CN102742604B - Application of fungus fimetariella rabenhorstii A20 in inducing aquilaria sinensis to generate guaiol - Google Patents
Application of fungus fimetariella rabenhorstii A20 in inducing aquilaria sinensis to generate guaiol Download PDFInfo
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- CN102742604B CN102742604B CN201210167198.3A CN201210167198A CN102742604B CN 102742604 B CN102742604 B CN 102742604B CN 201210167198 A CN201210167198 A CN 201210167198A CN 102742604 B CN102742604 B CN 102742604B
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Abstract
The invention discloses an application of a fungus F.rabenhorstii A20 in inducing aquilaria sinensis to generate guaiol. The fungus F.rabenhorstii A20 provided by the invention can induce aquilaria sinensis to generate guaiol. The invention has important theoretical and practical significance in analyzing Chinese eaglewood formation mechanism, and in researching artificial formation technologies.
Description
Technical field
The invention belongs to biotechnology and agriculture field, be specifically related to fungi (Fimetariella rabenhorstii) A20 and produce the application in agalloch eaglewood constituent guaiol at induction suspension culture of Aquilaria sinensis (Aquilaria sinensis (Lour.) Gilg).
Background technology:
Agalloch eaglewood is a kind of rare Chinese medicine, and its medicinal beginning is loaded in " Mingyi Bielu ", classifies as top grade.Its taste is pungent, bitter, slightly warm in nature, and tool promoting qi circulation and relieving pain, warming middle energizer to arrest vomiting, promoting inspiration to relieve asthma effect, be used for the treatment of the vexed pain of chest abdominal distension, gastrofrigid vomiting hiccup, the circulation of vital energy in the wrong direction of suffering from a deficiency of the kidney and breathe heavily the diseases (NF, 2010) such as anxious.In addition it or a kind of and tradition extensive use famous and precious natural perfume material very popular in Asia.The chemical composition of agalloch eaglewood mainly comprises sesquiterpenoids and 2-(2-phenethyl) chromone class, and wherein sesquiterpenoids has (Naef R, 2011) such as guaiol (its structural formula is suc as formula shown in I), agalloch eaglewood spiral alcohol, baimuxinic acid, baimuxinals.Agalloch eaglewood is a kind of black resin that suspension culture of Aquilaria sinensis trunk is secreted in the situation that sustaining damage or stimulate, suspension culture of Aquilaria sinensis (Aquilaria sinensis (Lour.) Gilg)) claim again buta-buta, belong to Thymelaeceae eaglewood, unique plant origin for the distinctive torrid zone of China and subtropical evergreen arbor ，Shi China production agalloch eaglewood.At present, suspension culture of Aquilaria sinensis forms concrete reason and the process unclear of agalloch eaglewood.
Much studies show that, eaglewood Edgeworthia chrysantha is relevant to fungi.From the thirties in 20th century, the fungi being separated to from eaglewood Edgeworthia chrysantha position has (Zhang Zheng etc., 2010) such as grape seat chamber bacterium (Botryosphaeria rhodina), look two spore bacterium (Diplodia sp.), sickle-like bacteria (Fusarium sp.), aspergillus (Aspergillus sp.), Mucor (Mucor sp.), mould (Penicillium sp.), wood mould (Trichoderma sp.), schizophyllum communes (Schizophyllum sp.).At present, more existing fungies can induce suspension culture of Aquilaria sinensis to produce the report of agalloch eaglewood.In patent " method of using fungi to elicit Aquilariasinensis tree to produce agilawood ", relate to fungi Fusarium Fusarium sp.A-11 and cephalosporium Cephalosporium sp.A-17 and can induce the secretion of suspension culture of Aquilaria sinensis trunk to produce filemot agalloch eaglewood.Article " two batches of bacterination process produce gas chromatography-mass spectrometry analysis of agalloch eaglewood chemical composition of volatile oil " (Lin Feng etc., 2010) find, suspension culture of Aquilaria sinensis trunk inoculation Menanotus flavoliven M.flavolivens, after half a year, 1 year, has sesquiterpenoids, aromatic compound and fatty acid in extracted by ether volatile oil.In patent " a kind of Tambac inducer and preparation method thereof ", provide the preparation method of agalloch eaglewood inducible strain Lasiodiplodia theobromae Lasiodiplodia theobromae as Tambac inducer.
Summary of the invention:
First object of the present invention is to provide fungi F.rabenhorstii A20 and produces the application in guaiol at induction suspension culture of Aquilaria sinensis.
The present invention found through experiments, inoculated fungi F.rabenhorstii A20 on suspension culture of Aquilaria sinensis (Aquilaria sinensis (Lour.) Gilg) trunk, after cultivating after a while, again sample is carried out pre-treatment and carries out GC-MS analysis, find that fungi F.rabenhorstii A20 can induce suspension culture of Aquilaria sinensis to produce agalloch eaglewood component guaiol.
Therefore, second object of the present invention is to provide a kind of method of inducing suspension culture of Aquilaria sinensis to produce guaiol, it is characterized in that, fungi Fimetariella rabenhorstii A20 is inoculated on suspension culture of Aquilaria sinensis trunk, after cultivating, suspension culture of Aquilaria sinensis can produce guaiol.
In the suspension culture of Aquilaria sinensis that described Xinyi City, fungi Fimetariella rabenhorstii A20Cong Guangdong Province gathers, separation obtains, through ITS sequence analysis contrast, be accredited as F.rabenhorstii, gene login sequence number is EU781677, and culture presevation, in Guangdong institute of microbiology, is numbered A20.
Fungi F.rabenhorstii A20 of the present invention can induce suspension culture of Aquilaria sinensis to produce guaiol, and this forms mechanism to dissecting agalloch eaglewood, and studying artificial Edgeworthia chrysantha technology has most important theories and practice significance.
Fungi F.rabenhorstii A20 of the present invention belongs to bacterial strain in prior art, it is disclosed in scientific and technical literature: Tao Meihua, Li Dongli, Zhang Weimin, Tan Jianwen, Wei Xiaoyi, white banksia rose endogenetic epiphyte Fimetariella rabenhorstii chemical constitution study, traditional Chinese medicine, 2011,34(02), 221～223. this bacterial strain the applicant also hold, and guarantee in 20 years, to the public, to provide from the applying date.
Accompanying drawing explanation:
Fig. 1 is that after acting on 20 days, bacterial strain F.rabenhorstii A20 is inoculated into the experimental group of suspension culture of Aquilaria sinensis branch and the total ion current figure of two control groups, and wherein I is the experimental group that bacterial strain F.rabenhorstii A20 is inoculated into suspension culture of Aquilaria sinensis branch; II for cultivating the bacterial strain F.rabenhorstii A20 control group of 7d in PDA culture dish; III is for not meeting fungi F.rabenhorstii A20, the fresh white banksia rose branch control group of same treatment;
Fig. 2 is that after acting on 30 days, bacterial strain F.rabenhorstii A20 is inoculated into the experimental group of suspension culture of Aquilaria sinensis branch and the total ion current figure of two control groups, and wherein I is the experimental group that bacterial strain F.rabenhorstii A20 is inoculated into suspension culture of Aquilaria sinensis branch; II for cultivating the bacterial strain F.rabenhorstii A20 control group of 7d in PDA culture dish; III is for not meeting fungi F.rabenhorstii A20, the fresh white banksia rose branch control group of same treatment;
Fig. 3 is mass spectrum comparison chart, and wherein I is guaiol in NIST05 mass spectral database; II is that in embodiment 2 and 1, retention time is the sample at 21.15,21.20min place.
Embodiment:
Following examples are to further illustrate of the present invention, rather than limitation of the present invention.
Embodiment 1:
The endogenetic fungus F.rabenhorstii A20 being obtained by the separation of suspension culture of Aquilaria sinensis root is preserved in 4 ℃ of low temperature refrigerators, and the slant strains of a small amount of preservation of picking is inoculated on autoclaved PDA plate medium, cultivates 7d for 28 ℃ and carries out actication of culture.The fresh branch of the thick about 0.5-1cm of suspension culture of Aquilaria sinensis is cut into the segment of about 8cm left and right, first in 0.1% mercuric chloride solution, soak 5min, after taking out, use rinsed with sterile water 3 times, put into again volume fraction 75% ethanol water and soak 5min, rinsed with sterile water 3 times, then punches on suspension culture of Aquilaria sinensis branch with sterilized card punch.Suspension culture of Aquilaria sinensis branch after punching is put on the flat board of moistening aseptic filter paper, by 1ml aseptic water washing fungi F.rabenhorstii A20 slant strains, large head straw is drawn bacterium liquid and is injected in suspension culture of Aquilaria sinensis branch Shang hole, and inoculation rear plate is with preservative film sealing and place 20d in dark, 27 ℃ of environment.Suspension culture of Aquilaria sinensis branch after cultivating is put into triangular flask, add volume fraction 95% ethanol water soaked overnight, extract Rotary Evaporators is evaporated to dry, after dissolving, crosses 0.22 μ m miillpore filter with 1.5ml chloroform, then filtrate is carried out to gas chromatograph-mass spectrometer analysis.Wherein GC conditions is Agilent capillary chromatographic column DB-5MS(30m * 0.25mm * 0.25 μ m); Chromatographic column temperature programming condition: 60 ℃ of initial column temperatures (keeping 2min), rise to 150 ℃ with 5 ℃/min, then rise to 160 ℃ (keeping 15min) with 1 ℃/min, then rise to 250 ℃ (keeping 20min) with 8 ℃/min; 250 ℃ of transmission line temperature, carrier gas is high-purity helium, flow velocity 1.0ml/min; Sample size 1 μ l, temperature programming vaporization Splitless injecting samples, 60 ℃ of initial temperatures, 14.5 ℃/min is warming up to 250 ℃.Mass spectrum condition is electronics bombardment (EI) ion gun, 230 ℃ of ion source temperatures; Quality of scanning scope (m/z) 40～500.Each experiment component adopts the retrieval of NIST05 mass spectral database to carry out qualitative.Arrange and do not meet fungi F.rabenhorstii A20 simultaneously, the fresh white banksia rose branch control group of same treatment and cultivate the bacterial strain F.rabenhorstii A20 control group of 7d in PDA culture dish, each experimental group arrange 3 parallel, the material extraction of culture is identical with experimental group with analysis.Result as shown in Figure 1, from Fig. 1, can find, the experimental group that bacterial strain F.rabenhorstii A20 is inoculated into suspension culture of Aquilaria sinensis branch is that 21.20min place can detect agalloch eaglewood constituent guaiol (retention time is that in the mass spectrogram of 21.20min cut and NIST05 mass spectral database guaiol mass spectrum comparison chart is as shown in Figure 3 in retention time, by Fig. 3, illustrated, retention time is that the cut of 21.20min is guaiol), and do not meet fungi F.rabenhorstii A20, the fresh white banksia rose branch control group of same treatment and the bacterial strain F.rabenhorstii A20 control group of cultivating 7d in PDA culture dish are finds this component, illustrate thus fungi F.rabenhorstii A20 is connect to bacterium to suspension culture of Aquilaria sinensis branch, can induce suspension culture of Aquilaria sinensis to produce agalloch eaglewood constituent guaiol.
Embodiment 2
The endogenetic fungus F.rabenhorstii A20 being obtained by the separation of suspension culture of Aquilaria sinensis root is preserved in 4 ℃ of low temperature refrigerators, and the slant strains of a small amount of preservation of picking is inoculated on autoclaved PDA plate medium, cultivates 5d for 28 ℃ and carries out actication of culture.The fresh branch of the thick about 0.5-1cm of suspension culture of Aquilaria sinensis is cut into the segment of about 8cm left and right, first in 0.1% mercuric chloride solution, soak 7min, after taking out, use rinsed with sterile water 4 times, put into again volume fraction 75% ethanol water and soak 7min, rinsed with sterile water 4 times, then punches on suspension culture of Aquilaria sinensis branch with sterilized card punch.Suspension culture of Aquilaria sinensis branch after punching is put on the flat board of moistening aseptic filter paper, by 1ml aseptic water washing fungi F.rabenhorstii A20 slant strains, large head straw is drawn bacterium liquid and is injected in suspension culture of Aquilaria sinensis branch Shang hole, and inoculation rear plate is with preservative film sealing and place 30d in dark, 27 ℃ of environment.The suspension culture of Aquilaria sinensis branch that covers with bacterium after cultivating is put into triangular flask, add volume fraction 95% ethanol water soaked overnight, extract Rotary Evaporators is evaporated to dry, after dissolving, crosses 0.22 μ m miillpore filter with 1.5ml chloroform, then carries out gas chromatograph-mass spectrometer analysis.Wherein GC conditions is Agilent capillary chromatographic column DB-5MS(30m * 0.25mm * 0.25 μ m); Chromatographic column temperature programming condition: 60 ℃ of initial column temperatures (keeping 2min), rise to 150 ℃ with 5 ℃/min, then rise to 160 ℃ (keeping 15min) with 1 ℃/min, then rise to 250 ℃ (keeping 20min) with 8 ℃/min; 250 ℃ of transmission line temperature, carrier gas is high-purity helium, flow velocity 1.0ml/min; Sample size 1 μ l, temperature programming vaporization Splitless injecting samples, 60 ℃ of initial temperatures, 14.5 ℃/min is warming up to 250 ℃.Mass spectrum condition is electronics bombardment (EI) ion gun, 230 ℃ of ion source temperatures; Quality of scanning scope (m/z) 40～500.Each experiment component adopts the retrieval of NIST05 mass spectral database to carry out qualitative.Arrange and do not meet fungi F.rabenhorstii A20 simultaneously, the fresh white banksia rose branch control group of same treatment and cultivate the bacterial strain F.rabenhorstii A20 control group of 7d in PDA culture dish, each experimental group arrange 3 parallel, the material extraction of culture is identical with experimental group with analysis.Result as shown in Figure 2, the experimental group that can find the bacterial strain F.rabenhorstii A20 to be inoculated into suspension culture of Aquilaria sinensis branch from Fig. 2 is that 21.15min can detect agalloch eaglewood constituent guaiol (retention time is that in the mass spectrogram of 21.15min cut and NIST05 mass spectral database guaiol mass spectrum comparison chart is as shown in Figure 3 in retention time, by Fig. 3, illustrated, retention time is that the cut of 21.15min is guaiol), and do not meet fungi F.rabenhorstii A20, the fresh white banksia rose branch control group of same treatment and the bacterial strain F.rabenhorstii A20 control group of cultivating 7d in PDA culture dish are finds this component, illustrate thus fungi F.rabenhorstii A20 is connect to bacterium to suspension culture of Aquilaria sinensis branch, can induce suspension culture of Aquilaria sinensis to produce agalloch eaglewood constituent guaiol.
Claims (2)
1. fungi Fimetariella rabenhorstii A20 produces the application in guaiol at induction suspension culture of Aquilaria sinensis Aquilaria sinensis (Lour.) Gilg, in the suspension culture of Aquilaria sinensis that described Xinyi City, fungi Fimetariella rabenhorstii A20Cong Guangdong Province gathers, separation obtains, through ITS sequence analysis contrast, be accredited as F.rabenhorstii, gene login sequence number is EU781677, culture presevation, in Guangdong institute of microbiology, is numbered A20.
2. a method of inducing suspension culture of Aquilaria sinensis to produce guaiol, it is characterized in that, fungi F.rabenhorstii A20 is inoculated on suspension culture of Aquilaria sinensis trunk, after cultivating, suspension culture of Aquilaria sinensis can produce guaiol, in the suspension culture of Aquilaria sinensis that described Xinyi City, fungi Fimetariella rabenhorstii A20Cong Guangdong Province gathers, separation obtains, through ITS sequence analysis contrast, be accredited as F.rabenhorstii, gene login sequence number is EU781677, culture presevation, in Guangdong institute of microbiology, is numbered A20.
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| CN104430555B (en) * | 2014-10-31 | 2017-05-31 | 北京全才中医医学研究院 | A kind of Edgeworthia chrysantha agent and its method for production agalloch eaglewood |
| CN107034252B (en) * | 2016-02-03 | 2020-09-11 | 北京中医药大学 | Method for inducing aquilaria sinensis callus cells to generate 2- (2-phenethyl) chromone components |
| CN106069499A (en) * | 2016-06-22 | 2016-11-09 | 云南爱尼沉香技术有限公司 | A kind of method improving Rhizoma Atractylodis macrocephalae's XIANGSHU Edgeworthia chrysantha Lindl. efficiency |
| CN106069256A (en) * | 2016-06-22 | 2016-11-09 | 云南爱尼沉香技术有限公司 | A kind of method of stopper strain induction Rhizoma Atractylodis macrocephalae's XIANGSHU Edgeworthia chrysantha Lindl. |
| CN116262902A (en) * | 2021-12-15 | 2023-06-16 | 中国中医科学院中药研究所 | Fungus capable of continuously inducing agilawood accumulation and application thereof |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2002094002A2 (en) * | 2001-05-24 | 2002-11-28 | Regents Of The University Of Minnesota | Cultivated agarwood |
| CN101781623A (en) * | 2009-01-15 | 2010-07-21 | 中国医学科学院药用植物研究所 | Method for using fungi to elicit Aquilariasinensis tree to produce agilawood |
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2002094002A2 (en) * | 2001-05-24 | 2002-11-28 | Regents Of The University Of Minnesota | Cultivated agarwood |
| CN101781623A (en) * | 2009-01-15 | 2010-07-21 | 中国医学科学院药用植物研究所 | Method for using fungi to elicit Aquilariasinensis tree to produce agilawood |
Non-Patent Citations (4)
| Title |
|---|
| 王磊 等.白木香内生真菌的分离及分子鉴定.《菌物研究》.2009,第7卷(第1期),37-42. |
| 白木香内生真菌Fimetariella rabenhorstii化学成分研究;陶美华 等;《中药材》;20110228;第34卷(第2期);221-223 * |
| 白木香内生真菌的分离及分子鉴定;王磊 等;《菌物研究》;20090331;第7卷(第1期);37-42 * |
| 陶美华 等.白木香内生真菌Fimetariella rabenhorstii化学成分研究.《中药材》.2011,第34卷(第2期),221-223. |
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