CN114015493B - Method for extracting grease from algae - Google Patents
Method for extracting grease from algae Download PDFInfo
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- CN114015493B CN114015493B CN202111126436.1A CN202111126436A CN114015493B CN 114015493 B CN114015493 B CN 114015493B CN 202111126436 A CN202111126436 A CN 202111126436A CN 114015493 B CN114015493 B CN 114015493B
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- 241000195493 Cryptophyta Species 0.000 title claims abstract description 198
- 239000004519 grease Substances 0.000 title claims abstract description 59
- 238000000034 method Methods 0.000 title claims abstract description 15
- 238000012258 culturing Methods 0.000 claims abstract description 5
- 238000005286 illumination Methods 0.000 claims description 28
- 238000009423 ventilation Methods 0.000 claims description 18
- 241000342018 Aegagropila linnaei Species 0.000 claims description 12
- 238000000605 extraction Methods 0.000 abstract description 10
- 230000008569 process Effects 0.000 abstract description 5
- 230000000694 effects Effects 0.000 abstract description 3
- 238000003306 harvesting Methods 0.000 abstract description 3
- 230000001580 bacterial effect Effects 0.000 abstract description 2
- 150000002632 lipids Chemical class 0.000 description 68
- 239000002028 Biomass Substances 0.000 description 28
- 239000003921 oil Substances 0.000 description 27
- 235000019198 oils Nutrition 0.000 description 27
- 238000005273 aeration Methods 0.000 description 10
- 238000005259 measurement Methods 0.000 description 10
- 240000004808 Saccharomyces cerevisiae Species 0.000 description 9
- 238000004519 manufacturing process Methods 0.000 description 8
- 244000081822 Uncaria gambir Species 0.000 description 6
- 238000001035 drying Methods 0.000 description 6
- 230000000243 photosynthetic effect Effects 0.000 description 6
- 239000003225 biodiesel Substances 0.000 description 5
- 230000007423 decrease Effects 0.000 description 5
- 238000002073 fluorescence micrograph Methods 0.000 description 5
- FGIUAXJPYTZDNR-UHFFFAOYSA-N potassium nitrate Chemical compound [K+].[O-][N+]([O-])=O FGIUAXJPYTZDNR-UHFFFAOYSA-N 0.000 description 4
- 239000002994 raw material Substances 0.000 description 4
- 241000196252 Ulva Species 0.000 description 3
- 241000157352 Uncaria Species 0.000 description 3
- 230000003247 decreasing effect Effects 0.000 description 3
- 230000001788 irregular Effects 0.000 description 3
- 230000010355 oscillation Effects 0.000 description 3
- 244000251953 Agaricus brunnescens Species 0.000 description 2
- 235000001674 Agaricus brunnescens Nutrition 0.000 description 2
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- 241000208811 Flaveria Species 0.000 description 2
- 241000233866 Fungi Species 0.000 description 2
- 241000157373 Uncaria rhynchophylla Species 0.000 description 2
- 230000000844 anti-bacterial effect Effects 0.000 description 2
- 230000033228 biological regulation Effects 0.000 description 2
- 235000014113 dietary fatty acids Nutrition 0.000 description 2
- 239000008157 edible vegetable oil Substances 0.000 description 2
- 238000002474 experimental method Methods 0.000 description 2
- 229930195729 fatty acid Natural products 0.000 description 2
- 239000000194 fatty acid Substances 0.000 description 2
- 150000004665 fatty acids Chemical class 0.000 description 2
- 238000009730 filament winding Methods 0.000 description 2
- 238000002955 isolation Methods 0.000 description 2
- 238000011031 large-scale manufacturing process Methods 0.000 description 2
- 244000005700 microbiome Species 0.000 description 2
- 235000010333 potassium nitrate Nutrition 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 241000222518 Agaricus Species 0.000 description 1
- 241000894006 Bacteria Species 0.000 description 1
- 241001478778 Cladophora Species 0.000 description 1
- 235000004237 Crocus Nutrition 0.000 description 1
- 241000596148 Crocus Species 0.000 description 1
- 230000005526 G1 to G0 transition Effects 0.000 description 1
- 241001048891 Jatropha curcas Species 0.000 description 1
- 241000721605 Mougeotia Species 0.000 description 1
- OAICVXFJPJFONN-UHFFFAOYSA-N Phosphorus Chemical compound [P] OAICVXFJPJFONN-UHFFFAOYSA-N 0.000 description 1
- 235000019484 Rapeseed oil Nutrition 0.000 description 1
- 241000196294 Spirogyra Species 0.000 description 1
- 241000196301 Spirogyra sp. Species 0.000 description 1
- 241000199474 Tribonema Species 0.000 description 1
- 241000908225 Tribonema utriculosum Species 0.000 description 1
- 241000196253 Ulva prolifera Species 0.000 description 1
- 241000200212 Vaucheria Species 0.000 description 1
- 241001482202 Vaucheria hamata Species 0.000 description 1
- 241000206764 Xanthophyceae Species 0.000 description 1
- 238000009825 accumulation Methods 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- WORJEOGGNQDSOE-UHFFFAOYSA-N chloroform;methanol Chemical compound OC.ClC(Cl)Cl WORJEOGGNQDSOE-UHFFFAOYSA-N 0.000 description 1
- 230000001276 controlling effect Effects 0.000 description 1
- 238000012136 culture method Methods 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 238000000855 fermentation Methods 0.000 description 1
- 230000004151 fermentation Effects 0.000 description 1
- 235000013305 food Nutrition 0.000 description 1
- 239000002803 fossil fuel Substances 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 230000007246 mechanism Effects 0.000 description 1
- 230000004060 metabolic process Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 231100000252 nontoxic Toxicity 0.000 description 1
- 230000003000 nontoxic effect Effects 0.000 description 1
- 238000000643 oven drying Methods 0.000 description 1
- 229910052698 phosphorus Inorganic materials 0.000 description 1
- 239000011574 phosphorus Substances 0.000 description 1
- 235000020777 polyunsaturated fatty acids Nutrition 0.000 description 1
- 230000001105 regulatory effect Effects 0.000 description 1
- 230000000284 resting effect Effects 0.000 description 1
- 235000012424 soybean oil Nutrition 0.000 description 1
- 239000003549 soybean oil Substances 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 235000020238 sunflower seed Nutrition 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C11—ANIMAL OR VEGETABLE OILS, FATS, FATTY SUBSTANCES OR WAXES; FATTY ACIDS THEREFROM; DETERGENTS; CANDLES
- C11B—PRODUCING, e.g. BY PRESSING RAW MATERIALS OR BY EXTRACTION FROM WASTE MATERIALS, REFINING OR PRESERVING FATS, FATTY SUBSTANCES, e.g. LANOLIN, FATTY OILS OR WAXES; ESSENTIAL OILS; PERFUMES
- C11B1/00—Production of fats or fatty oils from raw materials
- C11B1/02—Pretreatment
- C11B1/04—Pretreatment of vegetable raw material
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02E—REDUCTION OF GREENHOUSE GAS [GHG] EMISSIONS, RELATED TO ENERGY GENERATION, TRANSMISSION OR DISTRIBUTION
- Y02E50/00—Technologies for the production of fuel of non-fossil origin
- Y02E50/10—Biofuels, e.g. bio-diesel
Landscapes
- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Oil, Petroleum & Natural Gas (AREA)
- Wood Science & Technology (AREA)
- Organic Chemistry (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
The invention provides a method for extracting grease from algae organisms, which comprises the following steps: setting culture parameters of the uncinate algae, culturing the uncinate algae of the uncinate algae according to the culture parameters, and extracting grease from the uncinate algae of the uncinate algae after the culture is finished. According to the invention, the uncinate algae is selected for grease extraction, and because the uncinate algae can generate algae ball growth stress effect in the growth process, the characteristics of the uncinate algae break through the contradiction problem between stress conditions and grease yield when microalgae produce oil, and meanwhile, the characteristics of large filamentous algae also reduce risks such as bacterial pollution and high harvesting cost.
Description
Technical Field
The invention relates to the technical field of bioscience, in particular to a method for extracting grease from algae.
Background
With the increasing energy demands of the world in recent years, fossil fuels are exhausted, and renewable energy sources are increasingly focused. Biodiesel has been used as an environment-friendly alternative energy source which is sulfur-free, nontoxic and biodegradable. The first generation biodiesel mainly takes edible vegetable oil as raw materials, such as soybean oil, rapeseed oil, sunflower seed oil and the like. The second generation biodiesel mainly takes non-edible oil as raw materials, such as jatropha curcas, rape and the like. However, these raw materials require large areas of land planting, which is impractical for highly urban countries.
Under this demand, scientists have found that some microorganisms occupy little space, have high lipid content, and do not require large areas of agricultural land due to their unique short growth cycle. Microbiologically, oleaginous microorganisms can be divided into microalgae, fungi (mold and yeast) and bacteria, which have poor lipid-producing ability and only synthesize specific lipids and polyunsaturated fatty acids. Thus, microalgae and fungi are considered to be major grease producers.
At present, the filamentous microalgae for producing oil at home and abroad are mainly concentrated in the phylum xanthophyta, and the filamentous microalgae for producing oil are few in species, but recently, the filamentous microalgae are paid attention to gradually. Wang et al found a strain of filamentous oleaginous microalgae, tribonema sp, which can be used as a source of biodiesel. Xu Zijun and the like are studied on the growth condition, the oil content and the fatty acid composition and content of the flaveria saccorica (Tribonema utriculosum SAG 22.94.22.94), and the result shows that the oil content and the fatty acid content of the flaveria saccorica are very high. Therefore, the filamentous algae has higher research value and development potential in the production of raw materials of biodiesel and food oil, and the characteristics of high lipid content, easy harvesting, strong antibacterial property and the like, and the filamentous oleaginous algae can become a break for large-scale production research of oleaginous algae.
Disclosure of Invention
The invention provides a method for extracting grease from algae, which provides a new direction for algae oil production and grease accumulation regulation and control and lays a foundation for further revealing the oil production metabolism mechanism and industrialized culture of the uncinate algae because the uncinate algae has the characteristics of higher grease content, strong antibacterial and anti-predation capability, low collection cost and the like.
The invention provides a method for extracting grease from algae organisms, which comprises the following steps:
setting culture parameters of the uncinate algae;
culturing the uncinate algae according to the culture parameters, and extracting oil from the uncinate algae after the culture is finished.
Further, the culture parameters include a culture temperature, the culture temperature being 20 ℃.
Further, the culture parameters include an illumination intensity of 20. Mu. Mol.m -2 ·s -1 。
Further, the culture parameters include an initial density of 2 g.L -1 。
Further, the culture parameters include generalA ventilation amount of 1 V.V -1 ·min -1 。
Further, the culture parameters include a culture speed of 160rpm.
Further, the culture parameters include KNO3 concentration, the KNO3 concentration is 0.05g.L -1 。
Further, the culture parameters include K 2 HPO 4 Concentration of said K 2 HPO 4 The concentration is 20 mg.L -1 。
Further, the culture parameters include algae size, which is 40mg per ball -1 。
Further, the grease content of the uncinate algae is more than or equal to 30%.
According to the invention, the uncinate algae is selected for grease extraction, and because the uncinate algae can generate algae ball growth stress effect in the growth process, the characteristics of the uncinate algae break through the contradiction problem between stress conditions and grease yield when microalgae produce oil, and meanwhile, the characteristics of large filamentous algae also reduce risks such as bacterial pollution and high harvesting cost.
Drawings
FIG. 1 is a photograph of a uncinate algae under a microscope according to an embodiment of the present invention;
FIG. 2a shows biomass of the uncinate algae provided by the embodiment of the invention at different temperatures;
fig. 2b shows the morphology of the uncinate algae provided by the embodiment of the invention at different temperatures;
FIG. 2c shows total lipid content of the uncinate algae provided by the embodiment of the invention at different temperatures;
FIG. 2d shows total lipid yields of the Uncaria agaricus bisporus at different temperatures according to the present invention;
fig. 2e is a fluorescence image of grease of the uncinate algae provided by the embodiment of the invention at different temperatures;
FIG. 3a shows biomass of the uncinate algae provided by the embodiment of the invention under different illumination intensities;
fig. 3b shows the morphology of the uncinate algae provided by the embodiment of the invention under different illumination intensities;
FIG. 3c shows total lipid content of the uncinate algae provided by the embodiment of the invention under different illumination intensities;
FIG. 3d shows total lipid yields of the Uncaria gambir aphanidermatum provided by the example of the present invention under different illumination intensities;
fig. 3e is a fluorescence image of grease under different illumination intensities of the uncinate algae provided by the embodiment of the invention;
FIG. 4a is a graph showing biomass of the uncinate algae provided by the embodiment of the invention at different initial densities;
FIG. 4b shows the morphology of the uncinate algae according to the embodiment of the present invention at different initial densities;
FIG. 4c shows total lipid content of the uncinate algae provided by the embodiment of the invention at different initial densities;
FIG. 4d shows total lipid yields of the uncinate algae provided by the examples of the present invention at different initial densities;
fig. 4e is a fluorescence image of grease of the uncinate algae provided by the embodiment of the invention under different initial densities;
FIG. 5a shows biomass of the Uncaria rhynchophylla in different culture modes according to the embodiment of the invention;
FIG. 5b shows the morphology of the uncinate algae according to the embodiment of the present invention under different cultivation modes;
FIG. 5c shows the total lipid content of the uncinate algae according to the embodiment of the invention in different culture modes;
FIG. 5d shows total lipid yields of the Uncaria rhynchophylla according to the example of the present invention in different culture modes;
FIG. 5e is a fluorescence image of the oil of the uncinate algae provided by the embodiment of the invention in different culture modes;
FIG. 6a shows different KNO of the uncinate algae according to the embodiment of the present invention 3 Biomass at concentration;
FIG. 6b shows different KNO of the uncinate algae according to the embodiment of the present invention 3 Algae morphology at concentration;
FIG. 6c shows different KNO of the uncinate algae according to the embodiment of the present invention 3 Total lipid content at concentration;
FIG. 6d shows different KNO of the uncinate algae according to the embodiment of the present invention 3 Total lipid yield at concentration;
FIG. 6e shows different KNO of the uncinate algae according to the embodiment of the present invention 3 Grease fluorescence pictures under concentration;
FIG. 7a shows different K values of the uncinate algae according to the embodiment of the present invention 2 HPO 4 Biomass at concentration;
FIG. 7b shows different K values of the uncinate algae according to the embodiment of the present invention 2 HPO 4 Algae morphology at concentration;
FIG. 7c shows different K values of the uncinate algae according to the embodiment of the present invention 2 HPO 4 Total lipid content at concentration;
FIG. 7d shows different K values of the uncinate algae according to the embodiment of the present invention 2 HPO 4 Total lipid yield at concentration;
FIG. 7e shows different K values of the uncinate algae according to the embodiment of the present invention 2 HPO 4 Grease fluorescence pictures under concentration;
FIG. 8a shows biomass of the uncinate algae provided by the embodiment of the invention under different algae ball sizes;
FIG. 8b shows the morphology of the uncinate algae according to the embodiment of the present invention under different algae sizes;
FIG. 8c shows total lipid content of the uncinate algae provided by the embodiment of the invention under different algae ball sizes;
FIG. 8d shows total lipid yields of the Uncaria agaricus bisporus of the present invention under different algae ball sizes;
fig. 8e is a fluorescence image of grease of the uncinate algae provided by the embodiment of the invention under different algae ball sizes.
Detailed Description
For the purpose of making the objects, technical solutions and advantages of the present invention more apparent, the technical solutions in the embodiments of the present invention will be clearly and completely described in the following in conjunction with the embodiments of the present invention, and it is apparent that the described embodiments are some embodiments of the present invention, but not all embodiments. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
Example 1
According to the invention, 7 kinds of filamentous algae (Cladophora sp.), spirogyra (Spirogyra sp.), enteromorpha (Enteromorpha prolifera), enteromorpha (Enteromorpha sp.), aphanidermatum (Mougeotia sp.), rhynchophylla (Vaucheria hamata), and multiple-egg aphaniderm (Vaucheria Walzi) are collected in the wild, and the total lipid content of the rhynchophylla is measured by adopting chloroform methanol fermentation, and the total lipid content of the rhynchophylla is measured to be the highest and reaches 30%, which means that the lipid content of the rhynchophylla is higher than that of other filamentous algae, and the total lipid content of the rhynchophylla is large-sized filamentous algae, so that the problem of too high collection cost in the large-scale production process of the oleaginous microalgae can be solved, and the growth speed and the lipid content of the oleaginous microalgae can be further improved by only regulating and controlling the culture conditions.
Example 2 culture of Uncaria gambir aphanidermatum and extraction of oil and fat under different temperature culture conditions
In this example, four temperature gradients were set, 12℃at 16℃at 20℃at 24℃and the culture of uncinate algae was carried out at different temperatures by aeration and illumination, the results are shown in FIG. 2a and FIG. 2 b. a-D in fig. 2 b: the forms of algae are respectively at the 7 th, 14 th, 21 th and 28 th days under the culture condition of 12 ℃; E-H: the forms of algae are respectively at the 7 th, 14 th, 21 th and 28 th days under the culture condition of 16 ℃; I-L: the forms of algae are respectively at the 7 th, 14 th, 21 th and 28 th days under the culture condition of 20 ℃; M-P: the algal forms were obtained at days 7, 14, 21 and 28 under the culture conditions of 24 ℃. The observation shows that the growth speed of the uncinate algae is slow under the condition of 12 ℃, the algae body is in random flocculence, when the temperature reaches 20 ℃, the growth speed reaches the highest, the algae body is in an ellipsoid, the density of the algae body is increased, and when the temperature is increased to 24 DEG CWhen the growth rate of the uncinate algae is reduced. In this experiment, the biomass reached the highest at 20℃for 14 days, 4.66 g.multidot.L -1 The temperature is increased by 122.32 percent relative to the temperature at 16 ℃. At the moment, the ordering of the rhynchophylla-free biomass at other temperatures is sequentially 24 ℃ to 16 ℃ to 12 ℃ which are 3.214 g.L respectively -1 、2.096g·L -1 、0.9865g·L -1 . In conclusion, the growth state of the uncinate algae is optimal under the ventilation condition at 20 ℃.
Collecting fixed phase unchecked algae under different temperature conditions, oven drying, extracting total lipid, and measuring content. The total lipid content, yield measurement results and algae filament lipid fluorescence shooting results are shown in fig. 2c, 2D and 2e, and in fig. 2e, A-D: grease fluorescent pictures at the 7 th, 14 th, 21 th and 28 th days under the culture condition of 12 ℃ respectively; E-H: grease fluorescent pictures at the 7 th, 14 th, 21 th and 28 th days under the culture condition of 16 ℃ respectively; I-L: grease fluorescent pictures at the 7 th, 14 th, 21 th and 28 th days under the culture condition of 20 ℃ respectively; M-P: grease fluorescence pictures at the 7 th, 14 th, 21 th and 28 th days under the culture condition of 24 ℃ respectively. The observation shows that under the action of different temperatures, the total lipid content of the uncinate algae of the rhynchophylla gradually rises along with the temperature rise, reaches the highest value of 42.25 percent at 20 ℃, then the lipid content is reduced along with the temperature rise, and simultaneously the variation trend of the total lipid yield and the total lipid content is basically consistent along with the temperature rise, and the highest value of 14.08 mg.L at 20 DEG C -1 ·d -1 . In conclusion, the temperature of 20 ℃ is more suitable for oil production culture of the uncinate algae.
Example 3 cultivation of Uncaria gambir aphanidermatum under different illumination intensity cultivation conditions and oil extraction
The present example sets 4 illumination intensities of 10. Mu. Mol.m -2 ·s -1 ，20μmol·m -2 ·s -1 ，60μmol·m -2 ·s -1 ，120μmol·m -2 ·s -1 Aeration photosynthetic culture is carried out on the uncinate algae under different illumination conditions, and the results are shown in fig. 3a and 3b, and A-D in fig. 3b are as follows: respectively 10 mu mol m -2 ·s -1 Grease fluorescent pictures on days 7, 14, 21 and 28 under the illumination condition; E-H: 20 mu mol m respectively -2 ·s -1 Grease fluorescent pictures on days 7, 14, 21 and 28 under the illumination condition; I-L: 60 mu mol m respectively -2 ·s -1 Grease fluorescent pictures on days 7, 14, 21 and 28 under the illumination condition; M-P is 120 mu mol M respectively 2 ·s -1 Grease fluorescence pictures on days 7, 14, 21 and 28 under illumination conditions. As a result of observation, the content of the uncinate algae was 10. Mu. Mol.m -2 ·s -1 The growth speed is slow under the illumination condition, and is 20 mu mol.m -2 ·s -1 The growth speed is the fastest under the condition that the algae bodies are compact ellipsoids and are 60 mu mol.m -2 ·s -1 And 120. Mu. Mol.m -2 ·s -1 The growth speed is higher under the illumination condition, and the algae is loose flocculent. In the first 14 days of the culture process, the growth speed of the uncinate algae and the non-spaced algae of the uncinate algae is firstly increased and then reduced along with the increase of the light intensity, and is 20 mu mol.m -2 ·s -1 At a maximum of 4.66 g.L -1 The biomass sizes under other illumination conditions are sequentially ordered as follows: 60 mu mol m -2 ·s -1 ＞120μmol·m -2 ·s -1 ＞10μmol·m -2 ·s -1 Biomass was 3.44 g.L -1 、3.14g·L -1 、1.6g·L -1 . In conclusion, 20. Mu. Mol.m -2 ·s -1 The conditions are more suitable for the biomass increase of the rhynchophylla.
Collecting the uncinate algae under different illumination conditions, drying, extracting total fat and measuring the content. The total lipid content, yield measurement result and fluorescence shooting result of the algae filament oil are shown in FIG. 3c, FIG. 3D and FIG. 3e, wherein A-D in FIG. 3e are respectively 10 mu mol.m -2 ·s -1 Grease fluorescent pictures at 7 th, 14 th, 21 th and 28 th days under culture conditions, wherein E-H is 20 mu mol.m respectively -2 ·s -1 Grease fluorescent pictures on days 7, 14, 21 and 28 under culture conditions, wherein I-L are respectively 60 mu mol.m -2 ·s -1 Grease fluorescent pictures on days 7, 14, 21 and 28 under culture conditions, wherein M-P is 120 mu mol.m respectively -2 ·s -1 Grease fluorescence pictures on days 7, 14, 21 and 28 under culture conditions. The observation shows that under different illumination conditions, the total lipid content of the uncinate algae of the rhynchophylla increases firstly with the increase of the illumination intensity, and is 20 mu mol.m -2 ·s -1 At a maximum of 48.76%, and then the total lipid content decreases as the light intensity continues to rise, with the lipid content under other lighting conditions being 27.16% (10. Mu. Mol. M) -2 ·s -1 )、24.38(60μmol·m -2 ·s -1 )、24.78％(120μmol·m -2 ·s -1 ) The total lipid yield of the uncinate algae is increased firstly along with the increase of the illumination intensity and is 20 mu mol.m -2 ·s -1 At the highest value of 16.26 mg.L -1 ·d -1 . Subsequently, total lipid yield decreased with increasing light intensity, and total lipid yield was ranked under other light conditions: 60 mu mol m -2 ·s -1 ＞120μmol·m -2 ·s -1 ＞10μmol·m -2 ·s -1 6.68 mg.L -1 ·d -1 、4.48mg·L -1 ·d -1 、3.24mg·L -1 ·d -1 . In conclusion, the illumination intensity is 20 mu mol.m -2 ·s -1 Is more suitable for the oleaginous culture of the uncinate algae.
Example 4 culture and oil extraction of Uncaria agaricus in culture conditions of different initial Density
The present example sets 3 initial densities of 0.5 g.L -1 ，1g·L -1 ，2g·L -1 Aeration photosynthetic culture was performed on the anocelia crocus under different initial densities, and the results are shown in fig. 4a and 4b, and a-E in fig. 4 b: initial density of 0.5 g.L -1 Algae forms on days 3, 6, 9, 12, 15 under culture conditions; f-J: initial density of 1 g.L -1 Algae forms on days 3, 6, 9, 12, 15 under culture conditions; k-N: respectively the initial density is 2 g.L -1 Algal morphology at days 3, 6, 9, 12, 15 under culture conditions. The observation shows that the biomass growth times of the uncinate algae are gradually reduced along with the increase of the initial density, and are respectively 6.90 times, 5.33 times and 4.03 times, the algae are ellipsoidal in the first 6 days, then the algae are mutually wound and combined to form larger irregular nodular algae, and the algae are easier to combine with each other as the initial density is higher. However, from the viewpoint of biomass, the biomass of the uncinate algae is positively related to the initial density in the culture process, and the living beingsThe quantities and sizes are sequentially ordered as follows: 2 g.L -1 ＞1g·L -1 ＞0.5g·L -1 Maximum biomass of 8.06 g.L respectively -1 、5.33g·L -1 、3.45g·L -1 In summary, the initial density is 2g.L -1 The conditions are more suitable for oleaginous culture of the uncinate algae of the rhynchophylla.
Collecting the uncinate algae under different initial density conditions, drying, extracting total fat and measuring the content. The total lipid content, yield measurement results and algae filament lipid fluorescence photographing results are shown in fig. 4c, 4d and 4E, and in fig. 4E, A-E: initial density of 0.5 g.L -1 Grease fluorescence pictures on days 3, 6, 9, 12 and 15 under culture conditions; f-J: initial density of 1 g.L -1 Grease fluorescence pictures on days 3, 6, 9, 12 and 15 under culture conditions; k-O: respectively the initial density is 2 g.L -1 Grease fluorescence pictures on days 3, 6, 9, 12 and 15 under culture conditions. It was found that the total lipid content of the uncinate algae decreased with increasing initial density under different initial density conditions, and the lipid content was 50.19% (0.5 g.L) -1 )、44.92％(1g·L -1 )，40.61％(2g·L -1 ) However, the total lipid yield of the uncinate algae increases with the increase of the initial density, namely 11.55mg.L -1 ·d -1 、15.96mg·L -1 ·d -1 、27.28mg·L -1 ·d -1 . In conclusion, the initial density is 2g.L -1 The method is more suitable for oil production culture of the uncinate algae.
Example 5 cultivation of Uncaria gambir aphanidermatum and extraction of oil and fat in different cultivation modes
In this example, three culture methods of resting, aeration and shaking are adopted, and aeration rate is set to 1 V.V -1 ·min -1 And 2 V.V -1 ·min -1 Two gradients, the shaking rotation speed is set to be 120rpm and 160rpm, and the result of photosynthetic culture is shown in FIG. 5a and FIG. 5b, and A-E in FIG. 5 b: the ventilation rates are respectively 0 V.V -1 ·min -1 Algae forms on days 3, 6, 9, 12, 15 under culture conditions; f-J: the ventilation rates are respectively 1 V.V -1 ·min -1 3 rd, 6 th, 9 th, 12 th, 15 th under culture conditionsThe shape of the algae on the sky; k-N: the ventilation rates are respectively 2 V.V -1 ·min -1 Algae forms on days 3, 6, 9, 12, 15 under culture conditions; P-T: the algae forms at the 3 rd, 6 th, 9 th, 12 th and 15 th days under the culture condition of the shaking rotation speed of 120rpm respectively; U-Y: the algal forms were obtained on days 3, 6, 9, 12 and 15 under culture conditions of aeration rate oscillation at 160rpm, respectively. As a result of observation, the ventilation was 0 V.V -1 ·min -1 When the algae cells grow slowly, only some algae filaments grow, the algae filaments among the algae bodies are entangled in the growth period, but the algae body intervals are clear, and the algae bodies are not combined to form one algae body. When the ventilation is 1 V.V -1 ·min -1 And 2 V.V -1 ·min -1 When the algae are in a spherical shape independently in a hysteresis period, and then in a growth period, the algae are intertwined and combined with each other to form larger irregular nodular algae. The growth rate of the non-compartmental algae is ordered to be 1 V.V -1 ·min -1 ＞2V·V -1 ·min -1 ＞0V·V -1 ·min -1 Maximum biomass in stationary phase of 8.06 g.L respectively -1 、7.55g·L -1 、1.74g·L -1 . Under the oscillation condition, the uncinate algae basically grow independently, and the inter-algal filament winding of the uncinate algae rarely occurs. When the rotation speed is 120rpm, the biomass of the uncinate algae of the hooked yeast reaches the maximum value of 5.54 g.L at the 12 th day of culture -1 . When the rotation speed is 160rpm, the biomass of the uncinate algae of the hooked yeast reaches the maximum value of 2.98 g.L at 15 days of culture -1 . In summary, the ventilation is 1 V.V -1 ·min -1 The condition is more suitable for the biomass culture of the uncinate algae.
Collecting the uncinate algae of the uncinate algae in different culture modes, and carrying out total fat extraction and content measurement after drying. The total lipid content, yield measurement results and algae filament lipid fluorescence photographing results are shown in fig. 5c, 5d and 5E, and in fig. 5E, A-E: the ventilation rates are respectively 0 V.V -1 ·min -1 Grease fluorescence pictures on days 3, 6, 9, 12 and 15 under culture conditions; f-J: the ventilation rates are respectively 1 V.V -1 ·min -1 Grease fluorescence pictures on days 3, 6, 9, 12 and 15 under culture conditions; k-N: the ventilation rates are respectively 2 V.V -1 ·min -1 3 rd, 6 th, 9 th, 12 th, 1 st under culture conditionsGrease fluorescent pictures for 5 days; P-T: grease fluorescent pictures at 3 rd, 6 th, 9 th, 12 th and 15 th days under the culture condition of the shaking rotation speed of 120rpm respectively; U-Y: grease fluorescence pictures at 3 rd, 6 th, 9 th, 12 th and 15 th days under the culture condition of aeration rate oscillation rotation speed of 160rpm are respectively. It was observed that the total lipid content of the rhynchophylla increased with increasing ventilation under ventilation conditions, 30.00%, 40.61% and 48.10%, respectively. The total lipid yield increased with increasing ventilation and then decreased, respectively, was 3.61 mg.L -1 ·d -1 、27.28mg·L -1 ·d -1 、25.30mg·L -1 ·d -1 . Under the shaking condition, the total lipid content of the uncinate algae of the hooked yeast is 36.44 percent and 68.55 percent respectively at 120rpm and 160rpm, and the total lipid yield is 16.85 mg.L respectively -1 ·d -1 And 13.67 mg.L -1 ·d -1 . In summary, both high ventilation and high rotational speed are advantageous for increasing grease content without marine algae, but ventilation is 1 V.V when biomass and grease yield are taken into consideration -1 ·min -1 The method is more suitable for oil production culture of the uncinate algae.
Example 6 uncinate algae with different concentrations KNO 3 Culturing under the condition and extracting oil
The embodiment sets 5 KNO 3 Concentration gradient of 0 g.L -1 ，0.01g·L -1 ，0.05g·L -1 ，0.1g·L -1 ，0.2g·L -1 Different KNO is carried out on the uncinate algae of the hooked yeast 3 Aeration photosynthetic culture under concentration conditions, results are shown in FIG. 6a and FIG. 6b, A-D in FIG. 6 b: respectively KNO 3 Concentration of 0 g.L -1 Algal morphology at 3, 6, 9, 12 days under culture conditions; E-H: respectively KNO 3 Concentration of 0.01 g.L -1 Algal morphology at 3, 6, 9, 12 days under culture conditions; I-L: respectively KNO 3 Concentration of 0.05 g.L -1 Algal morphology at 3, 6, 9, 12 days under culture conditions; M-P: respectively KNO 3 Concentration is 0.1 g.L -1 Algal morphology at 3, 6, 9, 12 days under culture conditions; Q-T: respectively KNO 3 Concentration of 0 g.L -1 Algal morphology at days 3, 6, 9, 12 under culture conditions. As can be seen from the observation, the uncinate algae is 0 g.L -1 KNO 3 The growth speed is slow under the condition that the algae wires are wound between algae bodies to form irregular floccules, and when KNO 3 The concentration is increased to 0.1 g.L -1 When the growth speed is increased, the algae body is not integrated, and the algae body is treated as KNO 3 The concentration is 0.05-0.2 g.L -1 When the algae grow, the algae are combined into a whole body in a lump or strip shape. With KNO 3 The concentration is increased, the growth speed of the uncinate algae of the rhynchophylla is increased and then reduced, and the maximum biomass is 4.04 g.L respectively -1 、5.05g·L -1 、7.13g·L -1 、8.061g·L -1 、3.73g·L -1 However, in the present experiment, it was found that the content of the uncinate algae was 0.05g.L -1 KNO 3 Under the conditions, the highest biomass was reached at day 9 of culture. Is better than 0.1 g.L in terms of biological growth rate -1 KNO 3 Conditions. In conclusion, 0.05 g.L -1 KNO 3 The conditions are more suitable for the biomass increase of the rhynchophylla.
Collecting different KNOs 3 The hooked yeast under the concentration condition has no algae isolation, and the total fat extraction and the content measurement are carried out after the drying. The total lipid content, yield measurement results and algae filament lipid fluorescence photographing results are shown in fig. 6c, 6D and 6e, and in fig. 6e, A-D: respectively KNO 3 Concentration of 0 g.L -1 Grease fluorescence pictures on 3 rd, 6 th, 9 th and 12 th days under culture conditions; E-H: respectively KNO 3 Concentration of 0.01 g.L -1 Grease fluorescence pictures on 3 rd, 6 th, 9 th and 12 th days under culture conditions; I-L: respectively KNO 3 Concentration of 0.05 g.L -1 Grease fluorescence pictures on 3 rd, 6 th, 9 th and 12 th days under culture conditions; M-P: respectively KNO 3 Concentration is 0.1 g.L -1 Grease fluorescence pictures on 3 rd, 6 th, 9 th and 12 th days under culture conditions; Q-T: respectively KNO 3 Concentration of 0 g.L -1 Grease fluorescence pictures on days 3, 6, 9 and 12 under culture conditions. As can be seen from the observation, at different KNOs 3 Under the concentration condition, the total lipid content of the uncinate algae is along with the illumination intensity KNO 3 The concentration rise is increased first, at 0.05 g.L -1 Reaches a maximum value of 59.47%, and then the total lipid content is along with KNO 3 The concentration continues to rise and decrease, other KNOs 3 The grease content under the concentration condition is 39.33 percent (0 g.L) -1 )、35.78％(0.01g·L -1 )、40.61％(0.1g·L -1 )、36.70％(0.2g·L -1 ) The total lipid yield of the uncinate algae is increased along with the increase of the illumination intensity and is 0.05g.L -1 When the maximum value reaches 29.75mg.L -1 ·d -1 The total lipid yield then follows KNO 3 The concentration is increased and reduced, other KNO 3 Yield under the concentration condition is 11.66 mg.L respectively -1 ·d -1 (0g·L -1 )、20.99mg·L -1 ·d -1 (0.01g·L -1 )、27.28mg·L -1 ·d -1 (0.1g·L -1 )、11.55mg·L -1 ·d -1 (0.5g·L -1 ). In conclusion, 0.05 g.L - 1 KNO 3 The conditions are more suitable for the oleaginous culture of the uncinate algae.
EXAMPLE 7 Uncaria gambir aphanidermatum at different concentrations K 2 HPO 4 Culturing under the condition and extracting oil
The present embodiment sets 4K 2 HPO 4 Concentration gradient of 0 mg.L -1 ，10mg·L -1 ，20mg·L -1 ，40mg·L -1 Different K is carried out on the uncinate algae of the hooked yeast 2 HPO 4 Aeration photosynthetic culture under concentration conditions, results are shown in FIG. 7a and FIG. 7b, A-E in FIG. 7 b: respectively K 2 HPO 4 Concentration of 0 mg.L -1 Algae forms on days 3, 6, 9, 12, 15 under culture conditions; f-J: respectively K 2 HPO 4 Concentration of 10mg.L -1 Algae forms on days 3, 6, 9, 12, 15 under culture conditions; k-N: respectively K 2 HPO 4 Concentration of 20 mg.L -1 Algae forms on days 3, 6, 9, 12, 15 under culture conditions; P-T: respectively is K 2 HPO 4 Concentration of 40 mg.L -1 Algal morphology at days 3, 6, 9, 12, 15 under culture conditions. As can be seen from the observation, the uncinate algae is 0m g.L -1 K 2 HPO 4 The growth speed is slow under the condition, the algae filament winding among algae bodies is less, and the algae filament basically takes a small lump shape. When K is 2 HP0 4 The concentration is increased to 10 mg.L -1 -20mg·L -1 When the growth speed is increased, the algae is wound gradually into a wholeThe whole of each cluster is in a cluster shape or a strip shape. When K is 2 HPO 4 The concentration is increased to 40 mg.L -1 In this case, the growth rate is rather lowered. With K 2 HPO 4 The concentration is increased, the growth speed of the uncinate algae of the rhynchophylla is increased and then reduced, and the maximum biomass under each condition is 4.30 g.L respectively -1 、6.17g·L -1 、8.06g·L -1 、5.10g·L -1 In summary, 20 mg.L -1 K 2 HPO 4 The conditions are more suitable for the biomass increase of the rhynchophylla.
Collecting different K 2 HPO 4 The hooked yeast under the concentration condition has no algae isolation, and the total fat extraction and the content measurement are carried out after the drying. The total lipid content, yield measurement results and algae filament lipid fluorescence photographing results are shown in fig. 7c, 7d and 7E, and in fig. 7E, a-E: respectively K 2 HPO 4 Concentration of 0 mg.L -1 Grease fluorescence pictures on days 3, 6, 9, 12 and 15 under culture conditions; f-J: respectively K 2 HPO 4 Concentration of 10mg.L -1 Grease fluorescence pictures on days 3, 6, 9, 12 and 15 under culture conditions; k-N: respectively K 2 HPO 4 Concentration of 20 mg.L -1 Grease fluorescence pictures on days 3, 6, 9, 12 and 15 under culture conditions; P-T: respectively is K 2 HPO 4 Concentration of 40 mg.L -1 Grease fluorescence pictures on days 3, 6, 9, 12 and 15 under culture conditions. From observations, it can be seen that at different K 2 HPO 4 Under the concentration condition, the total lipid content of the uncinate algae is along with the illumination intensity K 2 HPO 4 The concentration rise is increased first, at 20 mg.L -1 At a maximum of 40.61% and then the total lipid content with K 2 HPO 4 The concentration continues to rise and decreases, other K 2 HPO 4 The grease content under the concentration condition is 28.21 percent (0 mg.L) -1 )、33.05％(10mg·L -1 )26.95％(40mg·L -1 ) The total lipid yield of the uncinate algae is increased along with the increase of the illumination intensity and is 20 mg.L -1 At the highest value of 27.28 mg.L -1 ·d -1 The total lipid yield then follows K 2 HPO 4 The concentration continues to rise and decreases, other K 2 HPO 4 Under the concentration conditionThe rates were 10.12 mg.L -1 ·d -1 、16.99mg·L -1 ·d -1 、9.16mg·L -1 ·d -1 . In conclusion, 20 mg.L -1 K 2 HP0 4 The conditions are more suitable for the oil production culture of the uncinate algae.
Example 8 culture of Uncaria gambir and oil extraction under different algal ball size culture conditions
In this example, 4 algae balls were set to have a gradient of 20mg per ball -1 、40mg·per ball -1 、80mg·per ball -1 、200mg·per ball -1 Aeration photosynthetic culture is carried out on the uncinate algae under the condition of different algae sizes, and the results are shown in fig. 8a and 8b, and A-E in fig. 8 b: the algae balls are respectively 20mg per ball -1 Algae forms on days 3, 6, 9, 12, 15 under culture conditions; f-J: the algae balls are respectively 40mg per ball -1 Algal forms at the following days 3, 6, 9, 12, 15; k-N: the algae balls are respectively 80mg per ball -1 Algae forms on days 3, 6, 9, 12, 15 under culture conditions; P-T: the algae balls are respectively 200mg per ball -1 Algal morphology at days 3, 6, 9, 12, 15 under culture conditions. The observation shows that the growth speed of the uncinate algae is basically the same in the growth period under the condition of different algae ball sizes, and the algae filaments are wound in small clusters among the algae bodies and are gradually combined into a whole in the stable period. Maximum biomass under each condition is 7.17 g.L -1 、7.00g·L -1 、8.06g·L -1 、6.67g·L -1 In summary, 80mg per ball -1 The conditions are more suitable for the biomass increase of the rhynchophylla.
Collecting uncinate algae of the hooked yeast under different algae ball sizes, drying, extracting total fat and measuring content. The total lipid content, yield measurement results and algae filament lipid fluorescence photographing results are shown in fig. 8c, 8d and 8E, and in fig. 8E, a-E: the algae balls are respectively 20mg per ball -1 Grease fluorescence pictures on days 3, 6, 9, 12 and 15 under culture conditions; F-J: the algae balls are respectively 40mg per ball -1 Grease fluorescence pictures on the following 3 rd, 6 th, 9 th, 12 th and 15 th days; K-N: the algae balls are respectively 80mg per ball -1 Grease fluorescence pictures on days 3, 6, 9, 12 and 15 under culture conditions; P-T: the algae balls are respectively 200mg per ball -1 Grease fluorescence pictures on days 3, 6, 9, 12 and 15 under culture conditions. The observation shows that under the condition of different algae sizes, the total lipid content of the uncinate algae increases with the increase of the algae size, and the total lipid content is 40mg per ball -1 The maximum value reaches 49.57%, then the total lipid content is reduced along with the continuous increase of the algae ball size, and the lipid content under the condition of other algae ball sizes is 40% (20 mg per ball respectively -1 )、40.61％(80mg·per ball -1 )、34.77％(200mg·per ball -1 ) The total lipid yield of the uncinate algae is increased along with the increase of the algae ball size, and is 40mg per ball -1 At the highest value of 33.32 mg.L -1 ·d -1 Then the total lipid yield decreases as the algae size continues to rise, and the yields under the condition of other algae sizes are 25.96 mg.L respectively -1 ·d -1 、27.28mg·L -1 ·d -1 、23.67mg·L -1 ·d -1 . In summary, 40mg per ball -1 The conditions are more suitable for the oleaginous culture of the uncinate algae.
The present invention tabulates the oil yield and productivity of the above culture conditions, and as shown in table 1, under the field condition, the oil yield of the uncinate algae is 30%, while the culture conditions provided by the present invention all contribute to the improvement of the oil yield; in addition, vibration has important regulation and control effects on oil production of the uncinate algae of the rhynchophylla, and the nitrogen concentration, the initial density, the diameter of the algae balls, the illumination, the ventilation, the temperature and the phosphorus concentration are the next.
TABLE 1 optimal culture conditions provided in examples 1-8
Finally, it should be noted that: the above embodiments are only for illustrating the technical solution of the present invention, and not for limiting the same; although the invention has been described in detail with reference to the foregoing embodiments, it will be understood by those of ordinary skill in the art that: the technical scheme described in the foregoing embodiments can be modified or some or all of the technical features thereof can be replaced by equivalents; such modifications and substitutions do not depart from the spirit of the invention.
Claims (2)
1. A method of extracting grease from algae, comprising the steps of:
setting culture parameters of the uncinate algae;
the culture parameters comprise a culture temperature, wherein the culture temperature is 20 ℃;
the culture parameters comprise illumination intensity of 20 mu mol m -2 ·s -1 ；
The culture parameters include an initial density of g.L -1 ；
The culture parameters include ventilation of 1V.V -1 ·min -1 ；
The culture parameters comprise a culture rotating speed, wherein the culture rotating speed is 160 rpm;
the culture parameters comprise KNO 3 Concentration of KNO of 3 The concentration is 0.05 g.L -1 ；
The culture parameters include K 2 HPO 4 Concentration of said K 2 HPO 4 The concentration is 20 mg.L -1 ；
The culture parameters comprise the size of algae balls, and the size of algae balls is 40mg per ball -1 ；
Culturing the uncinate algae according to the culture parameters, and extracting oil from the uncinate algae after the culture is finished.
2. The method of claim 1, wherein the grease content of the uncinate algae is greater than or equal to 30%.
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