CN105087410A - Novel aquaculture probiotics, and production method and use thereof - Google Patents
Novel aquaculture probiotics, and production method and use thereof Download PDFInfo
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- 238000009360 aquaculture Methods 0.000 title claims abstract description 27
- 244000144974 aquaculture Species 0.000 title claims abstract description 26
- 239000006041 probiotic Substances 0.000 title claims abstract description 21
- 235000018291 probiotics Nutrition 0.000 title claims abstract description 21
- 238000004519 manufacturing process Methods 0.000 title claims abstract description 9
- 230000001580 bacterial effect Effects 0.000 claims abstract description 14
- 241000894006 Bacteria Species 0.000 claims abstract description 12
- 244000063299 Bacillus subtilis Species 0.000 claims abstract description 11
- 235000014469 Bacillus subtilis Nutrition 0.000 claims abstract description 8
- 238000000855 fermentation Methods 0.000 claims abstract description 5
- 230000004151 fermentation Effects 0.000 claims abstract description 5
- 235000001855 Portulaca oleracea Nutrition 0.000 claims abstract description 4
- 239000001963 growth medium Substances 0.000 claims abstract description 3
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- 239000003795 chemical substances by application Substances 0.000 claims description 12
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Abstract
The invention relates to novel aquaculture probiotics, and a production method and use thereof, and belongs to the technical field of feed additives, aquaculture and biology. Bacterial strains of the novel aquaculture probiotics are bacillus subtilis SZX11 bacterial strains, and are preserved in a general microbiology center of a microbial preservation management committee in China, and the preservation number is CGMCC No. 8188. The novel aquaculture probiotics is a culture medium compounded by inorganic salts, vitamin, microelement, and extracts of lycium barbarm, purslane and euphorbia humifusa; SZX11 strains are inoculated on the probiotics to perform oxidation fermentation; and the probiotics with viable content greater than or equal to 2x109CFU/mL is prepared, and contains abundant active polysaccharide, abundant living bacteria and metabolism product full components. Moreover, when mixed into fodder for penaeus monodon, the novel aquaculture probiotics has a good prevention effect on intestinal diseases and immunity dysfunction of the penaeus monodon. Aquaculture survival rate and yield can be improved obviously, and the novel aquaculture probiotics has a wide application prospect.
Description
Technical field
The present invention relates to a kind of compound bio extract through subtilis (
bacillussubtilis) SZX11 strain fermentation makes the production method of active bacteria formulation and the application in prawn culturing thereof, belongs to fodder additives, aquaculture and biological technical field.
Background technology
The Penaeus vannamei of China in 2005 is light, the total cultured area of seawater reaches 1,500,000 mu, just reaches 1,400,000 mu to 2010 areas being only freshwater aquiculture Penaeus vannamei; Add nearly 3,000,000 mu of sea farming, output 90-100 ten thousand tons.But since 2011, the success ratio of culture of Penaeus vannamei is more and more lower, and disease mortality ratio is more and more higher, the row pool, most area rate more than 50%; To the first half of the year in 2011 in Hainan, the output in Guangdong and these provinces, Guangxi have dropped more than 80% than the same period, brings massive losses to white shrimp aquaculture.Its major cause is white shrimp leukodermia, gi tract disease and liver Pancreatic Necrosis syndromes spread, and makes this industry encounter unprecedented difficulty.
Subtilis (
bacillussubtilis) be the rod-shaped bacterium of a class aerobic, the degeneration-resistant spore of interior life, size (0.8 ~ 1.2) μm × (1.5 ~ 4.0) μm, Gram-positive, can produce pod membrane, raw or near middle raw in gemma, bacterium colony is coarse, energy liquefy gelatin, peptonized milk, reduction nitrate, hydrolyzed starch, for typical aerobic bacteria is extensively present in soil, lake, ocean and vegeto-animal body surface, self is not pathogenic.Along with deepening continuously of studying subtilis, find that it has widespread use and is worth in industry, agricultural, medical and health, health care of food, aquaculture etc.Such as: in Chinese invention patent (application number: 201110116641.X), disclose a bacillus subtilis, deposit number is CGMCCNo.4628, there is good resistance function and prebiotic function: can pH2.0 be tolerated, 1% pepsic simulated gastric fluid, the artificial cholate of 0.3% can be tolerated, the pelleting temperature of 80 DEG C can be tolerated; Have comparatively high inhibition effect to E.coli K88, K99 and streptococcus aureus, cellulase-producing ability is comparatively strong, can degraded cellulose; This bacterial strain, for the preparation of fodder additives, can improve animal immunizing power and feed conversion rate.
The technology utilizing Chinese herb feed additive to prevent and treat shrimp disease also has bibliographical information, such as: Chinese invention patent (application number: the Chinese herb feed additive disclosing a kind of prawn feed 201010109404.6), its composition comprises: Herba Andrographis 1-9 part, exceedingly high rattan 1-9 part, Radix Gentianae 2-8 part, Motherwort Herb 3-20 part, Herba Houttuyniae 1-9 part; This Chinese herb feed additive not only has adjustment prawn organism balance and physiological function, improve prawn body specificity and non-specific immunity, raising is digested and assimilated, the effect of growth promoting effects, and be rich in prawn and grow required mineral substance and a small amount of carbohydrate and protein, and can not resistance be produced, drug residue free, the healthy of the consumer group and environmental safety can not be affected.
Employing Chinese herbal medicine extract is main medium raw material by the present invention, with subtilis SZX11 bacterial strain for probiotics fermention bacterial classification, prepares the novel fodder additive of culture of Penaeus vannamei.
Summary of the invention
The object of this invention is to provide a kind of novel aquaculture probiotic agent and production thereof and using method.
Bacterial strain used in the present invention be by contriver from the strain out of nature separation screening have animal prebiotic effect subtilis (
bacillussubtilis) SZX11 bacterial strain, this bacterial strain is deposited in China Committee for Culture Collection of Microorganisms's common micro-organisms center on September 16th, 2013, deposit number is: CGMCCNo.8188, depositary institution address: No. 3, Yard 1, BeiChen xi Road, Chaoyang District, Beijing City, Institute of Microorganism, Academia Sinica, postcode: 100101.
The production method of Aquaculture probiotic agent of the present invention is as follows:
(1) preparation of liquid spawn: bacterium culture medium compound method (for 1L) used: potassium primary phosphate 1g, dipotassium hydrogen phosphate 0.5g, calcium chloride 50mg, ammonium sulfate 0.5g, magnesium sulfate 300mg, manganous sulfate 4mg, copper sulfate 2mg, Sodium orthomolybdate 0.6mg, para-amino benzoic acid 0.25mg, the each 0.2mg of vitamin H, VITMAIN B1, vitamin B6, niacinamide, glucose 6.2g, distilled water 1 liter; Adjust pH to 6.8-7.8 with sodium hydroxide solution, be dispensed in 500mL triangular flask, every bottle of 200mL substratum, in 121 DEG C of sterilizing 20min; The substratum of sterilizing is cooled to 30-35 DEG C, at the bacterial classification that aseptic operating platform inoculation inclined-plane is preserved, is placed in constant temperature oscillator, constant temperature 30 DEG C-35 DEG C; Rotating speed 150r/min, lucifuge concussion is cultivated 30h and is made liquid spawn, species Bacillus subtilis content 4 × 10
9cFU/mL.
(2) subtilis compound Chinese medicine used nutrient solution preparation method (for 1000L): choose without the wolfberry fruit rotting, damage by worms, go mouldy, mushroom, purslane, each 15kg of Humifuse Euphorbia Herb high-quality dry product, render in 2000L Chinese medicine extracting tank, add water to 1100-1200L, soak 50-60 after 1h to turn/min stirs and heats, heat to 100 DEG C of stirring 1h, be cooled to 40 DEG C-50 DEG C filtrations, collect filtrate and be about 1000L.Take potassium primary phosphate 600g, dipotassium hydrogen phosphate 300g, calcium chloride 30g, ammonium sulfate 300g, magnesium sulfate 180g, glucose 1800g, lactic acid 1000mL, join filtrate stirring, add water to 1000L as during insufficient total amount 1000L, adjust pH to 6.8-7.8 with sodium hydroxide solution; 115 DEG C of sterilizing 20min.
(3) the compound Chinese medicine nutrient solution after sterilizing is cooled to the subtilis liquid spawn that 30 DEG C-35 DEG C inoculation (1) steps obtain; Inoculum size 5%-10%, in the fermentor tank of constant temperature 30 DEG C-35 DEG C, passes into the fermentation 28-30h that purifies air; Make liquid bacteria suspension, bacterium liquid bacillus subtilis bacterial content>=2 × 10
9cFU/mL, namely obtains a kind of novel aquaculture probiotic agent.Filling after quality inspection, preserve in shady and cool dry place.
The using method of Aquaculture probiotic agent of the present invention is as follows:
1, above-mentioned Aquaculture probiotic agent is added in feed by the 1%-3% of feed relative, mixes;
2,2 times are added every day; Early, respectively mix and feed once evening, 1-2 hour premix, makes microbial inoculum fully penetrate in feed in advance;
Directly render to mixing mixed uniform feed on the material platform of cultivating pool when 3, throwing something and feeding.
embodiment:
Embodiment 1: the production method of Aquaculture probiotic agent
(1) preparation of liquid spawn: take potassium primary phosphate 1g, dipotassium hydrogen phosphate 0.5g, calcium chloride 50mg, ammonium sulfate 0.5g, magnesium sulfate 300mg, manganous sulfate 4mg, copper sulfate 2mg, Sodium orthomolybdate 0.6mg, para-amino benzoic acid 0.25mg, the each 0.2mg of vitamin H, VITMAIN B1, vitamin B6, niacinamide, glucose 6.2g, distilled water 1 liter; Adjust pH to 7.5 with sodium hydroxide solution, be dispensed in 500mL triangular flask, every bottle of 200mL substratum, in 121 DEG C of sterilizing 20min; The substratum of sterilizing is cooled to 35 DEG C, at the bacterial classification that aseptic operating platform inoculation inclined-plane is preserved, is placed in constant temperature oscillator, constant temperature 35 DEG C; Rotating speed 150r/min, lucifuge concussion is cultivated 30h and is made liquid spawn, species Bacillus subtilis content 4 × 10
9cFU/mL.
2, subtilis compound Chinese medicine used nutrient solution preparation method (for 1000L): choose without the wolfberry fruit rotting, damage by worms, go mouldy, mushroom, purslane, each 15kg of Humifuse Euphorbia Herb high-quality dry product, render in 2000L Chinese medicine extracting tank, add water to 1100L, after soaking 1h, 50 turns/min stirs and heats, heat to 100 DEG C of stirring 1h, be cooled to 50 DEG C of filtrations, collect filtrate and be about 1000L.Take potassium primary phosphate 600g, dipotassium hydrogen phosphate 300g, calcium chloride 30g, ammonium sulfate 300g, magnesium sulfate 180g, glucose 1800g, lactic acid 1000mL, join filtrate stirring, add water to 1000L as during insufficient total amount 1000L, adjust pH to 7.5 with sodium hydroxide solution; 115 DEG C of sterilizing 20min.
3, the compound Chinese medicine nutrient solution after sterilizing is cooled to the subtilis liquid spawn that 35 DEG C of inoculation (1) steps obtain; Inoculum size 10%, in the fermentor tank of constant temperature 35 DEG C, passes into the fermentation 28h that purifies air; Make liquid bacteria suspension, bacterium liquid bacillus subtilis bacterial content>=2 × 10
9cFU/mL, namely obtains a kind of novel aquaculture probiotic agent.Filling after quality inspection, preserve in shady and cool dry place.
Embodiment 2: Penaeus vannamei feeds experiment in 80 days
Place: sea-gull island, Fanyu District, Guangdong Province beam builds new shrimp field, the Low-salinity soil intensive culture pool, the pool.
Experimental technique: adopt with batch shrimp seedling, throw on the same day.Select honest feed, setting experiment 1 by 1.5%, experiment 2 by 2.0%, experiment 3 mixes by 2.5% and feeds above-mentioned fodder additives, every day sooner or later mixes and feeds twice.Control group is mixed and is fed isodose warm water, collects dead shrimp counting the dirty pipe box net in every day from the row end.
The average salinity 1.5/1000 of aquaculture water; Average temperature 27 DEG C; Time: 9.20-12.10 in 2013
Embodiment 3, Penaeus vannamei feed experiment in 90 days
Place: the high-order culture pond of yellow scape prawn field, lake-light town, Mazhang District, Zhanjiang, Guangdong Province high salinity.
Experimental technique: adopt with batch positive prawn seedling, select honest feed.Setting experiment 1 by 1.5%, experiment 2 by 2.0%, experiment 3 mixes by 2.5% and feeds above-mentioned fodder additives, every day sooner or later mixes and feeds twice.Control group is mixed and is fed isodose warm water.Dead shrimp counting is collected the dirty pipe box net in every day from the row end.
The average salinity 14/1000 of aquaculture water; Average temperature 29.6 DEG C; Time: 8.16-11.17 in 2013
Embodiment 4, Penaeus vannamei feed experiment in 90 days
Place: the high-order winter canopy culture pond of salinity in white horse with a black mane small lobsters field, forest farm, East Sea Island, Zhanjiang, Guangdong Province.
Experimental technique: adopt with batch positive prawn seedling, select Guangdong Yue Hai feed, setting experiment 1 by 1.5%, experiment 2 by 2.0%, experiment 3 mixes by 2.5% and feeds above-mentioned fodder additives, every day sooner or later mixes and feeds twice.Control group is mixed and is fed isodose warm water, collects dead shrimp counting the dirty pipe box net in every day from the row end.
The average salinity 8/1000 of aquaculture water; Average temperature 24.5 DEG C; On January 20 ,-2014 years on the 19th October in 2013
。
Claims (2)
1. Aquaculture probiotic agent and a production method thereof, is characterized in that: the production method of this microbial inoculum comprises the steps:
(1) preparation of liquid spawn: bacterium culture medium compound method (for 1L) used: potassium primary phosphate 1g, dipotassium hydrogen phosphate 0.5g, calcium chloride 50mg, ammonium sulfate 0.5g, magnesium sulfate 300mg, manganous sulfate 4mg, copper sulfate 2mg, Sodium orthomolybdate 0.6mg, para-amino benzoic acid 0.25mg, the each 0.2mg of vitamin H, VITMAIN B1, vitamin B6, niacinamide, glucose 6.2g, distilled water 1 liter; Adjust pH to 6.8-7.8 with sodium hydroxide solution, be dispensed in 500mL triangular flask, every bottle of 200mL substratum, in 121 DEG C of sterilizing 20min; The substratum of sterilizing is cooled to 30-35 DEG C, at the bacterial classification that aseptic operating platform inoculation inclined-plane is preserved, is placed in constant temperature oscillator, constant temperature 30 DEG C-35 DEG C; Rotating speed 150r/min, lucifuge concussion is cultivated 30h and is made liquid spawn, species Bacillus subtilis content 4 × 10
9cFU/mL;
(2) subtilis compound Chinese medicine used nutrient solution preparation method (for 1000L): choose without the wolfberry fruit rotting, damage by worms, go mouldy, mushroom, purslane, each 15kg of Humifuse Euphorbia Herb high-quality dry product, render in 2000L Chinese medicine extracting tank, add water to 1100-1200L, soak 50-60 after 1h to turn/min stirs and heats, heat to 100 DEG C of stirring 1h, be cooled to 40 DEG C-50 DEG C filtrations, collect filtrate and be about 1000L; Take potassium primary phosphate 600g, dipotassium hydrogen phosphate 300g, calcium chloride 30g, ammonium sulfate 300g, magnesium sulfate 180g, glucose 1800g, lactic acid 1000mL, join filtrate stirring, add water to 1000L as during insufficient total amount 1000L, adjust pH to 6.8-7.8 with sodium hydroxide solution; 115 DEG C of sterilizing 20min;
(3) the compound Chinese medicine nutrient solution after sterilizing is cooled to the subtilis liquid spawn that 30 DEG C-35 DEG C inoculation (1) steps obtain; Inoculum size 5%-10%, in the fermentor tank of constant temperature 30 DEG C-35 DEG C, passes into the fermentation 28-30h that purifies air; Make liquid bacteria suspension, bacterium liquid bacillus subtilis bacterial content>=2 × 10
9cFU/mL, namely obtains a kind of novel aquaculture probiotic agent; Filling after quality inspection, preserve in shady and cool dry place.
2. the using method of Aquaculture probiotic agent according to claim 1, is characterized in that: be added in feed by Aquaculture probiotic agent according to claim 1 by the 1%-3% of feed relative, mix; Add 2 every day; Early, respectively mix and feed once evening, 1-2 hour premix, makes microbial inoculum fully penetrate in feed in advance; Directly render to mixing mixed uniform feed on the material platform of cultivating pool when throwing something and feeding.
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| CN117598398A (en) * | 2023-11-27 | 2024-02-27 | 山东省海洋科学研究院(青岛国家海洋科学研究中心) | Non-resistance compound feed additive for aquatic animals |
| CN117598398B (en) * | 2023-11-27 | 2024-05-24 | 山东省海洋科学研究院(青岛国家海洋科学研究中心) | Non-resistance compound feed additive for aquatic animals |
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