CN110295116A - One plant of endogenetic fungal bacterial strain for producing a variety of fatty acid and its application - Google Patents

One plant of endogenetic fungal bacterial strain for producing a variety of fatty acid and its application Download PDF

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CN110295116A
CN110295116A CN201910680319.6A CN201910680319A CN110295116A CN 110295116 A CN110295116 A CN 110295116A CN 201910680319 A CN201910680319 A CN 201910680319A CN 110295116 A CN110295116 A CN 110295116A
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aspergillus
acid
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microbial inoculum
grease
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CN110295116B (en
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尹静
肖佳雷
詹亚光
徐志强
李林夕
王宇
张玉琦
李影
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Northeast Forestry University
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    • C12P7/64—Fats; Fatty oils; Ester-type waxes; Higher fatty acids, i.e. having at least seven carbon atoms in an unbroken chain bound to a carboxyl group; Oxidised oils or fats
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    • C12P7/00—Preparation of oxygen-containing organic compounds
    • C12P7/64—Fats; Fatty oils; Ester-type waxes; Higher fatty acids, i.e. having at least seven carbon atoms in an unbroken chain bound to a carboxyl group; Oxidised oils or fats
    • C12P7/6409—Fatty acids
    • C12P7/6427—Polyunsaturated fatty acids [PUFA], i.e. having two or more double bonds in their backbone
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Abstract

The endogenetic fungal bacterial strain for producing a variety of fatty acid the invention discloses one plant and its application.Present invention firstly provides one plant of aspergillus, the aspergillus is aspergillus (Aspergillus sp.) C1T2-4, is CCTCC NO:M 2019393 in the deposit number of China typical culture collection center.Invention further provides the microbial inoculum comprising above-mentioned aspergillus and its applications.Aspergillus (Aspergillus sp.) C1T2-4 of the invention can generate 15 kinds of fatty acid, and there is efficient alpha-linolenic acid synthesis capability, it is separated from the grease that aspergillus (Aspergillus sp.) C1T2-4 is produced and obtains 7.15% alpha-linolenic acid.Aspergillus (Aspergillus sp.) C1T2-4 provides strain excellent material for the exploitation of the following health care product, cosmetics and industrial use, has important value.

Description

One plant of endogenetic fungal bacterial strain for producing a variety of fatty acid and its application
Technical field
The invention belongs to technical field of bioengineering, and in particular to one plant of endogenetic fungal bacterial strain for producing a variety of fatty acid and its Using.
Background technique
During long-term biological evolution, plant and endogenetic fungus have gradually formed good mutualism relationship, and one Various nutriments needed for aspect plant provides existence for endogenetic fungus, another aspect endogenetic fungus generate secondary metabolites and mention High plant photosynthetic rate improves plant resistance to promote vine growth and development.Fungi shows relatively strong in the ecosystem Two packing spaces sexual clorminance, it is abundanter than other microbe groups in terms of functional diversity and genetic diversity, and have compared with The ability of strong antibiont and abiotic stress.Therefore, fungal species diversity (Speeies under different growing environment is probed into Diversity) and functional diversity (Functional diversity) become numerous focus of attention important topic.
Microbial oil is a kind of biodiesel raw material of great development prospect, and the high fat content bacterial strain of high frequency zone is real The key of existing microbial oil recovery energy.Since finding oleaginous microorganism for the first time from Germany scientist, scientists are ground Study carefully and never stops.After the nineties, how the emphasis of a very long time is all being obtained on functional grease.Linolenic acid is that human body must The fatty acid needed, and it is present, and linolenic main source is in the vegetable oil such as linolenic oil.Therefore, to from plant endogenesis Endogenetic fungus carry out bio-oil production either obtain fatty acid needed by human (such as linolenic acid) still industrial use Biodiesel, all have important realistic meaning.
Summary of the invention
The technical problems to be solved by the invention are to provide one plant of endogenetic fungal bacterial strain, can generate a variety of fatty acid, especially It is high yield alpha-linolenic acid.
To solve the above problems, the aspergillus is aspergillus (Aspergillus sp.) the present invention provides one plant of aspergillus C1T2-4 is CCTCC NO:M 2019393 in the deposit number of China typical culture collection center.
The ITS sequence of above-mentioned aspergillus (Aspergillus sp.) C1T2-4 is as shown in sequence 1 in sequence table.
Microbial inoculum containing above-mentioned aspergillus (Aspergillus sp.) C1T2-4 is also within protection scope of the present invention.
In above-mentioned microbial inoculum, the microbial inoculum can be the microbial inoculum of Lipid-producing；The grease includes myristic acid, pentadecane acid, palm Sour, cis- palmitoleic acid, 17 carbonic acid, heptadecenoic acid, stearic acid, elaidic acid, octadecenoic acid (trans- 9), octadecene Acid (cis- 9), linoleic acid, alpha-linolenic acid, two hendecoic acids, two ficocerylic acids, tetracosa carbon acid.
In above-mentioned microbial inoculum, the active constituent of the microbial inoculum can be aspergillus (Aspergillus sp.) C1T2-4, aspergillus The culture of the metabolin of (Aspergillus sp.) C1T2-4 and/aspergillus (Aspergillus sp.) C1T2-4, it is described The active constituent of microbial inoculum can also contain other biological ingredient or/and abiotic component, the other active components ability of the microbial inoculum Field technique personnel can be actually needed according to the present invention and be determined.
In above-mentioned microbial inoculum, aspergillus (Aspergillus sp.) C1T2-4 with spore, mycelia or can contain spore And/or the form of the culture of mycelia exists.
In above-mentioned microbial inoculum, the culture of aspergillus (Aspergillus sp.) C1T2-4 is by aspergillus The substance in culture vessel that (Aspergillus sp.) C1T2-4 is cultivated in microbiological culture media；Such as fermentation liquid.
In above-mentioned microbial inoculum, the metabolin of aspergillus (Aspergillus sp.) C1T2-4 can be from aspergillus It is obtained in the culture of (Aspergillus sp.) C1T2-4.
In above-mentioned microbial inoculum, the microbial inoculum also contains carrier in addition to the active constituent.The carrier can be led for grease production Domain is common and is being biologically inert carrier.The carrier can be solid carrier or liquid-carrier；The solid carrier It can be mineral material, vegetable material or high-molecular compound；The mineral material can for clay, talcum, kaolin, montmorillonite, At least one of white carbon, zeolite, silica and diatomite；The vegetable material can in corn flour, bean powder and starch at least It is a kind of；The high-molecular compound can be polyvinyl alcohol and/or polyglycols；The liquid-carrier can be organic solvent or water；Institute Stating organic solvent can be decane and/or dodecane.
In above-mentioned microbial inoculum, the dosage form of the microbial inoculum can be a variety of dosage forms, as liquor, emulsion, suspending agent, pulvis, granule, Wettable powder or water dispersible granules.
As needed, surfactant (such as polysorbas20, Tween 80), adhesive, stabilization can be also added in the microbial inoculum Agent (such as antioxidant), pH adjusting agent.
Invention further provides above-mentioned aspergillus (Aspergillus sp.) C1T2-4, aspergillus The culture of (Aspergillus sp.) C1T2-4, the metabolin of above-mentioned aspergillus (Aspergillus sp.) C1T2-4 or on Microbial inoculum is stated following 1) -4) it is any in application:
1) application in production grease；
2) application in alpha-linolenic acid is being prepared；
3) application in health care product is being prepared；
4) application in cosmetics is being prepared.
5) application in preparation chemical industry product.
The present invention also provides a kind of methods for producing grease.
The method that the present invention produces grease includes collecting culture with the above-mentioned aspergillus of microbiological culture media culture, from described Grease is obtained in culture.
In the above method, the grease include myristic acid, pentadecane acid, palmitinic acid, cis- palmitoleic acid, 17 carbonic acid, Heptadecenoic acid, stearic acid, elaidic acid, octadecenoic acid (trans- 9), octadecenoic acid (cis- 9), linoleic acid, alpha-linolenic acid, two Hendecoic acid, two ficocerylic acids and/or tetracosa carbon acid.
In the above method, the culture is by aspergillus (Aspergillus sp.) C1T2-4 in microbiological culture media The middle substance cultivated in obtained culture vessel；Such as fermentation liquid.
In the above method, the microbiological culture media can be the PDB culture medium after PDB culture medium or optimization.
In the above method, the PDB culture medium after optimization can be that the glucose in the PDB culture medium is replaced with other Carbon source such as starch, sucrose and/or inositol；It can also be to add nitrogen source in the PDB culture medium, the nitrogen source is tryptose Peptone, yeast extract, ammonium nitrate and/or acid hydrolyzed casein；Salt can also be added in the PDB culture medium, the salt is dense Degree is 0-9g/100ml；It can also be above-mentioned nitrogen source and salt in the PDB culture medium.
The carbon source of the i.e. described microbiological culture media is starch, glucose, sucrose or inositol, the nitrogen of the microbiological culture media Source is tryptone, yeast extract, ammonium nitrate or acid hydrolyzed casein, and the salinity of the microbiological culture media is 0-9g/ 100ml。
In the above method, the pH value of the microbiological culture media is 3-9.
In the above method, the time of the culture is 10-12 days；Specific is 10 days.
In the above method, more specifically, the carbon source of the microbiological culture media is glucose, and nitrogen source is sour water solution junket egg It is white, salinity 6g/100ml, pH 3.
Invention further provides a kind of methods for producing alpha-linolenic acid.
A method of producing alpha-linolenic acid, comprising the following steps: the grease for obtaining the method for above-mentioned production grease into Row isolates and purifies, and obtains alpha-linolenic acid.
The present invention has obtained 3 plant heights by screening layer by layer from 112 plants of endogenetic fungus and has produced oil-producing endogenetic fungus, selects aspergillus Belong to C1T2-4 and carried out preservation and analysis, shows that aspergillus (Aspergillus sp.) C1T2-4 can produce rich and varied rouge Fat acid (15 kinds), and aspergillus (Aspergillus sp.) C1T2-4 with alpha-linolenic acid synthesis capability is (raw from the bacterial strain Separated in the grease of production and obtain 7.15% alpha-linolenic acid), the exploitation for following health care product, cosmetics and chemical industry product provides Strain excellent material has important value.
Biomaterial information
Strain name: aspergillus (Aspergillus sp.) C1T2-4
Preservation mechanism: China typical culture collection center
Preservation mechanism abbreviation: CCTCC
Deposit number: CCTCC NO:M 2019393
Address: No. 299 Wuhan Universitys of Wuhan City, Hubei Province Wuchang District Bayi Road Wuhan University's collection in the school
Detailed description of the invention
Fig. 1 is that endogenetic fungus adds Nile red and is not added with comparison of the bacterium colony of Nile red under ultraviolet；Wherein, A is not The control group plate of Nile red is added, B is the experimental group plate for adding Nile red.
Fig. 2 is the colony morphology characteristic of endogenetic fungus C1T2-4.
Fig. 3 is the microstructure morphological feature of endogenetic fungus C1T2-4.
Fig. 4 is oil-producing bacterial strain C1T2-4 oil productivity and oil production and biomass time graph.
The fatty acid profile that Fig. 5 is oil-producing bacterial strain C1T2-4 is infused.
Specific embodiment
Embodiment below facilitates a better understanding of the present invention, but does not limit the present invention.Used in following embodiments Experimental method unless otherwise specified, be conventional method.The materials, reagents and the like used in the following examples, such as without special theory It is bright, it is commercially available.
The culture medium used in following embodiments and preparation method thereof is as follows:
PDA (abbreviation of potato dextrose agar, i.e. Potato Dextrose Agar) culture medium
It include potato 200g, glucose 20g, agar 15-20g in the culture medium of 1000ml, remaining is water.
The preparation method comprises the following steps: potato cleans peeling, weighs 200g and be cut into small pieces, add boiling is rotten (to boil 20-30 minutes, energy Poked by glass bar), it with eight layers of filtered through gauze, then needs according to actual experiment plus 15-20g agar, it is mixed to continue heating stirring It is even, after agar has dissolved, glucose 20g is added, stirs evenly, supplies moisture again after slightly cooling down to 1000 milliliters, packing tries Pipe or conical flask, jump a queue, wrap up, and 115 DEG C sterilize 20 minutes, store after cooling spare.
PDB culture medium: the fluid nutrient medium of PDA culture medium removal agar.
Embodiment 1, the separation screening of aspergillus C1T2-4 and identification
One, the separation screening of endophyte
Chinese tamarisk branch is acquired, with tap water continuous flushing capturing material, carries out explant disinfection and endogenetic fungus separation training Support, specific method with reference to Gao Jian (Gao Jian mangrove endophytic fungus diversity and its ecologicaI distribution [D] Guangdong Ocean University, 2013.).Using Zhejiang University Yuan Zhilin (at the beginning of Yuan Zhilin oryza meyeriana endogenetic fungus excavating resource, Phylogenetic Analysis and function Visit [D] Zhejiang University, 2010.) method carry out endogenetic fungus separation and culture, obtain endogenetic fungus C1T2-4.The present invention Inventor other 111 plants of endogenetic fungus have also been separated with the branch of white birch, blade, bark from soybean, Chinese tamarisk branch, bark. Following oil-producing screening is carried out to this 112 plants of endogenetic fungus:
1, Nile red flat band method primary dcreening operation oil-producing bacterial strain
Nile red is added after sterilizing with PDA culture medium, and (content reaches 0.5 μ g/cm3) plate be test group, to be not added with The plate of Nile red be control group, two groups from inclined-plane streak inoculation plant endogenesis epiphyte, and on each plate respectively inoculation refer to Show bacterium, be protected from light 3~4d of incubated at room temperature and be placed under ultraviolet lamp (280~300nm of wavelength) and observe, is oil-producing if having fluorescence appearance Strain, and can be determined compared with indicator bacteria in the more significant bacterial strain of apparently oil-producing according to its fluorescence intensity.
36 plants of endogenetic fungus have occurred different degrees of under the conditions of ultraviolet in 112 plants of endogenetic fungus being inoculated with as the result is shown Fluorescence developing, the bacterium colony and un-added bacterium colony for adding Nile red form sharp contrast, glimmering under ultraviolet according to Nile red plate Light intensity is weak, be divided into it is strong, in, weak three gradients, having chosen fluorescence intensity is that 12 plants of strong endogenetic fungus carry out oil productivity measurement.
2, oil productivity Preliminary detection
Grease is extracted using acid heat method to 12 plants of oil-producing endogenetic fungus that preliminary screening obtains, oil productivity is calculated.
Wherein, acid heat method extracts the step of grease are as follows:
Bacteria cake is taken from the bacterium colony plate of 12 plants of oil-producing endogenetic fungus be linked into be placed in 120r/min in PDB culture medium and shake It ferments dark culture (masking foil package) under the conditions of 25 DEG C on bed, fermentation liquid is centrifuged 5min in 4000r/min, collects thallus, presses The hydrochloric acid of 5ml 4mol/L is added in every gram of thallus, is put in 180r/min shaking table concussion 6h, then boiling water bath 10min, -20 DEG C of coolings 30min is repeated 3 times, and 2 times of volume of chloroform-methanol (1: 1) solution are added, and after fulling shake, 4000r/min is centrifuged 5min, takes bottom Portion's chloroform layer adds isometric 0.1% sodium chloride solution, mixes, and 4000r/min is centrifuged 5min, takes chloroform layer, and volatilization chloroform obtains oily Rouge weighing is oil production.Oil-producing fermented liquid 4000r/min is centrifuged 5min, collects thallus, and thallus is placed on 70 DEG C It is dried in baking oven, claims its weight up to thallus dry measure (Fungal biodiversity).Endogenetic fungus oil productivity is calculated: oil productivity=production Oil mass (g)/Fungal biodiversity (g).
The biomass of 12 plants of oil-producing endogenetic fungus is between 3-12g/L as the result is shown, oil production between 0.7-3.1g/L, Oil productivity is 20% or more, and oil productivity has three plants in 30% or so bacterial strain, and endogenetic fungus C1T2-4 is selected to reflect Fixed and preservation.
Two, the Morphological Identification of endogenetic fungus
The colony morphology characteristic of endogenetic fungus C1T2-4, interior life are observed in culture endogenetic fungus C1T2-4 single colonie 5-15 days The mycelia of fungi C1T2-4 is more, but mycelia is thicker, and entire body is purple, and the white spore for having a circle indistinct above generates, back Portion is faint yellow (Fig. 2).Using optical microphotograph sem observation mycelia micro-morphology, the mycelia of endogenetic fungus C1T2-4 has point Branch, have every, have in cluster spherical spore generate (Fig. 3).
Three, the Molecular Identification of endogenetic fungus C1T2-4
CTAB method extracts the genomic DNA of endogenetic fungus C1T2-4, with universal primer ITS1 (5`- CTTGGTCATTTAGACGAAGTAA-3`) and ITS4 (5`-GCATATCAATAAGCGGAGGA-3`) PCR amplification ITS sequence, instead Answer program are as follows: 94 DEG C of denaturation 40s, 55 DEG C of annealing 50s, 72 DEG C of extension 1min, totally 35 recycle.PCR product electrophoresis detection, sequencing ITS sequence that C1T2-4 is expanded is obtained as shown in sequence 1 in sequence table.Log in NCBI (https: // Blast.ncbi.nlm.nih.gov/ the sequence) is subjected to sequence ratio (Blast): endogenetic fungus C1T2-4 matching degree higher order The Gene BankNumber of column is MH345902.1, matching degree 98%.
To sum up, determine that endogenetic fungus C1T2-4 is aspergillus (Aspergillus by Morphological Identification and Molecular Identification sp.).Endogenetic fungus C1T2-4 is named as aspergillus (Aspergillus sp.) C1T2-4, hereinafter referred to as aspergillus C1T2- 4.Aspergillus C1T2-4 is preserved in China typical culture collection center (abbreviation CCTCC on May 24th, 2019；Address: in State Wuhan, Wuhan University；Postcode: 430072), deposit number is CCTCC NO:M 2019393.
The optimization of 2 aspergillus C1T2-4 fermentation system of embodiment
It can be seen that aspergillus C1T2-4 oil productivity is lower by oil productivity Preliminary detection result in embodiment 1, work be not achieved Therefore the desirable of industry production is optimized improvement on basic condition of culture to reach the requirement of superior strain.
One, oil-producing endogenetic fungal bacterial strain incubation time optimizes
Using above-mentioned aspergillus C1T2-4 as aimed strain, 3 bacteria cakes of uniform size are taken from the uniform plate of bacterium colony Be linked into PDB culture medium and be placed on 120r/min shaking table under the conditions of 25 DEG C fermentation dark culture (masking foil package), divide in 4d, 6d, 8d, 10d, 12d are sampled, and are extracted grease using acid heat method described in embodiment 1, are drawn oil productivity and Fungal biodiversity Time graph.
As a result as shown in figure 4, aspergillus C1T2-4 oil production before 6d is lower, and change less, oil production after 6d It quicklys increase, is reached most preferably when reaching 10d, start to be declined later again.Therefore, as can be seen from the figure aspergillus C1T2-4 The best oil-producing time between 10d, in the later period of the logarithmic growth phase of endogenetic fungal bacterial strain cell, oil production at this time and Fungal biodiversity all reaches saturation state, is optimal extraction time, if the time is too long with base consumption oil production and interior Raw fungal bacterial strain growth is slack-off, and the cell meeting broken wall of aging, lipid intracellular can spill into culture medium, can not collect, therewith Lead to the decline of oil productivity and oil production.
Two, oil-producing endogenetic fungal bacterial strain training systern
On the basis of determining aspergillus C1T2-4 best incubation time, improvement further is optimized to culture medium, if L16 as shown in Table 1 (4^4) orthogonal optimization test is set, investigates different pH, carbon source, nitrogen source and salinity to aspergillus C1T2- The influence of 4 oil productivity.
It is basic culture medium with PDB culture medium, has chosen pH, carbon source kind, nitrogen source type and the salt of culture medium respectively Concentration is factor, and four gradients are arranged in each factor, and specific as shown in table 1, i.e. pH is 3,5,7,9；Carbon source kind is respectively to form sediment Powder, glucose, sucrose, inositol；Nitrogen source type is respectively tryptone, yeast extract, ammonium nitrate, acid hydrolyzed casein；Salt Concentration is set as 0g/100ml, 3g/100ml, 6g/100ml, 9g/100ml.It is inoculated with aspergillus C1T2-4 (inoculum concentration 5ml/ 100ml), fermentation dark culture (masking foil package) 9d under the conditions of 25 DEG C is placed on 120r/min shaking table, using institute in embodiment 1 It states acid heat method and extracts grease, to investigate influence of the culture medium to biomass and oil productivity.Wherein each sample carries out 3 weights It is multiple.
Table 1 L16 (4^4) orthogonal optimization
The oil productivity of aspergillus C1T2-4 after optimization is calculated, the results showed that the Nei Shengzhen in different culture mediums The oil productivity of bacteria strain is different: the oil productivity of aspergillus C1T2-4 reaches maximum value 61.19% in the 4th group of experiment；Biomass Reach maximum value 3.477g/L in the 10th group of experiment；Oil production reaches maximum value 1.054g/L at the 10th group.
Range analysis is carried out to orthogonal optimization, analysis shows pH (A), carbon source (B), nitrogen source (C), salinity (D) are respectively Influence factor to aspergillus C1T2-4 oil production is D > C > A > B, influences maximum to be salinity on aspergillus C1T2-4, For orthogonal optimization as a result, aspergillus C1T2-4 is A1B2C4D3 (pH:3, carbon source: glucose, nitrogen source: acid hydrolyzed casein, salt Concentration: 6%), can high Lipid-producing.As it can be seen that lower pH value and glucose and acid hydrolyzed casein and 6% salinity are conducive to song Mould category C1T2-4 oil productivity improves.
Three, oil-producing bacterial strain oil fatty acid constituent analysis
Bacteria cake is taken from the bacterium colony plate of aspergillus C1T2-4 be linked into PDB culture medium be placed on 120r/min shaking table It ferments dark culture (masking foil package) under the conditions of 25 DEG C, fermentation liquid is centrifuged 5min in 4000r/min, thallus is collected, by every gram The hydrochloric acid of 5ml 4mol/L is added in thallus, is put in 180r/min shaking table concussion 6h, then boiling water bath 10min, -20 DEG C of coolings 30min is repeated 3 times, and 2 times of volume of chloroform-methanol (1: 1) solution are added, and after fulling shake, 4000r/min is centrifuged 5min, takes bottom Portion's chloroform layer adds isometric 0.1% sodium chloride solution, mixes, and 4000r/min is centrifuged 5min, takes chloroform layer, and volatilization chloroform obtains bent The mould grease for belonging to C1T2-4.Utilize the content of unsaturated fatty acid in GC-MS analysis grease (mark product are 35 kinds of fatty acid-mixed marks).
Wherein, GC-MS is analyzed method particularly includes:
Sample hydrolysis: it is appropriate to weigh uniform sample, and about 100mg pyrogallic acid is added, and several zeolites are added, add 95% ethyl alcohol of 2mL mixes.Hydrochloric acid solution 10mL is added, mixes.Flask is put into 70 DEG C of -80 DEG C of water-baths and hydrolyzes 40min.Often Flask is vibrated every 10min, is mixed into the particulate matter being attached in flask walls in solution.After the completion of hydrolysis, it is cold to take out flask But to room temperature.
The saponification of fat and methyl esterification of fatty acid: in fat-extraction object, it is molten to continuously add 2% sodium hydroxide methanol of 2mL 14% boron trifluoride methanol solution of 3mL, the water-bath in 85 DEG C of water-baths is added in liquid, water-bath 30min in 85 DEG C of water-baths 30min.After the completion of water-bath, etc. temperature drop to room temperature, in centrifuge tube be added 1mL n-hexane, concussion extraction 2min after, stand One hour, wait layering.100 μ L of supernatant liquor is taken, with n-hexane constant volume to 1mL.Examination with computer after crossing film with 0.45 μm of filter membrane.
Instrumental method
Chromatographic column: CD-2560 (100m × 0.25mm × 0.20 μm)；
Temperature program: 130 DEG C of holding 5min are warming up to 240 DEG C with the rate of 4 DEG C/min, keep 30min.
Injector temperature: 250 DEG C；Flow rate of carrier gas: 0.5mL/min；
Split sampling shunts: 10: 1；
Detector: FID；Detector temperature: 250 DEG C
The fatty acid profile of aspergillus C1T2-4 is shown in Fig. 5, and constituent analysis is shown in Table 2: 15 kinds are detected in aspergillus C1T2-4 Fatty acid, including it is myristic acid, pentadecane acid, palmitinic acid, cis- palmitoleic acid, 17 carbonic acid, heptadecenoic acid, stearic acid, anti- Oleic acid, octadecenoic acid (trans- 9), octadecenoic acid (cis- 9), linoleic acid, alpha-linolenic acid, two hendecoic acids, two ficocerylic acids, Tetracosa carbon acid illustrates that aspergillus C1T2-4 has more rich and varied fatty acid, it is possible to provide various rouge needed by human The bio-oil of fat acid and industrial use；All higher fatty acid of content has palmitinic acid in the grease that aspergillus C1T2-4 is generated 24.29%, linoleic acid 24.28%, oleic acid 14.52%, stearic acid 9.0%.In addition alpha-linolenic acid is also detected, content is 7.15%.
The above result shows that aspergillus C1T2-4 can generate a variety of fatty acid, and there is efficient alpha-linolenic acid synthesis capability, Strain excellent material is provided for the exploitation of the following health care product, cosmetics, chemical products, there is important value.
The Analysis of Fatty Acids Composition of 2 aspergillus C1T2-4 of table
The present invention is had been described in detail above.To those skilled in the art, do not depart from spirit of the invention and Range, and without carrying out under unnecessary experimental conditions, can synchronization parameters, concentration and under the conditions of, it is real in a wider range Apply the present invention.Although The present invention gives particular embodiments, it is understood that, the present invention can be improved further. In short, pressing the principle of the present invention, the application is intended to include any change, purposes or improvement of the present invention, including departing from this Shen Please in the open scope, and the change carried out with routine techniques known in the art.By the range of following attached claims, It can carry out the application of some essential characteristics.
SEQUENCE LISTING
<110>Northeast Forestry University
<120>one plants of endogenetic fungal bacterial strains for producing a variety of fatty acid and its application
<130> GNCFY191603
<160> 1
<170> PatentIn version 3.5
<210> 1
<211> 562
<212> DNA
<213>aspergillus (Aspergillus sp.)
<400> 1
aggttagggg tctagcgagc ccaacctccc acccgtgttt actgtacctt agttgcttcg 60
gcgggcccgc cattcatggc cgccgggggc tctcagcccc gggcccgcgc ccgccggaga 120
caccacgaac tctgtctgat ctagtgaagt ctgagttgat tgtatcgcaa tcagttaaaa 180
ctttcaacaa tggatctctt ggttccggca tcgatgaaga acgcagcgaa atgcgataac 240
tagtgtgaat tgcagaattc cgtgaatcat cgagtctttg aacgcacatt gcgccccctg 300
gtattccggg gggcatgcct gtccgagcgt cattgctgcc catcaagcac ggcttgtgtg 360
ttgggtcgtc gtcccctctc cgggggggac gggccccaaa ggcagcggcg gcaccgcgtc 420
cgatcctcga gcgtatgggg ctttgtcacc cgctctgtag gcccggccgg cgcttgccga 480
acgcaaatca atctttttcc aggttgacct cggatcaggt agggataccc gctgaactta 540
agcatatcaa taagcggagg aa 562

Claims (10)

1. one plant of aspergillus, it is characterised in that: the aspergillus is aspergillus (Aspergillus sp.) C1T2-4, in Chinese allusion quotation The deposit number of type culture collection is CCTCC N0:M 2019393.
2. a kind of microbial inoculum, it is characterised in that: the microbial inoculum is containing aspergillus (Aspergillus sp.) described in claim 1 ClT2-4。
3. microbial inoculum according to claim 2, it is characterised in that: the microbial inoculum is the microbial inoculum of Lipid-producing.
4. aspergillus (Aspergillus sp.) C1T2-4 described in claim 1 or microbial inoculum as claimed in claim 2 are as follows 1) application in -4) any:
1) application in production grease；
2) application in alpha-linolenic acid is being prepared；
3) application in health care product is being prepared；
4) application in cosmetics is being prepared；
5) application in industrial products is being prepared.
5. a kind of method for producing grease, it is characterised in that: the method includes with microbiological culture media culture claim 1 institute The aspergillus stated collects culture, obtains grease from the culture.
6. according to the method described in claim 5, it is characterized by: the carbon source of the microbiological culture media be starch, glucose, Sucrose or inositol, the nitrogen source of the microbiological culture media are tryptone, yeast extract, ammonium nitrate or acid hydrolyzed casein, The salinity of the microbiological culture media is 0-9g/100ml；Specifically, the carbon source of the microbiological culture media is glucose, institute The nitrogen source for stating microbiological culture media is acid hydrolyzed casein, and the salinity of the microbiological culture media is 6g/100ml.
7. according to the method described in claim 5, it is characterized by: the pH value of the microbiological culture media is 3-9；Specifically, The pH value is 3.
8. according to the method described in claim 5, it is characterized by: the time of the culture is 10-12 days.
9. microbial inoculum according to claim 3, or, application as claimed in claim 4, or, described in claim 5-8 is any Method, it is characterised in that:
The grease includes myristic acid, pentadecane acid, palmitinic acid, cis- palmitoleic acid, 17 carbonic acid, heptadecenoic acid, tristearin Acid, elaidic acid, octadecenoic acid (trans- 9), octadecenoic acid (cis- 9), linoleic acid, alpha-linolenic acid, two hendecoic acids, 23 Carbonic acid and/or tetracosa carbon acid.
10. a kind of method for producing alpha-linolenic acid, which comprises the following steps: claim 5-8 is any described The grease that method obtains is isolated and purified, and alpha-linolenic acid is obtained.
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Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN106047956A (en) * 2016-08-05 2016-10-26 广东海洋大学 Method for preparing gamma-linolenic acid-rich grease through sea squirt-associated Penicillium citrinum Asc2-4 fermentation

Patent Citations (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN106047956A (en) * 2016-08-05 2016-10-26 广东海洋大学 Method for preparing gamma-linolenic acid-rich grease through sea squirt-associated Penicillium citrinum Asc2-4 fermentation

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
叶思特等: "产油微生物的筛选", 《华南农业大学学报》 *
黄昌旭等: "普里兹湾沉积物样品产油真菌筛选及胞内脂肪酸成分分析", 《应用海洋学学报》 *

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