CN109999103A - Tobacco active component (TEP), extracting method and its application - Google Patents
Tobacco active component (TEP), extracting method and its application Download PDFInfo
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- CN109999103A CN109999103A CN201910167624.5A CN201910167624A CN109999103A CN 109999103 A CN109999103 A CN 109999103A CN 201910167624 A CN201910167624 A CN 201910167624A CN 109999103 A CN109999103 A CN 109999103A
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- 235000002637 Nicotiana tabacum Nutrition 0.000 title claims abstract description 101
- 238000000034 method Methods 0.000 title claims abstract description 34
- 244000061176 Nicotiana tabacum Species 0.000 title 1
- 241000208125 Nicotiana Species 0.000 claims abstract description 100
- 238000000605 extraction Methods 0.000 claims abstract description 59
- 230000000844 anti-bacterial effect Effects 0.000 claims abstract description 35
- 239000002904 solvent Substances 0.000 claims abstract description 31
- 239000000463 material Substances 0.000 claims abstract description 25
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 25
- 239000000284 extract Substances 0.000 claims abstract description 15
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims abstract description 13
- 238000012545 processing Methods 0.000 claims abstract description 13
- 229940088710 antibiotic agent Drugs 0.000 claims abstract description 4
- 239000000843 powder Substances 0.000 claims description 17
- 239000013049 sediment Substances 0.000 claims description 13
- 239000000470 constituent Substances 0.000 claims description 11
- 230000008569 process Effects 0.000 claims description 10
- 238000001035 drying Methods 0.000 claims description 9
- 239000006228 supernatant Substances 0.000 claims description 9
- 238000005119 centrifugation Methods 0.000 claims description 6
- 239000008213 purified water Substances 0.000 claims description 6
- 238000001816 cooling Methods 0.000 claims description 5
- 238000010298 pulverizing process Methods 0.000 claims description 5
- 230000001376 precipitating effect Effects 0.000 claims description 4
- 230000001476 alcoholic effect Effects 0.000 claims description 3
- 238000004108 freeze drying Methods 0.000 claims description 3
- 239000007788 liquid Substances 0.000 claims description 3
- 238000002360 preparation method Methods 0.000 claims description 3
- 238000001556 precipitation Methods 0.000 claims description 2
- 235000019504 cigarettes Nutrition 0.000 claims 1
- 238000005138 cryopreservation Methods 0.000 claims 1
- 230000000694 effects Effects 0.000 claims 1
- 238000002474 experimental method Methods 0.000 abstract description 24
- 235000019441 ethanol Nutrition 0.000 abstract description 7
- 239000002798 polar solvent Substances 0.000 abstract description 4
- 238000007781 pre-processing Methods 0.000 abstract description 2
- 238000000746 purification Methods 0.000 abstract description 2
- 235000019505 tobacco product Nutrition 0.000 abstract description 2
- 238000009509 drug development Methods 0.000 abstract 1
- 239000002547 new drug Substances 0.000 abstract 1
- 241000894006 Bacteria Species 0.000 description 17
- 229920001817 Agar Polymers 0.000 description 16
- 239000008272 agar Substances 0.000 description 16
- 230000005764 inhibitory process Effects 0.000 description 12
- 239000000243 solution Substances 0.000 description 11
- 238000012360 testing method Methods 0.000 description 11
- 230000002401 inhibitory effect Effects 0.000 description 8
- 239000000047 product Substances 0.000 description 8
- 239000007787 solid Substances 0.000 description 8
- 239000000022 bacteriostatic agent Substances 0.000 description 7
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 description 6
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 description 5
- 244000063299 Bacillus subtilis Species 0.000 description 5
- 235000014469 Bacillus subtilis Nutrition 0.000 description 5
- 241001264174 Cordyceps militaris Species 0.000 description 5
- 241000588724 Escherichia coli Species 0.000 description 5
- 239000003814 drug Substances 0.000 description 5
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 4
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 4
- 241000736199 Paeonia Species 0.000 description 4
- 235000006484 Paeonia officinalis Nutrition 0.000 description 4
- 239000001888 Peptone Substances 0.000 description 4
- 108010080698 Peptones Proteins 0.000 description 4
- 241000191967 Staphylococcus aureus Species 0.000 description 4
- 230000003385 bacteriostatic effect Effects 0.000 description 4
- 238000009395 breeding Methods 0.000 description 4
- 230000001488 breeding effect Effects 0.000 description 4
- 229940041514 candida albicans extract Drugs 0.000 description 4
- 238000010586 diagram Methods 0.000 description 4
- 238000010790 dilution Methods 0.000 description 4
- 239000012895 dilution Substances 0.000 description 4
- 239000008103 glucose Substances 0.000 description 4
- 230000002147 killing effect Effects 0.000 description 4
- VLKZOEOYAKHREP-UHFFFAOYSA-N n-Hexane Chemical compound CCCCCC VLKZOEOYAKHREP-UHFFFAOYSA-N 0.000 description 4
- 235000019319 peptone Nutrition 0.000 description 4
- 238000007711 solidification Methods 0.000 description 4
- 230000008023 solidification Effects 0.000 description 4
- 230000001954 sterilising effect Effects 0.000 description 4
- 238000004659 sterilization and disinfection Methods 0.000 description 4
- 239000012138 yeast extract Substances 0.000 description 4
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 description 3
- 230000007480 spreading Effects 0.000 description 3
- 238000003892 spreading Methods 0.000 description 3
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 description 2
- 239000011248 coating agent Substances 0.000 description 2
- 238000000576 coating method Methods 0.000 description 2
- 229960004756 ethanol Drugs 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 239000003208 petroleum Substances 0.000 description 2
- 241001478240 Coccus Species 0.000 description 1
- 241001646834 Mesona Species 0.000 description 1
- 238000000944 Soxhlet extraction Methods 0.000 description 1
- 235000009754 Vitis X bourquina Nutrition 0.000 description 1
- 235000012333 Vitis X labruscana Nutrition 0.000 description 1
- 240000006365 Vitis vinifera Species 0.000 description 1
- 235000014787 Vitis vinifera Nutrition 0.000 description 1
- 150000001298 alcohols Chemical class 0.000 description 1
- 229930013930 alkaloid Natural products 0.000 description 1
- 150000003797 alkaloid derivatives Chemical class 0.000 description 1
- 230000000259 anti-tumor effect Effects 0.000 description 1
- 230000000840 anti-viral effect Effects 0.000 description 1
- 230000003078 antioxidant effect Effects 0.000 description 1
- 239000007864 aqueous solution Substances 0.000 description 1
- 150000001720 carbohydrates Chemical class 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 229960000935 dehydrated alcohol Drugs 0.000 description 1
- ZPWVASYFFYYZEW-UHFFFAOYSA-L dipotassium hydrogen phosphate Chemical compound [K+].[K+].OP([O-])([O-])=O ZPWVASYFFYYZEW-UHFFFAOYSA-L 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 239000004744 fabric Substances 0.000 description 1
- 150000004676 glycans Chemical class 0.000 description 1
- 125000000623 heterocyclic group Chemical group 0.000 description 1
- 150000002632 lipids Chemical class 0.000 description 1
- 229910052943 magnesium sulfate Inorganic materials 0.000 description 1
- 235000019341 magnesium sulphate Nutrition 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 239000002366 mineral element Substances 0.000 description 1
- 150000007524 organic acids Chemical class 0.000 description 1
- 230000032696 parturition Effects 0.000 description 1
- 150000002989 phenols Chemical class 0.000 description 1
- 239000000049 pigment Substances 0.000 description 1
- 229920001282 polysaccharide Polymers 0.000 description 1
- 239000005017 polysaccharide Substances 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 238000000194 supercritical-fluid extraction Methods 0.000 description 1
- 150000003505 terpenes Chemical class 0.000 description 1
- 235000007586 terpenes Nutrition 0.000 description 1
- 231100000331 toxic Toxicity 0.000 description 1
- 230000002588 toxic effect Effects 0.000 description 1
Classifications
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
- A61K36/18—Magnoliophyta (angiosperms)
- A61K36/185—Magnoliopsida (dicotyledons)
- A61K36/81—Solanaceae (Potato family), e.g. tobacco, nightshade, tomato, belladonna, capsicum or jimsonweed
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
- A61P31/04—Antibacterial agents
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
- A61P31/10—Antimycotics
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/10—Preparation or pretreatment of starting material
- A61K2236/15—Preparation or pretreatment of starting material involving mechanical treatment, e.g. chopping up, cutting or grinding
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/30—Extraction of the material
- A61K2236/33—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones
- A61K2236/331—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones using water, e.g. cold water, infusion, tea, steam distillation or decoction
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/30—Extraction of the material
- A61K2236/33—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones
- A61K2236/333—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones using mixed solvents, e.g. 70% EtOH
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/30—Extraction of the material
- A61K2236/39—Complex extraction schemes, e.g. fractionation or repeated extraction steps
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- Health & Medical Sciences (AREA)
- Natural Medicines & Medicinal Plants (AREA)
- Life Sciences & Earth Sciences (AREA)
- Pharmacology & Pharmacy (AREA)
- Chemical & Material Sciences (AREA)
- Veterinary Medicine (AREA)
- Public Health (AREA)
- General Health & Medical Sciences (AREA)
- Animal Behavior & Ethology (AREA)
- Medicinal Chemistry (AREA)
- Communicable Diseases (AREA)
- Oncology (AREA)
- Organic Chemistry (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- General Chemical & Material Sciences (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Medical Informatics (AREA)
- Alternative & Traditional Medicine (AREA)
- Engineering & Computer Science (AREA)
- Biotechnology (AREA)
- Botany (AREA)
- Mycology (AREA)
- Epidemiology (AREA)
- Microbiology (AREA)
- Agricultural Chemicals And Associated Chemicals (AREA)
Abstract
The present invention provides a kind of tobacco active component (TEP), extracting method and its applications, belong to tobacco product processing extractive technique field.The present invention is by pre-processing tobacco material, and for pretreated tobacco material successively carry out room temperature extraction, high temperature extract, ethyl alcohol extraction operate three times, and use highly polar solvent purification water, ethyl alcohol as Extraction solvent, by using different solvents various extracting conditions, it can guarantee to realize the tobacco active component in tobacco material and sufficiently extract, improve the extraction content of tobacco active component.Experiment finds that the purpose component has antibacterial functions, therefore has antibacterials potentiality to be exploited, provides theoretical foundation for new drug development.
Description
Technical field
The present invention relates to tobacco products to process extractive technique field, and in particular to a kind of tobacco active component (TEP) is extracted
Method and its application.
Background technique
Tobacco is also referred to as mesona, is Chinese medicine traditional simply, its inorganic constituents includes water, mineral element；Organic principle
It mainly include carbohydrate, alkaloid, heterocyclic, pigment, phenols, terpene, organic acid, lipid, alcohols, aldoketones etc..Wherein,
Organic principle has antibacterial, antiviral and antitumor, antioxidant activity, removes the multiple functions such as interior free yl in tobacco.
Currently, generally using means of supercritical extraction, Soxhlet extraction, ultrasonic wave to realize the extraction to tobacco active component
Extracting mode.Using dehydrated alcohol, n-hexane, acetone, ethyl acetate, petroleum ether, methylene chloride as Extraction solvent.It is wherein low
Polar solvent such as petroleum ether, n-hexane, middle polar solvent acetone, methylene chloride, these solvents are mostly volatile, inflammable, toxic
Solvent requires height to the equipment of industrial product, personnel etc., and the content of obtained tobacco active component is few, this is just greatly limited
The further research and application of tobacco active component.
Summary of the invention
In view of the above-mentioned problems, the present invention provides a kind of tobacco active component (TEP), extracting method and its application, to improve
The extracted amount of tobacco active component.
In a first aspect, the present invention provides a kind of tobacco active component (TEP), extracting method, this method includes following step
It is rapid:
(1) it pre-processes: drying and processing being carried out to tobacco material, and the tobacco material progress tobacco after drying and processing is thick
It crushes, is finally pulverized again, obtain tobacco Ultramicro-powder；
(2) normal-temperature water mentions: Extraction solvent being added in tobacco Ultramicro-powder, extracts operation under normal temperature conditions；To mentioning
Tobacco Ultramicro-powder after extract operation carries out centrifugal treating, obtains tobacco normal-temperature water and mentions sediment and supernatant, by supernatant low temperature
It saves, freeze-drying obtains tobacco normal-temperature water and puts on clear liquid freeze-dried powder；Extraction solvent in step (2) is purified water；
(3) high-temperature water mentions: Extraction solvent is added in the sediment collected in step (2), in 90-100 DEG C of extraction temperature
Degree is lower to carry out concentration extraction operation；Cooling processing, centrifugal treating are carried out to the concentrated extracting solution obtained after concentration extraction operation, and
Sediment after collecting centrifugal treating；Extraction solvent in step (3) is purified water；
(4) alcoholic extraction: Extraction solvent is added in the supernatant obtained in step (3), under 0-4 DEG C of Extracting temperature
Extraction extraction operation is carried out, centrifugal treating is carried out to Soakage extraction object, and collect the precipitating after centrifugal treating, by the precipitating of collection
Drying and processing is carried out, tobacco active component tobacco polysaccharide extract TEP is obtained；The ethyl alcohol that Extraction solvent in step (4) is 85%.
Preferably, the ultramicro crushing treatment in step (1) is that the tobacco material after drying and processing is used 50-80 mesh
The pulverizer of net carries out pulverization process, and to the tobacco material after pulverization process, is carried out using the pulverizer of 200-400 mesh screen
Pulverization process obtains the tobacco Ultramicro-powder.
Preferably, the middle quality that Extraction solvent is added of step (2) is 2-4 times of the quality of the tobacco Ultramicro-powder, is extracted
Time is 0.5-3h.
Preferably, the centrifugal treating in step (2) refers to: centrifuging temperature is 8-12 DEG C, centrifugation time 0.5-1.5h, is turned
Speed is 3000-6000rpm/min.
Preferably, the middle quality that Extraction solvent is added of step (3) is 1.2-2.4 times of the quality of sediment.
Preferably, the centrifugal treating in step (3) refer to will cooling treated that the concentrated extracting solution is placed in centrifugal bottle
In, centrifuging temperature is 8-12 DEG C, centrifugation time 0.5-1.5h, revolving speed 3000-6000rpm/min.
Preferably, the middle quality that Extraction solvent is added of step (4) is 1.5-2.5 times of the quality of sediment.
Preferably, the Soakage extraction operation in step (4) refers to that low temperature alcohol precipitation is stayed overnight.
Preferably, the centrifugal treating in step (4), which refers to, is placed in the Soakage extraction object in centrifugal bottle, centrifuging temperature
Are as follows: 3-5 DEG C, centrifugation time 0.5-1.5h, revolving speed is 3000-6000 rpm/min.
Second aspect, the present invention provides a kind of applications according to the tobacco active component in preparation antibacterials.
The present invention have it is following the utility model has the advantages that
1, the present invention is by pre-processing tobacco material, and carries out room temperature for pretreated tobacco material and mention
It takes, use highly polar solvent purification water as Extraction solvent, the tobacco active component in tobacco material can be realized and sufficiently be mentioned
It takes, improves the extraction content of tobacco active component；
2, Extraction solvent be ethyl alcohol, aqueous solution, it is low in cost；
3, the present invention can be used as antibacterials exploitation by the extract that specific process extracts to tobacco material, have wide
Application prospect.Meanwhile the preparation method of tobacco extract of the invention is simple, is suitble to industrialized production.
Detailed description of the invention
Shown in FIG. 1 is the flow chart of tobacco active component (TEP) extracting method that the embodiment of the present invention 1 obtains；
Shown in Fig. 2 is that the obtained tobacco active component (TEP) of the embodiment of the present invention 2 inhibits fungi-Cordyceps Militaris growth
Schematic diagram (inhibition zone that black arrow meaning generates in figure for tobacco active component)；
Shown in Fig. 3 is that the obtained tobacco active component (TEP) of the embodiment of the present invention 3 inhibits bacterium-Escherichia coli raw
Long schematic diagram (inhibition zone that black arrow meaning generates in figure for tobacco active component)；
Shown in Fig. 4 is that the obtained tobacco active component (TEP) of the embodiment of the present invention 4 inhibits bacterium-golden yellow grape
Coccus grows schematic diagram (inhibition zone that black arrow meaning generates in figure for tobacco active component)；
Shown in fig. 5 is that the obtained tobacco active component (TEP) of the embodiment of the present invention 5 inhibits bacterium-bacillus subtilis
Bacterium grows schematic diagram (inhibition zone that black arrow meaning generates in figure for tobacco active component)；
The above description is only an overview of the technical scheme of the present invention, in order to better understand the technical means of the present invention,
And can be implemented in accordance with the contents of the specification, below with presently preferred embodiments of the present invention.
Specific embodiment
The present invention is further described in detail with reference to embodiments.It is noted that detailed description below is all to illustrate
Property, it is intended to further instruction is provided to the present invention.Unless otherwise indicated, all scientific and technical terms that the present invention uses
With with the normally understood identical meanings of the technical field of the invention personnel.
Embodiment 1: the extraction of tobacco antibacterial active constituents
1, the extraction step of tobacco antibacterial active constituents is as follows:
(1) it pre-processes: drying and processing being carried out to tobacco material, and the tobacco material progress tobacco after drying and processing is thick
It crushes, is finally pulverized again, obtain tobacco Ultramicro-powder；
(2) normal-temperature water mentions: Extraction solvent being added in tobacco Ultramicro-powder, extracts operation under normal temperature conditions；To mentioning
Tobacco Ultramicro-powder after extract operation carries out centrifugal treating, obtains tobacco normal-temperature water and mentions sediment and supernatant, by supernatant low temperature
It saves, freeze-drying obtains tobacco normal-temperature water and puts on clear liquid freeze-dried powder；Extraction solvent in step (2) is purified water；
(3) high-temperature water mentions: Extraction solvent is added in the sediment collected in step (2), in 90-100 DEG C of extraction temperature
Degree is lower to carry out concentration extraction operation；Cooling processing, centrifugal treating are carried out to the concentrated extracting solution obtained after concentration extraction operation, and
Sediment after collecting centrifugal treating；Extraction solvent in step (3) is purified water；
(4) alcoholic extraction: Extraction solvent is added in the supernatant obtained in step (3), under 0-4 DEG C of Extracting temperature
Extraction extraction operation is carried out, centrifugal treating is carried out to Soakage extraction object, and collect the precipitating after centrifugal treating, is dried to obtain tobacco
Active component；The alcohol that Extraction solvent in step (4) is 50-100%.
Embodiment 2: tobacco active component inhibits fungi-Cordyceps Militaris growth experiment
1, experiment purpose:
By bacteriostatic experiment, the inhibitory effect of tobacco extract can be measured.
2, experimental method:
Agar punch method
3, experimental principle:
It is raw to influence bacterium from hole to agar medium scattering and permeating and the inhibitory or killing effect on test organisms for antibacterial material
Long breeding, makes bacterium colony form inhibition zone, the power of antibacterial material antibacterial ability is determined according to the size of inhibition zone.This test is suitable for
The identification of bacteriostatic agent and the antibacterial product of stripping property.This test is suitable for the identification of bacteriostatic agent and the antibacterial product of stripping property.
4, experimental strain:
Cordyceps Militaris
5, experimental procedure:
(1) solid medium: yeast extract 0.2g, peptone 0.5g, glucose 0.2g, dipotassium hydrogen phosphate is configured
In 100mL, high pressure sterilization falls flat when temperature is suitable in superclean bench for 0.1g, magnesium sulfate 0.05g, agar 1.5g constant volume
Plate uses after solidification；
(2) Cordyceps Militaris is inoculated with: Cordyceps Militaris being inoculated into solid medium culture dish, is coated with uniformly, is covered with spreading rod
Culture dish；
(3) agar punches: making a call to three holes with 1mL pipette tips on the culture dish for be coated with bacterium solution；
(4) it adds tobacco active component: having beaten hole and chosen the partial medium that pipette tips in hole are not taken out of with sterile medicine Chinese herbaceous peony,
Tobacco active component is diluted with pure water, concentration 0.2g/mL pipettes 200 μ L dilutions to the three of bacterium solution culture dish with liquid-transfering gun
In a hole, culture dish carries out Cordyceps Militaris label and experiment date；
(5) it cultivates: culture dish being put into incubator 27 DEG C, is cultivated 48-96 hours；
(6) experimental result is as shown in Figure 2: using the culture dish seen after piece machine checks culture, not giving birth to around hole
Strain is grown, therefore shows that tobacco active component is inhibited to Cordyceps Militaris.
Embodiment 3: tobacco active component inhibits the experiment of bacterium-Escherichia coli Growth
1, experiment purpose:
By bacteriostatic experiment, the inhibitory effect of tobacco extract can be measured.
2, experimental method:
Agar punch method
3, experimental principle:
It is raw to influence bacterium from hole to agar medium scattering and permeating and the inhibitory or killing effect on test organisms for antibacterial material
Long breeding, makes bacterium colony form inhibition zone, the power of antibacterial material antibacterial ability is determined according to the size of inhibition zone.This test is suitable for
The identification of bacteriostatic agent and the antibacterial product of stripping property.
4, experimental strain:
Escherichia coli
5, experimental procedure:
(1) solid medium: yeast extract 0.2g, peptone 0.5g, glucose 0.2g, agar 1.5g constant volume is configured
In 100mL, high pressure sterilization, when temperature is suitable, the inverted plate in superclean bench uses after solidification.
(2) Escherichia coli are inoculated with: Escherichia coli are inoculated into solid medium culture dish, are coated with uniformly with spreading rod,
Cover culture dish.
(3) agar punches: making a call to three holes with 1mL pipette tips on the culture dish for be coated with bacterium solution.
(4) it adds tobacco active component: having beaten hole and chosen the partial medium that pipette tips in hole are not taken out of with sterile medicine Chinese herbaceous peony,
Tobacco active component is diluted with pure water, concentration 0.2g/mL pipettes 200L dilution to three of bacterium solution culture dish with liquid-transfering gun
Kong Zhong, culture dish carry out Escherichia coli label and experiment date.
(5) it cultivates: culture dish being put into incubator 37 DEG C, is cultivated 48-96 hours.
(6) experimental result is as shown in Figure 3: seeing the culture dish after piece machine checks culture, does not grow around hole
Strain, therefore show that tobacco active component is inhibited to Escherichia coli.
Embodiment 4: tobacco active component inhibits the experiment of bacterium-staphylococcus aureus growth
1, experiment purpose:
By bacteriostatic experiment, the inhibitory effect of tobacco extract can be measured.
2, experimental method:
Agar punch method
3, experimental principle:
It is raw to influence bacterium from hole to agar medium scattering and permeating and the inhibitory or killing effect on test organisms for antibacterial material
Long breeding, makes bacterium colony form inhibition zone, the power of antibacterial material antibacterial ability is determined according to the size of inhibition zone.This test is suitable for
The identification of bacteriostatic agent and the antibacterial product of stripping property.This test is suitable for the identification of bacteriostatic agent and the antibacterial product of stripping property.
4, experimental strain:
Staphylococcus aureus
5, experimental procedure:
(1) solid medium: yeast extract 0.2g, peptone 0.5g, glucose 0.2g, agar 1.5g constant volume is configured
In 100mL, high pressure sterilization, when temperature is suitable, the inverted plate in superclean bench uses after solidification；
(2) S. aureus Inoculate: by S. aureus Inoculate to solid medium culture dish, with coating
Stick coating uniformly, covers culture dish；
(3) agar punches: making a call to three holes with 1mL pipette tips on the culture dish for be coated with bacterium solution；
(4) it adds tobacco active component: having beaten hole and chosen the partial medium that pipette tips in hole are not taken out of with sterile medicine Chinese herbaceous peony,
Tobacco active component is diluted with pure water, concentration 0.2g/mL pipettes 200L dilution to three of bacterium solution culture dish with liquid-transfering gun
Kong Zhong, culture dish carry out staphylococcus aureus label and experiment date；
(5) it cultivates: culture dish being put into incubator 37 DEG C, is cultivated 48-96 hours；
(6) experimental result is as shown in Figure 4: seeing the culture dish after piece machine checks culture, does not grow around hole
Strain, therefore show that tobacco active component is inhibited to staphylococcus aureus.
Embodiment 5: tobacco active component inhibits bacterium-bacillus subtilis growth experiment
1, experiment purpose:
By bacteriostatic experiment, the inhibitory effect of tobacco extract can be measured.
2, experimental method:
Agar punch method
3, experimental principle:
It is raw to influence bacterium from hole to agar medium scattering and permeating and the inhibitory or killing effect on test organisms for antibacterial material
Long breeding, makes bacterium colony form inhibition zone, the power of antibacterial material antibacterial ability is determined according to the size of inhibition zone.This test is suitable for
The identification of bacteriostatic agent and the antibacterial product of stripping property.This test is suitable for the identification of bacteriostatic agent and the antibacterial product of stripping property.
4, experimental strain:
Bacillus subtilis
(1) solid medium: yeast extract 0.2g, peptone 0.5g, glucose 0.2g, agar 1.5g constant volume is configured
In 100mL, high pressure sterilization, when temperature is suitable, the inverted plate in superclean bench uses after solidification；
(2) bacillus subtilis is inoculated with: bacillus subtilis being inoculated into solid medium culture dish, is applied with spreading rod
Cloth is uniform, covers culture dish；
(3) agar punches: making a call to three holes with 1mL pipette tips on the culture dish for be coated with bacterium solution；
(4) it adds tobacco active component: having beaten hole and chosen the partial medium that pipette tips in hole are not taken out of with sterile medicine Chinese herbaceous peony,
Tobacco active component is diluted with pure water, concentration 0.2g/mL pipettes 200mL dilution to the three of bacterium solution culture dish with liquid-transfering gun
In a hole, culture dish carries out bacillus subtilis label and experiment date；
(5) it cultivates: culture dish being put into incubator 37 DEG C, is cultivated 48-96 hours；
(6) experimental result is as shown in Figure 5: seeing the culture dish after piece machine checks culture, does not grow around hole
Strain, therefore show that tobacco active component is inhibited to bacillus subtilis.
The above is merely a preferred embodiment of the present invention, it is noted that for those skilled in the art
For, under the premise of not departing from core of the invention technology, improvements and modifications can also be made, these improvements and modifications are also answered
Belong to scope of patent protection of the invention, and any change in the comparable meaning and scope of claims of the present invention, all
It is considered as including in Claims scope.
Claims (10)
1. a kind of tobacco active component (TEP), extracting method, which comprises the following steps:
(1) it pre-processes: drying and processing being carried out to tobacco material, and the tobacco material after drying and processing is subjected to tobacco coarse powder
It is broken, it is finally pulverized again, obtains tobacco Ultramicro-powder；
(2) normal-temperature water mentions: Extraction solvent being added in tobacco Ultramicro-powder, extracts operation under normal temperature conditions；It is grasped to extracting
Tobacco Ultramicro-powder after work carries out centrifugal treating, obtains tobacco normal-temperature water and mentions sediment and supernatant, by supernatant cryo-conservation,
Freeze-drying, obtains tobacco normal-temperature water and puts on clear liquid freeze-dried powder；Extraction solvent in step (2) is purified water；
(3) high-temperature water mentions: Extraction solvent is added in the sediment collected in step (2), under 90-100 DEG C of Extracting temperature
Carry out concentration extraction operation；Cooling processing, centrifugal treating are carried out to the concentrated extracting solution obtained after concentration extraction operation, and collected
Sediment after centrifugal treating；Extraction solvent in step (3) is purified water；
(4) alcoholic extraction: Extraction solvent is added in the supernatant obtained in step (3), is carried out under 0-4 DEG C of Extracting temperature
Extraction operation is extracted, centrifugal treating is carried out to Soakage extraction object, and collect the precipitating after centrifugal treating, is dried to obtain tobacco activity
Component；The alcohol that Extraction solvent in step (4) is 50-100%.
2. the method according to claim 1 for extracting antibacterial active constituents from tobacco, it is characterised in that: the step
(1) ultramicro crushing treatment in is to carry out the tobacco material after drying and processing at crushing using the pulverizer of 50-80 mesh screen
Reason, and to the tobacco material after pulverization process, pulverization process is carried out using the pulverizer of 200-400 mesh screen, obtains the cigarette
Careless Ultramicro-powder.
3. the method according to claim 1 for extracting antibacterial active constituents from tobacco, it is characterised in that: the step
(2) quality of addition Extraction solvent is 2-4 times of the quality of the tobacco Ultramicro-powder, extraction time 0.5-3h in.
4. the method according to claim 1 for extracting antibacterial active constituents from tobacco, it is characterised in that: the step
(2) centrifugal treating in refers to: centrifuging temperature is 8-12 DEG C, centrifugation time 0.5-1.5h, revolving speed 3000-6000rpm/
min。
5. the method according to claim 1 for extracting antibacterial active constituents from tobacco, it is characterised in that: the step
(3) quality of addition Extraction solvent is 1.2-2.4 times of the quality of sediment in.
6. the method according to claim 1 for extracting antibacterial active constituents from tobacco, it is characterised in that: the step
(3) centrifugal treating in refer to will cooling treated that the concentrated extracting solution is placed in centrifugal bottle, centrifuging temperature is 8-12 DEG C,
Centrifugation time is 0.5-1.5h, revolving speed 3000-6000rpm/min.
7. the method according to claim 1 for extracting antibacterial active constituents from tobacco, it is characterised in that: the step
(4) quality of addition Extraction solvent is 1.5-2.5 times of the quality of sediment in.
8. the method according to claim 1 for extracting antibacterial active constituents from tobacco, it is characterised in that: the step
(4) the Soakage extraction operation in refers to that low temperature alcohol precipitation is stayed overnight.
9. the method according to claim 1 for extracting antibacterial active constituents from tobacco, it is characterised in that: the step
(4) centrifugal treating in, which refers to, is placed in the Soakage extraction object in centrifugal bottle, centrifuging temperature are as follows: 3-5 DEG C, centrifugation time is
0.5-1.5h, revolving speed 3000-6000rpm/min.
10. application of the tobacco active component in preparation antibacterials according to claim 1.
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| CN110050798A (en) * | 2019-03-06 | 2019-07-26 | 贵州省烟草科学研究院 | Tobacco active component (TMS), extracting method and its application |
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| CN110946798A (en) * | 2019-12-24 | 2020-04-03 | 贵州贵安精准医学研究院股份有限公司 | Development and application of natural skin health-care powder |
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