CN101341888A - Bolete extract, preparation and application thereof - Google Patents
Bolete extract, preparation and application thereof Download PDFInfo
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- CN101341888A CN101341888A CNA2008101179831A CN200810117983A CN101341888A CN 101341888 A CN101341888 A CN 101341888A CN A2008101179831 A CNA2008101179831 A CN A2008101179831A CN 200810117983 A CN200810117983 A CN 200810117983A CN 101341888 A CN101341888 A CN 101341888A
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Abstract
The invention discloses bolete extract and a preparation method thereof; the method mainly comprises the following steps: bolete is dried under 30 DEG C to 40 DEG C, crushed, sieved by a 60 mesh to 500 mesh sieve, extracted by phosphate buffer under 0 DEG C to 4 DEG C and centrifugated for 20min to 30min under 0 DEG C to 4 DEG C at 10000rpm to 12000rpm; supernatant fluid is collected and the bolete extract is obtained. The invention provides an effective method for preventing tobacco mosaic virus; the inhibition rate of the bolete extract to the tobacco mosaic virus reaches 67.49 percent to 92.51 percent; the bolete extract belongs to natural extract, has no harm to the human beings, is environment friendly; the production method of the invention is simple, and the cost is low.
Description
Technical field
The invention belongs to bio-extract, relate in particular to a kind of bolete extract and preparation method thereof, and the application of this extract on controlling plant diseases.
Background technology
Viral diseases of plants is the plant disease that is only second to fungal disease, causes serious harm in agricultural production.According to estimates, the loss that whole world every year causes because of viral diseases of plants is above 15,000,000,000 dollars, wherein the harm of tobacco mosaic virus disease (Tobacco mosaic virus) cause every year with a toll of more than 100,000,000 dollars (Wu Yunfeng. world agriculture, 1995, (5): 35～36).At present, also there is not an effectively preventing method for tobacco mosaic virus disease evil.Therefore, seek safety and the biogenic antiviral active substance of tool antiviral activity or the research focus that disease-resistant gene is current viroses of plant prevention and control field.
In the seventies in 19th century, people just find multiple basidioma, edible fungi submerged fermentation liquid and solid culture infusion to plant virus have inhibitory action (Wang Jingying etc. edible fungi of china, 1997,16 (2): the high wooden health of 6-7. to etc. plant disease day and report, 1977,43:211-214. the king is earlier refined etc. microorganism journal, 1986,26 (4): 363-365).In edible mushroom, also find the various active composition, as containing agglutinin in about 30～50 the edible mushroom kind, also find edible mushroom contain Antiphytoviral albumen (Wang Hexiang etc. edible fungi of china, 1997,16 (1): 3-4), research report from the yellow mushroom (Plearotus citrinopileatus) of elm (Fu Mingjia etc. Chinese virology, 2002,17 (4): 350-353), Asparagus (Flammulina velutipes) (Fu Mingjia etc. University Of Agriculture and Forestry In Fujian's journal (natural science edition), 2003,2 (1): 84-88), Xingbao mushroom (Pleurotus eryngii) (Fu Mingjia etc. the microorganism journal, 2003,43 (1): 29-34.), willow mushroom (Agrocybe aegerita) (Sun Hui etc. Acta Biochimica et Biophysica Sinica, 2001,33 (3): 351-354), ash flower tree (Grifola fiondoca) (Chen Ning etc. Acta Biochimica et Biophysica Sinica, 2004,31 (3): 283-286), shaggy cap (Coprinus cornatus) (Wu Liping etc. Chinese virology, 2004,19 (1): 54-57) etc. be separated to protein in the multiple eating bacterium with anti-TMV activity.Pixie stool (Cantharellus minor Peck), mushroom (Agaricns campestris L:Fr.), mushroom [Lentinu edodes (Berk.) Sing.], flat mushroom (Pluribus), tea tree mushroom [Agrocybeaegerita (Brig.) Sing.], coprinus comatus (Coprinus comattts), Hypsizygus marmoreus (Hypsizygusmarmoreus), 8 kinds of edible mushroom extracts of Asparagus (Flammulina velutipes) all have passivation virus (TMV) and suppress the effect that virus (TMV) infects external; Except that pixie stool (Cantharellus minorPeck), other 7 kinds of edible mushroom extracts have in various degree preventive and therapeutic effect (hippology duckweed etc., Yunnan Prov Agriculture University's journal, 2007,22 (2): 1-2) to TMV.Therefore, screening can suppress the macro fungi of plant virus and isolate the effective way that wherein antiviral active substance composition may be the control viral diseases of plants.
The amino acid composition that bolete contains abundant mineral element and is fit to has reducing blood lipid and antioxidation; Contain more polysaccharose substance, enrich blood in addition, the effect of ease constipation, hair care with antitumaous effect.But its effect aspect control of plant disease does not see that report is arranged.
Summary of the invention
The object of the invention is to provide a kind of bolete extract, and this extract can be used for preventing and treating tobacco mosaic virus etc.
Second purpose of the present invention provides above-mentioned bolete preparation method of extract.
The 3rd purpose of the present invention provides the purposes of above-mentioned bolete extract.
Realize that the object of the invention technical scheme is as follows:
A kind of bolete extract is prepared as follows:
(1) bolete is dried, pulverizes, crosses 60～200 mesh sieves down at 30～40 ℃, get bolete dry powder;
(2) under 0～4 ℃, be that 1: 10～20 ratio adds phosphate buffer according to w/v in step (1) gained bolete dry powder, the concussion mixing, mixed liquor; Described phosphate buffer is 20～30mmol/L, pH7.0～8.0;
(3) with step (2) gained mixed liquor centrifugal 20～30min under 0～4 ℃, 10000～12000rpm; Abandon precipitation, collect supernatant, be the bolete extract.
Bolete described in the above-mentioned bolete extract is meant black cattle liver bacterium (Boletus aereus) or Boletus luridus (Boletus auritpes) etc.
The available tissue mashing machine of pulverizing described in the above-mentioned bolete extract step (1) carries out.
Above-mentioned bolete extract can place-20 ℃ of freezing preservations standby.
Above-mentioned bolete preparation method of extract comprises the steps:
(1) bolete is dried, pulverizes, crosses 60～200 mesh sieves down at 30～40 ℃; Get bolete dry powder;
(2) under 0～4 ℃, be that 1: 10～20 ratio adds phosphate buffer according to w/v in the bolete dry powder of step (1) gained, the concussion mixing, mixed liquor; Described phosphate buffer is 20～30mmol/L, pH7.0～8.0;
(3) with step (2) gained mixed liquor centrifugal 20～30min under 0～4 ℃, 10000～12000rpm; Abandon precipitation, collect supernatant, promptly get the bolete extract.
Bolete described in the above-mentioned preparation method is meant black cattle liver bacterium (Boletus aereus) or Boletus luridus (Boletus auritpes) etc.
The available tissue mashing machine of pulverizing described in above-mentioned preparation method's step (1) carries out.
Above-mentioned bolete extract is in the application that prevents and treats on the tobacco mosaic virus.Its concrete usage is as follows:
It is 10.00～40.00 μ g/ml that above-mentioned bolete extract is diluted with water to protein concentration, with dilution length is put in order the strain spraying to the tobacco of 8～10 true leaves then and gets final product.
Above-mentioned bolete extract is in the application that promotes on the Chinese cabbage blastogenesis length.Concrete grammar is as follows: it is 10.00～40.00 μ g/ml that the bolete extract is diluted with water to protein concentration, and the bolete extract with dilution got final product the Chinese cabbage seeds seed soaking in 6～8 hours then.
Advantage that the present invention has and beneficial effect: (1) the present invention provides a kind of new way for preventing and treating tobacco mosaic virus, and the present invention can reach 92.51% to the inhibiting rate of tobacco mosaic virus; (2) the present invention can promote the growth of Chinese cabbage bud; (3) the invention belongs to natural extract, can not cause environmental pollution, harmless to the people, belong to environmentally friendly; (4) production method of the present invention is simple, cost is low.
Embodiment
The preparation of embodiment 1 black cattle liver fungus extract
(1) with commercially available black cattle liver bacterium 40 ℃ of oven dry, take by weighing 10g, smash with tissue mashing machine, cross 60 mesh sieves, black cattle liver bacterium dry powder;
(2) (pH8.0 25mmol/L), shakes mixing, and operation is all carried out on ice, gets mixed liquor to add the 100ml phosphate buffer to step (1) gained black cattle liver bacterium dry powder under 4 ℃;
(3) with the mixed liquor of step (2) gained centrifugal 20min under 4 ℃, 12000rpm; Abandon precipitation, collect supernatant, promptly get black cattle liver fungus extract 90ml ,-20 ℃ of freezing preservations are standby.
With the protein concentration of BCA protein concentration kit (U.S. Pierce company) detection black cattle liver fungus extract, assay method carries out according to the specification of kit, and the protein concentration of the above-mentioned black cattle liver fungus extract of result is: 7.9507mg/ml.
The preparation of embodiment 2 Boletus luridus extracts
(1) with commercially available Boletus luridus 40 ℃ of oven dry, take by weighing 10g, smash with tissue mashing machine, cross 60 mesh sieves, dry powder;
(2) (PH8.0 25mmol/L), shakes mixing, operates on ice and carries out, and gets mixed liquor add the 100ml phosphate buffer under 4 ℃ in step (1) gained Boletus luridus dry powder;
(3) with the mixed liquor of step (2) gained centrifugal 20min under 4 ℃, 12000rpm; Abandon precipitation, collect supernatant, promptly get Boletus luridus extract 90ml ,-20 ℃ of freezing preservations are standby.
With the protein concentration of BCA protein concentration kit (U.S. Pierce company) detection Boletus luridus extract, assay method carries out according to the specification of kit, and the protein concentration of the above-mentioned Boletus luridus extract of result is: 7.3425mg/ml.
Embodiment 3 black cattle liver fungus extracts are to the antitoxin activity test of tobacco mosaic virus
(1) handle: it is 10.00 μ g/ml, 20.00 μ g/ml, three concentration of 40.00 μ g/ml that the black cattle liver fungus extract water of embodiment 1 preparation liquid successively is diluted to protein concentration, does contrast with water;
The processing of pinching during (2) with the withered spot three lives tobacco leaf length to 8 of seedling age unanimity～10 true leaves keeps the 5th to the 8th true leaf, and withered spot three lives cigarette is put in order the strain spraying, and three repetitions are done in each processing, and each repeats three strains, three leaves of every strain;
(3) induce 72 hours after, the tobacco leaf that infects the common cigarette of tobacco mosaic virus is removed master pulse, mix at 1: 20 according to w/v with the 0.01M phosphate buffer of pH7.0, every 20ml adds 0.015g diamond dust, blade is ground, the withered spot three lives cigarette of inducing is carried out frictional inoculation, use flushing with clean water after the inoculation immediately, in hot-house culture;
(4), inoculation observes it and produces withered spot number amount after 72 hours, be calculated as follows out withered spot inhibiting rate.
X＝(CK-Y)/CK×100
Wherein X is withered spot inhibiting rate, unit: %; CK is the average withered spot number of clear water contrast blade, unit: individual; Y is for inducing the average withered spot number of handling rear blade, unit: individual through the black cattle liver fungus extract.
(5), result's (seeing Table 1) protein concentration average withered spot inhibiting rate of being respectively the black cattle liver fungus extract of 10.00 μ g/ml, 20.00 μ g/ml, 40.00 μ g/ml is respectively 79.94%, 90.62%, 92.51%.Illustrate that the black cattle liver fungus extract has the better prevention effect to tobacco mosaic virus.
Table 1 black cattle liver fungus extract resisting tobacco mosaic virus result of the test
Embodiment 4 Boletus luridus extracts are to the resistant proof of tobacco mosaic virus
(1) handle: with the Boletus luridus extract water of embodiment 2 preparations liquid diluted protein concentration successively is 10.00 μ g/ml, 20.00 μ g/ml, three concentration of 40.00 μ g/ml, does contrast with water;
The processing of pinching during (2) with the withered spot three lives tobacco leaf length to 8 of seedling age unanimity～10 true leaves keeps the 5th to the 8th true leaf, and withered spot three lives cigarette is put in order the strain spraying, and three repetitions are done in each processing, and each repeats three strains, three leaves of every strain;
(3) induce 72 hours after, the tobacco leaf that infects the common cigarette of tobacco mosaic virus is removed master pulse, mix with 1: 20 ratio of w/v with the 0.01M phosphate buffer of pH7.0, every 20ml adds 0.015g diamond dust, blade is ground, the withered spot three lives cigarette of inducing is carried out frictional inoculation, use flushing with clean water after the inoculation immediately, in hot-house culture;
(4), inoculation observes it and produces withered spot number amount after 72 hours, be calculated as follows out withered spot inhibiting rate.
X＝(CK-Y)/CK×100
Wherein X is withered spot inhibiting rate, unit: %; CK is the average withered spot number of clear water contrast blade, unit: individual; Y is for inducing the average withered spot number of handling rear blade, unit: individual through the black cattle liver fungus extract.
(5), result's (seeing Table 2) protein concentration average withered spot inhibiting rate of being respectively the Boletus luridus extract of 10.00 μ g/ml, 20.00 μ g/ml, 40.00 μ g/ml is respectively 53.47%, 67.49%, 67.44%.Illustrate that the Boletus luridus extract has the better prevention effect to tobacco mosaic virus.
Table 2 Boletus luridus extract resisting tobacco mosaic virus result of the test
5 Ns of bacillus extracts of embodiment promote the growth test of Chinese cabbage bud
(1) handle: the Chinese cabbage cultivar of test is Beijing 106.It is 10.00 μ g/ml, 20.00 μ g/ml, three concentration of 40.00 μ g/ml that the black cattle liver fungus extract of embodiment 1 preparation is diluted to protein concentration with distilled water liquid successively, is contrast with distilled water.
(2) soaked seed 6 hours with each concentration dilution liquid and contrast distilled water, afterwards seed is evenly put in the culture dish that 2 metafiltration paper are arranged 50 in every ware; Each handles 3 repetitions; In time replenish dilution every day in ware, keep filter paper moistening, and Ce Liang Xia Pei axial length and radicle are long respectively after 5 days, are calculated as follows out growth rate:
A＝(B-CK)/CK×100
Wherein A is a growth rate, unit: %; B is that De Xia Pei axial length or radicle are long after the black cattle liver fungus extract is handled, unit: cm; CK is long for contrast distilled water De Xia Pei axial length or radicle, unit: cm.
(3) result's (seeing Table 3) compared with the control, after the black cattle liver fungus extract that protein concentration is respectively 10.00 μ g/ml, 20.00 μ g/ml, 40.00 μ g/ml was handled, the length average growth rate of its radicle was respectively 57.26%, 118.53%, 106.19%; The average growth rate of Qi Xia Pei axle is respectively 4.21%, 15.77%, 13.09%.Illustrate that the bolete extract has obvious facilitation to the growth of Chinese cabbage bud.
Table 3 black cattle liver fungus extract is to the long result of the test that promotes of Chinese cabbage blastogenesis
Claims (6)
1, a kind of bolete extract is prepared as follows:
(1) bolete is dried, pulverizes, crosses 60～200 mesh sieves down at 30～40 ℃, get bolete dry powder;
(2) under 0～4 ℃, be that 1: 10～20 ratio adds phosphate buffer according to w/v in step (1) gained bolete dry powder, the concussion mixing, mixed liquor; Described phosphate buffer is 20～30mmol/L, pH7.0～8.0;
(3) with step (2) gained mixed liquor centrifugal 20～30min under 0～4 ℃, 10000～12000rpm; Abandon precipitation, collect supernatant, be the bolete extract.
2,, it is characterized in that described bolete is meant black cattle liver bacterium or Boletus luridus according to the described bolete extract of claim 1.
3, the described bolete preparation method of extract of claim 1 comprises the steps:
(1) bolete is dried, pulverizes, crosses 60～200 mesh sieves down at 30～40 ℃; Get bolete dry powder;
(2) under 0～4 ℃, be that 1: 10～20 ratio adds phosphate buffer according to w/v in the bolete dry powder of step (1) gained, the concussion mixing, mixed liquor; Described phosphate buffer is 20～30mmol/L, pH7.0～8.0;
(3) with step (2) gained mixed liquor centrifugal 20～30min under 0～4 ℃, 10000～12000rpm; Abandon precipitation, collect supernatant, promptly get the bolete extract.
4,, it is characterized in that described bolete is meant black cattle liver bacterium or Boletus luridus according to the described preparation method of claim 3.
5, the described bolete extract of claim 1 is in the application that prevents and treats on the tobacco mosaic virus.
6, the described bolete extract of claim 1 is in the application that promotes on the Chinese cabbage blastogenesis length.
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| CN2008101179831A CN101341888B (en) | 2008-08-19 | 2008-08-19 | Bolete extract, preparation and application thereof |
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| FR2961692A1 (en) * | 2010-06-25 | 2011-12-30 | Sothys Auriac | Cosmetic use of extract of edible mushroom stem including extract of mushroom or bolete, for treating skin aging, by applying topical composition containing extract and vehicle, preferably for providing antiglycation effect |
| CN102578522A (en) * | 2009-12-14 | 2012-07-18 | 楚雄宏桂绿色食品有限公司 | Boletus edulis dry powder with special flavour and preparation method of boletus edulis dry powder |
| CN101715948B (en) * | 2009-12-14 | 2012-09-26 | 楚雄宏桂绿色食品有限公司 | Bolete extracting solution and beverages as well as preparation methods thereof |
| CN102920016A (en) * | 2012-11-09 | 2013-02-13 | 成都宏亿实业集团有限公司 | Porcini extract and preparation method and application thereof |
| CN105326866A (en) * | 2015-11-25 | 2016-02-17 | 苏州普罗达生物科技有限公司 | Method for carrying out superfine wall breaking and smashing on boletus |
| CN109315758A (en) * | 2018-07-31 | 2019-02-12 | 云南农业大学 | Bolete is adjusting the application in intestinal flora |
| CN110563823A (en) * | 2019-10-14 | 2019-12-13 | 中华全国供销合作总社昆明食用菌研究所 | Preparation method of Boletus luteus lectin with high agglutination activity |
Family Cites Families (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1027785C (en) * | 1993-03-20 | 1995-03-08 | 辽宁省出口食用菌研究所 | Agricultural chemical prepared from edible fungus for curing and preventing plant virus |
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2008
- 2008-08-19 CN CN2008101179831A patent/CN101341888B/en not_active Expired - Fee Related
Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102578522A (en) * | 2009-12-14 | 2012-07-18 | 楚雄宏桂绿色食品有限公司 | Boletus edulis dry powder with special flavour and preparation method of boletus edulis dry powder |
| CN101715948B (en) * | 2009-12-14 | 2012-09-26 | 楚雄宏桂绿色食品有限公司 | Bolete extracting solution and beverages as well as preparation methods thereof |
| FR2961692A1 (en) * | 2010-06-25 | 2011-12-30 | Sothys Auriac | Cosmetic use of extract of edible mushroom stem including extract of mushroom or bolete, for treating skin aging, by applying topical composition containing extract and vehicle, preferably for providing antiglycation effect |
| CN102920016A (en) * | 2012-11-09 | 2013-02-13 | 成都宏亿实业集团有限公司 | Porcini extract and preparation method and application thereof |
| CN102920016B (en) * | 2012-11-09 | 2014-12-10 | 成都宏亿实业集团有限公司 | Porcini extract and preparation method and application thereof |
| CN105326866A (en) * | 2015-11-25 | 2016-02-17 | 苏州普罗达生物科技有限公司 | Method for carrying out superfine wall breaking and smashing on boletus |
| CN109315758A (en) * | 2018-07-31 | 2019-02-12 | 云南农业大学 | Bolete is adjusting the application in intestinal flora |
| CN110563823A (en) * | 2019-10-14 | 2019-12-13 | 中华全国供销合作总社昆明食用菌研究所 | Preparation method of Boletus luteus lectin with high agglutination activity |
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