Method for producing tetrodotoxin from microorganisms by commensalism culture fermentation
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A technology of tetrodotoxin and microorganisms, applied in the field of microorganisms, can solve the problems of low yield, difficulty in large-scale industrial production, unstable genetic characteristics of toxin-producing strains, etc., achieve high yield and solve the effect of generally low yield
Inactive Publication Date: 2012-01-25
NANCHANG UNIV
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Problems solved by technology
Although some progress has been made in the production of tetrodotoxin by microbial fermentation, the yield of tetrodotoxin produced by microbial method is only 0.11-0.73MU / mL due to the unstable genetic characteristics of the toxin-producing strain and low yield in single-strain pure culture fermentation. making it difficult to implement large-scale industrial production
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Embodiment 1
[0024] Example 1: Isolation of puffer fish ovary microbial community and symbiotic fermentation culture to produce tetrodotoxin.
[0025] Step 1, isolating a single effective strain in the microbial community from puffer fish ovary, the specific operation is as follows.
[0026] 1) During the puffer spawning period in spring, fresh wild puffer fish that are in the spawning period are selected from the vicinity of the Yangtze River Basin.
[0027] 2) Fresh wild puffer fish are scrubbed with pure water, then scrubbed and disinfected with 95% alcohol, placed in a dissection tray under sterile conditions, dissected with tweezers and a scalpel, and the ovaries are taken out and placed in sterile seawater 1g of the ovary was weighed and added to a small amount of sterile seawater for grinding to obtain an ovary homogenate.
[0028] 3) After the ovary homogenate is diluted 1-10 times with sterile seawater, draw 1mL of serial dilutions and spread it on a solid plate, incubate at 26°C...
Embodiment 2
[0040] Example 2: Tetrodotoxin production by commensal fermentation of fugu ovary and liver microbial communities.
[0041] The preservation slant of each effective microbial strain isolated from puffer fish ovary and liver and identified by species was activated, and these strains were respectively.
[0042] 1 Puffer fish ovary microbial community: the toxin-producing bacteria include Serratia ( Serratia sp. ) 1 strain and Shewanella ( Shewanella sp. ) 2 strains; the non-toxin-producing symbiotic bacteria include yeast ( Saccharomyces sp. ) 1 strain, actinomycetes ( Actinomyces sp. ) 2 strains, Bacillus ( Bacillus sp. ) 2 strains, Pseudomonas ( Pseudomonas sp. ) 1 plant.
[0043] 2 Puffer fish liver microbial community: the toxin-producing bacteria include Fusobacterium ( Bacillus fusiformis ) 1 strain, Aeromonas ( Aeromonas sp. ) 1 strain, Microbacterium ( Microbacterium sp. ) 2 strains; the non-toxin-producing symbiotic bacteria include Bacillus ( Bacillu...
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Abstract
The invention provides a method for producing tetrodotoxin from microorganisms by commensalism culture fermentation. The method is characterized by comprising the following steps: preparing microorganism strains which are separated from veneniferous tissues of a wild puffer into bacterial suspensions by using sterile distilled water, and mixing the bacterial suspensions and inoculating into a same liquid culture medium to carry out seed liquid culture for 1-3 days; and inoculating 5-10% inoculation amount of seed liquid into a fermentation culture medium to carry out fermentation culture for 2-7 days, wherein the liquid culture medium comprises the following components by weight percent: 0.1-0.5% of yeast extract, 0.1-0.5% of beef extract, 0.1-0.5% of peptone, 0.5-2% of glucose, 0-1% of sugar and the balance of seawater, and the pH value is 6.5-7.2; and the seed liquid is subjected to fermentation culture under the conditions that the temperature is 24-30 DEG C and the rotation speed is 150-200 rpm (revolutions per minutes). According to the method provided by the invention, a micro-ecological environment in which the microorganisms producing toxicity in the puffer is located can be well simulated, thereby being beneficial to the adaptability of the microorganisms producing toxicity to culture in vitro, finally being beneficial to growth and toxicity production of the microorganisms producing toxicity and solving the bottleneck that at present, the yield of the tetrodotoxin produced by microbial fermentation is low in general.
Description
technical field [0001] The invention belongs to the technical field of microorganisms, and in particular relates to a method for isolating symbiotic microorganisms from various toxic tissues of wild puffer fish and fermenting them. technical background [0002] Tetrodotoxin (TIX) is a highly toxic small molecule non-protein neurotoxin, because it can bind highly specifically to sodium ion channel receptors in nerve cell membranes, block sodium ion channels, and thereby inhibit nerve impulses conduction. TTX has an excellent nerve analgesic effect, its anesthetic effect is more than 1000 times stronger than that of general anesthetics, and it has the advantage of non-addiction, it can replace chemical drugs and Chinese herbal medicines for drug detoxification. At the same time, it also has the effects of sedation, anti-arrhythmia, prevention of renal function, energy failure and lowering blood pressure. Therefore, it has great medicinal value. Because tetrodotoxin has a st...
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