CN101633894B - Culture medium of euglena gracilis and open type high-density culture method - Google Patents

Culture medium of euglena gracilis and open type high-density culture method Download PDF

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CN101633894B
CN101633894B CN2009100910114A CN200910091011A CN101633894B CN 101633894 B CN101633894 B CN 101633894B CN 2009100910114 A CN2009100910114 A CN 2009100910114A CN 200910091011 A CN200910091011 A CN 200910091011A CN 101633894 B CN101633894 B CN 101633894B
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substratum
thin euglena
culture medium
euglena
vitamins
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CN2009100910114A
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CN101633894A (en
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单东杰
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北京芳能科技有限公司
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Abstract

The invention provides a culture medium of euglena gracilis, which is added with different inorganic nutrient substances, in particular to nutrient components such as a mixture of ammonium chloride and urea, and the like. In an open culture process, the culture medium not only overcomes the defects that euglena gracilis is easy to pollute by microorganisms and bacteria so as to pollute the environment, and the like in the prior art, but also can ensure that the euglena gracilis quickly utilizes the inorganic nutrient components to accelerate cell division under the condition of not adding an organic carbon source. The culture medium of the invention shortens the grown cycle of the euglena gracilis by 2-5 days compared with a conventional culture medium and has a biomass accumulation being1-1.5 times higher than the conventional culture medium. The culture medium removes the organic carbon source of the conventional culture medium and supplements the carbon source in the mode of CO2, thus the culture medium not only can achieve the purpose of avoiding microorganism pollution, but also can increase the capacity of absorbing the CO2 of the euglena gracilis. In the process of the mass production and culture of the euglena gracilis, the culture medium can adsorb a large amount of industrial CO2 waste gas, thereby having important meaning to energy saving, emission reduction and environmental protection.

Description

Substratum of a kind of very thin Euglena and open type high-density cultural method thereof

Technical field

The present invention relates to a kind of substratum of very thin Euglena and method that open type high-density is cultured very thin Euglena of culturing.

Background technology

Very thin Euglena (Euglena gracilis) belongs to Euglenophyta, is the unicellular eukaryote of fresh water, and cell exposes no wall, is fusiform, long 35-66 μ m, wide 8-18 μ m has 8-28 sheet or discoid chromatoplast, 1 flagellum is about the long 0.15-1 of body doubly, and eyespot is obvious.It is the comparison equilibrated that the proteinic amino acid of very thin Euglena is formed, wherein the necessary 10 seed amino acid content of aquatic animal account for about 50% of its total amino acid content, and being rich in higher unsaturated fatty acid and basic lipid acid, is the high-quality bait of sea water and fresh water biology.Secondly, very thin Euglena can also be used for producing high value added products such as vitamins C, vitamin-E and β-Hu Luobusu.In addition, very thin Euglena be because growth rapidly, can tolerate sour environment, and well-grown when pH3.5 can tolerate the CO of high density 2, therefore, can be used as absorption industrial discharge CO 2Very thin Euglena resource is developed cultivation.

Generally, the cultural method of very thin Euglena has autotrophy cultivation and heterotrophism to cultivate two kinds.Autotrophy incubation growth speed is slow, is unfavorable for the accumulation of high-biomass.Heterotrophism is cultivated the at present Euglena of employing artificial mediums more, wherein contain organic nutrient substances such as yeast extract, extractum carnis and peptone, though the very thin Euglena speed of growth is very fast in this substratum, when open culture, very easily be subjected to microbial contaminations such as bacterium, be difficult to realize suitability for industrialized production.

Summary of the invention

For the cell density and the speed of growth that improves very thin Euglena, prevent that very thin Euglena is subjected to microbiological contamination such as bacterium and causes environmental pollution in the open culture process, the invention provides the different types of inorganic nutrient substance of a kind of interpolation, the nutritive ingredients such as mixture that particularly add ammonium chloride and urea, in the open culture process, feed certain density CO 2Gas, for very thin Euglena growth provides inorganic carbon source, the cell number accumulating level when reaching the heterotrophism cultivation is realized the open type high-density breed.

First purpose of the present invention provides a kind of substratum that very thin Euglena is grown fast.

Second purpose of the present invention provides a kind of method of using the very thin Euglena of culture medium culturing provided by the present invention.

The substratum that very thin Euglena is grown fast provided by the invention mainly comprises nitrogenous source and phosphorus source, and a spot of inorganic salt, trace element, VITAMIN and water.Wherein nitrogenous source is the mixture of ammonium chloride and urea, the phosphorus source can be potassium primary phosphate, inorganic salt can be one or more of molysite, sodium salt, calcium salt, magnesium salts, trace element can be in Manganous chloride tetrahydrate, rose vitriol, zinc sulfate, Sodium orthomolybdate and the copper sulfate one or more, and substratum of the present invention can also add other useful nutritive ingredients according to different purposes.

Very thin Euglena substratum called after EM provided by the present invention, its prescription is

Ammonium chloride 0.2～1.6g/L urea 30～120mg/L

Potassium primary phosphate 0.4～1.5g/L sal epsom 0.2～0.8g/L

Calcium chloride 0.01～0.05g/L Trisodium Citrate 0～1.0g/L

Ferric sulfate 3～30mg/L vitamins B 10.1～3mg/L

Trace element 0.5～2ml/L vitamins B 120.5～25 μ g/L

Disodium ethylene diamine tetraacetate 0.1～5mg/L.

Wherein Wei Liangyuansu compound method is:

Manganous chloride tetrahydrate 1.4～1.8g rose vitriol 0.3～1.6g

Zinc sulfate 0.3～0.5g Sodium orthomolybdate 0.1～0.4g

Copper sulfate 0.01～0.4g adding distil water is settled to 1L.

Very thin Euglena substratum provided by the present invention, prescription is preferably:

Ammonium chloride 0.5～1.4g/L urea 30～90mg/L

Potassium primary phosphate 0.8～1.3g/L sal epsom 0.4～0.6g/L

Calcium chloride 0.02～0.04g/L Trisodium Citrate 0～0.8g/L

Ferric sulfate 10～25mg/L vitamins B 10.5～2mg/L

Trace element 0.5～2ml/L vitamins B 122～15 μ g/L

Disodium ethylene diamine tetraacetate 1～3mg/L.

Wherein Wei Liangyuansu compound method is:

Manganous chloride tetrahydrate 1.5～1.6g rose vitriol 0.6～1.2g

Zinc sulfate 0.3～0.5g Sodium orthomolybdate 0.2～0.3g

Copper sulfate 0.05～0.2g adding distil water is settled to 1L.

The substratum of very thin Euglena provided by the present invention, its optimum formula is:

Ammonium chloride 1.0g/L urea 60mg/L

Dipotassium hydrogen phosphate 1.1g/L sal epsom 0.5g/L

Calcium chloride 0.03g/L Trisodium Citrate 0.8g/L

Ferric sulfate 20mg/L disodium ethylene diamine tetraacetate 2mg/L

Trace element 2ml/L vitamins B 11.5mg/L

Vitamins B 1210 μ g/L

Wherein Wei Liangyuansu prescription is as follows:

Manganous chloride tetrahydrate 1.5g rose vitriol 1.0g

Zinc sulfate 0.4g Sodium orthomolybdate 0.25g

Copper sulfate 0.1g adding distil water is settled to 1L.

If not refer in particular to, the chemical ingredients described in the culture medium prescription of the present invention is analytically pure commercial goods.

Use the method that EM substratum provided by the present invention is cultured very thin Euglena, comprise the steps:

1, preparation EM substratum;

2, be that 1: 4～1: 10 ratio is inoculated into very thin Euglena in the substratum in very thin Euglena and culture volume ratio in biological photoreactor system, liquid amount is 60-90%;

3, feeding concentration with the flow of 0.5-1.5L/min is 5～20% CO 2, the system pH of making is controlled at 3～5;

4, with the flow rate control of algae liquid at 10-18cm/s;

5, temperature is controlled at 22～27 ℃;

6, be 5000-30000LUX in intensity of illumination, light application time is under 10-14 hour/day the condition, cultivates can reach the cultivation terminal point in 6～9 days.

The described bioreactor of this patent method is a closed conduct formula bioreactor.

Use the method that the EM substratum is cultured very thin Euglena, culture condition is preferably: in very thin Euglena and culture volume ratio is that 1: 6～1: 9 ratio is inoculated into very thin Euglena in the substratum, and liquid amount is 70-90%; Feeding concentration with the flow of 0.8-1.3L/min is 15～20% CO 2, the system pH of making is controlled at 3～4; With the flow rate control of algae liquid at 14-16cm/s; In temperature is 24-26 ℃, and intensity of illumination is 13000-25000LUX, and light application time is under 11-13 hour/day the condition, cultivates can reach the cultivation terminal point in 6～8 days.

Use the method that the EM substratum is cultured very thin Euglena, optimal culture condition is: in very thin Euglena and culture volume ratio is that 1: 8 ratio is inoculated into very thin Euglena in the substratum, and liquid amount is 85%; Feeding concentration with 1.2L/min is 18% CO 2, the system pH of making is controlled at 3.5; With the flow rate control of algae liquid at 15cm/s; In temperature is 25-26 ℃, and intensity of illumination is 15000-20000LUX, and light application time is 12 hours/day, cultivates can reach the cultivation terminal point in 7 days.

Substratum of very thin Euglena provided by the present invention and cultural method, owing to added the ammonium chloride that is specially adapted to the breeding of very thin Euglena and the mixture of urea, both overcome in the prior art and to have cultured very thin Euglena and be subject to microorganism, bacterial contamination and bring defectives such as pollution to environment, can make very thin Euglena again under the situation of not adding organic carbon source, utilize the nutritive ingredient of substratum fast, impel cell to quicken division, growth cycle has shortened 2～5 days than traditional substratum, and being very beneficial for the accumulation of very thin Euglena biomass, the accumulation of the biomass of the very thin Euglena of culturing with substratum of the present invention is higher 1～1.5 times than using traditional substratum.Substratum provided by the invention has also been removed the organic carbon source in traditional heterotrophism substratum, to feed CO 2The form supplementary carbon source, can effectively reach the purpose of avoiding microbial contamination, can increase very thin Euglena again and absorb CO 2Ability, culture in the process of very thin Euglena in scale operation, absorb a large amount of CO that Industrial processes produced 2Waste gas has great importance to energy-saving and emission-reduction, protection environment.

Description of drawings

The growth curve of Fig. 1 for cultivating in the different substratum of very thin Euglena.: EM substratum; △: Euglena substratum;

The growth curve that Fig. 2 cultivates in the EM of different nitrogen sources content substratum for very thin Euglena.A, B, C, D, E, F and G represent respectively and add CO (NH 2) 20,20,40,60,80,100 and 120mg/L.

Fig. 3 be very thin Euglena in the 400L photo-bioreactor system, with complete EM culture medium culturing, feed 18%CO 2Cell number growth curve under the culture condition and dry weight growth curve.

Fig. 4 is that very thin Euglena is at the 400L photo-bioreactor system, with cultivating feeding 5%CO in the EM substratum of simplifying 2Cell number growth curve under the culture condition and dry weight growth curve.

Fig. 5 be very thin Euglena in the 400L photo-bioreactor system, use the EM culture medium culturing, feed 10%CO 2Cell number growth curve under the culture condition and dry weight growth curve.

Fig. 6 be very thin Euglena in the 400L photo-bioreactor system, use the EM culture medium culturing, feed 15%CO 2Cell number growth curve under the culture condition and dry weight growth curve.

Embodiment

Embodiment 1

1, the prescription of EM substratum is as follows:

Ammonium chloride 0.8g/L urea 60mg/L

Dipotassium hydrogen phosphate 1.0g/L sal epsom 0.2g/L

Calcium chloride 0.02g/L ferric sulfate 3mg/L

Disodium ethylene diamine tetraacetate 0.48mg/L

Trace element 1ml/L, wherein Wei Liangyuansu compound method is: get Manganous chloride tetrahydrate 1.8g, rose vitriol 1.5g, zinc sulfate 0.4g, Sodium orthomolybdate 0.2g, copper sulfate 0.02g, adding distil water is settled to 1L.

Vitamins B 10.1mg/L vitamins B 120.5 μ g/L

With the above-mentioned substance dissolving, be settled to 1L with distilled water, adjust pH to 3.5,121 ℃ of autoclaving 20min with hydrochloric acid.

Shake adding 100ml EM substratum in the bottle at 250ml, inoculum density is 5 * 10 5Individual cells/ml at 25 ℃ of temperature, light: dark=12: 12, light intensity 5000LUX, is cultivated under the conditions such as shaking speed 150r/min.Cultivated through 7 days, the maximum biomass of very thin Euglena can reach 3.6 * 10 7Individual cells/ml (see figure 1).

2, the Euglena culture medium prescription is as follows:

Sodium acetate 1g/L

Beef extract 1g/L

Peptone 2g/L

Yeast extract 2g/L

Calcium chloride 0.01g/L

With the above-mentioned substance dissolving, be settled to 1L with distilled water, transfer pH to 3.5,121 ℃ of autoclaving 20min with hydrochloric acid.Shake adding 100ml Euglena substratum in the bottle at 250ml, inoculum density is 5 * 10 5Individual cells/ml at 25 ℃ of temperature, light: dark=12: 12, light intensity 5000LUX, is cultivated under the conditions such as shaking speed 150r/min.Cultivate after 7 days, the maximum biomass of very thin Euglena can reach 2.4 * 10 7Individual cells/ml (see figure 1).

Result according to above-mentioned two kinds of very thin Euglenas of culture medium culturing can learn, use EM substratum provided by the present invention, can make very thin Euglena under light autotrophy condition, growth cycle has shortened 2～5 days than traditional substratum, and the accumulation of its biomass is higher 1～1.5 times than using traditional heterotrophism substratum.Shortcomings such as very thin Euglena poor growth of traditional light autotrophy culture medium culturing and biomass are extremely low have been overcome.Can obtain than using the very thin Euglena biomass of traditional light autotrophy cultivation more than 5 times with the EM substratum.

EM substratum provided by the present invention is compared with Euglena heterotrophism substratum, do not contain organic nutrient substances such as extractum carnis, peptone, yeast extract in its prescription, when extensive open culture, be not subject to the infecting of microorganism of bacterium etc., but also can accumulate higher biomass, reach the effect of high-density culture.

Embodiment 2

The EM culture medium prescription is as follows:

Ammonium chloride 0.2g/L

Urea is respectively 0,20,40,60,80,100 and 120mg/L

Dipotassium hydrogen phosphate 1.2g/L sal epsom 0.2g/L

Calcium chloride 0.02g/L ferric sulfate 3mg/L

Disodium ethylene diamine tetraacetate 0.48mg/L

Trace element 0.5ml/L, wherein Wei Liangyuansu compound method is: get

Manganous chloride tetrahydrate 1.8g, rose vitriol 1.6g, zinc sulfate 0.5g, Sodium orthomolybdate 0.2g, copper sulfate 0.02g, adding distil water is settled to 1L.

Vitamins B 10.25mg/L vitamins B 1220 μ g/L

With the above-mentioned substance dissolving, be settled to 1L with distilled water, transfer pH to 3.5,121 ℃ of autoclaving 20min with hydrochloric acid.Shake the EM substratum that adds 100ml in the bottle, wherein CO (NH at 250ml 2) 2Starting point concentration be respectively 0,20,40,60,80,100 and 120mg/L, inoculum density is 4.5 * 10 5Individual cells/ml is 12: 12, light intensity 5000lux in 25 ℃ of temperature, photoperiod, cultivates under the shaking speed 150r/min condition.The cell enlargement situation as shown in Figure 2, A, B, C, D, E, F, G, H and I represent respectively and add CO (NH among the figure 2) 2Amount is respectively 0,20,40,60,80,100 and the growing state of 120mg/L.Wherein, do not containing urea and containing in the substratum of higher concentration urea (120mg/L), this very thin Euglena growth is slower, and biomass is also lower, cultivates after 7 days, can only reach 0.9 * 10 7With 1.2 * 10 7Individual cells/ml.But along with the increase of the urea amount that adds, the growth of very thin Euglena is rapider, and when the urea content in the substratum reached 60mg/L, very thin Euglena growth was the fastest, promptly enters logarithmic phase in 3 days, and after cultivating in 7 days, the maximum biomass of very thin Euglena reaches 2.63 * 10 7Individual cells/ml.But along with the further increase of urea amount (＞60mg/L), it is slow that the growth of very thin Euglena becomes on the contrary.As seen, add the accumulation that suitable urea (is good with 60mg/L) can promote very thin Euglena biomass greatly, but excessive urea also can suppress the growth of very thin Euglena.

Embodiment 3

The prescription of EM substratum is as follows:

Ammonium chloride 1.6g/L urea 60mg/L

Dipotassium hydrogen phosphate 1.5g/L sal epsom 0.4g/L

Calcium chloride 0.02g/L ferric sulfate 10mg/L

Disodium ethylene diamine tetraacetate 5mg/L

Trace element 1.5ml/L, wherein Wei Liangyuansu prescription is: Manganous chloride tetrahydrate 1.8g, rose vitriol 1.5g, zinc sulfate 0.4g, Sodium orthomolybdate 0.2g, copper sulfate 0.02g

Vitamins B 1The 3mg/L vitamins B 1225 μ g/L;

In the biological photoreactor of 400L system, add the EM substratum, liquid amount is 90%, inoculation algae kind density is 2 * 10 5Individual cells/ml, feeding concentration with the flow of 0.6-1L/min is 18% CO 2Gas, the system pH of making is controlled at 3～5, and temperature is controlled at 25 ℃, and illumination is controlled at 18000-30000LUX, and light application time was controlled at 12 hours/day.The result shows that very thin Euglena is well-grown in this photo-bioreactor system.Cultivating and promptly entered exponential phase of growth on the 3rd day, in the time of the 6th day, reach maximum cell density, is 3.83 * 10 7Individual cells/ml (see figure 3), dry weight are the 2.65g/L (see figure 3).

Embodiment 4

The prescription of EM substratum is as follows:

Ammonium chloride 1.6g/L urea 60mg/L

Dipotassium hydrogen phosphate 1.5g/L sal epsom 0.4g/L

Calcium chloride 0.04g/L ferric sulfate 30mg/L

Manganous chloride tetrahydrate 1.5mg/L vitamins B 1225 μ g/L;

In the biological photoreactor of 400L system, add the EM substratum, liquid amount is 80%, inoculation algae kind density is 3 * 10 5Individual cells/ml, feeding concentration with the flow of 1-1.5L/min is 5% CO 2Gas, the system pH of making is controlled at 3～5, and temperature is controlled at 25 ℃, and illumination is controlled at 10000-30000LUX, and light application time was controlled at 11 hours/day.Cultivating and entered exponential phase of growth on the 5th day, in the time of the 8th day, reach maximum cell density, is 2.53 * 10 7Individual cells/ml (see figure 4), dry weight are the 1.75g/L (see figure 4).

Embodiment 5

The prescription of EM substratum is as follows:

Ammonium chloride 1.0g/L urea 60mg/L

Potassium primary phosphate 0.5g/L sal epsom 0.8g/L

Calcium chloride 0.01g/L Trisodium Citrate 1.0g/L

Ferric sulfate 15mg/L vitamins B 11mg/L

Trace element 1.8ml/L vitamins B 120.5 μ g/L

Disodium ethylene diamine tetraacetate 5mg/L.

Wherein Wei Liangyuansu compound method is:

Manganous chloride tetrahydrate 1.5g rose vitriol 0.4g

Zinc sulfate 0.4g Sodium orthomolybdate 0.2g

Copper sulfate 0.02g adding distil water is settled to 1L.

In the biological photoreactor of 400L system, add the EM substratum, liquid amount is 75%, inoculation algae kind density is 5 * 10 5Individual cells/ml, feeding concentration with the flow of 0.8-1.2L/min is 10% CO 2Gas, the system pH of making is controlled at 3.5～4, and temperature is controlled at 23 ℃-26 ℃, and illumination is controlled at 6000-20000LUX, and light application time was controlled at 13 hours/day.Cultivating and entered exponential phase of growth on the 4th day, in the time of the 7th day, reach maximum cell density, is 3.53 * 10 7Individual cells/ml (see figure 5), dry weight are the 2.47g/L (see figure 5).

Embodiment 6

The prescription of EM substratum is as follows:

Ammonium chloride 1.4g/L urea 60mg/L

Dipotassium hydrogen phosphate 0.6g/L sal epsom 0.6g/L

Calcium chloride 0.05g/L Trisodium Citrate 0.2g/L

Ferric sulfate 20mg/L disodium ethylene diamine tetraacetate 2.5mg/L

Trace element 1.2ml/L vitamins B 1225 μ g/L.

Wherein micro-compound method is as follows:

Manganous chloride tetrahydrate 1.5g rose vitriol 0.8g

Zinc sulfate 0.4g Sodium orthomolybdate 0.4g

Copper sulfate 0.3g

Adding distil water is settled to 1L.

In the biological photoreactor of 400L system, add the EM substratum, liquid amount is 85%, inoculation algae kind density is 9 * 10 5Individual cells/ml, feeding concentration with the flow of 0.9-1.4L/min is 15% CO 2Gas, the system pH of making is controlled at 3～4, and temperature is controlled at 24 ℃, and illumination is controlled at 8000-15000LUX, and light application time was controlled at 14 hours/day.Cultivating and entered exponential phase of growth on the 3rd day, in the time of the 6th day, reach maximum cell density, is 4.53 * 10 7Individual cells/ml (see figure 6), dry weight are the 3.13g/L (see figure 6).

Claims (7)

1. the substratum of the very thin Euglena of high-density breeding, comprise nitrogenous source, phosphorus source, small amounts of inorganic salt, trace element, VITAMIN and water, it is characterized in that described nitrogenous source is the mixture of ammonium chloride and urea, described phosphorus source is a potassium primary phosphate, described inorganic salt are one or more of molysite, sodium salt, calcium salt, magnesium salts, described trace element is one or more in Manganous chloride tetrahydrate, rose vitriol, zinc sulfate, Sodium orthomolybdate and the copper sulfate, and described VITAMIN is a vitamins B 1, vitamins B 12One or more, its prescription is:
Ammonium chloride 0.2～1.6g/L urea 30～120mg/L
Dipotassium hydrogen phosphate 0.4～1.5g/L sal epsom 0.2～0.8g/L
Calcium chloride 0.01～0.05g/L Trisodium Citrate 0～1.0g/L
Ferric sulfate 3～30mg/L disodium ethylene diamine tetraacetate 0.1～5mg/L
Trace element 0.5～2ml/L vitamins B 10.1～3mg/L
Vitamins B 120.5～25 μ g/L.
Wherein trace element formula is:
Manganous chloride tetrahydrate 1.4～1.8g rose vitriol 0.3～1.6g
Zinc sulfate 0.3～0.5g Sodium orthomolybdate 0.1～0.4g
Copper sulfate 0.01～0.4g adding distil water is settled to 1L.
2. the substratum of the very thin Euglena of breed according to claim 1 is characterized in that prescription is:
Ammonium chloride 0.5～1.4g/L urea 30～90mg/L
Potassium primary phosphate 0.8～1.3g/L sal epsom 0.4～0.6g/L
Calcium chloride 0.02～0.04g/L Trisodium Citrate 0～0.8g/L
Ferric sulfate 10～25mg/L vitamins B 10.5～2mg/L
Trace element 0.5～2ml/L vitamins B 122～15 μ g/L
Disodium ethylene diamine tetraacetate 1～3mg/L.
Wherein Wei Liangyuansu prescription is as follows:
Manganous chloride tetrahydrate 1.5～1.6g rose vitriol 0.6～1.2g
Zinc sulfate 0.3～0.5g Sodium orthomolybdate 0.2～0.3g
Copper sulfate 0.05～0.2g adding distil water is settled to 1L.
3. the substratum of the very thin Euglena of breed according to claim 1 is characterized in that prescription is:
Ammonium chloride 1.0g/L urea 60mg/L
Dipotassium hydrogen phosphate 1.1g/L sal epsom 0.5g/L
Calcium chloride 0.03g/L Trisodium Citrate 0.8g/L
Ferric sulfate 20mg/L disodium ethylene diamine tetraacetate 2mg/L
Trace element 2ml/L vitamins B 11.5mg/L
Vitamins B 1210 μ g/L
Wherein Wei Liangyuansu prescription is as follows:
Manganous chloride tetrahydrate 1.5g rose vitriol 1.0g
Zinc sulfate 0.4g Sodium orthomolybdate 0.25g
Copper sulfate 0.1g adding distil water is settled to 1L.
4. method of culturing very thin Euglena with the described substratum of claim 1-3 is characterized in that adopting following steps:
(1), preparation substratum;
(2), in biological photoreactor system, be that 1: 4～1: 10 ratio is inoculated into very thin Euglena in the substratum in very thin Euglena and culture volume ratio, liquid amount is 60-90%;
(3), feeding concentration with the flow of 0.5-1.5L/min is 5～20% CO 2, the system pH of making is controlled at 3～5;
(4), with the flow rate control of algae liquid at 10-18cm/s;
(5), temperature is controlled at 22～27 ℃;
(6), be that 5000-30000LUX, light application time are under 10-14 hour/day the condition in intensity of illumination, cultivated 6～9 days.
5. the method for the very thin Euglena of breed according to claim 4 is characterized in that adopting following steps:
(1), preparation substratum;
(2), in biological photoreactor system, be that 1: 6～1: 9 ratio is inoculated into very thin Euglena in the substratum in very thin Euglena and culture volume ratio, liquid amount is 70-90%;
(3), feeding concentration with the flow of 0.8-1.3L/min is 15～20% CO 2, the system pH of making is controlled at 3～4;
(4), with the flow rate control of algae liquid at 14-16cm/s;
(5), temperature is controlled at 24～26 ℃;
(6), be 13000-25000LUX in intensity of illumination, light application time is under 11-13 hour/day the condition, to cultivate 6～8 days.
6. the method for the very thin Euglena of breed according to claim 5 is characterized in that adopting following steps:
(1), preparation substratum;
(2), in biological photoreactor system, be that 1: 8 ratio is inoculated into very thin Euglena in the substratum in very thin Euglena and culture volume ratio, liquid amount is 85%;
(3), feeding concentration with the flow of 1.2L/min is 18% CO 2, the system pH of making is controlled at 3.5;
(4), with the flow rate control of algae liquid at 15cm/s;
(5), temperature is controlled at 25～26 ℃;
(6), be 15000-20000LUX in intensity of illumination, light application time is under 12 hours/day the condition, to cultivate 7 days.
7. the method for the very thin Euglena of breed according to claim 5 is characterized in that described bioreactor is the closed conduct formula.
CN2009100910114A 2009-08-20 2009-08-20 Culture medium of euglena gracilis and open type high-density culture method CN101633894B (en)

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