CN102492744B - Application of ethanolamine in improving lipid content in microalgae - Google Patents
Application of ethanolamine in improving lipid content in microalgae Download PDFInfo
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- CN102492744B CN102492744B CN2011103847834A CN201110384783A CN102492744B CN 102492744 B CN102492744 B CN 102492744B CN 2011103847834 A CN2011103847834 A CN 2011103847834A CN 201110384783 A CN201110384783 A CN 201110384783A CN 102492744 B CN102492744 B CN 102492744B
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- thanomin
- ethanolamine
- substratum
- scenedesmus obliquus
- microalgae
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Abstract
The invention discloses application of ethanolamine in improving lipid content in microalgae. Ethanolamine is added into a medium used for culturing microalgae. According to results of experiments, when ethanolamine is added into the medium used for culturing microalgae, total lipid content is improved by 10.2 to 15.2%; according to analysis from the perspective of fatty acid, addition of ethanolamine enables the content (18:1) of oleic acid to be increased, thereby enabling the quality of biodiesel to be improved. Embodiments in the invention fully illustrate that ethanolamine has a great application value in the field of production for biodiesel by using microalgae.
Description
Technical field
The invention belongs to the biofuel field, relate to a kind of purposes of thanomin.
Background technology
, along with the continuous consumption of fossil oil and to the destruction of environment, seek renewable energy source and enjoy countries in the world to pay close attention to.Biomass energy occupies an important position in whole energy resource system as a kind of renewable energy source.Little algae biofuel is a kind of biomass energy with large development potentiality, compare with the biofuel that animals and plants are raw material preparation, it have be easy to cultivate, do not occupy cultivated land, produce oil efficiency advantages of higher.Present little algae biofuel has obtained remarkable progress at home and abroad, but still face a lot of challenges, as high in production cost and lipid acid output is relatively low etc., and therefore, increasing people is devoted to study training mode and the high oily little algae aspect of screening of setting up high efficiency, low cost.
Summary of the invention
The purposes that the purpose of this invention is to provide ethanolamine in improving lipid content in microalgae.
Technical scheme of the present invention is summarized as follows:
The purposes of ethanolamine in improving lipid content in microalgae, join thanomin in the substratum of cultivating little algae.
Described little algae is scenedesmus obliquus Scenedesmus obliquus or chlorella Chlorella sorokiniana.
The concentration of described thanomin in the substratum of cultivating little algae is 0.5mM～5mM.
Experiment showed, that when adding thanomin in the substratum of cultivating little algae, total lipid content improves 10.2%-152%,, from the angle analysis of lipid acid, add after thanomin the content that has improved oleic acid (18: 1), therefore also improved the quality of biofuel.Proved absolutely that thanomin is utilizing little algae production field of biodiesel oil that very large purposes is arranged.
Description of drawings
Fig. 1 is the variation of 0.5mM, 2mM or 5mM fat content for adding respectively thanomin to make its concentration in the substratum to 3 parts of cultivation scenedesmus obliquus.
Fig. 2 is the variation of 0.5mM, 2mM or 5mM fat content for adding respectively thanomin to make its concentration in the substratum to 3 parts of cultivation chlorellas.
Fig. 3 is the changing conditions of 0.5mM lipid acid profile for adding thanomin to make its concentration in cultivation scenedesmus obliquus substratum
Fig. 4 is the changing conditions of 2mM lipid acid profile for adding thanomin to make its concentration in cultivation scenedesmus obliquus substratum
Embodiment
Embodiments of the invention are in order to make those skilled in the art can understand the present invention, to be not used in the present invention is imposed any restrictions.
Scenedesmus obliquus Scenedesmus obliquus or chlorella Chlorella sorokiniana all can buy in Inst. of Hydrobiology, Chinese Academy of Sciences's algae kind storehouse, and its preserving number is respectively FACHB-417 and FACHB-275.
The substratum of cultivating little algae is the BG11 substratum, specifically fills a prescription as follows:
First configure following mother liquor:
Mother liquor 1 (1L): NaNO
330g; K
2HPO
43H
2O 0.8g; Na
2CO
30.4g, with distilled water, be settled to 1L
Mother liquor 2 (100ml): citric acid 0.3g; Ferric ammonium citrate 0.3g; EDTA 0.05g, be settled to 100ml with distilled water
Mother liquor 3 (100ml): CaCl
22H
2O 1.8g, be settled to 100ml with distilled water
Mother liquor 4 (100ml): MgSO
47H
2O 3.75g, be settled to 100ml with distilled water
Mother liquor 5 (100ml): H
3BO
32.86g; MnCl
24H
2O 1.81g; ZnSO
47H
2O 0.222g; Na
2MoO
42H
2O0.39g; CuSO
45H
2O 0.079g; Co (NO
3)
26H
2O 0.04947g, be settled to 100ml with distilled water
During the BG11 substratum of preparation 1L, respectively. add mother liquor 1,2,3,4,5 volume is followed successively by 50ml, 2ml, 2ml, 2ml, 1ml, then add the distilled water polishing to 1L, transfers finally pH to 7.0-7.5 (various embodiments of the present invention adopt be pH to 7.0).
The present invention is further illustrated below in conjunction with specific embodiment.
Add ethanolamine solutions in the substratum of scenedesmus obliquus Scenedesmus obliquus, making the concentration of thanomin is 0.5mM, comprises following step:
(1) scenedesmus obliquus Scenedesmus obliquus is inoculated in the BG11 substratum, adds the thanomin aqueous solution, the concentration that makes thanomin is 0.5mM; (improve scenedesmus obliquus fat content in order to verify thanomin, establish three parallel, three BG11 culture medium culturing scenedesmus obliquus that do not add thanomin are set in addition);
Take a sample while (2) being cultured to stationary phase, centrifugal 5 minutes of 4000rpm, after the cell that obtains is cleaned three times with distilled water, freeze-drying in low-temperature freeze-drying machine, be stored in-80 ℃ stand-by.
Add ethanolamine solutions in the substratum of scenedesmus obliquus Scenedesmus obliquus, the concentration that makes thanomin is 2mM, and grid algae fat content improves obviously.Comprise following step:
(1) scenedesmus obliquus Scenedesmus obliquus is inoculated in the BG11 substratum, adds the thanomin aqueous solution, the concentration that makes thanomin is 2mM; (improve scenedesmus obliquus fat content in order to verify thanomin, establish three parallel, three BG11 culture medium culturing scenedesmus obliquus that do not add thanomin are set in addition);
(2) with embodiment 1;
Add ethanolamine solutions in the substratum of scenedesmus obliquus Scenedesmus obliquus, the concentration that makes thanomin is 5mM, and grid algae fat content improves nearly one times.Concrete steps are as follows:
(1) scenedesmus obliquus Scenedesmus obliquus is inoculated in the BG11 substratum and adds the thanomin aqueous solution, the concentration that makes thanomin is 5mM; (improve scenedesmus obliquus fat content in order to verify thanomin, establish three parallel, three BG11 culture medium culturing scenedesmus obliquus that do not add thanomin are set in addition);
(2) with embodiment 1;
Embodiment 4
To another green alga---add ethanolamine solutions in the substratum of chlorella Chlorella sorokiniana, the concentration that makes thanomin is 0.5mM, and fat content obviously improves.Concrete steps are as follows:
(1) chlorella Chlorella sorokiniana is inoculated in the BG11 substratum and adds the thanomin aqueous solution, the concentration that makes thanomin is 0.5mM; (improve chlorella fat content in order to verify thanomin, establish three parallel, three BG11 culture medium culturing chlorellas that do not add thanomin are set in addition);
(2) with embodiment 1;
Add ethanolamine solutions in the substratum of chlorella Chlorella sorokiniana, the concentration that makes thanomin is 2mM, and fat content is doubled many.Concrete steps are as follows:
(1) chlorella Chlorella sorokiniana is inoculated in the BG11 substratum and adds the thanomin aqueous solution, the concentration that makes thanomin is 2mM; (improve chlorella fat content in order to verify thanomin, establish three parallel, three BG11 culture medium culturing chlorellas that do not add thanomin are set in addition);
(2) with embodiment 1;
Embodiment 6
Add ethanolamine solutions in the substratum of chlorella Chlorella sorokiniana, the concentration that makes thanomin is 5mM, and fat content improves again.Concrete steps are as follows:
(1) chlorella Chlorella sorokiniana is inoculated in the BG11 substratum and adds the thanomin aqueous solution, the concentration that makes thanomin is 5mM; (improve chlorella fat content in order to verify thanomin, establish three parallel, three BG11 culture medium culturing chlorellas that do not add thanomin are set in addition);
(2) with embodiment 1;
Embodiment 7
Extract the total fat of cell, specific as follows:
The freeze drying cell 20mg that step (2) in embodiment 1 to 6 is obtained is placed in respectively the vial of 15ml, adds 2ml methyl alcohol and 1ml chloroform, after under 25 ℃ standing 24 hours, and vortex 2 minutes; Add the 1ml chloroform, vibrated 1 minute, then add 1.8ml distilled water, vortex 2 minutes, with solution 2000rpm, centrifugal two minutes, take off layer, with filter paper filtering of Whatman to the clean bottle of weigh in advance (being designated as W1); After the water-bath evaporation, residue further is dried to constant weight under 104 ℃.Weigh, be designated as W2.Total lipid content=W2-W1 (%dcw)
As seen from Figure 1, in embodiment 1, to after adding final concentration to be the thanomin of 0.5mM in the scenedesmus obliquus substratum, the scenedesmus obliquus total lipid content increases, but improves not obviously (12.5% to 13.8%), may be because the concentration that adds is too low; In embodiment 2, to after adding final concentration to be the ethanolamine solutions of 2mM in the scenedesmus obliquus substratum, the total lipid content of scenedesmus obliquus obviously improves (12.5% to 20.3%), illustrates that accumulation has obvious promoter action to fat for the thanomin of this concentration; After adding final concentration to be the ethanolamine solutions of 5mM to the scenedesmus obliquus substratum in embodiment 3, the total lipid content of scenedesmus obliquus is doubled (12.5% to 22.7%), illustrates that the thanomin of this concentration accumulates larger promoter action is arranged fat.
As seen from Figure 2, in embodiment 4, to after adding final concentration to be the ethanolamine solutions of 0.5mM in the chlorella substratum, the total lipid content of chlorella obviously improves (11.3% to 17.7%); In embodiment 5, to after adding final concentration to be the thanomin of 2mM in the chlorella substratum, the total lipid content of chlorella further improves (11.3% to 24.6%); In embodiment 6, to after adding final concentration to be the thanomin of 5mM in the chlorella substratum, the total lipid content of chlorella obviously improves (11.3% to 26.5%) again.
Embodiment 8
Extract and detect lipid acid, analyze and to add the lipid acid that content improves after thanomin.Concrete steps are as follows:
1. take the KOH-CH that freeze drying cell 10mg that step in embodiment 1 to 2 (2) obtains adds 1.5M
3OH solution 2ml,, in 85 ℃ of water-baths 1.5 hours, add interior mark margaric acid 10ug after vibration mixes after taking-up, get 500 μ L after fully mixing and carry out subsequent experimental;
2. add HCl-CH
3OH solution (1: 1, V/V) vibration is 2 minutes, makes it complete reaction, gets supernatant after centrifugal;
3. add normal hexane to extract, the centrifugal rear extraction liquid of drawing from upper strata;
4. the GC-MS detection is carried out in bottling
Adopt GC-MS to carry out qualitative, quantitative to lipid acid, condition is as follows:
Chromatographic column: DB-5 gas chromatographic column (30m*0.25mm, 0.25 μ m);
Sample size: 1 μ L;
Splitting ratio: 10: 1;
Injector temperature: 280 ℃;
GC interface temperature: 270 ℃;
Helium flow velocity: constant voltage, 91KPa;
Heating schedule: 70 ℃ keep 2min, with 5 ℃ of min
-1Speed be raised to 290 ℃, and at 290 ℃, keep 6min;
Ionization voltage: 70eV;
Source temperature: 250 ℃;
Sweep limit: 50-800m/z;
Sweep velocity: 2scans
-1
Fig. 3 shows, in embodiment 1, to after adding final concentration to be the thanomin of 0.5mM in the scenedesmus obliquus substratum, the content of linolic acid (18: 2) obviously increases.
Fig. 4 shows, in embodiment 2 to after adding final concentration to be the ethanolamine solutions of 2mM in the scenedesmus obliquus substratum, the content of hexadecadienoic acid (16: 2), oleic acid (18: 1) and linolic acid (18: 2) obviously increases, especially the content of oleic acid improves a lot, according to bibliographical information, the amount that contains oleic acid in biofuel is more, and the biofuel quality is better.Therefore, after adding thanomin, the biofuel quality of utilizing little algae to produce also is improved.
The present invention preferred scenedesmus obliquus Scenedesmus obliquus and chlorella Chlorella sorokiniana, experiment showed, that thanomin also can improve other little algae fat content.
Experiment showed, in cultivating other substratum of little algae and add thanomin, when to make the concentration of thanomin in the substratum of cultivating little algae be 0.5mM～5mM, also can improve fat content.
Claims (1)
1. the purposes of ethanolamine in improving lipid content in microalgae, is characterized in that, thanomin is joined in the substratum of cultivating little algae, and described little algae is scenedesmus obliquus Scenedesmus obliquus or chlorella Chlorella sorokiniana; The concentration of thanomin in the substratum of cultivating little algae is 0.5mM～5mM.
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| CN2011103847834A CN102492744B (en) | 2011-11-28 | 2011-11-28 | Application of ethanolamine in improving lipid content in microalgae |
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| WO2016138869A1 (en) * | 2015-03-04 | 2016-09-09 | Hubei University Of Technology | Methods for production of polyunsaturated fatty acids from coccolithophorid microalga pleurochrysis carterae |
| CN108410939B (en) * | 2018-07-12 | 2020-05-01 | 烟台大学 | Method for increasing astaxanthin content in haematococcus pluvialis |
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Non-Patent Citations (6)
| Title |
|---|
| Christopher R. McMaster et al..Modulation of phosphatidylethanolamine biosynthesis by exogenous ethanolamine and analogues in the hamster heart.《Molecular and Cellular Biochemistry》.1992,第116卷69-73. |
| Modulation of phosphatidylethanolamine biosynthesis by exogenous ethanolamine and analogues in the hamster heart;Christopher R. McMaster et al.;《Molecular and Cellular Biochemistry》;19921231;第116卷;69-73 * |
| 冯福应 等.海洋微藻Chlorella sp. 1061的脂类特征与分类探讨.《海洋与湖沼》.2004,第35卷(第5期),453-458. |
| 夏金兰 等.微藻生物柴油的现状与进展.《中国生物工程杂志》.2009,第29卷(第7期),118-126. |
| 微藻生物柴油的现状与进展;夏金兰 等;《中国生物工程杂志》;20091231;第29卷(第7期);118-126 * |
| 海洋微藻Chlorella sp. 1061的脂类特征与分类探讨;冯福应 等;《海洋与湖沼》;20040930;第35卷(第5期);453-458 * |
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