CN103181401B - Algistat - Google Patents
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- CN103181401B CN103181401B CN201310059352.XA CN201310059352A CN103181401B CN 103181401 B CN103181401 B CN 103181401B CN 201310059352 A CN201310059352 A CN 201310059352A CN 103181401 B CN103181401 B CN 103181401B
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- bulbus allii
- allii fistulosi
- anhydrous propanone
- extract
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- SVPKNMBRVBMTLB-UHFFFAOYSA-N 2,3-dichloronaphthalene-1,4-dione Chemical compound C1=CC=C2C(=O)C(Cl)=C(Cl)C(=O)C2=C1 SVPKNMBRVBMTLB-UHFFFAOYSA-N 0.000 title abstract 2
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 claims abstract description 93
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims abstract description 41
- 239000000287 crude extract Substances 0.000 claims abstract description 41
- 230000002401 inhibitory effect Effects 0.000 claims abstract description 35
- 239000000284 extract Substances 0.000 claims abstract description 21
- 241000195493 Cryptophyta Species 0.000 claims description 24
- ULIKDJVNUXNQHS-UHFFFAOYSA-N 2-Propene-1-thiol Chemical compound SCC=C ULIKDJVNUXNQHS-UHFFFAOYSA-N 0.000 claims description 14
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 14
- WVDYBOADDMMFIY-UHFFFAOYSA-N Cyclopentanethiol Chemical compound SC1CCCC1 WVDYBOADDMMFIY-UHFFFAOYSA-N 0.000 claims description 13
- 238000000034 method Methods 0.000 claims description 13
- 239000003795 chemical substances by application Substances 0.000 claims description 10
- 239000000706 filtrate Substances 0.000 claims description 6
- 238000004821 distillation Methods 0.000 claims description 3
- 239000012153 distilled water Substances 0.000 claims description 3
- 239000003480 eluent Substances 0.000 claims description 3
- 238000010828 elution Methods 0.000 claims description 3
- 238000011068 loading method Methods 0.000 claims description 3
- 239000000463 material Substances 0.000 claims description 3
- 238000005406 washing Methods 0.000 claims description 3
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 abstract description 38
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 abstract description 32
- 230000000694 effects Effects 0.000 abstract description 13
- 241000234282 Allium Species 0.000 abstract description 8
- 230000005764 inhibitory process Effects 0.000 abstract description 8
- 235000002732 Allium cepa var. cepa Nutrition 0.000 abstract description 7
- 239000003960 organic solvent Substances 0.000 abstract description 4
- 241000192710 Microcystis aeruginosa Species 0.000 abstract description 3
- 239000003208 petroleum Substances 0.000 abstract 3
- 238000000746 purification Methods 0.000 abstract 1
- 235000019441 ethanol Nutrition 0.000 description 16
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 13
- 241000196324 Embryophyta Species 0.000 description 9
- 244000291564 Allium cepa Species 0.000 description 7
- 235000010167 Allium cepa var aggregatum Nutrition 0.000 description 7
- 239000000243 solution Substances 0.000 description 7
- 230000002195 synergetic effect Effects 0.000 description 4
- 238000004458 analytical method Methods 0.000 description 3
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 description 3
- 238000002290 gas chromatography-mass spectrometry Methods 0.000 description 3
- XLYOFNOQVPJJNP-ZSJDYOACSA-N Heavy water Chemical compound [2H]O[2H] XLYOFNOQVPJJNP-ZSJDYOACSA-N 0.000 description 2
- OAICVXFJPJFONN-UHFFFAOYSA-N Phosphorus Chemical compound [P] OAICVXFJPJFONN-UHFFFAOYSA-N 0.000 description 2
- 244000249201 Scenedesmus obliquus Species 0.000 description 2
- 235000007122 Scenedesmus obliquus Nutrition 0.000 description 2
- 239000003627 allelochemical Substances 0.000 description 2
- 150000001875 compounds Chemical class 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 238000012851 eutrophication Methods 0.000 description 2
- 238000011156 evaluation Methods 0.000 description 2
- 238000002474 experimental method Methods 0.000 description 2
- 238000001914 filtration Methods 0.000 description 2
- 238000005286 illumination Methods 0.000 description 2
- 238000002386 leaching Methods 0.000 description 2
- 239000007788 liquid Substances 0.000 description 2
- 244000005700 microbiome Species 0.000 description 2
- 239000012982 microporous membrane Substances 0.000 description 2
- 238000002156 mixing Methods 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 238000001543 one-way ANOVA Methods 0.000 description 2
- 229910052698 phosphorus Inorganic materials 0.000 description 2
- 239000011574 phosphorus Substances 0.000 description 2
- 239000000419 plant extract Substances 0.000 description 2
- 239000000047 product Substances 0.000 description 2
- 239000011347 resin Substances 0.000 description 2
- 229920005989 resin Polymers 0.000 description 2
- 239000012488 sample solution Substances 0.000 description 2
- 231100000331 toxic Toxicity 0.000 description 2
- 230000002588 toxic effect Effects 0.000 description 2
- ZRKMQKLGEQPLNS-UHFFFAOYSA-N 1-Pentanethiol Chemical compound CCCCCS ZRKMQKLGEQPLNS-UHFFFAOYSA-N 0.000 description 1
- DYSJMQABFPKAQM-UHFFFAOYSA-N 1-benzothiophene-2-carboxylic acid Chemical compound C1=CC=C2SC(C(=O)O)=CC2=C1 DYSJMQABFPKAQM-UHFFFAOYSA-N 0.000 description 1
- 244000257727 Allium fistulosum Species 0.000 description 1
- 235000008553 Allium fistulosum Nutrition 0.000 description 1
- 241000192700 Cyanobacteria Species 0.000 description 1
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 1
- 206010020880 Hypertrophy Diseases 0.000 description 1
- 241000234280 Liliaceae Species 0.000 description 1
- NINIDFKCEFEMDL-UHFFFAOYSA-N Sulfur Chemical compound [S] NINIDFKCEFEMDL-UHFFFAOYSA-N 0.000 description 1
- 239000000654 additive Substances 0.000 description 1
- 230000000996 additive effect Effects 0.000 description 1
- 230000002581 algistatic effect Effects 0.000 description 1
- 239000007864 aqueous solution Substances 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- 230000008033 biological extinction Effects 0.000 description 1
- 238000010170 biological method Methods 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 239000012141 concentrate Substances 0.000 description 1
- 239000000470 constituent Substances 0.000 description 1
- 229960001760 dimethyl sulfoxide Drugs 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 238000003912 environmental pollution Methods 0.000 description 1
- 150000002148 esters Chemical class 0.000 description 1
- 230000003203 everyday effect Effects 0.000 description 1
- 229960002413 ferric citrate Drugs 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 238000002309 gasification Methods 0.000 description 1
- 230000009036 growth inhibition Effects 0.000 description 1
- 238000010438 heat treatment Methods 0.000 description 1
- 150000002500 ions Chemical class 0.000 description 1
- NPFOYSMITVOQOS-UHFFFAOYSA-K iron(III) citrate Chemical compound [Fe+3].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O NPFOYSMITVOQOS-UHFFFAOYSA-K 0.000 description 1
- 230000009916 joint effect Effects 0.000 description 1
- 238000012417 linear regression Methods 0.000 description 1
- 238000004949 mass spectrometry Methods 0.000 description 1
- 235000015097 nutrients Nutrition 0.000 description 1
- 235000016709 nutrition Nutrition 0.000 description 1
- 230000035764 nutrition Effects 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 238000012545 processing Methods 0.000 description 1
- 230000035755 proliferation Effects 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 150000003839 salts Chemical class 0.000 description 1
- 238000005070 sampling Methods 0.000 description 1
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- 239000002904 solvent Substances 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 229910052717 sulfur Inorganic materials 0.000 description 1
- 239000011593 sulfur Substances 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
- 239000003053 toxin Substances 0.000 description 1
- 231100000765 toxin Toxicity 0.000 description 1
- 108700012359 toxins Proteins 0.000 description 1
- 230000007306 turnover Effects 0.000 description 1
Landscapes
- Medicines Containing Plant Substances (AREA)
- Agricultural Chemicals And Associated Chemicals (AREA)
Abstract
The invention discloses an algistat, which is an absolute methanol crude extract of onion stalks, an absolute acetone crude extract of the onion stalks, an absolute petroleum ether crude extract of the onion stalks or an ethanol crude extract of the onion stalks. Compared with the prior art, the crude extracts from the onion stalks by the organic solvents perform an inhibitory effect to microcystis aeruginosa from high to low as follows: the inhibitory effect of absolute acetone is larger than that of 75% ethanol, the inhibitory effect of the 75% ethanol is larger than that of absolute petroleum ether, and the inhibitory effect of the absolute petroleum ether is larger than that of absolute methanol. The inhibition ratio of the absolute acetone pure extract of the onion stalks is enhanced along with the time, and a concentration effect is shown. The inhibition ratio of a high dose group on the fourth day can be up to 97.7%. EC50 (50% effective concentration) computed on the fifth day is 0.03g/L, which proves that the algal inhibition effect of the purified absolute acetone extract is more obvious than that before purification.
Description
Technical field
The invention belongs to this technical field of algae-inhibiting agent.
Background technology
In recent years, along with the increase of mankind's activity, lake, Reservoir Eutrophication are day by day serious, and cause wawter bloom frequently to break out, the usefulness that utilizes of water resource reduces, and therefore aquatic products industry economy be subject to huge loss, and therefore water environment safety is also faced with serious threat.Therefore, effectively suppress the excess growth of planktonic algae in eutrophication water and prevent wawter bloom from becoming study hotspot and the forward position of current environmental area.In the method for numerous control algal grown, the method for traditional physics, Chemical Inhibition algal grown can inevitably be destroyed the ecological balance and cause environmental pollution.And biological method especially utilizes Allelopathic Effect in Plants algal control to be considered to a kind of novel biological algal control technology, there is the features such as efficient, ecological security is good, thus quite concerned.At present to utilizing Allelopathic Effect in Plants algal control mainly to concentrate on heavy water, very water and phytoplankton
?research, but nutrient salt in water directly coerce the extinction that can cause submerged plant with toxic action; The phosphorus concentration maintaining shallow lake clear water state is generally 0.05-0.15 mg/L, when the total phosphorus concentration in water body is more than 0.15 mg/L, lake can start from careless type turnover ecosystem be the algae type ecosystem.Moreover, due to planktonic algae raised growth, water plants not only can be subject to the impact of various envirment factor, and the impact of the aspect such as nutrition, illumination can be subject to and cause withering away, therefore the cover degree that will reach performance effect of algae restraint is more difficult realization, thus effectively cannot control the excessive proliferation of harmful algae.Meanwhile, blue-green algae also can affect the allelopathic potential of water plants by the secretion of the allelochemicals such as Algae toxins.In addition, water plants is subject to again the restriction of season and weather, and therefore the algal control of water plants allelopathic has certain limitation in actual applications.
Summary of the invention
Technical problem to be solved by this invention is to provide a kind of algae-inhibiting agent containing terrestrial plant extract.
The technical scheme of technical solution problem of the present invention is: a kind of algae-inhibiting agent containing plant extracts, described algae-inhibiting agent is absolute methanol crude extract, the anhydrous propanone crude extract of Bulbus Allii Fistulosi, the dry oil ether crude extract of Bulbus Allii Fistulosi, the alcohol extracts of Bulbus Allii Fistulosi of Bulbus Allii Fistulosi.
The absolute methanol crude extract of described Bulbus Allii Fistulosi is prepared by the following method:
Be 100g:400mL in the ratio of Bulbus Allii Fistulosi and absolute methanol, room temperature lixiviate is filtered two days later, and collect filtrate, reduced pressure concentration volatilizes methyl alcohol, is dried to constant weight.
The anhydrous propanone crude extract of described Bulbus Allii Fistulosi is prepared by the following method:
Be 100g:400mL in the ratio of Bulbus Allii Fistulosi and anhydrous propanone, room temperature lixiviate is filtered two days later, and collect filtrate, reduced pressure concentration volatilizes anhydrous propanone, is dried to constant weight.
The absolute ether crude extract of described Bulbus Allii Fistulosi is prepared by the following method:
Be 100g:400mL in the ratio of Bulbus Allii Fistulosi and absolute ether, room temperature lixiviate is filtered two days later, and collect filtrate, reduced pressure concentration volatilizes absolute ether, is dried to constant weight.
The alcohol extracts of described Bulbus Allii Fistulosi is prepared by the following method:
By Bulbus Allii Fistulosi and 75%(ml/ml) proportion of ethanol be 100g:400mL, room temperature lixiviate is filtered two days later, collect filtrate, reduced pressure concentration volatilizes ethanol, is dried to constant weight.
By the anhydrous propanone crude extract of Bulbus Allii Fistulosi, dilute with anhydrous propanone, load in dropping funel, by dropping funel to chromatographic column (macroreticular resin XAD1180) drip, after loading is complete, with distillation washing post, till the distilled water flowed out in chromatographic column mixes with 95% ethanol (ml/ml) and do not produce muddiness, then carry out wash-out with the acetone of 2BV, in chromatographic column, the elution rate of eluent is constant is 0.5BV/h, volatilize anhydrous propanone through reduced pressure concentration again, the pure extract of anhydrous propanone of Bulbus Allii Fistulosi can be obtained.
A kind of algae-inhibiting agent, described algae-inhibiting agent is allyl mercaptan, one or several mixture of cyclopentanethiol.
Shallot (Allium fistulosum L.var. giganteam Makino) is common terrestrial plant, belongs to Liliaceae allium, is the raw herbaceous plant of 2-3 that is product with the false stem of the hypertrophy of leaf sheath composition and tender leaf; Shallot reached for more than 3000 years at the cultivation history of China, was one of country of Main Cultivation shallot in the world, and it is made up of Bulbus Allii Fistulosi and green onion leaf.
Compared with prior art, the Bulbus Allii Fistulosi crude extract of organic solvent to the inhibition of microcystic aeruginosa is sequentially from high to low: anhydrous propanone >75% ethanol > dry oil ether > absolute methanol in the present invention.The inhibiting rate of the pure extract of anhydrous propanone of Bulbus Allii Fistulosi increases along with the increase of time, and shows concentration effect.High dose group reaches 97.78% at the inhibiting rate of the 4th day; EC50 calculated by the 5th day, and result is 0.03g/L, and the crude extract effect of algae restraint namely after purifying comparatively purifying is significantly front.
Embodiment
Below in conjunction with embodiment, the present invention is described in detail.
Experiment material:
1.1
Microcystic aeruginosa (Microcystis aeruginosa) and scenedesmus obliquus (Scenedesmus obliquus) are provided by Inst. of Hydrobiology, Chinese Academy of Sciences's algae kind storehouse; Adopt BG-11 medium culture.
Fresh shallot, is purchased from food market, Wuhu City.
It is pure that absolute methanol, anhydrous propanone, dry oil ether and absolute ethyl alcohol, allyl mercaptan, cyclopentanethiol are analysis.
BG-11 medium main component
NaNO
3, 1.5 g/L; K
2hPO
43H
2o, 0.04 g/L; MgSO
47H
2o, 0.075 g/L; CaCl
22H
2o, 0.036 g/L; Citric acid, 0.006 g/L; Ferric citrate, 0.006 g/L; EDTA, 0.001 g/L; Na
2cO
3, 0.02 g/L; A
5solution, 0.1 mL;
A
5solution fills a prescription: H
3bO
3, 2.86 g/L; MnCl
24H
2o, 1.86 g/L; ZnSO
47H
2o, 0.22 g/L; Na
2moO
42H
2o, 0.39 g/L; CuSO
4. 5H
2o, 0.08 g/L; Co (NO
3)
26H
2o, 0.05 g/L.
1.2
Cultivate with carrying out expansion to microcystic aeruginosa before experiment.Condition of culture is: intensity of illumination 4000 lx, and Light To Dark Ratio is 12h:12h, temperature (25 ± 1) DEG C.Shake 3 ~ 5 every day, make Microcystis aeruginosa Strains enter exponential phase.
1.3 assay method
Microcystic aeruginosa density sepectrophotofluorometer counts; The constituent analysis of shallot Bulbus Allii Fistulosi uses Shimadzu GC-MS QP2010plus gas chromatograph-mass spectrometer.
1.4 data processing
Inhibiting rate (inhibition rate) formula of allelochemical to algae is:
IR?=?(1-N/N
0)×100%
In formula, IR-inhibiting rate; The fluorescent value of N-add leaching liquor group; N
0the fluorescent value of-control group.Shallot leaching liquor ultimate density and growth inhibition ratio are made one-variable linear regression, obtains half effect concentration EC
50(mL/L).
Data acquisition Excel 2010 software process, each group difference adopts one-way analysis of variance (one-way ANOVA), and P<0.05 indicates significant difference, and P<0.01 indicates pole significant difference.
Embodiment 1:
Get fresh shallot Bulbus Allii Fistulosi to remove the peel, grind, be 100g:400mL in the ratio of Bulbus Allii Fistulosi and organic solvent (absolute methanol, anhydrous propanone, dry oil ether, 75% ethanol).Lixiviate is filtered two days later, collect filtrate, volatilize organic solvent through reduced pressure concentration, crude extract is put in the beaker of constant weight, volatilize to constant weight, obtain the absolute methanol crude extract of Bulbus Allii Fistulosi, the anhydrous propanone crude extract of Bulbus Allii Fistulosi, the dry oil ether crude extract of Bulbus Allii Fistulosi, the ethanol crude extract of Bulbus Allii Fistulosi.
Embodiment 2:
By the anhydrous propanone crude extract of Bulbus Allii Fistulosi, 10% is diluted to anhydrous propanone, load in dropping funel, by dropping funel to chromatographic column (macroreticular resin XAD1180) drip, after loading is complete, with distillation washing post, till the distilled water flowed out in chromatographic column mixes with 95% ethanol (ml/ml) and does not produce muddiness, wash-out is carried out again with the acetone of 2BV, in chromatographic column, the elution rate of eluent is constant is 0.5BV/h, anhydrous propanone is volatilized again through reduced pressure concentration, obtain the pure extract of anhydrous propanone of Bulbus Allii Fistulosi.
Embodiment 3:
Made for embodiment 1 four kinds of Bulbus Allii Fistulosi crude extracts (the dry oil ether crude extract of the absolute methanol crude extract of Bulbus Allii Fistulosi, the anhydrous propanone crude extract of Bulbus Allii Fistulosi, Bulbus Allii Fistulosi, the ethanol crude extract of Bulbus Allii Fistulosi) are dissolved in (DMSO) in methyl-sulfoxide respectively, adding BG-11 medium again, to be made into concentration be 0.1g/L to control DMSO final concentration <l% (solvent DMSO aqueous solution inhibiting rate is under test conditions 1.06%, and this value has been included in error when inhibiting rate calculates).Aseptically pass through the filtering with microporous membrane of 0.22 μm to eliminate microorganism, obtain four kinds of sample solutions.
The microcystic aeruginosa of taking the logarithm vegetative period puts into sterilized 100mL conical flask respectively, and frustule ultimate density is 5.5 × 10
5cells/mL, adds four kinds of sample solution solution respectively, and make final volume be 50mL, ultimate density is 0.1g/L.Establish the blank group (CK) not adding crude extract simultaneously, often kind of processed group establish 3 parallel, condition of culture is with 1.2.Carry out the mensuration of fluorescent value every 24h, survey 5 days continuously.
Its result is as shown in table 1, table 2:
The crude extract of table 1 four kinds of Bulbus Allii Fistulosi is to the inhibiting rate (%) of microcystic aeruginosa
The EC that the crude extract of table 2 four kinds of Bulbus Allii Fistulosi suppresses microcystic aeruginosa
50
What table 1 showed is under microcystic aeruginosa initial density the same terms, and the anhydrous propanone crude extract of Bulbus Allii Fistulosi reaches 90.32% at the 4th day inhibiting rate, and within the 5th day, microcystic aeruginosa is almost killed completely; 75% alcohol extracts of the absolute methanol crude extract of the Bulbus Allii Fistulosi that concentration is identical, the dry oil ether crude extract of Bulbus Allii Fistulosi, Bulbus Allii Fistulosi is respectively 81.66%, 82.35% and 87.28% at the inhibiting rate of the 5th day.This illustrates, 75% alcohol extracts of the absolute methanol crude extract of Bulbus Allii Fistulosi, the dry oil ether crude extract of Bulbus Allii Fistulosi, Bulbus Allii Fistulosi will differ from the inhibition of microcystic aeruginosa.That table 2 shows is the EC that four kinds of Bulbus Allii Fistulosi crude extracts suppress microcystic aeruginosa
50value.Result shows, the EC of the anhydrous propanone crude extract of Bulbus Allii Fistulosi
50minimum, be 2.49g/L.The EC of 75% alcohol extracts of the absolute methanol crude extract of Bulbus Allii Fistulosi, the dry oil ether crude extract of Bulbus Allii Fistulosi, Bulbus Allii Fistulosi
50be more or less the same, and be all greater than the anhydrous propanone crude extract of Bulbus Allii Fistulosi.Again prove that the anhydrous propanone crude extract algistatic activity of Bulbus Allii Fistulosi is best.
Embodiment 4:
Claim the made pure extract of embodiment 2 to be dissolved in DMSO, then add BG-11 medium wiring solution-forming, aseptically pass through the filtering with microporous membrane of 0.22 μm to eliminate microorganism, obtain the pure extract solution of anhydrous propanone of Bulbus Allii Fistulosi.
The microcystic aeruginosa of taking the logarithm vegetative period puts into sterilized 100mL conical flask respectively, and frustule ultimate density is 5.5 × 10
5cells/mL, adds the pure extract solution of anhydrous propanone of Bulbus Allii Fistulosi, and makes final volume be 50mL.In conical flask, the anhydrous propanone extract solution ultimate density of Bulbus Allii Fistulosi is respectively 0.5g/L, 1.0g/L, 2.0g/L, arranges the blank group (CK) not adding extract simultaneously, often group establish 3 parallel, condition of culture is with 1.2.Carry out the mensuration of fluorescent value every 24h, survey five days continuously.
Its result is as shown in table 3, table 4:
The pure extract of anhydrous propanone of table 3 Bulbus Allii Fistulosi is to the inhibiting rate (%) of microcystic aeruginosa
The pure extract of anhydrous propanone of table 4 Bulbus Allii Fistulosi is to the EC suppressed microcystic aeruginosa
50
Table 3 shows, and along with the increase inhibiting rate of time is increase tendency, and has concentration effect.At the 4th day, the inhibiting rate of high dose group just reached 97.78%, almost completely suppressed at the 5th day microcystic aeruginosa; Other processed group are respectively at the inhibiting rate of the 5th day: 58.93% and 80.57%.Table 4 shows, along with the increase of concentration, and EC
50value reduces gradually.Show the anhydrous propanone of the Bulbus Allii Fistulosi after purifying pure extract effect of algae restraint highly significant.
Embodiment 5
Gas chromatography mass spectrometry (GC-MS) analysis is carried out to the pure extract of the anhydrous propanone of Bulbus Allii Fistulosi.GC conditions is, injector temperature: 260 DEG C of gasifications, flow velocity: 1.50mL/min, split ratio: 100:1, post type: RTX-5,30 × 0.25 × 0.25, heating schedule: initial temperature 40 DEG C, after keeping 2min, under 10 DEG C/min speed, be heated to 250 DEG C, keep 5min; Mass Spectrometry Conditions is, ion source temperature: 200 DEG C, interface temperature: 250 DEG C, detector voltage: 0.81 kV, sweep interval: 0.5Sec, scanning charge-mass ratio: 400-700aum, sampling volume: 1 μ L.Application NIST mass spectrometric data storehouse, analyzes mass spectrogram, analyzes each component materials.Go out the higher compound of 6 kinds of content by GC-MS Analysis and Identification, be mainly sulfur-containing compound, be respectively: allyl mercaptan
, cyclopentanethiol
, benzothiophene-2-carboxylic acid
, 3,4-thioxenes
in addition ether and ester is also had.
Embodiment 6:
Added by the microcystic aeruginosa algae liquid of exponential phase in sterilized 100 mL conical flasks, the final densities of frustule is 3.0 × 10
5cells/mL, and make volume finally remain 50mL.In conical flask, allyl mercaptan, cyclopentanethiol ultimate density that is independent and two kinds of alcohol mixing (1:1) are respectively 0,0.01,0.03,0.09,0.27 and 0.81g/L.Wherein, the half when concentration of often planting alcohol during the mixing of two kinds of alcohol is independent, establishes the blank group (CK) only adding algae liquid simultaneously, often group establish 3 parallel, each group be placed in 1.2 the same terms under cultivate, carry out the mensuration of fluorescent value every 24h.
Its result is as shown in table 5,6,7,8,9,10:
Table 5 allyl mercaptan is to the inhibiting rate (%) of microcystic aeruginosa
Table 6 cyclopentanethiol is to the inhibiting rate (%) of microcystic aeruginosa
To the inhibiting rate (%) of microcystic aeruginosa when table 7 allyl mercaptan and cyclopentanethiol
The EC that table 8 allyl mercaptan suppresses microcystic aeruginosa
50
The EC that table 9 cyclopentanethiol suppresses microcystic aeruginosa
50
EC when table 10 allyl mercaptan and the associating of ring change amyl hydrosulfide, microcystic aeruginosa suppressed
50
Table 5,6 and 7 displays, separately and associating time to the prolongation along with the time of the inhibiting rate of microcystic aeruginosa, the increase of concentration, all in increase tendency.During independent algal control, allyl mercaptan reaches 90.51% the inhibiting rate of the 3rd day is maximum, and cyclopentanethiol reaches 90.11% the inhibiting rate of the 4th day is maximum; Associating algal control reaches 97.90% the inhibiting rate of the 3rd day is maximum; Microcystic aeruginosa all almost killed at the 5th day by independent and synergy completely.During independent algal control, the two does not all reach 50% at the inhibiting rate of first day and second day, therefore 503nhibiting concentration calculates according to other skies, and that table 8,9 and 10 shows is independent and the EC of associating algal control
50, EC during allyl mercaptan independent role
50be respectively 0.22,0.08 and 0.04 g/L; EC during cyclopentanethiol independent role
50be respectively 0.21,0.08 and 0.05g/L; During the two synergy, the 503nhibiting concentration of five days is 0.56,0.35,0.13,0.05 and 0.02 g/L respectively.Result shows, during independent role, the effect of algae restraint of allyl mercaptan is better than cyclopentanethiol, and effect of algae restraint during associating algal control is better than independent algal control.
Table 11 synergy evaluating
Can find out from table 11, according to toxic unit method, M<1, the method evaluation result is synergistic effect; According to Additive Index, the AI=1/M-1 as M<1, calculates AI>0, is synergistic effect; According to Similarity Parameter method, calculate λ >1 by trial and error method, be evaluated as synergistic effect.By above-mentioned 3 kinds of methods, joint effect evaluation is carried out to allyl mercaptan and cyclopentanethiol, all show as synergistic effect, absolutely prove that allyl mercaptan and cyclopentanethiol have synergy to microcystic aeruginosa, to the inhibition of microcystic aeruginosa will get well when Combined Ration is independent both demonstrating simultaneously.
Claims (2)
1. an algae-inhibiting agent, is characterized in that: described algae-inhibiting agent is the anhydrous propanone extract of Bulbus Allii Fistulosi, containing allyl mercaptan, cyclopentanethiol two kinds of materials;
The anhydrous propanone method for preparing extractive of described Bulbus Allii Fistulosi is: by the anhydrous propanone crude extract of Bulbus Allii Fistulosi, dilute with anhydrous propanone, load in dropping funel, by dropping funel to chromatographic column drip, after loading is complete, with distillation washing post, till the distilled water flowed out in chromatographic column mixes with 95% ethanol (ml/ml) and does not produce muddiness, wash-out is carried out again with the acetone of 2BV, in chromatographic column, the elution rate of eluent is constant is 0.5BV/h, anhydrous propanone is volatilized again through reduced pressure concentration, obtain the anhydrous propanone extract of Bulbus Allii Fistulosi.
2. a kind of algae-inhibiting agent according to claim 1, is characterized in that: the anhydrous propanone crude extract of described Bulbus Allii Fistulosi is prepared by the following method:
Be 100g:400mL in the ratio of Bulbus Allii Fistulosi and anhydrous propanone, room temperature lixiviate is filtered two days later, and collect filtrate, reduced pressure concentration volatilizes anhydrous propanone, is dried to constant weight.
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