CN106967752A - A kind of method for preparing insecticide through microbial fermentation by raw material of Siberian cocklebur - Google Patents

A kind of method for preparing insecticide through microbial fermentation by raw material of Siberian cocklebur Download PDF

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CN106967752A
CN106967752A CN201710243501.6A CN201710243501A CN106967752A CN 106967752 A CN106967752 A CN 106967752A CN 201710243501 A CN201710243501 A CN 201710243501A CN 106967752 A CN106967752 A CN 106967752A
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fermentation
siberian cocklebur
insecticide
water
commercially available
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廖祥儒
窦欣
田乔鹏
蔡宇杰
管政兵
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Jiangnan University
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Jiangnan University
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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
    • C12P1/00—Preparation of compounds or compositions, not provided for in groups C12P3/00 - C12P39/00, by using microorganisms or enzymes
    • C12P1/04—Preparation of compounds or compositions, not provided for in groups C12P3/00 - C12P39/00, by using microorganisms or enzymes by using bacteria
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N63/00—Biocides, pest repellants or attractants, or plant growth regulators containing microorganisms, viruses, microbial fungi, animals or substances produced by, or obtained from, microorganisms, viruses, microbial fungi or animals, e.g. enzymes or fermentates
    • A01N63/10—Animals; Substances produced thereby or obtained therefrom
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N65/00—Biocides, pest repellants or attractants, or plant growth regulators containing material from algae, lichens, bryophyta, multi-cellular fungi or plants, or extracts thereof
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
    • A01N65/00—Biocides, pest repellants or attractants, or plant growth regulators containing material from algae, lichens, bryophyta, multi-cellular fungi or plants, or extracts thereof
    • A01N65/08—Magnoliopsida [dicotyledons]
    • A01N65/12—Asteraceae or Compositae [Aster or Sunflower family], e.g. daisy, pyrethrum, artichoke, lettuce, sunflower, wormwood or tarragon
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
    • C12N1/20—Bacteria; Culture media therefor
    • C12N1/205—Bacterial isolates
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12R—INDEXING SCHEME ASSOCIATED WITH SUBCLASSES C12C - C12Q, RELATING TO MICROORGANISMS
    • C12R2001/00—Microorganisms ; Processes using microorganisms
    • C12R2001/01—Bacteria or Actinomycetales ; using bacteria or Actinomycetales
    • C12R2001/07—Bacillus

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  • Virology (AREA)
  • Natural Medicines & Medicinal Plants (AREA)
  • General Chemical & Material Sciences (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Pest Control & Pesticides (AREA)
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  • Medicinal Chemistry (AREA)
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Abstract

The invention provides a kind of technology for preparing biological source insecticide.Its process characteristic is that, using Siberian cocklebur as raw material, using being stored in China typical culture collection center, deposit number is：CCTCCNO：M2013283 bacillus amyloliquefaciens (Bacillus amyloliquefaciens) H47 and commercially available bacillus pumilus and commercially available Dipel mixed fermentation.Raw material adds strain 100mL after suitably smashing, and add water 10L, 25 DEG C -30 DEG C 3 12d of fermentation, and finished product is obtained after filtering.Preparation method of the present invention and during non-pollution safety it is reliable, the fermentation time cycle is short, and process procedure is few, simple to operation, can reduce investment outlay and operating cost.Compared with traditional chemical medicament, it has green safety, the advantage such as environment-friendly, and Promoting bacteria also has preferable disinsection prophylaxis and growth-promoting functions in itself, gathers around and has broad application prospects.

Description

A kind of method for preparing insecticide through microbial fermentation by raw material of Siberian cocklebur
Technical field
The present invention relates to one plant of bacillus amyloliquefacienses (Bacillusamyloliquefaciens) H47 of solution and short and small bud Spore bacillus and bacillus thuringiensis microbiological fermentation Siberian cocklebur prepare the production method of plant insecticide, belong to microorganism and fermentation skill Art field.
Background technology
Siberian cocklebur (Xanthium sibiricum Patrin ex Widder) (former mutation) annual herb, high 20- 90cm.Root fusiform, not branch or branch.Stem upright not branch or rare branch, lower cylindrical shape, footpath 4-10mm, top has vertical Ditch, by canescence strigose.Leaf triangular shape is avette or heart, long 4-9cm, wide 5-10cm, nearly full edge, or has 3-5 substantially not shallow Split, top is sharp or blunt, base portion slightly heart-shaped or section shape, with petiole junction into equal wedge shape, there is irregular rough sawn tooth at edge, There are three bases to go out arteries and veins, lateral vein arc, go directly leaf margin, close by strigose on arteries and veins, above green, below pale asphyxia, by strigose；Petiole Long 3-11cm.The capitulum of male is spherical, footpath 4-6mm, with or without peduncle, the long round shape lanceolar of phyllary, long 1- 1.5mm, by pubescence, holder column, supporting spring falls lanceolar, is about 2mm, top point has microtriche, the male flowers for having majority, corolla Bell, there are 5 wide slivers pipe portion upper end；The long round shape of flower pesticide is linear；The capitulum ellipse of female, outer layer phyllary is small, drapes over one's shoulders pin Shape, is about 3mm, by pubescence, and internal layer phyllary is combined into cryptomere, width egg shape or ellipse, green, sometimes pistac or band Bronzing, hard in the ripe time-varying of achene, together with beak minister 12-15mm, wide 4-7mm, there are the hook-shaped thorn of the tool of sparsely grow, thorn in outside Superfine and straight, the micro- thickening of base portion or several not thickenings, long 1-1.5mm, base portion often have gland point by pubescence, or all hairless；Beak is hard Firmly, taper, upper end is slightly in sickle shaped, and long 2.5mm, normal Length discrepancy is rare to be combined into 1 beak.Achene 2, obovate.Florescence 7- August, the fruiting period 9-10 months.
Containing strumaroside (xanthostrumarin).Leaf contains xanthanol (xanthanol), isoxanthanol (jsoxanthanol), xanthumin (xanthumin) etc..Fruit contains strumaroside, fat oil, resin, alkaloid, vitamin C.It is grey Ear glucoside (Strumaroside), i.e. B- sitosterol glycoside), xanthinin (Xanthinin), (Xanthumin is xanthumin The stereoisomer of xanthinin), 8- (△ 2- isopentene groups) -5,7,3,4- kaempferols [8- (△ 2-Isopen-tenyl) - 5,7,3,4-Tetrahydroxy-flavone], and the secondary metabolite such as caffeic acid and 1,4-Dicaffeoylquinic acid, tool There is certain insecticidal effect, especially have preferable killing action to fly maggot class and black peach aphid.
The features such as plant insecticide has safe efficient, low toxicity, low-residual, meets the requirement of insect green preventing and treating, will New lover as researcher.Using suitable plant growth-promoting bacteria fermentation Siberian cocklebur, the extraction of physiological activator is not only contributed to, also The inactivation of active material is not easily caused, and Promoting bacteria also has preferable disinsection prophylaxis and growth-promoting functions in itself.Be conducive to improving plant Growth and enhancing insecticidal effect, are conducive to the effective utilization of environmental protection and Siberian cocklebur plant in itself.
The content of the invention
In view of the foregoing defects the prior art has, the present invention is raw material using Siberian cocklebur plant, is protected with June 24th, 2013 China typical culture collection center is stored in, deposit number is：CCTCCNO：M2013283 bacillus amyloliquefaciens (bacterial strain is hereinafter referred to as by (Bacillus amyloliquefaciens) H47：Bacillus amyloliquefaciens H47CCTCCM2013283) and commercially available bacillus pumilus and commercially available Dipel fermentation prepare plant insecticide, favorably In improving plant growth and enhancing insecticidal effect, be conducive to environmental protection.
The fermentation technique scheme of invention：
1) strain prepares：By bacillus amyloliquefaciens H47CCTCCM2013283 and commercially available bacillus pumilus, commercially available Soviet Union Cloud gold bacillus is inoculated in seed culture medium respectively, in 30 DEG C, 250r/min concussion and cultivates 12h；The seed culture based component is pressed G/L is calculated as：Peptone 10, yeast extract 5, NaCl 10, remaining composition is water, and 121 DEG C of autoclaving 15min adjust pH to 7.0. Seed is done in mixing by a certain percentage.
2) zymotic fluid prepares：From superior in quality Siberian cocklebur, rinsed with water, 70% -75% alcohol-pickled 3min uses water Clean, it is stand-by after being blended through pulper.
3) fermentation ullage disappears and air duct sterilization：Fermentation ullage is carried out by instructions book to disappear.
4) zymotic fluid disappears in fact：Step 2 gained Siberian cocklebur is put into fermentation tank, supplement running water to final volume is 10L, by phase Close specification and carry out real elimination bacterium.
5) inoculated and cultured：When fermentation jar temperature is down to 37 DEG C, by step 1 gained seed by volume 1% inoculum concentration, It is seeded in the fermentation tank equipped with Siberian cocklebur, is cultivated 3-12 days under the conditions of 25-30 DEG C, throughput and the holding of technological requirement is set Tank pressure, sets other each culture parameters cultures.
6) discharge：Culture is finished, and releases zymotic fluid, is filtered, and gained zymotic fluid is insecticide finished product after filtering.The present invention Beneficial effect：
A kind of method of biological source insecticide is made the invention provides fermentation Siberian cocklebur.Preparation method of the present invention and during Non-pollution safety is reliable, and the fermentation time cycle is short, and process procedure is few, simple to operation, can reduce investment outlay and operating cost.With biography System chemical agent is compared, its tool green safety, the advantage such as environment-friendly, and Promoting bacteria also has preferable disinsection prophylaxis and rush in itself Raw effect, gathers around and has broad application prospects.
Specific implementation method：
With reference to embodiment, the present invention is described further.Following examples are only several specific realities of the present invention Example is applied, but the design concept of the present invention is not limited thereto, all changes for carrying out unsubstantiality to the present invention using this design, The behavior for invading the scope of the present invention should be belonged to.
Embodiment 1
1) strain prepares：By bacillus amyloliquefaciens H47CCTCCM2013283 and commercially available bacillus pumilus, commercially available Soviet Union Cloud gold bacillus is inoculated in seed culture medium respectively, in 30 DEG C, 250r/min concussion and cultivates 12h；The seed culture based component is pressed G/L is calculated as：Peptone 10, yeast extract 5, NaCl 10, remaining composition is water, and 121 DEG C of autoclaving 15min adjust pH to 7.0. By 2:2:Seed is done in 1 mixing.
2) zymotic fluid prepares：From superior in quality Siberian cocklebur, rinsed with water, 70% -75% alcohol-pickled 3min uses water Clean, it is stand-by after being blended through pulper.
3) fermentation ullage disappears and air duct sterilization：Fermentation ullage is carried out by instructions book to disappear.
4) zymotic fluid disappears in fact：Step 2 gained Siberian cocklebur is put into fermentation tank, supplement running water to final volume is 10L, by phase Close specification and carry out real elimination bacterium.
5) inoculated and cultured：When fermentation jar temperature is down to 37 DEG C, by step 1 gained seed by volume 1% inoculum concentration, It is seeded in the fermentation tank equipped with Siberian cocklebur, is cultivated 6 days under the conditions of 25 DEG C, the throughput of technological requirement is set and tank pressure is kept, if Put other each culture parameters cultures.
6) discharge：Culture is finished, and releases zymotic fluid, is filtered, and gained zymotic fluid is insecticide finished product after filtering.
Embodiment 2
1) strain prepares：By bacillus amyloliquefaciens H47CCTCCM2013283 and commercially available bacillus pumilus, commercially available Soviet Union Cloud gold bacillus is inoculated in seed culture medium respectively, in 30 DEG C, 250r/min concussion and cultivates 12h；The seed culture based component is pressed G/L is calculated as：Peptone 10, yeast extract 5, NaCl 10, remaining composition is water, and 121 DEG C of autoclaving 15min adjust pH to 7.0. By 1:2:Seed is done in 1 mixing.
2) zymotic fluid prepares：From superior in quality Siberian cocklebur, rinsed with water, 70% -75% alcohol-pickled 3min uses water Clean, it is stand-by after being blended through pulper.
3) fermentation ullage disappears and air duct sterilization：Fermentation ullage is carried out by instructions book to disappear.
4) zymotic fluid disappears in fact：Step 2 gained Siberian cocklebur is put into fermentation tank, supplement running water to final volume is 10L, by phase Close specification and carry out real elimination bacterium.
5) inoculated and cultured：When fermentation jar temperature is down to 37 DEG C, by step 1 gained seed by volume 1% inoculum concentration, It is seeded in the fermentation tank equipped with Siberian cocklebur, is cultivated 6 days under the conditions of 27 DEG C, the throughput of technological requirement is set and tank pressure is kept, if Put other each culture parameters cultures.
6) discharge：Culture is finished, and releases zymotic fluid, is filtered, and gained zymotic fluid is insecticide finished product after filtering.
Embodiment 3
1) strain prepares：By bacillus amyloliquefaciens H47CCTCCM2013283 and commercially available bacillus pumilus, commercially available Soviet Union Cloud gold bacillus is inoculated in seed culture medium respectively, in 30 DEG C, 250r/min concussion and cultivates 12h；The seed culture based component is pressed G/L is calculated as：Peptone 10, yeast extract 5, NaCl 10, remaining composition is water, and 121 DEG C of autoclaving 15min adjust pH to 7.0. By 1:2:Seed is done in 2 mixing.
2) zymotic fluid prepares：From superior in quality Siberian cocklebur, rinsed with water, 70% -75% alcohol-pickled 3min uses water Clean, it is stand-by after being blended through pulper.
3) fermentation ullage disappears and air duct sterilization：Fermentation ullage is carried out by instructions book to disappear.
4) zymotic fluid disappears in fact：Step 2 gained Siberian cocklebur is put into fermentation tank, supplement running water to final volume is 10L, by phase Close specification and carry out real elimination bacterium.
5) inoculated and cultured：When fermentation jar temperature is down to 37 DEG C, by step 1 gained seed by volume 1% inoculum concentration, It is seeded in the fermentation tank equipped with Siberian cocklebur, is cultivated 6 days under the conditions of 30 DEG C, the throughput of technological requirement is set and tank pressure is kept, if Put other each culture parameters cultures.
6) discharge：Culture is finished, and releases zymotic fluid, is filtered, and gained zymotic fluid is insecticide finished product after filtering.

Claims (7)

1. a kind of method that Siberian cocklebur of fermenting prepares biological source insecticide.It is characterized in that using Siberian cocklebur as raw material, adding strain fermentation system Standby insecticide, this method comprises the following steps：
1) strain prepares：By bacillus amyloliquefaciens H47CCTCCM2013283 and commercially available bacillus pumilus, commercially available Su Yun gold Bacillus is inoculated in seed culture medium respectively, in 30 DEG C, 250r/min concussion and cultivates 12h；The seed culture based component is based on g/L For：Peptone 10, yeast extract 5, NaCl 10, remaining composition is water, and 121 DEG C of autoclaving 15min adjust pH to 7.0.By one Seed is done in certainty ratio mixing.
2) zymotic fluid prepares：From superior in quality Siberian cocklebur, rinsed with water, 70% -75% alcohol-pickled 3min is eluted with water, It is stand-by after being blended through pulper.
3) fermentation ullage disappears and air duct sterilization：Fermentation ullage is carried out by instructions book to disappear.
4) zymotic fluid disappears in fact：Step 2 gained Siberian cocklebur is put into fermentation tank, supplement running water to final volume is 10L, by mutually speaking on somebody's behalf Bright book carries out real elimination bacterium.
5) inoculated and cultured：When fermentation jar temperature is down to 37 DEG C, by step 1 gained seed by volume 1% inoculum concentration, inoculation Into the fermentation tank equipped with Siberian cocklebur, cultivated 3-12 days under the conditions of 25-30 DEG C, the throughput of technological requirement is set and tank is kept Pressure, sets other each culture parameters cultures.
6) discharge：Culture is finished, and releases zymotic fluid, is filtered, and gained zymotic fluid is insecticide finished product after filtering.
2. the method that fermentation Siberian cocklebur according to claim 1 prepares biological source insecticide, it is characterized in that：The strain is independent Seed culture medium culture 12h mix by a certain percentage after as fermentation seed.
3. the method that fermentation Siberian cocklebur according to claim 1 prepares biological source insecticide, it is characterized in that：Strain (solution starch Bacillus H47CCTCCM2013283 and commercially available bacillus pumilus, commercially available Dipel) be with seed culture based formulas (g/L)：Peptone 10, yeast extract 5, NaCl 10, remaining composition is water, and 121 DEG C of autoclaving 15min adjust pH to 7.0.
4. the method that fermentation Siberian cocklebur according to claim 1 prepares biological source insecticide, it is characterized in that：Siberian cocklebur processing method For：From superior in quality Siberian cocklebur, rinsed with water, 70% -75% alcohol-pickled 3min is eluted with water, after being blended through pulper It is stand-by.
5. the method that fermentation Siberian cocklebur according to claim 1 prepares biological source insecticide, it is characterized in that：The addition of strain For the 1% of fermentation system.
6. the method that fermentation Siberian cocklebur according to claim 1 prepares biological source insecticide, it is characterized in that：Raw material and strain are mixed Close uniform, 25-30 DEG C of fermentation temperature, fermentation time 3-12d.
7. the method that fermentation Siberian cocklebur according to claim 1 prepares biological source insecticide, it is characterized in that：The fermentation time cycle Short, process procedure is few, and the insecticide of preferable disinsection prophylaxis and growth-promoting functions can be obtained after fermentation.
CN201710243501.6A 2017-04-14 2017-04-14 A kind of method for preparing insecticide through microbial fermentation by raw material of Siberian cocklebur Pending CN106967752A (en)

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Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN108041102A (en) * 2017-12-05 2018-05-18 台州职业技术学院 A kind of method that diamondback moth biological agent is killed in fermented tcm compound material preparation
CN113817634A (en) * 2021-09-07 2021-12-21 江西省科学院微生物研究所 Bacillus amyloliquefaciens Bam-6 and preparation and application thereof

Citations (3)

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Publication number Priority date Publication date Assignee Title
CN1265834A (en) * 1999-03-07 2000-09-13 宛吉斌 Process for preparing natural plant biological biocide and biological fertilizer
CN101697740A (en) * 2009-04-04 2010-04-28 冯子龙 Novel traditional Chinese medicine pesticide and production technology thereof
CN103451128A (en) * 2013-07-12 2013-12-18 江南大学 Bacillus amyloliquefaciens and application thereof in preventing and controlling peach bleeding disease

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1265834A (en) * 1999-03-07 2000-09-13 宛吉斌 Process for preparing natural plant biological biocide and biological fertilizer
CN101697740A (en) * 2009-04-04 2010-04-28 冯子龙 Novel traditional Chinese medicine pesticide and production technology thereof
CN103451128A (en) * 2013-07-12 2013-12-18 江南大学 Bacillus amyloliquefaciens and application thereof in preventing and controlling peach bleeding disease

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Title
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Cited By (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN108041102A (en) * 2017-12-05 2018-05-18 台州职业技术学院 A kind of method that diamondback moth biological agent is killed in fermented tcm compound material preparation
CN108041102B (en) * 2017-12-05 2020-04-17 台州职业技术学院 Method for preparing plutella xylostella killing biological agent by fermenting traditional Chinese medicine composite raw materials
CN113817634A (en) * 2021-09-07 2021-12-21 江西省科学院微生物研究所 Bacillus amyloliquefaciens Bam-6 and preparation and application thereof
CN113817634B (en) * 2021-09-07 2023-01-17 江西省科学院微生物研究所 Bacillus amyloliquefaciens Bam-6 and preparation and application thereof

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