CN106520559A - High-efficiency light autotrophic culture method for chlorella - Google Patents
High-efficiency light autotrophic culture method for chlorella Download PDFInfo
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- CN106520559A CN106520559A CN201611231357.6A CN201611231357A CN106520559A CN 106520559 A CN106520559 A CN 106520559A CN 201611231357 A CN201611231357 A CN 201611231357A CN 106520559 A CN106520559 A CN 106520559A
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- chlorella
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M21/00—Bioreactors or fermenters specially adapted for specific uses
- C12M21/02—Photobioreactors
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/12—Unicellular algae; Culture media therefor
Abstract
The invention provides a high-efficiency light autotrophic culture method for chlorella. The method comprises the following steps: adding a culture solution into a photobioreactor, inoculating with chlorella, ensuring that the alga cell density is no less than 1 million/milliliter, and naturally illuminating, wherein the light intensity at day time is 10-130 Klx, the illumination period is 12/12, the air introduction amount is 1.0-10.0 L/min, the carbon dioxide gas introduction amount is 0.05-0.5 L/min, the alga solution temperature is 26-33 DEG C, and the pH value is kept at 7.0-9.0. According to the invention, high-efficiency large-scale production with the advantages of long continuous culture time, high yield and high nutritive salt utilization ratio is realized; and in the whole culture period, the collected alga dry basis is 22.131-23.181 kilograms, the alga protein content is up to 55% or above, and the nutritive salt utilization ratio is up to 90% or above.
Description
Technical field
The present invention relates to technical field of microalga biology, and in particular to a kind of chlorella high efficiency light autotrophy cultural method.
Background technology
Chlorella is the general natural disposition monoplast green alga of Chlorophyta Chlorella, is a kind of spherical, oval unicellular alga, directly
3～8 microns of footpath, is one of life earliest on the earth, before occurring in more than 20 hundred million years, is a kind of efficient photosynthetic plant, with light
Autophyting growth breeding is closed, is distributed all over the world, is moved in less shallow water more, also there are marine breeds.It is individual under natural endowment
It is less, artificial culture amount reproduction.Intracellular protein, fat and carbohydrate content are all very high, have various dimension lifes again
Element, edible and as bait.
Photosynthesis simply can be summarised as being mainly chlorophyllous plant cell and antibacterial containing photosynthetic pigments, in the sunlight
Using inorganic matters (CO2、H2O、H2S etc.) synthesis of organic substance (C6H12O6), and discharge oxygen (O2) or other materials (such as S etc.) mistake
Journey.Photosynthesis are the organic synthesis of the maximum carried out on the earth simultaneously.Energy from the sun to the earth is about 1.5 daily
×1022KJ, wherein about 1% is absorbed by photosynthetic organism, is converted into the chemical energy of molecular forms by photosynthesis, and is passed through food
Chain is utilized by other members of biosphere.Therefore, sunlight is the final source of nearly all biological required energy on the earth.
At present, according to chlorella metabolic type, the main training method of chlorella is divided into autotrophic type, simultaneous foster type and heterotrophism
Type.There is the shortcomings of culture density is low, growth cycle is long in autotrophy culture；Mixotrophic cultivation is present to be needed to provide ready-made organic carbon source
The energy that grows as which is originated the consumption problem of the caused energy, meanwhile, heterotrophism is transitioned into the autotrophy stage and easily causes frond
Discomfort, so as to cause frond occur a part of phenomena of mortality the shortcomings of；Heterotrophic culture is equally existed and consumes ready-made organic carbon source
Situation, and the nutrient substance accumulated in the chlorella cells of heterotrophism such as protein, chlorophyll, vitamin etc. are generally than certainly
The relevant material of foster chlorella cylinder accumulation is low.
The content of the invention
For the problem that above-mentioned 3 kinds of training methods are present, the present invention is provided in a kind of short time and reaches continuous incubation time
The high chlorella light autotrophy cultural method of length, high efficiency output, nutritive salt utilization rate, the cultural method comprise the steps：
(1) in tap water add nutritive salt fully to dissolve and obtain culture fluid, culture fluid is entered using ultraviolet sterilization device
Row sterilization processing, in making sterilized rear culture fluid, content of microorganisms is less than 100cfu/ milliliters；
(2) culture fluid is added in bioreactor, access chlorella, and ensure algae cell density >=1,000,000/milli
Rise, carry out light autotrophy culture；
(3) culture adopts natural lighting, and daytime, light intensity was in 10-130Klx, periodicity of illumination 12/12, blowing air amount 1.0-
10.0L/min, carbon dioxide intake 0.05-0.5L/min, algae solution temperature are 26-33 DEG C, and pH is maintained at 7.0-9.0；
(4), in incubation, start harvesting when intensity of illumination decays to 50% in algae solution, every time 50% algae solution of harvesting, then
Fresh medium is supplemented, such operation was repeated once every 1 day, and until chlorella culture enters phase of decline, culture terminates.
The method of the supplementary culture fluid is added using at the uniform velocity mode, and once addition finishes use time with algae solution circulation one
It is identical the time required to all.
The nutritive salt composition includes：Carbamide 0.38-0.47g/L, MgSO4·7H2O 0.25-0.5g/L, KH2PO4
0.095-0.12g/L, ferrous sulfate 0.005-0.006g/L, CaCl2·2H2O 0.02-0.03g/L, A5 liquid microelement 0.8-
1mL/L；Wherein A5 trace element formula of liquid is H3BO3 2.86g/L、MnCl2·4H2O 1.86g/L、ZnSO4·7H2O
0.22g/L、Na2MoO4·2H2O 0.39g/L、CuSO4·5H2O 0.08g/L and Co (NO3)2·6H2O 0.05g/L。
Chlorella to harvesting carries out being centrifuged, cleans, is dried acquisition dry algae powder, determines its algae butt；Frond internal protein
Assay is adopted《The measure of Protein in Food》(GB 5009.5 2010) Kjeldahl nitrogen determination；It is total in culture water body
Phosphorus is determined using the national standard method (GB/T for adding potassium peroxydisulfate (or nitric acid-perchloric acid) to aoxidize in neutral conditions
11893-89)；The national standard that determination of total nitrogen content is cleared up at 120-124 DEG C using alkaline chitinase, uv-spectrophotometric is quantitative
Method (GB/T11894-1989).
Compared with prior art, the present invention has the following advantages：(1) chlorella culture water is through sterilization treatment, its matter
Amount meets existing National Standard of the People's Republic of China《Drinking water sanitary standard》Relevant regulations；(2) improve bead
Algal nutrient formula so as to meet the optimal proportion of chlorella physiologically desired nutritional salt；(3) in light autotrophy incubation,
A part is harvested when intensity of illumination decays to 50% in algae solution, in remaining algae solution, adds fresh medium, single-cell algae again
Life continued, it is ensured that the vigor of frustule, improve frustule growth rate, extend continue cultivation cycle.This culture
Method realizes high efficiency large-scale production, and in 5.5 tons of bioreactors, chlorella continuously can be cultivated 20 days, from the 6th
It rises, and harvests once per 2 days, every time 50% algae solution of harvesting, and the algae solution to harvesting carries out being centrifuged, cleans, is dried acquisition dry algae powder,
Measure algae butt and frond internal protein content, whole culture period, the algae butt of harvesting is 25.203-25.226 kilogram, frond
Protein content reaches more than 69%, and nutritive salt utilization rate reaches more than 90%.
Specific embodiment
The present invention is further described by specific embodiment.
Embodiment 1
Table 1 below component nutritive salt is added first in tap water, and after each component fully dissolves, Jing ultraviolet sterilizations are added to
In 5 tons of bioreactor, chlorella is inoculated with, and ensures algae cell density for 1,000,000/milliliter, beginning chlorella light autotrophy；
Autotrophy condition of culture is：Using natural lighting, daytime, light intensity was in 10-130Klx, periodicity of illumination 12/12, blowing air amount 1.0L/
Min, carbon dioxide intake 0.05L/min, algae solution temperature are 26-33 DEG C, and pH is maintained at 7.0-9.0；In culture the 6th day
Rise, every harvesting in 1 day once, the algae solution of harvesting 50%, is supplemented with fresh medium and continues culture every time, until the 20th day, training
Support and terminate.Chlorella to harvesting carries out being centrifuged, cleans, is dried acquisition dry algae powder, determines its algae butt and frond internal protein
Content, whole culture period, the algae butt of harvesting is 25.226 kilograms (being shown in Table 4), and frond internal protein is 69.2%-70.9%
(being shown in Table 5)；The total nitrogen total phosphorus consumption rate situation of detection culture water body, total nitrogen total phosphorus utilization rate respectively reach 90.05%-
91.54% and 90.12%-91.56% (being shown in Table 6).
Table 1
| Carbamide | 0.38g/L |
| MgSO4·7H2O | 0.25g/L |
| KH2PO4 | 0.095g/L |
| Ferrous sulfate | 0.005g/L |
| CaCl2·2H2O | 0.02g/L |
| A5 liquid microelements | 0.8mL |
Remarks：Growth analyses, calculate average biological yield daily, and formula is as follows：K=(InN2–InN0)/(T2–T0), its
Middle K goes out efficiency for bio, used as one of harvesting basis for estimation；T0、T2For corresponding incubation time；N1And N2Respectively T0、T2
When Biomass；T0For being inoculated with the same day.
Embodiment 2
Table 2 below component nutritive salt is added first in tap water, and after each component fully dissolves, Jing ultraviolet sterilizations are added to
In 5 tons of bioreactor, chlorella is inoculated with, and ensures algae cell density for 1,200,000/milliliter, beginning chlorella light autotrophy；
Autotrophy condition of culture is：Using natural lighting, daytime, light intensity was in 10-130Klx, periodicity of illumination 12/12, blowing air amount
10.0L/min, carbon dioxide intake 0.5L/min, algae solution temperature are 26-33 DEG C, and pH is maintained at 7.0-9.0；In culture
50% algae solution from 6th day, every harvesting in 1 day once, is harvested every time, is supplemented with fresh medium and is continued culture, until the 20th
My god, culture terminates.Chlorella to harvesting carries out being centrifuged, cleans, is dried acquisition dry algae powder, determines its algae butt with egg in frond
White matter content, whole culture period, the algae butt of harvesting is 25.203 kilograms (being shown in Table 4), and frond internal protein is 69.2%-
70.9% (being shown in Table 5)；The total nitrogen total phosphorus consumption rate situation of detection culture water body, total nitrogen total phosphorus utilization rate respectively reach 90.12%-
91.9% and 90.22%-91.89% (being shown in Table 6).
Table 2
Embodiment 3
Table 3 below component nutritive salt is added first in tap water, and after each component fully dissolves, Jing ultraviolet sterilizations are added to
In 5 tons of bioreactor, chlorella is inoculated with, and ensures algae cell density for 1,400,000/milliliter, beginning chlorella light autotrophy；
Autotrophy condition of culture is：Using natural lighting, daytime, light intensity was in 10-130Klx, periodicity of illumination 12/12, blowing air amount 5.0L/
Min, carbon dioxide intake 0.2L/min, algae solution temperature are 26-33 DEG C, and pH is maintained at 7.0-9.0；In culture the 6th day
Rise, every harvesting in 1 day once, the algae solution of harvesting 50%, is supplemented with fresh medium and continues culture every time, until the 20th day, training
Support and terminate.Chlorella to harvesting carries out being centrifuged, cleans, is dried acquisition dry algae powder, determines its algae butt and frond internal protein
Content, whole culture period, the algae butt of harvesting is 25.216 kilograms (being shown in Table 4), and frond internal protein is 69.2%-70.9%
(being shown in Table 5)；The total nitrogen total phosphorus consumption rate situation of detection culture water body, total nitrogen total phosphorus utilization rate respectively reach 91.21%-
92.01% He
91.24%-92.09% (is shown in Table 6).
Table 3
| Carbamide | 0.40g/L |
| MgSO4·7H2O | 0.50g/L |
| KH2PO4 | 0.10g/L |
| Ferrous sulfate | 0.005g/L |
| CaCl2·2H2O | 0.025g/L |
| A5 liquid microelements | 0.9mL |
Table 4 harvests algae butt situation
Table 5 detects frond protein content after harvesting
Table 6 cultivates the total nitrogen total phosphorus situation of water body
Claims (5)
1. a kind of chlorella high efficiency light autotrophy cultural method, it is characterised in that the method comprises the steps：
(1) in tap water add nutritive salt fully to dissolve and obtain culture fluid, culture fluid is killed using ultraviolet sterilization device
Bacterium is processed, and in making sterilized rear culture fluid, content of microorganisms is less than 100cfu/ milliliters；
(2) culture fluid is added in bioreactor, access chlorella, and ensure algae cell density >=1,000,000/milliliter, enter
Row light autotrophy culture；
(3) culture adopts natural lighting, and daytime, light intensity was in 10-130Klx, periodicity of illumination 12/12, blowing air amount 1.0-
10.0L/min, carbon dioxide intake 0.05-0.5L/min, algae solution temperature are 26-33 DEG C, and pH is maintained at 7.0-9.0；
(4) in incubation, when intensity of illumination decays to 50% in algae solution, start harvesting, 50% algae solution of harvesting, is supplemented with every time
Fresh medium, such operation were repeated once every 1 day, and until chlorella culture enters phase of decline, culture terminates.
2. a kind of chlorella high efficiency light autotrophy cultural method as claimed in claim 1, it is characterised in that the supplementary culture
The method of liquid adopts at the uniform velocity mode to add, and once addition finishes use time and one week required time of algae solution circulation is identical.
3. a kind of chlorella high efficiency light autotrophy cultural method as claimed in claim 1, it is characterised in that the nutritive salt group
Into including：Carbamide 0.38-0.47g/L, MgSO4·7H2O 0.25-0.5g/L, KH2PO40.095-0.12g/L, ferrous sulfate
0.005-0.006g/L, CaCl2·2H2O 0.02-0.03g/L, A5 liquid microelement 0.8-1mL/L.
4. a kind of chlorella high efficiency light autotrophy cultural method as claimed in claim 1, it is characterised in that the nutritive salt group
Into including：Carbamide 0.4g/L, MgSO4·7H2O 0.5g/L, KH2PO40.1g/L, ferrous sulfate 0.005g/L, CaCl2·2H2O
0.025g/L, A5 liquid microelement 0.9mL/L.
5. a kind of chlorella high efficiency light autotrophy cultural method as claimed in claim 1, it is characterised in that the micro unit of the A5
Plain formula of liquid is H3BO3 2.86g/L、MnCl2·4H2O 1.86g/L、ZnSO4·7H2O 0.22g/L、Na2MoO4·2H2O
0.39g/L、CuSO4·5H2O 0.08g/L and Co (NO3)2·6H2O 0.05g/L。
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Cited By (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2019076004A (en) * | 2017-10-20 | 2019-05-23 | 清水建設株式会社 | Algae culture method and algae culture plant |
| CN110218653A (en) * | 2019-06-14 | 2019-09-10 | 阿尔格生命科学(江苏)有限公司 | A kind of chlorella animal feed production technology |
| CN116555039A (en) * | 2023-05-13 | 2023-08-08 | 华南理工大学 | Quick culture method of high-quality and high-biomass chlorella pyrenoidosa |
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| CN102021208A (en) * | 2010-11-16 | 2011-04-20 | 华东理工大学 | Method for rapidly accumulating micro-algae intracellular grease |
| CN103805514A (en) * | 2014-02-25 | 2014-05-21 | 中国科学院水生生物研究所 | Microalga photosynthetic aerobic high-density fermentation culture method utilizing inorganic nitrogen source and application |
| CN105985989A (en) * | 2015-01-30 | 2016-10-05 | 中国石油天然气股份有限公司 | Method for producing biodiesel from chlorella |
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Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102021208A (en) * | 2010-11-16 | 2011-04-20 | 华东理工大学 | Method for rapidly accumulating micro-algae intracellular grease |
| CN103805514A (en) * | 2014-02-25 | 2014-05-21 | 中国科学院水生生物研究所 | Microalga photosynthetic aerobic high-density fermentation culture method utilizing inorganic nitrogen source and application |
| CN105985989A (en) * | 2015-01-30 | 2016-10-05 | 中国石油天然气股份有限公司 | Method for producing biodiesel from chlorella |
Cited By (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JP2019076004A (en) * | 2017-10-20 | 2019-05-23 | 清水建設株式会社 | Algae culture method and algae culture plant |
| CN110218653A (en) * | 2019-06-14 | 2019-09-10 | 阿尔格生命科学(江苏)有限公司 | A kind of chlorella animal feed production technology |
| CN116555039A (en) * | 2023-05-13 | 2023-08-08 | 华南理工大学 | Quick culture method of high-quality and high-biomass chlorella pyrenoidosa |
| CN116555039B (en) * | 2023-05-13 | 2024-01-26 | 华南理工大学 | Quick culture method of chlorella pyrenoidosa |
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Application publication date: 20170322 Assignee: HAINAN JUNENG TECHNOLOGY INNOVATION RESEARCH INSTITUTE Co.,Ltd. Assignor: HAINAN GREEN ALGAE WORLD BIOTECHNOLOGY Co.,Ltd. Contract record no.: X2020980009684 Denomination of invention: An efficient photoautotrophic culture method for Chlorella vulgaris Granted publication date: 20190531 License type: Common License Record date: 20201221 |
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