CN113040075B - Method for culturing seawater rotifer by shrimp head fermentation liquor - Google Patents

Method for culturing seawater rotifer by shrimp head fermentation liquor Download PDF

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CN113040075B
CN113040075B CN202110263172.8A CN202110263172A CN113040075B CN 113040075 B CN113040075 B CN 113040075B CN 202110263172 A CN202110263172 A CN 202110263172A CN 113040075 B CN113040075 B CN 113040075B
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rotifers
bacillus
fermentation liquor
seawater
culturing
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CN113040075A (en
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汤保贵
张静
韩满强
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Guangdong Ocean University
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Guangdong Ocean University
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    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01K—ANIMAL HUSBANDRY; CARE OF BIRDS, FISHES, INSECTS; FISHING; REARING OR BREEDING ANIMALS, NOT OTHERWISE PROVIDED FOR; NEW BREEDS OF ANIMALS
    • A01K61/00—Culture of aquatic animals
    • A01K61/40—Culture of aquatic animals of annelids, e.g. lugworms or Eunice
    • 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
    • A23K10/16—Addition of microorganisms or extracts thereof, e.g. single-cell proteins, to feeding-stuff compositions
    • A23K10/18—Addition of microorganisms or extracts thereof, e.g. single-cell proteins, to feeding-stuff compositions of live microorganisms
    • 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
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23K—FODDER
    • A23K20/00—Accessory food factors for animal feeding-stuffs
    • A23K20/10—Organic substances
    • A23K20/163—Sugars; Polysaccharides
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23K—FODDER
    • A23K20/00—Accessory food factors for animal feeding-stuffs
    • A23K20/20—Inorganic substances, e.g. oligoelements
    • A23K20/22—Compounds of alkali metals
    • A—HUMAN NECESSITIES
    • A23—FOODS OR FOODSTUFFS; TREATMENT THEREOF, NOT COVERED BY OTHER CLASSES
    • A23K—FODDER
    • A23K50/00—Feeding-stuffs specially adapted for particular animals
    • A23K50/80—Feeding-stuffs specially adapted for particular animals for aquatic animals, e.g. fish, crustaceans or molluscs

Abstract

The invention provides a method for culturing seawater rotifers by shrimp head fermentation liquor, which is characterized in that shrimp heads are fermented and are cultured with lactic acid bacteria and bacillus to obtain fermentation liquor serving as bait of the seawater rotifers, so that the culture density and the yield of the rotifers are improved, the nutritional value of the rotifers is enhanced, and the requirements of larvae in fish and crustacean seedling culture on the seawater rotifers are met. The fermentation liquor provided by the invention has mild preparation conditions, does not need large-scale equipment, can be used for trial production of shrimp heads and shrimp shells in various places, and has strong operability and low production cost.

Description

Method for culturing seawater rotifer by shrimp head fermentation liquor
Technical Field
The invention belongs to the technical field of aquaculture, and particularly relates to a method for culturing seawater rotifers by using shrimp head fermentation liquor.
Background
The Brachionus plicata is a typical representative of inland brine rotifers, is smiling in body shape, is generally 233.1-277.6um long and 170.3-201.7um wide, is a high-quality initial live bait widely applied to aquatic animal larvae, and is widely applied to seawater larvae and juvenile fish cultivation, shrimp and crab seedling production and cultivation of rare ornamental aquatic animals.
Currently, in the mass-production culture of rotifers, yeast is mainly used as a suitable feed, and the rotifers cultured with yeast as a feed lack ω 3-series polyunsaturated fatty acids (ω 3-PUFA), particularly docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA), in their bodies. The results of research on aquatic animal nutrition have shown that: omega 3-PUFA is necessary nutrient for aquatic animal larva and is a main source of energy required by larva, and the composition and content of the omega 3-PUFA directly influence the normal growth and survival of the larva. Especially, DHA and EPA in omega 3-PUFA have important functions of maintaining normal physiological functions and neural development of cell membranes of the larvae and the fish and are essential fatty acids for maintaining normal growth and development of marine fishes. In addition, the rotifer can also be reinforced by nutrition reinforcing methods such as microalgae, grease type reinforcing agent and microcapsule reinforcing agent. Microalgae enrichment is the most traditional rotifer nutrition enrichment mode, but microalgae production is easily affected by factors such as culture environment conditions, culture technology and the like, and some algae such as Chlorella vulgaris (Chlorella vulgaris) are lack of DHA, so that microalgae enrichment has more defects. Both the oil-fat type reinforcer (such as refined fish oil, emulsified fish oil, soybean oil and the like) and the microcapsule reinforcer (such as 50DE and the like) can play a role in nutrition reinforcement, but the survival rate of rotifers is reduced because the oil-fat type reinforcer is easy to oxidize and pollute water, and the microencapsulation treatment process is complex and high in cost, so that the products are limited in practical application due to the reasons.
At present, the seed sources of rotifers mainly come from wild rotifers, the wild rotifers are obtained by river mouth and artificial pond fermentation cultivation, rough culture, the quality and the supply of the wild rotifers are unstable, and the rotifers cultivated by artificial outdoor fermentation are obtained by organic matter fermentation rich water (such as pig manure, chicken manure, chemical fertilizer and the like), various plankton are usually generated in the cultivation mode, the population and the number of the species are difficult to control, the yield in actual production is unstable, and the rotifers have a large number of pathogenic organisms such as parasites and bacteria, so that the outbreak of larval fish diseases and the spread of the diseases are easily caused. And outdoor weather influences are large, and if rain falls on a continuous cloudy day, the yield is reduced. Moreover, the addition of exogenous leavening often introduces harmful flora, pathogens and the like, and greatly increases the risk of breeding the marine fish. The seed source of the rotifer after the incubation contains a large amount of impurities and bacteria, which affects the culture of the rotifer and the survival rate of aquatic seedlings.
In view of the above background, it is still an urgent research topic to provide a culture method of rotifers with high rotifer survival rate and good nutrition enhancement. Through a large number of searches, no relevant report of applying shrimp head fermentation liquor to rotifer culture exists at present.
Disclosure of Invention
One of the purposes of the invention is to provide a method for culturing seawater rotifers by shrimp head fermentation liquor, and the shrimp head fermentation liquor, lactic acid bacteria and bacillus are cultured to obtain the fermentation liquor which is used as bait of the seawater rotifers, so that the culture density and yield of the rotifers are improved, the nutritional value of the rotifers is enhanced, and the requirements of larvae in fish and crustacean seedling culture on the seawater rotifers are met.
In order to achieve the purpose, the invention adopts the technical scheme that:
a preparation method of fermentation liquor for culturing seawater rotifers comprises the following steps:
(1) crushing shrimp heads, adding brown sugar, salt and water, uniformly stirring, adding citric acid to adjust the pH value, and preparing a mixed solution A;
(2) sterilizing the mixed solution A at high temperature;
(3) cooling the mixture at room temperature, adding lactobacillus and bacillus, and culturing in a shaking table;
(4) standing for 7-16 days, measuring pH value, and measuring colony count of lactobacillus and Bacillus;
(5) and filtering the fermentation liquor by using 60-mesh bolting silk to obtain the fermentation liquor.
In a preferred embodiment of the invention, the shrimp heads in the step (1) are prawn heads, and the weight ratio of the prawn heads to the brown sugar to the salt to the water is 2-10:10-20:0.5-2: 50-100.
In a preferred embodiment of the invention, the weight ratio of the prawn heads, the brown sugar, the salt and the water in the step (1) is 7.5:17.5:1.5:62.5, and all indexes are balanced in the proportioning, so that the optimal effect can be achieved when the prawn heads, the brown sugar, the salt and the water are used as a cultivation formula of the seawater rotifers.
In a preferred embodiment of the present invention, the pH value in the step (1) is 4-6.
In a preferred embodiment of the present invention, the pH value in step (1) is 5.8, and at the pH value, each index of the formula is nutritionally balanced.
In a preferred embodiment of the present invention, the temperature in the step (2) is 100 ℃ to 150 ℃, preferably 121 ℃.
In a preferred embodiment of the present invention, the time for high temperature sterilization in step (2) is 1 to 5 hours, preferably 2 hours.
In a preferred embodiment of the present invention, in the step (3), the temperature is cooled to 37 ℃, and the bacillus is selected from bacillus subtilis, bacillus licheniformis and bacillus pumilus, and the bacillus can be obtained by commercial means.
In general, lactic acid bacteria can inhibit the growth and reproduction of harmful bacteria by secreting inhibitory substances, competing for customized sites of the host or by the principle of competitive exclusion; meanwhile, organic substances in the aquaculture water body can be directly or indirectly decomposed and utilized, the content of ammonia nitrogen and nitrite is reduced, the breeding of harmful bacteria is inhibited, and the water body environment is improved; the bacillus is an aerobic gram-positive bacterium capable of forming spores, has the characteristics of acid resistance, salt resistance, high temperature resistance, quick reactivation, capability of secreting various enzymes capable of degrading organic matters and the like, and can also directly utilize nitrate and nitrite in water to decompose organic matters at the bottom of the pool, reduce the eutrophication degree of the culture water and optimize the water environment.
In a preferred embodiment of the invention, the weight ratio of the addition amount of the lactic acid bacteria and the bacillus to the brown sugar is 1: 300-500.
In a preferred embodiment of the invention, the lactic acid bacteria are 100 hundred million/g and the bacillus is 1000 hundred million/g.
In a preferred embodiment of the present invention, the cultivation time in the step (4) is preferably 14 days, the pH value is 3-5, and the best effect is achieved when the pH value is 3.7.
In a preferred embodiment of the present invention, the total number of colonies in the step (4) is 5X 109The effect is best when cfu/ml is used.
The invention also aims to provide a method for culturing seawater rotifers, which comprises the step of mixing the fermentation liquor and bait algae for use, and comprises the following steps: the fermentation liquor and bait algae are mixed and put into a seawater rotifer culture pond.
In a preferred embodiment of the present invention, the bait algae include, but are not limited to, one or more of chlorella, Platymonas, Nitzschia closterium, Nannochloropsis sp.
In a preferred embodiment of the invention, the nutrient enrichment effect on rotifers is optimal when the fermentation broth is used in combination with chlorella.
The invention solves the defects in the background technology, and has the following beneficial effects:
(1) the invention takes the prawn heads as the raw material, greatly reduces the production cost, increases the utilization way of the prawn head waste and changes waste into valuable.
(2) The fermentation liquor can obviously improve the survival rate and the reproduction rate of the seawater rotifer, and can also achieve the aim of strengthening the nutrition of the seawater rotifer.
(3) The fermentation liquor provided by the invention has mild preparation conditions, does not need large-scale equipment, can be used for trial production of shrimp heads and shrimp shells in various places, and has strong operability and low production cost.
Detailed Description
In order that the above objects, features and advantages of the present invention can be more clearly understood, the following detailed description of the present invention is given in conjunction with the specific embodiments, which are provided for illustration of the basic concept of the present invention, and it is to be understood that the embodiments and features of the embodiments in the present application can be combined with each other without conflict. This invention may, however, be embodied in many different forms and should not be construed as limited to the embodiments set forth herein. Rather, these embodiments are provided so that this disclosure will be thorough and complete.
First, fermentation liquid preparation example
1. Example 1
A preparation method of shrimp head fermentation liquor for culturing seawater rotifer comprises the following steps:
(1) taking 150g of shrimp heads, crushing, adding 350g of brown sugar, 30g of salt and 1200mL of water, uniformly stirring, adding citric acid to adjust the pH value to 5.8, and preparing a mixed solution A;
(2) sterilizing the mixed solution A at 121 ℃ for 2 hours;
(3) cooling the mixture to 37 ℃ in the shade at room temperature, adding 0.5g of lactic acid bacteria (100 hundred million/g) and bacillus subtilis (1000 hundred million/g) respectively, and culturing in a shaker at 90rpm and 37 ℃ for 48 hours;
(4) standing and culturing at 37 ℃ for 14 days, and determining the colony number of lactobacillus and bacillus when the pH value of the product is 3.7;
(5) the fermentation liquid was filtered through 60 mesh bolting cloth to obtain the fermentation liquid of example 1.
2. Example 2
(1) Crushing 200g of shrimp heads, adding 400g of brown sugar, 30g of salt and 1200mL of water, uniformly stirring, adding citric acid to adjust the pH value to 6, and preparing a mixed solution A;
(2) sterilizing the mixed solution A at 121 ℃ for 2 hours;
(3) cooling the mixture to 37 ℃ in the shade at room temperature, adding 0.6g of lactic acid bacteria (100 hundred million/g) and bacillus subtilis (1000 hundred million/g) respectively, and culturing in a shaker at 90rpm and 37 ℃ for 48 hours;
(4) standing and culturing at 37 ℃ for 14 days, and determining the colony number of lactobacillus and bacillus when the pH value of the product is 3.7;
(5) the fermentation liquid was filtered through 60 mesh bolting cloth to obtain the fermentation liquid of example 2.
3. Example 3
(1) Taking 150g of shrimp heads, crushing, adding 400g of brown sugar, 30g of salt and 1200mL of water, uniformly stirring, adding citric acid to adjust the pH value to 6, and preparing a mixed solution A;
(2) sterilizing the mixed solution A at 121 ℃ for 2 hours;
(3) cooling the mixture to 37 ℃ in the shade at room temperature, adding 0.6g of lactic acid bacteria (100 hundred million/g) and bacillus subtilis (1000 hundred million/g) respectively, and culturing in a shaker at 90rpm and 37 ℃ for 48 hours;
(4) standing and culturing at 37 ℃ for 14 days, and determining the colony number of lactobacillus and bacillus when the pH value of the product is 3.7;
(5) the fermentation liquid was filtered through 60 mesh bolting cloth to obtain the fermentation liquid of example 3.
4. Comparative example 1
(1) 350g of brown sugar, 30g of salt and 1200mL of water are uniformly stirred, and citric acid is added to adjust the pH value to 5.8, so as to prepare a mixed solution A;
(2) sterilizing the mixed solution A at 121 ℃ for 2 hours;
(3) cooling the mixture to 37 ℃ in the shade at room temperature, adding 0.5g of lactic acid bacteria (100 hundred million/g) and bacillus subtilis (1000 hundred million/g) respectively, and culturing in a shaker at 90rpm and 37 ℃ for 48 hours;
(4) standing and culturing at 37 ℃ for 14 days, and determining the colony number of lactobacillus and bacillus when the pH value of the product is 3.7;
(5) the fermentation broth was filtered through 60 mesh bolting cloth to obtain the fermentation broth of comparative example 1.
5. Comparative example 2
A preparation method of shrimp head fermentation liquor for culturing seawater rotifer comprises the following steps:
(1) taking 150g of shrimp heads, crushing, adding 350g of brown sugar, 30g of salt and 1200mL of water, uniformly stirring, adding citric acid to adjust the pH value to 5.8, and preparing a mixed solution A;
(2) sterilizing the mixed solution A at 121 ℃ for 2 hours;
(3) cooling the mixture to 37 deg.C in shade at room temperature, adding 0.5g each of lactobacillus (100 hundred million/g) and yeast (50 hundred million/g), and culturing in a shaker at 90rpm and 37 deg.C for 48 hr;
(4) standing and culturing at 37 ℃ for 14 days, and determining the colony number of lactobacillus and bacillus when the pH value of the product is 3.7;
(5) the fermentation broth was filtered through 60 mesh bolting cloth to obtain the fermentation broth of comparative example 2.
Second, application example
1. Effect of different fermentation broths on Density of Rotifera Sesamatica
The fermentation liquid obtained in examples 1-3 and comparative examples 1 and 2 of the application is fed into different cement ponds, the actual usage amount of the fermentation liquid is converted into 1000 mL/mu, 12 hours of the fermentation liquid is put into the prepared water containers, the water solution treated by the fermentation liquid is put into the water containers, the flushed rotifer brachytaliphalus raw materials are put into the water containers in equal amount, the rotifer number under the equal amount of water solution is counted under a microscope after 24 hours of parallel dilution every 24 hours, and the test results are shown in Table 1.
TABLE 1 number of rotifers cultured with different treatments
Treatment of 24h 48h
Example 1 567 736
Example 2 554 698
Example 3 537 695
Comparative example 1 521 573
Comparative example 2 546 627
As can be seen from Table 1, the fermentation broth treated aqueous solutions of examples 1-3 cultured rotifers, compared to comparative examples 1, 2, and examples 1, 2 were significantly better than each of the other treatments in comparison of the number of rotifers at 24 hours within the sampling period; after 48h of culture, examples 1-3 were all better than comparative examples 1, 2; in particular, in example 1, the number of rotifers was significantly better than that of each of the other treatments during the sampling period of 48 hours; the reason for this analysis may be that the use of the respective raw materials in the preparation of the fermentation broth reaches an optimum level, especially for rotifers, their nutrition and the presence of microorganisms in the fermentation broth, each property being optimized.
2. Effect of fermentation broth on rotifer fortification
The fermentation liquor, chlorella and Platymonas as described in the most preferred embodiment 1 are combined to determine the influence of the fermentation liquor on the strengthening of the rotifers, the chlorella and the Platymonas are respectively used as a reference, 3 times of repetition are set for each group, 0h, 4h, 8h, 12h, 16h and 20h 6 time gradients are provided, the strengthening process keeps sufficient sunlight, oxygen is continuously added in the test process, a sample is placed into an EP tube which is well sterilized by ultraviolet irradiation after being sampled, the EP tube is placed in a refrigerator at the temperature of-20 ℃ for storage and to be determined, the content of fatty acid is determined by a gas phase spectrometer method, and the test results are shown in Table 2.
Content of rotifer fatty acid after strengthening at 220 h in table
Treatment of EPA content (mg/10g) DHA content (mg/10g)
Example 1 fermentation broth + Chlorella 14.7 11.6
Chlorella vulgaris 12.6 9.4
Content of rotifer fatty acid after strengthening at 220 h in table
Treatment of EPA content (mg/10g) DHA content (mg/10g)
Example 1 fermentation broth + Platymonas 13.7 5.6
Platymonas subcordiformis (Fr.) Kuntze 13.5 5.1
As can be seen from the data in tables 1 and 2, the shrimp head fermentation liquid in example 1 is combined with chlorella to strengthen the nutrition of rotifer, and at 20 hours, the content of EPA is increased by 16.7% and the content of DHA is increased by 23.4%, which indicates that the shrimp head fermentation liquid can strengthen the nutrition of specific chlorella after being combined with the specific chlorella, and the mechanism of strengthening the nutrition after being combined still needs to be further researched. After the shrimp head fermentation liquid in example 1 is combined with the Platymonas mellifera, the content of EPA is increased by 1.5% and the content of DHA is increased by 9.8% at 20h, and the strengthening effect after the combination is general compared with that of the bait algae only.
The above examples only show some embodiments of the present invention, and the description thereof is more specific and detailed, but not construed as limiting the scope of the invention. It should be noted that, for a person skilled in the art, several variations and modifications can be made without departing from the inventive concept, which falls within the scope of the present invention. Therefore, the protection scope of the present patent shall be subject to the appended claims.

Claims (6)

1. A preparation method of fermentation liquor for culturing seawater rotifers is characterized by comprising the following steps:
(1) crushing shrimp heads, adding brown sugar, salt and water, uniformly stirring, adding citric acid to adjust the pH value to be 4-6, and preparing a mixed solution A;
(2) sterilizing the mixed solution A at high temperature of 100-150 ℃ for 1-5 hours;
(3) cooling the mixed solution after high-temperature sterilization in the shade at room temperature, adding lactic acid bacteria and bacillus, and culturing in a shaking table, wherein the weight ratio of the added lactic acid bacteria and bacillus to the brown sugar is 1: 300-500;
(4) standing for 7-16 days, measuring pH value, and measuring colony count of lactobacillus and Bacillus;
(5) filtering the fermentation liquor by using 60-mesh bolting silk to obtain the fermentation liquor;
wherein the weight ratio of the shrimp heads to the brown sugar to the salt to the water is 2-10:10-20:0.5-2: 50-100.
2. The method of claim 1, wherein the weight ratio of the shrimp heads, the brown sugar, the salt and the water is 7.5:17.5:1.5: 62.5.
3. The method according to claim 1, wherein the pH of the fermentation broth used in step (1) is 5.8.
4. The method of claim 1, wherein the elevated temperature is 121 ℃ and the sterilization time is 2 hours.
5. The method of claim 1, wherein the bacillus is selected from the group consisting of bacillus subtilis, bacillus licheniformis and bacillus pumilus.
6. A method for cultivating seawater rotifers, characterized in that the fermentation liquor according to any one of claims 1 to 5 is used and is fed into a seawater rotifer culture pond.
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