CN102533630B - Wuyiencin spore powder and production technology thereof - Google Patents
Wuyiencin spore powder and production technology thereof Download PDFInfo
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- CN102533630B CN102533630B CN 201110352110 CN201110352110A CN102533630B CN 102533630 B CN102533630 B CN 102533630B CN 201110352110 CN201110352110 CN 201110352110 CN 201110352110 A CN201110352110 A CN 201110352110A CN 102533630 B CN102533630 B CN 102533630B
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Abstract
The invention discloses wuyiencin spore powder and a production technology thereof. The production technology comprises liquid culture, solid culture, drying, crushing and separation so as to obtain the wuyiencin spore powder, wherein spore number of the wuyiencin spore powder is greater than or equal to 50.0*10<8>, the water content is less than or equal to 8%, the pH value is 4.0-7.0, and the resolution ratio of the effective components is less than or equal to 5%. In the invention, through the expansion culture of liquid phase before solid phase, the spore powder content of the obtained wuyiencin spore powder is greater than 50*10<8>/g, thus the stress resistance of the spores is strong, thereby being favorable for storage and transportation; the wuyiencin spore powder can be used in various modes by directly spraying powder or spraying after being prepared into a liquid preparation, thereby being convenient to use; and the wuyiencin spore powder is particularly suitable for preventing diseases of woods and fruit trees, and has an obvious prevention effect.
Description
Technical field
The present invention relates to a kind of Wuyiencin, specifically a kind of Wuyiencin spore powder and production technique thereof, utilize the Wuyiencin spore to be applied to the prevention and control of plant diseases, pest control of plant as biological pesticide, belongs to field of biological pesticide.
Background technology
Traditional sterilant major part is chemical pesticide, and wherein some biological pesticide is to utilize submerged fermentation, and the metabolic substd that beneficial microorganism is produced has the effect (also referred to as biochemicals) of bactericidal.Traditional chemical pesticide sterilant, the main effective constituent of its sterilization be difficult to be decomposed at occurring in nature, and residual period, is long, biology is harmful to, though do not advocate, but still be main products in plant protection technology.
The most of toxicity of biochemical pesticide is lower, and Environmental compatibility is larger, is technology and the product of advocating now.And biological pesticide is also found the weakness of some problems in promoting the use of: the one, and production cost is high, and the use cost of bringing is high; The 2nd, biochemical metabolite has breakthrough development than the technology difficulty that valency improves; The 3rd, the biochemical products metabolite also can impel the disease worm to develop immunity to drugs in preventing and treating the process of crop pest, and must strengthen working concentration, has affected actual popularization; The 4th, water aqua type biochemical pesticide principal market sprays on protection ground booth vegetable etc., fills with root, and having strengthened temperature has reduced effect, prevents and treats disease also influenced.The difference of various environment is also brought the difference of result of use.
Wuyiencin is the biological pesticide product with preventing disease production-increasing effect of Chinese Academy of Agricultural Sciences development, its derive from not streptomyces hygroscopicus Wuyi mutation (
Streptomyces anhygroscopicus var.wuyiensis) a kind of microbiotic (microorganism journal of producing, 24:401-402,1984: agricultural antibiotic BO-10 produces the evaluation of bacterium), be mainly used in preventing and treating the farm crop fungus disease, the multiple fungal diseases such as the Powdery Mildew of plant, graw mold of tomato, leaf muld of tomato, scab of cucumber are had good prevention effect, and its main component is the nucleoside antibiotics with cytidine skeleton, to the person poultry safety, therefore, have broad application prospects in the ecological agriculture and pollution-free food production.
Summary of the invention
In order to overcome the limitation of above-mentioned biological pesticide, the invention provides a kind of Wuyiencin spore powder and production technique thereof, utilize deep fermentation and the processing method that is connected two stages of solid fermentation to reach to turn out the Wuyi bacterium spore powder biological pesticide of the high-content of 5,000,000,000/gram Wuyi bacterium spore, adopt suitability for industrialized production, be beneficial to biological pesticide and enlarge scale.
Technical scheme of the present invention is:
A kind of production technique of Wuyiencin spore powder comprises the following steps:
(1) liquid culture: the Wuyi bacterial classification is pressed the 5-10% inoculation of liquid nutrient medium quality, utilized cradle vibrate fermentation 2-3 days under 28 ℃ of constant temperature, obtain containing the mycelial liquid in Wuyi;
(2) solid culture: be that 1 ︰ 3-4 mixes with the Wuyi mycelium that obtains of fermentation in above-mentioned steps (1) liquid fermentation medium by weight with the solid fermentation substratum, the solid fermentation cultivation is 3-5 days under temperature 28-30 ℃, the condition of humidity 60-80%;
(3) drying: the cultivation material that contains abundant spore after above-mentioned steps (2) solid fermentation is carried out drying, and 45 ℃ of drying temperatures time 10-30 minute, make material moisture extremely lower than 8%;
(4) pulverize separation: the dried dry thing that contains a large amount of Wuyis bacterium spore of above-mentioned steps (3) is pulverized, then utilized 80-100 purpose screen cloth, after separating stock culture material and Wuyi mycelium, obtain Wuyi bacterium spore powder.
Before above-mentioned steps (1) inoculation, Wuyi bacterium bacterium is activated, the bacterial classification of preservation state is put into suitable activation medium, enlarged culturing is in order to obtain pure and strong Wuyi bacterial classification step by step, the 2-3 rejuvenation process in generation generally need to be arranged, allow bacterial classification adapt to gradually the culture environment of making to produce bacterial classification.
Wherein, the consisting of of described liquid fermentation medium: potato 150-200 gram, glucose 15-20 gram, potassium primary phosphate 1-2 gram, 1000 milliliters of sal epsom 0.5-1 grams, yeast extract paste 0.5 gram, peptone 0.5 gram, sodium-chlor 0.5 gram, water, pH value 6.5.
The massfraction of described solid fermentation substratum consists of: soybean cake powder 20-30%, Semen Maydis powder 5-20%, wheat bran 5-10%, potassium primary phosphate 0.08-0.2%, trimagnesium phosphate 0.2%, calcium carbonate 0.3%, water complementary.
The thickness of the solid fermentation substratum of described step (2) is 6-8 centimetre, when media surface is covered with mycelia, all turns over and makes for one time culture evenly accept up and down air and moisture in culturing process, so that Wuyi bacterium spore enlarges increment in media surface.
The drying plant of described step (3) is belt vacuum and low temperature continuous drier.
Adopt the spore quantity of the spore goods that the inventive method obtains reach 5,000,000,000/more than gram, biphasic fermentation is cultivated the Wuyi spore powder, at first guarantee the purity of Wuyi bacterial classification at fermentation, avoided interference and the Wuyi bacterium spore " growths of how much level multiples " of other miscellaneous bacteria; After changing again solid-state cultivation over to, under good gaseity, facilitate Wuyi mycelia and spore to grow up more rapidly, the characteristic that more favourable recovery is original and growing way.Due to the strong stress resistance of spore, therefore be beneficial to preservation, transportation and several different methods and use, the particularly disease control of forest, fruit tree.
The spore quantity of the spore goods that employing the inventive method obtains 〉=50.0 * 10
8, water content≤8%, pH value=4.0-7.0, effective constituent rate of decomposition≤5%, heat storage stability is qualified.
Wuyi bacterium spore powder is long volution, general 5-12 circle spiral, and spore is circular or oval, and size is even, and spore surface is barbed, and the living bacterium powder of gas is canescence or grey on the PDA substratum, and in base, the bacterium powder is yellow.
Find with test after deliberation, utilize Wuyiencin to produce bacterium as biocontrol microorganisms, can directly prevent and treat soil-borne disease of crop, such as the verticillium of control eggplant, the blight of cotton etc. has obvious effect, be better than that other agricultural chemicals have more efficiently, the characteristics of wide spectrum, low toxicity, environmental safety
To be live body secrete microbiotic to the diseases prevention principle of Wuyiencin spore powder after diseased plant blade face, root growth, ecological niche competition and parasitize the eubiosis that crop surface is kept plant growth.Do not produce resistance and residual toxicity after chemical pesticide uses.
Be improved by spore reproduction ability after the formula of adjusting substratum, it is Wuyi bacterium solid phase body fermentation efficiency 500,000,000/g spore of living that the fermentation of particularly taking to optimize adds method that the native place spreads cultivation, more than bringing up to 5,000,000,000/g.The Wuyi spore powder biological pesticide that utilizes this product to make, its diseases prevention principle are ecological niche competition, parasitic and secretion microbiotic.Because used medium formula of the present invention is different from the conventional art formula, particularly the keeping mode of solid-state cultivation is improved, and it is very competent that the Wuyi spore produces spore, and the Product industrialization cost is low.Field test proves, Wuyi bacterium spore powder can adopt and spray, dresses seed, topdresses, spreads manuer in holes etc.In its viable bacteria metabolite, effectively the antimicrobial substance component content is high after crop body surface or root soil parasitism for its viable bacteria, and to rob the mass point positive with germ.Form simultaneously and stimulate the mechanism of making the spontaneous degeneration-resistant disease and pest of deposits yields, be consistent with " plant health is advocated theory " that international academic community was advocated in recent years.
The invention has the advantages that: more than utilizing first liquid phase the Wuyi bacterium spore powder content that obtains of solid phase enlarged culturing reaching 5,000,000,000/gram again, strong stress resistance because of spore, therefore, be more conducive to preserve, transport, and using method is versatile and flexible, can adopt pulvis directly to spray, also can be made into liquid preparation and spray, easy to use; Not only raw material environmental protection simultaneously, and make front, also environmental protection when making the neutralization use, reach the requirement of low-carbon (LC) and green, have very high social benefit and environmental benefit, can be widely used in the multiple diseases control of the crops such as vegetables, fruit tree, forestry, prevention effect is remarkable.
Embodiment
Below the preferred embodiments of the present invention are described, should be appreciated that preferred embodiment described herein only is used for description and interpretation the present invention, is not intended to limit the present invention.
Embodiment 1
A kind of production technique of Wuyiencin spore powder comprises the following steps:
1, bacterium inclined-plane, the Wuyi seed after activating is 8% to be linked in liquid fermentation medium by mass percentage, keeps 28 ℃ of temperature, and the shaking table concussion is cultivated, and ferments 3 days.
Wherein, consisting of of liquid fermentation medium used: potato 180 grams, glucose 16 grams, potassium primary phosphate 1.5 grams, sal epsom 1 gram, yeast extract paste 0.5 gram, peptone 0.5 gram, sodium-chlor 0.5 gram, 1000 milliliters, water, pH value 6.5.
2, be that 1 ︰ 3 mixes by weight with cultured Wuyi bacterium mycelium in liquid fermentation medium and solid fermentation substratum, (can scatter as standard after agglomerating to hold) stirs, carry out the solid phase fermentation under 28 ℃, humidity 60% condition, under solid state fermentation conditions, the thickness of substratum is 6 centimetres, in time watering keep humidity when the material surface is covered with the bacterium powder all upset make solid-state culture material evenly accept temperature and moisture, air, enlarged culturing effect.Cultivate the gas that produced in 4 days near natural form and give birth to Wuyi bacterium spore.
Wherein, the massfraction of solid fermentation substratum consists of: soybean cake powder 20%, Semen Maydis powder 20%, wheat bran 5%, potassium primary phosphate 0.08%, trimagnesium phosphate 0.2%, calcium carbonate 0.3%, water complementary.
3, the cultivation material that contains abundant spore after above-mentioned solid fermentation is utilized PLG series continuous vacuum dryer (changzhou city development drying plant company limited, specification type1500/12 type power 11kw) carry out drying, adjust Vacuumdrier at 45 ℃ of whole fermentation materials of oven dry.
4 pulverize the solid fermented material after above-mentioned oven dry.
5 utilize mechanical vibration to separate obtains Wuyi bacterium spore powder.
Adopt the ZD6*5 vibrating screen classifier to make spore and mycelium and cultivation feed separation, obtain the Wuyi spore powder of 5,000,000,000 of content/more than gram through microscopy and biological assay.
It is as follows that Wuyi of the present invention bacterium spore powder is controlled index:
Simultaneous test: protect ground leaf muld of tomato Control experiment
Leaf muld of tomato is the Deuteromycotina disease; be multiple disease in hot and humid protection ground cultivation; adopt Wuyi bacterium spore powder biotechnological formulation (the spore powder powder that the present embodiment makes and the preparation of the actual content that obtains after filling auxiliary material diatomite mixes at 200,000,000/gram; auxiliary material diatomite can dilute spore powder; allow simultaneously spore powder be attached on stopping composition, be convenient to agricultural use convenient.) with derosal the same terms of 800 times of liquid under, foliage-spray is weekly, uses altogether effect such as following table 3 times:
Result: compared with the control, this product has antagonism, nutrient competition effect to leaf mold, Wuyi bacterium spore to leaf mold can identify, contact, penetrate, parasitic use, controlled the generation of leaf muld of tomato.
Embodiment 2
A kind of production technique of Wuyiencin spore powder comprises the following steps:
1, bacterium inclined-plane, the Wuyi seed after activating is 5% to be linked in liquid fermentation medium by mass percentage, keeps 28 ℃ of temperature, and the shaking table concussion is cultivated, and ferments 2 days.
Wherein, consisting of of liquid fermentation medium used: potato 200 grams, glucose 15 grams, potassium primary phosphate 2 grams, sal epsom 0.5 gram, yeast extract paste 0.5 gram, peptone 0.5 gram, sodium-chlor 0.5 gram, 1000 milliliters, water, pH value 6.5.
2, be that 1 ︰ 3 mixes by weight with cultured Wuyi bacterium mycelium in liquid fermentation medium and solid fermentation substratum, (can scatter as standard after agglomerating to hold) stirs, carry out the solid phase fermentation under 30 ℃, humidity 70% condition, under solid state fermentation conditions, the thickness of substratum is 8 centimetres, in time watering keep humidity when the material surface is covered with the bacterium powder all upset make solid-state culture material evenly accept temperature and moisture, air, enlarged culturing effect.Cultivate the gas that produced in 5 days near natural form and give birth to Wuyi bacterium spore.
Wherein, the massfraction of solid fermentation substratum consists of: soybean cake powder 22%, Semen Maydis powder 10%, wheat bran 10%, potassium primary phosphate 0.2%, trimagnesium phosphate 0.2%, calcium carbonate 0.3%, water complementary.
3, the solid-phase culture material with a large amount of mycelia of above-mentioned generation, spore carries out drying, adjusts Vacuumdrier at 45 ℃ of whole fermentation materials of oven dry.
4 pulverize the solid fermented material after above-mentioned oven dry.
5 utilize mechanical vibration to separate obtains Wuyi bacterium spore powder.
It is as follows that Wuyi of the present invention bacterium spore powder is controlled index:
Simultaneous test: the preventive and therapeutic effect test of cotton wilt
Method is with embodiment 1
Result: compared with the control, this product to cotton wilt have antagonism, penetrate, parasitization, controlled the generation of cotton wilt.
Embodiment 3
A kind of production technique of Wuyiencin spore powder comprises the following steps:
1, bacterium inclined-plane, the Wuyi seed after activating is 7% to be linked in liquid fermentation medium by mass percentage, keeps 28 ℃ of temperature, and the shaking table concussion is cultivated, and ferments 3 days.
Wherein, consisting of of liquid fermentation medium used: potato 200 grams, glucose 15 grams, potassium primary phosphate 1 gram, sal epsom 1 gram, yeast extract paste 0.5 gram, peptone 0.5 gram, sodium-chlor 0.5 gram, 1000 milliliters, water, pH value 7.5.
2, be that 1 ︰ 4 mixes by weight with cultured Wuyi bacterium mycelium in liquid fermentation medium and solid fermentation substratum, stir (can scatter as standard after agglomerating to hold),
30 ℃, carry out the solid phase fermentation under humidity 80% condition, under solid state fermentation conditions, the thickness of substratum is 8 centimetres, in time watering keep humidity when the material surface is covered with the bacterium powder all upset make solid-state culture material evenly accept temperature and moisture, air, enlarged culturing effect.Cultivate the gas that produced in 5 days near natural form and give birth to Wuyi bacterium spore.
Wherein, the massfraction of solid fermentation substratum consists of: soybean cake powder 30%, Semen Maydis powder 5%, wheat bran 10%, potassium primary phosphate 0.2%, trimagnesium phosphate 0.2%, calcium carbonate 0.3%, water complementary.
3, the solid-phase culture material with a large amount of mycelia of above-mentioned generation, spore carries out drying, adjusts Vacuumdrier at 45 ℃ of whole fermentation materials of oven dry.
4 pulverize the solid fermented material after above-mentioned oven dry.
5 utilize mechanical vibration to separate obtains Wuyi bacterium spore powder.
It is as follows that Wuyi of the present invention bacterium spore powder is controlled index:
Simultaneous test: the preventive and therapeutic effect test of protection Viola grypoceras A. Gray gray mold
Method is with embodiment 1
Result: compared with the control, this product to gray mold of cucumber have antagonism, penetrate, parasitization, controlled the generation of gray mold of cucumber.
Embodiment 4
A kind of production technique of Wuyiencin spore powder comprises the following steps:
1, bacterium inclined-plane, the Wuyi seed after activating is 10% to be linked in liquid fermentation medium by mass percentage, keeps 28 ℃ of temperature, and the shaking table concussion is cultivated, and ferments 2 days.
Wherein, consisting of of liquid fermentation medium used: potato 150 grams, glucose 20 grams, potassium primary phosphate 1 gram, sal epsom 1 gram, yeast extract paste 0.5 gram, peptone 0.5 gram, sodium-chlor 0.5 gram, 1000 milliliters, water, pH value 6.5.
2, be that 1 ︰ 3 mixes by weight with cultured Wuyi bacterium mycelium in liquid fermentation medium and solid fermentation substratum, (can scatter as standard after agglomerating to hold) stirs, carry out the solid phase fermentation under 30 ℃, humidity 70% condition, under solid state fermentation conditions, the thickness of substratum is 7 centimetres, in time watering keep humidity when the material surface is covered with the bacterium powder all upset make solid-state culture material evenly accept temperature and moisture, air, enlarged culturing effect.Cultivate the gas that produced in 3 days near natural form and give birth to Wuyi bacterium spore.
The solid fermentation substratum consist of soybean cake powder 20%, Semen Maydis powder 10%, wheat bran 10%, potassium primary phosphate 0.2%, trimagnesium phosphate 0.2%, calcium carbonate 0.3%, water complementary.
3, the solid-phase culture material with a large amount of mycelia of above-mentioned generation, spore carries out drying, adjusts Vacuumdrier at 45 ℃ of whole fermentation materials of oven dry.
4 pulverize the solid fermented material after above-mentioned oven dry.
5 utilize mechanical vibration to separate obtains Wuyi bacterium spore powder.
It is as follows that Wuyi of the present invention bacterium spore powder is controlled index:
Simultaneous test: the preventive and therapeutic effect test of protection Fruit of Common Pratia gray mold
Method is with embodiment 1
Result: compared with the control, this product to grey mould of egg plant have antagonism, penetrate, parasitization, controlled the generation of grey mould of egg plant.
In each embodiment of the present invention, chemical industry material used is the raw material commonly used in the production of this area, all can obtain from market, and can not exert an influence for the production result; The equipment of using in each operation all adopts conventional equipment used in current production, there is no special feature.
It should be noted that at last: the above only is the preferred embodiments of the present invention, be not limited to the present invention, although with reference to previous embodiment, the present invention is had been described in detail, for a person skilled in the art, it still can be modified to the technical scheme that aforementioned each embodiment puts down in writing, and perhaps part technical characterictic wherein is equal to replacement.Within the spirit and principles in the present invention all, any modification of doing, be equal to replacement, improvement etc., within all should being included in protection scope of the present invention.
Claims (1)
1. the production technique of a Wuyiencin spore powder, is characterized in that, comprises the following steps:
(1) liquid culture: the Wuyi bacterial classification is pressed the 5-10% inoculation of liquid nutrient medium quality, utilized cradle vibrate fermentation 2-3 days under 28 ℃ of constant temperature;
(2) solid culture: by weight for 1:3-4 mixes, the solid fermentation cultivation is 3-5 days under temperature 28-30 ℃, the condition of humidity 60-80% with the Wuyi mycelium that obtains of fermentation in above-mentioned steps (1) liquid nutrient medium and solid fermentation substratum;
(3) drying: the cultivation material after above-mentioned steps (2) solid fermentation is carried out drying, and 45 ℃ of drying temperatures time 10-30 minute, make material moisture extremely lower than 8%;
(4) pulverize separation: the dried dry thing that contains a large amount of Wuyis bacterium spore of above-mentioned steps (3) is pulverized, then utilized 80-100 purpose screen cloth, after separating stock culture material and Wuyi mycelium, obtain Wuyi bacterium spore powder;
Before above-mentioned steps (1) inoculation, the bacterial classification of Wuyi bacterium is activated;
The thickness of the solid fermentation substratum of described step (2) is 6-8 centimetre, when media surface is covered with mycelia, all turns over and makes for one time culture evenly accept up and down air and moisture in culturing process, so that Wuyi bacterium spore enlarges increment in media surface;
Consisting of of described liquid fermentation medium: potato 150-200 gram, glucose 15-20 gram, potassium primary phosphate 1-2 gram, 1000 milliliters of sal epsom 0.5-1 grams, yeast extract paste 0.5 gram, peptone 0.5 gram, sodium-chlor 0.5 gram, water, pH value 6.5;
The massfraction of described solid fermentation substratum consists of: soybean cake powder 20-30%, Semen Maydis powder 5-20%, wheat bran 5-10%, potassium primary phosphate 0.08-0.2%, trimagnesium phosphate 0.2%, calcium carbonate 0.3%, water complementary;
The drying plant of described step (3) is belt vacuum and low temperature continuous drier.
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| CN101748171B (en) * | 2010-01-14 | 2012-05-09 | 潍坊万胜生物农药有限公司 | Process method for extracting wuyiencin oligosaccharide from wuyiencin submerged fermentation liquid |
| CN101948766B (en) * | 2010-08-16 | 2012-01-25 | 潍坊万胜生物农药有限公司 | Process method for producing Wuyi biocontrol bacteria by utilizing waste residues generated by deeply fermenting Wuyiencin |
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