CN102899166B - Biological grease in algae cell and preparation method and application of biological grease - Google Patents
Biological grease in algae cell and preparation method and application of biological grease Download PDFInfo
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Abstract
The invention discloses a method for preparing biological grease in algae cell. The method comprises the following steps of: firstly, crushing the algae cell, collecting mixed liquor of crushed algae cell, performing centrifugation, taking supernatant, performing first-time liquid-liquid extraction by using a normal hexane/ethyl acetate mixed organic solvent, and collecting aqueous phase and organic phase; secondly, performing second-time liquid-liquid extraction on the aqueous phase obtained by the first-time liquid-liquid extraction by using the normal hexane/ethyl acetate mixed organic solvent, and collecting aqueous phase and organic phase; thirdly, performing third-time liquid-liquid extraction on the aqueous phase obtained by the second-time liquid-liquid extraction by using the normal hexane/ethyl acetate mixed organic solvent, and collecting aqueous phase and organic phase; fourthly, combining the organic phases obtained by the three-time extraction, and performing vacuum evaporation to obtain the biological grease in algae cell; and finally adding a KOH-containing methanol solution into the biological grease in algae cell, stirring violently, and performing esterification reaction to obtain biodiesel which is in line with 'GB/T 20828-2007 Bildiesel Blend Stock (BD100) for Diesel Engine Fuels'.
Description
Technical field
The present invention relates to bio-oil and the application aspect preparation biofuel thereof in a kind of alga cells.
Background technology
Along with the acceleration consumption of fossil oil in world wide and petroleum-based energy and various pollutions and the environmental problem of bringing thus, reproducible bioenergy obtains people's attention day by day.Under the encouragement and support of every policy, the annual production of whole world biofuel (comprising biofuel and bio-ethanol etc.) has reached approximately 35,000,000,000 liters.
Biofuel is to be raw material with bio-oil (generally including vegetation fat, animal tallow etc.) with alcohols material (being generally methyl alcohol, ethanol), the longer chain fatty acid methyl esters obtaining by transesterification reaction.Methyl alcohol at a certain temperature, with the triglyceride level generation transesterification reaction in bio-oil, produces glycerine and fatty acid methyl ester, and after glycerine is separated, the fatty acid methyl ester obtaining is exactly biofuel.In transesterification reaction, the theoretical molar ratio of triglyceride level and methyl alcohol is 1:3, but when industrial production, conventionally adds excessive methyl alcohol to carry out to the direction that generates methyl esters to guarantee reaction.In transesterification reaction, can make catalyzer with acid, alkali or enzyme and carry out catalysis.Biofuel can replace traditional diesel fuel applications of being extracted by oil in existing diesel engine, its Stability Analysis of Structures, use safety, in combustion processes, than traditional diesel oil, give off harmful atmospheric polluting material still less as oxysulfide, arene compound, oxycarbide, particulate matter etc., and due to it, possess the advantages such as nontoxic, biodegradable, therefore demonstrate very potential application prospect.Correlative study shows, the biofuel of equivalent in combustion processes on average than conventional diesel few particulate matter that discharges 53%-69%, the flying dust of 79%-83%, the hydrocarbon polymer of 45%-67% and the carbon monoxide of 4%-16%.Among physical environment, most biofuel can be degraded within 21-28 days, its degradation speed is conventional diesel 4 times.In addition, biofuel also has good lubricity and better combustionproperty.
The Vegetable oil lipoprotein squeezing with oil crops, discarded cooking oil and animal grease are that the practice of waste biofuel has had the history that surpasses 50 years.In the U.S., the main raw material of biofuel is soybean oil, but its industrial scale and output have been difficult to meet huge demand.The microbial oil of take also has the history of nearly 30 years as the research of waste biofuel, yet until has just been subject in recent years more and more attracting attention.Wherein, the main source that algae is microbial oil.Algae be the low grade of a class to put oxygen autotrophic type biological, it is of a great variety, distribution is on earth extremely extensive, 40% in the biomass that tellurian biology is produced by photosynthesis every year all should give the credit to the photosynthesis of algae.Micro-algae refers to that unicellular, the colony of some microcosmic or frond are thread algae, and most micro algae growths breedings, in water surrounding, can not competed with agricultural land and animal husbandry land used mutually.
Microbial oil is microorganism to utilize the synthetic glyceryl ester of carbohydrate, its lipid acid to form close with vegetables oil as microalgae etc., is mainly tri-glyceride.Fat content in microorganism cells body is generally greater than 40% to 50%, and some even can reach more than 80% through fat content in the microalgae cell of genetically engineered improvement.Take microbial oil as raw material, through refining, transesterify, the processes such as separation and purification, can produce the biofuel of high-quality.
From algae, extracting bio-oil must one of experience carry out pretreated process to alga cells, and the object of this process, for destroying its cell walls and cytolemma, makes intracellular lipid flow out and be extracted.Preprocessing process generally comprises dry, several steps such as extraction of alga cells fragmentation and microbial oil.The breaking method of alga cells mainly contains: homogenizer is broken, and bead mill grinds, ultrasonic disruption, and broken microwave, the autoclave pressurization of heating is broken, and multigelation method is broken, organic solvent cracking, osmotic pressure impact grinding, enzymatic lysis etc.
In current worldwide, energy scarcity, fossil fuel is about to exhaust, and the exploitation of new forms of energy is extremely urgent.Biofuel refers to that take the waterplant greases such as oil crops, Wild oil plant and engineering microalgae and animal grease, food garbage wet goods is stock oil, the reproducibility fuel of the replaced petrifaction diesel of making by ester exchange process.Biofuel has good environmental protection characteristic, low-temperature startup performance, lubricity, good safety performance and combustionproperty etc.The bio-oil extracting in alga cells body, will be a kind of good biodiesel raw material.
Summary of the invention
One of object of the present invention is to provide the preparation method of the bio-oil in a kind of alga cells, and this preparation method can quick crashing alga cells, to extract the bio-oil in alga cells.
The application of bio-oil in a kind of alga cells that two of object of the present invention is to provide above-mentioned in preparation biofuel.
Technical scheme of the present invention
The preparation method of the bio-oil in a kind of alga cells, first that alga cells is broken, then collect broken alga cells mixed solution centrifugal, get supernatant liquor and carry out liquid-liquid extraction for the first time with n-hexane/ethyl acetate mixed solvent, collect water and organic phase;
The water that liquid-liquid extraction is for the first time obtained carries out liquid-liquid extraction for the second time with n-hexane/ethyl acetate mixed organic solvents, collects water and organic phase;
The water that extracts for the second time gained is carried out to liquid-liquid extraction for the third time with n-hexane/ethyl acetate mixed organic solvents, collect water and organic phase;
The organic phase vacuum-evaporation that merges three extraction gained is removed the product of gained after n-hexane/ethyl acetate mixed organic solvents and is the bio-oil in alga cells.
The preparation method of the bio-oil in above-mentioned a kind of alga cells, specifically comprises the steps:
(1), by algae liquid in airtight stainless steel cauldron, at the uniform velocity stir, keep chlorella to be suspended in solution;
(2), utilize ozonizer and air compressor machine that ozone and air are calculated by volume, be ozone: the ratio that air is 1:150 passes into the mixed gas of ozone and air composition in airtight reactor, until pressure reaches 0.4～0.8 Mpa in reactor, then open rapidly the purging valve of reactor, relief pressure is to normal atmosphere, and this process is called a pressurization-decompression cycle;
(3), the pressurization-decompression cycle 40～80 times of repeating step (2) repeatedly, finally obtain broken alga cells mixed solution;
(4), take out the broken alga cells mixed solution in closed reactor, after centrifugal, get supernatant liquor, calculate by volume, be supernatant liquor: n-hexane/ethyl acetate mixed organic solvents is 1:0.3, in supernatant liquor, add n-hexane/ethyl acetate mixed organic solvents, carry out liquid-liquid extraction for the first time, after layering, collect respectively water 1 and organic phase 1;
Described n-hexane/ethyl acetate mixed organic solvents, is calculated by normal hexane and ethyl acetate, i.e. normal hexane: the ratio that ethyl acetate is 1:1 mixes by volume;
(5) in the water 1 of, collecting in step (4), calculate by volume, be water 1: the ratio that n-hexane/ethyl acetate mixed organic solvents is 1:0.2 adds n-hexane/ethyl acetate mixed organic solvents, carry out liquid-liquid extraction for the second time, after layering, collect respectively water 2 and organic phase 2;
Described n-hexane/ethyl acetate mixed organic solvents, is calculated by normal hexane and ethyl acetate, i.e. normal hexane: the ratio that ethyl acetate is 1:1 mixes by volume;
(6) in the water 2 of, collecting in step (5), calculate by volume, be water 2: the ratio that n-hexane/ethyl acetate mixed organic solvents is 1:0.1 adds n-hexane/ethyl acetate mixed organic solvents, carry out liquid-liquid extraction for the third time, after layering, collect respectively water 3 and organic phase 3;
Described n-hexane/ethyl acetate mixed organic solvents, is calculated by normal hexane and ethyl acetate, i.e. normal hexane: the ratio that ethyl acetate is 1:1 mixes by volume;
(7) after organic phase 1, organic phase 2 and organic phase 3 that, combining step (4), (5) and (6) three extractions are collected, insert in vacuum rotary evaporator, under normal temperature, be evaporated to normal hexane and the evaporation of ethyl acetate mixed organic solvents completely, obtain the bio-oil in alga cells.
The application of bio-oil in above-mentioned a kind of alga cells in preparation biofuel, in the bio-oil in alga cells, adding concentration is the methanol solution of the KOH of 2 mol/L, vigorous stirring is carried out esterification, in reaction process, mixing speed is 2800 rpm, time is 2 min, esterification is converted into fatty acid methyl ester by lipid acid, stratification, and upper strata products therefrom is biofuel;
Described concentration is that the add-on of methanol solution of the KOH of 2 mol/L accounts for bio-oil mass ratio in alga cells by it and calculates, i.e. bio-oil in alga cells: concentration is that the methanol solution of the KOH of 2 mol/L is 1:10-20.
Beneficial effect of the present invention
The preparation method of the bio-oil in a kind of alga cells of the present invention, in its preparation process, utilize the auxiliary By Ozone of pressure to alga cells broken wall, take ozone as medium, by gas-liquid reaction, act on algae liquid, under agitator effect, make ozone mix and then broken alga cells with algae liquid, therefore there is equipment flowsheet simple, shell-broken effect is good, easily is automated the advantages such as control, is a kind of effective alga cells breaking method.
Further, the preparation method of the bio-oil in a kind of alga cells of the present invention, owing to utilizing cell walls and the cytolemma of the oxidation susceptibility oxidation algae of ozone, assist and destroy the cell of algae with pressure, make the composition outflows such as lipid acid in alga cells, and with an organic solvent in the mode of liquid-liquid extraction, obtain lipid acid and alkane.Experiment showed, that at system pressure be 0.8 MPa, under the condition that pressurization-decompression cycle number is 80, the bio-oil percentage extraction of chlorella can reach 16.7-26.7%, and the degree of saturation of product increases.So, the flow process of preparing biofuel with the bio-oil in alga cells provided by the invention, possess simple to operate, raw material sources are extensive, output is high, low price, be easy to the advantages such as mass-producing, are a kind of effective novel method for preparing biodiesel.The biofuel of final gained, its main component is that C16:0 and C18:0 are respectively 27.8%-45.9% and 34.7%-48.6%, meets national GB/T 20828-2007 " diesel-fuel is in harmonious proportion by biofuel (BD100) " standard.
Accompanying drawing explanation
Fig. 1, be the preparation method's of the bio-oil in a kind of alga cells of the present invention process flow sheet.
Embodiment
Below by specific embodiment, also describe in further detail by reference to the accompanying drawings and to the present invention, but do not limit the present invention.
Bio-oil in a kind of alga cells, by the preparation method who comprises the steps, be prepared, the process flow diagram of its preparation process as shown in Figure 1, be first that algal species cultivation and algae liquid obtain, then the algae liquid of results being put into autoclave stirs, pass into is ozone by volume: the ozone that air is 1:150 and the mixed gas of air also stir, be forced into 0.4～0.8 Mpa, maintained for 1～2 s time, open reactor vent valve relief pressure to normal pressure, at this moment oversaturated gas discharges from algae liquid;
Repeat above-mentioned stirring, pass into mixed gas, the pressurization of ozone and air, release of pressure, 40～80 circulations of process that oversaturated gas discharges from algae liquid, have completed the fragmentation of alga cells;
Collect broken alga cells mixed solution, get supernatant liquor after centrifugal, with n-hexane/ethyl acetate mixed organic solvents, carry out liquid-liquid extraction for the first time, after layering, collect respectively water and organic phase;
The water that liquid-liquid extraction is for the first time obtained carries out liquid-liquid extraction for the second time with n-hexane/ethyl acetate mixed organic solvents again, collects respectively water and organic phase after layering;
The water that extracts for the second time gained is carried out to liquid-liquid extraction for the third time with n-hexane/ethyl acetate mixed organic solvents again, after layering, collect respectively water and organic phase;
The organic phase that merges three extraction gained is inserted in vacuum rotary evaporator, is evaporated to the evaporation of n-hexane/ethyl acetate mixed organic solvents completely under normal temperature, obtains the bio-oil in alga cells.
The methanol solution that adds KOH in bio-oil in alga cells, vigorous stirring is carried out esterification, and products therefrom is biofuel.
embodiment 1
A bio-oil in alga cells, is prepared by a method comprising the following steps:
(1), the algae liquid that measures 200 ml results in the autoclave of 1 L, under 120 r/min conditions, stir 5 min;
(2), open ozonizer and air compressor machine, be ozone by volume: the ratio control ozone that air is 1:150 and the total flux of air are 2 L/min, the mixed gas of ozone and air composition is passed in autoclave, continuation is at the uniform velocity stirred algae liquid with 120 r/min, and the ozone, air and the algae liquid that pass into are evenly mixed;
(3), controlling autoclave pressure is 0.6 MPa, when reaching 0.6 Mpa, the vent valve of opening autoclave makes Pressure Drop to normal pressure, and then boost to 0.6 MPa, when reaching 0.6 Mpa, the vent valve of again opening autoclave makes Pressure Drop to normal pressure, and circulation is boosted, step-down process obtains broken alga cells 40 times;
(4), open reactor, take out algae liquid, get supernatant liquor after centrifugal, utilize the mixed organic solvents being formed by normal hexane and ethyl acetate that 25 mL volume ratios are 1:1 to carry out liquid-liquid extraction for the first time, separatory, collect respectively water 1 and organic phase 1;
(5), take out the water 1 that step (4) obtains, utilize the mixed organic solvents being formed by normal hexane and ethyl acetate that 15 mL volume ratios are 1:1 to carry out liquid-liquid extraction for the second time, separatory, collect respectively water 2 and organic phase 3.
(6), take out the water 2 that step (5) obtains, utilize the mixed organic solvents being formed by normal hexane and ethyl acetate that 10 ml volume ratios are 1:1 to carry out liquid-liquid extraction for the third time, separatory, collection organic phase 3; .
(7), organic phase 1, organic phase 2 and the organic phase 3 of combining step (4), (5), (6) three step gained insert in vacuum rotary evaporator device, rotary evaporation 30 min, obtain the bio-oil in alga cells, its productive rate is the dry algae of 17.6g/100g.
In the bio-oil extracting to the algae liquid of above-mentioned gained, adding concentration is the methanol solution of the KOH of 2 mol/L, the mass ratio that its add-on accounts for the bio-oil in alga cells by it calculates, be the bio-oil in alga cells: concentration is that the methanol solution of the KOH of 2 mol/L is about 1:17, vigorous stirring is carried out esterification, in esterification reaction process, controlling mixing speed is 2800 rpm, time is 2 min, obtains biofuel after having reacted.
The biofuel of above-mentioned gained, be dissolved in the laggard row gas chromatography-mass spectrum of 1 mL hexane solution and detect discovery, its main component is C16:0 and C18:0, according to national standard analytical procedure (GB/T 22223-2008), determine that its relative percentage composition is respectively 27.8% and 34.7%, meet national GB/T 20828-2007 " diesel-fuel is in harmonious proportion by biofuel (BD100) " standard.
embodiment 2
A bio-oil in alga cells, is prepared by a method comprising the following steps:
(1), the algae liquid that measures 200 mL results in the autoclave of 1 L, under 120 r/min conditions, stir 5 min;
(2), open ozonizer and air compressor machine, be ozone by volume: the ratio control ozone that air is 1:150 and the total flux of air are 2 L/min, the mixed gas of ozone and air composition is passed in autoclave, continuation is at the uniform velocity stirred algae liquid with 120 r/min, and the ozone, air and the algae liquid that pass into are evenly mixed;
(3), controlling autoclave pressure is 0.6 MPa, when reaching 0.6 Mpa, the vent valve of opening autoclave makes Pressure Drop to normal pressure, and then boost to 0.6 MPa, when reaching 0.6 Mpa, the vent valve of again opening autoclave makes Pressure Drop to normal pressure, and circulation is boosted, step-down process obtains broken alga cells 80 times;
(4), open reactor, take out algae liquid, get supernatant liquor after centrifugal, utilize the mixed organic solvents being formed by normal hexane and ethyl acetate that 25 mL volume ratios are 1:1 to carry out liquid-liquid extraction for the first time, separatory, collect respectively water 1 and organic phase 1;
(5), take out the water 1 that step (4) obtains, utilize the mixed organic solvents being formed by normal hexane and ethyl acetate that 15 mL volume ratios are 1:1 to carry out liquid-liquid extraction for the second time, separatory, collect respectively water 2 and organic phase 3.
(6), take out the water 2 that step (5) obtains, utilize the mixed organic solvents being formed by normal hexane and ethyl acetate that 10 ml volume ratios are 1:1 to carry out liquid-liquid extraction for the third time, separatory, collection organic phase 3; .
(7), organic phase 1, organic phase 2 and the organic phase 3 of combining step (4), (5), (6) three step gained insert in vacuum rotary evaporator device, rotary evaporation 30 min, obtain the bio-oil in alga cells, its productive rate is the dry algae of 26.7g/100g.
In the bio-oil that the algae liquid of above-mentioned gained is extracted, adding concentration is the methanol solution of the KOH of 2 mol/L, the bio-oil mass ratio that its add-on accounts in alga cells by it calculates, be the bio-oil in alga cells: concentration is that the methanol solution of the KOH of 2 mol/L is about 1:10, vigorous stirring is carried out esterification, in esterification reaction process, controlling mixing speed is 2800 rpm, time is 2 min, obtains biofuel after having reacted.
The biofuel of above-mentioned gained, be dissolved in the laggard row gas chromatography-mass spectrum of 1 mL hexane solution and detect discovery, its main component is C16:0 and C18:0, according to national standard analytical procedure (GB/T 22223-2008), determine that its relative percentage composition is respectively 45.9% and 46.3%, meet national GB/T 20828-2007 " diesel-fuel is in harmonious proportion by biofuel (BD100) " standard.
embodiment 3
A bio-oil in alga cells, is prepared by a method comprising the following steps:
(1), the algae liquid that measures 200 mL results in the autoclave of 1 L, under 120 r/min conditions, stir 5 min;
(2), open ozonizer and air compressor machine, be ozone by volume: the ratio control ozone that air is 1:150 and the total flux of air are 2 L/min, the mixed gas of ozone and air composition is passed in autoclave, continuation is at the uniform velocity stirred algae liquid with 120 r/min, and the ozone, air and the algae liquid that pass into are evenly mixed;
(3), controlling autoclave pressure is 0.8 MPa, when reaching 0.8 Mpa, the vent valve of opening autoclave makes Pressure Drop to normal pressure, and then boost to 0.8 MPa, when reaching 0.8 Mpa, the vent valve of again opening autoclave makes Pressure Drop to normal pressure, and circulation is boosted, step-down process obtains broken alga cells 80 times;
(4), open reactor, take out algae liquid, get supernatant liquor after centrifugal, utilize the mixed organic solvents being formed by normal hexane and ethyl acetate that 25 mL volume ratios are 1:1 to carry out liquid-liquid extraction for the first time, separatory, collect respectively water 1 and organic phase 1;
(5), take out the water 1 that step (4) obtains, utilize the mixed organic solvents being formed by normal hexane and ethyl acetate that 15 mL volume ratios are 1:1 to carry out liquid-liquid extraction for the second time, separatory, collect respectively water 2 and organic phase 3.
(6), take out the water 2 that step (5) obtains, utilize the mixed organic solvents being formed by normal hexane and ethyl acetate that 10 ml volume ratios are 1:1 to carry out liquid-liquid extraction for the third time, separatory, collection organic phase 3; .
(7), organic phase 1, organic phase 2 and the organic phase 3 of combining step (4), (5), (6) three step gained insert in vacuum rotary evaporator device, rotary evaporation 30 min, obtain the bio-oil in alga cells, its productive rate is the dry algae of 24.3g/100g.
In the bio-oil extracting to the algae liquid of above-mentioned gained, adding concentration is the methanol solution of the KOH of 2 mol/L, the mass ratio that its add-on accounts for the bio-oil in alga cells by it calculates, be the bio-oil in alga cells: concentration is that the methanol solution of the KOH of 2 mol/L is about 1:13, vigorous stirring is carried out esterification, in esterification reaction process, controlling mixing speed is 2800 rpm, time is 2 min, obtains biofuel after having reacted.
The biofuel of above-mentioned gained, be dissolved in the laggard row gas chromatography-mass spectrum of 1 mL hexane solution and detect discovery, its main component is C16:0 and C18:0, according to national standard analytical procedure (GB/T 22223-2008), determine that its relative percentage composition is respectively 40.4% and 48.6%, meet national GB/T 20828-2007 " diesel-fuel is in harmonious proportion by biofuel (BD100) " standard.
embodiment 4
A bio-oil in alga cells, is prepared by a method comprising the following steps:
(1), the algae liquid that measures 200 ml results in the autoclave of 1 L, under 120 r/min conditions, stir 5 min;
(2), open ozonizer and air compressor machine, be ozone by volume: the ratio control ozone that air is 1:150 and the total flux of air are 2 L/min, the mixed gas of ozone and air composition is passed in autoclave, continuation is at the uniform velocity stirred algae liquid with 120 r/min, and the ozone, air and the algae liquid that pass into are evenly mixed;
(3), controlling autoclave pressure is 0.4 MPa, when reaching 0.4 Mpa, the vent valve of opening autoclave makes Pressure Drop to normal pressure, and then boost to 0.4 MPa, when reaching 0.4 Mpa, the vent valve of again opening autoclave makes Pressure Drop to normal pressure, and circulation is boosted, step-down process obtains broken alga cells 80 times;
(4), open reactor, take out algae liquid, get supernatant liquor after centrifugal, utilize the mixed organic solvents being formed by normal hexane and ethyl acetate that 25 mL volume ratios are 1:1 to carry out liquid-liquid extraction for the first time, separatory, collect respectively water 1 and organic phase 1;
(5), take out the water 1 that step (4) obtains, utilize the mixed organic solvents being formed by normal hexane and ethyl acetate that 15 mL volume ratios are 1:1 to carry out liquid-liquid extraction for the second time, separatory, collect respectively water 2 and organic phase 3.
(6), take out the water 2 that step (5) obtains, utilize the mixed organic solvents being formed by normal hexane and ethyl acetate that 10 ml volume ratios are 1:1 to carry out liquid-liquid extraction for the third time, separatory, collection organic phase 3; .
(7), organic phase 1, organic phase 2 and the organic phase 3 of combining step (4), (5), (6) three step gained insert in vacuum rotary evaporator device, rotary evaporation 30 min, obtain the bio-oil in alga cells, its productive rate is the dry algae of 22.5 g/100g.
To adding concentration in the bio-oil of above-mentioned gained, it is the methanol solution of the KOH of 2 mol/L, the mass ratio that its add-on accounts for the bio-oil in alga cells by it calculates, be the bio-oil in alga cells: concentration is that the methanol solution of the KOH of 2 mol/L is about 1:20, vigorous stirring is carried out esterification, in esterification reaction process, controlling mixing speed is 2800 rpm, time is 2 min, obtains biofuel after having reacted.
The biofuel of above-mentioned gained, be dissolved in the laggard row gas chromatography-mass spectrum of 1 mL hexane solution and detect (manufacturer: U.S. Agilent company, model: 7890) find, its main component is C16:0 and C18:0, according to national standard analytical procedure (GB/T 22223-2008), determine that its relative percentage composition is respectively 40.8% and 39.2%, meet national GB/T 20828-2007 " diesel-fuel is in harmonious proportion by biofuel (BD100) " standard.
The above is only giving an example of embodiments of the present invention; it should be pointed out that for those skilled in the art, do not departing under the prerequisite of the technology of the present invention principle; can also make some improvement and modification, these improve and modification also should be considered as protection scope of the present invention.
Claims (5)
1. the preparation method of the bio-oil in an alga cells, first alga cells is broken, then collect broken alga cells mixed solution centrifugal, get supernatant liquor and carry out liquid-liquid extraction for the first time with n-hexane/ethyl acetate mixed organic solvents, collect water and organic phase;
The water that liquid-liquid extraction is for the first time obtained carries out liquid-liquid extraction for the second time with n-hexane/ethyl acetate mixed organic solvents, collects water and organic phase;
The water that extracts for the second time gained is carried out to liquid-liquid extraction for the third time with n-hexane/ethyl acetate mixed organic solvents, collect water and organic phase;
The organic phase vacuum-evaporation that merges three extraction gained is removed the product of gained after n-hexane/ethyl acetate mixed organic solvents and is the bio-oil in alga cells;
It is characterized in that first described alga cells fragmentation be that algal species cultivation and algae liquid obtain, then the algae liquid of results being put into autoclave stirs, pass into the mixed gas of ozone and air and stir, be forced into 0.4～0.8Mpa, maintain 1～2s time, open reactor vent valve relief pressure to normal pressure, at this moment oversaturated gas discharges from algae liquid;
Repeat above-mentioned stirring, pass into mixed gas, the pressurization of ozone and air, release of pressure, 40～80 circulations of process that oversaturated gas discharges from algae liquid, obtain broken alga cells.
2. the preparation method of the bio-oil in a kind of alga cells as claimed in claim 1, is characterized in that described ozone and air gas mixture are ozone by volume: air is 1:150, and total flux is 2 L/min.
3. the preparation method of the bio-oil in a kind of alga cells as claimed in claim 2, while it is characterized in that liquid-liquid extraction for the first time, calculates, i.e. supernatant liquor: n-hexane/ethyl acetate mixed organic solvents is 1:0.3 by volume;
Described n-hexane/ethyl acetate mixed organic solvents, is calculated by normal hexane and ethyl acetate, i.e. normal hexane: the ratio that ethyl acetate is 1:1 mixes by volume.
4. the preparation method of the bio-oil in a kind of alga cells as claimed in claim 3, while it is characterized in that liquid-liquid extraction for the second time, calculates by volume, i.e. the water of liquid-liquid extraction gained for the first time: n-hexane/ethyl acetate mixed organic solvents is 1:0.2;
Described n-hexane/ethyl acetate mixed organic solvents, is calculated by normal hexane and ethyl acetate, i.e. normal hexane: the ratio that ethyl acetate is 1:1 mixes by volume.
5. the preparation method of the bio-oil in a kind of alga cells as claimed in claim 4, while it is characterized in that liquid-liquid extraction for the third time, calculates by volume, i.e. the water of liquid-liquid extraction gained for the second time: n-hexane/ethyl acetate mixed organic solvents is 1:0.1;
Described n-hexane/ethyl acetate mixed organic solvents, is calculated by normal hexane and ethyl acetate, i.e. normal hexane: the ratio that ethyl acetate is 1:1 mixes by volume.
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| CN102533879A (en) * | 2010-12-17 | 2012-07-04 | 中国科学院大连化学物理研究所 | Microbial oil extraction method |
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