band of the PCR product of the gyrB gene is about 1200bp, and the electrophoretic band can be obtained from the graph shown in figure 4 and figure 5, and the PCR amplification bands of the 16S rDNA and the gyrB are clear and bright and are consistent with the target band.
(3) Analysis of 16S rDNA sequencing results
The 16S rDNA gene fragment size of strain LGHK2 was 1464bp, and the strain LGHK2 was up to 99% identical to lactococcus garvieae (accession No. AP017373.1) when compared to the BLAST software at the NCBI accession.
(4) Analysis of sequencing results for gyrB
The size of the gyrB gene fragment of the strain LGHK2 is 607bp, the similarity comparison can be carried out by using BLAST software when the strain is registered to NCBI, the consistency of the strain LGHK2 and lactococcus garvieae (accession number: LC377166.1) is as high as 98%, and the sequence shown in SEQ ID NO: 1.
the strain LGHK2 can be determined to be accurately identified as lactococcus garvieae (Lactocousgarvieae) by combining the morphological characteristics of the strain, the physiological and biochemical test results and the 16S rDNA and gyrB gene sequence analysis. The Lactococcus garvieae belongs to the phylum Firmicutes, the class Bacillales (Bacillus), the order Lactobacillales (Lactobacillus), the family Streptococcaceae (Streptococcus), and the genus Lactococcus (Lactobacillus). The cells are spherical or oval, 0.5-1.2 μm is multiplied by 0.5-1.5 μm, and are paired and short-chained in a liquid culture medium. No spore, gram positive. No movement, no capsule. Facultative anaerobic. Chemoheterotrophic, fermentative metabolism, which produces acid by fermentation of some carbohydrates, mainly L (+) -lactic acid, without producing gas. The nutrition requirement is complex, catalase negative and oxidase negative. The optimum temperature is 30 ℃ and can grow at 10 ℃ without growing at 45 ℃ in 0.5% NaCl, usually Lancefield serogroup N. Found in dairy and vegetable products.
According to the invention, the lactobacillus garvieae LGHK2 is used for fermenting and treating the shrimp waste, so that the autolysis time for treating the shrimp heads and the shrimp shells can be prolonged, the residual protein in the shrimp waste can be efficiently degraded, the subsequent chitin production process is simplified, and a new practical method is provided for further extracting high-added-value products such as chitin.
In addition, the invention utilizes Lactococcus garvieae LGHK2 to ferment and treat the shrimp waste, solves the problems that the quality and efficiency of obtaining high value-added products by treating the shrimp waste by the existing chemical method are not high, the environmental pollution is caused and the like, has simple preparation method and high effect, and has wide application development and market prospect in the fields of preparation of breeding feeds and microecologics, preparation of chitin and the like.
Preferably, the carbon source in the MRS medium in S1 is one or more of glucose, sucrose, or lactose.
More preferably, the composition of the MRS medium in S1 is as follows: 20.0g of carbon source, 10.0g of peptone, 10.0g of beef extract, 5.0g of yeast extract, 5.0g of anhydrous sodium acetate, 2.0g of dipotassium hydrogen phosphate, 2.0g of diamine hydrogen citrate, 0.58g of MgSO4.7H2O0, 0.25g of MnSO4.H2O and 15.0g of agar, 1000mL of distilled water, adjusting pH to 6.8, and sterilizing at 121 ℃ for 20 min.
Preferably, the fermentation temperature in S2 is 34-40 ℃.
Preferably, the shrimp waste in S1 is one or more of a shrimp head or a shrimp shell.
Preferably, when the shrimp waste is shrimp shells, the solid-to-liquid ratio in S1 is 1: 5-20.
More preferably, the solid-to-liquid ratio in S1 is 1: 5.
Preferably, when the shrimp waste is shrimp shells, the pH value of the S1 is adjusted to 6.8-7.0.
Preferably, when the shrimp waste is shrimp shells, the mass ratio of the shrimp shells to the MRS culture medium in S1 is 5: 2.
Preferably, when the shrimp waste is shrimp shells, the inoculation amount of the Lactococcus garvieae LGHK2 in S2 is 4.08X 108Per gram.
Preferably, when the shrimp waste is shrimp heads, the solid-to-liquid ratio in S1 is 1: 1-5;
more preferably, the solid-to-liquid ratio in S1 is 1: 4.
Preferably, when the shrimp waste is shrimp heads, the pH value in S1 is adjusted to 6.8-7.0.
Preferably, when the shrimp waste is shrimp heads, the mass ratio of the shrimp heads to the MRS culture medium in S1 is 5: 3.
Preferably, when the shrimp waste is shrimp heads, the inoculation amount of the Lactococcus garvieae LGHK2 in S2 is 1.02X 108Per gram.
Compared with the prior art, the invention has the following beneficial effects:
(1) the treatment method provided by the invention utilizes Lactococcus garvieae LGHK2 to ferment, is simple to operate and is easy for large-scale production; the lactococcus lactis has the advantages of high growth speed, easy culture, strong activity and strong acid production capability, is used for fermenting fresh shrimp shells, and has the decalcification rate of 69.47-77.75%; when the shrimp heads are used for fermenting fresh shrimp heads, the pH value of fermentation liquor can be as low as 3.3, the apparent hydrolysis degree can reach 40.74 percent at most, the protein removal rate reaches 97.0 percent, and the shrimp heads have high-efficiency calcium and protein dissolving capacity; the processing method provided by the invention has the advantages of high utilization rate of the shrimp waste and better environmental compatibility. .
(2) According to the invention, the lactobacillus garvieae LGHK2 is used for fermenting and treating the shrimp waste, so that the autolysis time for treating the shrimp heads and the shrimp shells can be prolonged, the residual protein in the shrimp waste can be efficiently degraded, the subsequent chitin production process is simplified, and a new practical method is provided for further extracting high-added-value products such as chitin.
(3) The method utilizes Lactococcus garvieae LGHK2 to treat the shrimp waste by fermentation, solves the problems that the quality and efficiency of obtaining high value-added products by treating the shrimp waste by the existing chemical method are not high, the environmental pollution is caused and the like, has simple preparation method and high effect, and has wide application development and market prospect in the fields of preparation of breeding feeds and microecologics, preparation of chitin and the like.
Drawings
FIG. 1 is a colony map (A) and a gram stain map (B) of Lactococcus garvieae LGHK 2;
FIG. 2 shows the results of physiological and biochemical experiments with Lactococcus garvieae LGHK 2;
FIG. 3 shows the results of genomic DNA extraction of Lactococcus garvieae LGHK 2;
FIG. 4 is an electrophoretic band of the 16S rDNA PCR product;
FIG. 5 is an electrophoretic band of PCR product of gyrB gene.
Detailed Description
The invention is further illustrated by the following examples. These examples are intended to illustrate the invention and are not intended to limit the scope of the invention. Experimental procedures without specific conditions noted in the examples below, generally according to conditions conventional in the art or as suggested by the manufacturer; the raw materials, reagents and the like used are, unless otherwise specified, those commercially available from the conventional markets and the like. Any insubstantial changes and substitutions made by those skilled in the art based on the present invention are intended to be covered by the claims.
Example 1 Strain isolation, purification and characterization
Separation and purification of Lactococcus garvieae LGHK2
Purchasing fresh Penaeus vannamei (Penaeus vannamei) in the market, washing with clear water, peeling off shrimp shell, cutting into pieces, putting into a sterilized conical flask, adding a proper amount of distilled water, sealing and fermenting for several days; with 2% CaCO3The MRS culture medium is used for carrying out conventional separation, purification, screening and identification on the shrimp-derived lactococcus lactis; the conventional glycerol cryopreservation of active and functional lactococcus species.
Second, preliminary identification of Lactococcus garvieae LGHK2
(1) Morphological characteristics
Lactococcus garvieae LGHK2 did not form a transparent ring on MRS agar medium containing 2% calcium carbonate, and the colony was small, milky white or pale yellow, and had circular protrusions on the surface (FIG. 1A). Gram staining was positive (+), cells were spherical or oval, often paired, and had no spores (FIG. 1B).
(2) Physiological and biochemical characteristics
Physiological and biochemical characteristics of strain LGHK2 are shown in FIG. 2 and Table 1, physiological and biochemical characteristics of strain LGHK2 are substantially similar to those of lactococcus garvieae, and strain LGHK2 was preliminarily determined to be lactococcus garvieae.
TABLE 1 results of physiological and biochemical tests of Strain LGHK2
Note: +: positive; -: and (4) negativity.
Third, molecular identification results and analysis
(1) Genomic DNA extraction results
Total DNA of Lactococcus garvieae LGHK2 was extracted using the bacterial genomic DNA rapid extraction kit, and the results are shown in FIG. 3, M: DL5000 Marker; 1: lactococcus garvieae LGHK 2.
(2) PCR amplification results
The electrophoretic band of the 16S rDNA PCR product is 1500bp, the electrophoretic band of the PCR product of the gyrB gene is about 1200bp, and the electrophoretic band can be obtained from the graph shown in figure 4 and figure 5, and the PCR amplification bands of the 16S rDNA and the gyrB are clear and bright and are consistent with the target band.
(3) Analysis of 16S rDNA sequencing results
The 16S rDNA gene fragment of strain LGHK2 was 1464bp in size, and the strain LGHK2 was aligned with lactococcus garvieae (accession No.: AP017373.1) for up to 99% using the BLAST software at the NCBI accession.
GTGAGAGGCGGGTGCCTATACATGCAAGTCGAGCGATGATTGAAGATAGCTTGCTATTTTCATGAAGAGCGGCGAACGGGTGAGTAACGCGTGGGAAATCTGCCGAGTAGCGGGGGACAACGTTTGGAAACGAACGCTAATACCGCATAACAATGAGAATCGCATGATTCTTATTTGAAAGAAGCAATTGCTTCACTACTTGATGATCCCGCGTTGTATTAGCTAGTTGGTAGTGTAAAGGACTACCAAGGCGATGATACATAGCCGACCTGAGAGGGTGATCGGCCACACTGGGACTGAGACACGGCCCAGACTCCTACGGGAGGCAGCAGTAGGGAATCTTCGGCAATGGGGGCAACCCTGACCGAGCAACGCCGCGTGAGTGAAGaAAGGTTTTCgGGATCGTAAAACTCtTGTTGTTAGAGAAGAACGTTAAGTAGAgGTGGAAAATTACTTAAGTGACGGTATCTAACCAGAAAGGGACGGCTAACTACGTGCCAGCAGCCGCGGTAATACGTAGGTCCCAAGCGTTGTCCGGATTTATTGGGCGTAAAGCGAGCGCAGGTGGTTTCTTAAGTCTGATGTAAAAGGCAGTGGCTCAACCATTGTGTGCATTGGAAACTGGGAGACTTGAGTGCAGGAGAGGAGAGTGGAATTCCATGTGTAGCGGTGAAATGCGTAGATATATGGAGGAACACCGGAGGCGAAAGCGGCTCTCTGGCCTGTAACTGACACTGAGGCTCGAAAGCGTGGGGAGCAAACAGGATTAGATACCCTGGTAGTCCACGCCGTAAACGATGAGTGCTAGCTGTAGGGAGCTATAAGTTCTCTGTAGCGCAGCTAACGCATTAAGCACTCCGCCTGGGGAGTACGACCGCAAGGTTGAAACTCAAAGGAATTGACGGGGGCCCGCACAAGCGGTGGAGCATGTGGTTTAATTCGAAGCAACGCGAAGAACCTTACCaAGGTCTTGACATACTCGTGCTATCCTTAGAGATAAgGGAGTTCCTTCGGGACACGGGATACAGGTGGTGCATGGtTTGTCGTCAGCTCGTGTCGTGAGAWKKTGGGtTTAAGTCCCGCAACGAGCGCAACCCTTATTACYWRKTKSCATCATTAAGTTGGGCACTCTAGTGAGACTGCCGGTGATAAACCGGAGGAAGGTGGGGATGACGTCAAATCATCATGCCCCTTATGACCTGGGCTACACACGTGCTACAATGGATGGTACAACGAGTCGCCAACCCGCGAGGGTGCGCTAATCTCTTAAAACCATTCTCAGTTCGGATTGCAGGCTGCAACTCGCCTGCATGAAGTCGGAATCGCTAGTAATCGCGGATCAGCACGCCGCGGTGAATACGTTCCCGGGCCTTGTACACACCGCCCGTCACACCACGGAAGTTGGGAGTACCCAAAGTAGGTTGCCTAACCGCAAGGAGGGCGCTCCTAAGTAGACCGATTGTG。
(4) Analysis of sequencing results for gyrB
The size of the gyrB gene fragment of strain LGHK2 was 607bp, and it was obtained by similarity comparison using BLAST software registered at NCBI, and the identity of strain LGHK2 with lactococcus garvieae (accession number: LC377166.1) was as high as 98%, as follows.
GCTATATGTTTCTGGTGGTTTACACGGTGTTGGTTCATCCGTCGTGAATGCTCTCTCTACACAGTTAGATGTTACCGTACATAAAGATGGTCAAAAATATTATCAAGAATATCATCGTGGTGTTGTTGTCGAAGATTTGGCTATTGTCGGTGAAACAGAGAAACGTGGTACGGTCGTTCACTTTACACCAGACCCAACGATTTTCACTGAGACACAAATCTTTGACTATGATAAGTTGGTGACACGTGTCCGTGAATTAGCTTTCTTAAATCGTGGCTTACGTATCTCTATTACCGACAAACGTGAAGGGCAAGAAGACAATCATGCTATGTTCCACTATGAAGGTGGGATTCAATCTTATGTTTCCTTCATTAATGAAAATAAGGAAGTTATTTTTGACACACCCATCTATACAGAAGGTGAATTAAATGGTATTACTGTAGAAGTTGCTATGCAATATACAGGGACTTACCATTCGACAATCATGTCTTTTGCAAATAATATTAATACACACGAAGGTGGTACGCATGAACAAGGTTTCCGTACTGCTTTAACGCGTGCGATAAACAACTATGCAAAAGCTCAAAAGCTTCTTAAAGATAATGAA。
The strain LGHK2 can be determined to be accurately identified as lactococcus garvieae (Lactocousgarvieae) by combining the morphological characteristics of the strain, the physiological and biochemical test results and the 16S rDNA and gyrB gene sequence analysis. The Lactococcus garvieae belongs to the phylum Firmicutes, the class Bacillales (Bacillus), the order Lactobacillales (Lactobacillus), the family Streptococcaceae (Streptococcus), and the genus Lactococcus (Lactobacillus). The cells are spherical or oval, 0.5-1.2 μm is multiplied by 0.5-1.5 μm, and are paired and short-chained in a liquid culture medium. No spore, gram positive. No movement, no capsule. Facultative anaerobic. Chemoheterotrophic, fermentative metabolism, which produces acid by fermentation of some carbohydrates, mainly L (+) -lactic acid, without producing gas. The nutrition requirement is complex, catalase negative and oxidase negative. The optimum temperature is 30 ℃ and can grow at 10 ℃ without growing at 45 ℃ in 0.5% NaCl, usually Lancefield serogroup N. Found in dairy and vegetable products.
Example 2 one of the decalcifying effects of Lactococcus garvieae LGHK2 on shrimp shells
Weighing several parts of fresh minced Penaeus vannamei Boone shell, each part 5g, respectively filling into sterilized conical flask, taking the shrimp shell as raw material, adding 30% glucose, fermenting for 36h at 37 deg.C and 60r/min at initial pH of 6.8, inoculating 4mL of 5.1 × 108The decalcification effect was measured at a solid-to-liquid ratio (the ratio of the mass of the solid phase to the mass of the liquid phase in the culture medium, for example, 10g of the mass of the solid matter (shrimp heads, sucrose, etc.) in the culture medium, and the solid-to-liquid ratio of the culture medium was 1:4 when the mass of the solvent water was 40g, and the maximum decalcification rate of Lactococcus garvieae LGHK2 was 69.47% and the pH was 3.3 when the solid-to-liquid ratio was 1: 5) per mL of Lactococcus garvieae LGHK2, as shown in Table 2.
TABLE 2 influence of solid-liquid ratio on decalcification effect of lactococcus LGHK2
EXAMPLE 3 decalcification Effect of Lactococcus lactis LGHK2 on shrimp shells two aliquots of minced fresh Penaeus vannamei Boone shells, 5g each, were individually filled in sterilized Erlenmeyer flasks and the shrimp shells were usedAdding 30% glucose into the raw material at a solid-to-liquid ratio of 1:5, initial pH of 7.0, temperature of 37 deg.C, rotation speed of 60r/min, inoculating 4mL of 5.1 × 108The effect of different fermentation times on pH and decalcification rates of shrimp shell fermentation broth was determined for each mL of Lactococcus garvieae LGHK2 and the results are shown in Table 3.
As can be seen from Table 3, the pH of the fermentation broth rapidly decreased to 3.3 within 0 to 36 hours of shrimp shell fermentation, and the decalcification rate of the fermentation broth was the greatest at 36 hours, reaching 77.75%. After that, the decalcification rate was maintained at a high level, and as a result, the optimum fermentation time of Lactococcus garvieae LGHK2 was 36 hours.
TABLE 3 influence of fermentation time on pH and decalcification rates of shrimp Shell fermentation broths
Example 4 one of the deproteinizing effects of Lactococcus garvieae LGHK2 on shrimp heads
Taking fresh heads of Penaeus vannamei Boone, removing eyes and whiskers, cutting the heads for later use, respectively putting 20g of each part into a sterilized conical flask, taking the heads as a raw material, adding 15% of glucose, wherein the solid-to-liquid ratio is 1:10, the initial pH is 6.8, the fermentation temperature is constant at 37 ℃, and the inoculation amount of Lactococcus garvieae LGHK2 is 4mL (5.1 × 10)8Pieces/ml) was added, and the mixture was fermented in a constant temperature shaking incubator at 60 rpm. Setting fermentation time as a single variable factor, keeping other fermentation conditions unchanged, fermenting in a constant-temperature shaking table at 37 ℃ for 12h, 24h, 36h, 48h and 60h, respectively measuring pH and amino acid nitrogen in the fermentation liquor, and calculating apparent hydrolysis degree. As shown in Table 4, the pH of the fermentation broth rapidly decreased to 3.3 within 36 hours, and the change was gradual, and the content of amino acid nitrogen increased significantly to 3.61 g/L.
TABLE 4 Effect of fermentation time on Deproteinization Effect of lactococcus lactis LGHK2 fermented shrimp heads
Example 5 two of the deproteinizing effects of Lactococcus garvieae LGHK2 on shrimp heads
Taking fresh heads of Penaeus vannamei Boone, removing eyes and whiskers, cutting the heads into pieces for later use, respectively putting 20g of each piece of heads into a sterilized conical flask, taking the heads as a raw material, adding 15% of glucose, wherein the solid-to-liquid ratio is 1:10, the fermentation temperature is constant at 37 ℃, and the inoculation amount of lactococcus lactis is 4mL (5.1 multiplied by 10)8Pieces/ml) and fermented for 36h in a constant temperature shaking incubator at 60 rpm. Setting initial pH as single variable factor, keeping other fermentation conditions unchanged, fermenting the initial pH respectively 5.5, 6.0, 6.5, 7.0 and 7.5 in a constant temperature shaking table for 36h, respectively measuring pH and amino acid nitrogen in the fermentation liquor, and calculating apparent hydrolysis degree. As a result, as shown in Table 5, the pH of the fermentation broth was minimized (3.3) and the amino acid nitrogen content (4.31g/L) and the apparent degree of protein hydrolysis (40.51%) were maximized at an initial pH of 7.0.
TABLE 5 Effect of initial pH on the deproteinizing Effect of lactococcus lactis LGHK2 fermented shrimp heads
Example 6 three effects of Lactococcus garvieae LGHK2 on deproteinization of shrimp heads
Taking fresh heads of Penaeus vannamei Boone, removing eyes and whiskers, cutting the heads for later use, respectively putting 20g of each part into a sterilized conical flask, taking the heads as a raw material, adding 15% glucose, wherein the initial pH is 7.0, the fermentation temperature is constant at 37 ℃, and the inoculation amount of lactococcus lactis is 4mL (5.1 × 10)8Pieces/ml) and fermented for 36h in a constant temperature shaking incubator at 60 rpm. The solid-liquid ratio is set as a single variable factor, and other fermentation conditions are unchanged, so that the influence of the solid-liquid ratio on the deproteinization effect is researched. The solid-liquid ratio was 1:1, 1:2, 1:3, 1:4, and 1:5, respectively, and after 36 hours, 1mL of the solution was taken to measure the pH and the amino acid nitrogen content, respectively, and the apparent degree of hydrolysis was calculated, and the results are shown in Table 6, in which the amino acid nitrogen reached the maximum (4.33g/L) at a fermentation solution solid-liquid ratio of 1:4, and the shrimp content was the maximumThe apparent hydrolysis degree of the cephalin reaches 40.74 percent at most, the protein is fully liquefied and separated, and the converted deproteinization rate can reach 97.0 percent.
TABLE 6 influence of solid-liquid ratio on deproteinizing effect of fermented shrimp heads by lactococcus lactis LGHK2
The above-mentioned embodiments are intended to illustrate the objects, technical solutions and advantages of the present invention in further detail, and it should be understood that the above-mentioned embodiments are merely exemplary embodiments of the present invention, and are not intended to limit the scope of the present invention, and any modifications, equivalent substitutions, improvements and the like made within the spirit and principle of the present invention should be included in the scope of the present invention.
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Claims (10)
1. A method for efficiently treating shrimp wastes is characterized by comprising the following steps:
s1: crushing the shrimp waste, sterilizing, adding an MRS culture medium and water to prepare a culture medium with a solid-to-liquid ratio of 1: 1-20, and adjusting the pH to 5.5-7.5;
s2: inoculating Lactococcus garvieae LGHK2, and performing shaking fermentation for at least 36 h; the Lactococcus lactis garvieae LGHK2 is preserved in Guangdong province microbial strain preservation center in 2018, 9 and 5, with the preservation number being GDMCC No.60443 and the preservation address being No. 59 floor 5 of Michelia Tokyo 100, Guangzhou, Guangdong province.
2. The high-efficiency treatment method according to claim 1, wherein the carbon source in the MRS medium in S1 is one or more of glucose, sucrose or lactose.
3. The efficient treatment method according to claim 1, wherein the fermentation temperature in S2 is 34-40 ℃.
4. The efficient processing method according to claim 1, wherein the shrimp waste in S1 is one or both of shrimp heads and shrimp shells.
5. The efficient treatment method according to claim 4, wherein when the shrimp waste is shrimp shells, the solid-to-liquid ratio in S1 is 1: 5-20; adjusting the pH value to 6.8-7.0 in S1.
6. The efficient processing method according to claim 5, wherein the solid-to-liquid ratio in S1 is 1: 5.
7. The efficient processing method according to claim 4, wherein the inoculation amount of Lactococcus garvieae LGHK2 in S2 is 4.08X 10 when the shrimp waste is shrimp shells8Per gram.
8. The efficient treatment method according to claim 4, wherein when the shrimp waste is shrimp heads, the solid-to-liquid ratio in S1 is 1: 1-5; adjusting the pH value to 6.8-7.0 in S1.
9. The efficient processing method according to claim 8, wherein the solid-to-liquid ratio in S1 is 1: 4.
10. The efficient processing method according to claim 4, wherein the inoculation amount of Lactococcus garvieae LGHK2 in S2 is 1.02 x 10 when the shrimp waste is shrimp heads8Per gram.
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| CN103156049A (en) * | 2011-12-08 | 2013-06-19 | 上海红马饲料有限公司 | Method for extracting protein and chitin by fermenting shrimp heads and shrimp shells |
| CN102586378A (en) * | 2012-03-12 | 2012-07-18 | 中国海洋大学 | Method for extracting substances from fermented shrimp head shells |
| CN109837224A (en) * | 2018-06-06 | 2019-06-04 | 长沙学院 | Cray processing waste microbial fermentation preparation and its preparation method and application |
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